Revilija Mozuraityte

Quality changes of salmon by-products during storage: Assessment and quantification by NMR

Safe utilization of fish by-products is an important task due to increasing fish consumption. It can provide new valuable food/feed and will increase the economical profit and sustainability of the fishery industry. NMR spectroscopy is a reliable tool able to monitor qualitative and quantitative changes in by-products. In this work the trichloroacetic acid extracts of salmon backbones, heads and viscera stored at industrially relevant temperatures (4 and 10°C) were studied using NMR. Twenty-five metabolites were detected and the possibility of salmon by-products utilization as a source of anserine, phosphocreatine and taurine was discussed. Statistical data elaboration allowed determining the main processes occurring during by-products storage: formation of trimethylamine and biogenic amines, proteolysis and different types of fermentations. By-products freshness was evaluated using a multi-parameter approach: the trimethylamine and biogenic amines concentration changes were compared with Ki and H-values and safe temperatures and times for storage of salmon by-products were proposed.

Kinetic studies of lipid oxidation induced by hemoglobin measured by consumption of dissolved oxygen in a liposome model system.

The effect of hemoglobin (Hb) and lipid concentration, pH, temperature, and different antioxidants on heme-mediated lipid oxidation of liposomes from marine phospholipids was studied. The rate of lipid oxidation was measured by consumption of dissolved oxygen. Heme-mediated lipid oxidation at different Hb and lipid concentrations was modeled by Michaelis-Menten kinetics. The maximum rate (V(max)) for the reaction with cod and bovine Hb as a pro-oxidant was 66.2 +/- 3.4 and 56.6 +/- 3.4 microM/min, respectively. The Michaelis-Menten constant (K(m)) for the reaction with cod and bovine Hb was 0.67 +/- 0.09 and 1.2 +/- 0.2 microM, respectively. V(max) for the relationship between the oxygen uptake rate and lipid concentration was 43.2 +/- 1.5 microM/min, while the K(m) was 0.93 +/- 0.14 mg/mL. The effect of the temperature followed Arrhenius kinetics, and there was no significant difference in activation energy between cod and bovine Hb. The rate of lipid oxidation induced by bovine Hb was highest around pH 6. Ethylenediaminetetraacetic acid (EDTA) had no significant effect on heme-mediated lipid oxidation, but alpha-tocopherol and astaxanthin worked well as antioxidants. Kinetic differences were found between iron and Hb as pro-oxidants, and the efficacy of the antioxidants depended upon the pro-oxidant in the system.

Bioactivities of fish protein hydrolysates from defatted salmon backbones

Graphical abstract

The effect of dietary antioxidants on iron-mediated lipid peroxidation in marine emulsions studied by measurement of dissolved oxygen consumption

