Characterization of aqueous components in chicken breast muscle as inhibitors of hemoglobin-mediated lipid oxidation

Unheated press juice (PJ) obtained from chicken breast muscle was a potent inhibitor of hemoglobin-mediated lipid oxidation in washed cod muscle. The <1 kDa fraction had a negligible effect on the rate of lipid oxidation. The high-molecular-weight (HMW) fraction was mildly inhibitory when added alone and highly inhibitory in the presence of <1 kDa components. Proteins of the HMW fraction were further fractionated by ammonium sulfate precipitation. Proteins in the 80% fraction were most inhibitory compared with other precipitated fractions on an equal protein basis. Inhibition by PJ was substantially decreased due to treatment with ascorbate oxidase. Adding ascorbate to the HMW fraction did not increase its inhibition, which suggested the presence of a complex ascorbate-reducing system in PJ consisting of HMW and low-molecular-weight (LMW) components. The ability of added ceruloplasmin to inhibit lipid oxidation was remarkably enhanced by addition of ascorbate or the <1 kDa fraction. Heated and centrifuged PJ had 8 times more LMW iron compared to unheated PJ. Adding heated PJ to washed cod containing hemoglobin slightly increased the rate and extent of lipid oxidation.     

Deoxyhemoglobin-mediated lipid oxidation in washed fish muscle

Deoxyhemoglobin-mediated lipid oxidation was studied by comparing the pro-oxidative activity of anodic and cathodic hemoglobins from trout in a washed cod muscle model system. At pH 6.3, cathodic hemoglobins were nearly fully oxygenated while anodic hemoglobins were poorly oxygenated. Anodic hemoglobins initiated lipid oxidation in washed cod muscle much more rapidly than cathodic hemoglobins, as measured by thiobarbituric acid reactive substances (TBARS) formation. Moreover, anodic hemoglobins appeared to oxidize more rapidly as compared to cathodic hemoglobins in the washed cod muscle model system, as measured by a decrease in redness (a value). A more pronounced pro-oxidative activity of deoxyhemoglobin as compared to oxyhemoglobin was confirmed by accelerated lipid hydroperoxide and TBARS formation in the washed cod muscle model system upon combined addition of anodic hemoglobins and adenosine triphosphate, which is known to lower the oxygenation of anodic hemoglobins at pH 7.2, as compared to only addition of anodic hemoglobins to the washed cod muscle. These studies suggest that deoxyhemoglobin is more pro-oxidative than its oxygenated counterpart at pH values found in postmortem fish muscle.     

Contributions of blood and blood components to lipid oxidation in fish muscle.

There was a wide variation in the amounts of hemoglobin extracted from the muscle tissue of bled and unbled fish. Averaged values suggested that the residual blood level in the muscle of bled fish was substantial. Myoglobin content was minimal as compared to hemoglobin content in mackerel light muscle and trout whole muscle. Hemoglobin made up 65 and 56% of the total heme protein by weight in dark muscle from unbled and bled mackerel, respectively. Bleeding significantly reduced rancidity in minced trout whole muscle, minced mackerel light muscle, and intact mackerel dark muscle but not minced mackerel dark muscle stored at 2 degrees C. The reduction was in the number of fish that had a longer shelf life; muscle from certain bled fish had rancidity that was comparable to the rancidity in unbled controls. The soluble contents of erythrocytes accounted for all of the lipid oxidation capacity of whole blood added to washed cod muscle. Limiting lysis of erythrocytes delayed lipid oxidation, which was likely due to keeping hemoglobin inside the erythrocyte. Apparent breakdown of lipid hydroperoxides occurred only when a critical level of hemoglobin was present. Blood plasma was slightly inhibitory to oxidation of washed cod lipids. These studies suggest that blood-mediated lipid oxidation in fish muscle depends on various factors that include hemoglobin concentration, types of hemoglobin, plasma volume, and erythrocyte integrity.     

