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Title: Anti-inflammatory and phytochemical properties oftwelve medicinal plants used for treating gastro-intestinalailments in South Africa
Authors: O.A. Fawole, A.R. Ndhlala, S.O. Amoo, J.F. Finnie,J. Van Staden
PII: S0378-8741(09)00153-6DOI: doi:10.1016/j.jep.2009.03.012Reference: JEP 5470
To appear in: Journal of Ethnopharmacology
Received date: 23-12-2008Revised date: 5-3-2009Accepted date: 11-3-2009
Please cite this article as: Fawole, O.A., Ndhlala, A.R., Amoo, S.O., Finnie, J.F.,Van Staden, J., Anti-inflammatory and phytochemical properties of twelve medicinalplants used for treating gastro-intestinal ailments in South Africa, Journal ofEthnopharmacology (2008), doi:10.1016/j.jep.2009.03.012
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Anti-inflammatory and phytochemical properties of twelve medicinal 1
plants used for treating gastro-intestinal ailments in South Africa2
3
O.A. Fawole, A.R. Ndhlala, S.O. Amoo, J.F. Finnie, J. Van Staden*4
5
Research Centre for Plant Growth and Development, School of Biological and Conservation Sciences, 6
University of KwaZulu-Natal Pietermaritzburg, Private Bag X01, Scottsville 3209, South Africa7
* Corresponding author, e-mail address: [email protected] (J. Van Staden)8
Tel: +27 33 2605130; Fax: +27 33 26058979
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Received………………………..11
12
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Abstract14
15
Ethnopharmacological relevance: The investigated medicinal plants are commonly used for 16
the treatment of pains and cramps related to gastro-intestinal tract infections in South African 17
traditional medicine.18
Aims of the study: This study aimed to evaluate the ability of the plant extracts to inhibit 19
cyclooxygenase enzymes. Phytochemical analysis was also carried out in the quest to 20
determine some plant metabolites that may be responsible for the observed anti-inflammatory 21
activity.22
Materials and methods: The cyclooxygenase assay was used to test for the anti-inflammatory 23
activity of the plant extracts using cyclooxygenase-1 and -2 (COX-1 and COX-2) enzymes. 24
Total phenolic compounds including condensed tannins, gallotannins and flavonoids were 25
* Manuscript
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quantitatively determined using spectrophotometric methods. Qualitative tests for alkaloids 26
and saponins were also carried out.27
Results: Most of the plant extracts evaluated showed dose dependent activity against COX-1 28
and/or COX-2 enzymes. Agapanthus campanulatus root dichloromethane extract showed the 29
highest COX-2 inhibitory activity (83.7%) at 62.5 µg/ml. The presence and/or amounts of 30
phenolics, condensed tannins, gallotannins, flavonoids, alkaloids and saponins varied with 31
plant parts and species.32
Conclusion: The results support the use of the investigated plant in treating pain and cramp 33
related to gastro-intestinal tract infections. The observed anti-inflammatory activity could to 34
some extent be attributed to the various plant secondary metabolites detected in the plant 35
materials.36
37
Keywords: Anti-inflammatory; Cyclooxygenase; Gastro-intestinal ailments; Secondary 38
metabolites39
40
41
1. Introduction42
43
Gastro-intestinal ailments are associated with inflammation of the gastro-intestinal tract44
resulting in abdominal pains and cramps of varying degree (Barbara, 1998). Naik and Sketh 45
(1976) defined inflammation as a complex, vascular lymphatic and local tissue reaction 46
elicited in animals by the presence of viable and non-viable irritants. The intestine is 47
vulnerable to muscle spasm in patients suffering from gastro-intestinal infections, and most48
patients suffering from such conditions often complain of abdominal cramps and pains49
(Sleisenger and Fordtrand, 1993). Escherichia coli and other gastro-intestinal pathogens 50
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associated with food poisoning produce enterotoxins that induce watery diarrhoea and 51
abdominal tissue damage through plasmid-encoded invasion factors, resulting in acute or 52
chronic abdominal pains and cramps (Naik and Sketh, 1976; Sleisenger and Fordtrand, 1993).53
Non-steroidal anti-inflammatory drugs (NSAIDs) typically relieve inflammation and 54
associated pain by inhibiting cyclooxygenase enzymes involved in the production of 55
prostaglandins. These enzymes exist in two isoforms (COX-1 and COX-2) coded by distinct 56
genes on different chromosomes (Polya, 2003). The two isoforms show about 50% homology 57
and have similar catalytic activity, but are physiologically distinct (Pasinetti, 2001). 58
Compounds that inhibit COX enzymes could therefore be considered to be potential anti-59
inflammatory drugs. However, many of the commonly used anti-inflammatory agents are 60
becoming less acceptable due to serious adverse reactions such as gastric intolerance, bone 61
marrow depression and water and salt retention, resulting from prolonged use (Xiao et al., 62
2005). This necessitates the continued search for potent anti-inflammatory agents with 63
reduced or no side-effects. Studies based on the ethnobotanical use of plants have often 64
