Journal

of

Ethnopharmacology.

129

129-133

33 (1991)

Elsevier Scientific Publishers Ireland Ltd.

Antiinflammatory

activity of a Ghanaian preparation: II

antiarthritic

herbal

G. Kweifio-Okai Depurtmenf

of Anatomy

and Physiology,

Phillip Insritute of Technology,

Bundoora 3083 (Australia,l

(Accepted December 31. 1990) A boiling water extract from a mixture of Alsronia boonei and Rauvolfia vomitoria root barks and EIaeis guineensis nuts without pericarp was tested for its antiinflammatory activity by measuring over a period of 17 days the changes in rat ankle diameter caused by subpIantar injection of complete Freund’s adjuvant. The extract fed in drinking water ad libitum reduced ipsilateral ankle adjuvant swelling by an average of 16% for the period of +4 to +17 days and improved weight gain. Key words: arthritis; antiinflammatory;

adjuvant; triterpenes.

Introduction

Despite intensive research into rheumatoid arthritis and the development of numerous new antiarthritic agents over the past two decades, cures of the condition remain elusive (Brune, 1989). The use of inhibitors of phospholipid-derived mediators of inflammation, mainly the cyclooxygenase inhibitors, is limited by their often deleterious side effects. Dietary management has provided only transient benefits (Kremer et al., 1985) while immunomodulation based on demonstrated antibodies to joint components (Hayner et al., 1986; Mollenhauer and Brune, 1988) remains a promising but largely unexplored area of arthritis management. It is therefore useful to evaluate the effects and mode of action of traditional herbal cures in a search for more effective and less deleterious agents in arthritic therapy. In Ghana, West Africa, an aqueous extract of Alstonia boonei de Wild. (Apocynaceae), Rauvolfia vomitoria Afz. (Apocynaceae) and Elaeis guineensis Jacq. (Palmae) is used in treating rheumatoid arthritis (Koranteng et al., personal communication). A. boonei is reCorrespondence to: G. Kweitio-Okai, Department of Anatomy and Physiology, Phillip Institute of Technology, Bundoora 3083, Australia.

0 1991 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland

0378-8741/$03.50

ported to be responsible for the antirheumatoid effect and can be used alone (Dalziel, 1937; Koranteng et al., personal communication). When used in combination with the other two plants, R. vomitoria provides sedative cover (Sofowora, 1982) while E. ~i~eensis reduces the toxicity of R. vomitoria (Boye, G.L., personal communication). In the present paper, a crude extract made from the above three plants is tested for its antiinflammatory activity in CFA-induced polyarthriti~ rats by measuring ankle diameter changes over a period of 17 days. Adjuvant arthritis, developed by Stoerk et al., (1954) and Pearson (1956), has been widely used to screen for drugs useful in human arthritis. Rainsford (1982) has discussed the validity of this technique as an experimental model of human arthritis. Both adjuvant and human arthritis are characterized by T-cell dependent delayed hypersensitivity reactions, both show similar patterns of fibrin deposition and leucocyte infiltration of synovium and bone marrow and both show pannus formation, a granulomatous outgrowth of synovial tissue into joint space which acts as the major factor for joint destruction and fusion in the chronic stages of each condition. However, in adjuvant rats there are no demonstrable immune complexes, e.g. rheumatoid factor in the sera, and no true lymphoid follicles in the synovial tissue. Both of these factors are

130

pathognomic of human arthritis. The implication of these differences in the screening of antiarthritic drugs in adjuvant rats is unclear. Materials and Methods Extract

preparation

A powdered mixture (150 g) containing 90% A. boonei (root bark), 5% R. vomitoria (root bark) and 5% E. guineensis (nuts without pericarp) was added to 1.5 1 of warm water, boiled for 15 min, allowed to settle at room temperature, filtered and 1.25 1 of filtrate diluted with 1.25 I of water. This product is referred to as the drug extract in this paper. The original plant materials were authenticated by G.D. Koranteng, a Ghana Government recognized herbalist and botanist. Animal stock

Three-week-old outbred male Wistar rats, 9&-l 15 g (Monash University Animal House, Clayton, Australia) were used. Animals were kept in groups of 5 at room temperature on a 12: 12 1ight:dark cycle and had free access to food and water prior to the experiments. Introduction

of arthritis

A total of 10 rats was innoculated with complete Freund’s adjuvant (CFA) (Difco, Laboratories, Detroit, MI) containing additional 10 mg/ml of Mycobacterium tuberculosis H37 Ra (Difco). Each rat received a subplantar injection of 150 ~1 of CFA in the midline mid-metatarsal region of the right hindfoot pad. Five control rats received an equivalent amount of saline. All were given free access to food after the subplantar injections. The control rats and 5 adjuvant rats had free access to water while the other 5 adjuvant rats had free access to drug extract. Because the drug extract had a pH of 5.2 + 0.05, the pH of the water was adjusted to 5.2. Anteroposterior diameters of the ankles on the injected and contralateral limbs were measured before and over 17 days after subplantar injections of saline or adjuvant using a sliding vernier scale. Body weights were followed over the same period. Data were expressed as a percentage change from pre-injection values and were analysed using unpaired Student’s t-test.

