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1 Research Paper Antipyretic, antidiarrhoeal, hypoglycaemic and hepatoprotective activities of ethyl acetate extract of Acacia catechu Willd. in albino rats D. Ray, Kh. Sharatchandra*, I.S. Thokchom** ABSTRACT Objective: To evaluate the antipyretic, antidiarrhoeal, hypoglycaemic and hepatoprotective Department of Pharmacology, effects of the ethyl acetate extract of Acacia catechu in experimental animal models. Agartala Govt. Medical College, Materials and Methods: Ethyl acetate extract of Acacia catechu was evaluated for antipyretic Po. Kunjavan, activity in yeast induced pyrexia and for antidiarrhoeal activity in castor oil induced Agartala , Tripura, diarrhoea in albino rats. Hypoglycaemic activity was studied in both normal and alloxan *Department of Pharmacology, (120 mg/kg, s.c.) induced diabetic albino rats. The hepatoprotective potential of Acacia Regional Institute of Medical catechu was evaluated by CCl Sciences, Imphal, Manipur induced hepatotoxicity in albino rats. Results: Single administration of the ethyl acetate extract of Acacia catechu at doses of 004 and **Department of 250 and 500 mg/kg, p.o. showed significant antipyretic activity (P<0.01) in albino rats. Pharmacology, Sikkim-Manipal Institute of Medical Sciences, Acacia catechu at a dose of 250 mg/kg, p.o., (single dose) has been found to possess 5 th Mile, Tadong highly significant antidiarrhoeal property (P<0.001) in respect of latent period of onset of diarrhoea, average number of stool passed and purging index. Significant reduction of Received: blood glucose level was observed in nondiabetic albino rats following single dose treatment Revised: with the test drug at a dose of 500 mg/kg, p.o. (P<0.01). Significant reduction of blood Accepted: glucose level was also evident in diabetic rats at doses of 250 and 500 mg/kg (P<0.001). Highly significant hepatoprotective activity was also observed when the extract of Acacia Correspondence to: catechu (250 mg/kg) was administered prophylactically for seven days (P<0.001). D. Ray dr_raydebasis@yahoo.co.in Introduction Conclusion: The present study shows that ethyl acetate extract of Acacia catechu (cutch/ katha) has significant antipyretic, antidiarrhoeal, hypoglycaemic and hepatoprotective properties. KEY WORDS: Antidiabetic, cutch, pyrexia, liver. The diverse culture of our country is a rich source of traditional medicines, many of which are of plant origin. Scientific data on such plant derivatives could be of clinical use. [1] Catechu or cutch (Katha in Hindi and Manipuri), the extract prepared from the hard wood of Acacia catechu, has been used for treating fever, diarrhoea, leucorrhoea, piles and erysipelas. [2] The juice of its fresh bark has been used in treatment of haemoptysis and gonorrhoea. Catechuic acid found in cavities of the wood of the Acacia catechu tree (Leguminosae) was valued for facilitating expectoration in chest infection. [3] Catechu contains catechuic acid, catechutannic acid (25% 33%), acacatechin (10% 12%), catechu red, quercetin, catechin (2% 12%), epicatechin, phlebotanin (25% 33%), gummy matter, quercitrin, quercitin and moisture. Quercitin is a phenolic flavonoid and catechu of acacia is a pseudotanin. Catechu [3, 4] and epicatechin usually accompany other flavonoids. It is reported that Acacia catechu has hypoglycaemic activity [5] and catechu, a product of Acacia catechu, has hepatoprotective, antipyretic and digestive properties. [6,7] Cyanidanol, an active principle of Acacia catechu, is claimed to be effective in treating liver diseases. [8] Catechu was used in the treatment of diarrhoea and throat infection [9] because the tannin and polyphenols present in it impart astringent activity. Considering the above facts, the present study was undertaken to evaluate the antipyretic, antidiarrhoeal, hepatoprotective and hypoglycaemic properties of Acacia catechu in experimental animal models. Materials and Methods Collection and preparation of extract Catechu (250 g) was collected (from the local market during the month of October), identified and differentiated from [10, 11] Uncaria gambier by gambier fluorescin test. 408 Indian J Pharmacol December 2006 Vol 38 Issue

