Antioxidant and anti-inflammatory study of Rubus fruticosus and Verbascum thapsus medicinal plants collected from Dir (L) N.W.F.P. Pakistan

Abstract

The aim of this study is not only to preserve the traditional information but also to update and expand this knowledge of medicinal plants according to modern parameters. In this regard two species Rubus fruticosus (Rosaceae) and Verbascum thapsus (Scrophulariaceae) were collected from Northern area Dir (KPK formally called NWFP) of Pakistan which is very rich in medicinal plants, not only to evaluate but to validate their traditional uses according to advanced screening techniques. Methanolic extract of each part of both species was investigated and evaluated for anti-inflammatory, antioxidant potentials, pharmacognostic, neuropharmacological, gross behavioral studies, toxicity studies and diuretic action. Pharmacognostic studies The presence of carbohydrates, triterpenes, tannins, flavonoids and sterols were detected in R. fruticosus and V. thapsus during chemical screening. Thin layer chromatographic plates were also developed for the extract of each part of the plant. The Rf value of each spot was calculated during experiment, for R. fruticosus fruit (0.38,0.42,0.59,0.66,0.73,0.81) for leaves (0.33,0.62,0.64,0.72,0.75) for root (0.38, 0.43) and for stem extract (0.36, 0.43) in Ethyl acetate, methanol and water system. The Rf values in the same solvent system were calculated for V. thapsus fruit (0.26,0.32, 0.46,0.59,0.78) for leaves (0.14,0.22,0.32) for root (0.36,0.46,0.48,0.52, 0.59,0.78) and for stem extract (0.17, 0.39,0.4,0.52, 0.59,0.69,0.78). Antioxidant activity Antioxidant studies using DPPH, ABTS and Nitric oxide free radical spectrophotometeric methods showed the antioxidant capacity order on % free radical scavenging basis for R. fruticosus various parts extracts as fruit > leaves > root > stem and for V. thapsus root > fruit > leaves > stem. The % radical scavenging capacity using 0.5 mg/ ml of sample in DPPH method were 96%, 92.8%, 90.07% and 89.88% for R. fruticosus fruit, leaves, root and stem respectively. Using same method and concentration gave % radical scavenging for V. thapsus fruit, leaves, root and stem 89.40, 87.73, 96.13 and 85.05 respectively. ix Anti-inflammatory activity Formalin induced inflammation in mice and Carrageenan induced inflammation in rats were used to evaluate anti-inflammatory activity. Dose dependent anti-inflammatory effects were observed for both R. fruticosus and V. thapsus at dose of 100, 300 and 500 mg/kg. The fruit and leaves extract of R. fruticosus showed significant anti-inflammatory activity. Leaves extract showed different however the extract of fruit showed the same inhibition in both phases of Formalin test. The root and stem extracts showed no significant anti-inflammatory effect. The reduction capacity of paw edema by R. fruticosus various extracts followed the order; leaves > fruit > root > stem. The order of potency in formalin test and on the basis of reducing the rat paw edema for V. thapsus various parts using 100, 300 and 500 mg/kg doses were observed as fruit > leaves > root > stem. Analgesic activity Leaves and fruit exhibited higher of analgesic effect compared to root and stem extracts of R. fruticosus using hot plate, tail flick and writhing test. The order of potency for analgesic effect for various parts of V. thapsus was observed as leaves > fruit > root > stem. The analgesic effect was observed in dose dependent fashion and the applied doses were 100, 300 and 500 mg/kg. Diuretic activity Albino mice were used to monitor the diuretic activity of R. fruticosus and V. thapsus. Fruit and to some extent leaves of R. fruticosus showed diuretic action but the rest parts showed no significant action. Similarly fruit of V. thapsus was observed for significant diuretic effect but leaves, root and stem showed no significant diuretic activity. Antibacterial activity and Antifungal activity E. coli, S. typhi, S. aureus, M. luteus, Citrobacter, B. subtilis, P. aeruginosa and P. mirabilis were use in antibacterial studies and significant results were obtained for extracts of both plants using agar disc diffusion method. The order of activity for R. fruticosus was stem > leaves > fruit > root and for V. thapsus was leaves > fruit > root > stem. The stem extract of R. fruticosus was found the most effective with MIC of 20μg comparable with standard. No significant antifungal activity was observed for methanolic extracts of both plants. Gross behavioral studies At dose of 100, 300 and 500 mg/kg the gross behaviors such as sweating, salavation, piloerection and increased respiration etc. were observed. Slight urination was found in fruit of the R. fruticosus. A slight increase in overall performance was observed. Plioerection and cough reflex was observed for leaves extract of V. thapsus and overall decrease in performance was observed. Neuropharmacological activity Slight anxiolytic effect without sedation was observed with the administration of extract of R. fruticosus and no muscle relaxing effect. Antidepressant effect was observed after forced swimming test. The order of antidepressant effect for various parts of R. fruticosus were as fruit > root > leaves > stem. Sedative effect was observed with V. thapsus especially with fruit and leaves extracts which resulted decrease in motor performance. There was no muscle relaxing effect. Similarly antidepressant effects were produced by leaves and fruit in forced swimming test comparative to root and stem. Toxicity studies Various parts extracts of R. fruticosus and V. thapsus showed no significant insecticidal activity however the anthelmintic activity was remarkable especially for fruit and leaves of V. thapsus at dose of 50, 75 and 100mg. The brine shrimp toxicity test also point out leaves and fruit of V. thapsus the most cytotoxic. The R. fruticosus was comparatively safer.