Mechanisms of heme protein-mediated lipid oxidation using hemoglobin and myoglobin variants in raw and heated washed muscle

The hemoglobin variant rHb 0.1, which possesses a decreased ability to form subunits, stimulated lipid oxidation in washed fish muscle less effectively as compared to wild-type hemoglobin (rHb 0.0). This could be due to the lower hemin affinity and more rapid autoxidation rate of subunits as compared to tetramers. To differentiate between hemin affinity and autoxidation effects, ferrous V68T Mb was compared to ferrous wild-type myoglobin (WT Mb). WT Mb has a more rapid hemin loss rate (25-fold) than does V68T, while V68T autoxidized more rapidly than did WT Mb (60-fold). Ferrous WT Mb promoted TBARS and lipid peroxide formation more rapidly than did ferrous V68T (p < 0.01). This indicated hemin loss rate was more critical in determining onset of lipid oxidation as compared to autoxidation rate. Hemin alone was capable of stimulating lipid oxidation. Albumin enhanced the ability of hemin to promote lipid oxidation. MetMb promoted lipid oxidation more effectively than did ferrous Mb, which could be due to the lower hemin affinity of metMb as compared to that of ferrous Mb. EDTA, an iron chelator, had no effect on the rate or extent of lipid oxidation mediated by Mb in the cooked system. Variants with a 975-fold range of hemin affinities promoted lipid oxidation with equivalent efficacy in cooked washed cod contrary to results in uncooked washed cod. The cooking temperatures apparently denature the globin and release hemin reactant to such an extent that the impact of hemin affinity on lipid oxidation observed in the raw state is negated in the cooked state. These studies collectively suggest released hemin is of primary importance in promoting lipid oxidation in raw and cooked washed fish muscle.     

Effects of fish heme protein structure and lipid substrate composition on hemoglobin-mediated lipid oxidation

Hemoglobin (Hb) promoted lipid oxidation more effectively in washed tilapia as compared to washed cod in spite of a 2.8-fold higher polyenoic index in the washed cod. This suggested that increasing the fatty acid unsaturation of the substrate did not accelerate the onset of lipid oxidation. Substantial phospholipid hydrolysis in the washed cod was observed, which has the potential to inhibit lipid oxidation. MetHb formation and lipid oxidation occurred more rapidly at pH 6.3 as compared to pH 7.4. Trout Hb autoxidized faster and was a better promoter of lipid oxidation as compared to tilapia Hb. The greater ability of trout Hb to promote lipid oxidation was attributed in part to its lower conformational and structural stability based on secondary and tertiary structure, acid-induced unfolding, and thermal aggregation measurements. It is suggested that the structural instability and lipid oxidation capacity of trout Hb were at least partly due to low hemin affinity. Trout and tilapia Hb were equivalent in their ability to cause lipid oxidation in washed cod muscle heated to 80 degrees C. Apparently, these high temperatures denature both trout and tilapia Hb to such an extent that any differences in conformational stability observed at lower temperatures were negated.     

Comparative analysis of different hemoglobins: autoxidation,reaction with peroxide,and lipid oxidation

Beef hemoglobin (Hb) had lower levels of deoxyHb and autoxidized much slower as compared to trout Hb at pH 6.3. Chicken Hb autoxidized at a rate intermediate between beef and trout Hb. In the presence of hydrogen peroxide, metHb formed rapidly from trout Hb whereas beef Hb was essentially nonreactive with hydrogen peroxide. The autoxidation rate of perch Hb was more rapid than trout Hb despite the low deoxyHb content of perch Hb. Perch Hb was a better catalyst of lipid oxidation than trout Hb when added to washed cod muscle based on formation of lipid hydroperoxides and thiobarbituric acid reactive substances. These studies indicate that autoxidation rate does not always increase with increasing deoxyHb content. The role of heme crevice volume in heme protein autoxidation is discussed. Among other factors, these studies suggest that rates of lipid oxidation in various muscle foods may depend on the relative ability of hemoglobins from different animal species to promote lipid oxidation.     

Studies with myoglobin variants indicate that released hemin is the primary promoter of lipid oxidation in washed fish muscle

Variants of sperm whale myoglobin (Mb) were used to assess the mechanism of heme protein-mediated lipid oxidation in washed cod muscle. A myoglobin variant with high hemin affinity (V68T) was an exceptionally poor promoter of lipid oxidation, while a Mb variant with low hemin affinity (H97A) was a potent promoter of lipid oxidation. V68T releases hemin slowly due to the ability of threonine to hydrogen bond with coordinated water and the distal histidine within the heme crevice. H97A rapidly releases hemin because the relatively small alanine residue creates a channel for water to easily enter the heme crevice which weakens the covalent linkage of hemin to the proximal histidine. A variant sensitive to heme degradation (L29F/H64Q) was a weaker promoter of lipid oxidation compared to wild-type Mb. This suggests that degrading the heme ring and releasing iron decreased the ability of Mb to promote lipid oxidation. Free radicals resulting from hemin-mediated decomposition of lipid hydroperoxides have the capacity to propagate lipid oxidation and degrade hemin catalyst. This may explain why heme proteins behave as reactants rather than "catalysts" of lipid oxidation in washed cod. Collectively these studies strongly suggest that released hemin is the critical entity that drives heme protein-mediated lipid oxidation in washed fish muscle.     

