MODIFIED ATMOSPHERE PACKAGING OF READY TO COOK IDLI
Transcription
MODIFIED ATMOSPHERE PACKAGING OF READY TO COOK IDLI
MODIFIED ATMOSPHERE PACKAGING OF READY TO COOK IDLI BATTER THESIS Submitted to the Pondicherry University for the award of the degree DOCTOR OF PHILOSOPHY In FOOD SCIENCE AND NUTRITION By M. DURGADEVI DEPARTMENT OF FOOD SCIENCE AND TECHNOLOGY PONDICHERRY UNIVERSITY PUDUCHERRY-605 014 DECEMBER – 2011 Pondicherry University Department of Food Science and Technology Dr. H. PrathapKumar Shetty Reader and Head (i/c) CERTIFICATE This is to certify that this thesis entitled “Modified atmosphere packaging of ready to cook idli batter” is a record of research work done by the candidate, Miss. M. Durgadevi, during the period of her study in the Department of Food Science and Technology, Pondicherry University, Puducherry under my supervision. This research work has not previously been formed the basis for award of any degree, diploma, associateship or fellowship or any other similar title and that it represents entirely an independent work of the candidate. Signature of Guide Signature of H.O.D Signature of Dean R.V. Nagar, Kalapet, Puducherry- 605 014, India Ms. M. Durgadevi Department of food Science and Technology Pondicherry University Puducherry-605 014 DECLARATION I hereby declare that the thesis entitled “Modified atmosphere packaging of ready to cook idli batter” submitted to Pondicherry University in partial fulfillment of the requirement for the award of Doctor of Philosophy in Food Science and Nutrition, is my original work and has not been previously formed the basis for award of any degree, diploma, associateship, fellowship or any other similar title. Puducherry Date: (M. Durgadevi) ACKNOWLEDGEMENT I express my humble gratitude to the Vice Chancellor, Pondicherry University, Dr. J.A.K.Tareen for granting me permission to carry out my research work in the Department of Food Science and Technology, Pondicherry University as the first Scholar of the department. I express my thanks to Dean of Life Sciences, Dr. P.P.Mathur for providing all necessary facilities to the Department to carry out my research. I express my profound gratitude to my supervisor Dr. H. PrathapKumar Shetty, Reader and Head (i/c), Department of Food Science and Technology, Pondicherry University who explored in me the art of doing research and helped me advance in my independent thinking. I appreciate the freedom given to me throughout the research period and the ceaseless and dynamic guidance. I also thank him for the necessary facilities and arrangements provided to carry out my research successfully. I thank my Doctoral committee members Dr. Jeevarathinam (Head, Department of Microbiology) and Dr. Shakthivel (Head, Department of Biotechnology) for providing valuable suggestions and constructive criticism on my work. I would like to express my gratefulness to my Doctoral committee member Dr. John Don Bosco, Reader Department of Food Science and Technology for his guidance, granting permission to operate the equipments in his Product Development Lab. I also thank him for his moral support and encouragement to complete my work successfully. I am happy to thank Dr. Narayanasamy Sangeetha, Assistant Professor, Department of Food Science and Technology for her continuous support, and for granting permission to carry out my experiments in Food and Nutrition lab. I sincerely thank Dr. S.Hari priya, Assistant Professor, Department of Food Science and Technology for her valuable suggestions and comments on my work progress and for helping me throughout the study period. I thank Dr. K.V.Sunooj, Assistant Professor, Department of Food Science and Technology for motivating me throughout the study period and helping me in operating texture analyzer. I express my sincere gratitude to Dr. R.Ravi, Department of Sensory Sciences, CFTRI, for guiding me in statistical analysis, for making himself available to clarify all queries whenever required, for the patience shown towards my work progress and for supporting me to complete the study successfully. I express my gratitude to Dr. Vishnuvardhan, Assistant Professor, Department of Statistics, Pondicherry University and Miss. Anusuya, guest lecturer, Department of Food Science and technology for supporting my study with statistical guidance. I thank my non-teaching staffs Mrs. Gomathi, Mr. Chakravarthy, Mrs. Kolanchiammal, Miss. Chandra, Mrs. Valliammal, and Mr. Angappin for helping me with the needed amenities. I thank Dr. Sundar for inspiration and guidance given to carry out my research. I thank all my colleagues S. Uma Maheswari, D. Sumitha, C.Saravanan, Sanjay Prathap Singh, P. Vasanthakumari, M. Pushpadevi, M. Shakthi Kandamoorthy, K.Devi, S.J.Cynthia, Soumya Bhol, Ravindra Kumar Agarwal, Ch.Koteeswara Reddy, Shabir and P.Vandarkuzhali, for their help rendered during my research period and for the moral support given. I also thank research assistants Santhalakshmi and Venkadesaperumal for their timely help. I thank to Dr. Kumari who encouraged me to do my research work at Pondicherry University. I thank Dr. Saraswathi, Dr. K.S. Pushpa, Dr. S.S. Vijayanchali and Mrs. Kavitha Mythili, Mr. Devanarayanan, Dr. C. K. Venil for their encouragement. I express my sincere thanks to University Grants Commission for providing me Junior Research Fellowship without which I must not have carried out my research work. I express my thanks to Tamil Nadu Rice Research Institute, TNAU, and Aduthurai for providing me ADT3 variety black gram dhal throughout my study period. I also express my thanks to SID, IISc, Bangalore for helping me to carry out LC-MS analysis for my samples. I thank all the respondents of my study who patiently supported my survey. I thank Dr.Victor Anand raj and family, Dr. Bhusan Sudhakar and family, Dr. Vellari and family, Dr.Shiva Sankar and family, Dr. Tokozu and family, Dr.Sherry and family, Dr.Hannah Rachael Vasanthi, Dr.Rejena, Anandh uncle, Rena aunty, Ellan anna, Sujitha akka, Pastor Kima, Deepa aunty and family, Phelix uncle and family and all PUCF friends for encouraging my research work through prayers and for the care shown in my personal life. My special thanks to my dear friends Sharen Elizabeth Thomas, Gita, Aliza Princy, Ithayamalar, Venkataramaiah, Murali, Srikanth, Ravindra, Ramachitra and family, Shanmuga Sundari, Indhumathi, Kayalvizhi, Suman, Rajinish, Vijaya Bharathy, Vikram, Vinayagam, Mayilvaganan, Supriya, Maya and Vijayalakshmi for their continuous support to carry out my work. My humble thanks to my parents Mr.R.Manoharan and Mrs.M.Janaki , my sister Mrs.M.Ahiladevi, brother-in-law Mr.J.Arokiaraj, my brother Mr. M. Prasanna Venkateshwaran, my sister-in-law Mrs.Jothi , my niece Baby. A. Thilaka Catheriene and Mr. K. Durga Prasad and family for all moral support, financial assistance, love, care, and encouragement showered on me throughout my study period. Above all, I owe my humble gratefulness and faithfulness to my Lord Almighty for His continuous grace and blessings to finish my research work successfully. ABBREVATIONS ADT3 - Aduthurai ANOVA - Analysis of Variance BMI - Body Mass Index BV - Biological Value CA - Controlled Atmosphere CCRD - Central Component Rotatable design DC - Digestibility Co-efficient FAA - Free Amino Acids FAN - Free α-amino Nitrogen FER - Feed Efficiency Ratio FOS - Fructo-oligosaccharide HM - High Molecule HUDCO - Housing and Urban Development Corporation IR20 - International Rice 20 LCMS - Liquid Chromatography Mass Spectrometry LDPE - Low Density Poly Ethylene MAP - Modified Atmosphere Packaging MS - Mild Steel NDOs - Non Digestible oligosaccharides NPU - Net Protein Utilization NSP - Non Starch polysaccharides PCA - Principal Component Analysis PER - Protein Efficiency Ratio PP - Poly Propylene QDA - Quantitative Descriptive Analysis RA - Relative abundance RPER - Relative Protein Efficiency Ratio RSM - Response Surface Methodology SCFA - Short Chain fatty Acid TAG - Triacylglycerol TPA - Texture Profile Analysis VLDL - Very Low density Lipoprotein WHO - World Health Organization v/v - volume/volume w/w - weight/weight h - Hour Pa - Pascal mm - Milli meter mL - Milli liter g - Gram µL -Micro liter N - Newton Ns - Newton second CONTENTS 1 1.1 1.2 1.2.1 1.2.2 1.2.3 1.2.3.1 1.2.3.2 1.2.4 1.2.5 1.2.6 1.2.6.1 1.2.6.2 1.2.6.3 1.2.6.4 1.2.7 Introduction and review of literature Introduction Review of literature Significance of fermented foods Positive health outcomes of breakfast consumption Idli and its properties Nutritional composition of idli Physico- chemical parameters of idli Rice - a staple food grain in idli making Black gram- a protein source in idli making Oligosaccharides in foods Conversion of polysaccharides into oligosaccharides Physiological properties of oligosaccharides Animal studies on oligosaccharides Applications of FOS in food formulations Modified atmosphere packaging 1 1 2 2 4 5 5 6 9 12 13 14 14 17 18 18 2 23 2.1 2.2 2.2.1 2.2.2 2.2.3 2.2.4 2.3 2.3.1 2.3.2 2.3.3 2.3.4 2.4 Transition in the preparation and consumption of idli among the population of Puducherry Introduction Materials and Methods Selection of area Selection of tool for data collection Selection of respondents Data analysis Results and discussion Socio-economic profile of the selected respondents Consumption pattern of breakfast among the selected respondents Preparation of idli at household level Preference for commercial idli batter against home-made batter Conclusion 3 3.1 3.2 3.2.1 3.2.2 3.2.3 Texture optimization of idli Introduction Materials and methods Materials Preparation of idli Experimental design 36 36 38 38 38 38 23 23 23 24 24 24 24 24 27 29 33 35 3.2.3.1 3.2.3.2 3.2.3.3 3.2.3.4 3.2.4 3.3 3.3.1 3.3.2 3.3.3 3.3.4 3.3.5 3.3.6 3.3.7 3.4 Response surface methodology Optimization of idli Instrumental color measurement Texture profile analysis (TPA) Statistical analysis Results and discussion Effect of rice varieties on rice batter volume Effect of black gram on batter volume Effect of ratios of rice to black gram dhal on batter volume Response surfaces Instrumental color measurement of idli Texture parameters Simultaneous optimization Conclusion 38 40 40 41 45 45 45 46 47 50 50 54 63 64 4 4.1 4.2 4.2.1 4.2.2 4.2.3 4.2.3.1 4.2.3.2 4.2.3.3 4.2.3.4 4.2.4 4.2.4.1 4.3 4.3.1 4.3.1.1 4.3.1.2 4.3.1.3 4.3.2 4.3.2.1 4.3.2.2 4.3.3 4.3.4 4.3.5 4.3.6 4.4 Process optimization of idli using sensory attributes Introduction Materials and methods Materials Preparation of idli Experimental design Response surface methodology Optimization of idli using RSM Sensory analysis of idli Quantitative descriptive analysis (QDA) Statistical analysis of data Principal component analysis (PCA) Results and discussion Desirable parameters of idli Color Fluffiness and sponginess of idli Fermented aroma Negative drivers of liking Stickiness of the idli Sourness of idli Overall quality of the idli Simultaneous optimization Principal component analysis (PCA) Optimization of texture and sensory attributes Conclusion 65 65 66 66 66 67 67 67 68 68 68 69 70 70 70 71 74 74 78 78 80 80 81 82 83 5 5.1 5.2 5.2.1 5.2.2 5.2.3 5.3 5.3.1 5.3.2 5.4.3 5.4 Nutritional composition of optimized idli Introduction Materials and methods Nutritional composition of the idli Determination of fatty acids and alcohols Determination of oligosaccharides Results and discussion Nutritional composition of idli Fatty acids and alcohols in optimized idli Disaccharides and oligosaccharides in optimized idli Conclusion 84 84 84 84 84 85 85 85 87 92 97 6 6.1 6.2 6.2.1 6.2.2 6.2.2.1 6.2.2.2 6.2.2.3 6.2.2.4 6.2.2.5 6.3 6.3.1 6.3.2 6.3.3 6.3.4 6.4 Improving the shelf-life of ready to cook idli batter Introduction Materials and methods Materials Methods Preparation of batter Selection of packaging materials Experiment I Experiment II Experiment III Results and discussion Respiration dynamics Experiment I Experiment II Experiment III Conclusion Executive summary and conclusion Practical implications / recommendations References Annexures List of publications 98 98 98 98 99 99 99 102 102 103 104 104 105 111 128 131 132 135 LIST OF PLATES 2.1 3.1 3.2 3.3 3.4 6.1 6.2 City map of Puducherry showing selected areas for the study Color flex Cutting idli with the designed mould One inch cubic mould and SMS/75mm compression probe Texture analyzer Modified atmosphere packaging machine Respirometer connected to gas analyzer 23 41 43 44 44 100 101 LIST OF TABLES 2.1 2.2 2.3 2.4 2.5 2.6 3.1 3.2 3.3 3.4 3.5 3.6 3.7 3.8 3.9 4.1 4.2 4.3 4.4 4.5 4.6 4.7 5.1 5.2 5.3 6.1 6.2 6.3 6.4 6.5 6.6 Age and sex wise distribution of the selected respondents (N=300) Educational and economic status of the selected respondents (N=300) Details on breakfast consumption (N=300) Idli preparation at household level (N=300) Fermentation time and measures followed to control fermentation of idli batter at households (N=300) Details about purchase of commercial idli batter (N=300) Central composite rotatable design: coded and actual values of independent variables Effect of rice varieties on the batter volume after fermentation Effect of black gram (var. Adt 3) on the batter volume after fermentation Idli batter volume characteristics as affected by parboiled rice and black gram dhal (without husk) Experimental design: CCRD with actual levels of independent variables for color parameters Experimental design: CCRD with coded and actual levels of independent variables for TPA Regression co-efficient for dependent TPA parameters Analysis of variance (ANOVA) for dependent TPA parameters: f values Simultaneous optimization of process parameters by desirability approach Sensory attributes used for sensory analysis of idli Experimental designs and mean scores of desirable sensory attributes Experimental designs and mean scores of undesirable sensory attributes Regression co-efficient for sensory parameters Regression co-efficient for overall quality of idli Simultaneous optimization of process parameters by desirability approach Combined analysis of texture and sensory attributes Proximate composition of optimized idli List of fatty acids and alcohols List of disaccharides and oligosaccharides Thickness of packaging materials map of idli batter Gas treatment used in experiment II Gas treatment used in experiment III Change in gas mixture over storage period Concentration of gases in LDPE (0.014mm) during the storage period Concentration of gases in LDPE (0.012mm) during the storage period 25 27 28 29 33 34 40 47 48 48 51 56 62 62 64 69 71 75 79 79 81 83 86 87 93 99 103 103 106 112 113 6.7 6.8 6.9 6.10 6.11 6.12 6.13 2 2.1 2.2 2.3 3.1 3.2a 3.2b 3.2c 3.3a 3.3b 3.3c 3.3d 3.4a 3.4b 3.5a 3.5b 3.5c 3.5d 3.5e 3.5f Concentration of gases in LDPE (0.009mm) during the storage period Sensory scores of the product made from batter packaged in LDPE (0.014mm) Sensory scores of the product made from batter packed in LDPE (0.012mm) Sensory scores of the product made from batter packaged in LDPE (0.009mm) Comparison of gas mixture on the first day and seventh day of storage TPA parameters of idli made from map batter Overall quality of idli LIST OF FIGURES Socio-economic profile of the selected respondents Details on idli preparation at household level Details on fermentation time and measures to fermentation Flowchart showing work design for TPA of idli A. Effect of rice varieties on batter volume after fermentation B. Effect of type of black gram dhal on batter volume after fermentation C. Effect of ratios of rice to black gram dhal on batter volume after fermentation Response surface graph showing relation between independent parameters on L* Response surface graph showing relation between independent parameters on a* Response surface graph showing relation between independent parameters on b* Response surface graph showing relation between independent parameters on chroma Fig.3.3.a texture profile of idli made of ratio 3:1.25 at 12 h fermentation time Fig.3.3.b texture profile of idli made of ratio 3:2 at 12 h fermentation time Fig.3.3.a response surface graph showing relation between independent parameters on hardness Response surface graph showing relation between independent parameters on adhesiveness Response surface graph showing relation between independent parameters on springiness Response surface graph showing relation between independent parameters on cohesiveness Response surface graph showing relation between independent parameters on chewiness Response surface graph showing relation between independent parameters 114 125 125 126 129 129 130 26 31 32 42 49 49 49 52 52 53 53 55 55 58 58 59 59 60 61 4.1a 4.1b 4.1c 4.1d 4.2a 4.2b 4.2c 4.2d 4.3 4.4 5.1 5.2 5.3 5.4 5.5 5.6 6.1 6.2a 6.2b 6.3a 6.3b 6.4a 6.4b 6.5a 6.5b 6.6a 6.6b 6.7a 6.7b 6.8a on resilience Response surface graph for color Response surface graph for fluffiness Response surface graph for sponginess Response surface graph for fermented aroma Response surface graph for compactness Response surface graph for firmness Response surface graph for stickiness Response surface graph for sourness Response surface graph showing the overall quality of the idli Principal component analysis (PCA) biplot of experimental design points over sensory attributes of idli Typical chromatogram and mass spectra showing fatty acids in nonfermented batter Typical chromatogram and mass spectra showing fatty acids and alcohols in fermented batter Typical chromatogram and mass spectra showing fatty acids in optimized idli Typical chromatogram and mass spectra showing disaccharides and oligosaccharides in non-fermented batter Typical chromatogram and mass spectra showing disaccharides and oligosaccharides in fermented batter Typical chromatogram and mass spectra showing disaccharides and oligosaccharides in optimized idli Change in gas concentration during its fermentation time Treatment 1 (0% CO2) showing change in CO2 level (%) among different packaging material Treatment 1 (0% CO2) showing change in O2 level (%) among different packaging material Treatment 2 (5% CO2) showing change in CO2 level (%) among different packaging material Treatment 2 (5% CO2) showing change in O2 level (%) among different packaging material Treatment 4 (15% CO2) showing change in CO2 level (%) among different packaging material Treatment 4 (15% CO2) showing change in O2 level (%) among different packaging material Treatment 1 (0% CO2 and 15% O2) showing percentage of CO2 (%) Treatment 1 (0% CO2 and 15% O2) showing percentage of O2 (%) Treatment 2 (5% CO2 and 15% O2) showing percentage of CO2 (%) Treatment 2 (5% CO2 and 15% O2) showing percentage of O2 (%) Treatment 3 (10% CO2 and 15% O2) showing percentage of CO2 (%) Treatment 3 (10% CO2 and 15% O2) showing percentage of O2 (%) Treatment 4 (15% CO2 and 15% O2) showing percentage of CO2 (%) 72 72 73 73 76 76 77 77 80 82 88 89 90 94 95 96 104 108 108 109 109 110 110 115 115 116 116 117 117 118 6.8b 6.9a 6.9b 6.10a 6.10b 6.11a 6.11b 6.12a 6.12b 6.13a 6.13b 6.14a 6.14b 6.15 6.16 Treatment 4 (15% CO2 and 15% o2) showing percentage of O2 (%) Treatment 5 (0% CO2 and 17.5% O2) showing percentage of CO2 (%) Treatment 5 (0% CO2 and 17.5% O2) showing percentage of O2 (%) Treatment 8 (15% CO2 and 17.5% O2) showing percentage of CO2 (%) Treatment 8 (15% CO2 and 17.5% O2) showing percentage of O2 (%) Treatment 9 (0% CO2 and 20% O2) showing percentage of CO2 (%) Treatment 9 (0% CO2 and 20% O2) showing percentage of O2 (%) Treatment 10 (5% CO2 and 20% O2) showing percentage of CO2 (%) Treatment 10 (5% CO2 and 20% O2) showing percentage of O2 (%) Treatment 11 (10% CO2 and 20% O2) showing percentage of CO2 (%) Treatment 11 (10% CO2 and 20% O2) showing percentage of O2 (%) Treatment 12 (15% CO2 and 20% O2) showing percentage of CO2 (%) Treatment 12 (15% CO2 and 20% O2) showing percentage of O2 (%) Comparison of sensory scores of idli made from treatment 1 (0% CO2 and 15% O2) Comparison of sensory scores of idli made from treatment 6 (5% CO2 and 15% O2) 118 119 119 120 120 121 121 122 122 123 123 124 124 127 128 INTRODUCTION AND REVIEW OF LITERATURE 0 MODIFIED ATMOSPHERE PACKAGING OF READY TO COOK IDLI BATTER 1.1 INTRODUCTION Fermented cereals, pulses and meat are consumed throughout the world both as means of preservation by identifying their texture, aroma and flavour in addition to their health benefits. Idli, one of the most common traditional cereal-pulse based fermented breakfast product is consumed mostly in the southern part of India and Sri Lanka. Idli is the most preferred breakfast product due to its soft texture, mild pleasant flavour and aroma, easy digestibility and known health and nutritional benefits. Even with rapid social transition, idli still remains to be the choice of breakfast for the population either at home or homeaway. With rapid urbanization, idli is one of the most served products in the restaurants and catering establishments. Idli being a lactic acid bacteria fermented product, is traditionally prepared by rice and dhal soaked, ground and fermented before steamed and consumed. With rapid urbanization, Ready to cook, packaged fermented batter is made available in the cities by household vendors, frequently with quality and safety problems. In spite of heavy demand there has not been proper commercialization of the product due to lack of set quality parameters as well as issues with the shelf life of the product. There has also been an effort made to develop starter cultures aimed at preparing the final product with consistent sensory parameters. However these starter cultures did not become popular due to their inferior sensory characteristics. Efforts are being made by various research groups to develop appropriate and acceptable starter cultures for idli. In spite of heavy demands organized food industries have not taken up the commercialization of ready to cook idli batter in view of short shelf life. Even the commercial prospects of scientifically developed starter cultures will remain curtailed till the shelf life of ready to cook batter is considerably extended. In the pursuit of extension of shelf life of ready to cook idli batter, a microbiologically dynamic fermentation medium with other several factors needs to be kept in mind. The product should be close to the natural as any change in the dynamics of the fermentation flora could lead to unacceptable product characteristics. Although only a few fermented 1 products or preserved with Modified Atmosphere Packaging (MAP), most of them are packaged after the fermentation/maturation process. In this work an effort has been made to scientifically optimize the process of preparation of ready to cook idli batter and packaging the ground product in the initial stages of fermentation with optimized gas combinations supporting slow but desirable fermentation process extending the product shelf life without affecting texture and sensory characteristics. 1.2 REVIEW OF LITERATURE The review of literature of the current study is done under the following headings: 1.2.1. Significance of fermented foods 1.2.2. Positive health outcomes of breakfast consumption 1.2.3. Idli and its properties 1.2.4. Rice - A staple food grain in idli making 1.2.5. Black gram - A protein source in idli making 1.2.6. Oligosaccharides in foods 1.2.6. Shelf life of fermented foods 1.2.1. SIGNIFICANCE OF FERMENTED FOODS Fermented foods are those foods which have been subjected to the action of microorganisms or their enzymes to produce desirable biochemical changes and results in significant modification to the food. Fermented foods provide variety to the diet supplying nutrients predominantly proteins and amino acids. The process of fermentation also aids in detoxification (Campbell-Platt, 1994). Fermentation plays diverse roles like enhancing the diet with wide range of flavours, aromas and textures, preserving substantial amounts of food through lactic acid, alcoholic, acetic acid, alkaline fermentations, enriching food substrates with nutrients and also reducing cooking times 2 and fuel requirements (Steinkraus, 1994). Lactic acid fermented foods are common in tropical countries and these foods give improved organoleptic qualities (Cookey et al., 1987). Fermentation affords a natural way to reduce the volume of the material to be transported, abolish undesirable components, enhance the nutritive value and improve appearance of the food, decrease the energy required for cooking and make a safer product (Simango, 1997). Fermented foods are produced worldwide by various manufacturing techniques, raw materials and microorganisms. However, there are only four main fermentation processes which include alcoholic, lactic acid, acetic acid and alkali fermentation (Soni and Sandhu, 1989). Alcohol fermentation results in the production of ethanol, and yeasts are the predominant organisms (e.g. wines and beer), fermented milks and cereals are mainly conceded out by lactic acid bacteria. A second group of bacteria significant in food fermentation is the acetic acid producers (Acetobacter species). Acetobacter sp. converts alcohol to acetic acid in the presence of excess oxygen (McKay and Baldwin, 1990). Likewise, fermentation significantly improves the protein quality as well as the level of amino acid particularly lysine in maize, millet, sorghum, and other cereals (Hamad and Fields, 1979). Fermentation also leads to improvement in the shelf life, texture, taste and aroma of the final product. During cereal fermentation a number of volatile compounds are formed, which contribute to a composite blend of flavours in the products (Chavan and Kadam, 1989). The presence of aromas representative of acetic acid and butyric acid make fermented cereal based products more appetizing. Traditional fermented foods prepared from most common types of cereals (such as rice, wheat, corn or sorghum) are well known in various parts of the world. Some are utilized as colorants, spices, beverages and breakfast or light meal foods, while a few of them are used as key foods in the diet. The microbiology of many of these fermented products is quite complex and not known. In most of these products, fermentation is natural and involves mixed cultures of yeasts, bacteria and fungi. Some microorganisms may participate in parallel, while others act in a sequential manner with exchanging dominant flora during the course of the fermentation. The common bacteria involved in fermentation are species of Leuconostoc, 3 Lactobacillus, Streptococcus, Pediococcus, Micrococcus and Bacillus. The fungi genera Aspergillus, Paecilomyces, Cladosporium, Fusarium and Saccharomyces (yeast) are most often found in certain products (Blandino et al, 2003). 