proved to be a more efficient method of drug discovery than random plant screening (Slish et 65
al., 1999; Khafagi and Dewedar, 2000). Some plant secondary metabolites such as alkaloids, 66
phenols, tannins, glycosides, terpenoids, saponins, flavonoids and steroids have been 67
implicated in their ability to inhibit the formation of pro-inflammatory signalling molecules 68
such as prostaglandin or leukotrienes (Polya, 2003). In the present study, we evaluated twelve69
medicinal plants used traditionally in the treatment of pain associated with gastro-intestinal 70
infections. The phytochemical components of these plants such as flavonoids, gallotannins, , 71
condensed tannins, other phenolic compounds, alkaloids and saponins were also evaluated.72
The antimicrobial and genotoxicity evaluation of the same plant materials have earlier been 73
reported (Fawole et al., 2009).74
75
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76
2. Material and methods77
78
2.1. Plant material79
80
Twelve traditional medicinal plants that are commonly used for the treatment of gastro-81
intestinal ailments (Table 1) were collected between November (2007) and February (2008)82
from Mt. Gilboa (29º 16.766' S, 30º 17.627' E), Midmar (29º 29.703' S, 30º 12.417' E), 83
University of KwaZulu-Natal Botanical Garden and Pietermaritzburg National Botanical 84
Garden in KwaZulu-Natal Province, South Africa. Due to availability and consideration of 85
potential sustainable harvesting of medicinal plants, the leaves of some plant species were 86
substituted for their roots. Voucher specimens were identified by and lodged in the University 87
of KwaZulu-Natal Herbarium, Pietermaritzburg. Plant materials were oven-dried at 50 ºC, 88
ground into powders and stored in airtight containers at room temperature in the dark.89
90
2.2. Anti-inflammatory activity91
92
2.2.1. Preparation of extracts93
Ground plant materials (5 g) were sequentially extracted with 100 ml of petroleum ether 94
(PE), dichloromethane (DCM) and 70% ethanol (EtOH) in a sonication bath (Julabo GMBH, 95
West Germany) at room temperature for 1 h each. The extracts were then filtered under 96
vacuum through Whatman No.1 filter paper. Water extracts were prepared non-sequentially 97
and freeze-dried while organic extracts were concentrated in vacuo using a rotary evaporator 98
at 30 ºC. The resultant extracts were air-dried at room temperature.99
100
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2.2.2. Cyclooxygenase assays101
The cyclooxygenase assays (COX-1 and COX-2), as described by Eldeen and Van 102
Staden (2008), were used to evaluate the anti-inflammatory activity of the extracts. Crude 103
extracts were screened at a concentration of 250 µg/ml for organic extracts and 2 mg/ml for 104
aqueous extract. Extracts showing good (≥ 50%) COX-2 inhibitory activity were then further 105
evaluated at concentrations of 125 µg/ml and 62.5 µg/ml in both COX-1 and COX-2 assays. 106
Indomethacin (Sigma) (5 µM for COX-1, 200 µM for COX-2) was used as a positive control, 107
while background in which the enzyme was inactivated with HCl before adding [14C] 108
arachidonic acid, and solvent blank were used as negative controls. Percentage inhibition by 109
the extracts was calculated by comparing the amount of radioactivity present in the sample to 110
that in the solvent blank using the equation below:111
112
COX Inhibition (%) = 113
114
where DPMsample = Disintegrations per minute for plant extract115
DPMbackground = Disintegrations per minute in which the enzyme was inactivated 116
DPMblank = Disintegrations per minute for solvent used in dissolving plant extracts117
118
Results are expressed as means ± standard errors of two independent experiments, each 119
experiment in duplicate.120
121
2.3. Phytochemical analysis122
123
2.3.1. Preparation of extracts124
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Phenolic compounds were extracted from plant materials as described by Makkar125
(1999). Dried plant samples (2 g) were extracted with 10 ml of 50% aqueous methanol by 126
sonication in cold water for 20 min. The extracts were then filtered under vacuum through 127
Whatman No.1 filter paper.128
129
2.3.2. Determination of total phenolics130
The amount of total phenolics in plant samples was determined using the Folin 131
Ciocalteu (Folin C.) assay for total phenolics as described by Makkar (1999). Fifty microlitres 132
of each extract from the plant samples were transferred into test tubes and 950 µl distilled 133
water were added to make up to 1 ml, followed by 1 N Folin C. reagent (500 µl) and 2% 134
sodium carbonate (2.5 ml). A blank that contained aqueous methanol instead of plant extracts 135
was also prepared. The test mixtures were incubated for 40 min at room temperature and the 136
absorbance read at 725 nm using a UV- visible spectrophotometer (Varian). Each extract had 137
three replicates and total phenolic concentrations were expressed as gallic acid equivalents138
(GAE).139
140
2.3.3. The butanol-HCl assay for condensed tannins141
Three millilitres of butanol-HCl reagent (95:5 v/v) were added to 500 µl of each extract, 142