Long chain omega-3 polyunsaturated fatty acids (LC omega-3 PUFA) are vital for physiological functions and have therapeutic and health benefits. The consumption of LC PUFA in the Western world has been below recommended intake levels the past decades, despite promotion of seafood and omega-3 supplements. Incorporation of the LC PUFA into processed food consumed on a daily basis might therefore bridge the gap between the recommended and actual consumption of LC omega-3 PUFA. Unfortunately, the development of omega-3 enriched food is hampered by a very high susceptibility of LC PUFA to oxidative deterioration. Furthermore, oxidised lipids are believed to create health risks. It has also been suggested that gastric juice may deteriorate the healthy LC PUFA after they are ingested. Important lipid oxidation promoters in food are low molecular weight (LMW) iron (Fe) and methemoglobin (metHb). To incorporate the LC omega-3 PUFA safely into food with respect to oxidation, it is necessary to understand both Fe- and metHb-mediated oxidation of PUFA and how the oxidation is influenced by conditions and dietary antioxidants.The main objective of this thesis is therefore to study Fe- and metHb-mediated lipid oxidation in food-related lipid model systems – emulsions stabilised with phospholipids and liposomes made of phospholipids – containing LC omega-3 PUFA from fish. The focus was on clarifying the reaction mechanisms and the impact of different factors, including dietary antioxidants and gastric juice, on the prooxidant activity of Fe and metHb. Measurement of the consumption rate of the essential substrate for lipid oxidation – oxygen – by the LC PUFA was used for assessment of lipid oxidation.The continuous measurement of the dissolved oxygen concentration has been shown to be a robust method for direct and instantaneous monitoring of peroxidation in both the liposomes and emulsions. The method was especially useful for measurement of the oxygen consumption kinetics in the lipid systems. The determination of oxygen uptake rates (OUR) enabled screening and evaluation of the impact of the different factors and antioxidants on the prooxidant activity of Fe and metHb.Pre-formed lipid hydroperoxides (LOOH) were shown to be essential for the prooxidant activity of both Fe and metHb, and the prooxidant activity of metHb was not affected by the lack of light. The oxygen uptake kinetics revealed that iron behaved as a catalyst in lipid oxidation while the prooxidant activity of metHb weakened over time, presumably due to degradation of meHb molecule during lipid oxidation. MetHb was shown to be a stronger prooxidant than Fe, but the strong prooxidative activity was facilitated by a complete structure of the metHb molecule. The prooxidant mechanism of both Fe and metHb was not limited by the level of dissolved oxygen, as long as oxygen was present, or the level of pre-formed LOOH and double bonds in fatty acids, as long as they were present in higher concentrations than the prooxidant.The extent of the prooxidative activity of Fe was shown to vary in dependence on:The total surface area: Smaller liposomal vesicles with lower lipid content were more prone to oxidation than larger emulsion droplets with a higher lipid content, presumably due to more frequent interactions of Fe with pre-formed LOOH in the interphase.The amount of phospholipid emulsifier: Higher levels of phospholipids resulted in the formation of smaller droplets. The highest OUR were measured for emulsifier concentrations ranging from 5 – 10% (w/w lipid base).pH of the aqueous phase: Fe-mediated oxidation was highest at pH interval 4.5 – 5.5.Dissolved compounds: Sodium chloride (NaCl) and 0.2% of xanthan gum dissolved in the aqueous phase inhibited Fe-mediated oxidation in a concentration dependent manner.Electrostatic retention of Fe by phosphate groups within phospholipid heads has been suggested to facilitate the contact between pre-formed LOOH and Fe, and to create competitive reactions for iron precipitation at pH > 5 and iron complexation by chelating compounds.The activity of dietary antioxidants has been shown to be affected by the type of prooxidant in the lipid system. Ascorbic acid, caffeic acid, propyl gallate, astaxanthin, ascorbyl palmitate, α-tocopherol, and δ-tocopherol inhibited metHb-mediated oxidation in concentration dependent manners. EDTA had a minor effect on metHb-mediated oxidation.In Fe-mediated oxidation, caffeic acid, ascorbic acid and α-tocopherol were prooxidants. They directly interacted with Fe, reducing Fe3+ to the more catalytically active Fe2+. The magnitude of the pro-oxidative behaviour was dependent on the Fe-to-antioxidant ratio, antioxidant concentration and pH. Ascorbic acid was depleted by interactions with Fe, and decreased the pro-oxidative activity of α-tocopherol. EDTA and citric acid inhibited Fe-mediated oxidation completely at twice the ratio to Fe and pH > 3.5. Propyl gallate efficiently inhibited Fe-mediated oxidation, while astaxanthin and β-carotene had only minor effects. In addition, chemical structure and physical location of the antioxidants determined their effects.The work in this thesis shows that for correct interpretation of the effects of antioxidants it is important to assess what types of prooxidants are present in the system.Both gastric juice and hydrochloric acid solution (HCl) did not prevent oxidation of marine lipids in emulsions and liposomes (pH 4.0). Furthermore, gastric juice did not inhibit metHb-mediated oxidation, but it was capable of reducing the prooxidant activity of dietary LMW iron, compared to HCl solution. Berry juice, green tea, red wine, and caffeic acid reduced the OUR in the acidic environments while coffee, ascorbic acid and orange juice increased the OUR. Therefore, beverages accompanying foods rich in marine lipids will affect the course of post-prandial lipid oxidation.

Oxidation of Food Components

This article gives basic introduction into the mechanisms of oxidation of food lipids and proteins. It contains key chemical background for each of the mechanisms where lipid oxidation products are introduced. The main factors affecting lipid oxidation rates are presented, as well as the concept of pro- and antioxidants involved in lipid oxidation. For protein oxidation, the main reactions and processes taking place as well as the effect of oxidation on the properties of proteins are described.

