Objective: The objective of the present work is to isolate, purify, and fractionate anthocyanin from selected Osbeckia species and also to compare the analgesic and anti-inflammatory potentiality using animal models.Methods: Methodologies include extraction of anthocyanin from the in vitro callus culture of Osbeckia aspera and Osbeckia reticulata, purification using amberlite column chromatography, and fractionation by liquid chromatography-tandem mass spectrometry. The analgesic activity was determined by tail immersion method, analgesy meter, hot plate, and acetic acid-induced writhing test. Anti-inflammatory activity was evaluated by carrageenan-induced paw inflammation in mice.Results: Anthocyanin-producing callus cultures were established in MS medium fortified with various combinations of phytohormones and sucrose. Optimal callus formation in O. aspera was initiated on cultures containing 0.5 mg/L of 2, 4-D, and 0.5 mg/L 6-benzylaminopurine (BA). In O. reticulata callus was initiated in the presence of 1.2 mg/L BA and 1.4 mg/L naphthalene acetic acid. The same hormonal combination on extended treatments turned the white friable callus into red compact callus. Anthocyanins obtained from Osbeckia species were purified and fractionated containing malvidin-3-diglucoside, delphinidin, cyanidin aglycone, and peonidin. Purified anthocyanin of O. aspera at the concentrations 50, 100, 150, and 200 mg/kg after 120 min exhibited significant analgesic activity by tail immersion method, in comparison to O. reticulata. However, with hot plate method, anthocyanin of O. reticulata produced a significant analgesic activity even at lower doses (50 and 100 mg/kg) after 120 min. However, in writhing test, theepurified anthocyanin of O. aspera significantly stopped the number of writhes at a dose of 200 mg/kg and also by the anthocyanin of O. reticulata. In the evaluation of anti-inflammatory effect using plethysmometer, anthocyanin at doses of 100,150 and 200 mg/kg started producing anti-inflammatory effect after 30 min, which lasted until 120 min.Conclusion: It is concluded from the present study that the purified anthocyanin of Osbeckia possesses potent analgesic and anti-inflammatory activities.