Anti-inflammatory and in vitro antioxidant activity of Desmodium triquetrum (L.).

Sir,

Desmodium triquetrum contains a wide variety of free radical scavenging molecules, such as phenolic and nitrogen compounds, terpenoids, and carotenoids that are rich in antioxidant activity.[1] The chloroform and alcohol extracts of D. triquetrum were reported for their antibacterial activity. Epidemiological studies have shown that many of these antioxidant compounds possess anti-inflammatory, anti-atherosclerotic, antitumor, anti-mutagenic, antibacterial, antiviral, anti-carcinogenic, and anti-allergic activities. Thus, on the basis of its chemical constituents, the present study was undertaken to evaluate D. triquetrum for anti-inflammatory and in vitro antioxidant activity. Leaves of D. triquetrum were collected, authenticated, shade dried, coarsely powdered, and extracted with 95% ethanol using Soxhlet extractor. The preliminary phytochemical analysis of the extract confirmed the presence of flavonoids, glycosides, steroids, saponins, phenolic compounds, and amino acids.

Adult albino Wistar rats of either sex, weighing about 150-180 g were used for the study. They were maintained on standard animal diet and water ad libitum. The study was approved by the Institutional Animal Ethics Committee. All chemicals and reagents used were of the analytical grade.

Acute toxicity studies were carried out as per the Organization for Economic Co-operation and Development (OECD) guidelines 423. Animals were divided into five groups of six each. They were fasted overnight, but allowed free access to water. Group 1 served as normal control, received 1 ml of vehicle orally. Animals of group 2, 3, 4 received ethanol extract of D. triquetrum leaf (DTE) in the dose of 100, 200 and 300mg/kg body weight p.o., respectively, and group 5 standard drug, diclofenac sodium, 15 mg/kg body weight orally. Paw volume of all the animals were measured using plethysmometer (UGO Basile, Italy). After 30 min of dosing, all the animals were injected 0.1 ml of 1% w/v suspension of carrageenan in saline in the sub plantar region of left hind paw. The paw volume was measured at one hour interval till 3 h and compared with standard group.[2]

Antioxidant activity of DTE was tested in vitro by following methods. The free radical scavenging activity of the samples was measured by 1, 1-diphenyl-2-picryl-hydrazil (DPPH) using the method described by Shimada et al.[3] The ability of samples to scavenge H2O2was determined according to the method of Ruch et al.[4] The interaction of samples with NO was assessed by the nitrite detection method.[5]

All tests were performed in triplicate. Ascorbic acid was used as positive control.

The results obtained were analyzed using one-way analysis of variance (ANOVA) followed by Dunnett's multiple comparison tests. Differences between the data were considered significant at P< 0.05.

DTE was found to be safe up to at a dose of 2000 mg/kg. Hence, the doses of 100, 200, and 300 mg/kg were selected for the present study. The results of the present investigation suggest that DTE produced significant anti-inflammatory [Table 1] and antioxidant activity. Carragenan administration to the control group resulted in increase in paw volume at 30 and 60 min and gradually decreased after 120 min. The treatment with DTE in all the doses showed significant decrease in the paw volume compared to control (P<0.05). The maximum inhibition of paw edema was observed at 60 min at the dose of 300 mg/kg body weight. The results were comparable with the standard drug.

Table 1

Effect of ethanol extract of D. triquetrum leaf on carrageenan induced paw edema in rats

A significant (P<0.01) decrease the concentration of DPPH· radical due to the scavenging ability of DTE and ascorbic acid. The results indicated that the test extract has a significant dose dependent effect on scavenging of free radicals. The DTE had strong H2O2 scavenging activity when compared with control (P<0.01). Moreover, this activity was close to ascorbic acid. The maximum H2O2 scavenging activity was observed at 50 μg/ml of the test extract. NO is a potent pleiotropic mediator of physiological processes. DTE (25 to 75μg/ml) also moderately inhibited nitric oxide in a dose dependent manner. Standard ascorbic acid was found to have 76.82% activity at 75 μg/ml.

Carrageenan induced inflammatory process is believed to be biphasic.[6] DTE showed a significant anti-inflammatory activity in both phases of inflammation. Increase in cyclic adenosine monophosphate (cAMP)-phosphodiesterase (cAMP-PDE) activity in edematous tissue after carrageenan injection paralleled the increase in migrated cells as reported.[7] It has been reported that alcohol extract of D. triquetrum was found to inhibit cAMP-PDE activity.[8] Therefore, the possible mechanism for significant anti-inflammatory activity of DTE may be by inhibition of cAMP-PDE activity and also it can be attributed to its antioxidant activity as evidenced by the presence of flavonoids. Further, the isolation of the compounds responsible for the claimed activities has to be taken up to support the present findings.