followed by 100 µl ferric reagent (2% ferric ammonium sulphate in 2 N HCl). The test 143
combination was vortexed and placed in a boiling water bath for 60 min. The absorbance was 144
then read at 550 nm using a UV- visible spectrophotometer (Varian) against a blank prepared 145
by mixing extract (500 µl) with butanol-HCl reagent (3 ml) and ferric reagent (100 µl), but 146
without heating. Each extract had three replicates. Condensed tannin (% per dry matter) was147
calculated as equivalent amount of leucocyanidins using the formula developed by Porter et 148
al. (1986):149
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150
Condensed tannin = (A550 nm × 78.26 × dilution factor) ∕ (% dry matter)151
152
2.3.4. Rhodanine assay for gallotannins153
Plant extracts (50 µl) were made up to 1 ml with distilled water. One hundred 154
microlitres of 0.4 N sulphuric acid and 600 µl rhodanine were added to the diluted extracts.155
After 5 min, 200 µl of 0.5 N potassium hydroxide were added and 4 ml distilled water after 156
2.5 min. The mixtures were left for a further 15 min at room temperature, after which the 157
absorbance at 520 nm was read using a UV- visible spectrophotometer against a blank test 158
that contained methanol instead of sample. Each extract was evaluated in replicates and 159
gallotannin concentrations were expressed as gallic acid equivalents (GAE) (Makkar, 1999).160
161
2.3.5. Vanillin assay for flavonoids162
Plant extracts (50 µl) (in triplicate), were made up to 1 ml with distilled water in test 163
tubes before adding 2.5 ml methanolic-HCl (95:5 v/v) and 2.5 ml vanillin reagent (1 g/100 164
ml). Similar preparations of a blank that contained methanol instead of plant extracts were 165
made. After 20 min at room temperature, absorbance at 500 nm was read using a UV- visible 166
spectrophotometer (Varian Cary 50) against the blank. The flavonoids in the plant extracts 167
were expressed as catechin equivalents (Makkar, 1999).168
169
2.3.6. Thin layer chromatography for alkaloid detection170
Ten microlitres each of PE, DCM, EtOH and water extracts (50 mg/ml) (prepared as 171
described in Section 2.2.1.) were spotted on thin layer chromatographic (TLC) plates (Silica 172
gel 60 F254, Merck, Germany). The plates were developed using hexane:benzene:ethyl acetate 173
(5:2:3) for PE and DCM extracts, while ethyl acetate:methanol:water (100:16.5:13.5) was 174
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used for EtOH and water extracts. After development, the plates were dried, viewed under UV 175
(254,366 nm) and the fluorescence noted. The presence of alkaloids was indicated by red 176
coloration when the plates were sprayed with Dragendorff reagent (Robert, 1962; Wilfred and 177
Ralph, 2006).178
179
2.3.7. Froth test for saponins180
Distilled water (10 ml) was added to 0.1 g of ground plant sample in a test tube. The test 181
tube was corked and vigorously shaken for 2 min. The appearance of stable foam on the 182
liquid surface for 45 min indicated the presence of saponins (Makkar, 1999). To confirm the 183
presence of saponins, ten drops of olive oil were added to the aqueous extract (2 ml) in a test 184
tube. The test tube was then corked and vigorously shaken. The formation of an emulsion 185
confirmed the presence of saponins (Tadhani and Subhash, 2006).186
187
188
3. Results and discussion189
190
3.1. Anti-inflammatory activity191
192
The percentage inhibition of COX-1 and COX-2 by all the extracts at 250 µg/ml is193
presented in Table 2. Plant extracts showing a minimum inhibition of 50% are considered to 194
have good activity (Eldeen and Van Staden, 2008). All the PE and DCM extracts of the plant195
material (with the exception of Antidesma venosum leaf and Protea simplex bark) showed 196
good activity against both COX-1 and COX-2 (inhibition of prostaglandin synthesis ranging 197
from 58.8 to 103%). Generally, all the ethanol (except Diospyros lycioides leaf and Watsonia 198
tabularis corm) and water extracts showed weak or no activity (inhibition < 50%) against 199
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COX-2. However, water extracts of Agapanthus campanulatus leaf, Becium obovatum root, 200
Cyperus textilis root, Protea simplex leaf and bark, and Watsonia tabularis corm exhibited 201
good activity mainly against COX-1 enzyme with inhibition ranging from 50.3 to 90.5%.202
Most of the plant extracts evaluated showed dose dependent activity against COX-1 203
and/or COX-2 enzymes (Figs 1 and 2 respectively). Interestingly, at 62.5 µg/ml concentration, 204
16 out of 28 PE and DCM extracts evaluated still showed good activity (> 50%) against the 205
COX-2 enzyme. Generally, most of the extracts showed higher percentage inhibition for 206
COX-1 than for COX-2 at the highest screening concentration (250 µg/ml). Although COX-2 207
specific inhibitors have been suggested to be potential classical non-steroid anti-inflammatory 208
drugs due to their reduced or no side effects, some authors have reported that they also have a 209
non-negligible risk of gastro-intestinal toxicity in some patients (MacAulay and Blackburn, 210
