FABAD J. Pharm. Sci., 22, 153-158, 1997
RESEARCH AR11CLES / BİLiMSELARAŞTlRMALAR
A Comparative Chemotaxonomic Study on Eleven Rumex Species Growing in Turkey
L. Ömiir DEMİREZER*0, Ayşe KURUÜZÜM*
A Coınparative Cheınotaxonomic Study on Eleven Ruınex
Species Growing in 1'urkey
Su1n1nary : in tlıis study, tlıe roots of the -eleven Runıex spe- cies growing in Turkey were studied vvitlı regard to clıe111.o
faxononıy. Major substaııce~· identified -..vere anthraquinones and tannins. Naphthaiene {/{'rfvatives and flavonoids rverc found in sonıe species. Only one species contairıed an-
ıhocyanidol and counıarin ,vvhilst nane oftlıe species studied l-Vere found to contain. iridoids and cardiac glycosides. The an1ou11t of tannins and flavonoids ~vere detennined as 1.97- 8.48 % and 0.01-0.09 %, respectively.
Key words: Runıex spec.; Polygo11aceae; clıe11ıotaxon.onıy
Received Revised Accepted
3.12.1996 9.6.1997 16.6.1997
INTRODUCTION
The roots of Ruınex species are often ernployed in Turkish folk rnedicine asa Jaxative, diuretic and chol- agogue. lt is known !o contain anthraquinone de- rivatives and tannins as the bioactive principlesl.
Moreover, the diuretic effect of the planı may be rc- lated with its fla vonoid content. in a previous study, the quantities of anthraquinone glycosides in 10 Ruın
ex species werc determined by using spectro- photometry2. However, !here is no report concerning the tannin and flavonoid content of !he selected Rum- ex species.
The aim of !his study is !o examine !he methanolic extracts of Rnmex rooıs for the presence of an- thraquinones, tannins, flavonoids and other bio- genetically related compounds, and to discuss the re- sults obtained from the chemotaxonomic point of view to establisl1 a coı111ectio11 betvveen_ chenlical con- stituents and taxonomy of plants.
1'iirkiye'de Yetişen 11Rıanex1'iirüniin Karşılaştırınalı
Bir Keınotaksonoınik Çalışunasz
Özet: Bu çalışnıada, Türkiye'de yetişen 11 Runıex türünün kökleri keınotaksononıik açıdan incelennıi_ştir. Bitkilerde
ınajör olarak antrakiııon ve tanen. saptann11ştır. Bazr tür- lerde najiale11 ve flavonoit bulunnıu:ştur. Sadece bir türde antosiyanidol ve kıonari11 teşhis e(Ul!ne:sine rağnı.en, kar- diyoaktif heterozit ve iridoite hiç rastlannıanuştır. Yapılan nıiktar tayinleri sonucunda % 1,97-8,48 arasında tanen, % 0,01-0 ,09 arasında flavonoit buh11unu.ştur.
Anahtar kelilneler: Rıunex türleri, Polygonaceae,
keJnotaksononıi.
Far !his purpose, we colkcted 11 species from differ- ent localities in Turkey, identified a11d categorizcd
!hem by their morphological characters. The separa- tiÜn and identificatio11 of the main co11stitue11ts of
Rıımex extracts are described by means of thin layer chromatography and colour reactions.
The second goal of this study is to compare thc dala obtained frorn the quantitative analysis of the tannins and flavonoids present in the roots of 11 Rwnex spc- cies growing in Turkey according to chemotaxonomic distribution.
MATERIALS and METHODS
l'lant Material : Al! plants were collected from differ- ent localities in Turkey. Rııınex acetosella L., Ti.. an-
gııstifoliııs Campd. subsp. angııstifolius, R. gracilescens Rech. and R. tmoleııs Boiss. were collected from Sivas- Tokat (alt. 1600 m -2000 m). R. conglomeratııs Murr., R. dentatus L. subsp. lıalacsyi, R. sangııineııs L. were
*
Hacettepe University, Faculty of Pharınacy, Departınent of Pharnıacognosy 06100 Ankara/ Turkey Correspondenc.eDe-ınirezer, Kuruüzürn
collected from Aydın-Kuşadası-Davutlar (alt. O -10 m), R. alpiııııs L. frorn İzrnir-Öderniş-Bozdağ (alt.
1650 rn), R. crispııs L. from Ankara-Lalahan (alt. 1100 m), R. patientia L. frorn Niğde-Bor (alt. 1050 m), R.
scutatus L. from Yozgat-Çekerek (alt. 1200 m).
Voucher specimens were deposited in the herbarium of the Faculty of Pharmacy, Hacettepe University, Ankara (Turkey). Completely dried rooı material was powdered wilh an electric grinder and stored in well closed cellophane bags at room temperalure.
Sample preparalion and colour reac!ion: Sample preparations and color reactions were made as de- scribed in the litera!ure (3,4,5,6). For anthocyanidol acid and base reactions, for anthraquinone Borntraeg- er reaction, for flavoı1oid Shi11oda rcaction, for iridoid Trim-Hill reaction, for cournarin NaOH reaction, for saponin Salkowski reaction, for tannin FeCl3 and gel- atin-salt block test were applied.
