NEUTRAL AND ALKALINE MUSCLE PROTEASES OF MARINE FISH AND INVERTEBRATES A REVIEW

Muscle proteases active at neutral and alkaline pH's include calcium-activated neutral proteases, heat-activated thiol proteases, serine proteases, and metallo-proteases. They participate to different extents in postmortem degradation of fish muscle myofibrillar and scaffold proteins. Their activity in fish, the presence of endogenous activators and inhibitors, pH, and temperature. Recent studies indicate that neutral and alkaline proteases have more impact on the postmortem deterioration in quality of fish muscles than the cathepsins active at acid pH. Significant quality losses are caused by enzymatic collagen degradation in raw tissues and by heat-activated enzymes in fish gels.     

PROPERTIES OF CHITIN DEACETYLASE FROM CRUDE EXTRACTS OF MUCOR ROUXII MYCELIUM

The catalytic properties of chitin deacetylase from Mucor rouxii were studied with the aim of using the results for control of the properties of chitosan prepared by enzymatic deacetylation. A crude deacetylase extract from the mycelium of Mucor rouxii exhibited maximal activity at pH 5.8 with both water-soluble and acid-soluble chitosan. The extracellular enzyme of the culture medium exhibited maximal activity at pH 4.8. The deacetylase in the crude extract was stable over the pH range of 4.1–8.9 at 25C, retaining 85–100% of maximal activity at 40C. The extract was most active towards acid-soluble chitosan at 50C and pH 5.8. Chitin deacetylase was stable to 40C when incubated without substrate, with 85% of the activity retained at 50C. Ca²⁺, Mn²⁺, and Zn²⁺ had no effect on activity, while EDTA, Fe²⁺, and Fe³⁺ caused partial inhibition in extracts incubated without substrate for 1 h at 25C and pH 5.8. The deacetylase in the mycelial extract and in the culture medium was slightly activated by Co²⁺. The differences in temperature optima and thermal stability of deacetylase and hydrolytic enzymes can be used for controlling the degree of hydrolysis of chitosan, while the difference in pH stability has been used for prepurification, resulting in 3-fold increase in specific activity of deacetylase .     

EFFECT OF SQUID LIVER EXTRACT ON PROTEINS AND ON THE TEXTURE OF COOKED SQUID MANTLE

ABSTRACT The liver extract of squid Illex argentinus has a proteolytic activity at pH 2–8, with optimum at pH 2.6–3.1. At pH 6 the enzymes retain about 30 and 55% of the maximum proteolytic activity towards hemoglobin and casein, respectively. In squid mantle meat treated with the extract 24 h at 4C and 20C, the sarcoplasmic and myofibrillar proteins were hydrolyzed to low molecular fractions, as was shown by SDS-PAGE. Soaking isolated collagen fibers in the liver extract resulted after 24 h in almost complete solubilization of the fibers in buffered SDS/urea solutions. The solubilized product contained only low molecular weight components. A significant hydrolysis of the proteins in the squid mantle, treated with the liver extract at 20 or 4C, was accompanied by a decrease in toughness of the cooked product by about 65 and 40%, respectively, as compared to the toughness of untreated cooked samples. Cooking of the mantle directly in the liver extract had practically no tenderizing effect.     

PROTEOLYTIC ACTIVITY OF CRUDE ENZYME EXTRACTS OF SQUID ILLEX ARGENTINUS LIVER

The extract obtained from the acetone powder of frozen stored squid liver had optimum proteolytic activity against hemoglobin at pH 2.6 to 4.0. About 90% of the activity at pH 3.0 was inhibited by pepstatin and about 15% was inhibited by phenylmethanesulfonyl fluoride or iodoacetamide. The activity was increased by dithiothreitol and cysteine by about 55 and 40%, respectively. At pH 3 the enzyme extract had a temperature optimum of 45 to 50C; at 5C the activity was about 15% of that at 45 to 50C. At pH 7 the optimum activity was at about 45C. Heating of the extract without substrate at pH 3 and 7 caused a rapid decrease in proteolytic activity above 35 and 30C, respectively. A water extract of the liver was used to treat isolated bovine myofibrils at pH 5.5 and 0C. After 20 h a fragment of slightly lower m.w. than that of the myosin heavy chain was detected. After treatment at pH 7 and 0C, little degradation of the myofibrillar proteins was evident. At 20C the myofibrils were hydrolyzed to several products.     

Malondialdehyde-Nitrite Interactions in Meat and Model Systems

Nitrite retarded formation of carbonyl compounds in meat when added before cooking, but had no effect after cooking and storage even though the 2-thiobarbituric acid (TBA) number was very low due to reaction of malondialdehyde with nitrous acid. Although tripolyphosphate/ascorbate reacted similary preventing formation of carbonyl compounds, they had no effect on the TBA number when added after cooking. In a model system, nitrite reacted with malondialdehyde at pH 1.3 at room temperature. The percentage of reacting malondialdehyde decreased with higher pH and increased with nitrite concentration. The reaction of malondialdehyde with nitrite yielded high-molecular-weight products. Sulfanilamide prevented the reaction of malondialdehyde with nitrite, but only when added before nitrite.     

Behavior of Glass Fibers in Strong Acidic and Alkaline Media

The properties of basaltic, glass wool, quartz, and E-glass fibers were investigated under strong acid (HF:H₂O=l:3, HF:H₂O=1:1, 10% H₂SO₄+3% HF:H₂O=1:1) and alkaline (1 N NaOH + 1 N Na₂CO₃) treatment. The multiple-oxide fibers were found to be composed of several layers which exhibit different morphology and composition. These findings are in accordance with some proposals in the literature.     

The proteolytic activity of the crude enzyme extracts of Baltic cod (Gadus morhua) alimentary tract

The crude enzyme extract from Baltic cod alimentary tract had maximum activity of aspartic proteinases towards haemoglobin at pH 2.0 and 3.4, and of serine proteinases with casein as the substrate at pH 8.3 and 10.4. With bovine myofibrils as the substrate the proteolysis at pH 5.0-8.0 was maximum at the lowest and highest pH values. the optimum temperature for the proteolytic activity at pH 3.4 and 10.4 was 30–45°C and at pH 8.3 it was 40–50°C. Heating the crude extract for 10 min at pH 3.4,8.3, and 10.4 in the absence of the substrate had no effect on the activity of the acid and alkaline proteinases up to about 35–40°C. the stability at higher temperature decreased gradually and total inactivation occurred at 55–60°C. In the pH range 5.5–7.5 the proteolytic activity against bovine myofibrils was low at 0°C but brought about significant loss of myosin heavy chain at 20°C.     

Texture of Cooked Mantle of Squid lllexargentinus as Influenced by Specimen Characteristics and Treatments

Boiling frozen stored squid in 2% NaCl solution caused 30% weight loss after 5 min but changes in protein solubility and softening of the meat continued for at least 45 min. The influence of variability within the species on the texture of the cooked product was more significant than that of frozen storage. In discolored squid significant proteolysis was shown by electrophoresis. Such meat was less tough than that from nondiscolored specimens. Acetic acid cure induced proteolysis without improving texture, while polyphosphates softened the product without detectable proteolysis. The decisive effect on texture was displayed by pH and polyphosphates. Heating at 80° and 90°C left an uncooked sensory note.