1.2.2. POSITIVE HEALTH OUTCOMES OF BREAKFAST CONSUMPTION Studies done by Agostoni, et al., (2010) disclose that breakfast represents a healthy habit and association with positive health outcomes proves breakfast should be consistent with local and family dietary behaviours. Policies and interventions supportive of breakfast consumption are therefore encouraged. According to neurobehavioral data, the good example of parents and access to a variety of palatable and pleasant breakfast foods should drive children to choose self select breakfast models with balanced composition, while respecting recommended dietary allowances. A balanced macronutrient composition, the proposition of a variety of models leading to a total energy density preferably within lower ranges (< 1 to 1.5), as well as glycemic indices in the lower range for the same food class, could emphasize the positive short and long term health outcomes which is now attributable to breakfast. Regular breakfast consumption can have a multitude of positive health benefits, yet young people are more likely to skip breakfast than any other meal. Given the evidence that dietary behaviours established in childhood and adolescence track into adulthood along with evidence that breakfast skipping increases with age, identifying correlates of children's and adolescent's breakfast behaviour is imperative. Few studies have examined the same specific family correlates of breakfast consumption, limiting the possibilities of drawing strong or consistent conclusions. Parental breakfast eating and living in twoparent families were the correlates supported by the greatest amount of evidence in association with adolescent's breakfast consumption. The results suggest that parents should be encouraged to be positive role models to their children by targeting their own dietary behaviours and that family structure should be considered when designing programmers to promote healthy breakfast behaviours (Pearson et al., 2009). Eating breakfast is important for the health and development of children and adolescents. Reports on the findings of an Australian survey of 699 thirteen year old concerning the 4 extent of skipping breakfasts, indicated approximately 12 percent of the sample skipped breakfast. Gender was the only statistically significant socio demographic variable, with females skipping at over three times the rate of males. Skippers were more likely to be dissatisfied with their body shape and to have been on a diet to lose weight than were those who ate breakfast (Shaw, 1998). Wesnes et al., (2003) reported in their study that a typical breakfast of cereal rich in complex carbohydrates can help maintain mental performance over the morning. Frequency of breakfast and cereal consumption decreased with age. Days eating breakfast were associated with higher calcium and fiber intake in all models, regardless of adjustment variable. After adjusting for energy intake, cereal consumption was related to increased intake of fiber, calcium, iron, folic acid, vitamin C, zinc, and decreased intake of fat and cholesterol. Cereal consumption as part of an overall healthful lifestyle may play a role in maintaining a healthful Body Mass Index (BMI) and adequate nutrient intake among adolescent girls (Barton et al., 2005). 1.2.3. IDLI AND ITS PROPERTIES 1.2.3.1. Nutritional composition of idli Idli, a very popular fermented breakfast food consumed in the Indian subcontinent, consists mainly of rice and black gram. Idli fermentation was carried out in the conventional way in a batter having rice to black gram in the ratios of 2:1, 3:1 and 4:1 at room temperature. It makes an important contribution to the diet as a source of protein, calories and vitamins, especially B-complex vitamins, compared to the raw unfermented ingredients. It can be produced locally and used as a dietary supplement in developing countries to treat people suffering from protein calorie malnutrition and kwashiorkor (Nagaraju and Manohar, 2000). Adding Saccharomyces cerevisiae, along with natural bacterial flora of the ingredients, was the best method for standardizing idli fermentation in terms of improved organoleptic characteristics, leavening and nutritional constituents. Traditional idli fermentation involves several bacteria and yeasts, contributed by the ingredients rice (Oryza sativa), black gram (Phaseolus mungo) and the environment, with overall 5 dominance of the former in bringing about various changes. Idli fermentation is accompanied by an increase in total acids, batter volume, soluble solids, reducing sugars, non protein nitrogen, free amino acids, amylases, proteinases and water soluble vitamins B1, B2 and B12 contents, thus accounting for improved digestibility and nutritional value of the staples. Novel idli batter prepared by replacing conventional black gram with other legumes, revealed significant change but with difference in the levels of some biochemical constituents (Soni and Sandhu, 1989). Idli, Dhokla, Nan, Kulcha, Bread, Jalebi, Bhatura, Bhalla, Dosa, Gulgule and Wadian were prepared in the laboratory using traditional fermentation techniques. The fermented batter of idli and dosa contained higher amount of available lysine, cystine and methionine. After processing, maximum retention of lysine, methionine and cystine was observed in steamed idli (Riat and Sadana, 2009). Growth and nitrogen balance feeding trials were conducted with rats to determine the protein quality of idli, a fermented steamed cake prepared from beans (Phaseolus vulgaris) and rice. Feed Efficiency Ratio (FER), Protein Efficiency Ratio (PER) and Relative PER (RPER) of fermented idli diets were significantly lower (p<0.05) than the FER, PER and PER of unfermented idli diets. The Digestibility Coefficient (DC) and Net Protein Utilization (NPU) of fermented idli diets were significantly lower (p<0.05) than the DC and NPU of unfermented idli diets. Biological Value (BV) of fermented and unfermented idli diets was similar to the BV of a caesin control diet. Fermentation does not improve the protein quality of idli prepared from beans and rice (Joseph and Swanson, 1994). 1.2.3.2. Physico chemical parameters of idli Balasubramanian and Viswanathan (2007a) has shown that idli batter was prepared from soaking polished parboiled rice and decorticated black gram. The blend a ratio of 2:1, 3:1 and 4:1 (v/v) and the batter was allowed for fermentation (0, 6, 12, 18 and 24 h) adding two percent of salt. Other legumes such as soybeans and Great Northern beans could be substituted for black gram in preparation of idli (Reddy et al., 1981). Fermentation time of the batter varies from 14 to 24 h with overnight fermentation being the most frequent 6 time interval. The ingredients for idli are carefully washed, soaked in water separately, grounded, mixed, and finally allowed to ferment overnight. When the batter has been raised sufficiently, it is cooked by steaming and served hot. The product has a very soft and spongy texture and a desirably sour flavour and taste. The black gram was washed several times, first with tap water and finally with distilled water to remove the surface microorganisms. These were found to produce off flavour in the idli unless they were washed out (Mukherjee et al., 1965). Mukherjee et al., (1965), studied the fermentation of idli batter. The microorganisms responsible for the characteristic changes in the batter were isolated and identified. Although there is a sequential change in the bacterial flora, the predominant microorganism responsible for souring, as well as for gas production, was found to be Leuconostoc mesenteroides. In the later stages of fermentation, growth of Streptococcus faecalis and, followed by Pediococcus cerevisiae became significant. The fermentation of idli demonstrates a leavening action caused by the activity of the hetero fermentative lactic acid bacterium, L. mesenteroides. As far as is known, this is the first record of a leavening action produced exclusively by the activity of a lactic acid bacterium. Idli is traditional fermented rice and black gram based breakfast food of South India. Idli batter was prepared from soaking polished parboiled rice and decorticated black gram for 4 h at 30 ± 1oC in water. The soaked mass was ground to 0.5 to 0.7 mm particle size batter using wet grinder with adequate amount of water. The idli batter parameters such as bulk density, pH, total acidity, flow behaviour index and consistency coefficients were studied for different fermentation times and blend ratios. The bulk density, pH and percentage total acidity of batter during different fermentation times and blend ratios ranged between 0.94 and 0.59 g/cm3, 5.9 and 4.1 and 0.443 and 0.910%, respectively. The consistency coefficient at any fermentation time shows increasing trend as the rice to black gram ratio increased. The flow behaviour index indicated strong non-Newtonian fluid behaviour (pseudoplastic) of idli batter at different fermentation times and blend ratios (Balasubramanian and Viswanathan 2007a). The rheology of the idli batter was assessed using a Brookfield viscometer having disc spindles. Power law model with yield stress adequately fitted the data. Yield stress values 7 were in the range of 13-43 Pa and reached a maximum value at 7 h of fermentation. Flow behaviour indices were in the range of 0.287-0.605. Flow behaviour indices at 23 h were significantly lower than those at 0 h. Consistency index values, at any fermentation time, increased as the rice to black gram ratio increased. Mean particle size ranged from 500 to 600 micro meter and there was no definite trend noticed with respect to time of fermentation and rice to black gram ratio. There was a steep change in volume increase after 4-h fermentation (Nagaraju and Manohar, 2000). The idli batter comprises lactic acid bacteria and yeasts and causes an improvement in the nutritional, textural and flavour characteristics of the final product. The desirable flavour compounds such as ketones, diols and acids were found to be present up to eight days of storage, whereas undesirable flavours like sulphurous and oxazolidone compounds, ethanone and thiazole appeared in the batter subsequent to six days of storage. The sensory attributes of idli (final product) prepared from the stored batter related well to the determined flavour profile (Agrawal et al., 2000). The work done by Nisha et al., (2005) stabilized the idli batter at room temperature (2830°C) and refrigerated storage (4-8°C) by using various hydrocolloids and some surfaceactive agents. The batter was evaluated in terms of decrease in volume, and whey separation. While hydrocolloids gave good stabilization, surface-active agents failed to stabilize the batter and they reduced whey separation. Among the various hydrocolloids, 0.1percent guar gave best batter stabilization, and idli made after ten days of room temperature and 30 days of refrigerated storage of batter were found to be of acceptable quality. Reduction in the fermentation time of the idli batter is of great commercial significance for large-scale idli production and can be potentially achieved by addition of enzymes. The study done by Iyer and Anathanarayan, (2008) was undertaken to explore the possibility of expediting the idli batter fermentation process by adding an exogenous source of α-amylase enzyme. 5, 15 and 25 U per 100 g batter of amylase added to the idli batter was allowed to ferment. Different parameters were monitored and sensory attributes were also studied and compared with that of the control set. The fermentation 8 time was reduced from a conventional 14 h to 8 h and the sensory attributes of the final product were also successfully maintained. Texture Profile Analysis (TPA) test was performed for idli, making cylinder samples (13.5 mm diameter, 10 mm long) of idli. In Pearson correlation matrix, majority of the parameters were positively correlated at p<0.01 and p<0.05. The firmness value positively correlated with gumminess and chewiness, which depicts the soft nature of idli (Balasubramanian and Viswanathan 2007b). 1.2.4. RICE - A STAPLE FOOD GRAIN IN IDLI MAKING Cereal grains particularly rice, form a major source of dietary nutrients for all people, particularly those in the developing countries. However, compared with animal foods, nutritional quality of cereal grains is inferior due to lower protein content, deficiency of certain essential amino acids, lower protein and starch availabilities, and the presence of some antinutritional factors. Fermentation of cereals for a limited period of time improves amino acid composition and vitamin content, increases protein and starch availabilities, and lowers the levels of antinutrients. The traditional foods prepared by fermentation of cereals in different parts of the world are briefly described and future research needs to improve their nutritional contribution are addressed (Chavan et al., 1989). Cereals are deficient in lysine, but are rich in cysteine and methionine. Legumes, on the other hand, are rich in lysine but deficient in sulphur containing amino acids. Thus, by combining cereal with legumes, the overall protein quality is improved (Camphell-Platt, 1994). Fermented foods prepared from cereals and legumes are an important part of the human diet in Southeast Asia and parts of East Africa. The popularity of legume based fermented foods is due to desirable changes including texture and organoleptic characteristics. Improvement in digestibility and enhancement of keeping quality, partial or complete elimination of anti-nutritional factors or natural toxins, increased nutritive value, and reduced cooking time (Joseph, 1994). Cereal grains constitute a major source of dietary nutrients all over the world. Although cereals are deficient in some basic components, fermentation may be the most simple and 9 economical way of improving their nutritional value, sensory properties, and functional qualities. Products produced from different cereal substrates (sometimes mixed with other pulses) fermented by lactic acid bacteria, yeast and/or fungi (Blandino et al., 2003). Rice colour changes from white to amber during parboiling (soaking and steaming). Colour parameters indicated that, during soaking, yellow bran pigments leaches out in the water. The levels of the Maillard precursors (i.e., reducing sugars and free α-amino nitrogen (FAN)) depends on soaking temperature and time: leaching of RS was compensated by enzymatic formation for long soaking times (>60 min), while proteolytic activity was too low to compensate for FAN leaching. Parboiled rice soaking under nitrogen, oxygen, or ambient conditions and determination of polyphenol oxidase activity allowed to conclude that the effect of enzymatic colour changes on the soaked rice colour was rather small. Colour measurements of brown and milled mildly, intermediately, and severely parboiled rice samples showed that both brown and milled rice samples were darker and more red and yellow after parboiling and that the effect depended on the severity of parboiling conditions. Furthermore, steaming affected the rice colour more and in a way opposite to that observed in soaking (Lamberts et al., 2006). Parboiled brown rice contained considerably more Reducing Sugars (RS) but less sucrose and Free Amino Acids (FAA) than raw brown rice. On milling, there was considerable loss of sucrose and FAA from raw rice, but very little from parboiled rice; reducing sugars changed little in either. Processing conditions affected the contents of sugars and FAA. Maximum increase in RS and decrease in sucrose content occurred after soaking at 60C. Controlled incubation of rice flour, intact grain, separated germ and deemed rice in water showed that considerable changes in sugars and FAA occurred in all cases, the magnitude depending on the circumstances, but a greater part of the sugars leached out into the water during soaking (Ali and Bhattacharya, 1980). Grinding characteristics of raw and parboiled rice were evaluated in various wet grinding systems like, mixer grinder, stone grinder and colloid mill. The duration of grinding had inverse effect on the particle size and direct impact on the starch damage as well as energy consumption in batch grinders. Stone grinder was the least energy efficient and specific energy consumption for grinding raw rice (160.6 kJ/kg) was nearly twice as that 10 of mixer grinder (74.9 kJ/kg). Parboiled rice required longer duration of grinding compared to raw rice, consequently specific energy consumption was higher (∼220 kJ/kg) (Sharma et al., 2008). Wet grinding is a critical step in the preparation of batter based traditional food products. It involves both physical and chemical changes while dry grinding is a mere size reduction operation. In wet grinding of cereals, the protein matrix holding the starch granules is destroyed, releasing the starch granules from the protein network (Kent and Evers, 1994). A colloid mil was comparatively evaluated with domestic wet grinding systems, namely, a mixer grinder and a stone grinder for grinding of raw rice, parboiled rice and black gram. The wet ground samples were finer in particle size compared with dry ground samples. The starch damage was the least in black gram followed by raw rice and parboiled rice in dry grinding. In wet grinding, the starch damage in black gram as well as raw rice remained more or less same whereas the parboiled rice showed greater damage. Parboiled rice required 2.5 to 3 times more energy (216-252 kJ/kg) as that of raw rice (72-108 kJ/kg) for grinding in the mixer grinder and the stone grinder (Solanki et al., 2005) The nutritional quality of wild rice tends to be comparable with other cereals characterized by a high content of starch and protein and a low fat content. As a whole grain, wild rice is also a good source of dietary fibre (Qiu et al., 2010).The presence of Streptococcus faecalis in the fermented batter, the presence of pharmacological active amines such as thiamine was expected but they were not detected (VanVeen, et al., 2008). Parboiled brown rice contained considerably more (RS) but less sucrose and FAA than raw brown rice. On milling, there was considerably loss of sucrose and FAA from raw rice, but very little from parboiled rice; reducing sugars and FAA. Maximum increase in RS and decrease in sucrose content occurred after soaking at 600 C (Ali et al., 2007). Larsen et al., (2000) opines that rice is an important crop, forming a staple food for many of the world‘s population. A study showed there was an effect of severely pressure parboiled rice reduced the glycaemic index. Brown rice malt from Indica and Japonica type rice were prepared and their nutrient composition as well as Non-Starch Polysaccharide (NSP) contents and also some of the 11 physicochemical characteristics were determined. The activity of α- and β-amylases in the un-germinated (native) rice was negligible but increased considerably on germination. Malting altered the chemical composition of both Indica and Japonica rice to a small extent but caused noticeable changes in the pasting characteristics. Controlled germination or malting causes considerable changes in the physicochemical and biochemical properties of both Indica and Japonica rice (Mohan et al., 2010). Whole grain rice is rich in phenolic compounds. The effect of γ-irradiation on the main phenolic compounds in the rice grains of three genotypes (black, red, and white) was investigated. Three phenolic acids (p-coumaric acid, ferulic acid, and sinapinic acid) were identified as major phenolic compounds in all rice samples, while two anthocyanins (cyanidin-3glucoside and peonidin-3-glucoside) were identified in pigmented grain samples (Zhu et al., 2010). 1.2.5. BLACK GRAM A PROTEIN SOURCE IN IDLI MAKING Blackgram (Phaseolus mungo) is a pulse traditionally used in the preparation of South Indian breakfast foods, such as idli, which is relished for its soft and spongy texture (Susheelamma and Rao, 1979a). The components responsible for these properties are the surface active proteins that generate a foam and as a result impart a porous structure to the food, and the viscogenic mucilaginous polysaccharide (~6%) that stabilizes the porous structure against thermal disruption during steaming. The overall carbohydrate composition (Bhat and Tharanathan, 1986) and the structure function characteristics of the total polysaccharides of black gram have been reported. During fermentation of black gram, for the preparation of leavened foods, it was found that the mucilaginous polysaccharide undergoes compositional and rheological changes (Muralikrishna et al.,1987). Here, the fermentation is due to the activities of endogenous microflora (endophytes) in black gram, in particular Leuconostoc mesenteroides, yeasts, lactic acid bacteria and coliforms. More than one oligosaccharide was observed as in green gram (stachyose, maltohexaose), sorghum (stachyose, maltotriose), barley (stachyose, raffinose), wheat (stachyose, raffinose) and black gram (stachyose, raffinose). In ragi, bajra and rice malt oligosaccharides were absent. Germination of seeds for 48 h resulted in complete loss of 12 stachyose and raffinose in cereals and pulses. The maltotriose content in pulses completely disappeared on germination but among cereals, 45.1 and 57.3 percent loss was observed in sorghum and maize, respectively (Sampath et al., 2008). In black gram after fermentation, apparent viscosity of cold paste increased. Some of the properties such as intrinsic viscosity, swelling and solubility after fermentation were reported by them. Fermentation and steaming approximately 40 per cent reduction in oligosaccharides resulting in reduced flatulence in the body (Koh and Singh, 2009). Nutritional benefits are produced in legume fermentations, when microorganisms break down the flatulence causing indigestible oligosaccharides, such as stachyose and verbascose are broken down into the absorbable monosaccharaides and disaccharides. Biosynthesis of B vitamins in food fermentations has been recognized to be of major nutritional significance, particularly in Africa where high-carbohydrate diets, particularly maize diets can be deficient in essential B vitamins, the significance of B vitamin synthesis during fermentation to maize and sorghum beers in southern Africa was recognized by the use of the term ‗biological ennoblement‘ by Platt (1964). 1.2.6. OLIGOSACCHARIDES IN FOODS Carbohydrates are classified into monosaccharide, disaccharides, oligosaccharides and polysaccharides. Oligosaccharides are low molecular weight carbohydrates consisting 3 to 10 sugar monomers (Voragen, 1998). Oligosaccharides withstand salivary hydrolysis and digestive enzymes of human animal intestine so these oligosaccharides are not absorbed in the upper digestive tract and are able to reach the colon unaltered. In colon, oligosaccharides interact with the microflora and affecting immunomodulation (Reiffova and Nemcova, 2006). The non-digestible oligosaccharides promote the growth of beneficial bacteria in the colon, chiefly the Bifidobacteria sp., and are thus recognized as prebiotics (Mussatto and Mancilha, 2006). Most of the known prebiotics and prebiotic candidates are nondigestible oligosaccharides, obtained by extraction from plants (e.g., chicory inulin), followed by enzymatic hydrolysis (e.g., oligofructose from inulin) and by synthesis (by trans-glycosylation reactions) from mono and/or disaccharides such as sucrose (fructooligosaccharides) and lactose (trans-galactosylated oligosaccharides/galactooligosaccharides) (Crittenden and Playne, 1996). Among the 13 prebiotics, inulin and oligosaccharides are the most studied and have been recognized as dietary fibre worldwide (Moshfegh et al., 1999). 