2002; Bertin, 2004; Warner and Mitchell, 2008). However, the observed activity in many of 211
the extracts supports their uses in South African traditional medicine. In the case of extracts 212
showing weak or no activity in these assays, high dosages of extract are often used in 213
traditional medicine which may result in COX inhibition. Some of these extracts might be 214
active at other sites in the inflammatory pathways and/or contain compounds showing better 215
activity in vivo as they undergo metabolic transformation (McGaw et al., 1997; Garcia et al., 216
2003). In the human inflammatory process, for example, anti-inflammatory activity of 217
medicinal plants could be manifested in the inhibition of nuclear transcriptase factor (NFкB) 218
mediated signalling pathway in immune cells that lead to the production of inducible nitric 219
oxide synthase (iNOS), pro-inflammatory cytokines and inducible cyclooxygenase (iCOX) 220
(Polya, 2003). Moreover, the presence of comparable activities in the leaves and root or stem 221
of Agapanthus campanulatus, Cyperus textilis, Diospyros lycioides, and Protea simplex222
supports the idea of plant part substitution for sustainable use of many highly threatened 223
plants (Zschocke and Van Staden, 2000).224
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225
3.2. Phytochemical analysis226
227
3.2.1. Total phenolics composition228
Figure 3 shows the total phenolic compounds’ concentrations of the investigated plant 229
species. All the plant species evaluated contained some phenolic compounds. The highest 230
concentration of total phenolics was detected in C. textilis leaf (84.5 mgGAE/g). In addition, 231
B. obovatum root, P. simplex leaf and bark, and C. textilis root contained total phenolic 232
concentrations ≥ 50 mgGAE/g. Phenolic compounds are of important pharmacological value,233
some having anti-inflammatory properties (Bruneton, 1995). Different types of phenolic 234
compounds such as flavonoids, condensed tannins, and gallotannins are known to inhibit 235
some molecular targets of pro-inflammatory mediators in inflammatory responses (Sharma et 236
al., 1994; Iwalewa et al., 2007). Specific types of phenolic compounds present in these 237
species were therefore investigated.238
239
3.2.2. Condensed tannins contents240
The amounts of condensed tannins expressed as percentage per dry matter are shown in 241
Fig. 4. Highest amounts of condensed tannins were detected in C. textilis leaves and roots242
while low levels were detected in Gladiolus dalenii. No condensed tannins were detected in 243
B. obovatum root, A. campanulatus root and Vernonia natalensis leaf. Condensed tannins 244
(proanthocyanidins) are essentially derived from (+) gallocatechin, (–) epicatechin, (+) 245
catechin and epigallocatechin, and their derivatives via carbon to carbon (C-C) links. These 246
compounds are antagonists of particular hormone receptors or inhibitors of particular 247
enzymes such as cyclooxygenase enzymes (Polya, 2003).248
249
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3.2.3. Rhodanine assay for gallotannins250
Figure 5 presents the amounts of gallotannins present in the investigated plants. With 251
the exceptions of A. campanulatus root and G. dalenii corm, gallotannins are detected in all 252
the investigated species. The highest amount of gallotannin was detected in P. simplex leaf253
(13.5 µg/g dry matter). Gallotannins exert various biological effects ranging from anti-254
inflammatory to anticancer and antiviral properties (Erde`lyi et al., 2005). The mechanisms 255
underlying the anti-inflammatory effect of tannins include the scavenging of radicals and 256
inhibition of the expression of inflammatory mediators, such as some cytokines, inducible 257
nitric-oxide synthase, and cyclooxygenase-2 (Polya, 2003; Erde`lyi et al., 2005). The high 258
amounts of gallotannins present in some of the evaluated plant materials could in part be 259
responsible for the observed high anti-inflammatory activity.260
261
3.2.4. Vanillin assay for flavonoids262
The flavonoid concentrations present in the investigated plant materials are shown in 263
Fig. 6. The highest (7.4 mg/g) and the lowest (0.24 mg/g) amounts were detected in C. textilis264
and Haworthia limifolia leaves respectively. According to Talhouk et al. (2007), flavonoids 265
are known to act on the inflammatory response via many routes and block molecules like 266
COX, iNOS, cytokines, nuclear factor-кB and matrix metalloproteinases. Some flavonoids 267
have been reported to be effective against acute inflammation in vivo using a carrageenin-268
induced mouse paw oedema model (Pelzer et al., 1998).269
270
3.2.5. Alkaloids detection271
The presence or absence of alkaloids in the investigated plant extracts are summarized 272
in Table 3. Twelve out of 48 extracts evaluated showed the presence of alkaloids. Previous 273
researchers have reported the presence of alkaloids in A. venosum, Vernonia sp., and D. 274
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lycioides (Watt and Breyer-Brandwijk, 1962; Hutchings et al., 1996; Ndukwe et al., 2004).275
Some alkaloids such as isoquinoline, indole and diterpene are known to have good anti-276