Fish Oils: Production and Properties

The global fish oil production is estimated to be about 1 million tons per year. Fish oil and meal is traditionally produced by using the wet rendering method. The process involves cooking of the raw material, pressing, and separation of the oil. Enzymatic hydrolysis is an alternative process for oil and protein production in which proteases are used to facilitate the degradation of fish tissue and the liberation of oil. Oil intended for human consumption is further processed after production to remove impurities and unwanted components.

Quality of Filleted Atlantic Mackerel (Scomber Scombrus) During Chilled and Frozen Storage: Changes in Lipids, Vitamin D, Proteins, and Small Metabolites, including Biogenic Amines

ABSTRACT Quality changes of vacuum-packed Atlantic mackerel (Scomber scombrus) fillets during 12 months’ frozen storage at −27°C and 9 days’ chilled storage at +4°C were evaluated. Freezing at −27°C preserved the long chain n-3 polyunsaturated fatty acids (LC n-3 PUFAs), both in light and dark muscle, vitamin D, and the low molecular weight metabolites (LMW) (studied by high resolution nuclear magnetic resonance spectroscopy, HR NMR). Protein oxidation took place, especially between 1 and 7 months, decreasing water holding capacity and protein extractability. During chilled storage, no lipid or protein oxidation was observed, but lipolysis increased, and several LMW metabolites relevant for sensory and nutritional quality degraded into non-favorable compounds. The content of biogenic amines was high at day 9 (e.g., 18 mg histamine/100 g), jeopardizing safety. Preservation of mackerel fillets by freezing at −27°C is thus a better option compared to prolonged chilled storage at +4°C; the quality was well preserved for 12 months’ frozen storage.

Two-stage processing of salmon backbones to obtain high-quality oil and proteins

Traditional processing technologies for fish by-products containing significant amounts of oils usually either give high amounts of oil or maximised solubilisation of proteins. Due to lower yields and insufficient quality, the proteins or the oil is considered as secondary products. The proposed concept combines a gentle thermal separation of oil followed by enzymatic hydrolysis of the remaining protein-rich fraction. The first stage, thermal treatment (40 °C) of fresh salmon backbones, separated up to 85% of the oil from the raw material and gave high-quality oil (PV = 0.2 0.0 meq kg , 0.16 0.05% free fatty acids). Separation of a significant part of the oil gave reduced mass flow into the enzymatic stage, which then requires less enzymes and reduced energy consumption. Among the tested enzymes: Trypsin, Corolase PP and Mixture of Papain and Bromelain gave the highest yield of fish protein hydrolysates (FPH), while use of Protamex and Corolase PP resulted in FPH with the best sensory properties leading to the lowest bitterness.

Oxidative Stability and Shelf Life of Fish Oil

Abstract Fish oils are rich in long-chain omega-3 polyunsaturated fatty acids that are highly prone to oxidation. Therefore, lipid oxidation is the limiting factor for the shelf life of fish oils and the knowledge of factors affecting oxidation is crucial. This chapter, together with cited literature, provides insight into the different approaches and strategies that have been developed by both industry and academia to sustain the oxidative stability of highly unsaturated fish oils, including omega-3 concentrates. The chapter also discusses some challenges related to the measurement of oxidative stability of fish oils by advanced and classic oxidation status methods.

Effect of dietary antioxidants on iron-mediated peroxidation in emulsions studied by dissolved oxygen consumption

