M. J. Alvarez*, C. J. López-Bote, A. Diez*, G. Corraze, J. Arzel, J. Dias, S. J. Kaushik, and J. M. Bautista*,3

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1 The Partial Substitution of Digestible Protein with Gelatinized Starch as an Energy Source Reduces Susceptibility to Lipid Oxidation in Rainbow Trout (Oncorhynchus mykiss) and Sea Bass (Dicentrarchus labrax) Muscle 1,2 M. J. Alvarez*, C. J. López-Bote, A. Diez*, G. Corraze, J. Arzel, J. Dias, S. J. Kaushik, and J. M. Bautista*,3 *Departamento de Bioquímica y Biología Molecular IV and Departamento de Producción Animal, Universidad Complutense de Madrid, Facultad de Veterinaria, Madrid, Spain; Fish Nutrition Laboratory, Unité Mixte INRA-IFREMER, Saint Pèe-sur-Nivelle, France; and Fish Nutrition Laboratory, Unité Mixte INRA-IFREMER, Centre de Brest, IFREMER, Plouzané, France ABSTRACT: We evaluated the influence of dietary gelatinized starch and protein on the fatty acid composition of muscle in rainbow trout and European sea bass and on the susceptibility of flesh to lipid peroxidation. The possibility that flesh peroxidation could be accounted for by lipogenesis and the deposition of fat was also explored. The inclusion of gelatinized starch in the diet of rainbow trout improved growth with respect to that observed in fish fed crude starch (P <.001). This was especially noticeable at the lowest concentration of dietary protein tested (P =.037), suggesting that gelatinized starch may partially replace protein in the production of energy without inducing a negative effect on growth. However, in European sea bass, the gelatinization of starch and dietary protein concentration showed no significant effect on final body weight. The intramuscular neutral lipid concentration of the sea bass was reduced by the gelatinization of dietary starch (P =.034). The highest dietary protein concentration increased the proportion of saturated fatty acids in the neutral (P =.0742) and polar (P =.0033) lipid fractions. The dietary inclusion of high levels of protein in rainbow trout led to a lower concentration of total (n-3) (P =.0457) and (n-6) (P =.0522) fatty acids and a higher concentration of total monounsaturated fatty acids (P =.0006). The inclusion of gelatinized starch led to a lower concentration of (n-3) fatty acids (P =.0034) and a higher concentration of saturated fatty acids (P =.0007). The polar fraction was hardly affected by the same treatment. A significantly lower susceptibility of the dorsal muscle to oxidation was observed in groups of European sea bass fed gelatinized starch (P <.01). A similar trend was observed in rainbow trout, although differences were not significant. The findings suggest that the digestible protein concentration of nutrientdense diets for rainbow trout and European sea bass can be reduced with a beneficial effect on tissue lipid oxidation and no negative effects on growth and muscle composition. Key Words: Rainbow Trout, Sea Bass, Dietary Fat, Protein, Starch, Oxidation 1999 American Society of Animal Science. All rights reserved. J. Anim. Sci : Introduction The high level of digestible protein (DP) in fish diets is associated with metabolic losses and nitrogen loading of the environment (Cho and Kaushik, 1990). Further, a high DP level is associated with a higher susceptibility of fish muscle to lipid oxidation (Alvarez et al., 1998). 1 This research was financed by grants from the European Commission (AIR-CT : Fat deposition in farmed rainbow trout and European sea bass; and AIR-CT : Dietary treatment and oxidative stability of muscle and meat products) and by a Spanish CICYT grant (AGF ). Received November 12, Accepted June 7, Optimal digestible protein:digestible energy (DP:DE) ratios for rainbow trout can be reduced if a complementary energy source is supplied to allow protein sparing (Kaushik and Médale, 1994). Increased DE is generally achieved by raising dietary oils, which are nutritionally essential to teleosts (Yu and Sinnhuber, 1979). However, high-fat diets for fish lead to a greater degree of fat deposition and enhanced susceptibility to lipid peroxidation than do high-dp diets (Alvarez et al., 1998). Carbohydrates are another 2 Thanks are due to S. Pérez-Benavente and L. Larroquet for their efficient technical assistance. 3 To whom correspondence should be addressed (phone: ; fax: ; bauchem@eucmax. sim.ucm.es). 3322