inflammatory activity (Barbosa-Filho et al., 2006).277
278
3.2.6. Saponins detection279
All the evaluated plant materials except H. limifolia, P. simplex, A. venosum and D. 280
princeps leaves tested positive for saponins. The anti-inflammatory activities of some saponin281
derivatives such as triterpenoids saponins have been reported (Sahu and Mahato, 1994).282
According to Sparg et al. (2004), many saponins extracted from plant sources produce an 283
inhibition of inflammation in the mouse carrageenan-induced oedema assay.284
285
286
4. Conclusions287
288
As far as we can ascertain, the anti-inflammatory activity and phytochemical properties289
of many of the investigated plant species have not been reported, yet they are extensively used 290
in traditional medicine. To a large extent, the results in this study validate the traditional 291
medicinal use of the evaluated plant species in treating stomach pains and cramps associated 292
with gastro-intestinal infections. The study of their phytochemical constituents might be 293
considered sufficient for further studies aimed at isolating and identifying the active 294
principle(s) as well as potential combination effects (if any) of the isolated compounds as 295
some of these plants are frequently included in multiple decoctions.296
297
298
Acknowledgements299
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300
Mrs Alison Young of the University of KwaZulu-Natal Botanical Garden and Gary 301
Stafford of the Research Centre for Plant Growth and Development, UKZN are thanked for 302
assistance in plant collection. The National Research Foundation and the University of 303
KwaZulu-Natal are gratefully acknowledged for financial assistance.304
305
306
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15
Naik, S.R., Sketh, U.K., 1976. Inflammatory process and screening methods for anti-349
inflammatory agents- A review. Journal of Postgraduate Medicine 22, 5-21.350
Ndukwe, K.C., Lamikanra, A., Okeke, I.N., 2004. Antibacterial activity in plants used as 351
chewing sticks in Africa. Drugs of the Future 29, 1221.352
Pasinetti, G.M., 2001. Cyclooxygenase and Alzheimer’s disease: implication for preventive 353
initiatives to slow the progression of clinical dementia. Archives of Gerontology and 354
Geriatrics 33, 13-28.355
Pelzer, L.E., Guardia, T., Osvaldo Juarez, A., Guerreiro, E., 1998. Acute and chronic anti-356
inflammatory effects of plant flavonoids. Farmaco 53, 421-424.357
Polya, G.M., 2003. Biochemical targets of plant bioactive compounds. A pharmacological 358
reference guide to sites of action and biological effects. CRC Press, Florida.359
Pooley, E., 1998. A Field Guide to Wild Flowers in KwaZulu-Natal and the Eastern Region. 360
Natal Flora Publication Trust, Durban.361
Porter, L.J., Hrstich, L.N., Chan, B.G., 1986. The conversion of procyanidins and 362
prodelphinidins to cyanidin and delphinidin. Phytochemistry 25, 223-230.363
Robert, F.R., 1962. Simple test for alkaloid-containing plants. Economic Botany 16, 171-172.364
Sahu, N.P., Mahato, S.B., 1994. Anti-inflammatory triterpene saponins of Pithecellobium 365
dulce: characterization of an echinocystic acid bisdesmoside. Phytochemistry 37, 1425-366
1427.367
Sharma, S., Stutzman, J.D., Kellof, G.J., Steele, V.E., 1994. Screening of potential 368
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54, 5848-5855.370
Sleisenger, M.H., Fordtrand, J.S., 1993. Gastrointestinal Disease: Pathophysiology, 371
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Slish, D.F., Ueda, H., Arvigo, R., Balick, M.J., 1999. Ethnobotany in the search for 373
vasoactive herbal medicines. Journal of Ethnopharmacology 66, 159-165.374
Sparg, S.G., Light, M.E., Van Staden, J., 2004. Biological activities and distribution of plant 375
saponins. Journal of Ethnopharmacology 94, 219-243.376
Tadhani, M., Subhash, R., 2006. Preliminary studies on Stevia rebaudiana leaves: proximal 377
composition, mineral analysis and phytochemical screening. Journal of Medical Science 378
6, 321-326.379
Talhouk, R.S., Karam, C., Fostok, S., El-Jouni, W., Barbour, E.K., 2007. Anti-inflammatory 380
bioactivities in plant extracts. Journal of Medicinal Food 10, 1-10.381
Warner, T.D., Mitchell, J.A., 2008. COX-2 selectivity alone does not define the 382
cardiovascular risks associated with non-steroidal anti-inflammatory drugs. Lancet 371, 383
270–273.384
Watt, J.M., Breyer-Brandwijk, M.G., 1962. The Medicinal and Poisonous Plants of Southern 385
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Wilfred, V., Ralph, N., 2006. Phenolic Compound Biochemistry. Springer, Netherlands.387
Xiao, J., Jiang, X., Chen, X., 2005. Antibacterial, anti-inflammatory and diuretic effect of 388
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Journal of Ethnopharmacology 71, 473-478.393
394
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Table 1395
Medicinal plants used against gastrointestinal problems in South Africa.396
Family Species Voucher number Traditional uses
Agapanthaceae Agapanthus campanulatus
Leighton
FAW 4 NU Root decoctions are taken orally or as enemas for stomach
problems in children (Watt and Breyer- Brandwijk, 1962).