2 Studies on A. catechu The ethyl acetate extract of Acacia catechu was obtained Antipyretic activity by the procedure as described by Jayasekhar et al, [6] and Antipyretic activity on albino rats was studied with fever Seikel. [12] The powdered catechu was defatted with a nonpolar induced by 20% Brewer s yeast as described. [14, 15] Albino rats organic solvent such as petroleum ether (40 0 C 60 0 C) to remove ( g) were fed uniformly till 24 h before giving drugs, the phenolic and nonpolar substances from the dried material. when food was withdrawn. After measuring rectal temperature After defatting, catechu was extracted with 95% ethanol. Then of the animals by introducing 1.5 cm of digital thermometer in the dried ethanol extract was again extracted with ethyl acetate rectum, pyrexia was induced by injecting, subcutaneously, 20% to concentrate the minute amounts of phenolic materials suspension of dried yeast in 2% gum acacia in normal saline present. The yield at the end of extraction was 29.2%. at a dose of 20 ml/kg of body weight. After 18 h of yeast Phytochemical studies injection, rats which showed a rise in temperature of at least Tests for presence of flavonoid compounds in the catechu 1 0 F (0.6 o C) were taken for the study. Animals in the various were done according to the methods described by Kokate [11] groups were treated as follows. and Seikel. [12] However, identification and estimation of different Group A: 3% aqueous suspension of gum acacia (1 ml/200 g) flavonoid compounds like catechin, quercetin and cyanidol were not done. Animals Colony bred, healthy, Wistar strain, albino rats of either sex weighing g were used for the study. They were housed in standard polypropylene cages, under room temperature (24±2 0 C); relative humidity (60% 70%) and exposed to 12:12 h light:dark cycle. The rats were fed with standard diet and water ad libitum. The ethics committee, Regional Institute of Medical Sciences, Imphal, approved the protocol of the present study. Drugs The following chemicals (analar/gr) and drugs were used: gum acacia, castor oil, tablet Lomotil (diphenoxylate 2.5 mg + atropine sulphate mg per tablet), acetyl salicylic acid, dried yeast, carbon tetrachloride (Qualizen), olive oil, silymarin (Micro Lab), glibenclamide (Aventis) and alloxan monohydrate. as vehicle, orally. Group B: Aqueous suspension of ethyl acetate extract of Acacia catechu 250 mg/kg (1 ml/200 g) with 3% gum acacia as 5% suspension, orally. Group C: Aqueous suspension of ethyl acetate extract of Acacia catechu 500 mg/kg (1 ml/200 g) with 3% gum acacia as 10% suspension, orally. Group D: Aqueous acetyl salicylic acid, 300 mg/kg (1 ml/200 g) with 3% gum acacia as 6% suspension, orally. Rectal temperature was recorded every hour for four hours after administration of drugs. Hepatoprotective activity Hepatoprotective activity of Acacia catechu was studied [6] using the method described by Jayasekhar et al. Blood was collected from the orbital sinus of albino rats and allowed to coagulate for 30 min. Serum was separated by centrifugation at 2500 rpm for 20 min and aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were