1.2.6.1Conversion of polysaccharides into oligosaccharides Polysaccharides are the major source of bioactive oligosaccharides and around twenty different types of non-digestible oligosaccharides (NDOs) are described for prebiotic activities. Fructooligosaccharides, galacto-oligosaccharides, xylo-oligosaccharides and galacturono-oligosaccharides are predominant NDOs with prebiotic characteristics. Oligosaccharides are recognized as non-cariogenic, non-digestible and low-calorie molecules and can be used as natural food preservatives. Chitosan and oligochitosans, inhibits growth of pathogens and extends shelf-life of food products (Barreteau et al., 2006). Glycosyl-hydrolases and polysaccharide lyases, are used to obtain oligosaccharide from polysaccharides. Polysaccharide chains were broken down by polysaccharide lyases and form a double bond at new reducing end (Michaud et al., 2003). Polysaccharide hydrolase cleave glycosidic bond by transfer of a H2O molecule and act as exohydrolases/endohydrolases (Boels et al., 2001; Bojarova and Kren, 2007). Enzyme used for the purpose must be specific to the substrate should be more efficient. The solution for having enzyme specificity is to use bacteria to produce particular enzyme. From the bacteria the specific enzyme of the polysaccharide can be isolated, purified and concentrated and used (Sutherland, 1999). 1.2.6.2. Physiological properties of oligosaccharides Oligosaccharides possess important physicochemical properties. They are used as food ingredients as their physiological properties were beneficial to human health. The NDOs can be used as low caricinogenic sugar surrogates in products like confectionery, chewing gums, yoghurts and drinks (Crittenden and Playne, 1996). Many NDOs are not digested by humans because the human body doesn‘t have the enzymes necessary to hydrolyze certain units of monosaccharides. Compounds include carbohydrates where fructose, galactose, glucose and/or xylose are the monosaccharides units. This property of NDOs makes the suitable for use in sweet, low-caloric diet foods, and for consumption 14 by individuals with diabetes (Crittenden and Playne, 1996; Rivero-Urgell and Santamaria-Orleans, 2001). Most oligosaccharides were quantitatively hydrolyzed in the upper part of the GIT. The resulting monosaccharides are transported through the portal blood to the liver and subsequently, to the circulatory system. These carbohydrates are important for health as they serve as both substrates and regulators for major metabolic pathways. Nevertheless, some oligosaccharides present specific physicochemical properties resist to the digestive process, reaching the caeco colon. In the caeco-colon, most of the nondigestible oligosaccharides were hydrolyzed to small oligomers and monomers and further metabolized by most of the anaerobic bacteria. Such a metabolic process, known as fermentation, not only serves the bacteria by providing energy for proliferation, but it also produces gases (H2, CO2, CH4), which are metabolically useless to the host, and small organic acids (Short-Chain Fatty Acids – SCFA) such as acetate, propionate, butyrate and L-lactate. Even though they do not provide the body with monosaccharaides, the non-digestible oligosaccharides are indirect energy substrates and metabolic regulators (Delzenne and Roberfroid, 1994). The amounts and types of SCFA produced in the colon depend on the type of NDO substrate as well as on the composition of the intestinal flora (Sako et al., 1999). Oligosaccharides serve as substrate for growth and proliferation of anaerobic bacteria, mainly the Bifidobacteria, which inhibit the growth of putrefactive and pathogenic bacteria present in the caeco-colon (Sangeetha et al., 2005). NDOs leads to decrease of pH in the colon and consequently, in faeces, resulting from the production of SCFA. Lower pH values inhibit the growth of certain pathogenic bacteria species while stimulating the growth of the bifidobacteria and other lactic acid species (Manning and Gibson, 2004). An increase in faecal dry weight excretion, which is related to the increased number of bacteria resulting from the extensive fermentation of NDOs (Bielecka et al., 2002) The indigestible quality of NDOs means that they have effects similar to dietary fibre, and thus prevent constipation. The end products of NDOs fermentation by colonic 15 bacteria, the SCFA, are efficiently absorbed and utilized by the human colonic epithelial cells, stimulating their growth as well as the salt and water absorption, increasing thus the humidity of the fecal bolus through osmotic pressure, and consequently improving the intestinal motility (Rivero-Urgell and Santamaria-Orleans , 2001). NDOs help in inhibition of diarrhea, especially when it is associated with intestinal infections (Roberfroid and Slavin, 2000). An increase in absorption of different minerals, such as iron, calcium, and magnesium take place due to the binding/sequestering capacity of the NDOs. The minerals that are bound/sequestered and, consequently, are not absorbed in the small intestine reach the colon, where they are released from the carbohydrate matrix and absorbed. The increase on calcium absorption, in particular, reduces the risk of osteoporosis since this mineral promotes an increase in the bone density and bone mass. The hypotheses most frequently proposed to explain this enhancing effect of NDOs on mineral absorption are the osmotic effect, acidification of the colonic content due to fermentation and production of SCFA, formation of calcium and magnesium salts of these acids, hypertrophy of the colon wall (Younes, 1996). Beneficial effect on the carbohydrates and lipids metabolism is that oligosaccharides lead to a decrease in the cholesterol, triglycerides and phospholipids concentration in the blood, reducing thus the risk of diabetes and obesity. Changes in the concentration of serum cholesterol have been related with changes in the intestinal microflora. Some strains of Lactobacillus acidophilus assimilate the cholesterol present in the medium, while others appear to inhibit the absorption of cholesterol through the intestinal wall. On the other hand, the changes in lipid metabolism were suggested to be a consequence of a metabolic adaptation of the liver that might be induced by SCFA (Daubiol et al., 2000). NDOs aid in reduction of cancer risk, mainly the gut cancer. This anti-carcinogenic effect appears to be related to an increase in cellular immunity, the components of the cell wall and the extra-cellular components of bifidobacteria. Faecal physiological parameters such as pH, ammonia, p-cresol, and indole are considered to be risk factors not only for colon cancer development but also for systemic disorders. It has been demonstrated in a human study that the intake of trans-galactosylated disaccharides reduces the faecal pH as well 16 as ammonia, p-cresol and indole concentrations with an increase in bifidobacteria and lactobacilli and a decrease in bacteroid populations. These alterations may be considered to be beneficial in reducing the risk of cancer development. A low colonic pH may also aid in the excretion of carcinogens (Delzenne and Roberfroid ,1994) 1.2.6.3 Animal studies on Oligosaccharides Feeding mice with diets supplemented with inulin and oligofructose increased activities of natural killer cells and phagocytes and enhanced T-lymphocyte functions compared to mice fed diets with cellulose or lacking fibre. These results are consistent with the observations of heightened resistance to systemic infections with Listeria spp. and Salmonella spp., the lower incidence and growth of tumours after exposure to carcinogens and transplanted tumour cells and are in agreement with enhanced innate and acquired immune functions provided by Lactobacillus and other LAB. Supplementing diets with FOS should increase production of SCFA, and particularly butyrate, and can be predicted to strengthen mucosal defences and enhance response to health challenges (Buddington et al., 2002). Colonic fermentation of FOS results in the synthesis of short chain fatty acids, which influences the lipid metabolism in human beings. Feeding male Wistar rats on a carbohydrate rich diet containing 10 percent FOS significantly lowers serum triacylglycerol (TAG) and phospholipid concentration (Delzenne et al., 2002). FOS reduces post-prandial triglyceridemia by 50% and avoids the increase in serum free cholesterol level occurring in rats fed with a Western type high fat diet. FOS protects rats against steatosis (liver TAG accumulation) induced by fructose, or occurring in obese Zucker fa/fa rats. FOS given at the dose of 10 percent in the diet of male Wistar rats for 30 days reduces postprandial insulinemia by 26 percent (Daubiol et al., 2000). Animal studies provide strong evidence that FOS inhibit secretion of TAG rich Very Low Density Lipoprotein (VLDL) particles via inhibition of de novo fatty acid synthesis. High levels of fat present inmost human diets mean that rates of hepatic de novo fatty acid synthesis are extremely low, since exogenous dietary fatty acids provide all the required substrate for hepatic triacylglycerol synthesis (Parks, 2002). 17 Dietary treatment with inulin/oligofructose (15 percent) incorporated in the basal diets for experimental animals resulted in (a) reduction of the incidence of mammary tumours induced in Sprague Dawley rats by methyl-nitrosourea (b) inhibited the growth of transplantable malignant tumours in mice and (c) decreased the incidence of lung metastases of a malignant tumour implanted intramuscularly in mice. It is reported that the dietary treatment with FOS/inulin significantly potentiated the effects of subtherapeutic doses of six different cytotoxic drugs commonly utilized in human cancer treatment (Taper and Roberfroid, 2002). Roberfroid and Slavin, (2000) has reported that feeding rats with FOS (10 percent) for a few weeks decreased uremia in both normal and nephrectomized rats. Dietary FOS enhanced faecal nitrogen excretion and reduced renal excretion of nitrogen in rats. This occurs because these fermentable carbohydrates serve as energy source for the intestinal bacteria, which during growth also require a source of nitrogen for protein synthesis. 1.2.6.4 Applications of FOS in food formulations Inulin and oligofructose are ingredients that deliver a number of important nutritional benefits as well as contribute functional properties that enhance shelf life and taste profile of various food products like nutrition bars (Izzo and Niness, 2001). FOS can be used as the sole sweetening agent and gives 34 percent calorie reduction compared with sucrose standard. Organoleptic characteristics of the products are claimed to be very similar, with the test sample having a lower sweetness and a softer texture. FOS can be used with inulin to replace all the sugar and reduce the fat content and give excellent mouth feel characteristics. Since the freezing point depression is less with oligo-fructose than with sugar, the texture can be harder. Hard candies, gums, and marshmallows can be made while achieving significantly reduced energy values (Murphy, 2001). 1.2.7 MODIFIED ATMOSPHERE PACKAGING The common perception that modified atmospheres are useful for improving storability has significant historical precedent. The written history of the use of modified atmospheres can actually be traced back at least 2000 years to the use of underground, sealed silos (Owen, 1800) where atmosphere modification was detected as ―foul air‖ that 18 was dangerous to enter and was likely a result of O2 depletion and CO2 accumulation due to the respiratory activity of the grain. The modified atmosphere was unintentional, although probably beneficial. The foul air in the storages would presumably control rodent and insect pests, thereby acting to preserve the quality and storage life of the grain. The potential for a positive impact from alteration in the respiratory gases O2 and CO2 became increasingly known through the early research of Berard (1821), Mangin (1896), Kidd and West (1914, 1927, 1945), and Blackman and Parija (1928). Gas modification technologies can be segregated into two classes based on the manner in which the atmospheres are generated and maintained. One class of technologies is referred to as Controlled Atmosphere (CA) storage, in which the atmosphere is either manually or mechanically controlled to achieve target headspace gas concentrations. In CA storages, O2 and CO2 concentrations can be modulated independently from one another. The second class of technologies is (MAP), in which a package possessing a film or foil barrier passively limits gas exchange by the living produce enclosed in the package, thereby altering the headspace atmosphere. In MAP, both oxygen and carbon dioxide are modified simultaneously and their concentrations at steady state are a function of one another. In MAP, the primary route of gas exchange may be through gaspermeable film, perforations in film, or both. In what is referred to as active or intelligent packaging techniques, packages may be flushed with specific gas mixtures designed to obtain a desired initial atmospheric composition, gases may be actively released or scavenged in the package, a partial vacuum can be imposed, biologically active materials can be incorporated in the package, sensors may be used to respond to the product or package conditions, and so on. The aim of MAP (passive, active, or intelligent in design) is to take advantage of physiological responses of the enclosed plant material or plant or human pathogens to the respiratory gases O2 and CO2. Presumably, MAP use is intended to maintain product quality, thereby ensuring appropriate value to the consumer and adequate cash flow back through the marketing and handling chain such that the production and marketing system is sustainable .Knowledge of the physiological responses to atmosphere modification is beneficial in terms of anticipating improved quality retention as a result of technology investment (Beaudry, 2008). 19 Atmosphere modification in a package requires a barrier through which gas exchange is restricted. Enclosing an actively respiring product within a package composed all or in part of a film that acts as a gas barrier reduces O2 and increases CO2, creating gradients across the film barrier. These gradients provide the driving force for gas flux into or out of the package. In passive MAP, a package always tends toward steady state, in which O2 and CO2 levels are constant and O2 uptake and CO2 production by the product are equal to those permeating through the package, a situation that exists only when the respiratory rate is constant or nearly so. The steady-state levels of O2 and CO2 within a package are dependent on the interaction of respiration of the product and the permeability properties of packaging film or micro- perforations (Beaudry et al., 1992; Cameron et al., 1989; Jurin and Karel, 1963; Kader et al., 1989). MAP should be carefully designed, as a system incorrectly designed may be ineffective or even shorten the shelf life of the product. The design should take into consideration not only steady-state conditions, but also the dynamic process, because if the product is exposed for a long time to unsuitable gas composition before reaching the adequate atmosphere, the package may have no benefit. The design of Modified atmosphere package depends on a number of variables, the characteristics of the product, its mass, the recommend atmosphere composition, the permeability of the packaging materials to gases and its dependence on temperature and the respiration rate of the product as affected by different gas composition and temperature. Since, respiration rate modelling is vital to the design of MAP for fresh fruits and vegetables (Fonseca et al., 2002). Temperature is exceptionally important in package design, continuous and perforated films differ in their response to temperature changes. The O2 and CO2 permeability of continuous films increases with temperature, while the diffusion of gases through perforations is extremely insensitive to temperature changes. O2 permeation over LDPE increases 200% in 0 to 15°C, an exchange of O2 over perforations increases only 11% at the same temperature range. Depends on the rate of respiration and transmission, the atmosphere modification can be achieved quickly or relatively slow. At lower temperatures, atmosphere modification will take several days, that some package systems cannot achieve steady-state environments prior to the end of their shelf-life. In many 20 cases, purging the package atmosphere with CO2, N2 or a combination of gases is often desirable during filling and sealing to rapidly obtain the maximum benefits of MAP. Product temperature affects storability more than any other factor. Pre-cooling and temperature maintenance during handling and shipping were critical in preserving quality. Temperature also significantly affects permeability of film and thereby the O2 and CO2 content of the package. The elevated rate of respiration at high temperature could be used to rapidly establish the desired package atmosphere, but this would only be useful in the few circumstances in which it would be more important to rapidly establish the atmosphere than to slow physiological processes, eg., to reduce cut-surface browning. Negative Responses in MAP show that respiration gets reduced as O2 becomes limiting, but there is usually a limit to which O2 can be reduced. The lower O2 limit is frequently considered to be the level of O2 that induces fermentation. This fermentation threshold is not always the lower O2 limit in commercial practice, however, because lower O2 levels may confer benefits that outweigh the loss in flavour or other quality parameters. Ethanol, acetaldehyde, ethyl acetate and lactate are products of fermentation that can contribute to the development of off-flavours as well as physical injury (Kays, 1997; Mattheis and Fellman, 2000). With regard to MAP study done by Fandos et.al (2000) on Cameros cheese, it was found that packaging in 50%CO2/50%N2 and 40%CO2/60%N2 were the most effective conditions for extending the shelf life of cheese with good sensory characteristics. MAP studies have been done in iced fresh hake slices (Pastoriza et al, 1996), Fresh cut mangoes (Aguliar et al, 2000), fresh-cut ice berg lettuce (Fan et al, 2003), refrigerated sea bass slices (Masniyom et al, 2002), blueberry (Song et al, 2002), minimally processed mango and pineapple fruits (Martínez-Ferrer et al, 2002), pomegranate (Artes et al, 2000). From the review of literature and state of the art it is clear that there are no studies on optimization of idli with respect to components and fermentation time taking into consideration, both instrument based texture analysis and 15mm rating scale and combination of RSM and PCA to evolve the optimized parameters for idli. In addition there are no studies which deal with improvement of shelf life of idli batter with modified 21 atmosphere packaging. In continuation with this, the present study has been planned with an objective to improve the shelf life of ready to cook idli batter using modified atmosphere packaging. The set objective is achieved through following the three major sub-objectives: 1. To understand the presently followed practices for the preparation of idli. 2. To optimize the process of preparation of the product with respect to ingredient ratios and fermentation time. 3. To improve the shelf-life of ready to cook idli batter by optimized process. 22 TRANSITION IN THE PREPARATION AND CONSUMPTION OF IDLI AMONG THE POPULATION OF PUDUCHERRY 2.1 INTRODUCTION Idli occupies a special place in the diets of Indians and is one of the predominant choices of food in the daily diet particularly as a breakfast food. Idli being a fermented food possesses a great significance as it provides aroma and soft texture and also act as a nutritious food. The current study was undertaken among the selected population to study the preference of idli, consumption pattern of breakfast, preparation of idli at household level and preference of commercial idli batter against homemade batter. 2.2 MATERIALS AND METHODS 2.2.1 Selection of area The area selected for the study was Union Territory of Puducherry which has a population of 12.44 lakhs (Puducherry population census, 2011) representing multilingual and multi-cultural population. Eight areas namely Kalapet, Muthaiyalpet, Villiyanur, Gorimed, Thattanchavadi, Lawspet, Vandrampet and Uppalam were selected for the study .Plate 2.1 shows selected areas in the city map of Puducherry. Plate 2.1 City map of Puducherry showing selected areas for the study 23 2.2.2 Selection of tool for data collection An oral interview schedule was formulated to collect information regarding the consumption of idli and preference of commercial idli batter. Interview schedule (Annexure - I) included questions regarding socio-economic status, preference for idli, preparation of idli at house hold level, consumption pattern of idli and commercial idli batter. This formulated tool was tested on a pilot population, based upon the suggestions the corrections were implemented and the questionnaire was used to collect information from the respondents. 2.2.3 Selection of respondents In total 300 respondents were randomly surveyed for the study irrespective of the cultural and linguistic background 2.2.4 Data Analysis The collected data were statistically treated for distributional analysis using SPSS Statistical software (18.0). 2.3 RESULTS AND DISCUSSION The results and discussion of the present study is discussed under the following 2.3.1 Socio-economic profile of the selected respondents 2.3.2 Consumption pattern of breakfast among the selected respondents 2.3.3 Methods of preparation of idli at household level 2.3.4 Preference for commercial idli batter against home-made batter 2.3.1 Socio-economic profile of the selected respondents The details on the age, sex, educational qualification, employment and economic status of the respondents are discussed. Table 2.1 shows the age (Fig. 2.1.a) and sex-wise distribution of the selected respondents. Table 2.1 showed that, 62 per cent of the selected respondents were in the age group of 21 to 40and only 2.66 per cent were in the age group of 61 to70 years. Among 300 respondents, four per cent people were male and 96 per cent people were female. From this data, it is clearly noted that the study is correlated with female respondents who are the majority home makers. Women are the integral part of family (Jan and Akhtar, 2008) and vital force in the decision making concerning child 24 growth, money management, health and nutrition, and socio-economic progress of the family. Table 2.1 Age and sex wise distribution of the selected respondents (N=300) Particulars Percentage (%) Age (in years) 21-30 31.66 31-40 31.66 41-50 26.33 51-60 07.66 61-70 02.66 Male 03.66 Female 96.33 Sex From Table 2.2 and Fig. 2.1.b it was noted that 82 per cent of the selected population were literates and 18 per cent were illiterates. According to HUDCO (2004), the total family income for low income group ranged from Rs. 2500 to Rs. 4500, for middle income Rs. 4501 to Rs.7500 and for high income Rs. 7501 and above. In the present study majority (50%) of the population fell under high income group, 22 per cent belonged to middle income group and 18.3 per cent fell below low income group (Fig.2.1.c). Hence the study covered the respondents from almost all income groups with regard to occupational status. Of the population majority (71%) was housewives and 22 per cent were labourer earning daily wages and six per cent were only self-employed (Fig.2.1.d) 25 a. Age wise (in years) distribution of the respondents c. Monthly income of the respondents b. Educational qualification of the respondents 26 d. Occupational status of the respondents Fig. 2.1 Socio-economic profile of the selected respondents Table 2.2 Educational and economic status of the selected respondents (N=300) Particulars Percentage (%) Educational qualification Illiterate 18.00 Primary school 12.00 Higher secondary 33.00 High school 16.33 Graduate 20.66 Monthly income (Rs.) <2500 18.3 2501-4500 09.7 4501-7500 22.0 >7501 50.0 Occupational status Housewife 71.33 Labourer 22.33 Self-employment 06.33 2.3.2 Consumption pattern of breakfast among the selected respondents Details about breakfast items preferred and consumed by the selected population are shown in Table 2.3. It was found that 85 per cent consumed breakfast regularly and fifteen per cent of the selected population has shown to skip breakfast. In the study done by Agostoni et al., (2010) it was reported that nearly 10-30 per cent of the breakfast skippers are found throughout the world which is in par with the current study. 