Asphodelaceae Haworthia limifolia Marloth FAW 3 NU Decoction made from the leaves is used for stomach trouble
(Hutchings et al., 1996).
Asteraceae Vernonia natalensis Sch. Bip. ex
Walp.
FAW 6 NU Decoctions from leaves and stems are used for stomach
cramps, nervous spasms of the stomach and other stomach
ailments (Hutchings et al., 1996).
Cucurbitaceae Cucumis hirsutus Sond FAW 2 NU Leaf and root decoctions are used for abdominal pain as
well as diarrhoea (Hutchings et al., 1996).
Cyperaceae Cyperus textiles Thunb. FAW 9 NU Root infusions are used as enemas for children with various
stomach pains and cramps (Hutchings et al., 1996).
Ebenaceae Diospyros lycioides Desf. FAW 10 NU Bark and root decoctions are taken for bloody faeces and
dysentery (Hutchings et al., 1996).
Euphorbiaceae Antidesma venosum E. Mey.ex
Tul.
FAW 7 NU Leaf decoctions are used for abdominal cramps and
dysentery. (Hutchings et al., 1996).
Iridaceae Gladiolus dalenii van Geel
FAW 12 NU
Corm ground down to a fine meal and taken this mixed
with warm water in small quantities to relieve dysentery,
diarrhoea and stomach cramps (Margaret, 1990).
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Watsonia tabularis Bak FAW 5 NU Corms are used for diarrhoea in humans and calves
(Hutchings et al., 1996).
Lamiaceae Becium obovatum E. Mey. Ex
Benth.
FAW 8 NU Warm water infusions of pounded roots and leaf are
administered as enemas to treat children with stomach
ailments as well as abdominal pain (Pooley, 1998).
Melastomataceae Dissotis princeps (Kunth) Triana FAW 11 NU Leaf infusions are administered as enemas for dysentery
and diarrhoea (Hutchings et al., 1996).
Proteaceae Protea simplex E. Phillips FAW 1 NU Decorticated root and bark infusions are used for dysentery,
diarrhoea and stomach pains in humans (Hutchings et al.,
1996).
397
398
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Table 2399
Inhibitory activity (COX-1 and COX-2) of different plant extracts evaluated at 250 µg/ml400
Plant species
Plant
part
Percentage inhibition
COX-1 COX-2
PE DCM EtOH Water PE DCM EtOH Water
Agapanthus
campanulatus L 92.6 ± 1.1 78.4 ± 7.2 12.8 ± 0.8 74.2 ± 5.9 72.3 ± 9.1 68.1 ± 3.8 16.9 ± 9.5 47.5 ± 3.7
R 97.7 ± 1.4 98.4 ± 1.0 48.1 ± 9.3 33.4 ± 3.3 78.0 ± 4.4 97.0 ± 1.2 9.1 ± 21.0 28.8 ± 4.0
Antidesma venosum L 103.0 ± 0.8 72.8 ± 4.3 84.3 ± 7.0 36.2 ± 6.1 46.6± 12.3 40.9 ± 9.9 40.9 ± 10.5 0.0
Becium obovatum R 78.5 ± 4.0 86.4 ± 2.6 4.1 ± 3.0 85.3 ± 1.9 76.6 ± 0.8 62.6 ± 9.3 0.0 1.2 ± 17.0
Cucumis hirsutus L 91.7 ± 2.1 101.8 ± 1.1 29.1 ± 5.0 26.0 ± 9.4 80.3 ± 3.5 81.5 ± 4.1 0.0 0.0
Cyperus textilis R 91.7 ± 5.0 88.5 ± 4.9 81.4 ± 8.9 61.3 ± 0.69 75.6 ± 9.8 73.5 ± 2.4 47.9 ± 24.3 0.0
L 86.3 ± 0.7 88.4 ± 0.5 79.8 ± 8.9 0.0 75.0 ± 1.8 83.0 ± 0.1 32.8 ± 1.0 0.0
Diospyros lycioides L 92.8 ± 0.9 94.0 ± 5.7 90.4 ± 4.3 37.1 ± 0.2 91.6 ± 1.9 84.8 ± 1.3 72.0 ± 2.3 0.0
S 73.8 ± 3.6 81.0 ± 2.1 70.6 ± 1.2 13.1 ± 5.7 67.7 ± 0.3 65.9 ± 1.7 37.9 ± 10.1 0.0
Dissotis princeps L 58.8 ± 1.2 82.7 ± 2.3 87.1 ± 4.0 4.8 ± 2.8 60.6 ± 7.7 67.2 ± 9.5 22.1 ± 1.1 0.0
Gladiolus dalenii C 88.3 ± 1.5 101.8 ± 0.3 53.1 ± 4.4 34.4 ± 0.1 68.4 ± 5.7 100.6 ±3.5 35.6 ± 5.7 0.0
Haworthia limifolia L 88.3 ± 3.8 83.5 ± 1.9 1.3 ± 4.1 30.7 ± 10.5 82.4 ± 2.3 72.3 ± 2.8 0.0 0.0
Protea simplex L 100.1 ± 0.8 80.6 ± 8.7 23.7 ± 6.1 57.8 ± 14.4 72.4 ± 1.1 68.4 ± 3.7 0.0 20.9 ± 6.9
