Anatomical structure of cladodes of Ruscus L. taxa (Liliaceae) in

FABAD J. Pharm. Sci., 36, 119-128, 2011
RESEARCH ARTICLE
Anatomical structure of cladodes of Ruscus L. taxa
(Liliaceae) in Turkey
Ayşegül GÜVENÇ*°, Maksut COŞKUN*, Okan ARIHAN**
Anatomical structure of cladodes of Ruscus L. taxa
(Liliaceae) in Turkey
Türkiye’deki Ruscus L. taksonlarının (Liliaceae) Kladot
Anatomisi
Summary
Özet
The genus “Ruscus” of Liliaceae family is represented by four
species (5 taxa) in Turkey; namely R. aculeatus L., R. hypoglossum L., R. colchicus P. F. Yeo, and R. hypophyllum L.
Two varietes of R. aculeatus (R. aculeatus var. aculeatus and
R. aculeatus var. angustifolius Boiss.) are also listed in the
Flora of Turkey.
Liliaceae familasında yer alan “Ruscus” cinsi, Türkiye’de R.
aculeatus L., R. hypoglossum L., R. colchicus P. F. Yeo ve
R. hypophyllum L. olmak üzere 4 tür (5 takson) ile temsil
edilmektedir. R. aculeatus (R. aculeatus var. aculeatus ve R.
aculeatus var. angustifolius Boiss.) iki varyete ile Türkiye
Flora’sında yer almaktadır.
Microscopical properties of transverse sections from the
cladodes of each taxa were illustrated and photographed. In
the cross section, it was not possible to differentiated upper
and lower epiderma of cladodes. In the mesophyll of cladodes
palisade and spongy parenchyma are not seen. There are a
few layers of parenchyma cells beneath the upper epidermis
and above the lower epidermis. Cross section of the midrib is
clearly convex in the lower surface. Vascular bundles have
been surrounded by sclerenchymatic bundles. The cladode is
isobilateral and both epiderma are similar in appearance being composed of generally long cells with, straight or slightly
sinuous walls. Numerous paracytic stomata are present on
both surfaces.
Her bir taksonun kladotlarından alınan enine kesitlerin
mikroskobik özellikleri, çizim ve fotoğraflarla verilmiştir.
Kladotların enine kesitlerinde alt ve üst epiderma farklılığını
belirlemek mümkün değildir. Kladot mezofilinde palizat
ve sünger parenkiması görülmemiştir. Üst epidermanın
altında ve alt epidermanın üstünde çok dar parenkimatik
hücre tabakası yer almaktadır. Orta damar, enine kesitte,
alt yüzde hafifçe konvekstir. İletim demetleri sklerenkimatik
bir halka ile kuşatılmıştır. Kladotlar isobilateraldir. Her iki
epiderma genellikle uzun hücrelerden oluşmuş ve birbirine
benzer görünüşte, düz veya hafifçe girintili çıkıntılı
çeperlidir. Her iki yüzde de çok sayıda parasitik tipte stoma
görülmektedir.
Key Words: Ruscus aculeatus, Ruscus hypoglossum, Ruscus
colchicus, Ruscus hypophyllum, cladodes, anatomy, Liliaceae.
Anahtar Kelimeler: Ruscus aculeatus, Ruscus hypoglossum,
Ruscus colchicus, Ruscus hypophyllum, kladot, anatomi,
Liliaceae.
Received: 01.10.2012
Revised: 09.01.2013
Accepted: 11.02.2013
* Ankara University, Faculty of Pharmacy, Department of Pharmaceutical Botany, Ankara, Turkey
** Yüzüncü Yıl University, Faculty of Arts and Sciences, Department of Biology, Zeve Campus-Van, Turkey
° Corresponding Author E-mail: aguvenc@ankara.edu.tr
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Güvenç, Coşkun, Arıhan
INTRODUCTION
Ruscus species are members of the Liliaceae
family, which are perennial, rhizomatus and
evergreen shrubs. The genus Ruscus is native to the
Mediterranean, Southern and Western Europe. Five
taxa of Ruscus grow naturally in Turkey. These are R.