27 Table 2.3 Details on breakfast consumption (N=300) Particulars Percentage (%) Breakfast intake Eating breakfast Skipping breakfast 84.66 15.33 Items preferred Idli 69.30 Dosa 21.00 Oats 09.70 Items consumed Idli 25.00 Dosa 20.00 Chapathi 18.00 Noodles 13.85 Poori 12.15 Oats porridge 05.40 Bread 05.60 Studies done by Reddy et.al (1981) and Balasubramanian and Viswanathan (2007a) showed that idli is a breakfast food and the current study reassures the same with 69 and 21 per cent of the selected population preferring to take idli and dosa respectively as breakfast item daily . Further it was noted that only 25 per cent consume idli daily and 20 per cent consume dosa daily. The interesting fact is that 99 per cent of the respondents have a liking for idli as breakfast. 28 2.3.3 Preparation of idli at household level In spite of 71 per cent being housewives among the selected population, Table 2.4 revealed that only seven per cent of the population grind idli batter at home daily whereas majority (58.33%) grinds only once in a week (Fig.2.2.a). Most of the respondents reveal it due to the reason that grinding idli batter is laborious. For idli making majority of the population (68%) prefer parboiled rice (Fig.2.2.b). Table 2.4 Idli preparation at household level (N=300) Particulars Percentage (%) Frequency of grinding idli batter Daily 07.00 Once in a week 58.33 Twice in a week 28.66 Once in a month 06.00 Variety of rice Parboiled rice 68.03 Ration rice (provided at PDS) 19.30 Mixed rice 12.66 Type of black gram dhal Husk removed (decorticated) 34 Husk removed after soaking 49 Both 17 Ratio of rice: black gram dhal 3:1 99.70 8:1 00.30 29 Only 34 per cent used decorticated black gram whereas 49 per cent used black gram with husk after soaking (Fig.2.2.c). Ratio of the raw ingredients in idli making is an important criterion for the texture of the idli. In the current study majority (99.7%) of the respondents used 3:1 ratio of rice and black gram dhal respectively. The findings regarding the preparation of idli at house hold level is supported by the study done by Balasubramanian and Viswanathan (2007b) who reported that idli batter was prepared from soaking polished parboiled rice and decorticated black gram for 4 hour at 30±1C in water and the soaked ingredients were ground to 0.5-0.7-mm particle size batter using wet grinder with adequate amount of water. The blend ratios of 2:1, 3:1, 4:1(v/v) batter were allowed for fermentation adding two percent of salt. From the survey it was also found that 100 per cent of the respondents added fenugreek as an additional ingredient in idli making. Generally after grinding idli batter, the batter is left for fermentation. Fermentation time varied between 5 h to 12 h at the selected households (Table 2.5 and Fig.2.3.a). Majority (71.3%) of them fermented the idli batter for 11 to 12 h. The texture of idli is influenced by many variables like raw material, quantity, soaking time, grinding conditions, fermentation time and temperature are adjuncts on quality of idli (Desikachar et al., (1960) and Radhakrishnamurthy et al., (1961). Fermentation of idli batter is an essential step because as reported by Mukherjee et al., (1965), Rajalakshmi and Vanaja (1967) the microorganisms present in black gram dhal helps in acidification and leavening of the batter by which gas, acid and several volatile compounds are formed during fermentation which contribute to a complex blend of flavours in the products (Chavan and Kadam, 1989). The household measures to control fermentation of idli batter to extend the shelflife are shown in Table 2.5 and Fig.2.2.b. Majority (73%) stored the idli batter in refrigerated condition, 19 per cent store idli batter over water tub, and rest of the selected population place betel leaves (4.33%), lady‘s finger (1.66%) and coconut slices (1%) over the idli batter respectively. The scientific reasons for these measures expect for refrigeration is still in dark. 30 a. Frequency of grinding idli batter at house hold level b. Variety of rice used by respondents for making idli c. Type of black gram dhal used for idli making Fig. 2.2 Details on idli preparation at household level 31 a. Fermentation time of batter at household level b. Measures to control fermentation at household level Fig.2.3 Details on fermentation time and measures to fermentation 32 Table 2.5 Fermentation time and measures followed to control fermentation of idli batter at households (N=300) Particulars Percentage (%) Fermentation time (h) 05-06 04.30 07-08 00.70 09-10 23.70 11-12 71.30 Measures to control fermentation Refrigerator 73.33 Placing over water 19.33 Betel leaves 04.33 Lady‘s finger 01.66 Coconut slices 01.00 Plantain leaves 00.33 2.3.4 Preference for commercial idli batter against home-made batter From Table 2.4 it was discussed that only seven per cent of the selected population ground idli batter daily, but Table 2.3 show that 45 per cent of the selected population consume idli and dosa every day for breakfast. This is because 49 per cent of the respondents have shown interest to purchase ready to cook idli batter every day (Table 2.6). The cost variation of the commercial idli batter may be due to the quality of ingredients used and the type of packaging. Majority (31.3%) purchase batter which range between Rs. 10 to Rs.12, 26 per cent purchase between Rs. 13 to Rs.16. Regarding colour of the commercial idli batter majority (59%) of the selected population expressed that the colour was pale and unappealing and 64 per cent criticized that the commercial 33 idli batter had thin consistency which gave poor quality of idli and so it was used for dosa making. Table 2.6 Details about purchase of commercial idli batter (N=300) Particulars Percentage (%) Frequency of purchase Daily 49.3 Weekly 21.0 Monthly 19.7 Cost in rupees 7-9 16.3 10-12 31.3 13-16 26.3 17-20 14.7 21-30 00.7 Nil 10.7 Colour Bright 41.00 Pale 58.99 Consistency Thick 35.33 Thin 64.00 Preference Like 47.23 Dislike 52.77 34 Among the selected population 53 per cent disliked commercial idli batter especially for its aroma, out of which 31 per cent expressed problems regarding quality and shelf-life of the batter. The study revealed that all the respondents (100%) will purchase ready to cook idli batter if the batter quality, fermented aroma and shelf-life are improved. 2.4 CONCLUSION The results of the survey indicates the practices currently followed by the population, not statistically representative is similar to the practices reported in literature such as variety of rice, type of black gram, ratio of ingredients used for idli making, fermentation time and shelf –life of the batter. 35 TEXTURE OPTIMIZATION OF IDLI 3.1 INTRODUCTION Indigenous or native fermented foods have been prepared and consumed for thousands of years, and are strongly linked to culture and tradition. The fermented foods are better in terms of nutrition and easy for digestion than the normal cooked foods. The fermentation process causes enrichment and improvement of food through flavour, aroma, and change in texture, preservation by providing organic acids, nutritional enrichment, reduction of exogenous toxins and reduction in the duration of cooking. During traditional fermentation process, locally available ingredients, which may be of plant or animal origin, are converted into edible products by the physiological activities of microorganisms and have distinct odour (Steinkraus 1996, Reddy and Salunkhe 1980) namely Lactobacillus sp. and Pediococcus sp. which produce organic acids such as lactic acid and acetic acid, alcohol and carbon dioxide (Caplice and Fitzgerald 1999) and reduce the pH, thereby inhibiting the growth of food spoiling microorganisms. These fermented foods can be preserved for several days (Tamang, 1998) and also have therapeutic properties (Sekar and Mariappan, 2007). There are different types of fermented foods in which a range of different substrates are metabolized by a variety of microorganisms to yield products with unique and appealing characteristics (Caplice and Fitzgerald 1999). Among all traditional fermented foods in India, idli is a white, fermented (acid leavened), steamed, soft and spongy texture product, widely popular and consumed in the entire South India. Idli is the resultant product from the naturally fermented batter made from washed and soaked rice (Oryza sativa L.) and dehusked black gram dhal (Phaseolus mungo L.). Apart from its unique texture properties, idli makes an important contribution to the diet as a source of protein, calories and vitamins, especially B-complex vitamins, compared to the raw unfermented ingredients (Reddy et al., 1982). Traditionally, rice and black gram in various proportions are soaked and ground adding water in mortar and pestle to yield a batter with the desired consistency. Parboiled rice is preferred over raw rice for idli and dosa with rice: black gram usually fermented at 3:1 36 (Steinkraus et al, 1967, Jama and Varadaraj, 1999) weight ratio for making soft and spongy textured idli (Nazni and Shalini 2010). Black gram, the leguminous component of idli batter, serves not only as effective substrate but also provides the maximum number of micro-organisms for fermentation (Balasubramanian and Viswanathan, 2007a). As a result of fermentation, (Padhye and Salunkhe, 1978) observed a significant increase in predicted biological value. Fermentation also improves the protein efficiency ratio (PER) of idli over the unfermented mixture (Van Veen et al, 1967). Idli preparation in the conventional manner takes at least 18 h. The available instant idli pre-mixes do not provide the desired textural characteristic and also lack the typical fermented aroma and on the other hand, idli prepared in different households do not have consistent quality (Nisha et al, 2005). Fermented foods in general have immense scope for commercialization as foods with improved nutritional value as well as functional foods. Fermented foods with scientifically developed starter cultures can aid the commercialization of these products. However scientific optimization of the process is the basic necessity for commercialization of any product including the fermented foods. Several researchers have used RSM successfully to optimize the conditions for making products like boondi (Ravi and Susheelamma, 2005), tandoori roti, puri and parotta (Saxsena and Haridasrao, 1996 and Vatsala 2001) .The current study is undertaken to set an optimized condition for the preparation of idli which will help the manufacturers at industrial level to produce idli with the desired textural property. This would also help to make proprietary products using proper starter culture. The main objectives of this study were to explore the effect of rice and black gram dhal and fermentation time on the texture of idli, analyzing the instrumental texture profile (TPA) parameters as a function of raw material composition and fermentation time and to find the optimum levels to maximize the desirable textural properties of idli using RSM. 37 3.2 MATERIALS AND METHODS 3.2.1 Materials Different rice varieties namely IR 20 idli rice, raw rice, broken rice, ration rice and red rice were procured from local market and black gram variety Aduthurai 3 (ADT3) which has 24.16 per cent protein content was procured from Tamil Nadu Rice Research Institute (TRRI), Aduthurai, Tamil Nadu, India. They were cleaned and stored at refrigerated conditions until use. 3.2.2 Preparation of idli Before framing the design using CCRD, preliminary trails were conducted to choose the ratios of rice to black gram dhal. The trails were done using the rice to black gram dhal ratios as 3:0.5, 3:1, 3:1.5, 3:2, 3:2.5 and 3:3 respectively where rice ratios were kept constant and the dhal ratios varied. The fermentation time varied between 10 to 14h. In the trial, idli made from the ratio 3:1 and 3:1.5 with a fermentation time between 11 to 12 h gave better results. Based on this, the maximum and minimum values for the independent variables were chosen to frame the model. The rice and black gram dhal were mixed at different ratios as per the CCRD (Table 2.1). The rice and dhal were soaked for 4 h and ground separately to a coarse consistency and mixed together with salt. The batter was left overnight (time based on the developed design) for fermentation. The fermented batter was mixed thoroughly to expel the gas formed due to the release of carbon-dioxide .The batter was poured in idli mould, and steamed in the idli steamer for 15 minutes. The idli were brought to room temperature and then used for instrumental texture profile. 3.2.3 Experimental design 3.2.3.1 Response surface Methodology RSM is a collection of statistical and mathematical techniques useful for developing, improving, and optimizing processes in which a response of interest is influenced by several variables and the objective is to optimize this response. RSM has important 38 application in the development and formulation of new products, as well as in the improvement of existing product. It helps to study the effect of the independent variables, alone or in combination, on the responses. In addition to analyzing the effects of the independent variables, it provides a mathematical model, which describes the relationships between the independent and dependent variables (Myers and Montgomery, 1995). RSM has been very popular for optimization studies in recent years. RSM reduces the number of experiment trials needed to evaluate multiple parameters and their interactions. The graphical perspective of the mathematical model has led to the term Response Surface Methodology. Generally an optimization study involving RSM has three stages. The first stage is the preliminary experimental trials, in which the determination of the independent variables and their limits are carried out. The second stage involves the selection of appropriate experimental design followed by prediction and verification of the model equation. The last stage is the generation of response surface plots as well as contour plots of the responses as a function of the independent parameters and determination of optimum conditions. The model used in RSM is generally a full quadratic equation or the diminished form of the equation. The second order model can be written as Eqn.1. …………….Eqn. 1 where Y is the predicted response, β0, β j, β jj and β ij are regression coefficients for intercept, linear, quadratic and interaction coefficients respectively, k is the number of independent variables and Xi and Xj are coded independent variables. Response surface methodology has been widely applied in the food industry optimizing complex processes and products (Wong et al, 2003, Lee et al, 2006 and Sin 2006). In the present study RSM was used to determine the optimum conditions of two independent variables (rice to black gram dhal ratio and fermentation time) on the TPA and colour attributes of idli. A CCRD was constructed using software package Statistica (1999) from StatSoft, OK, USA. Five levels of each predictor variable were incorporated into the developed design. Table 1 shows levels of predictor variables. 39 3.2.3.2 Optimization of idli The procedure was based on the hypothesis that quality attributes of idli were functionally related to rice to black gram dhal ratio and fermentation time, and attempts were made to fit multiple regression equations describing the responses. Two coded independent variables in the process were rice to black gram dhal ratio (X1) and fermentation time (X2). Five levels of each of the independent variable were chosen for the study (Table 3.1); thus, there were 13 combinations, including the replicates of the center point that were performed in random order, based on an experimental CCRD for two factors. The dependent variables were hardness, adhesiveness, springiness, cohesiveness, chewiness and resilience and colour attributes. Table 3.1 Central composite rotatable design: Coded and actual values of independent variables Experimental design points Rice : black gram Ratio (w / w) Actual 1 3 : 0.72 2 3 : 0.72 3 3 : 1.78 4 3 : 1.78 5 3 : 0.50 6 3 : 2.00 7 3 : 1.25 8 3 : 1.25 9 3 : 1.25 10* 3 : 1.25 *Centre point repeated 3 times Fermentation time (h) Coded Actual Coded -1.000 -1.000 1.000 1.000 -1.414 1.414 0.000 0.000 0.000 0.000 10.58 13.42 10.58 13.42 12.00 12.00 10.00 14.00 12.00 12.00 -1.000 1.000 -1.000 1.000 0.000 0.000 -1.414 1.414 0.000 0.000 3.2.3.3 Instrumental Colour Measurement The colour parameters of idli were measured using a Hunter Lab colour flex model A601012-312 (Hunter Associates laboratory, Reston, VA). The equipment was standardized each time with white and black standards. Samples were scanned to determine lightness (L*), red-green (a*) and yellow-blue (b*) colour components (Olajide, 2010). As in the 40 work done by (Ronald and Daniel, 1998) the hue angles were derived as the arctangent of b*/a* expressed as degrees and the chroma values were also calculated as the square root of the sum of the squared values of both CIE a* and CIE b*. The chroma and Hue angle were calculated by the formula Eqn.2 and Eqn. 3, respectively. …………….Eqn. 2 …………….Eqn. 3 Where a* indicated Red-Green colour components, while b* indicates yellow to blue colour components (Ali, 2008).Plate 3.1 shows the picture of colour flex. Plate 3.1 Color flex 3.2.3.4 Texture profile analysis (TPA) The TPA test consists of compressing a bite-size piece of idli two times in a reciprocating motion that imitates the action of the jaw. The idli was cooled to room temperature and was cut into an inch cube (Plate 3.3) using an inch cubic mould (Plate 3.2.a). The texture of each idli was analyzed using SMS/75mm (Plate 3.2.b) compression platen in Texture 41 Rice (IR20 idli rice) and Black gram dhal (ADT3) (ratio based on the experimental design) Soak (4h) and grind Ferment the ground batter (Based on the experimental design) Pour batter in idli mould and steam for 15 minutes Cool idli to room temperature and cut the centre using one inch cubic mould TPA of cut idli using SMS/75mm compression platen Statistical Analysis (RSM; Regression) 42 Fig.3.1 Flow chart showing work design for TPA of idli Analyzer (Stable Micro Systems, Surrey,UK). The extra top and bottom layers were sliced off to make the idli fit to the mould. The cut piece was placed on the heavy duty platform and the test speed was set to 5mm/sec and the probe compressed 50% of the idli to get the TPA of the idli. Based on the force deformation curves, several parameters like adhesiveness, springiness, cohesiveness, chewiness and resilience can be calculated. Plate 3.2 Cutting idli with the designed mould 43 a) b) Plate 3.3 One inch cubic mould and SMS/75mm compression probe Plate 3.4 Texture analyzer 44 3.2.4 Statistical Analysis The independent variables and dependent variables (responses) were fit to the secondorder polynomial function and examined for the goodness of fit. The R2 or coefficient of determination is defined as the ratio of explained variation to the total variation and is a measure of the degree of fit (Haber and Runyon, 1997). All experimental designs and statistical data were analyzed and response surfaces, ANOVA, regression analysis were reported using Statistica (StatSoft, OK, USA) statistical software. 3.3 RESULTS AND DISCUSSION The results of chapter 3 are discussed under the flowing heads: 3.3.1Effect of rice varieties on rice batter volume 3.3.2 Effect of black gram on batter volume 3.3.3 Effect of ratios of rice to black gram dhal on batter volume 3.3.4 Response surfaces 3.3.5 Instrumental Colour measurement of idli 3.3.6 Texture parameters 3.3.7 Simultaneous optimization 3.3.1 Effect of rice varieties on rice batter volume In the present study five varieties of rice namely ration rice, raw rice; broken rice, red rice and parboiled rice were used for idli making. The rise in CO2 production can be correlated with the increase in batter volume (Sridevi et al., 2010).The percentage of increase in batter volume was significant (p< 0.05) in the batter prepared with ration rice, followed by parboiled rice, raw rice, broken rice, and red rice. Though there is high increase in batter volume, after expulsion of gas the volume of batter gets significantly decreased in ration rice, whereas the batter volume did not show significant (p< 0.05) decrease in parboiled rice. Table 3.2 and Fig 3.2.a shows the effect of rice varieties on batter volume. The sensory score of idli showed variation with the variety of rice used. As the idli prepared from parboiled rice is very soft when compared with idli made with other varieties. Parboiled rice may be best suited for idli making which is in par with the 45 result reported by Juliano and Sakurai (1985) that parboiled rice is better suited than raw rice for producing idli , i.e., it is soft without becoming sticky. The idli prepared using very light coloured parboiled rice are preferred by consumers traditionally accustomed to eating raw rice. Sowbhagya et al., (1991) studied the effect of variety, parboiling and ageing of rice on the quality of idli and reported that the normal parboiled rice is best suited for making idli as shown by its higher scores for softness. In the present study the idli made of parboiled rice is soft and it may also be due to fact proved by Sharma et al., (2008) that the greater starch damage in parboiled rice during wet grinding, attribute to its greater susceptibility to undergo damage owing to its softness after soaking as well as to the longer duration of grinding favouring parboiled rice to be suited for idli making. Roy et al., (2010) noted that the hardness and adhesion of cooked rice were dependent not only on the moisture content but also on the forms and variety of rice. Roy et al., (2004), Roy et al., (2008), Islam et al., (2001) and Shimizu et al., (1997) reported that the hardness of the cooked rice depend on the moisture content of cooked parboiled and untreated rice. In case of idli, steaming increases the moisture content of idli and it is a major factor that makes idli made with parboiled rice softer and for the same reason that red rice has acquired more moisture which affected its texture losing firmness. 3.3.2 Effect of black gram on batter volume The percentage of increase in batter volume was significant (p< 0.05) at five per cent level (Table 3.3) for the batter made from parboiled rice and black gram used with husk, and thou the idli made from the same batter were spongy, the colour was unappealing to the panel members. The difference in batter volume was not significantly higher with the batter made from the black gram with husk removed. On the other hand, though, the percentage of increase in batter volume was low (38.9%) in the batter made from parboiled rice and black gram dhal with husk removed after soaking, 46 Table 3.2 Effect of rice varieties on the batter volume after fermentation Varieties of rice Batter characteristics Ration rice Raw rice Broken rice Red rice 211.1±1.55 b 218.6±0.98 c 200.5±0.14 a 238.2±0.14 e 299.7±1.41 d 275.8±2.61 b 248.8±2.61 a 293.7±3.53 c 38.1±1.27 c 42.0±0.70 d 26.2±0.35 b 24.1±0.21 a 23.3±0.28 a 202.0±0.56 e 150.8±0.84 a 181.0±0.70 c 193.0±1.83 d 158.0±1.41 b Batter volume decreased after expulsion of expulsion of gas (%) 34.0±1.41 b 49.7±0.00 d 34.4±0.63 b 22.4±1.41 a 46.1±0.0 c Sensory Rank I IV II III V Initial volume of the batter (cm3) Final volume of the batter (cm3) Batter volume increased after fermentation (%) Volume of the batter after expulsion of gas (cm3) Parboiled rice 221.6 ±2.05 d 306.1± 3.74 e mean values in a row with different letters differ significantly p<0.05 by LSD (n=3) the texture was very spongy and the colour was also appealing making this variation a better choice in terms of colour and texture on sensory basis. Fig 3.2.b shows the effect of variation in black gram dhal on batter volume. 