Trypsin from the Processing Waste of the Lane Snapper (Lutjanus synagris) and Its Compatibility with Oxidants, Surfactants and Commercial Detergents

Abstract

A trypsin from the viscera of the lane snapper (Lutjanus synagris) was purified by heat treatment, fractionation with ammonium sulfate and affinity chromatography. The molecular weight of the enzyme was estimated to be 28.4 kDa (SDS-PAGE). The purified enzyme was capable of hydrolyzing the specific substrate for trypsin benzoyl-arginine-p-nitroanilide (BApNA) and was inhibited by benzamidine and tosyl lysine chloromethyl ketone (TLCK), synthetic trypsin inhibitors and phenylmethylsulfonyl fluoride (PMSF), which is a serine-protease inhibitor. The enzyme exhibited maximal activity at pH 9.0 and 45 degrees C and retained 100% of the activity after incubation at the optimal temperature for 30 min. At a concentration of 10 mM, activity was slightly activated by Ca(2+) and inhibited by the following ions in decreasing order: Cd(2+) > Hg(2+) > Cu(2+) > Zn(2+) > Al(3+). The effects of Ba(2+), K(1+) and Li(1+) proved to be less intensive. Using 1% (w/v) azocasein as substrate, the enzyme revealed high resistance (60% residual activity) when incubated with 10% H(2)O(2) for 75 min. The enzyme retained more than 80% activity after 60 min in the presence of different surfactants (Tween 20, Tween 80 and sodium choleate). The alkaline protease demonstrated compatibility with commercial detergents (7 mg/mL), such as Bem-te-vi, Surf and Ala, retaining more than 50% of initial activity after 60 min at 25 degrees C and 30 min at 40 degrees C. The thermostability and compatibility of this enzyme with commercial detergents suggest a good potentiality for application in the detergent industry.

Metal-sensitive and thermostable trypsin from the crevalle jack (Caranx hippos) pyloric caeca: purification and characterization

“…The loss of enzymatic activity at pH values outside the range is probably caused by protein conformational changes as a result of charge repulsion [23]. Similar behavior was observed for trypsin from Nile tilapia [1], pirarucu [2], silver mojarra [3], lane snapper [4], Monterey sardine [9], sardine [10], jacopever and elkhorn sculpin [11], skipjack tuna [12], brownstripe red snapper [13], and grey triggerfish [24] and crude extract from crevalle jack [16]. According to Maurer [25], the optimum pH is a relevant parameter that indicates the potential utilization of enzymes in detergent formulations to be used in the alkaline pH range.…”

“…Fish trypsins have molecular masses between 23 and 28 kDa, e.g., Oreochromis niloticus (23.5 kDa) [1], Colossoma macropomum (23.9 kDa) [5], Gadus macrocephalus (24 kDa) [19], Pseudupeneus maculatus (24.5 kDa) [6], Sardina pilchardus (25 kDa) [10], Diapterus rhombeus (26.5 kDa) [3], Salaria basilisca (27 kDa) [20], Pterygoplichthys disjunctivus (27.5 kDa) [21], Arapaima gigas (28 kDa) [2], Pomatomus saltatrix (28 kDa) [22], and Lutjanus synagris (28.4 kDa) [4]. …”

“…Trypsins (EC 3.4.21.4) are present in the digestive tract of fish and have largely identical substrate specificities [9]. These enzymes are generally thermostable, showing high activity at alkaline pH and sensitivity to numerous metal ions [1-4,6,9-13]. …”

Purification, characterization and application of novel alkaline protease from new Bacillus cereus UV-15 mutant

“…Usefulness of alkaline proteases in the facilitation of blood stains removal from cotton cloth was reported (37,38). The addition of proteases to detergents considerably increases the cleaning effect by removing protein stains and the consumption of surface active substances, thereby decreasing the pollution load (39).…”

Effect of Four Commonly Used Dissolution Media Surfactants on Pancreatin Proteolytic Activity

“…On the other hand, Kubota et al reported that high concentrations of Triton X-100 (1-10%) did not effect, or had a negligible effect, on the activity of two carboxypeptidases isolated from the exocarp of mandarin oranges (27). At the same time, it has been shown that the activities of trypsin from Bacillus licheniformis and trypsin isolated from fish intestine (lane snapper) were inhibited by 20 and 14%, respectively, in the presence of this surfactant (28,29). Pancreatin, however, is a multicomponent enzyme mixture, and we cannot say what the influence of this surfactant on the individual enzymes is.…”