High Speed Counter Current Chromatography (HSCCC) of Pourouma guianensis stem bark extract

Pourouma guianensis (Urticaceae) is a large tree reaching 30 m high. In Brazil, it is found almost in all over the country and is popularly known as “embaúba benguê”. P. guianensis is reported to have leishmanicidal, larvicidal, antimicrobial and antioxidant activity. In folk medicine, the green fruits are used for wound healing and the stem bark infusion treats dysentery1. P. guianensis chemical profile comprises steroids, flavones and terpenes2. This work aims to isolate chemical compounds of dichloromethane extract of P. guianensis stems by means of High Speed Counter Current Chromatography (HSCCC). Dried and powdered plant material was extracted with dichloromethane using Soxhlet apparatus. Crude extract was submitted to Column Chromatography (CC) using silica gel as stationary phase and eluted with a solvent mixture of chloroform, ethyl acetate, methanol and water, in different concentrations. Two fractions were obtained and then submitted to HSCCC. One fraction used a gradient constituted with hexane: ethyl acetate: methanol: water (1:2:1:1, system 1), (1:2:1.5:1, system 2) and (1:2:2:1, system 3), and the system was chosen by shake-flask method. The stationary phase was the upper phase of system 1 and the mobile phase was the lower phase of each system. Stationary phase retention with the first eluent was by 67.2% and 207.8 mg of the sample was injected. Chromatographic separation was achieved at 1.5 mL flow rate by 1200 rpm. Spectroscopic analysis of the fractions obtained by CC and HSCCC allowed the identification of: triterpenes (friedelin and friedelanol); triterpenic acids (tormentic, arjunolic and 2β,3α,19α,23-tetraidroxiurs-12-en-28-oic): steroids (sitosterol); nor-sesquiterpene (vomifoliol). Pre fractionation using silica gel increased the capacity of separation of the dichloromethane extract by gathering compounds with similar polarity which facilitated the choice of solvent system. These results are in accordance with the chemical profile presented by species belonging to the same subfamily.