aculeatus var. aculeatus, R. aculeatus var. angustifolius,
R. hypoglossum, R. colchicus and R. hypophyllum (1).
The underground parts of the Ruscus aculeatus
have been used medicinally as diuretic and antiinflamatory agents, as well as for hemorroids
treatments and atherosclerosis and circulatory
insufficiency prevention. As a treatment for diseases
of circulatory system, R. aculeatus has a long tradition
of proven success in Europe. A written record of
its use as a phlebotherapeutic agent dates back at
least 2000 years. These species are called as “sılcan,
tavşanmemesi, tavşan kirazı, diken kökü, yandak
dikeni, kandak, or sıçan dikeni”. A decoction of roots
of R. aculeatus is widely used as a diuretic and for
the treatment of urinary system disorders, as well
as kidney stones in folk medicine in the vicinities of
Eskişehir, Hatay, Mersin in Turkey (2-4). The roots of
these species, which is exported around 200-1000 tons
per year from Turkey, either fresh or as dried roots.
Because of this reason, R. aculeatus is vulnerable in
Turkey (5).
In the chemical studies conducted on various parts of
the Ruscus species, flavonoids and steroidal saponins
particularly ruscine, ruscoside, neoruscogenin and
ruscogenin as well as many new steroidal saponins
have been identified. Intensive chemical studies have
been carried on the Ruscus species and isolated many
new steroidal saponins. The amounts of steroidal
saponins have also been determined by HPTLC,
HPLC, HPLC-ESI-MS, and UPLC methods (6-18).
Stem leaves in Ruscus species, are scale-like and
membranous, additionally, stems are reduced to
photosynthetic phylloclades. Being a metamorphic
form of stem, anatomical structure of the phylloclades
should be very interesting. In a study, Sachs et al.
investigated variable development and cellular
patterning in the epidermis of R. hypoglossum. This
study has concerned the presence and the meaning
of immature stomata in the epidermis of it. The
distribution of the stomata of Ruscus was remarkably
variable (19). Although a lot of work has been done on
R. aculeatus, especially morphological characteristics,
phylloclade development and systematic (20-22),
a few data are available in the literature related to
anatomical structure of the genus (19, 23). We studied
cross sections obtained from cladodes of R. aculeatus
which is in European Pharmacopoeia 2008 (24) and
the other three species growing in Turkey. We used
the term of cladodes, because this term is used in the
Flora of Turkey.
Materials and methods
Research materials were collected from various
locations in Turkey. Voucher specimens were
deposited in AEF (the Herbarium of Ankara
University, Faculty of Pharmacy) Locations of the
investigated plant samples are given in Table 1.
Plants materials were preserved in 70% ethanol.
Cross sections of the cladodes of the Ruscus taxa were
performed by hand from the preserved materials in
chloral hydrate solution and Sartur reagent. Sartur
Table 1. Locations of the studied Ruscus taxa
Species
Locality
Ruscus aculeatus var. aculeatus
A3 Sakarya: Karasu, 22.9.2001, M. Coşkun, A. Güvenç & C. S. Erdurak (AEF 22940).
R. aculeatus var. angustifolius
A6 Samsun: Terme, Sivaslar village, 14.3.2002, M. Coşkun & O. Arıhan (AEF 22939).
R. colchicus
A6 Ordu: Ünye, 15.3.2002, M. Coşkun & O. Arıhan (AEF 22934).
R. hypoglossum
A7 Giresun: Tirebolu, İnköy roadside, 15.3.2002, M. Coşkun & O. Arıhan (AEF 22930);
R. hypophyllum
A7 Giresun: Tirebolu to Espiye, 16.3.2002, M. Coşkun & O. Arıhan (AEF 22933);
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FABAD J. Pharm. Sci., 36, 119-128, 2011
reagent contains KI-I, aniline, sudan III, lactic acid,
alcohol and water (25). Illustrations were made
using a Leitz drawing prism attached to a LeitzWetzlar (45o) microscope. The cross sections were
photographed with an automatic camera attached to
Olympus BX 50 microscope.
Results
The anatomical features were observed in cross
sections of the cladodes (Figures 1-10) and the
comparisons of the anatomical characters of cladodes
of taxa are given in Table 2.
Cross section of cladode blade
In the cross-section, it was not possible to
differentiate upper and lower surfaces of the
cladodes; after a thick cuticle, epidermis is preceded
by parenchymatic cells. In the mesophyll of cladodes,
palisade and spongy parenchymas were not seen.
