Kinetic Studies of Purified Bromelain from Pineapple (Ananas comosus [L.]) Merr) Core with Hidroxyapatite and CM Sephadex C-50 Ion Exchange Chromatography

Bromelain is a protease enzyme that functions to break peptide bonds. in the health field bromelain can function as anti-inflammatory, anti-thrombotic and fibrinolytic.Here we purposed a new purification step to collect enzyme from pineaplle. The purification of the enzyme from Ananas comosuswas carried out by precipitation with varying concentration of acetone and followed by column chromatography using hidroxyapatite ion exchage and CM sephadex C-50 resin. The enzyme activities were evaluated using casein as substrate. The highest specific activity of bromelain was gained from acetone fractination as51,51U/mg in the range of 50-80% saturation with purity level of enzyme as11times from its crude extract. The highest specific activity fromlast fractionwhich runonhidroxyapatite ion-exchange column chromatography was87,49 units/mg withpurity level ofenzyme resulted19 times higher compared to the crude extract. Later on, the purification with CM sephadex C-50resulted in increasing the specific activity to 200U/mg with purity level of enzyme as45timeshigherfrom its crude extract. Hydrolysis of various casein concentration with purified bromelain was carried out at optimum reaction condition of pH 7,0 and 370C. The results obtained revealed the Km and Vmax value were 0,94% (w/v) and 0,023U/min respectively. Fromthe various purification steps that have been done, it can be observed the increasing of bromelain specific activity from each stage.