3.3.3 Effect of ratios of rice to black gram dhal on batter volume The percentage of increase in batter volume was high for the ratio 3:3.5 (w/w) of rice to black gram dhal respectively with 5% significance followed by other ratios such as 3:3, 3:2.5 and so on. When the texture of idli was compared on sensory basis, the idli made of ratio 3:1 was very spongy compared to idli made of other ratios of rice and black gram dhal showing that the proportions of compositions of the substrate also have an important role in the outcome of the product. Table 3.4 and Fig 3.2.c shows the effect of ratios of ingredients on batter volume. Hence for the further study parboiled rice namely IR 20, black gram variety namely ADT 3 with husk removed after soaking was used to find the effect of ingredients and descriptive sensory profile of idli. 47 Table 3.3 Effect of black gram (var. ADT 3) on the batter volume after fermentation Black gram BHR Batter characteristics Initial volume of the batter (cm3) BHRAS BWH 271.4 ±0.14 c 226.1 ±0.84 a 339.2±0.131 b 376.9 ±0.07 c 324.1 ±0.00a 28.6 ±0.00 a 38.9 ±0.07 b 043.3 ±0.07 c 248.7 ±0.35 b 256.3 ±0.42 c 211.1 ±0.07 a 26.7 ±0.00 a 32.0 ±0.00 b 34.9±0.14 c 263.8±0.14 b Final volume of the batter (cm3) Batter volume increased after fermentation (%) Volume of the batter after expulsion (cm3) Batter volume decreased after expulsion of gas (%) Sensory Rank II I III Mean values in a row with different letters differ significantly p<0.05 by LSD (n=3) BHR-black gram husk removed; BHRAS-black gram husk removed after soaking; BWH -black gram with husk Table 3.4 Idli batter volume characteristics as affected by parboiled rice and black gram dhal (without husk) Rice and black gram ratio (w/w) Batter characteristics Initial volume of the batter (cm3) Final volume of the batter (cm3) Batter volume increase after fermentation (%) Volume of the batter after expulsion (cm3) Batter volume decrease after expulsion of gas (%) Sensory Rank 3:1 3 : 1.5 3:2 3 : 2.5 3:3 3 : 3.5 150.7 ±0.07a 241.2±0.28c 301.5±0.21d 324.1 ±0.14e 339.2 ±1.13f 414.6 ±0.07g 248.7±1.13a 316.6 ±0.84c 422.7 ±0.07d 452.3 ±0.07e 467.4 ±0.00f 603.1 ±0.07g 65.0 ±0.0a 31.3 ±0.07c 40.2 ±0.07d 39.6 ±0.28e 37.8 ±0.35f 45.5 ±3.0.07g 158.3 ±0.21a 173.4 ±0.0b 233.7 ±0.14d 248.7 ±0.28e 256.3 ±0.14f 301.5 ±0.0g 36.3 ±0.28b 45.2 ±0.28d 44.7 ±0.28c 45.0 ±1.41d 44.2 ± 1.13c 50.0±0.14e I III II IV V VI Mean values in a row with different letters differ significantly p<0.05 by LSD (n=3) 48 Fig.3.2.a Effect of rice varieties on batter volume after fermentation Fig.3.2.b Effect of type of dhal on batter volume after fermentation Fig.3.2.c Effect of ratios of rice to black gram dhal on batter volume after fermentation 49 3.3.4 Response surfaces Several parameters namely raw material variety, quality, their proximate composition, raw material composition, particle size, temperature etc., affect the texture of idli but still, the texture of idli is very unique from the consumer point of view. Among all the parameters mentioned, fermentation time is one of the key factors which can affect the texture due to its air production and leavening action. The texture of the cooked idli is a subject of interest, to judge and optimize the production process of good textured idli with the selection of the ingredients and the process. The fermentation periods are slightly different for idli making owing to the difference in raw materials, composition, process and region (Balasubramanian and Viswanathan, 2007b). 3.3.5 Instrumental Colour measurement of idli Colour of the idli is one of the most important parameter for the acceptability of the product. The colour of the idli showed variation based on the ratio of rice and black gram dhal used. The L*, a*, b* values and graph are shown in Table 3.5 and Fig.3.3.a, b, c respectively. The L* value which correspond to lightness ranged from 73.40 to 75.99 indicating the difference in the proportion of black gram dhal used. The positive values of b* indicates yellowness in the idli, which may be due to the use of black gram with husk for soaking. The chroma (Fig.3.3.d) values are closer to the b* values. The hue angle value corresponds to whether the object is red, orange, yellow, green, blue, or violet (Ali et al, 2008). The negative values in the hue angle shows that the product deviates from the colour adding positive factor to the current study because lightness in the colour of the idli is an important factor in the view of customer perception. The intensity of chroma is low for the idli made with the ratio of 3:0.5 and is higher for the idli made from the ratio 3:2 showing that the ratio of rice and dhal used for idli making has an impact on the intense of chroma of the idli. 50 Table 3.5 Experimental design: CCRD with actual levels of independent variables for colour parameters Instrumental colour parameters Experimental design points L* a* b* Chroma 1 74.03 + 0.07 -0.44+0.021 11.52+ 0.064 11.56 Hue angle (°) -87.72 2 74.13 + 0.07 -0.57+0.007 10.60+0.035 10.59 -86.92 3 75.76 + 0.11 -0.25+0.028 12.21+0.085 12.15 -88.92 4 73.99 + 0.06 -0.24+0.021 13.57+0.007 13.56 -89.03 5 75.57 + 0.07 -0.76+0.035 10.01+0.360 9.936 -85.79 6 75.78 + 0.03 -0.02+0.070 15.97+0.085 16.03 -89.89 7 73.40 + 0.11 -0.43+0.014 13.09+0.177 12.96 -88.14 8 74.32 + 0.51 -0.13+0.014 11.88+0.205 11.74 -89.14 9 74.35 + 0.11 -0.40+0.007 10.56+0.163 10.44 -87.81 10* 74.36 + 0.05 -0.43+0.028 10.61+0.361 10.35 -87.73 *Centre point replicated 3 times 51 Fig. 3.3.a. Response surface graph showing relation between independent parameters on L* Fig. 3.3.b Response surface graph showing relation between independent parameters on a* 52 Fig.3.3.c Response surface graph showing relation between independent parameters on b* Fig.3.3.d Response surface graph showing relation between independent parameters on Chroma 53 3.3.6 Texture parameters The experimental values for the response variables of texture analysis are shown in Table 3.6. Figure 3.4.a and Figure 3.4.b shows the typical TPA graph. Hardness of idli is indicated by the maximum force required to compress the idli and usually represented by the first peak in the graph. The hardness of the idli (Fig.3.5.a) varies between a minimum force of 20.58 N to a maximum force of 44.19 N i.e., the minimum force was required to compress idli of ratio 3:0.72 at 13.42 h fermentation time and the maximum force for the ratio 3:1.78 at 10.58 h of fermentation time. This variation in the force is due to the variation in the ratio of the ingredients and fermentation time of the batter. Higher the force shows that harder is the idli. ANOVA results indicated that the ratio of rice and black gram dhal used for idli making (in the linear effect) is significant (P< 0.05) to the hardness of the idli. The co-efficient of regression is given in Table 3.7. The goodness of fit was high with R2 value =0.942. Adhesiveness of idli can be defined as the negative force area for the first bite and represents the work required to overcome the attractive forces between the surface of the cut piece of idli and the surface of the probe with which the idli comes into contact, i.e. the total force necessary to pull the compression plunger away from the food. The negative area in the graph is taken as the adhesiveness. The adhesiveness of the idli varies between -0.00051N s to -0.05127 N s. If the product is sticky, the adhesiveness will be higher. Ghasemi et al, (2009) reported that the adhesiveness may be due to the gelatinization and more fluidity of rice starch structure in the cooked samples. As idli is adhesive in nature, to optimize the product minimum adhesiveness can be considered. In the current study since the batter was coarse ground and cooking time was constant the adhesiveness must be due to the ratio of rice and dhal and the quality of the ingredient. The minimum adhesiveness is obtained for the idli made of ratio 3:0.5 at 12 h fermentation time and the maximum adhesiveness is obtained for the ratio 3:0.72 at 10.58h fermentation time. Fig. 3.5.b shows the response surface graph for adhesiveness. 54 Force (N) Time (sec) Fig.3.4.a Texture profile of idli made of ratio 3:1.25 at 12 h fermentation time Force (N) Time (sec) Fig.3.4.b Texture profile of idli made of ratio 3:2 at 12 h fermentation time 55 Table 3.6 Experimental design: CCRD with coded and actual levels of independent variables for TPA Experi mental design point 1 Dependent variables Hardness (N) Adhesiveness (N s) Springiness Cohesiveness Chewiness Resilience 23.73±2.01 -0.0512±0.0045 0.926±0.33 0.876±0.12 1963.61± 16.26 0.595 ± 0.12 2 20.58±1.42 -0.0337±0.0038 0.960±0.28 0.819±0.04 1650.89±14.05 0.562 ± 0.52 3 44.19±2.02 -0.0284±0.0042 0.809±0.41 0.643±0.09 2344.08±21.01 0.340 ± 0.41 4 36.57±2.24 -0.0005±0.0037 0.847±0.20 0.674±0.07 2127.97±16.42 0.404 ± 0.24 5 20.66±3.52 -0.0051±0.0069 0.854±0.32 0.912±0.17 1845.66±18.01 0.654 ± 0.42 6 32.47±4.13 -0.0290±0.0053 0.965±0.48 0.825±0.02 2333.37±14.01 0.511 ± 0.54 7 35.36±1.41 -0.0085±0.0075 0.733±0.24 0.526±0.04 1389.17±13.32 0.285 ± 0.10 8 24.12±2.14 -0.0008±0.0061 0.916±0.42 0.755±0.04 1701.18±12.42 0.483 ± 0.27 9 30.85±0.05 -0.0062±0.0047 0.928±0.31 0.876±0.02 2557.13±11.14 0.579 ± 0.13 10* 30.72±1.28 -0.0057±0.0039 0.913±0.31 0.885±0.06 2532.79±15.05 0.574 ± 0.41 *Centre point replicated 3 times Springiness is the height that the idli recovers during the time that elapses between the end of the first bite and the start of the second bite, usually in TPA the first compression and second compression. The difference between the first peak and the second peak in the graph is taken as springiness. The springiness of idli depends on the quantity of the dhal used because the soft spongy texture observed in the leavened steamed idli made out of black gram is due to presence of two components, namely surface active protein (globulin) and a polysaccharide (arabinogalactan) in black gram (Susheelamma and Rao 1974, 1979a, 1979b, 1980). The specialty of black gram in idli preparation is due to the mucilaginous property which helps in the retention of carbon-dioxide evolved during fermentation (Nazni and Shalini, 2010). In the current study the springiness varied from 0.733 to 0.965. The maximum springiness is obtained for the ratio 3:2 at 12 h 56 fermentation time. Hence the result reveals that the quantity of black gram dhal used has a major role in the springiness of the idli. The response surface graph in 3D is depicted in Fig.3.5.c showing the relation between rice to black gram dhal ratio and fermentation time on springiness. From the ANOVA table it is clear that the independent variables in the linear effect showed a significant influence on the springiness of the idli and the model showed high goodness of fit (R2 = 0.909) . Cohesiveness is defined as the ratio of the positive force area during the second compression to that during the first compression. Cohesiveness may be measured as the rate at which the material disintegrates under mechanical action. The cohesiveness is minimum (0.526) for the ratio 3:1.25 at 10 h fermentation time and maximum (0.912) for the ratio 3:0.5 at 12 h fermentation time. Both the independent variables namely rice to black gram dhal ratio in linear effect and fermentation time in quadratic effect is significant at 5 % level on the cohesiveness of the idli. The graph in Fig.3.5.d shows an initial increase in the cohesiveness as the fermentation time increases, but gradually decreases with further increase in fermentation time. 57 Fig.3.5.a Response surface graph showing relation between independent parameters on hardness Fig.3.5.b Response surface graph showing relation between independent parameters on adhesiveness 58 Fig.3.5.c Response surface graph showing relation between independent parameters on springiness Fig.3.5.d Response surface graph showing relation between independent parameters on cohesiveness 59 Fig.3.5.e Response surface graph showing relation between independent parameters on Chewiness 60 Fig.3.5.f Response surface graph showing relation between independent parameters on resilience 61 Table 3.7 Regression co-efficient for dependent TPA parameters Regression Co-efficient Independent variables Mean/Interaction 1. Rice : Dhal ratio (L) Rice : Dhal ratio (Q) 2. Fermentation time (L) Fermentation time (Q) 1L by 2L R2 Hardness Springiness Cohesiveness Chewiness Resilience 34.390 -2.254 -5.873 00.00 -4.602 31.132 1.981 -3.517 0.147 -1.645 0.942 -0.182 -0.001 0.525* -0.021 0.008 0.908 -0.529 -0.049 1.161* -0.049* 0.042 0.886 661.94 -378.64 4241.93* -178.51* 63.76 0.85 -0.603 -0.003 0.906* -0.038* 0.036 0.931 L - Linear effect; Q - Quadratic effect; *=P < 0.05 Table 3.8 Analysis of Variance (ANOVA) for dependent TPA parameters: F values Dependent parameters Independent variables 1. Rice : Dhal ratio (L) Rice : Dhal ratio (Q) 2. Fermentation time (L) Fermentation time (Q) 1L by 2L Hardness 000.000 241.174* 000.000 063.752 1050.770* Springiness Cohesiveness Chewiness Resilience 15.644* 0.001 15.404* 7.401 0.138 12.755* 0.228 5.074 11.447* 1.027 11.161* 1.134 0.487 12.628* 0.198 0.524* 96.244 3.823* 2.724* 31.967 L - Linear effect; Q - Quadratic effect; *=P < 0.05 Chewiness is defined as the product of hardness x cohesiveness x springiness and is therefore influenced by the change of any one of these parameters. Lower the chewiness softer is the idli. The chewiness of the idli varied between 1389.172 for the ratio 3:1.25 at10 h fermentation time to 2557.135 for the ratio 3:1.25 at 12 h fermentation time. It is proved by the ANOVA table (Table 3.8) that the ratio of rice to black gram dhal in linear effect and fermentation time in quadratic effect also have significant impact (P < 0.05) 62 on the chewiness of the idli. As hardness, springiness and cohesiveness show significant influence because of the independent variable hence the chewiness of the idli will also be affected by the both independent and dependent variables. The chewiness (Fig.3.5.e) of the idli varied for the same ratio of idli with difference in fermentation time which relates the decrease in cohesiveness with further increase in fermentation time. Resilience is a measurement of how the sample recovers from deformation both in terms of speed and forces derived. It is taken as the ratio of areas from the first probe reversal point to the crossing of the x-axis and the area produced from the first compression cycle. The resilience varies between 0.285 for the ratio 3:1.25 at 10 h fermentation time to 0.654 for the ratio3:0.50 at 12 h fermentation time. Lower resilience value shows that the product can recover faster from deformation proving the firmness of the product. The response surface graph in 3D is depicted in Fig.3.5.f showing the relation between rice to black gram dhal ratio to fermentation time on resilience of the idli. From the ANOVA table it is evident that the resilience of the idli is influenced significantly by rice to black gram dhal ratio in linear effect and by fermentation time both linear and quadratic effect. The closer the value of R2 approaches unity, the better the empirical model fit the actual data (Nuraliaa et al., 2010). As the R2 value for resilience (0.932) was closer to unity and the result of resilience fit to the actual data. 3.3.7 Simultaneous optimization Simultaneous optimization was performed on the TPA parameters like hardness, adhesiveness, springiness, cohesiveness, chewiness and resilience by imposing desirability constraints. In case of springiness, the softer idli shows high springiness. Hence the software take into account of the values of independent and dependent values and finally gives a maximum desirable score and the condition at which the maximum score can be obtained with some constraints by assigning maximal desirability score as one and minimal desirability score as zero. Table 3.9 shows the constraints imposed for good textured idli with the desirable value for both independent and dependant variables. The maximum desirable score that can be achieved with the desirable value will be 0.8279. On the basis of these calculations good textured idli could be made when 3:1.575 63 (mass) ratio of rice to black gram dhal respectively is fermented for 14 h. The optimum results were validated by performing the experiment at the optimized ratio and fermentation time by comparing the observed and the predicted values. The predicted values are shown in Table 3.9. The predicted values were insignificant with observed values indicating the appropriateness of the model developed. Table 3.9 Simultaneous optimization of process parameters by desirability approach Dependent variables Independent parameters Rice : dhal Fermentation ratio (w/w) time (h) 3 : 1.575 14.00 TPA Overall Constraints Predicted Observed imposed values values Hardness Minimum 19.340 019.92 ± 01.03 Adhesiveness Minimum -0.030 -0.032 ± 00.01 Springiness Maximum 0.947 0.930 ± 00.14 Cohesiveness Minimum 0.773 000.78 ± 00.02 Chewiness Minimum 1299.7 1286.8 ± 32.20 Resilience Maximum 0.555 0. 547 ± 00.030 L* (lightness) Maximum 75.16 075.21 ± 00.58 parameters and L* values Desirability score 0.8279 3.4 CONCLUSION The optimization results indicated that the optimum ratio of rice to black gram dhal is 3:1.575 (w/w), with 14 h of fermentation time will provide the product with maximum score for desirable textural parameters. 64 PROCESS OPTIMIZATION OF IDLI USING SENSORY ATTRIBUTES 4.1 INTRODUCTION Fermented foods are defined as foods that have been subjected to the action of selected microorganisms by which a biochemically and organoleptically modified substrate is produced, resulting in an acceptable product for human consumption (Tamang, 1998). There are different types of fermented foods, in which a range of different substrates are metabolized by a variety of microorganisms to yield products with unique and appealing characteristics (Campbell-Platt, 1994). Fermented foods supply important nutrients, particularly proteins and amino acids. People become familiar with particular fermented foods produced in their part of the world, and many of these foods became an integral part of the local diet (Caplice and Fitzgerald, 1999) and culture, and were regarded as essential for human consumption and nutrition. Idli is one such food, which is prepared from low cost staple crop, which helps to improve health. Its composition includes rice and black gram. In the traditional idli batter, fermentation takes place due to the microflora present in the raw materials and in the environment leading to the several changes that has impact on digestibility and nutritional value bringing about desirable changes (Soni and Sandhu, 1989). The example of idli illustrates the opportunities of co- fermentation of cereals (rice) and leguminous seeds (black gram) (Young and Pellet, 1994). A large proportion of the world cereal production is processed by fermentation prior to consumption. The enhancement of attractive flavour and texture and the improved shelflife and digestibility as a result of fermentation, are important reasons for fermenting cereals before consumption (Nout, 2009). Characteristic variables such as water content, i.e. before and after soaking or fermentation, duration and temperature affect the cereal fermentation (Hammes and Ganzle, 1998). This sour and spongy type breakfast food (idli) of India and Sri Lanka constitutes an important group of naturally fermented food (Ramakrishnan, 1979). 65 Legume contains more of proteins than cereals (Geervani and Theophilus, 1981). Changes in the nutritive value of proteins as a result of fermentation are particularly important for cereals and legumes. These sources of protein often are of lower nutritional quality than animal products, and they tend to be major dietary sources of protein for people with marginal and sub-marginal protein intake. Therefore, fermentation processes that consistently improve protein quality or availability of cereal or legumes could have a positive impact on the diets of people (McFeeters, 1988). Cereal and legume being the important component of idli, the present study is done to find out the interrelationship between the substrates during fermentation at different fermentation time and different ratios of the substrates on the sensory attributes of idli including desirable and non-desirable parameters with the objectives to select the ingredients for optimum desirable product characteristics and to identify the optimum levels of ingredients and fermentation time with respect to sensory attributes using Response Surface Methodology (RSM). 4.2 MATERIALS AND METHODS 4.2.1 Materials In the current study, the most commonly used local variety of rice namely IR 20idli rice and a protein rich black gram variety Aduthurai 3 (ADT3) which were chosen from the preliminary study were used. 4.2.2 Preparation of Idli Before framing the design using CCRD, preliminary trails were conducted to choose the best suited rice, variation of black gram and ratios of rice and black gram dhal. The varieties of rice chosen were parboiled rice, raw rice, ration rice, broken rice and red rice. The variations in black gram dhal were black gram with husk, husk removed and husk removed after soaking. The different ratios of rice to black gram dhal used were 3:1, 3:1.5, 3:2, 3:2.5, 3:3 and 4:1 respectively. The difference in batter volume after fermentation and the texture of idli based on sensory was used to screen the ingredients and ratios. The result of the preliminary study is discussed to show the reason for choosing the maximum and minimum values for the independent variables chosen to 66 frame the model. The rice and black gram dhal were mixed at different ratios as per the CCRD. To carry out the experiment framed using CCRD, the rice and black gram dhal were soaked for 4 h and ground separately to a coarse consistency and mixed together. The batter was left overnight (time based on the developed design) for fermentation with addition of salt. The fermented batter was mixed thoroughly to expel the gas formed due to the release of carbon-dioxide. The batter was poured in idli mould, and steamed in the idli steamer for 15 minutes. The cooked idlis were subjected to sensory analysis. 4.2.3 Experimental design 4.2.3.1 Response Surface Methodology A response surface methodology as explained by Box and Wilson (1951) was conducted to determine the relative contributions of two predictor variables (ratio of rice to black gram dhal and fermentation time) to the quality of the idli. RSM is an effective tool for optimizing complex processes and has been widely applied in the food industry (Wong et al, 2003; Lee et al, 2006; Sin et al, 2006). A CCRD was constructed using software package Statistica (1999). Maximum and minimum predictor values were chosen after carrying out preliminary cooking trails. Five levels of each predictor variable were incorporated into the design. Table 4.1 shows levels of predictor variables. RSM reduces the number of experiment trials needed to evaluate multiple parameters and their interactions. For idli preparation different ratios of rice to black gram dhal and fermentation time can be optimized using RSM keeping temperature constant (30o C). 4.2.3.2 Optimization of idli using RSM The procedure was based on the hypothesis that quality attributes (desirable and undesirable parameters) of idli were functionally related to ratios of rice to black gram dhal and fermentation time, and attempts were made to fit multiple regression equations describing the responses. Two coded independent variables in the process were rice to black gram dhal ratio (X1) and fermentation time (X2). Five levels of each of the independent variable were chosen for the study; thus, there were 15 combinations, including the replicates of the centre point that were performed in random order, based on an experimental CCRD for two factors as shown in Table 3.1 (Chapter 3). 67 4.2.3.3 Sensory analysis of idli Idli samples were coded and served to ten panel members for analysis. The desirable parameters included were colour, fluffiness, sponginess and fermented aroma. The undesirable parameters included were compactness, firmness, stickiness and sourness. The score card also had an option to give the score for overall quality of the sample. The attributes selected were shown in Table 3. The panelists evaluated three sets of samples at separate time. The first set included samples made with ratios 3:0.5 and 3:0.72 of rice and black gram dhal respectively, the second set included samples made with ratio 3:1.25 and the third set included samples made with ratios of 3:1.78 and 3:2 with the respective fermentation time as shown in Table 3.1. In each set 3 samples of idli were placed for evaluation. The panel members were given a fifteen point rating scale to evaluate the idli. The ranges of the quality of idli were given by panelist by marking a line on the rating scale. The marking in the rating scale was counted as the score by using a measurement scale. 4.2.3.4 Quantitative Descriptive Analysis (QDA) The principle of QDA is based on the ability to train panelists to measure specific attributes of a product in a reproducible manner to yield a comprehensive quantitative product description amenable to statistical analysis (Ghosh and Chattopadhyay, 2011). The panel members were selected and trained as how to evaluate the sample based on the desirable and undesirable parameters for idli. PCA of the fermented food sample was performed with the data collected from the panelists after scoring through 150 mm unstructured scale. The descriptive sensory attributes are shown in Table 4.1. 4.2.4 Statistical analysis of data The fitness of good was found through R2 or coefficient of determination (Haber and Runyon 1977). All experimental designs and statistical data were analyzed and response surface graphs, ANOVA, regression analysis were reported using Statistica (StatSoft, OK, USA) software. 68 Table 4.1 Sensory attributes used for sensory analysis of Idli Sensory attribute Description Range The colour of the idli range from pale yellow Low to high Color to white Appearance Fluffiness Compactness Texture Sponginess Firmness Stickiness Aroma Fermented Taste Sour Overall quality The extent of fluffy appearance after cooking the batter The lack of porous nature in the idli The soft feeling obtained by the panelist while touching the idli The rigid nature of idli experienced by the panelist by touch or bite The adhesiveness of the idli experienced on touch Low to high Low to high Low to high Low to high Low to high The characteristic aroma after the fermentation of rice and dhal Low to high The range showing the extent of fermentation on tasting Low to high The impact of the product based on other sensory attributes expressed by the panelist revealing the acceptability of the product Low to high 4.2.4.1 Principal Component Analysis (PCA) Principal component analysis (PCA) is a statistical technique that can be applied to QDA data to reduce the set of dependent variables (i.e., attributes) to a smaller set of underlying variables (called factors) based on patterns of correlation among the original variables (Lawless and Heymann, 1998). 