There were a few layers of parenchyma cells beneath
the upper epidermis and above the lower epidermis.
This parenchyma contains chloroplast and they
exist instead of palisade parenchyma. Between the
two parenchyma layers, one or two layers of large
and colourless cells are present, differing from one
species to other. This cell layer exists instead of
spongy parenchyma (Fig. 2 A, B; Fig.3 A; Fig. 4 A;
Fig. 5 A, B; Fig. 6 A, B; Fig. 9 A; Fig. 10 A).
Cross section of the midrib
Cross section of the midrib is clearly convex in
the lower surface (Fig. 1). Vascular bundles are
Figure 1. Transverse sections of the cladodes of Ruscus
species growing in Turkey. A- R. aculeatus var. aculeatus;
B- R. aculeatus var. angustifolius; C- R. colchicus; D- R.
hypoglossum; E- R. hypophyllum.
collateral and surrounded by sclerenchyma. The
number of vascular bundles differ from species
to species; there are numerous vascular bundles
in R. colchicus, R. hypoglossum, and R. hypophyllum
Table 2. Comparison of the cross sections of different Ruscus taxa native in Turkey.
Epidermal
cell
Upper
Lower
Colorless Vascular bundles in the
Section of
parenchyma parenchyma parenchyma
midrib
cladode margin
R. aculeatus var.
aculeatus
Square or
rectangular
3-4 (-5) layers
3-5 layers
2 layers
1, pith sclerenchymatic
rectangular
R. aculeatus var.
angustifolius
rectangular
3-4 layers
3- (-4) layers
2 layers
1, pith sclerenchymatic
rectangular
R. colchicus
rectangular
3-5 layers
2-4 layers
1- (-2) layers
6-7,pith sclerenchymatic
and parenchymatic
acute
R. hypoglossum
rectangular
2-3 layers
2-3 layers
(1-) -2 layers
10-11 (-13), pith
sclerenchymatic and
parenchymatic
rhombic
R. hypophyllum
rectangular
2-4 layers
2-3 layers
1- (-2) layers
10-11 (-13), pith
sclerenchymatic and
parenchymatic
acute
Species
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Güvenç, Coşkun, Arıhan
Figure 2. A-Cross sections of the cladodes margins of R. aculeatus var. aculeatus, A1- Surface of upper epidermis, A2-Surface
of lower epidermis; B- R. aculeatus var. angustifolius; B1-Surface of upper epidermis B2-Surface of lower epidermis. acuticle, b- epidermis, c- upper parenchyma, d- colorless parenchyma, e- lover parenchyma, f- sclerenchyma, g- phloem,
h- xylem.
however few in the R. aculeatus (Fig. 3B; Fig. 4B; Fig.
7; Fig. 8; Fig. 10B).
Vascular bundles other than midrib the cross section
of the vascular bundles of the cladode blade, phloem
is at the upper surface.
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Surface epidermis
Upper and lower epiderma are similar in appearance
being composed of generally long cells, with straight
or slightly sinuous walls. Numerous paracytic
stomata are present on both surface (Fig. 2 A1, A2, B1,
B2; Fig. 5 A1, A2, B1, B2; Fig. 9 A1, A2).
FABAD J. Pharm. Sci., 36, 119-128, 2011
Figure 3. Cross sections of the cladodes of R. aculeatus var.
aculeatus. A- leaf blade, B- midrib; a- cuticle, b- epidermis,
c- upper parenchyma, d- colorless parenchyma, e- lover
parenchyma, f- sclerenchyma, g- phloem, h- xylem.
Figure 4. Cross sections of the cladodes of R. aculeatus
var. angustifolius. A- leaf blade, B- midrib; a- cuticle, bepidermis, c- upper parenchyma, d- colorless parenchyma,
e- lover parenchyma, f- sclerenchyma, g- phloem, h- xylem.
Discussions
In this study, we have reported the anatomical
structure of cladodes of Ruscus taxa native to Turkey.
Anatomical characteristics of the cladodes of these
taxa have been studied and their structures have
been determined.
