IDENTIFICATION OF TWO MATRIX METALLOPROTEINASES IN THE SKELETAL MUSCLE OF PACIFIC ROCKFISH (SEBASTES SP.)

Two heat-stable metalloproteinases with collagenase activity have been identified in the skeletal muscle of Pacific rockfish (Sebastes sp.) and partially characterized. The 47 kDa and 95 kDa enzymes appear to be similar to mammalian matrix metalloproteinases (MMPs), with respect to molecular weight, pH-activity profile, substrate specificity, dependence on calcium for activity, and inhibition-activation profiles. The fish metalloproteinases readily hydrolyzed collagen and gelatin, but not β- or K-casein, in SDS-PAGE substrate zymograms. They also hydrolyzed collagen and gelatin in solution, but showed very low activity in solubilizing native fibrillar collagen. They did not hydrolyze solutions of β- or K-casein, azocasein, hide powder azure or synthetic substrates for trypsin, chymotrypsin or collagenase. To our knowledge, this is the first report where fish muscle proteinases have been identified as possible members of the family of MMPs. The rockfish muscle MMPs appear to be different from other currently identified fish muscle proteinases in regard to molecular weight, substrate specificity and activation-inhibition profiles. These two enzymes could be partly responsible for the degradation of collagen and other extracellular matrix proteins in fish muscle and for the texture softening of seafood products.     

DETERMINATION OF COLLAGEN CROSSLINKS IN ROCKFISH SKELETAL MUSCLE

The low collagen content in fish muscle makes the quantitative analysis of collagen crosslinks difficult. We have developed a method for the quantitative determination of reducible difunctional collagen crosslinks in samples containing as little as 0.5 picomoles of crosslink per microgram of hydroxyproline. The low collagen-containing tissue is first enriched by removing most of the non-collagenous protein with cold sodium hydroxide solution. The lyophilized alkali-insoluble extract is then reduced with NaB³H₄ and acid-hydrolyzed. Initial fractionation of hydrolysates on a Bio-Gel P-2 gel filtration column provides partial separation of components with a sharp peak of radioactivity containing more than 90% of the difunctional crosslinks. This peak is ultimately analyzed by HPLC for the quantitative determination of specific difunctional crosslinks. This method has been used to show that the difunctional crosslink composition of muscle collagen from two species of Pacific rockfish differs quantitatively from each other and from that of rat tail tendon.     

CHANGES IN FRESHNESS OF CHILIPEPPER ROCKFISH (SEBASTES GOODEI) DURING STORAGE AS MEASURED BY CHEMICAL SENSORS AND BIOSENSORS

ABSTRACT Trimethylamine (TMA) and ammonia contents in Chilipepper rockfishes were determined by ion specific electrodes, as a late indicator of fish freshness. After 9 days of storage in ice, TMA contents significantly increased, indicating that bacterial spoilage was in progress. The pattern of changes in ammonia contents was similar to that of TMA. Determination of ATP degradation products in Chilipepper rockfish by HPLC showed that AMP and hypoxanthine levels were low and did not change much during storage. The concentration of IMP initially increased and then continuously decreased as inosine accumulated. Only trace amounts of hypoxanthine were detected in rockfish tissues. Chilipepper rockfish appears to differ from other Sebastes species in that ATP degradation results in inosine accumulation rather than hypoxanthine.     

LOCALIZATION OF PARATROPOMYOSIN IN CHICKEN SKELETAL MUSCLE MYOFIBRILS AND ITS BEHAVIOR DURING POSTMORTEM STORAGE OF MUSCLES REVEALED BY IMMUNOELECTRON MICROSCOPY

Paratropomyosin is a minor myofibrillar protein which in freshly prepared myofibrils is exclusively localized at the A-I junction region of sarcomeres. We investigated the ultrastructural localization of paratropomyosin in intact and postrigor myofibrils by immunoelectron microscopy. Paratropomyosin was localized as two distinct stripes at the A-I junction in intact myofibrils. It also was localized at the position corresponding to the original A-I junction in thick filament-free myofibrils (I-Z-I brushes). However, following postmortem storage, paratropomyosin was found broadly distributed in thin filaments of myofibrils.     