B 94.2 ± 3.7 86.1 ± 7.1 89.2 ± 7.8 90.5 ± 1.2 41.0 ± 7.4 35.8 ± 2.4 20.0 ± 2.1 16.7 ± 0.3
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Vernonia natalensis L 88.5 ± 3.2 77.5 ± 3.4 51.2 ± 12.4 38.7 ± 3.3 86.7 ± 0.9 63.4 ± 0.5 0.0 0.0
Watsonia tabularis C 97.6 ± 5.3 73.9 ± 5.1 90.3 ± 3.4 50.3 ± 1.8 91.5 ± 0.9 80.5 ± 8.7 51.1 ± 8.8 0.0
Percentage inhibition of prostaglandin synthesis by indomethacin was 70 ± 3.3 for COX-1 and 68.9 ± 2.5 for COX-2.401
B- bark, C-corm, L-leaf, R-root, S-stem402
403
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Table 3404
Detection of alkaloids in twelve medicinal plants traditionally used for treating gastro-405
intestinal ailments in South Africa.406
Plant name Plant part Extracts
PE DCM EtOH
Agapanthus
campanulatus
Leaves - - +
Roots - -
Antidesma venosum Leaves + - -
Becium obovatum Roots - + -
Cucumis hirsutus Leaves - - +
Cyperus textiles Roots - + -
Leaves - - -
Diospyros lycioides Leaves + - -
Stems - - -
Dissotis princeps Leaves - + -
Gladiolus dalenii Corms - - -
Haworthia limifolia Leaves + - +
Protea simplex Leaves - - +
Bark - - +
Vernonia natalensis Leaves + - -
Watsonia tabularis Corms - - -
- = absence
+ = presence
407
408
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22
Figure Legends409
410
Fig. 1. Dose-dependent COX-1 percentage inhibition by different plant extracts. (A) 411
Petroleum ether extracts (B) Dichloromethane extracts, and (C) Ethanol extracts. Percentage 412
inhibition of prostaglandin synthesis by Indomethacin was 70 ± 3.3.413
A.c = Agapanthus campanulatus; B.o = Becium obovatum; C.h = Cucumis hirsutus; C.t = 414
Cyperus textilis; D.l = Diospyros lycioides; D.p = Dissotis princeps; G.d = Gladiolus dalenii; 415
H.l = Haworthia limifolia; P.s = Protea simplex; V.n = Vernonia natalensis; W.t = Watsonia 416
tabularis.417
418
Fig. 2. Dose-dependent COX-2 percentage inhibition by different plant extracts. (A) 419
Petroleum ether extracts (B) Dichloromethane extracts, and (C) Ethanol extracts. Percentage 420
inhibition of prostaglandin synthesis by Indomethacin was 68.9 ± 2.5.421
A.c = Agapanthus campanulatus; B.o = Becium obovatum; C.h = Cucumis hirsutus; C.t = 422
Cyperus textilis; D.l = Diospyros lycioides; D.p = Dissotis princeps; G.d = Gladiolus dalenii; 423
H.l = Haworthia limifolia; P.s = Protea simplex; V.n = Vernonia natalensis; W.t = Watsonia 424
tabularis.425
426
Fig. 3. Total phenolic compounds per dry matter (DM) of twelve medicinal plants 427
traditionally used for treating gastro-intestinal ailments.428
1-Haworthia limifolia (leaf), 2-Cucumis hirsutus (leaf), 3-Becium obovatum (root), 4-Protea 429
simplex (leaf), 5-Protea simplex (bark), 6-Agapanthus campanulatus (root), 7-Cyperus textilis430
(root), 8-Cyperus textilis (leaf), 9-Vernonia natalensis (leaf), 10-Watsonia tabularis (corm), 431
11-Antidesma venosum (leaf), 12- Diospyros lycioides (leaf), 13-Diospyros lycioides (stem), 432
14-Dissotis princeps (leaf), 15-Gladiolus dalenii (corm).433
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23
434
Fig. 4. Percentage condensed tannins as leucocyanidins equivalents of twelve medicinal 435
plants traditionally used for treating gastro-intestinal ailments.436
1-Haworthia limifolia (leaf), 2-Cucumis hirsutus (leaf), 3-Becium obovatum (root), 4-Protea 437
