Hydrolysis of bradykinin by stem bromelain.

During the course of studies on the anti-edema and anti-inflammatory actions of stem bromelain, a potent kininase activity was observed (1). The present paper describes the analysis of the bonds of bradykinin which are susceptible to the action of stem bromelain. Minoh, Osaka) dissolved in 3 ml of 5 mM phosphate buffer, pH 7.5, containing 2 mM (3 mercaptoethanol, was mixed with 3 ml of purified stem bromelain (7.2 nag: 0.218 lomoles) preincubated in the same buffer at 37'C for 20 min for full activation. Twenty microliter aliquots of the reaction mixture incubated at 37 °C were withdrawn every 5 min for assay of rat uterus smooth muscle contracting activity. The contracting activity decreased to 0.5 of the original after 15 min incubation. The resultant reaction mixture was stored at-20°C until required. Enzyme solution without added bradykinin was treated in parallel as control. An aliquot of the reaction mixture was heated in a boiling water bath for 10 min to inactivate the enzyme and dinitrophenylated at pH 9.0 at 40'C in the dark for 2 hr (2). The DNP derivatives were separated into ether-soluble and aqueous fractions by extracting with