Evaluation of antipyretic potential of Clitoria ternatea L. extract in rats.

Summary

The methanol extract of Clitoria ternatea L. root (MECTR) blue flowered variety (Family: Fabaceae), was evaluated for its anti-pyretic potential on normal body temperature and yeast-induced pyrexia in albino rats. Yeast suspension (10 ml/kg body wt.) increased rectal temperature after 19 hours of subcutaneous injection. The extract, at doses of 200, 300 and 400 mg/kg body wt., p.o., produced significant reduction in normal body temperature and yeast-provoked elevated temperature in a dose-dependent manner. The effect extended up to 5 hours after the drug administration. The anti-pyretic effect of the extract was comparable to that of paracetamol (150 mg/kg body wt., p.o.), a standard anti-pyretic agent.

Key words: Root of Clitoria ternatea (MECTR), Anti-pyretic

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Introduction

Clitoria ternatea L. (Family: Fabaceae) is a well-known perennial, twining herb, found abundantly in Indochina, the Phillipines and Madagascar. This plant is known commonly as "Aparajit" in Hindi and "Aparajita" in Bengali, and as Kakkattan in Tamil. The root has a sharp bitter taste. The root part of this plant is used traditionally for its laxative, diuretic, anti-inflammatory and anthelmintic activities. It is also useful in the treatment of severe bronchitis, asthma and hectic fever (Kirtikar and Basu, 1975), and is used by the local tribal people to cause abortion; paste is applied for curing abdominal swellings, sore throat, mucous disorders and fever (Asolkar et al. 1992; Nadkarni and Nadkarni 1976). Root juice is given with cold milk to remove phlegm in chronic bronchitis. Various parts of this plant, including the root, stem and flower, are recommended for the treatment of snake-bite. The Root juice is given for the relief of fever by rural people of Jhilimili, Bankura, West Bengal. The main objective of this work was to study the anti-pyretic activity of Clitoria ternatea L. root extract in an experimental animal model using rats to verify the action of the herb claimed by the rural people.

Materials and Methods

Plant material

Roots of Clitoria ternatea Linn. were collected from the Jhilimili, Bankura, West Bengal, India. Taxonomical identification of the plant was made by the Botanical Survey of India, Shibpur, Howrah, and the voucher specimen was preserved in our laboratory for future reference. The roots of the plants were dried under controlled temperature, powdered and passed through a # 40 mesh sieve and stored in an airtight container for further use.

Extraction procedure

The powdered roots were extracted using methanol in a Soxhlet extraction apparatus. The solvent was removed in vacuo to provide a dry extract (9.8% w/w, as compared to the powdered material). The chemical constituents of the methanol extract were identified by qualitative analysis and confirmed by thin layer chromatography for the presence of flavonoids, tannins, steroids and saponins. The extract was stored in a refrigerator and a weighed quantity was suspended in 2% Tragacanth solution for the experiment.

Animals used

Albino rats (Wistar strain) of either sex weighing 180-200 g each supplied by M/s. B.N. Ghosh and Co., Calcutta, India. were used in this study. The animals were maintained under suitable nutritional and environmental conditions throughout the experiment, and were maintained under standard laboratory conditions prior to the experiments.

Toxicity study

Acute toxicity relating to the determination of L[D.sub.50] value was performed with different doses of the extract according to the method described by Ghosh (1984).

Study on normal body temperature

Rats of either sex were divided into five groups of six each for this experiment. The body temperature of each rat was measured rectally at predetermined intervals before and for 5 h after administration of either 2% aqueous tragacanth solution (control) or MECTR at doses of 200, 300 and 400 mg/kg body wt., orally (Murugesan et al. 2000).

Induction of yeast-induced pyrexia

Rats were divided into five groups of six rats each. The normal body temperature of each rat was measured rectally at predetermined intervals and recorded. Fever was induced per the method described by Smith and Hambourger (1935). The rats were trained to remain quiet in a restraint cage. A thermister probe was inserted 3-4 cm deep into the rectum and fastened to the tail by adhesive tape. The temperature was measured on a thermometer. After measuring the basal rectal temperature, animals were given a subcutaneous injection of 10 ml/kg body wt. of 15% w/v yeast suspended in 0.5% w/v methyl cellulose solution. Rats were then returned to their housing cages. After 19 h of yeast injection, the animals were again restrained in individual cages for another recording of their rectal temperatures as described above.

Drug administration

After 19 h of yeast injection, the MECTR was administered orally at doses of 200, 300, and 400 mg/kg body wt. to three groups of animals, respectively. A similar volume (5 ml/kg body wt.) of 2% aqueous tragacanth solution was administered orally to the control group. The fifth group of animals received the standard drug paracetamol (150 mg/kg body wt.) orally. Rats were restrained for recording of their rectal temperatures at the nineteenthhour, immediately before MECTR, saline or paracetamol administration, and again at one-hour intervals up to the twenty-third hour after yeast injection.