Added triacylglycerols do not hasten hemoglobin-mediated lipid oxidation in washed minced cod muscle

Hemoglobin-mediated lipid oxidation in washed, minced cod muscle was related to the triacylglycerol to membrane lipid ratio. The same rapid development of thiobarbituric acid reactive substances (TBARS) and painty odor occurred with and without the presence of up to 15% menhaden oil. Without hemoglobin, development of TBARS and painty odor was slow, despite a high amount of hydroperoxides in samples with oil added (1135 micromol/kg muscle). This suggested that hemoglobin reacted by cleaving preformed hydroperoxides into secondary oxidation products. Nearly doubling the hemoglobin concentration approximately doubled the extent of lipid oxidation with and without added oil. This indicated that hemoglobin was limiting for the oxidation reaction. The noneffect of added oil suggests that membrane lipids and/or preformed membrane lipid hydroperoxides provided sufficient substrate in hemoglobin-catalyzed oxidation of washed minced cod muscle. Fe(2+-)ADP did not induce any oxidation of washed minced cod with/without added oil. Results suggest that lipid oxidation in fatty fish may be more related to the quantity and type of the aqueous pro-oxidant and the membrane lipids than to variations in total fat contents.     

Hemoglobin-mediated lipid oxidation and compositional characteristics of washed fish mince model systems made from cod (Gadus morhua), herring (Clupea harengus), and salmon (Salmo salar) muscle

The use of washed cod light muscle minces in mechanistic studies of hemoglobin (Hb)-mediated fish lipid oxidation has largely increased in the past 5 years. Although cod light muscle has a low level of intrinsic lipid oxidation catalysts, a prerequisite for a good oxidation model system, we believe it cannot fully mimic the oxidation kinetics taking place in other fish species being more susceptible to lipid oxidation. The aim of this study was to systematically investigate whether washed mince model systems useful in Hb-mediated oxidation studies could be prepared also from herring (Clupea harengus) and salmon (Salmo salar) light muscles. The kinetics of oxidation in the washed models was measured during ice storage (+/-Hb), and the results were related to compositional differences. Minces from cod, herring, and salmon light muscles were washed 3 times with 3 volumes of water and buffer. A 20 microM portion of Hb and 200 ppm streptomycin was then added, followed by adjustment of pH and moisture to 6.3 and 86%, respectively. Samples with or without Hb were then stored on ice, and oxidation was followed as peroxide value (PV), rancid odor, redness (a*) loss and yellowness (b*). Prior to storage, all minces and models were also analyzed for total lipids, fatty acids, alpha-tocopherol, proteins, Hb, Fe, Cu, and Zn. Hb-mediated lipid oxidation appeared within 2 days on ice in all models. Small differences in the oxidation rates ranked the models as herring > cod > salmon. These differences were ascribed to more preformed peroxides and trace elements in the herring model, and more antioxidants in the salmon model. Controls, without Hb, stayed stable in all cases except herring, where a very slight oxidation appeared, especially if the herring raw material had been prefrozen. In conclusion, fattier fish like dark muscle species and salmonoids are useful for making washed mince model systems and would be a better choice than cod if there is an interest in the oxidation kinetics of such species.     

Effect of pH on hemoglobin-catalyzed lipid oxidation in cod muscle membranes in vitro and in situ

The effect of pH and hemoglobin on oxidation of the microsomal lipids of cod was determined in isolated microsomes and in washed cod muscle. An increase of hemoglobin concentration from 0.5 to 15 microM accelerated lipid oxidation in both systems. In cod microsomes the rate of lipid oxidation increased in the order pH 6.8 > pH 7.6 > pH 8.4 > pH 6.0 > pH 3.5. However, in washed cod muscle a decrease of pH from 7.8 to 6.8 greatly increased the lag phase and decreased the rate of lipid oxidation. A further decrease in pH to 3.5 decreased the lag phase and increased the rate of lipid oxidation further. A decrease of pH from 7.6 to 6.4 greatly reduced the affinity of hemoglobin for oxygen. Formation of methemoglobin due to autoxidation occurred more rapidly at pH 6.0 than at pH 7.5. Structural changes of the isolated microsomal membranes could be the reason for the unexpected slow lipid oxidation in microsomes at pH 6.0 and below.     