69 4.3 RESULTS AND DISCUSSION The results are discussed under the following heads: 4.3.1 Desirable parameters of idli 4.3.2 Negative drivers of liking 4.3.3 Overall quality of the idli 4.3.4 Simultaneous optimization 4.3.1 Desirable parameters of Idli The desirable and undesirable parameters of idli were evaluated by sensory analysis because the evaluation of different cooked varieties of idli revealed the wide acceptance of the conventional product due to its attractive aroma, taste and consistency (Soni and Sandhu, 1989). The sensory parameters as shown in Table 4.1 were studied for the idli made from the parboiled rice and black gram dhal with husk removed after soaking with the ratios framed using CCRD at varying fermentation time. The idli showed large difference in the sensory parameters in relation to the ratios and timing of fermentation which is supported by the study done by Ghosh and Chattopadhyay (2011) who reported that the changes during fermentation affect the physical properties like appearance, texture, aroma, flavour and overall acceptability and these parameters are vital to assess the acceptability of the product in the consumer point of view. Table 4.2 shows the score given by panel members for desirable parameters. Table 4.4 shows the regression coefficient values for the desirable parameters. The R2 values for colour, fluffiness, sponginess were 0.953, 0.915 and 0.806 respectively which reaches unity favouring the product. 4.3.1.1 Colour The colour of the idli varied with the difference in ratios of the ingredients and change in fermentation time. There was improvement in the colour of idli with increase in fermentation time. As the ratio of black gram dhal increased there was gradual decrease in brightness of idli colour due to the black gram dhal content. The R2 value (Table 4.4) for colour was found to be 0.953. Fig.4.1.a to Fig 4.1.d shows the response surface graphs for desirable parameters. 70 Table 4.2 Experimental designs and mean scores of desirable sensory attributes Sensory attributes Overall quality Experimental Colour Fluffiness design points 1 8.7±0.22 08.2±0.26 2 9.2±0.14 07.4±0.38 3 6.9±0.21 10.1±0.38 4 7.3±0.29 10.7±0.41 5 8.6±0.19 08.0±0.39 6 5.2±0.42 11.4±0.67 7 8.6±0.23 09.6±0.51 8 8.3±0.49 10.2±0.54 9 9.1±0.46 09.7±0.43 10* 8.8±0.44 10.2±0.64 * Centre point repeated 3 times Sponginess 09.7±0.47 10.6±0.43 11.0±0.83 11.2±0.62 07.6±0.35 12.3±0.51 10.4±0.46 11.3±0.32 11.5±0.58 11.4±0.51 Fermented aroma 10.2±0.69 11.1±0.62 10.2±0.47 10.9±0.46 09.6±0.43 09.5±0.42 08.3±0.61 11.4±0.61 10.4±0.60 10.7±0.47 09.2±0.34 08.7±0.33 11.4±0.73 11.7±0.43 07.5±0.27 11.6±0.59 09.8±0.41 11.7±0.59 12.1±0.33 11.9±0.49 4.3.1.2 Fluffiness and sponginess of idli Texture of idli is very critical from consumer point of view, it should be spongy, soft and fluffy (Ramakrishnan 1979, Radhakrishnamurthy et al., 1961and Desikachar et al., 1960). The texture of idli is influenced by many variables like raw material, quantity, soaking time, grinding conditions, fermentation temperature and time and adjuncts on quality of idli (Desikachar et al., (1960); Radhakrishnamurthy et al., (1961). The fluffiness and sponginess increased with increase in the ratio of black gram dhal and fermentation time. The maximum score for fluffiness 71 Fig.4.1.a Response surface graph for colour Fig.4.1.b Response surface graph for fluffiness 72 Fig.4.1.c Response surface graph for sponginess Fig.4.1.d Response surface graph for fermented aroma 73 is 11.4 for the idli made of ratio 3:2 at 12 h fermentation time. The R2 value for fluffiness was 0.915. The important factor affecting the texture (sponginess, firmness and stickiness) of the idli is the starch content of the ingredients which is supported by the study done by Tharanathan and Mahadevamma (2003), that apart from its energy contribution, starch content is the major factor which governs the texture of idli and as a result, to the organoleptic properties of food. It was reported that the spongy texture of idli is also due to the presence of surface active proteins (globulin) that generate a foamy character resulting in the porous structure to the idli and this porous structure is stabilized even during steaming process by the presence of viscogenic mucilaginous polysaccharide called arabinogalactan (Susheelamma and Rao, 1979) proving that this viscosity and foam stabilizing properties of native polysaccharide is a special functional value of foods prepared from black gram (Tharanathan et al.,1994). As the starch content of IR20 rice is 79.5 per cent and that of black gram is 52 percent and the protein content of rice and dhal were 6.46 and 24.16 respectively the texture of the developed idli is found to be good. 4.3.1.3 Fermented aroma In case of fermented foods the shelf-life, texture, taste and aroma of the final product is improved because of fermentation. The changes in fermentation depend on the available nutrients in the starting materials, the unique metabolic abilities of the fermenting microorganisms and possible interactions among all of these elements (McFeeters, 1987). The response surface graph reveals that fermented aroma increased with increase in fermentation time. Mukherjee et al., (1965), Rajalakshmi and Vanaja (1967) have reported that black gram naturally possess L. mesenteroides, and the gas, acid and several volatile compounds are formed during fermentation which contribute to a complex blend of flavours in the products (Chavan and Kadam, 1989). 4.3.2 Negative drivers of liking The compactness was high for the idli made with ratio 3:1.25, followed by 3:0.5 and low for the idli made of ratio 3:2 (Table 4.3). This shows that the proportion of the 74 ingredients have a direct impact on the quality of the product. Fig.4.2.a shows that compactness decreased with increase in fermentation time and with high quantity of black gram dhal. The firmness (Fig.4.2.b) of the idli was high for the ratio 3:0.5 and it was noted that firmness decreased with increase in black gram dhal quantity. The undesirable parameter does not insist that the attributes are not required for the product but should have moderate effect on the product. The attributes such as compactness, firmness and stickiness when high are generally disliked by the consumers. Table 4.4 shows the regression co-efficient values for the undesirable parameters Table 4.3 Experimental designs and mean scores of undesirable sensory attributes Experimental Compactness Firmness Stickiness Sourness 1 8.7±0.57 9.2±0.47 11.3±0.46 8.5±0.40 2 8.5±0.46 9.6±0.53 11.1±0.50 9.6±0.58 3 9.2±0.58 5.8±0.42 9.8±0.44 8.9±0.45 4 8.1±0.55 6.2±0.43 9.2±0.41 10.3±0.38 5 9.4±0.74 11.5±0.50 8.3±0.43 8.2±0.37 6 7.8±0.65 5.3±0.35 7.4±0.35 8.1±0.53 7 9.7±0.65 5.8±0.52 7.1±0.35 6.4±0.29 8 8.1±0.53 4.9±0.52 7.1±0.38 9.7±0.42 9 8.5±0.68 5.7±0.44 7.7±0.43 9.1±0.44 10* 8.7±0.51 5.4±0.53 7.4±0.35 9.3±0.49 design points * centre point repeated 3 times 75 Fig.4.2.a Response surface graph for compactness Fig.4.2.b Response surface graph for firmness 76 Fig.4.2.c Response surface graph for stickiness Fig.4.2.d Response surface graph for sourness 77 4.3.2.1 Stickiness of the idli The physicochemical properties such as moisture content, adhesion and hardness are all induced by the processing conditions which affect the textural as well as eating quality of rice. The parboiling treatment given to rice decreases the stickiness (Roy et al., (2004), Kato et al., (1983), Biswas et al., (1988) and Islam et al., (2001). Rice with low amylose content is generally soft when cooked, whereas rice with high amylose content has higher grain hardness (Juliano, 1971). High-amylose rice has more long chains than lowamylose rice (Hizukuri et al., (1989) and Radhika et al., (1993). The more long chains, the firmer the rice is when cooked and vice-versa (Bhattacharya, 2004). Rice with high water binding capacity normally yields soft texture cooked rice (Mohapatra and Bal, 2006). In the present study the amylose content was low in the parboiled rice (32%) and black gram dhal (17%) compared to the amylopectin content (Table 5.1) hence the stickiness is due to the ratios of rice and black gram dhal and the fermentation time. An optimum ratio and fermentation time can yield a product with low to minimum stickiness. Fig 4.2.c shows the response graph for stickiness of idli. 4.3.2.2 Sourness of Idli Fig.4.2.d show that sourness increased with increase in fermentation time. The sourness was high (10.3) for the idli made of ratio 3:1.78 at 13.42 h fermentation time. Increase in fermentation time increases the acidity of the batter due to microbial growth which leads to increase in sourness of the batter. The R2 value for sourness was found to be 0.884. 78 Table 4.4 Regression co-efficient for sensory parameters Fermented aroma Compactness Firmness Stickiness Sourness -6.367 -2.919 -18.396 -6.970 16.848 27.570 36.448 -17.399 6.722 -0.551 10.968 3.152 3.359 -18.180 -15.245 0.959 -3.332* -1.068 -2.465 -0.974 -0.112 5.804 5.762 -0.77 2.019 1.899 3.4125 2.072 -1.357 -1.353 -2.905 3.677 -0.081 -0.101 -0.121 -0.061 0.059 0.055 0.125 -0.132 -0.033 0.465 -0.233 -0.066 -0.299 0.000 -0.132 0.099 0.953 0.915 0.806 0.646 0.941 0.948 0.853 0.884 Colour Sponginess Regression co-efficients Fluffiness Independent variables Mean/Interaction 1. Rice : Dhal ratio (L) Rice : Dhal ratio (Q) 2. Fermentation time (L) Fermentation time (Q) 1L by 2L R2 L = linear effect; Q = quadratic effect; *= p < 0.005 Table 4.5 Regression co-efficient for overall quality of idli Independent variables Regression co-efficients Mean/Interaction 1. Rice : Dhal ratio (L) Rice : Dhal ratio (Q) 2. Fermentation time (L) Fermentation time (Q) 1L by 2L R2 L = linear effect; Q = quadratic effect; *= p < 0.005 79 -39.697 10.062 -4.263* 7.0723 -0.299 0.266 0.952 4.3.3 Overall quality of the idli Table 4.4 shows that the overall quality of the idli in sensory attribute was high (12.1) for the ratio 3:1.25 at 12 h fermentation time. Fig 4.3 illustrates the surface graph showing the relation between ratios of rice and black gram dhal and fermentation time on the overall quality of idli taking into consideration all sensory attributes of idli. From Table 4.5 it was known that the overall quality of the idli was at 5% level of significance with the change in ratio of rice to dhal ratio in quadratic effect. The R2 value for overall quality was 0.952. Fig.4.3 Response surface graph showing the overall quality of the idli 4.3.4 Simultaneous optimization Simultaneous optimization was performed for sensory attributes parameters like colour, appearance, texture, taste, aroma and overall quality by imposing desirability constraints. In case of sponginess, the softer idli shows high sponginess. Hence the software finally gives a maximum desirable score and the condition at which the maximum score can be 80 obtained with some constraints by assigning maximal desirability score as 1 and minimal desirability score as 0. Table 4.5 shows the constraints imposed for idli with better sensory attributes with the desirable value for both independent and dependant variables. The maximum desirable score that can be achieved with the desirable value will be 0.7439. On the basis of these calculations good idli could be made when the rice to black gram dhal ratio is 3:1.475 (w/w), fermented for 10.2 h. The optimum results were validated by performing the experiment at the optimized ratio and fermentation time by comparing the observed and the predicted values. The predicted values are shown in Table 4.5. The observed and predicted values were not significantly different (P >0.05) which confirmed the optimization results. Table 4.5 Simultaneous optimization of process parameters by desirability approach Independent Dependent variables parameters Overall Rice : dhal Fermentation Sensory Constraints Predicted Observed ratio time (h) parameters imposed values values Colour Maximum 08.00 08.20±0.64 Fluffiness Maximum 09.81 09.60±0.72 Sponginess Maximum 11.01 10.70±1.42 Fermented aroma Optimum 09.35 08.70±0.30 Compactness Optimum 09.35 08.80±0.39 Firmness Minimum 05.26 05.60±0.42 Stickiness Optimum 07.87 08.10±0.32 Sourness Optimum 07.60 07.30±0.28 Overall Quality Maximum 10.89 10.60±0.51 Desirability score (w/w) 3 : 1.475 10.2 0.7439 4.3.5 Principal Component Analysis (PCA) Sensory scores were subjected to PCA analysis. The PCA analysis revealed that PC1 and PC2 accounted for 78 percent of the total variance in the data matrix. It is clear from the plot that sensory attributes like sponginess and fluffiness associated with each other 81 strongly on the positive side of the PC1 axis while firmness, compactness, stickiness were clustered together on the negative side of the PC1 axis. The third cluster is formed by fermented aroma and sourness on the positive side of PC2 axis. Sample from the experimental design point 6 was closely associated with desirable sensory attributes like sponginess and fluffiness followed by sourness and fermented aroma. On the other hand, design points 5, 1 and 2 were closely correlated with undesirable sensory attributes like firmness, compactness and stickiness (Fig.4.4). From the PCA biplot it is clear that PCA Fig.4.4 Principal Component Analysis (PCA) biplot of experimental design points over sensory attributes of idli (refer Table 3.1 for design points) is a powerful technique which can discriminate the samples and attributes within the data matrix, depending upon their inter relationships. 4.3.6 Optimization of texture and sensory attributes The optimization results indicated that the optimum ratio of rice to black gram dhal is 3:1.575 (w/w), with 14 h of fermentation time will provide the product with maximum 82 score for desirable textural parameters. On sensory analysis followed by RSM analysis of idli prepared from various combinations of ingredients fermented at different durations up to 14 h, the rice and dhal combination of 3:1.475 fermented to 10.2 h was found to be the best accepted product. The results of both texture and sensory were combined to get an optimized result which gave idli with the best instrumental texture quality as well as sensory attributes. The impact making attributes of the idli chosen were shown in table 4.6. From the table it was found that the observed values and the predicted values showed no significant difference (p > 0.05) and the model fits the design. Table 4.6 Combined analysis of texture and sensory attributes Independent parameters Dependent variables Sensory / Texture Constraints Imposed Predicted Values Colour Maximum 10.11 10.60±0.36NS Sponginess Stickiness Sourness Overall quality Hardness Springiness Maximum Minimum Minimum Maximum Minimum Springiness 11.70 07.01 07.42 12.14 25.15 0.892 11.31±0.67 NS 07.40±0.41 NS 07.70±0.19 NS 11.85±0.86 NS 24.83±1.07 NS 00.91±0.06 NS Resilience Maximum NS = No Significant difference, (p> 0.05) 0.641 00.62±0.03 NS Rice:dhal (w/w) 3:1.18 Fermentation time (h) 12.02 Observed values Overall Desirability score 3.4 CONCLUSION On sensory analysis followed by RSM analysis of idli prepared from various combinations of ingredients fermented at duration up to 14 h, rice and dhal combination of 3:1.475 fermented to 10.2 h was found to be the best accepted product. On merging TPA and sensory results (Chapter 3 and 4) an optimized ratio of 3:1.18 and fermentation time of 12.02 h was evolved. 83 0.714 NUTRITIONAL COMPOSITION OF OPTIMIZED IDLI 5.1 INTRODUCTION Adequate nutrition through food is necessary for human life. Essential micro and macronutrients required for growth, metabolic regulations and physiological functions is provided by foods. The World Health Organization (WHO) recognizes the importance of breakfast in human diet. Nutritional value of idli has been reported in numerous publications; however, there are few studies on optimized idli. In this study the nutritional composition of optimized idli and its ingredients were studied without any starter cultures. Total carbohydrates and fats were broken down by natural fermentation process into oligosaccharides and fatty acids. Oligosaccharides have been known for its prebiotic activity thereby it enhances the probiotic flora in the human gut. 5.2 MATERIALS AND METHODS 5.2.1 Nutritional composition of the idli Nutritional composition of the raw ingredients (idli rice - IR 20, black gram variety ADT 3) and optimized idli were determined. Nutrients like starch, amylose, total carbohydrates, total sugars by Sadasivam and Manickam (2008), protein, fat and crude fibre (Cunniff, 1995) were estimated. 5.2.2 Determination of fatty acids and alcohols Fatty acids and alcohols were determined for the unfermented, fermented idli batter and idli prepared after optimized fermentation time. One gram of the sample was weighed and suspended into 10 ml of methylene chloride. The extraction of fatty acids and alcohols was done using the method followed by Agrawal et al., (2000). The extracted samples were analyzed for fatty acids and alcohols using Liquid Chromatography – Mass Spectrophotometer (LC-MS) (Thermo Finnigan Surveyor and Thermo LCQ Deca XP MAX). The experimental column used in LC-M was BDS HYPERSIL C18 and the volume of sample injected was 10µL. 84 5.2.3 Determination of oligosaccharides The determination of fatty acids and alcohols were done for the unfermented, fermented idli batter and idli prepared after optimized fermentation time. The extraction of oligosaccharides was performed as per Carlsson et al., (1992). The extracted samples were analyzed using LC-MS (Thermo Finnigan Surveyor and Thermo LCQ Deca XP MAX). The experimental column used in LC-MS was BDS HYPERSIL C18 and the volume of sample injected was 10µL. 5.3 RESULTS AND DISCUSSION The results are discussed under the following heads: 5.4.1 Nutritional composition of idli 5.4.2 Fatty acids and alcohols in optimized idli 5.4.3 Disaccharides and oligosaccharides in optimized idli 5.3.1 Nutritional composition of idli Proximate analysis of idli showed 81.60 g% total carbohydrates, in which starch was 75.0 g % (Amylose - 31.00, Amylopectin - 44.00). Protein content of rice and black gram were 6.46 g %, and 24.16, and optimized idli was found to be 10.21 g%. The fat and crude fibre concentrations in optimized idli were 00.10 ± 0.01 and 00.28 ± 0.01 g% respectively (Table 5.1). The carbohydrate level was comparatively high in optimized idli than protein, fat and crude fibre. Presence of amylose and amylopectin considerably influences rice starch digestion in the gastrointestinal tract, influencing faecal excretion and constitution, post- prandial blood glucose response and total cholesterol. Amylose content is normally used to evaluate some properties of product consumption such as cohesion and softness and also aid the control of biologically relevant parameters such as blood glucose and triglyceride concentration (Denardin et al, 2007). Amylose and amylopectin are fermented in the gastro-intestinal tract by 72% of the human colonic bacteroid strains (Salyers et al, 1977a, b). This study shows that the amylose and amylopectin in idli will help in the growth of gut microflora supporting the starch polysaccharide as a prebiotic. The good 85 amount of starch in idli is attributed by its raw ingredients. Starch were broken down to form reducing sugars and oligosaccharides which led to reduction in starch content in idli compared to the raw ingredients. Parameters (g % ± SD) Table 5.1 Proximate composition of optimized idli Rice Black gram (Variety IR 20) (Variety ADT3) Idli (3:1.18) Starch 79.50 ± 2.90 52.00 ± 1.21 75.00 ± 2.84 Amylose 32.00 ± 1.60 17.00 ± 0.74 31.00 ± 1.42 Amylopectin 47.50 ± 2.37 35.00 ± 1.62 44.00 ± 2.08 Total carbohydrates 84.00 ± 3.52 65.80 ± 3.02 81.60 ± 3.42 Protein 06.46 ± 0.32 24.16 ± 1.20 10.21 ± 0.50 Fat 00.27 ± 0.13 00.87 ± 0.02 00.10 ± 0.01 Crude fibre 00.20 ± 0.01 00.70 ± 0.03 00.28 ± 0.01 The protein content of idli prepared from combination of different starter culture namely Pediococcus pentosacens, Enterococcus faecium MTCC 5153, Ent. faecium IB2 with Candida versatilis were 3.3, 3.2 and 3.2 respectively (Sridevi et al, 2010). In a study done by Nazni and Shalini (2010) the protein content of the developed idli prepared from pearl millet was found to be 9.16g and the corresponding standard idli had 7.0 g whereas the protein content of the optimized idli in the current study was high (10.21 g %) which is found to be nutritionally rich even without addition of starter culture or millets. The major contribution of protein to the idli was attributed by the variety of black gram used when compared to rice. Decrease in fat content of idli was noted when compared to raw ingredients which may be due to the degradation of fats into fatty acids during fermentation process by microorganisms. 86 5.3.2 Fatty acids and alcohols in optimized idli Fatty acids and alcohols were analyzed in unfermented batter (Fig.5.1), Fermented batter (Fig.5.2), and idli (Fig.5.3) using LC-MS. Pentacontanoic acid, Hexadecanoic acid, Pentadecanoic acid, Nonadecanoic acid, Hexacosane, Nonacosane, Nonacosanol, 9Decanal, Decyl decanoate were present in unfermented batter, in which Relative Abundance (RA) of Pentacontanoic acid was maximum (38 %). In fermented batter Pentacontanoic acid, Decanoic acid, Octadecanoic acid, Hexadecanoic acid, Pentadecanoic acid, Nonadecanoic acid, Hexacosane, Nonacosane, Nonacosanol, 9Decanal, Decyl decanoate, Propanol and 2-Pentanone were present. Fatty acids and alcohol profile of idli was similar to fermented batter but Pentadecanoic acid and Nonacosanol were absent in final idli. The relative abundance (RA) of Fatty acids and alcohols in unfermented batter, fermented batter and idli is given in Table 5.2. Table 5.2 List of fatty acids and alcohols Unfermented batter Acids RA (%) Pentacontanoic acid 38 Hexadecanoic acid 11 Pentadecanoic acid 21 Nonadecanoic acid 19 Hexacosane 11 Nonacosane 12 Nonacosanol 18 9-Decanal 6 Decyl decanoate 8 - Fermented batter Acids RA (%) Pentacontanoic acid 23 Decanoic acid 12 Octadecanoic acid 10 Hexadecanoic acid 100 Pentadecanoic acid 8 Nonadecanoic acid 19 Hexacosane 11 Nonacosane 19 Nonacosanol 26 9-Decanal 8 Decyl decanoate 20 Propanol 10 2-Pentanone 10 87 Idli Acids Decanoic acid Octadecanoic acid Hexadecanoic acid Pentadecanoic acid Nonadecanoic acid Hexacosane Nonacosane 9-Decanal Decyl decanoate Propanol 2-Pentanone RA (%) 58 20 100 19 10 19 12 20 10 10 Fig. 5.1 Typical chromatogram and mass spectra showing fatty acids in unfermented batter 88 Fig. 5.2 Typical chromatogram and mass spectra showing fatty acids and alcohols in fermented batter 89 Fig. 5.3 Typical chromatogram and mass spectra showing fatty acids in optimized idli Mahadevappa and Raina (1978), reported that among the five varieties of legumes namely cow pea, field gram, red gram, horse gram and black gram, the major saturated fatty acid was palmitic acid which constitutes 15-25% in the neutral lipids, 20-40% in the glycolipids, and 26-30% in the phospholipids. It was also found that in the 90 two black gram (P. mungo T9 and Khargan 3) varieties, the fatty acid profile is characterized by exceptionally high levels of linolenic acid in all lipid classes: 60% in the neutral lipids, 50% in the glycolipids, and 33% in the phospholipids, accompanied in all categories by about 10% of linoleic acid and 15-20% of oleic acid. High levels of the two unsaturated essential fatty acids could have nutritional implications. In the present study, Hexadecanoic acid which is known as palmitic acid was found to be 100% in relative abundance in idli when compared to unfermented batter. Octadecanoic acid which is called as stearic acid was absent in unfermented batter but appear after fermentation in fermented batter and as well with a relative abundance of 10 – 20 %. Stearic acid is a long chain fatty acid consisting of 18 carbon atoms without double bonds. Ahrens et al. (1957), Keys et al., (1965), Hegsted et al. (1965 and 1993), and Yu et al (1995) found that saturated fatty acids with chain lengths more than 10 carbon atoms generally raised blood cholesterol levels whereas polyunsaturated fatty acids – PUFA (primarily linoleic acid) lowered blood cholesterol levels; and monounsaturated fatty acids had either a neutral or mildly hypo-cholesterolemic effect on blood cholesterol levels. These investigators also found that the stearic acid, a saturated fatty acid did not increase blood total or low density lipoprotein (LDL) cholesterol levels (bad cholesterol). Study by Yu et al. (1995), reported that adults were fed controlled, whole-food diets, to evaluate the effect of stearic acid on blood lipid levels revealed that stearic acid, showed no effect on LDL, and high density lipoprotein (HDL). A metaanalysis done by Mensink (2003) consisting of 35 controlled trials showed that when stearic acid replaced carbohydrate in the diet it had a neutral effect on blood lipid and