assayed. AST and ALT levels were estimated from blood by the widely used method of Reitman and Frankel. [27] The protocol for the study Antidiarrhoeal studies Antidiarrhoeal activity was evaluated as described by Yegnanarayan and Shrotri [13] with slight modification to ensure of hepatoprotective activity is shown in Table 1. evenness of dose of castor oil in rats according to body weight. Fifty albino rats were screened initially by administering 1 ml Table 1 of castor oil orally and those (32) which developed diarrhoea were selected (consistency, i.e. loose stool, was the criterion for selection). The experimental set-up was as follows. Group A (Control):received 3% aqueous gum acacia suspension 1 ml/200 g, p.o., at 0 hour and castor oil 1 Group Treatment (1-7) days ml/150 g, p.o., one hour later. Protocol for study of hepatoprotective activity of Acacia catechu in albino rats Toxicant on 7 th day single dose Group B (Test): received ethyl acetate extract of Acacia Control Aqueous 5% gum acacia Olive oil catechu 250 mg/kg as 5% suspension in 3% 1 ml / 200 g 0.12 ml / 100 g gum acacia in DW (1 ml/200 g, p.o.) at 0 hour and castor oil 1 ml/150 g, p.o., one hour Toxic control Aqueous 5% gum acacia CCl 4 + Olive oil (1:1) later. 1 ml/200 g 0.25 ml /100 g Group C (Standard): received diphenoxylate 10 mg/kg (0.2%) with atropine sulphate (0.002%) suspension in 3% gum acacia in distilled water at a dose of 1ml/ 200 g, p.o. and castor oil 1ml/150 g, p.o, one hour later. Percent respondents X Average number of stools of respondents Purging index (PI) = Average latent period of onset of diarrhoea Standard 0.5% silymarin 25 mg/kg CCl 4 +Olive oil (1:1) 1 ml/200 g 0.25 ml/100 g Test drug A catechu extract 250 mg/kg CCl 4 +Olive oil (1:1) in 5% gum acacia 0.25 ml/100 g Estimation of AST and ALT was done on 8th day of experiment. All the drugs were administered orally. CCl 4 was used as toxicant with olive oil. n=6 in each group. Indian J Pharmacol December 2006 Vol 38 Issue

3 Ray et al. Study of hypoglycaemic activity of Acacia catechu Hypoglycaemic activity of Acacia catechu was studied both in normal and alloxan induced diabetic rats. Hypoglycaemic activity in normal rats Twenty-four albino rats weighing g were fasted for 18 h and were divided into four groups of six animals in each. The groups included i) vehicle control (5% gum acacia in normal saline, 1 ml/200 g rat); ii) test drug (250 mg/kg, p.o. 5% w/v, 1 ml/200 g rat); iii) test drug (500 mg/kg, p.o., 10% w/v, 1 ml/200 g rat) and iv) standard control, glibenclamide (0.5 mg/kg, p.o., 1% 1 ml/200 g rat). One millilitre of blood from the orbital sinus of each rat was collected at 0 hour. At two hours of treatment, blood samples were collected again from the treated animals and blood glucose was estimated by glucose oxidase method. [16] Hypoglycaemic activity in diabetic rats Albino rats (n = 44) were fasted for 48 h. Diabetes was induced by administering [5, 17-19, 25, 26] freshly prepared alloxan monohydrate 2.4% in normal saline subcutaneously at a dose of 120 mg/kg, body weight as single dose. [17] After 72 h of alloxan, 18 h fasting blood was collected from those that survived (n=34) [18] and blood sugar estimated by glucose oxidase method. Twenty-four diabetic rats with blood glucose level of mg% were selected and were divided into four groups of six each. The selected groups were treated with the vehicle (5% gum acacia, 1 ml/200 g), test drug (250 mg/ kg, p.o.), test drug (500 mg/kg, p.o.) and glibenclamide (0.5 mg/kg, p.o.), respectively, for seven days. [19] On the eighth day blood samples were collected after 18 h of fasting and blood glucose was