Henrik Hauch Nielsen

Purification and characterization of cathepsin D from herring muscle (Clupea harengus)

Cathepsin D was purified and concentrated 469-fold from a homogenate of Clupea harengus muscle. The purified enzyme is a monomer with a molecular weight of 38000-39000. It is inhibited by pepstatin and has optimal activity at pH 2.5 with hemoglobin as the substrate. The isoelectric point is at pH 6.8. Glycosidase treatment and binding to Concanavalin A indicated that the enzyme contains one N-linked carbohydrate moiety of the high-mannose type per molecule. The first 21 amino acid residues of the N-terminal showed high similarity to cathepsin D from antarctic icefish liver (Chionodraco hamatus) and trout ovary (Oncorhynchus mykiss). Digestion of the beta-chain of oxidized insulin resulted in preferential cleavage at Leu(15)-Tyr(16), (47%), Tyr(16)-Leu(17) (34%) and Ala(14)-Leu(15) (18%). Incubation with myofibrils from herring muscle at pH 4.23 showed that the enzyme mainly degraded myosin, actin and tropomyosin.

Oxidative changes during ice storage of rainbow trout (Oncorhynchus mykiss) fed different ratios of marine and vegetable feed ingredients

Recently fish meal and oil have increasingly been replaced with proteins and oils from vegetable sources in the diets of farmed salmonids, but the consequences for the oxidative stability of the resulting fish products have not been investigated. The aim of the present study was to evaluate the influence of feeding regime on composition of rainbow trout fillets, as well as on lipid and protein oxidation during storage on ice. Rainbow trout were fed six different diets, which differed in their levels of marine oil and proteins vs. vegetable oil and protein. Fish fillets were characterised by measurement of fatty acid and amino acid composition, primary and secondary lipid oxidation products, astaxanthin and tocopherol content. Protein oxidation was assessed by measuring protein carbonyl content, oxidised amino acids, sulfhydryl groups and immuno-blotting against carbonyl groups. Feeding regimes significantly influenced fatty acid composition. Replacement of fish oil with vegetable oil reduced formation of primary oxidation products, but the effect on secondary oxidation products differed between different types of volatiles. The differences in protein and amino acid composition were not significant, and there were no clear effects of diets on protein oxidation, but data indicated that compounds present in the marine ingredients might have had an effect on protein oxidation.

The effect of farmed trout on cardiovascular risk markers in healthy men

Increased intake of marine long-chain n-3 PUFA (n-3 LCPUFA) may decrease the risk of CVD and reduce mortality by lowering serum TAG and blood pressure (BP). Furthermore, n-3 LCPUFA may affect novel CVD risk markers related to inflammation and vascular function. The objective of the present study was to examine the effect of farmed trout on novel and traditional CVD risk markers in healthy men, and to evaluate whether this was affected by the aquacultural feed regime. We performed a parallel, 8-week intervention study in which sixty-eight healthy male volunteers were randomised to consume either a daily meal with 150 g farmed trout raised on either marine or vegetable-based feed, or a reference meal containing 150 g chicken. Twenty-four hour BP, pulse wave velocity, augmentation index, fatty acid composition of erythrocyte (RBC), and concentrations of TAG, HDL-cholesterol, LDL-cholesterol, glucose, insulin, C-reactive protein (CRP) and other markers of inflammation were measured at weeks 0 and 8. RBC content of total n-3 LCPUFA, both EPA and DHA, was significantly higher among men consuming trout raised on marine feed compared with men consuming the vegetable-fed trout or chicken. The three intervention groups did not differ significantly with respect to any of the other outcome variables, although there were trends towards associations between the changes in RBC n-3 LCPUFA and those in BP and CRP. In the present study, we conclude that we could not confirm the fish oil-induced reduction in CVD risk markers after daily consumption of trout with high or low n-3 LCPUFA content. However, trout raised on vegetable-based feed had less pronounced impact on RBC n-3 LCPUFA status.

Chemical Characterization, Antioxidant and Enzymatic Activity of Brines from Scandinavian Marinated Herring Products

Brines generated during the last marination step in the production of marinated herring (Clupea harengus) were chemically characterized and analyzed for antioxidant and enzyme activities. The end-products were vinegar cured, spice cured and traditional barrel-salted herring with either salt or spices. The chemical characterization encompassed pH, dry matter, ash, salt, fatty acids, protein, polypeptide pattern, iron and nitrogen. The antioxidant activity was tested with three assays measuring: iron chelation, reducing power and radical scavenging activity. The enzymatic activity for peroxidase and protease were also tested. Results revealed that the brine can contain up to 56.7 mg protein/ mL, up to 20.1 mg fatty acid/mL, good antioxidant activity, high amounts of the antioxidative amino acids lysine, alanine, and glycine, and high enzymatic activity. The potential of using the protein-rich fraction with biological activity from brines from the marinated herring production was demonstrated in this work.