Thin Layer Chromatography: 0.1 g dried material was extracted with 10 ml methanol. After filtration 10 µ! of aliquoı was applied to a 0.2 mm thin layer silica gel plate.
Solvenı systemsı:
Ethyl acelate:Methanol:Waler (100:13,5:10)
Ethyl acetate:Formic aC°ıd:GI. acetic acid:Water (100:11:11 :27)
Chloroform:Methanol:Water (64:50:1 O) Chloroform:Methanol:Water (80:20:2) Cyc lohexane:Elhylform iate: Melhylene chloride:Formic acid (35:30:30:5) C h 1 o rof o r m: Metha no 1 :Wa ter: Form i c acid (7:13:8:0.1)
Petroleum elher:Ethyl acelate:Acetic acid (75:24:1)
Detection6,7:
flavonoids, cardiac gly- cosides, coumarins, ir- idoids, lannins, an- thraquinones anlhocyanidols, flav- onoids
saponins
anlhraquinones (bound)
anthraquinorıes (free) naphlhalenes naphthalenes
1. The spols were first s!udied directly on !he chromatogram in daylight and under UV light (Camag)
2. The following spray reagents were used for vis- ualization:
a) Fası blue salt reagent: 5 % of 4,4'-bis-(2-melhyl benzol diazoniumchlorid) in water for naph- lhalene derivatives.
154
b) 5 % of KOH in methanol (50 % v /v) and heat- ed for 15 rnin at 100 °C for anthraguinone and coumarine derivatives.
c) Na!urstoff reagent: 1 % of diphenylboric acid arninoethyl ester in methanol for flavonoid de- rivatives.
d) NH3 vapour for flavonoid and anthocyanidol derivatives.
e) 5 % of FeCl3 in water and vanillin/ conc.
H2S04 for tannin derivatives.
f) Kedde reagent (3,5-dinitrobenzoic acid) far car- diac glycosides.
g) 30 % of H2S04 in w.ater and heated 100°C far saponin derivatives.
h) Trirn-Hill reagent for iridoid derivatives.
Quan!ila!ive Determinalion of Flavonoids and Tan- nins:
Flavonoid determination was made according to ref- erence 6 and 8.
Tannin determination was realized according to ref- erence 6.
RESULTS AND DISCUSSION
Ali species of Rumex genus investigated were screened with classical colour reactions for the pres- ence of anthraquinones, flavonoids, saponins, an- thocyanidols, tannins, coumarins, naphtha]enes, car- diac glycosides, leucoanthocyanidols and iridoids.
These results were confirmed with thin-layer cllfom- atography (tlc). The results of color reactions and tlc were surnmarized in Table 1.
Tannins were previously reported in the roots of R.
acetosella9,1D,1l, R. alpinııSJ,lO,l2,l3,l4, R. conglomerahıs 9,10,ıs, R. crispı159,10,15,ı6, R. patientia9,ıo,12,ıs, R. sangııin
eus15 and R. scııtah.ıs9,n in our study, ali screened spe- cies of Rı11nex genus were shown to contain taımins
and for the first time tannin presence was proven in 4 species (R. gracilescens, R. tmoleus, R. anguslifolius subsp. angustifolius, R. dentah.ıs subsp. halacsyi).
lt has been observed that, except R. tınoleus10, other species contained anthraquinones and naphthalene derivatives. This observation is very interesting from the chemotaxonomic point of view. Although Rııınex
species are known as anthraguinone containing plants, one of the species is completely devoid of the anthraguinones. ln previous s!udies, the presence of two naphtoguinone derivatives, nepodin and nep- oside, in the roots of R. alpimıs, R. crispııs and R. pa- tientia were reportedıo,ı7,ıs,ı9.
Table 1. (CR: Color reaclion; TLC: Thin Layer Chromalography; +: present; -: absent)
Anthocyanidol Anthraquinone Flavonoid İridoid Cardiac
Species glycoside
CR TLC CR TLC CR TLC CR TLC TLC
R. acetosella + + + + + +
- - -
R. alpinus -
-
+ + + +- - -
R. angııstifoliııs
-
- + +- - - -
-'
subsp.angustifolius
R. conglorneratııs
-
- + +- - - -
-R. crispus
- - -
+ + + +- -
-R. dentatııs -
-
+ +- - - - -
1 subsp. halacsyi
R. gracilescens
- -
+ + + +- -
--
R. patientia
- -
+ + + +- - -
R. sanguineus
- -
+ + + +- - -
R. scııtatus
- -
+ + + +- - -
1
R. trnoleus
- - -
- +...