lipoprotein levels. In foods like chocolate and lean red meats it was reported that their in-take does not increase the risk of cardiovascular disease because of high levels of stearic acid part in their saturated fatty acid. Similarly lean red meat (beef) and lean white meat (chicken, fish) are equally effective in reducing total and LDL cholesterol in adults fed lipidlowering diets (Ding et al., 2006, Davidson et al., 1999, Hunninghake et al., 2000, Scott et al., 1994 and Melanson et al., 2003) may be attributed in part to red meat‘s higher content of stearic acid compared to that in chicken or fish. The flavour characteristic known to be prevalent in idli batter appear from the combination of raw materials (rice 91 and black gram) and microbial starter cultures (natural inoculum) which helps in fermentation. Buttery et al., (1988) reported that ketones composed of ethanone, pentanone and butanones are found in rice. In the present study as the fermentation begins, it leads to the formation of 2-pentanone which is found to appear in fermented batter and not in unfermented batter. Polyunsaturated fatty acids play a primary role in the development of ketones and this source has been attributed to black gram dhal (Steinkraus et al. 1967). Microorganisms during fermentation lead to acidification of the raw material producing organic acids, mainly lactic acid. Also, their production of acetic acid, ethanol, aroma compounds, bacteriocin, exopolysaccharides and several enzymes improve shelf life, microbial safety, texture, and play a role increasing the pleasant sensory profile of the end product (Leroy and De Vuyst 2004). This finding supports the current study that the fermentation of idli batter helped in the formation of acetic acid, Dodecanol, Propanol and phenyl ethyl alcohol that improved the texture and sensory profile of the optimized idli. 5.4.3 Disaccharides and oligosaccharides in optimized idli Table 5.3 shows the list of disaccharides and oligosaccharides present in unfermented batter, fermented batter and in idli. Trehalose, maltose, melezitose, maltotriose, maltotetrose, maltopentose and maltohexose that were absent in unfermented batter appeared in fermented batter and idli which is due to the breakdown of polysaccharides into oligosaccharides and disaccharides during the process of fermentation. Typical chromatograms and mass spectra showing disaccharides and oligosaccharides for unfermented batter (Fig.5.4), fermented batter (Fig.5.5) and idli (Fig.5.6) are shown below. Crittenden & Playne, (1996) proved in their study that oligosaccharides in doses of <15 g/day increase bifidobacteria numbers in the colon. Authors suggest that a daily intake of 10 g of galacto-oligosaccharides is sufficient to cause a bifidogenic effect. Seeds of legumes, lentils, and mustard are rich source of raffinose oligosaccharides (Johansen et al., 1996; Sánchez-Mata et al., 1998). Hate et al (1983) reported that clinical data of Japanese researchers suggest a regular addition of fructooligosaccharides to diet lowers 92 total cholesterol and triglyceride in blood which is proved by Losada and Olleros (2002) that fructooligosaccharides increase production of volatile fatty acids by the action of gut microflora on oligosaccharides have effect on cholesterol in the liver. Table 5.3 List of Disaccharides and Oligosaccharides Compound Saccharose Trehalose Maltose Melezitose Raffinose Maltotriose Stachyose Maltotetrose Verbascose Maltopentose Maltohexose Non-Fermented batter + + + + - Fermented batter + + + + + + + + + + + Idli + + + + + + + + + + - Cereal grains consist of at least two types of oligosaccharides such as galactosyl derivatives and fructosyl derivatives. Galactosyl derivatives include sucrose, stachyose and raffinose and fructosyl derivatives include sucrose and fructo-oligosaccharides (Henry and Saini, 1989). Table 5.3 shows that the presence of raffinose in unfermented, fermented batter and idli was attributed by the raw ingredient black gram dhal. This finding was supported by the Voragen (1998) who indicated that raffinose and stachyose in soya bean and other pulses and leguminous seeds are examples of naturally occurring non-digestible oligosaccharides. 93 Fig. 5.4 Typical chromatogram and mass spectra showing Disaccharides and oligosaccharides in unfermented batter 94 Fig. 5.5 Typical chromatogram and mass spectra showing Disaccharides and oligosaccharides in fermented batter 95 Fig. 5.5 Typical chromatogram and mass spectra showing Disaccharides and oligosaccharides in optimized idli 96 Results showed that there was reduction in the abundance of disaccharides and oligosaccharides namely saccharose, trehalose, melezitose, maltotriose, maltopentose and raffinose in idli compared to fermented batter. Onigbinde and Akinyele (1983) also had proposed that decrease in the levels of raffinose, stachyose, and verbascose during cooking might be attributed to heat hydrolysis to disaccharides and monosaccharides or to the formation of other compounds. Basha (1992) showed that the rate of oligosaccharide breakdown increased with increasing acid concentration. This rope with the current study which shows that steaming of fermented batter to get idli may decrease the level of abundance of oligosaccharides in idli when compared to fermented batter. The acidic medium is caused by the action of microorganisms during fermentation process (Sridevi et al, 2010). The isomers of trehalose showed an increase in bifidobacteria population similar to fructooligosaccharides giving high prebiotic index in the in-vitro fermentation (Sanz et al, 2005). Carbohydrates can also act as prebiotics for selected bacterial group within the gut reducing the pathogen population by increasing immunity (Gibson, 1998). Presence of disaccharides and oligosaccharides in idli will possibly help to improve the health by improving the gut microflora. 5.4 CONCLUSION The fatty acids namely Decanoic acid, Octadecanoic acid and Hexadecanoic acid were of high relative abundance in idli compared to unfermented batter. Regarding the oligosaccharide profile, the sugars namely trehalose, maltose, melezitose, maltotriose, maltotetrose and maltopentose were formed during fermentation. The process of fermentation has led to the increase in nutrient content of the idli. 97 IMPROVING THE SHELF LIFE OF READY TO COOK IDLI BATTER 6.1 INTRODUCTION Ready to cook idli batter which was optimized to give desired quality parameters in the earlier chapters (3 and 4) could only be successfully commercialized as a viable product only its shelf-life is increased substantially from approximately one day to at least several days.Idli, unlike other ready to cook food products rapidly gets over fermented as it is a live actively growing bacterial medium, although common spoilage problems are less. This is a challenge in terms of its preservation and shelf life. Ready to cook idli batter is already available in the market as a packaged product. For several years prepared by local vendors as a perishable product sold on daily basis or stored and sold under refrigerated conditions. For the first time we have hypothesized that modification of gaseous environment in the packaged form could regulate the fermentation flora in the medium leading to longer shelf life. Second part of the hypothesis is that regulating the gaseous exchange with the external environment could support the modified atmosphere in the packaged product to work long. MAP helps to preserve foods by reducing microbial spoilage thereby increasing storability. MAP is done to maintain the freshness of the produce when purchased. Success of MAP packaged foods depends on the quality of raw material and hygienic practices followed during preparation and packaging, the gas mixture used for packaging and the packaging material. The gases used in MAP are CO2, O2 and N2. Researchers have successfully applied MAP to perishable foods like fruits, vegetables, flesh foods and certain dairy products. 6.2 MATERIALS AND METHODS 6.2.1 Materials 1. Modified Atmosphere Packaging machine (VAC Star-Swiss) 2. Head space analyzer (Dansensor, Italy) 3. Gas mixer (Dansensor, Italy) 4. Packaging materials- LDPE, PP, HM 98 6.2.2 Methods 6.2.2.1 Preparation of batter The selected IR20 idli rice and ADT3 variety black gram dhal were taken in the optimized ratio of 3:1.18. The quantity required varied for each set of experiment. Ingredients were soaked and ground and were immediately packaged in the packaging materials Fig.6.1 shows the research design of this chapter. 6.2.2.2 Selection of packaging materials Three packaging materials namely low density poly ethylene (LDPE), Poly propylene (PP), High Molecular (HM) were used for packing the idli batter. The thickness of the packaging materials is given in Table 6.1. Dimension of the packaging material was 6×10 inches. 100g of the batter was filled in each pack. Table 6.1 Thickness of packaging materials Packaging materials Thickness (mm) LDPE 0.009 LDPE 0.012 LDPE 0.014 PP 0.003 PP 0.005 HM 0.002 HM 0.006 6.2.2.3 MAP of idli batter MAP was done using Modified Atmosphere Packaging machine (VAC Star-Swiss). MAP machine consists of three gas cylinders viz., oxygen, carbon dioxide and nitrogen, each of which is connected to a gas mixer provided with a separate cylinder where the required combination of gases can be set and stored temporarily in buffer tank. Gas analyzer is another important component of the MAP machine which helps to check if the gas is mixed in the expected combination and the same is used for determining gas in the head 99 space of the packaged sample as and when required. Plate 6.1 shows the MAP machine. The batter in each packet was packaged with modified air of required combination. Plate 6.1 Modified Atmosphere Packaging (MAP) machine 6.2.3 Respiration dynamics of the idli batter Respiration dynamics was carried out to find the percentage of oxygen utilized and percentage of carbon dioxide released during fermentation of batter. Respirometer as designed by Bosco (1997) was used. Plate 6.2 shows the picture of respiration dynamics done for the idli batter using respirometer connected to gas analyzer. The respirometer consists of a glass jar without spout of capacity 250 mL resting on a flat MS plate and covered with another MS plate. Both the plates had hole at each corner through which bolt were inserted. By tightening the nuts of these bolts, the glass jar could be closed with the cover plate. The joint between the glass jar and the cover plate was made air tight by providing a neoprene rubber gasket. The cover plate had one hole at the centre where the gas septum had been fixed for sampling the gas. Gas tightness of Respirometer was verified by the respirometer ability to hold 50 mm vacuum for 15 minutes as done by 100 Brown (1922). For the experiment, 100g of fresh ground batter was taken in 250 ml beaker. Atmospheric air was maintained inside the beaker. The Respirometer was connected to the gas analyzer to monitor the change in gas environment every half an hour. The experiment was conducted for 12.02h (optimized fermentation time). Plate 6.2 Respirometer connected to gas analyzer 101 6.2.4 EXPERIMENT I Experiment I was carried out to find suitable packaging material for idli batter. The study was designed for 3 days (Sridevi et al., 2010), hence a total of 126 packets (7 packaging materials × 6 gas treatments × 3 days) were required. Based on the optimized ratio, batter was prepared. In each packaging material 100g of batter was packaged with gas treatment of 0% CO2, 5% CO2, 10% CO2, 15% CO2 and vacuum packaging and sealed. The package in which no treatment was done served as the control. The packaged batters were stored in room temperature (30 C). Each day 42 packets representing 7 packaging material and 5 gas treatments were analyzed for the gas mixture using gas analyzer. 6.2.5 EXPERIMENT II In experiment II, the idli batter was packaged and sealed with 12 gas treatments and three controls were used. Batter placed in vessel served as control I, batter packaged and sealed with ordinary sealing machine served as control II, the batter packaged in packaging material but not sealed served as control III. The gas treatments are shown in Table 6.2. The study was done using selected 3 packaging material. Hence a total of 215 samples were required. The MA packaged batter and control packs were stored in room temperature (30 C). Each day 43 samples stored in three different packaging materials with 12 gas treatments including 3 controls were analyzed for the gas concentrations (%) followed by sensory analysis of the idli cooked from packaged batter. Overall quality based on the colour, texture, fermented aroma of the idli was assessed. 102 Table 6.2 Gas treatment used in experiment II Treatments 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 6.2.6 CO2 (%) 00 05 10 15 00 05 10 15 00 05 10 15 Control I Control II Control III O2 (%) N2 (%) 15.0 85.0 15.0 80.0 15.0 75.0 15.0 70.0 17.5 82.5 17.5 77.5 17.5 72.5 17.5 67.5 20.0 80.0 20.0 75.0 20.0 70.0 20.0 65.0 Batter in vessel Batter in unsealed package Batter packaged with ambient air EXPERIMENT III In experiment III, selected one packaging material and 8 gas treatments and control were used. Gas treatments are shown in Table 6.3. In this experiment the batter packages were left to check the extended shelf life of the batter. Table 6.3 Gas treatment used in experiment III Treatments CO2 (%) O2 (%) N2 (%) 1 0 15.0 85.0 2 0 12.5 87.5 3 0 10.0 90.0 4 0 07.5 92.5 5 5 15.0 80.0 6 5 12.5 82.5 7 5 10.0 85.0 8 5 07.5 87.5 103 6.3 RESULTS AND DISCUSSION 6.3.1 Respiration dynamics Respiration is usually the measure oxygen uptake or the production of CO2, producing heat and water vapour. The results of respiration dynamics give an idea to flush the package with required atmosphere so that steady state conditions are reached immediately (Zagory and Kader, 1988). The O2 consumption and CO2 evolution differ based on the composition such as fatty acids, sugars or organic acids of the respiring sample (Dilley et al, 1990 and Platenius, 1942). Fig.6.1 Change in gas concentration during its fermentation time Fig.6.1 showed the O2% consumed and CO2 % evolved during the fermentation of idli batter. As fermentation began, the O2% declined from 21% to 13.9%. The increase in CO2% began after 2 h and gradually increased from 0% to 12.9%. The results showed that 100g of idli batter consumed 7.1% O2 and produced 12.9% of carbon dioxide. Based on the results of respiration dynamics MAP of ready to cook idli batter was done with different gas treatments and the results are discussed below. 104 6.3.2 EXPERIMENT I Table 6.4 shows the changes in atmosphere of the packaged idli batter. In treatment1 where the package was packed with lack of oxygen and carbon dioxide (0% O2 and 0% CO2) showed a gradual increase in CO2 over three days of storage. This increase in CO2 is a result of fermentation of idli batter inside the package. The O2 (%) remained zero in the LDPE packages but HM and PP showed increase in O2, which meant that the packaging material permitted permeability of air. Fig.6.2 shows the changes in gas mixture among different packaging material over three days of storage. In treatment 2 where the package was flushed with 5% CO2, it was found that there was increase in CO2 % from 5% to a maximum of 14.4 % CO2 in LDPE packaging material during the second day of storage. In the packaging material PP (0.003mm) there was decrease in CO2 to 0.8% on the third day. The O2 concentration in PP (0.003mm) and HM (0.002 mm) were not maintained inside the package. Fig.6.3 shows the changes in gas mixture among different packaging material over three days of storage. In treatment 3 where the 10% CO2 was flushed in the package, LDPE of higher thickness showed fermentation effect on the batter with increase in CO2, whereas in LDPE of lower thickness, PP and HM the change in gas system was not gradual. In treatment 4, the package was flushed with 15% CO2, increased in LDPE of medium and lower thickness and the concentration of CO2 varied in other packaging materials over the storage period. Fig.6.4 shows the changes in gas mixture among different packaging materials. Treatment 5 was vacuum packaging and over the storage period the gas concentrations were not analyzed as the pressure was too low to detect the gas in the head space. Treatment 6 was control with lack of gas treatments but package had ambient gas composition. The atmospheric air in package favoured fermentation of batter which led to decrease in O2 concentration. From the results of experiment I it was inferred that gas permeability differed with different packaging material. The O2 permeability was less than that of CO2 in LDPE of varying thickness helped to maintain the atmosphere within the package compared to other packaging materials. This result is supported by the study done by Bosco (1997), 105 who reported that, the O2 permeability of LDPE and PP was less than that of CO2 and the variation in permeability of a film is due to the fact that the film were purchased from the retail market at different places and might be from different batches of production Table 6.4 Change in gas mixture over storage period Packaging material LDPE (0.14 mm) Treatment 1 Treatment 2 Treatment 3 Treatment 4 Treatment 5 Treatment 6 LDPE (0.12 mm) Treatment 1 Treatment 2 Treatment 3 Treatment 4 Treatment 5 Treatment 6 LDPE (0.009 mm) Treatment 1 Treatment 2 Treatment 3 Treatment 4 Treatment 5 Treatment 6 PP (Thin) Treatment 1 Treatment 2 Treatment 3 Treatment 4 Treatment 5 Treatment 6 Day 1 CO2 (%) O2 (%) Day 2 Day 3 CO2 (%) O2 (%) CO2 (%) O2 (%) 0.114 5.6 10.5 14.2 0 0 0.009 0.1 0.008 21 11 6.3 13.8 12.4 2.1 0 1.07 0 0 14.8 11.1 10.9 15.5 8.9 4.2 0.017 0.009 0 19.4 7.15 0.114 5.6 10.5 14.2 0 0 0.009 0.1 0.008 21 10.3 10.6 12.6 14.9 4.1 0 5.4 0 0.011 7.56 10 10.8 15.4 14.9 3.1 0.265 5.2 0.09 0 12.6 0.114 5.6 10.5 14.2 0 0 0.009 0.1 0.008 21 10.3 14.4 12.9 10.5 3.3 0 0.484 0 0.095 8.91 12.8 15 5.2 12.6 0.3 0 1.2 0.036 0.015 19.8 0.114 5.6 10.5 14.2 0 0 0.009 0.1 0.008 21 9.9 13.5 11.3 13.4 0 0.112 19.5 19.1 0.908 20.3 12.7 0.8 1.2 15.2 0.3 0.112 19.5 19.1 0.908 19.8 106 PP (Thick) Treatment 1 Treatment 2 Treatment 3 Treatment 4 Treatment 5 Treatment 6 HM (Thin) Treatment 1 Treatment 2 Treatment 3 Treatment 4 Treatment 5 Treatment 6 HM (Thick) Treatment 1 Treatment 2 Treatment 3 Treatment 4 Treatment 5 Treatment 6 0.114 5.6 10.5 14.2 0 0 0.009 0.1 0.008 21 5.1 10.8 14.3 15.2 8.2 0.549 0.001 0.032 0.001 6.2 10.2 15 15 14.6 0 0.539 0.001 0 0.149 0.3 0.114 5.6 10.5 14.2 0 0 0.009 0.1 0.008 21 3.4 2.8 0 0.7 0.4 3.18 0.04 20.4 19.6 19.5 11.6 5.9 0.8 1 0 9.5 19.2 18.9 19.3 20.3 0.114 5.6 10.5 14.2 0 0 0.009 0.1 0.008 21 3.4 6 0.5 0.1 0.3 4 0.5 19.3 20.3 19.7 19.7 7.3 2.8 4.1 0.1 9.7 0.6 17.1 9.39 20.2 At low oxygen levels, anaerobic respiration can occur, resulting in production of substances that contribute to off-flavours and odours (Lee et al, 1995 and Zagory 1995). Hence the idli prepared from the batter were subjected to sensory analysis only for its texture. Idli made from the batter packaged in different packaging material scored very low rating which might be due to deterioration of the batter. The experiment was repeated with combination of both O2 and CO2 in LDPE of varying thickness. 107 Fig. 6.2.a Treatment 1 (0% CO2) showing change in CO2 level (%) among different packaging material Fig. 6.2.b Treatment 1 (0% CO2) showing change in O2 level (%) among different packaging material 108 Fig. 6.3.a Treatment 2 (5% CO2) showing change in CO2 level (%) among different packaging material Fig. 6.3.b Treatment 2 (5% CO2) showing change in O2 level (%) among different packaging material 109 Fig. 6.4.a Treatment 4 (15% CO2) showing change in CO2 level (%) among different packaging material Fig. 6.4.b Treatment 4 (15% CO2) showing change in O2 level (%) among different packaging material 110 6.3.3 Experiment II Table 6.5 to 6.8 shows the changes in gas concentrations in the packaging materials with idli batter. The 12 treatments showed the rate of fermentation of batter in different MAP system in different packaging material at room temperature (30 C). Treatment 1 to 12 showed increase in concentration of CO2% within 48h of packaging which is due to the evolution of CO2 during fermentation of the batter. Treatment 1 showed gradual increase in fermentation rate of the batter for five days of storage period compared to other treatments. The percentage of O2 decreased gradually from 15% to 1.75% (LDPE 0.12mm). Fig.6.5 showed the changes in concentration of CO2 and O2 among LDPE of varying thickness. The batter in treatments 5 to 12 which were flushed with 17.5% to 20% O2 led to complete consumption of O2 in LDPE of 0.014 and 0.012mm by the batter supporting fermentation and also whey separation. During storage of batter whey separation persisted (Nisha et al, 2005). The reason stated by Nisha et al was that idli batter is foam in which gas molecules are entrapped in a solidliquid phase. The batter collapse and whey separates when the high energy interface takes place during air-water interface. Fig.6.5 to 6.14 shows the depletion of O2 curve. LDPE of 0.009mm did not support the MAP system showing variations in gas concentration over the period of batter storage. The gas combination was not analyzed for control I and II as they were exposed to the atmospheric air. The O2 (%) concentration in control III was zero per cent and that of CO2 decreased from 11% (2nd day) to 5.7 % (5th day) during storage. It showed that the oxygen was consumed for the fermentation process and the carbon dioxide evolved was decreased due to its permeability through the packaging material. 