Characterization and expression levels of protease enzymes at different developmental stages of Siamese fighting fish (Betta splendens Regan, 1910)

Abstract

Characterization of total proteases, trypsin and chymotrypsin were performed at pH 2 12 and temperatures 20-80°C in three life stages of Siamese fighting fish (Betta splendens Regan, 1910). At least fourteen protease activities were detected, including seven acidic (pH levels 2, 4 and 5-6), one neutral (pH 7), and six alkaline (pH levels 8-12) activities in the optimal temperature range of 30-60°C. The neutral and alkaline pH profiles were different between sexes during the on-growing stage (1.5 month-old). The acidic proteases played a major role in digestion in the early life stage (10-day-old), while the alkaline proteases became more important toward older stages. Trypsin and chymotrypsin showed similar characteristics with at least five activities observed. For trypsin, activity was detected with pH 7 and 8 at 50°C, and pH 10 at 30, 50 and 60°C. For chymotrypsin, activity was found at pH levels 7 and 8 at 50°C, and pH 10 at 30, 40 and 50°C. Regardless of sex and age, the most suitable conditions for studying digestive enzyme activities in general were at pH 8 and 50°C for total proteases, pH 7-8 and 50°C or pH 10 and 30-35°C for trypsin and pH 7-8 and 50°C or pH 10 and 40°C for chymotrypsin. Trypsin and chymotrypsin are important proteases showing similar general optimal conditions to that of total proteases. This information provides elementary knowledge for studying protein digestibility by in vitro techniques and for determining growth performance quality using digestive enzyme markers in Siamese fighting fish.