--
-Coumarin Naphth- Saponin
cı-lene
CR TLC TLC CR TLC
+ + +
- -
- -
1
+ +
-
- -
1 + +-
1
-
-
+ +-
- -
+ +-
- -
+ --
- -
+ +-
- -
+'
+ --
- -
+ + -- -
+- -
-
- -
--
'
TanninCR TLC
+ +
+ +
+ +
'
+ i +
1
+ +
1 + +
1
1 + +
+
t
++ +
i-b
.,, ~
""
;;;:
"' ~
"'
'1.
w w
~
"'
'-"'""
_62
~
:::s
' l
Deınirezcr, Kurııüziinı
Moreover, naphthalene derivatives were dctected in 7 species (R. acetosella, R. angııstifolius subsp. angustifolius, R.
conglomerahıs, R. dentahıs subsp. halacsı;i, R. gracilescens, R.
sangııineus, R. scııtatus) far the first time. Previously sap- onins were reported in the roots of R. abyssinicusıo, R.
chalepensis20 and R. flexuosus21. We detemıincd saponins in 7 species of Ruınex by the classical colour reactions, but thcrc were no spots in tlc leading that the presence of saponins remained doubtfnl and nccds to be studied in delail. Although leucoanthocyanidols were previously reported in R. hymrnosepalus and R. confertus 10,22,23,24, thc existence of thcse typc of compounds were not clearly proven in this study. None of the species studied in !his study conlained iridoids or cardiac glycosides.
The detennination of flavonoids was ambiguous in tlc because of overlapping with other substances but col- our reactions have given positive results. For a reliab]e result, flavonoid contents were determined by the spectrophotometric method6·8. in this way in 8 sp;ecies flavonoid presence "\vas found to range from O.Ol ıo 0.09 %. Tamun and flavonoid percentages of the selected species are shown in Table 2. Up to the present flavonoids have been reported in 32 Rıunex
vvere found in R. dentatııs. Our findings showed that a subspecies of lhe plant, R. dentahıs subsp. halacsyi, con- tained no flavonoids in the roots.
Scheme 1
Q_
COOM cınn~mfc aci~ \
°' /
F
C"'lo-C 1 il 4-Coum"l)'I Col\
o
r"'Y'~
~ COUH
C,. CO<JIT],ı1ilC!~
/
W
~'" 1
Otl O
HO .y
1
Catochol J\~lh<>qooodol
Table 2. The Amounts of Tannins aııd Flavonoids with Standard deviation and Standard error.
Tannin
Species % Standard
Tannin deviation
R. acetosella 2.61
o
R. alpinus 4.94 ±0.14
R. angustifolius subsp. 1.97 ±0.17 angustifolius
R. conglomeratus 8.48
o
R. crispus 2.90 ±0.24
R. dentatus subsp. 6.18 ±1.23
halacsyi
R. gracilescens 3.23 ±0.65
R. patientia 3.95 ±0.30
R. sanguineus 7.80 ±0.42
R. scutatus 6.18 ±1.00
R. tmoleus 6.88 ±0.24
spedes in different studies. Accordiııg to these studies, the aerial parts of most R11mex species aııd undcr- ground parts of 5 Ruınex species contained flav- onoids21,25,26,27. There is a disputed result with R. den-
tatııs. According to Hegnauer21 quercetin and rutin
156
Flavonoid
Standard % Standard Standard
error Flavonoid deviation error
o
0.015 ±0.0195 ±0.0088±0.10 0.045 ±0.00008 ±0.000034
±0.12
o
- -o
() - -±0.17 0.055
o o
±0.87
o
- -±0.46 0.040 ±0.005 ±0.0022
±0.21 0.090 ±0.098 ±0.044
±0.29 0.010 ±0.0059 ±0.0026
±0.71 0.018 ±0.0063 ±0.0028
±0.17 0.026 ±0.00005 ±0.00003
Only the roots of R. acetosella contained anthocyanidol and coumarin. Coumarin in this plant was also re- ported previousJyll. When the biogenctic pathway (Scheme 1)28 from cinnamic acid is considcred, flav- onoids originatc frorn one of the pathways, and cou- marins are form'"'Cl from the oföer pathway. An-
FABiıD J. Plıarnı. Sci., 22, 153-158, 1997
thocyanidols derive from dihydroflavonol meanwhile catechol also appears by a different mechanism.
To contribute to chemotaxonorny, tannin contents were determined by thc titrimetric method6. AB spe- cies contained quite a high percentage of tannins. The amount of tamıins were found ıo vary from 1.97 to 8.48 % in these species. The results of quantitative analysis and their sta11dard error of tannins and flav- onoids present in the roots of select~d Rıımex species were also calculated.
Flavonoid and tannin percentages of Runıex acetosella were found ıo be considerably lowcr than those of other screened Rıımex species. Accordingly it can be concluded that cinnamic acid, which may contributc to flavonoid presence, was mostly utilized in cournar- in formation, therefore flavonoid arnounts in thi_s spe- cies was lower than in others. Similarly, the increase of anthocyanidol conversion frorn dihydroflavonol decreased catechol quantity in this plant.
shown.The anthraquinone percentages given here are extracted from a previous study2.
!n conclusion, on the basis of these results, the major substances of Rurnex genus are a.ı1thraquinones and tannins. The underground parts of only one species, Rumex acetosella, is proven to contain anthocyanidol and coumarin lor the lirst time. The presence of flav- onoids is of no significance.
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'"""ı
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~ Truının n
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Figure 1. Quantitative Comparison of Tannins, Flavonoids and Anthraquinones in the roots of 11 Rıunex species.
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