Multi-constituent synergism is responsible for anti-inflammatory effect of Azadirachta indica leaf extract

• Published in: Pharmaceutical biology Vol. 52; no. 11; pp. 1411 - 1422
• Main Authors: Umar, Muhammad Ihtisham, Asmawi, Mohd Zaini, Sadikun, Amirin, Abdul Majid, A. M. S., Atangwho, Item Justin, Khadeer Ahamed, Mohamed B., Altaf, Rabia, Ahmad, Ashfaq
• Format: Journal Article
• Language: English
• Published: England Informa Healthcare USA, Inc 01.11.2014; Taylor & Francis
• Subjects: Animals; Anti-Inflammatory Agents - isolation & purification; Anti-Inflammatory Agents - therapeutic use; Azadirachta; Bioactivity guided fractionation; carrageenan assay; cotton pellet assay; cyclooxygenase; cytokines; Dose-Response Relationship, Drug; Drug Synergism; Edema - blood; Edema - drug therapy; Edema - pathology; Female; Humans; interlukin-1; Male; Plant Extracts - isolation & purification; Plant Extracts - therapeutic use; Plant Leaves; Rats; Rats, Sprague-Dawley; Sheep; TNF-α; Treatment Outcome; carrageenan assay; TNF-α; interlukin-1; Bioactivity guided fractionation; cotton pellet assay; cyclooxygenase; cytokines
• DOI: 10.3109/13880209.2014.895017
• ISSN: 1388-0209

Abstract Context: Azadirachta indica A. Juss. (Meliaceaes) leaves have been used traditionally to treat swelling and rheumatism in Indian cultures. Objective: To fractionate A. indica leaf extracts using bioactivity guided manner for identification of the active anti-inflammatory principles. Materials and methods: Polarity-gradient sequential extracts (petroleum ether, chloroform, methanol, and water) of A. indica leaves were screened for their anti-inflammatory potential using the carrageenan-induced rat paw edema model (1 g/kg). The chloroform extract was sequentially fractionated to obtain n-hexane (F-1), n-hexane-chloroform (F-2), and chloroform (F-3) fractions and their inhibitory effect on rat paw edema was evaluated (500 mg/kg). Inhibitory effect of F-2 on granuloma formation, plasma interleukin (IL-1), and tumor necrosis factor (TNF-α) was assessed at the doses of 100, 200, and 400 mg/kg using the cotton pellet assay in rats. Three sub-fractions (SF-1, SF-2, and SF-3) were obtained upon chromatography of F-2, and their inhibitory effect on cyclooxygenase was assessed at 200 µg/mL concentration. The sub-fractions were subjected to gas chromatography-mass spectrometry (GC-MS). Results: All the extracts showed significant anti-inflammatory effect; however, chloroform extract was the most effective against paw edema (53.25% inhibition). The three fractions of chloroform extract showed significant effect, while F-2 being the most potent (51.02%). F-2 demonstrated dose-dependent inhibition of granuloma and cytokines. Interestingly, all the sub-fractions of F-2 inhibited COX-1 and COX-2 with almost equal potential. GC-MS revealed that chemically the sub-fractions were totally different from each other. Discussion and conclusion: Anti-inflammatory effect of A. indica is a result of cumulative and synergistic effects of diversified constituents with varying polarities that collectively exert the effect via suppression of cyclo-oxygenases and cytokines (IL-1 and TNF-α).