Statistical analysis

The data were analyzed for significance using the unpaired two-tailed student's t-test (Woodson, 1987).

Results

Toxicity study

In the L[D.sub.50] value determination, we observed that the extract is safe to use in animals even at a dose of 3.2 g/kg (see Table 1).

Effects of normal and yeast-induced pyrexia

The effect of the MECTR on normal body temperature in rats is presented in Table 2. It was found that the MECTR at doses of 200 mg/kg body wt. caused significant lowering of body temperature up to 4 h following its administration. This effect was maximal at 4 hrs and doses of 300 and 400 mg/kg body wt. in a dose-dependent manner and caused significant lowering of body temperature up to 5 h after its administration. The subcutaneous injection of a yeast suspension elevated the rectal temperature markedly after 19 h of administration. Treatment with the MECTR at doses of 200, 300 and 400 mg/kg body wt. decreased the rectal temperature of the rats in a dose-dependent manner. The antipyretic effect started as early as 1 h, and the effect was maintained for 4 h, after its administration. The standard drug paracetamol at 150 mg/kg body wt. reduced the yeast-provoked elevation of body temperature significantly. The results obtained for standard, drug-treated and MECTR-treated rats were compared with the control (2% aqueous tragacanth solution) group and we observed a significant reduction in the yeast-elevated rectal temperature (see Table 3).

The present results show that the methanol extract of Clitoria ternatea (MECTR) possesses a significant antipyretic effect in yeast-provoked elevation of body temperature in rats, and its effect is comparable to that of paracetamol (standard drug). Furthermore, the MECTR also reduced normal body temperature significantly, and this effect will be studied further to ascertain the exact mechanism of action.

Discussion

This study examined the antipyretic activity of a methanol extract of the root of Clitoria ternatea L. (MECTR) in an experimental animal model using rats. We observed that MECTR lowers the body temperature in a dose-dependent manner up to 5 h after its administration. Fever was induced as described by Smith and Hambourger (1935) and Murugesan et al. (2000). Fever may be a result of infection or one of the sequelae of tissue damage, inflammation, graft rejection, or other disease states. Antipyretics are drugs which reduce elevated body temperature. Regulation of body temperature requires a delicate balance between the production and loss of heat, and the hypothalamus regulates the set point at which body temperature is maintained.

In fever this set point is elevated, and drugs like paracetamol do not influence body temperature when it is elevated by factors such as exercise or increases in ambient temperature (Goodman and Gilman 1996). The present study reveals that the root extract of Clitoria ternatea L. causes a significant antipyretic effect in yeast-provoked elevation of body temperature (Table 3) as well as normal body temperature in rats (Table 2). In both the cases, the extract caused a significant lowering of body temperature, with the effect being comparable to that of paracetamol. Thus, the present pharmacological evidence provides support for the folklore claim as an anti-pyretic agent.

Table 1. Toxicity study of MECTR in mice.

Treatment  Dose (mg/kg body  No. of    No. of     No. of  L[D.sub.50]
           wt.)              animals   survivals  deaths  value

Control     Tween 80          20        20         0        -
            Solution
MECTR        100              20        20         0        -
             200              20        20         0        -
             400              20        20         0        -
             800              20        20         0        -
            1600              20        20         0        -
            3200              20        20         0        > 3.2 g/kg

MECTR -- Methanol Extract of Clitoria ternatea L. roots.

Table 2. Effect of MECTR on normal body temperature.

Treatment               Rectal temperature ([degrees]C) before and after
                        treatment
                        0 h                1 h

Control                 37.4 [+ or -] 0.1  37.2 [+ or -] 0.1
  (5 ml/kg body wt.)
MECTR                   37.2 [+ or -] 0.1  36.6 [+ or -] 0.2 (b)
  (200 mg/kg body wt.)
MECTR                   37.3 [+ or -] 0.1  36.2 [+ or -] 0.1 (a)
  (300 mg/kg body wt.)
MECTR                   37.1 [+ or -] 0.1  35.9 [+ or -] 0.2 (a)
  (400 mg/kg body wt.)

Treatment               Rectal temperature ([degrees]C) before and after
                        treatment
                        2 h                    3 h

Control                 37.2 [+ or -] 0.2      37.3 [+ or -] 0.2
  (5 ml/kg body wt.)
MECTR                   36.7 [+ or -] 0.1 (b)  36.6 [+ or -] 0.2 (b)
  (200 mg/kg body wt.)
MECTR                   36.2 [+ or -] 0.2 (a)  36.3 [+ or -] 0.2 (a)
  (300 mg/kg body wt.)
MECTR                   35.8 [+ or -] 0.1 (a)  35.7 [+ or -] 0.1 (a)
  (400 mg/kg body wt.)