EVALUATION OF TIME-TEMPERATURE RELATED QUALITY CHANGES IN ICE CREAM DURING STORAGE

The keeping quality of vanilla-flavored ice cream was investigated during 21 weeks of storage. Ice cream samples at 4 different temperature treatments were evaluated for 3 attributes by 14 judges using a deviation-from-reference scale. Sampling tools and presentation containers were designed to provide uniform samples and serving temperatures without melting. The ice cream kept at a variable storage treatment showed a trend toward becoming less firm and darker in yellow than the other samples from the 86th day of storage onward. Differences in creaminess, hardness, and vanilla flavor were noticed in ice cream after 170 days of storage. A commercial time-temperature indicator was used to monitor temperature exposure; but, since indicator response was slower than published, no correlations could be made with quality changes.     

QUANTITATIVE ANALYSIS OF TEXTURE CHANGE IN COD MUSCLE DURING FROZEN STORAGE

The textural deterioration of cod muscle during frozen storage was investigated by objective measurement using the Instron Universal Testing Machine. The rates of texture change as a function of storage time revealed that the increased toughness and decreased cohesiveness could be described by a first-order relationship. Higher storage temperatures resulted in more rapid rates of textural deterioration. Larger activation energy constants for toughness of cod muscle as compared to cohesiveness indicated that toughening was more temperature dependent than cohesiveness. The increased toughness and the decreased cohesiveness of frozen-stored fish appeared to be related to the protein denaturation of the cod muscle and the loss of water-holding capacity, respectively.     

CHANGES IN MOLECULAR SPECIES COMPOSITIONS OF GLYCEROPHOSPHOLIPIDS IN THE ADDUCTOR MUSCLE OF THE GIANT EZO SCALLOP PATINOPECTEN YESSOENSIS DURING FROZEN STORAGE

Changes in lipid components, particularly glycerophospholipids, in the adducto; muscle of giant ezo scallop during storage at −20C were investigated. During storage, the contents of total lipids (TL) and polar lipids (PL) decreased, but that of nonpolar lipids increased. Thiobarbituric acid reactive substances and peroxide values of the TL increased with duration of storage. Glycerylphosphorylcholine (GPC) and glycerylphosphrylethanolamine (GPE) decomposed considerably during storage, and decreased by 50% and 15% of the initial content, while lyso-PC (LPC) and free fatty acids (FFA) increased, respectively. The percentages of polyunsaturated fatty acids (PUFAs) such as 20:5n-3 and 22:6n-3 in the TL and PL during storage decreased, however, those of the PUPAs in the NL increased. Diacylglycerylphosphorylcholine (diacyl-GPC) and diacylglycerylophosphorylethanolamine (diacyl-GPE), major components in the GPC and GPE subclasses, showed a marked decrease with duration of storage. Particularly, diacyl-GPC decreased by 68% of the initial contents of the PL. In the alkenylacyl-GPE and diacyl-GPC, the percentages of the molecular species having longer hydrocarbon chains on the sn-l positions of glycerol moieties decreased at     

CHANGES IN TEXTURE AND MICROSTRUCTURE OF PRESSURE-TREATED FISH MUSCLE TISSUE DURING CHILLED STORAGE

Activities of four endogenous enzymes (cathepsin C, collagenase, chymotrypsin- and trypsin-like enzymes), as well as firmness/strength and elasticity of pressurized fish tissues were monitored over 3 weeks of storage (4–7C). Results indicate that pressurization of fish muscle at 1,000 atm increased firmness whereas pressurization at 2,000 atm or 3,000 atm caused an opposite effect. Changes in tissue elasticity also showed similar trends with correlation between firmness and elasticity. During storage, pressure-inactivated enzymes were reactivated to various extents depending on level of pressurization. Scanning electron microscopy of the tissues revealed some morphological changes with pressurization. At 1,000 atm, there were no significant changes in the myofibers while pressurization at 2,000 atm and 3,000 atm resulted in breakdown of myofibers and connective tissue networks. The results indicate that pressurization may be used to enhance and maintain fresh seafood texture during storage.     

COMPUTER SIMULATION OF VITAMIN DEGRADATION IN A DRY MODEL FOOD SYSTEM DURING STORAGE1

During storage of dry foods, oxidative mechanisms can result in nutrient degradation. Accurate prediction of the vitamin content for dry foods during storage is essential to provide the processor with the assurance of meeting label claims. Since appropriate shelf-life tests for measurement of vitamin degradation represents an alternative worth considering. A mathematical model of the relationship between the rate constant of ascorbic acid oxidation and water activity was established. Computer simulation to predict product moisture content and ascorbic acid degradation in a dry model food product was developed by considering the simultaneous influence of water activity and temperature on rate constants which change with storage time due to varying storage environments. The computer simulation predicts product moisture content and vitamin degradation in good agreement with the experimental data. The simulation will account for the influence of storage temperature and relative humidity as well as characteristics of the food product and package material.