estimated again. Statistical analysis The results were analysed for statistical significance using one-way ANOVA followed by Dunnett s test. Nonparametric data were analysed by Kruskal Wallis one-way ANOVA. P values <0.05 were considered significant. Results Antidiarrhoeal studies The extract of Acacia catechu (250 mg/kg) markedly reduced the percentage of animals that had diarrhoea (50%). Table 2 The latent period (5.25±0.31) was prolonged and average number of stool passed was significantly reduced (0.7±0.26), leading to very low purging index (16.28) compared to the control group. Both the standard and test drugs brought a highly significant (P<0.001) change in the latent period of onset of passing stool as well as in total number of stools passed. [Table 2] Antipyretic activity Table 3 shows the effect of Acacia catechu in yeast induced pyretic rats. There was no significant difference in the basal temperature at 0 hour between the different groups. The ethyl acetate extract of Acacia catechu and aspirin significantly decreased the temperature of pyretic rats at 2nd, 3rd and 4th h after drug or extract treatment. Hepatoprotective activity Significant increase in the levels of AST and ALT were found in the toxicant group (P<0.001) after 24 h of administration of CCl 4 orally. Pretreatment with Acacia catechu (250 mg/kg) and silymarin (25 mg/kg) in test group and standard group, respectively, daily for seven days showed highly significant (P<0.001) protective effect against CCl induced hepatotoxicity 4 when compared to toxicant alone group. There was no significant difference (P<0.5) between the protective ability of the test drug and the standard drug silymarin, when compared. [Table 4] Hypoglycaemic activity The hypoglycaemic activity of ethyl acetate extract of Acacia Catechu in normal (nondiabetic) and diabetic rats is shown in Table 4. The test drug, at a dose of 500 mg/kg, p.o. significantly lowered the blood glucose level (P<0.01) as compared to the control group, at 2 h. However, the activity of the standard drug, glibenclamide (0.5 mg/kg/day), was more pronounced (P<0.001). In alloxan induced diabetic albino rats, Acacia catechu at a dose of 250 and 500 mg/kg/day and standard drug glibenclamide (0.5 mg/kg/day) for seven days was highly significant (P<0.001) in comparison with the control group. However, in diabetic rats the hypoglycaemic effect of the test drug at 250 mg/kg was significantly less than the standard drug glibenclamide. Antidiarrhoeal activity of Acacia catechu in albino rats Group Dose and route Mean latent period Mean total no. of Purging index (h) stools passed Control 1 cc/200 g, p.o. 2± ± % Gum acacia Test 250 mg/kg, p.o. 5.25±0.31** 0.7±0.26** A. catechu Standard 10 mg/kg, p.o. 5.95±0.05** 0.2±0.13** 3.48 Diphenoxylate One-way F ANOVA P <0.001 < Values are mean±sem; n=10 in each group. df=2,27. **P<0.001 as compared to control. Latent period of the animals which did not pass stool within 6 h was considered as 6 h. 410 Indian J Pharmacol December 2006 Vol 38 Issue

4 Studies on A. catechu Table 3 Antipyretic activity of Acacia catechu in albino rats Group Drug and dose Temperature ( o F) at -18 h 0 h 1 h 2 h 3 h 4 h Control 3% gum acacia 99.07± ±0.42* ± ± ± ±0.13 in DW, 1ml/200 g Test A. catechu 99.11± ±0.25** ± ±0.33 a ±0.22 a 100.6±0.22 aa 250 mg/kg 500 mg/kg 99.20± ±0.41* 100.9± ±0.39 a 100.3±0.43 aa ±0.36 aa Standard Aspirin 98.98± ±0.27* ± ±0.29 aa 98.68±0.37 aaa 98.61±0.25 aaa 300 mg/kg One-way F ANOVA P < < Values are mean±sem; n=6 in each group, df=3,23. *P<0.01; **P<0.001 as compared to initial basal temperature. a P<0.05; aa P<0.001 as compared to respective control value. Table 4 Castor