Inhibition of Protease Activity in Muscle Extracts and Surimi from Pacific Whiting, Merluccius productus, and Arrowtooth Flounder, Atheresthes stomias

Muscle extracts of Pacific whiting, Merluccius productus, and arrowtooth flounder, Atheresthes stomias, were assayed for proteolytic activity using azocasein as a substrate. Pacific whiting extracts showed maximum activity at pH 5.0-5.2 and a temperature of 50°C, while arrowtooth flounder extracts had maximum activity at pH 5.5 and 55°C. Three sources of inhibitors (potatoes, egg white, beef plasma protein) were evaluated in vitro for inhibition of protease activity. All three were found to be effective inhibitors in crude muscle extracts. Further studies utilizing these inhibitors in surimi showed that potato was equivalent to both egg white and beef plasma protein in preserving the gel forming characteristics ofheated kamaboko in both species.

Inhibition of seafood-borne bacteria in cooked mackerel (Rastrelliger kanagurta) fish meat by lactic acid bacteria

Antagonistic activity of lactic acid bacteria (LAB) namely Streptococcus faecalis, Pediococcus cerevisiae and Lactobacillus casei was tested against seafood-borne bacteria such as Staphylococcus aureus, Bacillus cereus, Escherichia coli, Clostridium perfringens and Listeria monocytogenes. Three lactic acid bacteria such as Streptococcus faecalis, Lactobacillus casei and Pediococcus cerevisiae were coated on cooked mackerel meat, individually and in combination against fish-borne bacteria. S. faecalis inhibited C. perfringens in individual coat by 3.7 log units as compared to control, whereas L. casei did not inhibit C. perfringens. P. cerevisiae inhibited S. aureus by 5 log units. L. casei, inhibited L. monocytogenes by 3.3 log units on the third day of storage as compared to control. On the other hand, S. aureus and B. cereus were inhibited on the third and second day by 4.9 log and 5.2 log units respectively. B. cereus, S. aureus, L. monocytogenes were the most sensitive to all three LAB. C. perfringens was the least inhibited among all the seafood-borne bacteria tried. Multiple LAB or LAB strains in combination showed much earlier inhibitory activity on seafood-borne bacteria than single LAB coat.

Liver cytochrome oxidase P4501A1 purification of Acipenser persicus and designing ELISA for measuring

Moon oxygenases related to cytochrome P450s are the molecular Biomarkers which have important role in Biotransformation of endogenous and exogenous compounds and catalazyin of many biological reactions. One of the important isoenzyme is cytochrome P4501A. This isoenzyme involved in metabolism of environment pollutnts such as PAHs. Because of its inducibility, it has a key tool for impact assesment of contaminants in aquatic environment. In this study, at first, that fractions containing Acipenser persicus and Huso huso isoenzyme were purified, and after that Antibodies against them were prepared. For isolation of isoenzyme fraction, Microsomes were prepared from fish liver using differential centrifugation at high speeds. microsomes were solubiized by cholat sodium and Emulgen. Extraction of this isoenzyme was done with the combinatuion of ionexchange chromatography and gelfiltration or chromatofocusing chromatography. Ion exchange chromatography and gel filtration were applied in DEAE sepharose fast flow and sephacryl S200 respectively and chromatofocusing was done at poly buffer 74 and 94 exchanger. The results of SDS-PAGE Showed that the molecular weight of isoenzyme was about 58±1 KDa. Furthermore the inmunoblotting results confirmed this subject. Isoenzyme activigy based on EROD (Ethoxyresorofin o-deethylase) reaction showed about 20-26 fold increase in enzyme activity of treated fish than control fish. The results of Elisa, Using monoclonal anti cod P4501A demonstrated the inducibility and highly elevated of its activity in treated sample more than the control fish. Mean while, the fish sample were showed the strong reaction to polyclonal antibody against beluga P4501A1 prepared in our Lab compared to monoclonal anti body.