111 Table 6.5 Concentration of gases in LDPE (0.014mm) during the storage period Treatments 1 2 3 4 5 6 7 8 9 10 11 12 Day 0 CO2 (%) O2 (%) 00 15.0 05 15.0 10 15.0 15 15.0 00 17.5 05 17.5 10 17.5 15 17.5 00 20.0 05 20.0 10 20.0 15 20.0 Gas combinations Day 1 Day 2 Day 3 CO2 (%) O2 (%) CO2 (%) O2 (%) CO2 (%) O2 (%) 08.1 9.40 10.4 02.2 00 0.00 07.0 7.31 03.3 13.6 6.8 0.44 05.4 11.2 02.6 14.7 1.2 18.8 02.5 16.7 05.4 0.707 4.4 4.09 04.3 6.38 08.0 0.087 6.9 0.94 09.6 0.72 04.0 04.59 00 0.00 12.2 0.23 13.7 0.077 6.1 0.40 10.1 1.26 05.3 0.730 6.1 0.19 22.1 0.001 08.1 0.024 6.3 0.16 16.7 0.12 08.1 0.045 4.9 0.09 16.5 4.11 10.3 0.205 8.5 1.13 14.9 3.03 07.1 02.06 7.5 0.14 112 Day 4 CO2 (%) O2 (%) 6.2 1.500 5.9 0.453 8.3 2.740 4.7 0.424 4.3 2.660 5.9 0.085 6.6 0.000 4.6 0.172 5.4 0.076 4.9 0.134 2.8 9.460 6.0 0.361 Day 5 CO2 (%) O2 (%) 6.2 0.000 6.2 7.000 6.0 3.100 5.5 0.600 9.6 0.850 6.4 0.002 6.1 0.800 5.7 0.70 5.8 0.137 5.0 0.142 7.1 0.873 5.8 0.197 Table 6.6 Concentration of gases in LDPE (0.012mm) during the storage period Treatments 1 2 3 4 5 6 7 8 9 10 11 12 Day 0 CO2 (%) O2 (%) 00 15.0 05 15.0 10 15.0 15 15.0 00 17.5 05 17.5 10 17.5 15 17.5 00 20.0 05 20.0 10 20.0 15 20.0 Gas combinations Day 1 Day 2 Day 3 CO2 (%) O2 (%) CO2 (%) O2 (%) CO2 (%) O2 (%) 07.2 07.05 09.3 3.524 10.1 2.650 07.6 04.72 03.0 9.780 05.4 0.601 11.0 07.85 09.6 0.759 08.2 0.534 05.7 12.40 05.5 0.486 04.8 0.291 12.4 00.54 08.3 0.206 00.4 20.10 15.4 02.10 08.7 1.450 08.1 0.622 06.9 03.79 06.2 0.246 08.8 8.470 16.2 0.268 08.8 0.071 07.2 0.453 19.3 0.161 05.7 0.164 05.8 0.017 05.3 14.50 15.7 0.000 07.0 0.000 05.0 13.60 08.0 5.150 04.3 0.238 15.2 03.14 06.6 8.910 09.3 0.152 113 Day 4 CO2 (%) O2 (%) 11.7 1.95 05.7 0.71 08.2 3.16 03.9 0.26 08.4 0.113 07.2 0.044 05.4 0.197 04.9 0.194 05.4 0.367 05.1 0.161 07.5 0.252 04.9 0.185 Day 5 CO2 (%) O2 (%) 11.9 01.75 04.0 00.29 05.5 00.09 00.0 00.00 05.8 11.10 00.2 20.80 04.9 00.22 09.2 01.42 05.8 00.03 07.2 00.36 06.6 00.37 09.1 00.24 Table 6.7 Concentration of gases in LDPE (0.009mm) during the storage period Treatments 1 2 3 4 5 6 7 8 9 10 11 12 Day 0 CO2 (%) O2 (%) 00 15.0 05 15.0 10 15.0 15 15.0 00 17.5 05 17.5 10 17.5 15 17.5 00 20.0 05 20.0 10 20.0 15 20.0 Day 1 CO2 (%) O2 (%) 10.2 09.45 01.1 18.90 08.3 05.49 16.5 00.43 17.3 00.24 18.7 00.31 07.8 05.29 13.5 00.69 10.9 01.26 11.2 06.71 17.9 00.12 10.0 00.27 Gas combinations Day 2 Day 3 CO2 (%) O2 (%) CO2 (%) O2 (%) 09.0 0.08 1.7 18.1 08.9 2.54 4.9 0.12 04.0 1.87 6.0 6.61 05.4 9.69 0.2 20.5 09.3 0.16 1.0 20.2 06.2 0.42 7.8 1.21 04.5 1.96 8.9 1.03 08.9 0.06 5.3 11.3 11.3 0.91 4.6 0.19 12.5 1.22 5.9 0.17 09.5 0.29 10.4 0.65 05.4 0.77 3.8 2.37 114 Day 4 CO2 (%) O2 (%) 7.2 08.69 8.9 00.79 4.8 00.34 9.3 03.35 5.4 00.20 2.4 18.40 5.4 11.90 14 01.61 10.8 00.04 7.3 00.52 8.0 00.04 11.2 02.19 Day 5 CO2 (%) O2 (%) 05.6 00.16 09.3 00.12 07.7 00.09 04.6 014.3 06.6 00.29 08.5 01.79 06.1 06.20 10.0 00.39 05.7 00.09 10.5 01.65 04.4 13.20 07.7 00.09 Storage period Fig 6.5.a Treatment 1 (0% CO2 and 15% O2) showing percentage of CO2 (%) Storage period Fig.6.5. b Treatment 1 (0% CO2 and 15% O2) showing percentage of O2 (%) 115 Storage period Fig 6.6.a Treatment 2 (5% CO2 and 15% O2) showing percentage of CO2 (%) Storage period Fig 6.6.b Treatment 2 (5% CO2 and 15% O2) showing percentage of O2 (%) 116 Storage period Fig. 6.7.a Treatment 3 (10% CO2 and 15% O2) showing percentage of CO2 (%) Storage period Fig. 6.7.b Treatment 3 (10% CO2 and 15% O2) showing percentage of O2 (%) 117 Storage period Fig. 6.8.a Treatment 4 (15% CO2 and 15% O2) showing percentage of CO2 (%) Storage period Fig. 6.8.b Treatment 4 (15% CO2 and 15% O2) showing percentage of O2 (%) 118 Storage period Fig. 6.9.a Treatment 5 (0% CO2 and 17.5% O2) showing percentage of C O2 (%) Storage period Fig. 6.9.b Treatment 5 (0% CO2 and 17.5% O2) showing percentage of O2 (%) 119 Storage period Fig. 6.10.a Treatment 8 (15% CO2 and 17.5% O2) showing percentage of CO2 (%) Storage period Fig. 6.10.b Treatment 8 (15% CO2 and 17.5% O2) showing percentage of O2 (%) 120 Storage period Fig. 6.11.a Treatment 9 (0% CO2 and 20% O2) showing percentage of CO2 (%) Storage period Fig. 6.11.b Treatment 9 (0% CO2 and 20% O2) showing percentage of O2 (%) 121 Storage period Fig. 6.12.a Treatment 10 (5% CO2 and 20% O2) showing percentage of CO2 (%) Storage period Fig. 6.12.b Treatment 10 (5% CO2 and 20% O2) showing percentage of O2 (%) 122 Storage period Fig. 6.13.a Treatment 11 (10% CO2 and 20% O2) showing percentage of CO2 (%) Storage period Fig. 6.13.b Treatment 11 (10% CO2 and 20% O2) showing percentage of O2 (%) 123 Storage period Fig. 6.14.a Treatment 12 (15% CO2 and 20% O2) showing percentage of CO2 (%) Storage period Fig. 6.14.b Treatment 12 (15% CO2 and 20% O2) showing percentage of O2 (%) 124 Table 6.8 Sensory scores of the product made from batter packaged in LDPE (0.014mm) Treatments 1 2 3 4 5 6 7 8 9 10 11 12 Control I Control II Control III Day 1 Sensory scores Day 2 Day 3 Day 4 Day 5 8.1 7.5 5.2 9.3 8.5 8.5 8.1 6.4 8.8 8.3 8.1 8.0 6.7 8.0 8.2 9.1 8.5 8.5 8.3 7.3 7.7 5.7 7.2 7.7 8.2 8.0 8.0 5.2 7.4 1 5.2 1 5.4 1 1 1 1 1 3.5 3.7 3.0 - 7.1 5.1 1 1 7.2 6.2 1 1 6.1 7.3 4.3 7.3 - 1 5.1 1 1 7.7 7.0 1 1 1 7.1 1 1 - Table 6.9 Sensory scores of the product made from batter packaged in LDPE (0.012mm) Treatments 1 2 3 4 5 6 7 8 9 10 11 12 Control I Control II Control III Day 1 Sensory scores Day 2 Day 3 Day 4 Day 5 3.0 4.0 6.2 6.3 6.1 6.3 8.2 6.3 8.2 8.5 8.5 8.2 6.8 7.4 7.5 7.3 7.1 7.4 7.2 6.5 8.3 8.5 7.2 6.3 7.3 4.2 6.0 4.7 6.2 9.0 5.0 6.2 5.1 5.7 7.3 5.1 6.3 6.2 6.1 6.6 5.3 - 8.2 5.4 7.9 6.4 5.2 8.9 6.8 7.7 6.4 5.9 5.4 4.0 - Table 6.10 125 8.3 5.6 6.9 6.8 6.8 8.5 6.4 5.9 5.2 6.0 6.7 5.2 - Sensory scores of the product made from batter packaged in LDPE (0.009mm) Treatments 1 2 3 4 5 6 7 8 9 10 11 12 Control I Control II Control III Day 1 Sensory scores Day 2 Day 3 Day 4 Day 5 7.5 8.1 5.4 8.2 8.0 8.3 8.5 8.2 5.3 7.8 5.2 5.0 6.8 7.3 7.1 6.4 5.3 5.9 4.5 6.1 6.2 6.3 8.1 8.3 7.4 8.1 6.3 5.0 6.8 3.8 3.7 2.7 4.3 2.7 1.6 2.7 1.8 3.2 2.9 3.2 1.6 - 1.5 1.5 1.7 1.3 1.4 4.0 6.2 1.7 6.5 5.7 5.3 5.2 - 1.6 1.8 1.9 1.4 1.7 1.6 1.5 1.8 5.2 4.3 2.5 2.0 - Table 6.8 to 6.10 showed the sensory scores of idli made from batter treated with different gas combinations in different packaging material. The sensory score represented the overall quality of the idli. Sensory scores of idli prepared on the second day of storage showed high acceptability which might be due to the reason that the batter had been fermented and gave idli of high acceptability. The scores ranged from 4.2 to 9.1 (treatment 11 in LDPE -0.012 mm and treatment 1 in LDPE- 0.014 mm). The sensory scores on the third, fourth and fifth day showed poor acceptability of the product except for treatments in LDPE 0.012 which increased during the storage period. The highest score obtained was 9 for treatment 1 (Fig.6.15) followed by 7.3 for treatment 6 (Fig.6.16) on the fifth day of storage. The batter in control I was discarded due to over fermentation after 24 h followed by fungal contamination. The batter in control samples were evaluated for sensory for a maximum two days whereas the batter in packages was used for idli preparation and evaluated for sensory on all storage days in spite of its poor scores. Thou there was whey separation which made idli harder, the packaged batter used for idli preparation when compared to control. 126 The results of experiment II discussed from gas concentration in different treatments and different packaging material showed that LDPE of medium thickness (0.012 mm) may support to maintain the MAP system inside the packaged atmosphere when compared to LDPE of other thickness . The gas treatment of 0% CO2 with 15% O2 and 5% CO2 with 15% O2 were found to extend the shelf-life of the batter compared to other gas treated samples and control samples. Fig. 6.15 Comparison of sensory scores of idli made from treatment 1 (0% CO2 and 15% O2) 127 Fig. 6.16 Comparison of sensory scores of idli made from treatment 6 (5% CO2 and 15% O2) 6.3.4 Experiment III The observations of experiment III are shown in Table 6.11. It was found that the percentage of carbon dioxide increased in all treatments except for 8 which showed very low CO2% and low O2%. The consumption of O2 percentage was high showing complete decrease of O2 percentage in the package. During fermentation of batter oxygen is consumed and when the fall is below 1% may lead to anaerobic respiration (Lee et al, 1995 and Zagory 1995). In the table 6.11, on the seventh day of storage, all treatments showed poor concentrations of O2% except treatment 1 which showed 1.4% O2. Initial pH of the fresh batter 6.41 and change in pH of the batter in different treatments over seven days of storage showed decrease in pH. The change in pH is associated with the development of Streptococcus faecalis producing both lactic acid, which lowers the pH and carbon dioxide which leavens the batter (Balasubramanian and Viswanathan, 2007a). The control sample was discarded on the third day due to fungal contamination. 128 Table 6.11 Comparison of gas mixture on the first day and seventh day of storage Treatments Day 0 Day 7 CO2 (%) O2 (%) CO (%) O 2 2 (%) pH pH 1 0 15.0 6.41 11.3 1.4 4.32 2 0 12.5 6.41 12.6 0.4 4.10 3 0 10.0 6.41 10.3 0.7 4.31 4 0 07.5 6.41 12.4 0.9 4.31 5 5 15.0 6.41 12.9 0.0 4.28 6 5 12.5 6.41 14.6 0.5 4.00 7 5 10.0 6.41 13.1 0.6 4.10 8 5 07.5 6.41 3.4 0.0 4.22 Control Ambient atmosphere 6.41 - Table 6.12 TPA parameters of idli made from MAP batter Treatments Hardness (N) 1 17.020±4.68 2 Springiness Cohesiveness -07.257±5.58 0.874±0.07 0.671±0.02 1024.01±333.8 0.386±0.02 21.455±1.22 -09.924±5.52 0.814±0.05 0.622±0.04 1106.65±95.3 0.337±0.04 3 21.705±2.72 -09.907±3.39 0.829±0.02 0.645±0.00 1181.11±116.7 0.364±0.02 4 19.500±2.63 -15.461±13.27 0.852±0.11 0.663±0.09 1112.96±155.5S 0.369±0.06 5 27.315±2.55 -10.757±05.48 0.870±0.01 0.686±0.07 1045.67±35.7 0.383±0.03 6 28.955±0.45 -12.552±02.95 0.843±0.04 0.624±0.05 1229.40±131.8 0.356±0.05 7 28.571±0.21 -09.417±06.58 0.858±0.05 0.666±0.03 1339.51±34.7 0.377±0.03 8 27.152±0.52 -14.790±00.09 0.904±0.06 0.720±0.00 1761.41±9.2 0.336±0.01 Control - Adhesiveness (N s) - - - Chewiness - The texture of idli prepared from the treated batter was analyzed and Table 6.12 shows the TPA values of the idli. The hardness of the idli ranged between a minimum of 17.02 N (treatment 1) to a maximum hardness of 28.95 N (treatment 6). The hardness of idli was low for the treatments 1 to 4 when compared to other treatments which were due to the whey separation seen in treatment 5 to 8. The maximum springiness was found for the idli made from the treatment 8 followed by idli made from treatment 1 and 5. 129 Resilience - Cohesiveness was maximum for the idli made from the treatment 8 followed by 2 and 6. Resilience was maximum for the idli made of treatment 1 (0.386) followed by treatment 7 (0.377). The texture profile values when compared with the optimized value show that idli made from treatments 1 to 4 were soft compared to idli made from other treatments. Treatments Table 6.13 Overall quality of idli CO2 (%) O2 (%) Overall quality 1 0 15 8.6 ± 0.84 2 0 12.5 7.0 ± 0.00 3 0 10 7.0 ± 1.34 4 0 7.5 7.4 ± 2.4 5 5 15 5.8 ± 0.49 6 5 12.5 6.4 ± 0.49 7 5 10 6.6 ± 0.70 8 Control 5 7.5 Batter packaged with ambient air 4.0 ± 5.65 - Table 6.13 shows the scores of overall quality of the idli made from MAP treated batter. The maximum score was 8.6 (Treatment 1) followed by 7.4 (Treatment 4). The overall comparison of the sensory scores show that treatments 1 (8.6), 2 (7.0) and 4 (7.4) had high scores respectively when compared to treatments 5 to 8. Studies done by Day (1996) and Zagory and Kader, (1988) showed that by modifying the atmospheric oxygen level, particularly by lowering the oxygen concentration inside the package, the respiration rate of the packaged produce is slowed down and the sensory shelf life can be extended which cannot be applied to the current study. Idli batter being a live product which produces 12.9% of CO2 during fermentation requires oxygen in order to maintain aerobic condition and to sustain the aroma of the fermented batter and the final product when the batter is steamed. Hence the study done by Song et al., (1998) Mattheis and 130 Fellman, (2000) supports the current study who reported that production of aromatic compounds of many fruit, including apple, banana, pear, peaches, strawberries and others, can be adversely affected by low O2 and elevated CO2 i.e., synthesis of aroma compounds are generally suppressed. As mentioned in chapter 4, fermented aroma is one the criteria of idli which will be considered for sensory analysis and hence the treatment with high O2% may support the sensory quality of MAP packaged idli. The result of table 6.13 show that idli made from batter packaged with 0% to 5% CO2 and O2 ranging from 7.5% to 15% gave better results compared to all other treatments applied in the above experiments. 6.4 CONCLUSION From this study it can be concluded that ready to cook idli batter packaged in medium thickness (0.012 mm) LDPE flushed with 0% CO2 and 7.5 to 15% O2 could increase the shelf-life up to seven fold increase without compromising the sensory qualities at room temperature. 131 EXECUTIVE SUMMARY AND CONCLUSION Major objective of this work was to extend the shelf life of ready to cook idli batter using modified atmosphere packaging and the sub-objectives were 1. To understand the presently followed practices for the preparation of idli. 2. To optimize the process of preparation of the product with respect to ingredient ratios and fermentation time. 3. To improve the shelf-life of ready to cook idli batter by optimized process. First chapter was conceptualized with an objective to understand the presently followed practices for the preparation of idli. A survey based study was conducted in eight regions through an oral interview scheduled which covered a sample size of 300. The results of the survey indicated that at house hold level 68% of the selected population preferred parboiled rice. Only 34 per cent used decorticated black gram whereas 49 per cent used black gram with husk removed after soaking. Majority (99.7%) of the respondents used 3:1 ratio of rice and black gram dhal for preparing idli. Fermentation time varied between 5 h to 12 h at the selected households. Majority (71.3%) of them fermented the idli batter for 11 to 12 h and 73% stored the idli batter in refrigerated condition. The results were similar to the practices reported in literature such as variety of rice, type of black gram, ratio of ingredients used for idli making, fermentation time and shelf –life of the batter. Chapter 3 was aimed to optimize the process of preparation of the product with respect to ingredient ratios and fermentation time based on the instrumental texture profile of the idli using response surface methodology. Before framing the design using Central Composite Rotatable Design, preliminary trails were conducted to choose the best suited rice, variation of black gram and ratios of rice and black gram dhal. Five differently processed rice and ADT3 variety dhal were used for the preliminary study. Results of the preliminary study showed that IR20 parboiled rice and ADT3 variety black gram dhal with husk removed after soaking were best suited for idli making. The rice and black gram dhal were mixed at different ratios as per the CCRD. The independent parameters for this study were ratios of rice to black gram dhal and fermentation time. The dependent 132 parameters were the texture attributes namely hardness, adhesiveness, springiness, cohesiveness, chewiness and resilience. The results obtained were subjected to regression analysis and ANOVA. Based on the results certain constraints were imposed on the dependent parameters to get idli with better texture properties and the predicted values were obtained for the dependent parameters. From the study it was concluded that the optimum ratio of rice to black gram dhal is 3:1.575 with an optimum fermentation time of 14 h where a desirable value of 0.8279 will be obtained for the product. The results were validated by preparing idli at the optimized conditions. The results prove the designed model to be valid. Chapter 4 focused on the objective to identify the optimum ratios of ingredients and fermentation time with respect to sensory attributes using Response Surface Methodology (RSM). The desirable sensory attributes were colour, fluffiness, sponginess and fermented aroma. The undesirable parameters were compactness, stickiness, firmness and sourness. The idli were prepared according to the framed design. The semi-trained panel members evaluated the idli using a 15mm rating scale. Data were analyzed using RSM and constraints were imposed on the experimental results as in Chapter 3. On sensory analysis followed by RSM analysis of idli prepared from various combinations of ingredients fermented at different duration up to 14 h, rice and dhal combination of 3:1.475 fermented to 10.2 h was found to be the best accepted product. Principal Component Analysis was done to find the interrelationship between the sensory attributes of the idli. The PCA analysis revealed that PC1 and PC2 accounted for 78% of the total variance in the data matrix. It was clear that sensory attributes like sponginess and fluffiness associated with each other strongly on the positive side of the PC1 axis while firmness, compactness, stickiness were clustered together on the negative side of the PC1 axis. The third cluster is formed by fermented aroma and sourness on the positive side of PC2 axis. Sample from the experimental design point 6 was closely associated with desirable sensory attributes like sponginess and fluffiness followed by sourness and fermented aroma. On the other hand, design points 5, 1 and 2 were closely correlated with undesirable sensory attributes like firmness, compactness and stickiness. 133 The texture and sensory data were analyzed together by imposing constraints on the principal parameters of idli resulting in an optimized ratio of 3:1.18 with a fermentation time of 12.02 h. Chapter 5 dealt with the chemical components of nutritional importance in the optimized product. The final product was low in fat indicating the break down into fatty acids during fermentation. Analysis of fatty acid profile and oligosaccharide profile was done using LCMS. The fatty acids namely Decanoic acid, Octadecanoic acid and Hexadecanoic acid were of high relative abundance in idli compared to unfermented batter. Regarding the oligosaccharide profile, the sugars namely trehalose, maltose, melezitose, maltotriose, maltotetrose and maltopentose were formed during fermentation. Results showed that the process of fermentation has led to the increase in nutrient content of the idli. Chapter 6 aimed to improve the shelf-life of the batter using modified atmosphere packaging. Respiration dynamics was studied out to find the percentage of oxygen utilized and percentage of carbon dioxide released during fermentation of batter. Three packaging materials namely low density poly ethylene (LDPE), Poly propylene (PP), High Molecular (HM) of varying thickness were used for packing the idli batter. Twenty three gas combinations were used for MAP. The MAP packaged batter was stored at 30C and analyzed for gas mixture followed by texture and sensory analysis of the product during the storage period. The results of the respiration dynamics of the idli batter showed that the batter consumed 7% of O2 and evolved 12.6- 13% CO2 at 12.02 h fermentation time. From this study it can be concluded that RTC idli batter packaged in medium thickness LDPE flushed with 0% CO2 and 7.5 to 15% O2 could increase the shelf-life up to seven fold increase without compromising the sensory qualities at 30C. 134 Practical implications / Recommendations Based on the results and interpretations of this work following recommendations can be made to further improve the commercial prospects of ready to cook idli batter. 1. A detailed survey on the consumer perception and acceptance as well as problems associated with the currently available packaged ready to cook idli batter should be done to estimate the true potential of commercial ready to cook idli batter. 2. Detailed study on shelf-life of Modified atmosphere packaged batter under refrigerated conditions should be done. 3. 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(2010), Effect of γ-irradiation on phenolic compounds in rice grain, Food Chemistry, 120, 74-77. 184. http://www.census2011.co.in/census/state/puducherry.html 153 Annexure-I ORAL INTERVIEW SCHEDULE 1. Name : 2. Age : 3. Sex : 4. Address : a) Male b) Female 5. Educational qualification: 7. Occupation: 8. Monthly income: 9. Marital status: a) single 10. Type of family: a) nuclear b) married b) joint 11. Native place: 12. At what time do you take breakfast? a) 7-8 am b) 8-9 am) 9-10 am 13. Do you skip breakfast? a) Yes b) No 14. If yes, reasons to skip breakfast? 15. What are the food items you prefer for breakfast? a) Idly b) Dosai c) Poori d) Chapatti e) Oats 16. What are the breakfast items you usually prepare? 17. How often do you consume idly? a) Weekly b) Daily c) Monthly 18. How long years are you consuming Idli? 19. Do you like Idli? a)Yes b) No 154 20. Do you prepare Idli at home? a) Yes b) No 21. If no, from where do you get Idli? a)Hotel b) Idli stall 22. Do you buy as Idli batter? a) Yes b) No 23. Which is convenient to buy? a) Batter b) Idli 24. How often do you buy Idli batter? a) Daily b) Weekly c) Monthly 25. Where do you buy Idli batter? a) Household b) Departmental store 26. What is the brand name? 27. Do you buy Idli batter in the same or different shop? 28. Do you store the purchased batter or buy only when necessary? 29. Do you enquire about the ingredients and quality of the Idli batter while buying? a) Yes b) No 30. Do you like the quality of that Idli batter? a) Yes b) No 31. Does the Idli prepared from such batter taste good? a) Yes b) No 32. Do you prepare dosa from such batter? a) Yes b) No 33. Do you check the manufacturing date while buying? a) Yes b) No 155 34. Does the purchased Idli batter contain required amount of salt? a)Yes b) No 35. What will be consistency of the purchased Idli batter? a) Thick b) thin c) Normal 36. What will be the texture of the purchased Idli batter? a) Smooth b) coarse 37. Why do you prefer to buy Idli batter rather than preparing at home? a)Convenient b)Taste good c) Time saving d) Any other please specify 38. Do you buy Idli batter as cups measurement or in packet? a) Cup b) Packet 39. What is the cost of one cup of batter? 40. What is the cost of one packet of batter? 41. How many Idli can be made from one cup or one packet of batter? 42. Do you think it is hygienic to buy as batter? a)Yes b) No 43. What will be the colour of Idli prepared from purchased batter? 44. Do you grind Idli batter at home? a) Yes b) No 45. How often do you grind for Idli? a) Once in a week b) Twice a week c) Daily d) Once in a month 46. In what proportion you take rice and dhal? a) 3:1 b) 3:1.5 c) 3:2 156 47. Which variety of rice and dhal you use? 48. Do you always use same variety of rice and dhal? a) Yes b) No 49. Have you used with some other varieties of rice and dhal? Why? a) Yes b) No 50. How many times do you wash the rice and dhal before soaking? 51. What is the quantity of water you use to soak the rice and dhal? 52. For how many hours do you soak? 53. When do you usually grind the rice and dhal? a) Morning b) Afternoon c) Evening d) Night 54. Do you mix any additional ingredient along with dhal? a) Yes b) No 55. What are the additional ingredients you use? a) Rice flakes b) fenugreek c) Any other please specify 56. If yes, what is the proportion of the additional ingredient? 57. What is the purpose of the additional ingredient? 58. Which ingredient do you grind first? a) Rice b) Dhal 59. Which equipment you use to grind? a) Grinder b) Mixie c) Hand pound 60. Which do you think is the best equipment for grinding Idli batter? 61. Does the dhal that you use increase in volume while grinding? a) Yes b) No 157 62. If it does not increase in volume what will be the impact on Idli? 63. How long will you grind rice and dhal? 64. Do you grind them together or separately? 65. Till what consistency do you grind rice and dhal? a)Thick b)Thin c) Normal 66. What will be the texture of mixed batter? a)Coarse b) smooth 67. What will be the consistency of Idli batter after mixing? 68. Do you add salt before or after fermentation? 69. How long do you leave the batter for fermentation? 70. Do you add curd or yeast to favor fermentation? a) Yes b) No 71. What kind of vessel do you prefer to ferment the batter? a) Plastic b) Ever silver c) Mud pot d) Any other 72. Do you mix the batter with hand or ladle? 73. Whom do you prefer to mix batter and so why? 74. Does your batter get fermented overnight? a) Yes b) No 75. Does the batter volume increase after fermentation? a) Yes b) No 76. Do you get any respective flavour from fermented batter? a) Yes b) No 77. Usually what will be the colour of your Idli? a) White c) Light yellow 158 78. If the batter does not ferment do you get better Idli? a) Yes b) No 79. How do you control fermentation to increase the shelf life of the batter? 80. Do you prepare dosa from the same batter? a) Yes b) No 81. If the acidity increases, do you use that batter to prepare Idli? a) Yes b) No 82. Which utensil do you prefer to prepare Idli? a) Idli cooker b) Idli vessel 83. How many minutes do you steam? 84. How will you check if the Idli is cooked or not? 85. Do you give Idli as lunch for school going children? a) Yes b) No 86. How many times do you take Idli per day when the batter is available? 87. Do you prepare Idli regularly or only when some is sick at home? a) Yes b) No 88. Do you give Idli for the sick people? a) Yes b) No 89. Why do you prefer Idli during convalescent period? 90. Is your consumption of Idli increased when compared to the past years? 91. What do you think is the main reason? 92. Why don‘t go buy Idli as batter to prepare Idli? 93. If it is a hygienically prepared, healthy batter will you be ready to buy? a) Yes b) No 159 Annexure-II Sensory analysis score card for Idli Name: ___________ Date: ___________ -Taste the given samples and indicate intensity of perceived attributes by marking on the line at appropriate place. -Cleanse your palate with water in between samples. COLOUR ---- | ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ---- | ---Low High APPEARANCE Fluffiness ---- | ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ---- | ---Low High Compactness ---- | ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ---- | ---Low High TEXTURE Sponginess ---- | ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ---- | ---Low High Firmness ---- | ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ---- | ---Low High Stickiness ---- | ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ---- | ---Low High AROMA Fermented ---- | ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ---- | ---Low High 160 TASTE Sour ---- | ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ----- . ---- | ---Low High OVERALL QUALITY ---- | ----- . ----- . ----- . ----- . ----- . ----- . --|-- . ----- . ----- . ----- . ----- . ----- . ---- | ---Very Poor Fair Very Good Comments: Signature: 161 List of Publications 1. Transition in the Preparation and Consumption of Idli among the People of Puducherry, Durgadevi, M and Prathapkumar H. Shetty, Indian Journal of Nutrition and Dietetics (In Press). 2. Effect of Ingredients on Texture Profile of Fermented Food- Idli, Durgadevi, M and Prathapkumar H. Shetty APCBEES Procedia Journal (In Press). 3. Effect of Ingredients on Sensory Profile of Idli using Response Surface Methodology, Durgadevi, M and Prathapkumar H. Shetty, Journal of Food Science and Technology (Under revision). List of Proceedings 1. Presented a poster entitled ―Perception and consumption pattern of idli among the people of Puducherry‖ in the International conference on Traditional foods (Dec 1-3, 2010) conducted at Pondicherry University. 2. Presented a poster entitled ―Process optimization of the texture and sensory attributes of idli using response surface methodology (RSM)‖ in the National conference on Agro Food Processing Technologies (Nov 3-4, 2011) conducted at Pondicherry University. 3. Presented a poster entitled ―Effect on ingredients on the texture and sensory attributes of an Indian fermented food, Idli‖ in the Fifth International Conference on Fermented foods, health status and social well being: Challenges and Opportunities (Dec 15-16, 2011) at CFTRI, Mysore. 162