Development and validation of a new HPTLC method for quantification of conophylline in Tabernaemontana divaricata samples obtained from different seasons and extraction techniques: Insights into variation of pancreatic lipase inhibitory activity

Abstract

Tabernaemontana divaricata (L.) R. Br. Ex Roem. & Schult Apocynaceae) is an important indole alkaloid rich Indian medicinal plant with conophylline being a potential lead in the treatment of diabetes mellitus and obesity. Seasonal variations and extraction techniques are two major variables that play a crucial role in the extractive yield of phytochemical constituents. Till date, there are no reports on quantification of conophylline from the leaves of T. divaricata of Indian origin, and its variation due to season and extraction techniques. The present study reports for the first time, development and validation of a new HPTLC method for the quantification of conophylline. A resolved peak of conophylline was achieved in the chromatogram with mobile phase of chloroform: methanol (90:10% v/v), when developed on a 15 cm length TLC plate. Further, the leaf samples of T. divaricata were collected during the months of August, November, February and May, and subjected individually to different extraction techniques viz., ultrasonic extraction, hot percolation and cold maceration. A total of 12 alkaloid rich fractions were obtained, and the extractive value of conophylline was found to be highest in August sample, when subjected to ultrasonic extraction (35.57 mg per 1 g of alkaloid rich fraction). Pancreatic lipase inhibitory activity of the alkaloid rich fractions was in correlation to their respective conophylline content (Pearson’s r = −0.7152) with potent activity exhibited by August sample, that was obtained by ultrasonic extraction (IC50 = 7.86 μg/mL).