It has been observed in the anatomical structure of
the cladodes of various Ruscus taxa that, especially
midrib and section of cladode margins are quite
different from each other. The number of vascular
bundles in the midrib is one in R. aculeatus;
however, this number is six or more than the six in
the R. colchicus, R. hypoglossum and R. hypophyllum,
respectively. Section of the cladode margin is
rectangular in R. aculeatus, acute in R. colchicus and R.
hypophyllum, and rhombic in R. hypoglossum (Figures
1, 2, 5, 9; Table 2).
In Ruscus and Asparagus species (both from
Asparagaceae family), stem leaves become scalelike and membranous as a result of the reduction of
the stem to photosynthetic cladodes. In a previous
study, we have reported the anatomy of cladodes of
Asparagus species (26). In Asparagus species palisade
and spongy parenchyma are seen in the mesophyll
of cladodes. However, in Ruscus species these two
types of parenchyma are not observed. Vascular
bundles in the midrib of certain Ruscus taxa are
numerous. This property is characteristic for stems
of Monocotyledonae. Crystals of calcium oxalate are
not seen in the cladodes of Ruscus species.
References
1. Davis PH. Ruscus L. In: Davis PH. (ed.), Flora of
Turkey and The East Aegean Islands. Vol. 8:72-74,
Edinburgh University Press; Edinburgh; 1984.
2. Başer KHC, Honda G, Miki W. Herb Drugs and
Herbalists in Turkey. Instutute for the study of
Languages and Cultures of Asia and Africa.
Tokyo, p. 36, 231, 1986.
123
Güvenç, Coşkun, Arıhan
Figure 5. A-Cross sections of the cladodes margins of R. colchicus, A1- Surface of upper epidermis, A2-Surface of lower
epidermis; B- R. hypoglossum B1-Surface of upper epidermis B2-Surface of lower epidermis a- cuticle, b- epidermis, cupper parenchyma, d- colorless parenchyma, e- lover parenchyma, f- sclerenchyma, g- phloem, h- xylem, ı- stoma.
124
FABAD J. Pharm. Sci., 36, 119-128, 2011
Figure 6. Cross sections of the cladodes of A- R. colchicus. B- R. hypoglossum; a- cuticle, b- epidermis, c- upper parenchyma,
d- colorless parenchyma, e- lover parenchyma, f- stoma
Figure 7. Cross section of the cladodes midrib of R. colchicus.
a- cuticle, b- epider-mis, c- upper parenchyma, d- colorless
parenchyma, e- lover parenchyma, f- sclerenchyma, gphloem, h- xylem.
Figure 8. Cross section of the cladodes midrib of R.
hypoglossum. a- cuticle, b- epider-mis, c- upper parenchyma,
d- colorless parenchyma, e- lover parenchyma, fsclerenchyma, g- phloem, h- xylem.
125
Güvenç, Coşkun, Arıhan
Figure 9. A- Cross sections of the cladodes margins of R. hypophyllum. A1- Surface of upper epidermis, A2-Surface of lower
epidermis; a- cuticle, b- epidermis, c- upper parenchyma, d- colorless parenchyma, e- lover parenchyma, f- sclerenchyma,
g- phloem, h- xylem.
126
FABAD J. Pharm. Sci., 36, 119-128, 2011
Figure 10. Cross sections of the cladodes of R. hypophyllum. A- leaf blade, B-midrib; a- cuticle, b- epidermis, c- upper
parenchyma, d- colorless parenchyma, e- lover parenchyma, f- sclerenchyma, g- phloem, h- xylem.
3. Baytop T. “Türkiye’de Bitkiler ile Tedavi.
Geçmişte ve Bugün”. 2nd ed., Nobel Tıp
Kitabevleri, İstanbul, p 353, 1999.
4. Tabata M, Honda G, Sezik E. A report on
traditional medicine and medicinal plants
in Turkey (1986). Faculty of Pharmaceutical
Sciences, Kyoto University, pp. 52, 54, 1988.
5. Özhatay N, Koyuncu M, Atay S, Byfield A.
Türkiye’nin Doğal Tıbbi Bitkilerinin Ticareti
Hakkında Bir Çalışma. Doğal Hayatı Koruma
Derneği, İstanbul, Türkiye. p. 50, 1997.