Effect of low pH on the susceptibility of isolated cod (Gadus morhua) microsomes to lipid oxidation

During the extraction of muscle to produce protein isolates by acid or alkali solubilization, membranes are exposed to abnormally low or high pH. Low but not high pH treatment induces rapid oxidation of membrane phospholipids in the presence of hemoglobin. The goal of this research work was to study the oxidative stability of microsomes under the conditions met during acid solubilization. Isolated microsomes from cod muscle were used as a model system. At pH 5.3 or lower, 99% of isolated cod membranes sedimented at low centrifugation speeds. Isolated membranes that were exposed to pH 3.0 were less susceptible to hemoglobin-mediated lipid oxidation. Cod hemoglobin exposed to pH 3 was rendered less pro-oxidative than the untreated cod hemoglobin. However, when microsomes and hemoglobin were together exposed to low pH, oxidation was promoted. Citric acid and calcium chloride, as well as press juice isolated from cod muscle, were able to inhibit lipid oxidation of microsomal suspensions.     

Mercury distribution and lipid oxidation in fish muscle: effects of washing and isoelectric protein precipitation

Nearly all the mercury (Hg) in whole muscle from whitefish (Coregonus clupeaformis) and walleye (Sander vitreus) was present as methyl mercury (MeHg). The Hg content in whole muscle from whitefish and walleye was 0.04-0.09 and 0.14-0.81 ppm, respectively. The myofibril fraction contained approximately three-fourths of the Hg in whitefish and walleye whole muscle. The sarcoplasmic protein fraction (e.g., press juice) was the next most abundant source of Hg. Isolated myosin, triacylglycerols, and cellular membranes contained the least Hg. Protein isolates prepared by pH shifting in the presence of citric acid did not decrease Hg levels. Addition of cysteine during washing decreased the Hg content in washed muscle probably through the interaction of the sulfhydryl group in cysteine with MeHg. Primary and secondary lipid oxidation products were lower during 2 °C storage in isolates prepared by pH shifting compared to those of washed or unwashed mince from whole muscle. This was attributed to removing some of the cellular membranes by pH shifting. Washing the mince accelerated lipid peroxide formation but decreased secondary lipid oxidation products compared to that of the unwashed mince. This suggested that there was a lipid hydroperoxide generating system that was active upon dilution of aqueous antioxidants and pro-oxidants.     

The effect of acid and alkali unfolding and subsequent refolding on the pro-oxidative activity of trout hemoglobin

The pro-oxidative activity of trout hemoglobin was significantly increased at low pH (2.5-3.5) in a washed fish muscle (WFM) system. It was found that the more unfolded the hemoglobin was the more exposed its heme group was, which increased its pro-oxidative activity. The amount of oxidation products produced (TBARS) were, however, lower at low pH vs neutral pH. At pH 10.5-11, the pro-oxidative activity of hemoglobin was greatly suppressed. The conformation of hemoglobin was significantly more stable at high pH as compared to pH 7 as judged by its visible absorption spectrum. Hemoglobin readjusted from low pH to pH 7 had a higher pro-oxidative activity (i.e., more rapid oxidation) in WFM than native hemoglobin at pH 7, even though TBARS values were lower than in the untreated sample at pH 7. The results suggest that the WFM becomes slightly more susceptible to oxidation after low pH treatment but also produces less TBARS. The increased pro-oxidative activity after pH readjustment correlated well with an incomplete recovery in the native structure on pH readjustment. A longer unfolding time and a lower pH led to a less refolded hemoglobin with increased pro-oxidative activity. Hemoglobin was less pro-oxidative at low pH in the presence of 500 mM NaCl. The presence of salt did, however, increase the pro-oxidative properties of hemoglobin after readjustment to pH 7. The treatment of washed fish muscle at alkaline pH followed by adjustment to pH 7 led to a slight delay in hemoglobin-mediated lipid oxidation in WFM as compared to native hemoglobin at pH 7. The results suggest that WFM becomes less susceptible toward oxidation after pH readjustment from alkaline pH. These results clearly show that for muscle protein extraction/isolation processes requiring highly alkaline or acidic conditions, alkaline conditions are preferred if the lipid oxidation originating from hemoglobin is to be minimized.     

