Optimization of the protein fraction of microcapsules used in feeding of marine fish larvae using in vitro digestibility techniques

The development of artificial feeds for marine fish larvae is demanding and, taking into account the difficulties and costs associated with the rearing of the larvae of some fish, preliminary evaluation of feed ingredients using in vitro techniques may be an alternative to in vivo assays. Some proteins used in microfeeds for marine fish were tested in vitro for examination of their effect on the proteases of seabream larvae. Casein and cuttlefish meal did not affect protease activity of crude larval extracts, whereas ovalbumin produced a 60% inhibition. The use of a pH-stat for the assessment of the degree of hydrolysis (DH) of proteins confirmed the low nutritive value of ovalbumin, since a much lower value was obtained for microcapsules prepared using this protein source when compared with those prepared using casein or cuttlefish meal (3.2 vs. 7.3 and 7.6, respectively). Products resulting from such hydrolysis were analysed using sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE). Image analysis of such gels led to the formulation of an index named the coefficient of protein degradation (CPD) which expressed the extent to which the main protein fractions were hydrolysed by larval proteases in a given time. A significant correlation (r2 = 0.98, P < 0.05) was obtained between DH and CPD values measured for either protein sources or microcapsules. The combination of protease inhibition assays with measurements of DH and CPD is proposed as a preliminary evaluation of protein ingredients used in the formulation of artificial feeds for larval fish.