Kinetic studies of partially purified bromelain from Bogor pineapple (Ananas comosus [L.] Merr) core with ion exchange chromatography and in vitro evaluation of its antiplatelet activity

Bromelain is a mixture of the proteolitic enzymes found in pineapple plant tissues within the Bromeliaceous family. In this research the isolation and purification of bromelain from Bogor pineapple [Ananas comosus (L.) Merr] core were carried out using ammonium sulphate precipitation and Diethylaminoethy-Cellulose (DEAE-Cellulose) chromatography. The enzyme activities were evaluated using casein as substrate. The highest specific activity of bromelain from ammonium sulphate fractionation was obtained at 51.75 U/mg in the range of 20-80% saturation with a purity level of enzyme 24 times from its crude extract. Later on, the purification with DEAE-Cellulose resulted in the increasing of specific activity to 60.57 U/mg with a purity level of enzyme 28 times from its crude extract. Hydrolysis of various casein concentrations with purified bromelain was carried out at the optimum reaction condition of pH 7.0 and 37°C. The results obtained revealed the K_m and Vmax values were 0.61% (w/v) and 6.22 U/min, respectively. In vitro study of antiplatelet agent activity using human Platelet Rich Plasma (PRP) revealed that all bromelain fractions showed the activity as an antiplatelet agent. The highest inhibition was shown by DEAE-Cellulose fraction of 54.16% with IC₅₀ of 31.37μL/mL.