Long chain omega-3 polyunsaturated fatty acids (LC omega-3 PUFA) are vital for physiological functions and have therapeutic and health benefits. The consumption of LC PUFA in the Western world has been below recommended intake levels the past decades, despite promotion of seafood and omega-3 supplements. Incorporation of the LC PUFA into processed food consumed on a daily basis might therefore bridge the gap between the recommended and actual consumption of LC omega-3 PUFA. Unfortunately, the development of omega-3 enriched food is hampered by a very high susceptibility of LC PUFA to oxidative deterioration. Furthermore, oxidised lipids are believed to create health risks. It has also been suggested that gastric juice may deteriorate the healthy LC PUFA after they are ingested. Important lipid oxidation promoters in food are low molecular weight (LMW) iron (Fe) and methemoglobin (metHb). To incorporate the LC omega-3 PUFA safely into food with respect to oxidation, it is necessary to understand both Fe- and metHb-mediated oxidation of PUFA and how the oxidation is influenced by conditions and dietary antioxidants.The main objective of this thesis is therefore to study Fe- and metHb-mediated lipid oxidation in food-related lipid model systems – emulsions stabilised with phospholipids and liposomes made of phospholipids – containing LC omega-3 PUFA from fish. The focus was on clarifying the reaction mechanisms and the impact of different factors, including dietary antioxidants and gastric juice, on the prooxidant activity of Fe and metHb. Measurement of the consumption rate of the essential substrate for lipid oxidation – oxygen – by the LC PUFA was used for assessment of lipid oxidation.The continuous measurement of the dissolved oxygen concentration has been shown to be a robust method for direct and instantaneous monitoring of peroxidation in both the liposomes and emulsions. The method was especially useful for measurement of the oxygen consumption kinetics in the lipid systems. The determination of oxygen uptake rates (OUR) enabled screening and evaluation of the impact of the different factors and antioxidants on the prooxidant activity of Fe and metHb.Pre-formed lipid hydroperoxides (LOOH) were shown to be essential for the prooxidant activity of both Fe and metHb, and the prooxidant activity of metHb was not affected by the lack of light. The oxygen uptake kinetics revealed that iron behaved as a catalyst in lipid oxidation while the prooxidant activity of metHb weakened over time, presumably due to degradation of meHb molecule during lipid oxidation. MetHb was shown to be a stronger prooxidant than Fe, but the strong prooxidative activity was facilitated by a complete structure of the metHb molecule. The prooxidant mechanism of both Fe and metHb was not limited by the level of dissolved oxygen, as long as oxygen was present, or the level of pre-formed LOOH and double bonds in fatty acids, as long as they were present in higher concentrations than the prooxidant.The extent of the prooxidative activity of Fe was shown to vary in dependence on:The total surface area: Smaller liposomal vesicles with lower lipid content were more prone to oxidation than larger emulsion droplets with a higher lipid content, presumably due to more frequent interactions of Fe with pre-formed LOOH in the interphase.The amount of phospholipid emulsifier: Higher levels of phospholipids resulted in the formation of smaller droplets. The highest OUR were measured for emulsifier concentrations ranging from 5 – 10% (w/w lipid base).pH of the aqueous phase: Fe-mediated oxidation was highest at pH interval 4.5 – 5.5.Dissolved compounds: Sodium chloride (NaCl) and 0.2% of xanthan gum dissolved in the aqueous phase inhibited Fe-mediated oxidation in a concentration dependent manner.Electrostatic retention of Fe by phosphate groups within phospholipid heads has been suggested to facilitate the contact between pre-formed LOOH and Fe, and to create competitive reactions for iron precipitation at pH > 5 and iron complexation by chelating compounds.The activity of dietary antioxidants has been shown to be affected by the type of prooxidant in the lipid system. Ascorbic acid, caffeic acid, propyl gallate, astaxanthin, ascorbyl palmitate, α-tocopherol, and δ-tocopherol inhibited metHb-mediated oxidation in concentration dependent manners. EDTA had a minor effect on metHb-mediated oxidation.In Fe-mediated oxidation, caffeic acid, ascorbic acid and α-tocopherol were prooxidants. They directly interacted with Fe, reducing Fe3+ to the more catalytically active Fe2+. The magnitude of the pro-oxidative behaviour was dependent on the Fe-to-antioxidant ratio, antioxidant concentration and pH. Ascorbic acid was depleted by interactions with Fe, and decreased the pro-oxidative activity of α-tocopherol. EDTA and citric acid inhibited Fe-mediated oxidation completely at twice the ratio to Fe and pH > 3.5. Propyl gallate efficiently inhibited Fe-mediated oxidation, while astaxanthin and β-carotene had only minor effects. In addition, chemical structure and physical location of the antioxidants determined their effects.The work in this thesis shows that for correct interpretation of the effects of antioxidants it is important to assess what types of prooxidants are present in the system.Both gastric juice and hydrochloric acid solution (HCl) did not prevent oxidation of marine lipids in emulsions and liposomes (pH 4.0). Furthermore, gastric juice did not inhibit metHb-mediated oxidation, but it was capable of reducing the prooxidant activity of dietary LMW iron, compared to HCl solution. Berry juice, green tea, red wine, and caffeic acid reduced the OUR in the acidic environments while coffee, ascorbic acid and orange juice increased the OUR. Therefore, beverages accompanying foods rich in marine lipids will affect the course of post-prandial lipid oxidation.