Effects of oxidized dietary oil and vitamin E supplementation on lipid profile and oxidation of muscle and liver of juvenile atlantic cod (Gadus morhua)

The effects of oxidized dietary lipid and the role of vitamin E on lipid profile, retained tocopherol levels, and lipid oxidation of juvenile Atlantic cod (Gadus morhua) were evaluated following a 9-week feeding trial. Four isonitrogenous experimental diets containing fresh or oxidized (peroxide value of 94 mequiv/kg) fish oil with or without added vitamin E (alpha-tocopherol or mixed tocopherols) were fed to juvenile cod in duplicate tanks. There was no significant (P > 0.05) influence on major lipid classes of cod liver and muscle by diet with the exception of sterols. Sterols content was increased in liver but decreased in muscle by oxidized dietary oil in the absence of vitamin E. Dietary vitamin E supplementation decreased the sterols level in cod liver but with no significant (P > 0.05) effect on their level in the muscle. Fatty acid composition varied between lipid fractions in muscle tissue and was affected by the diet. Oxidized oil significantly (P < 0.05) decreased the deposition of alpha-tocopherol in liver but not in muscle. gamma- and delta-Tocopherols from dietary tocopherol mixtures were retained at very low levels in liver, but higher retention was observed in muscle tissue. The oxidative state of both liver and muscle, as measured by the 2-thiobarbituric acid reactive substances (TBARS) and headspace propanal, negatively correlated with tissue vitamin E levels. It is suggested that oxidized oil affected juvenile Atlantic cod by causing vitamin E deficiency in certain tissues and that these effects could be alleviated by supplementation of a sufficient amount of dietary vitamin E. The results also indicate that mixed tocopherols were good antioxidants for Atlantic cod, although less effective than alpha-tocopherol alone in many tissues with the exception of muscle, where gamma- and delta-tocopherols were deposited at relatively high levels.     

Site e14 in hemoglobins and myoglobins: a key residue that affects hemin loss and lipid oxidation capacity

Fish hemoglobins (Hbs) frequently contain glycine at site E14 while mammalian Hbs contain larger residues (e.g., alanine and serine). These differences were examined by creating structural variants at E14 using recombinant bovine myoglobin (Mb) as a model heme protein that contains alanine at E14. The Ala(E14)Gly mutation increased k(ox) and hemin loss 3-fold and 45-fold, respectively. Glycine at E14 creates a channel for solvent to enter the heme crevice, which enhances autoxidation and hemin loss rates. Hydration of the proximal heme pocket facilitates hemin loss because protonation of the proximal histidine weakens the linkage of the imidazole group to the iron atom of the hemin moiety. Ala(E14)Gly promoted lipid oxidation in washed fish muscle more rapidly during iced storage compared to wild type Mb at pH 5.7. This suggested that the rapid hemin loss from Ala(E14)Gly accelerated lipid oxidation. Ala(E14)Ser and Ala(E14)Val had little effect on k(ox) but somewhat accelerated net hemin loss. These studies suggest that enhanced access of solvent to the heme crevice of many fish Hbs at site E14 facilitates rapid hemin loss and moderately accelerates autoxidation. This likely is part of the reason fish Hbs promote lipid oxidation much more effectively compared to mammalian Hbs.     

Development of yellow pigmentation in squid (Loligo peali) as a result of lipid oxidation

The impact of lipid oxidation on yellow pigment formation in squid lipids and proteins was studied. When the squid microsomes were oxidized with iron and ascorbate, thiobarbituric acid reactive substance were observed to increase simultaneously with b values (yellowness) and pyrrole compounds concomitantly with a decrease in free amines. Oxidized microsomes were not able to change the solubility, sulfhydryl content, or color of salt-soluble squid myofibrillar proteins. Aldehydic lipid oxidation products were able to decrease solubility and sulfhydryl content of salt-soluble squid myofibrillar proteins but had no impact on color. Aldehydic lipid oxidation products increased b values (yellowness) and pyrrole compounds and decreased free amines in both squid phospholipid and egg yolk lecithin liposomes. The ability of aldehydic lipid oxidation products to change the physical and chemical properties of egg yolk lecithin liposomes increased with increasing level of unsaturation and when the carbon number was increased from 6 to 7. These data suggest that off-color formation in squid muscle could be due to nonenzymatic browning reactions occurring between aldehydic lipid oxidation products and the amines on phospholipids headgroups.     