oil induced model of diarrhoea incorporates both secretory and motility diarrhoea as a result of prostaglandin Hepatoprotective activity of Acacia catechu in albino rats release from the intestinal mucosa. [13] Acacia catechu Group AST on 8 th day ALT on 8 th day significantly decreased the number of stool passed (P<0.001). (i.u./l) (i.u./l) The percentage of respondents and purging index were also significantly decreased. Highly significant (P<0.001) increase Control 5% Gum acacia 14.7± ±2.69 in the latent period of onset of diarrhea, as compared to the in DW 1 cc/200 g, p.o. control group, was also produced. Parameters in the control Toxicant (CCl 4 ) 2.5 mg/kg, p.o ±19.23* 838.8±30.23* group of animals agree with the findings of previous workers. [13] The result shows that the ethyl acetate extract of Acacia A.Catechu 330.7±31.76** 343.5±38.4** a catechu (test drug) is as effective as the standard drug, diphenoxylate, in respect of total number of stool passed Test 250 mg/kg, p.o. Standard Sylimarin, (25 mg/kg) One-way ANOVA (P>0.05). The antidiarrhoeal property of the ethyl acetate 242.3±37.8** 263.5±36.77** extract of Acacia catechu appears to be due to its tannin [9] F P 0.99 < Data shown are mean ± SEM. n=6 in each group, df=3,23. a P<0.5 as compared to standard. *P<0.001 as compared to control, **P<0.001 as compared to toxic control. Discussion content, which has astringent property. Results of the present study shows that ethyl acetate extract of Acacia catechu (500 mg/kg) significantly decreases fasting glucose levels of normal rats (P<0.001). However, the reduction was found to be less effective than that of glibenclamide. [Table 5] The blood glucose levels in normal and diabetic albino rats before administration of the drugs correspond well with the findings of previous workers. [25] The test drug at doses of 250 and 500 mg/kg/day for seven days reduced the blood glucose level of diabetic rats significantly (P<0.001). Effect of the test drug at doses of 250 and 500 mg/kg, orally, on blood glucose level was comparable with that of the standard drug, glibenclamide. The hypoglycaemic effect of A. catechu may be due to presence of flavonoids which acts as insulin secretagogues. [18] Epicatechin, a flavonoid compound, is reported to promote regeneration of [1, 5, 19, 23] ß cells of the Islets of Langerhans. Carbon tetrachloride is a hepatotoxin commonly used for the production of experimental liver toxicity. The serum transaminase level is most widely used as a measure of hepatic injury, due to its ease of measurement and high degree of sensitivity. It is useful for the detection of early damage of hepatic tissue and requires less effort than that required for a In the present study, the mean initial basal rectal temperature of rats before yeast injection corresponds with findings of other workers. [20,21] In our study, the initial rise of temperature after 18 h of subcutaneous yeast injection was 1.71 F 2.1 F (0.95 o C to 1.67 o C), which corresponds with the findings of Hajare et al. [15] There was no significant difference between the initial mean basal temperature of the different groups and the mean temperature between the groups of pyrexia rats, after 18 h of yeast injection. Body temperatures of pyrexia rats were lowered significantly with the test drug. The antipyretic effect of the test drug may be due to presence of flavonoid compounds, as some flavonoids are predominant [4, 22, 23] inhibitors of cyclooxygenase or lipooxygenase. Indian J Pharmacol December 2006 Vol 38 Issue