simplex (leaf), 5-Protea simplex (bark), 6-Agapanthus campanulatus (root), 7-Cyperus textilis438
(root), 8-Cyperus textilis (leaf), 9-Vernonia natalensis (leaf), 10-Watsonia tabularis (corm), 439
11-Antidesma venosum (leaf), 12- Diospyros lycioides (leaf), 13-Diospyros lycioides (stem), 440
14-Dissotis princeps (leaf), 15-Gladiolus dalenii (corm).441
442
Fig. 5. Gallotannin concentrations per dry matter (DM) of twelve medicinal plants 443
traditionally used for treating gastro-intestinal ailments.444
1-Haworthia limifolia (leaf), 2-Cucumis hirsutus (leaf), 3-Becium obovatum (root), 4-Protea 445
simplex (leaf), 5-Protea simplex (bark), 6-Agapanthus campanulatus (root), 7-Cyperus textilis446
(root), 8-Cyperus textilis (leaf), 9-Vernonia natalensis (leaf), 10-Watsonia tabularis (corm), 447
11-Antidesma venosum (leaf), 12- Diospyros lycioides (leaf), 13-Diospyros lycioides (stem), 448
14-Dissotis princeps (leaf), 15-Gladiolus dalenii (corm).449
450
Fig. 6. Flavonoid concentration as catechin equivalent of twelve medicinal plants traditionally 451
used for treating gastro-intestinal ailments.452
1-Haworthia limifolia (leaf), 2-Cucumis hirsutus (leaf), 3-Becium obovatum (root), 4-Protea 453
simplex (leaf), 5-Protea simplex (bark), 6-Agapanthus campanulatus (root), 7-Cyperus textilis454
(root), 8-Cyperus textilis (leaf), 9-Vernonia natalensis (leaf), 10-Watsonia tabularis (corm), 455
11-Antidesma venosum (leaf), 12- Diospyros lycioides (leaf), 13-Diospyros lycioides (stem), 456
14-Dissotis princeps (leaf), 15-Gladiolus dalenii (corm).457
458
459
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460Pe
rcen
tage
Inhi
bitio
n
0
20
40
60
80
100
120
62.5 µg/ml 125 µg/ml 250 µg/ml
Leaf Root Corm Stem
Perc
enta
ge in
hibi
tion
0
20
40
60
80
100
120
Leaf Root Corm StemB
A
Plant species
Perc
enta
ge in
hibi
tion
0
20
40
60
80
100
D.l W.t
Leaf CormC
A.c C.h C.t D.l D.p H.l P.s V.n A.c B.o C.t G.d W.t D.l
A.c C.h C.t D.l D.p H.l P.s V.n A.c B.o C.t G.d W.t D.l
461
462
Figure 1463
464
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25
465Pe
rcen
tage
inhi
bitio
n
0
20
40
60
80
100
62.5 µg/ml 125 µg/ml 250 µg/ml
Perc
enta
ge in
hibi
tion
0
20
40
60
80
100
120
Plant species
Perc
enta
ge in
hibi
tion
0
20
40
60
80
D.l W.t
A.c C.h C.t D.l D.p H.l P.s V.n A.c B.o C.t G.d W.t D.l
A.c C.h C.t D.l D.p H.l P.s V.n A.c B.o C.t G.d W.t D.l
A Leaf Root Corm Stem
LeafB Root Corm Stem
C CormLeaf
466
Figure 2467
468
469
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470
1 2 3 4 5 6 7 8 9 10 11 12 13 14 150
25
50
75
100
Sample
Tot
al p
heno
lic
conc
entr
atio
n(m
g G
AE
/g D
M)
471
Figure 3472
473
474
475
476
477
478
479
480
481
482
483
484
485
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486
1 2 3 4 5 6 7 8 9 10 11 12 13 14 150.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
4.5
Sample
Con
dens
ed ta
nnin
s(%
per
dry
mat
ter)
487
Figure 4488
489
490
491
492
493
494
495
496
497
498
499
500
501
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502
1 2 3 4 5 6 7 8 9 10 11 12 13 14 150123456789
1011121314
Sample
Gal
lota
nnin
con
cent
rati
onµ
g G
AE
/g D
M
503
504
Figure 5505
506
507
508
509
510
511
512
513
514
515
516
517
518
519
520
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521
522
1 2 3 4 5 6 7 8 9 10 11 12 13 14 150
1
2
3
4
5
6
7
8
Sample
Flav
onoi
d co
ncen
trat
ion
(mg
Cat
echi
n eq
uiva
lent
/g D
M)
523
Figure 6524