Results Ankle diameter

changes

Control rats whose right hindpaws have been injected with saline showed an increase in right ankle diameter of 27% on the 17th day after injection (Table 1, control). The increase in ankle diameter is no doubt due to the rapid growth of the threeweek-old rats as seen by the doubling of body weights in 17 days (Table 3). In Table 1 (b - a), adjuvant increased right ankle diameter by at least 29% over the 17 day period. The greatest effect of adjuvant was observed early, a 56% increase on day 2. By the 17th day, the increase was 42”/u. In contrast, adjuvant increased left ankle diameters by 20% and 17% on days 2 and 4 and 9% on day 13. Glenn and Gray (1965) and Van Arman (1976) have identified the experimental conditions conductive to induction of adjuvant arthritis and it is likely that the strain and sex of rats used here may be responsible for the less successful induction of adjuvant arthritis in the contralateral left ankle. In the present paper, the antiinflammatory activity of the extract is evaluated using only the ipsilateral (right ankle) adjuvant swelling because of the magnitude and consistency of swelling over the duration of the experiment. Effects

of extract

Crucial to the interpretation of data on the effects of extract is whether adjuvant rats with free access to extract drank sufficiently to warrant conclusions on the effects of the extract. Table 2 shows that volume of extract drunk by adjuvant rats was marginally greater than the volume of water drunk by the other group of adjuvant rats, ranging from 1I9 to 1I6 ml of extract/kg BW/day compared to 108-150 ml of water/kg BW/day (compare columns 7 and 10, Table 2). Hence even though control rats drank more water per day, ranging from 117 to 188 ml/kg BW/day (column 4, Table 2) the fluid intake in the two groups of adjuvant rats was identical. In Table 1 (c - b) the drug extract had no effect on right ankle adjuvant swelling in the first 2 days. Thereafter, it suppressed adjuvant swelling by an average of 16%.

45 f 7 +56*** -10

5.94 f 0.21

-0.37 +0.21 +44*** -18*

46zt6

20 f 3 64*3

+6

+31*** -l8*

41 f 5

28 t 4 59 f 5

+8

+29*** -12*

50 f 3

33 * 3 62 f 4

+I1

+39*** -18*

50 f 5

29 zt 2 68 f 3

+13

.434*** -II**

57 f 5

34 f 3 68 f 4

+lS

+42*** -17*

52 f 3

27 f 3 69 f 5

+I7 days

-

+2 +4 +6 +8 +I! +I3 +I5 +I7

0

Day

.. -

_

.”

. 11

-

-

_.

_

,,

_

-



,,,

_

529.5 561.1 616.2 672.4 734 850.4 909.4 968.5 1030.1

-

-

508.3 571.0 651.7 712.3 779. I 838.4 941.3 991.8 1068.9 187 188 I70 168 158 I61 132 117

BW (g)

ml/kg BWlday

VOL (ml)

BW (g) 190 215 222 240 370 270 248 232

CFA

Control

,.

\._

.

.-

I55 140 I82 200 330 222 217 210

VOL (ml)

^

-

146 I25 I48 149 150 I31 119 108

-

..d^_

ml/kg BWtday

-7

503.4 536.4 614.9 673.9 742.0 857.7 925.6 999.5 1060.9

BW (g)

CFA + extract

--

_.

160 I78 190 216 364 234 248 238

A

--

-

-,>

159 166 I55 I60 164 136 134 119

ml/kg BWiday (ml)

VOL

-

Tabular values represent total weight of each group of 5 (BW), volume of fluid drank since previous measurement (VOL) and rate of fluid intake expressed as volume drank per weight at previous measurement per 24 h (ml/kg BW per day).

RATS WITH AND WlTHOUT EXTRACT

significant for comparison specified: *P < 0.05, **P < 0,Ol. ***P< 0.001.

+51*** -17*

45 f 6

II f 4 62 f 3

+4

FLUID INTAKE BY CONTROL AND A~JUVANT

TABLE 2

Stat&ally

-1 * 2 55 rfi 7

6.10 rt 0.11 5.73 f 0.14

(a) Control (b) CFA only (c) CFA + extract (b-a) (c-b)

+2

Ankle diameter change (%)

0 Time ankle diameter (mm)

Treatment

Tabular values represent mean =t S.E.M. changes from 0 time readings, N = S/group.

RIGHT ANKLE DIAMETER CHANGES IN CONTROL AND ADJUVANT RATS WITH AND WITHOUT EXTRACT

TABLE I

_-a

-

RATS WITH AND WIT~U~T

CFA f Extract

OIlI)!

+4 -5

101 * II

102 * II 106* 4

(g)

0 time weight

23 f 2 _13*** +7*

-7%” -1

29 4: 2 16 f 1

13 i I 6&l S*t

+4

+2

Body weight increase (‘XI

Statistically signi~~~~t for comparison specified: *P < 0.05,

(c - bf

(b - a)

(c)

(a) Control (b) CFA

Treatment

**P

c: 0.01,

-14** +8**

35 f 3

41 f4 27 f I

+6

***P

< O,Oal,

-I6* +11**

49 * 5

5.5 f $ 39 zk 2

+8

Tabular values represent mean rl; S&M. changes from 0 time readings, N = S/group.