5 Ray et al. Table 5 Hypoglycaemic activity of Acacia catechu in normal and diabetic albino rats Blood glucose - in normal rats Blood glucose - in diabetic rats Group fasting 2 h after treatment before treatment after 7 days treatment Control ± ± ± ± % Gum acacia Test I A. catechu ± ± ± ± 6.16** (250 mg/kg) Test II A. catechu ± ±3.75* ± ± 3.03** (500 mg/kg) Standard 75.0 ± ±2.24** ± ± 2.65** Glibenclamide (0.5 mg / kg) One-way F ANOVA P Values are (mg%) mean±sem. n=6 in each group, df=3,23. *P<0.01; **P<0.001 as compared to respective control. < histologic analysis, moreover without sacrifice of the animals. 3. Wallis TE. Cutch. Text book of Pharmacognosy. 5th ed. London: J. and A. In the present study, AST and ALT levels in normal control Churchill Ltd; group were in conformity with the findings of Kapoor et al. [28] 4. Trease GE, Evans WC. Flavone and related flavonoid glycoside. Pharmacognosy. 4 th ed. London: Bailliere Tindall; Seven days pretreatment with the test drug (250 mg/kg) 5. Singh KN, Mittal RK and Barthwal KC. Hypoglycemic activity of Acacia catprotected the animals significantly (P<0.001) from CCl 4 echu, Acacia suma, Albizzia odoratissima seed diets in normal albino rats. induced hepatotoxicity as compared to toxic control, and Indian J Med Res 1976;64: reflects the hepatoprotective activity of ethyl acetate extract 6. Jayasekhar P, Mohanan PV, Rathinam K. Hepatoprotective activity of ethyl acetate extract of Acacia catechu. Indian J Pharmacol 1997;29: of A. catechu. There was no significant difference between the 7. Chopra RN, Nayar SL, Chopra IC. Acacia catechu Willd. Glossary of Indian test drug group and standard drug group. The hepatoprotective Medicinal plants. New Delhi: Council of Scientific and Industrial Research; activity of A. catechu could be due to the presence of bioflavonoids which have hepatoprotective and antioxidant 8. Rage N, Dahanukar S, Karandikar SM. Hepatoprotective effect of cyanidanol [4, 8, 23] properties. against carbon tetrachloride induced liver damage. Indian Drugs 1984;22: In the present study, preliminary phytochemical screening Usher G. Acacia catechu Willd (Catechu, Dark catechu). A dictionary of plant. of Acacia catechu found the presence of flavonoids and reports 1st ed. Delhi: CBS publishers and distributors; substantiate the presence of catechutanic acid, [3] quercetin, [4] 10. Handa SS, Kapoor VK. Drugs containing tannins. Pharmacognosy. 2 nd ed. Delhi: [4, 22] catechin, pseudotannin, [4] phlabotannin, [4] polyphenols, [9] Vallabh Prakashan; epicatechin [22] and cyanidanol [8] compounds in A. catechu. The 11. Kokate CK. Plant constituents. Practical pharmacognosy. 4th ed. Delhi: Vallabh antidiarrhoeal and hepatoprotective properties of A. catechu Prakashan; Seikel MK. Isolation and identification of phenolic compounds in biological materials. In: Harborne JB editor. Biochemistry of phenolic compounds. Lon- don: Academic press; p Yegnanarayan R, Shrotri DS. Comparison of antidiarrhoeal activity of some drugs in experimental diarrhea. Indian J Pharmacol 1982;14: Loux JJ, Depalma PD, Yankell SL. Antipyretic testing of aspirin in rats. Toxicol Appl Pharmacol 1972;22: Hajare SW, Chandra S, Tandan SK, Sarma J, Lal J, Telang AG. Analgesic and antipyretic activities of Dalbergia sissoo leaves. Indian J Pharmacol 2000; 32: Barham D, Trinder P. An improved colour reagent for determination of blood glucose by oxidase system. Analyst 1972;97: Shoka AA. Effect of some oral antidiabetic drugs on glucose and mineral metabolism in alloxan diabetic rats pretreated with ethylene diamine tetra acetic acid disodium salt. Indian J Pharmacol 1992;24: Geetha BS, Mathew BC, Augusti KT. Hypoglycemic effects of Leucodelphinidin derivative isolated from Ficus bengalensis (Linn.). Indian J Physiol Pharmacol 1994;38: Singh N, Tyagi