Inhibition of hemoglobin-mediated oxidation of regular and lipid-fortified washed cod mince by a white grape dietary fiber

The effectiveness of a white grape dietary fiber concentrate (WGDF) against hemoglobin-mediated oxidation of washed cod mince, with and without 10% added herring oil, was evaluated during ice storage. WGDF was added at two different levels: 2 and 4% based on final weight. An ethanol extract with the ethanol extractable polyphenols (EPP) and the ethanol-extracted grape dietary fiber residue were also tested as antioxidants in the washed cod mince. The addition of WGDF to the model system completely and significantly (p 相似文献   

Loss of redness (a*) as a tool to follow hemoglobin-mediated lipid oxidation in washed cod mince

Instrumental measurement of redness loss (decrease in a* value) was evaluated as a tool to follow hemoglobin (Hb)-mediated lipid oxidation in fish muscle. Two washed cod mince model systems were used (prepared at pH 6.5 and 5.5), both fortified with 15 micromol/kg of trout Hb and adjusted to pH 6.5 and 81% moisture. The rate of oxidation was varied through pH alterations (pH 6.1 and 6.9) and addition of an antioxidative cod muscle press juice. During ice storage, TBARS, painty odor, and a* values were followed. In all "oxidizing" samples, a* values correlated well with TBARS and painty odor development; r = -0.95 and -0.77, respectively. In press juice containing samples, the correlation was lower (0.55 for a vs TBARS) because there was a slight a* value decrease even in the absence of measurable lipid oxidation. a* values distinguished between "oxidizing" and stable samples within 1 day, before any lipid oxidation products could be chemically detected. It was confirmed in an aqueous phosphate buffer model system that the redness loss corresponded to a buildup of brownish met-Hb at the expense of oxy- and deoxy-Hb. The a* value data were best used as a lipid oxidation index by calculating the rate of decrease (k value) in the "initial phase" of the redness loss (before accumulation of lipid oxidation products) or in the "differentiation phase" (during the exponential raise in TBARS/painty odor). Calibration to lipid oxidation products must, however, be made for each specific sample type. Washing method, pH, Hb-type, etc., all affected both k values and absolute a* readings. Small yellowness (b*) increases also occurred along with a value losses, possibly the result of polymerized Schiff bases.     

Pro-oxidative characteristics of trout hemoglobin and myoglobin: a role for released heme in oxidation of lipids

The molecular mass of trout myoglobin was 16017 Da based on electrospray ionization mass spectrometry. A Root effect (low oxygen affinity at pH 6.3) was determined in trout hemoglobin but not myoglobin. At pH 6.3, myoglobin autoxidized more rapidly (3.5-fold) as compared to anodic hemoglobin. Anodic hemoglobin was a better catalyst of lipid oxidation in washed cod muscle as compared to myoglobin at pH 6.3. This suggested that some process other than met heme protein formation was the rate-limiting step in lipid oxidation processes. Heme loss rates were determined using the apomyoglobin mutant H64Y prepared from sperm whale. Anodic hemoglobin released its heme group much more rapidly than myoglobin. In comparisons of anodic and cathodic hemoglobins, heme loss rate better predicted the onset of lipid oxidation than autoxidation rate. These studies collectively suggest that heme dissociation has a primary role in the ability of different heme proteins to promote lipid oxidation processes.     

Galloylated polyphenols as inhibitors of hemoglobin-catalyzed lipid oxidation in fish muscle

The influence of galloyl residues on the antioxidant mechanism of polyphenols to prevent hemoglobin-promoted lipid oxidation was investigated by using polyphenolic fractions with different degrees of galloylation: nongalloylated structures from pine bark (IVP), medium-galloylated from grape pomace (IVG), and high-galloylated from witch hazel bark (IVH). Hemoglobin (Hb) from the pelagic fish horse mackerel (Trachurus trachurus) was employed as a Hb standard. In vitro experiments showed an important increase in the deoxygenation and autoxidation of horse mackerel Hb at acidic pH values. All polyphenolic fractions significantly reduced the redox stability of Hb in buffer solutions, showing a greater deoxygenation and methemoglobin (metHb) formation in the presence of IVH, followed in decreasing order by IVG and IVP. However, galloylated polyphenols (IVH and IVG) were efficient to inhibit the oxidation of the oxygenated Hb (OxyHb) and the formation of lipid oxidation products in chilled washed fish muscle. This antioxidant activity of galloylated proanthocyanidins showed a positive relationship with the phenolic concentration. Polyphenols devoid of galloyl groups (IVP) were less active to prevent either Hb oxidation or lipid oxidation in fish muscle. The results draw attention to the potential role of galloyl residues to lessen Hb-catalyzed lipid oxidation in muscle and to maintain Hb in reduced and oxygenated states, which exhibit lower pro-oxidant activity as compared to the metHb and deoxyHb species.