Optimization of Classical Lipase Activity Assays for Fish Digestive Tract Samples

Abstract

Fish possess lipases from embryonic development to adulthood. Lipase activity methods vary and significantly differ in terms of the concentration of the substrate used, bile salt, Ca2+, temperature, pH, and type of lipase units, which limits comparative studies. The three most-used substrates are p-nitrophenyl (p-NP), β-naphthyl (β-N) derivates, and emulsified natural oils. These were selected to be redesigned in this study to measure lipase activity under temperature, pH, ion, and bile salt conditions closer to fish physiology, using the appropriate molar absorption coefficient to calculate the lipase units. Cynoscion parvipinnis (CP), Seriola rivoliana (SR), Centropomus viridis (CV), Elop affinis (EA), and Canthidermis maculate (CM) pyloric caeca-intestine extracts were studied. Sodium taurocholate showed the highest activity for intestinal lipases, and the fatty acid length in the substrates changed the lipase hydrolysis rate. The highest lipase activity was obtained with p-NP butyrate and p-NP caprylate in four fish species. Lipase activity was highly activated with Ca2+ (4–7 mM). The β-N absorption spectrum indicates a plateau between 534 and 554 nm for fish lipases. Salmon oil was identified as the most digestible lipid in the four fish species using the in vitro digestibility assay. The lipase zymogram showed an apparent size of 46.3 kDa for CP, 40.2 kDa for SR, 46.2 kDa for CM, 106.6 kDa for EA, and 58.3, 84.6, and 162.1 kDa for CV.

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