Determination of digestive enzyme kinetics: a new method to define trophic niches in freshwater snails

Abstract

 Fluorogenic substrate analogues (MUF substrates) are very sensitive in detecting hydrolytic enzymes. This method was adapted for the quantitative analysis of extracellular enzymes in snails and other animals. It was then used to determine cellobiase, chitobiase, protease, esterase, phosphatase and lipase in the digestive tract of Radix peregra and Bithynia tentaculata. The method was sensitive enough to determine the complete enzyme kinetics (K _m, V _max values) in the esophagus, stomach, digestive gland and intestine of single animals adapted to various natural foods. With the exception of lipase and phosphatase, K _m values were between 15 and 30 mmol l^–1, independent of prefeeding. Enzyme activities, in contrast, changed with food conditions: cellobiase-activities ranged from 80 to 8000 picokatal, esterase from 500 to 3000 pKat, chitobiase from 150 to 750 pKat and protease from 100 to 1100 pKat per animal. R. peregra had cellobiase activities 10 times higher than B. tentaculata. For chitobiase, esterase and protease the enzyme activities were in the same order of magnitude in both species. All enzymes differed significantly in activity between prefeeding regimes. This variability with prefeeding was much higher in Bithynia than in Radix. These results help to better define the trophic niches and feeding specializations of the two species studied.