Several attempts to further purify the bromelain powder obtained from acetone precipitation step were on dialysis, ultrafiltration, Duolite C-225 and Sephadex G-100 chromtography. By using those techniques, the purity of the enzyme were found to be insignificantly increased (~ 1.4-2 folds). The activity of bromelain powder was 281-604 CDU/mg powder. The activity of brome-lain from stem extracts could be raised by either cysteine hydrochloride or sodium metabisulphite. The activation effect of cysteine hydrochloride on the enzyme activity was additionally enhanced (~ 1.5 folds) when supplemented with ethylenediaminetetraacetic acid. Partial purification by polyacrylic acid yielded approximately 1.2 g bromelain powder/kg stem with the activity per mg powder (~ 1941 CDU) 4-5 folds higher than that from the acetone precipitation. The further purification of so obtained enzyme powder by ultrafiltration, CM-Cellulose chromatography and Sephadex G-100 chromatography or combination of the two later techniques gave no improvement of bromelain activity. Neverthe-less, results from PAGE analysis cleary illustrated the decrease in types of protein in the enzyme powder. This highly purified bromelain powder was not significantly different from the polyacrylic acid enzyme powder at the aspect of its properties to catalyze casein hydrolysis although its stability to temperature was found to be lowered. Bromelain could be immobilized on CM-Cellulose, either with or without glutaraldehyde. Both immobilized enzymes had the optimum temperature at 45℃ , while the free from optimally catalyzed reaction at 65℃. The immobilized was more stable at 30-50℃ than the free enzyme. Moreover the results also indicated that immobilization of bromelain on CM-Cellulose could enhance its capacity to hydrolyze substrate casein by 15-20 folds in comparison to the free enzyme.