BODY WEIGHT GAIN IN CONTROL AND A~J~VANT

TABLE 3

-6 +I2

73 j; 6

67 k 7 61 zt 3

i-11

EXTRACT

-14 +15**

87 * 7

88 f 9 12 f 4

+13

-16 +19**

102*

90 f 83*

+I5

8

10 4

11 5

-21 +20

115 * 10

115 f 94zt

+I7 days

133

Body weight changes

Table 3 (control) shows a natural weight gain of 115% by control rats on day 17. In Table 3 (b - a), adjuvant significantly reduced body weight gain through day 8, thereafter, up to the 17th day, body weight of adjuvant rats was consistently lower than control rats but the differences were not statistically significant. In Table 3 (c - b), adjuvant rats given the drug extract showed greater increases in body weights than adjuvant rats without the drug extract, with the improvement in weight gain increasing with time. Yet only the improvements in weight gain for day 4 through day 8 and day 13 through day 15 reached statistical significance. Discussion

The present experiments have shown that a boiling water extract prepared from a mixture of A. boonei root bark, R. vomitoria root bark and E. guineensis nuts (without pericarp), as used in Ghana for rheumatoid arthritis, reduced ankle swelling of adjuvant rats when provided in their drinking water. The magnitude of reduction was on average 16% over 17 days. Even though these observations were made on the ipsilateral ankle of the injected foot, it has been shown that inflammatory activity at both ipsilateral and contralateral ankles following subplantar injection of adjuvant tend to be substantially similar (Rainsford, 1982). The present experiments have also shown that at a period of rapid growth of the animals, the extract improved weight gain in adjuvant rats. The improvement in weight gain may be due to an imin the inflammatory condition, provement although Jacka et al. (1983) have shown that this may not necessarily be so. These workers showed that copper salicylate reduced ankle swelling in adjuvant rats at the expense of weight gain. The mechanisms of the extract’s antiinflammatory activity and improvement in weight gain

cannot be deduced from these preliminary studies. However the results should encourage further investigations into the therapeutic potential of this extract and its chemical constituents in arthritic management. Acknowledgement

This work was supported by a Phillip Institite of Technology Research and Development Grant. References Brune, K. (1989) Ten years research

on inflammation

revisited.

Agents und Actions 26, L-8.

Dalziel, J.M. ( 1937) The Useful Plunts of’ West Tropicul A,fiicu. Crown Agents for the Colonies, London, p. 614. Glenn, E.M. and Gray, J. (1965) Adjuvant-induced polyarthritis in rats. Biologic and histologic background. Americun Juurnul of Veterinury Reseurch 26, I I 80- I 194. Hayner, D.C., Gershwin, M.E., Robbins, D.L., Miller, III, J.J. and Costa, D. (1986) Autoantibody profiles in juvenile arthritis. Journul qf Rheumutolugy 13. 358-363. Jacka, T.. Bernard, C.C.A. and Singer. G. (1983) Copper salicylate as an antiinflammatory and analgesic agent in arthritic rats. Lifi Sciences 32, lO23--1030. Kremer, J.M., Michalek, A.Y., Lininger, L., Huyck, C., Bigauquette, J.. Timchalk, M.A., Rynes, R.I., Zieminsi, J. and Bartholomew, L.E. (1985) Effects of manipulation ofdietary fatty acids in clinical manifestations of rheumatoid arthritis. Luncet I, 18L-187. Mollenhauer, J. and Brune. K. (1988) Detection of autoimmuno reactive antibodies against cartilage cell surface proteins in the blood of rheumatic patients. Agents und Actiun,s 23. 48-49. Pearson, C.M. (1956) Development of arthritis, periarthritis and periostitis in rats given adjuvants. Proceeding.r ufrhe SOciery of Experimeniul

Biology und Medicine

Rainsford, K.D. (1982) Adjuvant polyarthritis satisfactory model for screening antiarthritic

9 I, 95-10 I. in rats: Is this a drugs? Agcws

und Actions 12, 452458. SOfOWOra,

A.

(

1982) Medicinal

Plunf.r und Trudilionul

in Africu. Wiley and Sons,

Mc,dicine

Chichester, p. 40, 75-76. Stoerk, H.C., Bielinski, T.C. and Budzilovich, T. (1954) Chronic polyarthritis in rats injected with spleen in adjuvants. Americun Journul oj Pathology 30. 616. Van Arman, C.G. (1976) Pathway to adjuvant arthritis. Federution Proceedings 35. 2442-2446.

Antiinflammatory activity of a Ghanaian antiarthritic herbal preparation: II.

A boiling water extract from a mixture of Alstonia boonei and Rauvolfia vomitoria root barks and Elaeis guineensis nuts without pericarp was tested fo...
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