Treatment               Rectal temperature ([degrees]C) before and after
                        treatment
                        4 h                    5 h

Control                 37.2 [+ or -] 0.2      37.3 [+ or -] 0.2
  (5 ml/kg body wt.)
MECTR                   36.4 [+ or -] 0.2 (b)  36.8 [+ or -] 0.2 (b)
  (200 mg/kg body wt.)
MECTR                   36.5 [+ or -] 0.2 (b)  36.6 [+ or -] 0.2 (b)
  (300 mg/kg body wt.)
MECTR                   35.7 [+ or -] 0.1 (a)  35.8 [+ or -] 0.1 (a)
  (400 mg/kg body wt.)

Each value represents mean [+ or -] SE (N = 6).
Control (2% aqueous tragacanth solution).
(a) P < 0.001, (b) P < 0.01 significant as compared to control values at
corresponding hour.
MECTR -- Methanol Extract of Clitoria ternatea L. roots.

Table 3. Effects of MECTR on Yeast-induced pyrexia in rats.

Treatment               Rectal temperature ([degrees]C) after Yeast
                        injection
                        0 h                 19 h

Control                 37.7 [+ or -] 0.02  39.8 [+ or -] 0.02
  (5 ml/kg body wt.)
Paracetamol             37.9 [+ or -] 0.01  39.8 [+ or -] 0.04
  (150 mg/kg body wt.)
MECTR                   37.7 [+ or -] 0.03  39.5 [+ or -] 0.02
  (200 mg/kg body wt.)
MECTR                   37.5 [+ or -] 0.02  39.7 [+ or -] 0.01
  (300 mg/kg body wt.)
MECTR                   37.6 [+ or -] 0.04  39.9 [+ or -] 0.02
  (400 mg/kg body wt.)

Treatment               Rectal temperature ([degrees]C) after Yeast
                        injection
                        20 h                    21 h

Control                 39.3 [+ or -] 0.07      39.1 [+ or -] 0.06
  (5 ml/kg body wt.)
Paracetamol             38.6 [+ or -] 0.02 (a)  38.2 [+ or -] 0.03 (a)
  (150 mg/kg body wt.)
MECTR                   39.1 [+ or -] 0.02 (b)  38.8 [+ or -] 0.01 (a)
  (200 mg/kg body wt.)
MECTR                   38.7 [+ or -] 0.04 (a)  38.1 [+ or -] 0.04 (a)
  (300 mg/kg body wt.)
MECTR                   38.4 [+ or -] 0.02 (a)  37.7 [+ or -] 0.03 (a)
  (400 mg/kg body wt.)

Treatment               Rectal temperature ([degrees]C) after Yeast
                        injection
                        22 h                    23 h

Control                 39.4 [+ or -] 0.05      39.4 [+ or -] 0.04
  (5 ml/kg body wt.)
Paracetamol             37.8 [+ or -] 0.04 (a)  37.2 [+ or -] 0.03 (a)
  (150 mg/kg body wt.)
MECTR                   38.2 [+ or -] 0.04 (a)  37.9 [+ or -] 0.05 (a)
  (200 mg/kg body wt.)
MECTR                   37.9 [+ or -] 0.05 (a)  37.7 [+ or -] 0.04 (a)
  (300 mg/kg body wt.)
MECTR                   37.5 [+ or -] 0.01 (a)  37.4 [+ or -] 0.04 (a)
  (400 mg/kg body wt.)

Each value represents mean [+ or -] SE (N = 6).
Control (2% aqueous tragacanth solution).
(a) P < 0.001, (b) P < 0.01 significant as compared to control values at
corresponding hour.
MECTR -- Methanol Extract of Clitoria ternatea L. roots.

Acknowledgement

The authors are grateful to University Grant Commission New Delhi for financial assistance. We are also thankful to Mr. V. Ramaswamy for collecting plant material.

References

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Murugesan T, Mandal SC, Bhakta T, Das J, Pal M, Saha BP (2000) Evaluation of anti-pyretic potential of Jussiaea suffruticosa L. extract in rats. Phytomedicine 7(3): 231-234

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Smith PK, Hambourger WE (1935) The ratio of the toxicity of acetanilamide to its antipyretic activity in rats. J Pharmacol Exp Ther 54: 346

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B. Parimaladevi, R. Boominathan, and Subhash C. Mandal

Division of Pharmacognosy and Phytochemistry, Department of Pharmaceutical Technology, Faculty of Engineering and Technology, Jadavpur University, Calcutta, India

Address

Subhash C. Mandal, Division of Pharmacognosy and Phytochemistry, Department of Pharmaceutical Technology, Faculty of Engineering and Technology, Jadavpur University, Calcutta-700 032, India

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