6. Elsohly AM, Doorenbos NJ, Quimby MW, Knapp JE,
Slatkin DJ, Schiff PL, Euparone, a new benzofuran
from Ruscus aculeatus L. J Pharm Sci 63: 1623-24, 1974.
7. Evans WC. Trease and Evans’ pharmacognosy.
Thirteenth edition, English Language Book
Society/ Bailliere Tindall, Oxford, p. 491, 1989.
8. Blumenthal M, Busse WR, Goldberg A,
Gruenwald J, Hall T, Riggins CW, Rister RS. The
complete German commission E monographs.
Therapeutic guide to herbal medicines. American
Botanical Council, Austin, Texas, pp. 99-100, 1998.
9. PDR for Herbal Medicines. Second edition.
Medical Economics Company, Montvale, New
Jersey, pp. 561-62, 2000.
10. Redman DA. Ruscus aculeatus (Butcher’s
Broom) as a potential treatment for orthostatic
hypotension, with a case report. J. Alternative
Complementary Med. 6: 539-49, 2000.
11. Panova Von D, Nikoloff St, Achtardjieff C.
Steroidsapogenine und sterole aus Ruscus
hypoglossum. Planta Medica 26: 90-3, 1974.
12. Peneva B, Krasteva I, Nikolov S, Minkov
E. HPTLC densitometric determination of
ruscogenins in dry extract of Ruscus aculeatus L.
Pharmazie 55: 103-6, 2000.
13. Bertani W, Forni GP. HPLC determination of
ruscogenin and neoruscogenin in raw material
and pharmaceutical preperations. Fitoterapia 55,
101-4, 1984.
14. Mimaki Y, Kuroda, Yokosuka A, Sashida Y. A
spirostanol saponin from the underground parts
of Ruscus aculeatus. Phytochemistry 51: 689-92,
1999.
15. De Combarieu E, Falzoni M, Fuzzati N, Gattesco
F, Giori A, Lovati M, Pace R. Idetification of
Ruscus steroidal saponins by HPLC-MS analysis.
Fitoterapia 73: 583-96, 2002.
16. Güvenç A, Şatır E, Coşkun M. Determination of
ruscogenin in Turkish Ruscus L. species by UPLC.
Chromatographia 66: 141-5, 2007.
17. Mimaki Y, Aoki T, Jitsuno M, Kılıç CS, Coşkun M.
Steroidal glycosides from the rhizomes of Ruscus
hypophyllum. Phytochemistry 69: 729-37, 2008.
18. Mimaki Y, Aoki T, Jitsuno M, Yokosuka A, Kılıç
CS, Coşkun M. Steroidal saponins from the
rhizomes of Ruscus hypophyllum. Natural Product
Communications 3: 1671-8, 2008.
19. Sachs T, Novoplansky N, Kagan ML. Variable
development and cellular patterning in the
epidermis of Ruscus hypoglossum. Annals of
Botany 71: 237-43, 1993.
20. Cooney-Sovetts C, Sattler R. Phylloclade
development in the Asparagaceae: an example of
127
Güvenç, Coşkun, Arıhan
homoeosis. Botanical Journal of the Linnean Society
94: 327-71, 1986.
21. Hirsch AN. A developmental study of the
phylloclades of Ruscus aculeatus L. Botanical
Journal of the Linnean Society 74: 355-65, 1977.
22. Rudall PJ, Conran JG, Chase MW. Systematics
of Ruscaceae/Convallariaceae: a combined
morphological and molecular investigation.
Botanical Journal of the Linnean Society 134: 73-92,
2000.
23. Schulte PJ, Gibson AC. Hydraulic conductance
and tracheid anatomy in six species of extatnt
seed plants. Can J Bot 66: 1073-9, 1987.
128
24. European Pharmacopoeia, 6th Ed., Vol. 2,
Strasbourg: Council of Europe pp. 1355-6, 2008.
25. Çelebioğlu S, Baytop T. Bitkisel tozların tetkiki
için yeni bir reaktif. Farmakognozi Enstitüsü
Yayınları, No. 10. Farmakolog 19, 301, İstanbul.
1949.
26. Güvenç A, Koyuncu M. Studies on the
anatomical structure of cladodes of Asparagus L.
species (Liliaceae) in Turkey. Israel Journal of Plant
Sciences 50: 51-65, 2002.