SD, Agarwal SC. Effects of long term feeding of acetone extract of Momordica charantia (whole fruit powder) on alloxan diabetic albino rats. Indian J Physiol Pharmacol 1989;33: Gupta MB. Palit TK, Singh N, Bhargava KP. Pharmacological studies to isolate the active constituents from Cyprus rotundus possessing anti-inflammatory, could be attributed to the presence of tannins [9] and cyanidanol [8] and quercetin, [23] respectively. The antipyretic and hypoglycaemic properties of Acacia catechu may be ascribed to the presence of flavonoids, which have been shown to inhibit cyclooxygenases [23] and promote ß-cell regeneration besides having insulin secretagogues and antioxidant [1, 5, 18, 19, 23] properties. The results of the present study suggest that ethyl acetate extract of Acacia catechu in doses of 250 and 500 mg/kg, significantly reduced the temperature of pyretic rats, and also illustrate significant hypoglycaemic activity. The extract of A. catechu, 250 mg/kg, p.o., has also shown to possess significant antidiarrhoeal and hepatoprotective property. References 1. Gupta SS. Prospects and perspectives of natural plants products in medicine. Indian J Pharmacol 1994;26: Kirtikar KR, Basu BD. Acacia (Tourn.) Linn. In: Blatter E, Caius JF, and Mhaskar K, editors. Indian Medicinal Plants. 2nd ed. Dehradun: International Book Distributors, Book Sellers and Publishers; p Indian J Pharmacol December 2006 Vol 38 Issue

6 Studies on A. catechu anti-pyretic and analgesic activities. Indian J Med Res 1971;59: try. 6th ed. Oxford: Heinemann professional publishing Ltd, p Vimala R, Nagarajan S, Alam M, Susan T, Joy S. Anti-inflammatory and anti- 25. Ghosh R, Sharatchandra KH, Thokchom IS. Hypoglycemic activity of Ficus pyretic activity of Michelia champaca Linn. (white variety), Ixora brachiata Roxb. hispida (bark) in normal and diabetic albino rats. Indian J Pharmacol 2004; and Rhynchosia cana (Willd) D.C. flower extract, Indian J Expt Biol 1997;35: 36: Anturlikar SD, Gopumadhavan S, Chauhan BL, Mitra SK. Effects of D-400, a 22. Mathew AG, Parpia HAB. Food browning as a polyphenol reaction. In: Chich- herbal formulation, on blood sugar of normal and alloxan-induced diabetic rats. ester CO, Mrak EM, Stewart GF,editors. Advances in food research. New York: Indian J Pysiol Pharmacol 1995;39: Academic Press; p Reitman S, Frankel S. A colorimetric method for the determination of serum 23. Rajnarayana K, Reddy MS, Chaluvadi MR, Krishna DR. Bioflavonoids classifi- glutamic oxalacetic and glutamic pyruvic transaminases. Am J Clin Path cation, pharmacological, biochemical effects and therapeutic potential. Indian 1957;28: J Pharmacol 2001;33: Kapur V, Pillai KK, Hussain SZ, Balani DK. Hepatoprotective activity of jigrine 24. McLauchlan DM. Glucose, other sugars and ketones. In: wengolock AH, on liver damage caused by alcohol, carbon tetrachloride and paracetamol in McMurray JR, McLauchlan DM, editors. Varley s Practical Clinical Biochemis- rats. Indian J Pharmacol 1994;26: CAL DVD This issue (December 2006) of IJP comes with a free DVD containing CAL software to support pharmacology teaching. IJP would like to thank the following for allowing us to include their software in the DVD: Prof. Henk van Wilgenburg, The Netherlands - Microlabs 2006 Prof. M. Saghaei, Iran - Random Allocation Software The release and the distribution of the DVD was made possible by the generous financial assistance from the following organizations. IJP would like to thank for their generous gesture: Emcure Pharmaceuticals Limited, Pune Pfizer Limited, Mumbai Wockhardt Limited, Mumbai Ranbaxy, New Delhi Zydus Cadila, Ahmedabad Indian J Pharmacol December 2006 Vol 38 Issue