RESUMEN
Gelatins from lizardfish and threadfin bream skins were extracted using distilled water at 45 and 60 °C and 4, 8 and 12 h. Gelatin recovered from both lizardfish and threadfin bream skins was in the range of 63.96-91.46%. As extraction temperature and duration increased, the turbidity of gelatin solution from both species increased. Gelatins isolated from either lizardfish or threadfin bream skins at 45 °C for 4 and 8 h showed the maximum bloom strength (245.03-320.85 g), which were also greater than commercial gelatin from bovine (208.55 g) (P < 0.05). The gelatin gels (6.67%, w/v) could set at 4 °C within 3 min and were able to set at room temperatures within 51.83 min. Gelatins extracted from both fish skins contained α1-, ß- and γ-chains as predominant protein components. The lightness of all gelatin gels faintly declined with an increase in extraction temperature and time. Among the various production conditions explored, lizardfish/threadfin bream skin gelatin developed at 45 °C and 8 h was found to be highly comparable to commercial bovine gelatin. Based on the results obtained, gelatin from both fish species could be used as a replacement for land animal counterparts and can be used in many different food and pharmaceutical products.
RESUMEN
Partitioning and recovery of proteases from stomach extract (SE) and acidified stomach extract (ASE) of lizardfish using a three-phase partitioning (TPP) system in combination with an aqueous two-phase system (ATPS) was optimized. The highest yield and purity were obtained in the interphase of the TPP system, which consisted of a SE or ASE to t-butanol ratio of 1.0 : 0.5 in the presence of 40% (w/w) (NH4)2SO4. Both TPP fractions were further subjected to ATPS. Phase compositions of ATPS including PEG molecular mass and concentrations as well as types and concentrations of salts influenced protein partitioning. The best ATPS conditions for protease partitioning into the top phase from TPP fractions of SE and ASE were 15% Na3C6H5O7-20% PEG1000 and 20% Na3C6H5O7-15% PEG1000, which increased the purity by 4-fold and 5-fold with the recovered activity of 82% and 77%, respectively. ATPS fractions of SE and ASE were subsequently mixed with several PEGs and salts for back extraction (BE). BE using 25% PEG8000-5% Na3C6H5O7 gave the highest PF and yield for both ATPS fractions. SDS-PAGE investigation revealed that the decrease in contaminating protein bands was observed after the combined partitioning systems. BE fractions of SE and ASE were quite stable at -20 and 0 °C up to 14 days. Therefore, the combination of TPP, ATPS and BE could be effectively applied to recover and purify proteases from the stomach of lizardfish.
RESUMEN
One major pepsinogen, PG-I, and two minor pepsinogens, PG-II and PG-III were purified from lizardfish stomach by ammonium sulfate precipitation and two chromatographic columns. The three purified PGs migrated as single bands in native-PAGE gels with molecular weights (MW) ranging from 36 to 38 kDa. Each PG was converted to pepsin (P) at pH 2.0, and the MW were determined as 32 kDa (for P-I), 31 kDa (for P-II) and 30 kDa (for P-III). The optimum pH and temperature of pepsins were 2.0-3.5, and 40-50 °C. All 3 pepsins were strongly inhibited by pepstatin A. Divalent cations slightly stimulated the pepsin activities, but ATP had no effect on the pepsins. Purified pepsins were effective in the hydrolysis of various proteins. Km and kcat of the three pepsins for hemoglobin hydrolysis were 107.64-276.61 µM and 18.30-32.68 s-1, respectively. The new pepsins have potential for use in protein food procession and modification.