Sensory properties of marinated herring ( Clupea harengus ) - influence of fishing ground and season

ABSTRACT The sensory properties of marinated herring produced immediately post mortem of raw material from different fishing ground and seasons were described and related to biological, biochemical and functional properties. Subtle variation was encountered in the appearance of whole marinated herring fillets. Fishing ground did not influence the odor, flavor or texture, but there was an apparent effect of season on the sensory profile. The sensory properties were influenced by body weight, but not by age, sex and gonad maturity. The influence of varying lipid content, water content and liquid holding capacity resulted in similar effects showing the high correlation between these properties. The results indicated that variation in sensory quality observed by the industry is not primarily due to the parameters fishing ground and season.

In situ activity of chymotrypsin in sugar-salted herring during cold storage

In situ activity of intestinal chymotrypsin in sugar-salted whole herring during cold storage was evaluated by analysing changes in the low-molecular-weight nitrogen fraction when a specific inhibitor was added. Addition of chymostatin (0.01 mM) to sugar-salted herring gave 100% inhibition of chymotrypsin activity compared to sugar-salted herring without chymostatin. Inhibition of chymotrypsin did not affect the profile of low-molecular-weight peptides analysed by capillary zone electrophoresis, but resulted in 22% lower content after 26 weeks of storage. Inhibition of chymotrypsin resulted in 17% lower content of free amino acids. The relative amount of single free amino acids was unaffected by the presence of inhibitor, except for valine, leucine, isoleucine, methionine, phenylalanine and tyrosine, which were formed in lower relative amount. Removal of intestines (gutted herring) and thereby all intestinal proteolytic activity did not change the free amino acid profile, except for the concentration of histidine which decreased more when intestines were present.

Purification and characterization of elastase from the pyloric caeca of rainbow trout (Oncorhynchus mykiss)

Abstract 1. 1. An elastase-like enzyme was purified from the pyloric caeca of rainbow trout by hydrophobic interaction, cation exchange and gel-filtration chromatography. 2. 2. The approximate molecular weight of the elastase was 27 kDa and the isoelectric point was remarkably basic. 3. 3. The pH optimum of this enzyme was 8.0, when assayed with Succinyl-Ala-Ala-Ala- p -Nitroanilide. 4. 4. When assayed with Succinyl-Ala-Ala-Ala- p -Nitroanilide, the enzyme activity had a temperature optimum of 45°C, and the enzyme was stable up to this temperature. 5. 5. The trout elastase exhibited a higher specific activity than porcine elastase against Succinyl-Ala-Ala-Ala- p -Nitroanilide and elastin-orcein. 6. 6. The trout elastase was inhibited by elastatinal, PMSF, TPCK, SBTI and Bowman-Birk inhibitor.

Chemical processes responsible for quality deterioration in fish.

Abstract: The chapter focuses on fish as seafood and introduces the characteristics of fish that make them different from muscle food of other animal origin. The major chemical degradation processes responsible for quality deterioration in fish including enzymatic degradation of proteins, lipids and trimethylamine-N-oxide (TMAO) as well as oxidative reactions are reviewed. Methods to prevent these reactions are discussed.

Influence of Dietary Lipid and Protein Sources on the Sensory Quality of Organic Rainbow Trout (Oncorhynchus mykiss) After Ice Storage

The influence of dietary protein and lipid sources on the quality of organic rainbow trout (Oncorhynchus mykiss) was studied. The protein and oil sources were fishmeal, fish oil, and organic vegetable protein and oils. Sensory profiling was performed during 3 to 14 days of ice storage along with lipid analyses of the fillet. Overall, the results showed that the sensory characteristics of the trout were affected in different ways during ice storage. The source of lipid seemed to affect the sensory quality at the beginning of the storage period, while the protein source seemed to have a more pronounced impact at the end of the storage period.

Vegetable based fish feed changes protein expression in muscle of rainbow trout ( Oncorhynchus mykiss )

Feed production for aquaculture of carnivore fish species relies heavily on protein and lipid from the limited resources of wild fish and other sea living organisms. Thus the development of alternative feeds replacing fish meal and oil with alternatives of e.g. vegetable origin is important for a sustainable production of fish from aquaculture (Glencross et al. 2007). Such a change in feed ingredients will affect the metabolic pathways in fish and during the last years genomic and proteomic techniques have contributed in obtaining a better understanding of the involved mechanisms (Panserat & Kaushik 2010; Martin et al. 2003; Vilhelmsson et al. 2004). A consequence of fish meal and fish oil replacement in the feed may also be a change of eating quality (Johnsen et al. 2011; Lesiow et al. 2009).