2 GELATINIZED STARCH AND OXIDATION IN FISH 3323 possible source of energy but are of limited use in some fish species due to the poor digestibility of crude starch (Singh and Nose, 1967), although cooked starch is of a higher digestibility (Smith, 1980). This has led to an increased interest in the use of higher levels of carbohydrates in fish diets to partially substitute fat or protein for energy purposes. However, modifications to a feeding system should be evaluated in terms of flesh fatty acid composition and susceptibility to oxidation. In mammals, the source of dietary energy other than fat is of importance in the susceptibility of muscle to lipid oxidation (Lopez-Bote et al., 1997). The postharvest quality of farmed fish in terms of flesh lipid oxidation related to dietary carbohydrate is yet to be established. The main objectives for this study were 1) to evaluate the effects of dietary gelatinized starch and protein on the fatty acid composition of fish muscle, 2) to relate these findings to the susceptibility of tissue to lipid peroxidation, and 3) to estimate whether metabolic activity related to lipogenesis and deposition of fat could account for different susceptibilities to flesh peroxidation. Materials and Methods Rainbow trout (Oncorhynchus mykiss) and European sea bass (Dicentrarchus labrax) were fed one of four dry, pelleted diets formulated to contain a low (LP) or high (HP) CP and a gelatinized or crude native starch source (Tables 1 and 2). The experimental diets used for sea bass contained slightly more protein than the NRC (1993) recommendations for this species. Rainbow trout growth trials were conducted at the INRA experimental fish farm in Donzacq (Landes, France) at a constant water temperature of 17 ± 1 C for a period of 12 wk. Sea bass growth trials were performed at the seawater (salinity: 35 ppt) facilities of IFREMER, Centre de Brest (France), where the water temperature was maintained at 20 C. Juvenile rainbow trout (mean body weight: 73 g) were distributed into groups of 100 fish in plastic tanks (volume: 4 m 3 ). Triplicate groups were hand-fed twice daily to apparent satiety. European sea bass (mean body weight: 130 g) were assigned to groups of 45 fish in plastic tanks ( m) for a 12-wk period. Triplicate groups were also hand-fed twice daily to apparent satiety. In both trials, fish were group-weighed and counted every 3 wk to follow growth and feed intake. To calculate the DE and DP of feeds, apparent digestibility was estimated by means of an indigestible tracer (10 g/kg Cr 2 O 3 ). To this end, triplicate groups of 10 fish of each species were adapted over more than a fortnight to the diets. Feces were collected (Choubert et al., 1982) daily for 6 d and were frozen and freeze-dried prior to analysis. After the final end-of-trial weighing, 10 fish from each experimental diet group were stunned to obtain the dorsal muscles, which were frozen and stored in liquid nitrogen. Liver samples were obtained from a further six fish from each group and immediately processed for the determination of enzymatic activity within 36 h. The proximate compositions of the dietary ingredients, diets, and feces were estimated using routine laboratory procedures (Lopez-Bote et al., 1997). Lipid oxidation and fatty acid composition were determined within 1 wk and 1 mo after slaughter, respectively. Lipids from muscle samples were obtained according to the method developed by Marmer and Maxwell (1981) and were subsequently methylated using 2 M alcoholic methanol-koh in a.7 M HCl methanol solution as described elsewhere (Lopez-Bote et al., 1997). Fatty acids were separated using a 5890 Hewlett Packard gas chromatograph equipped with a split injector and flame ionization detector (FID) (50:1 split ratio) and a 30-m.32-mm.25-µm crosslinked polyethylene glycol capillary column (Hewlett Packard, Avondale, PA). Samples were analyzed under the following operating conditions: injector/detector temperature 250 C, helium gas flow 3 ml/min, and temperature gradient 170 to 245 C. Individual fatty acids were identified by comparison with known standard mixtures. Tricosanoic acid (Sigma Chemical Co., St Louis, MO) was used as the internal standard. The susceptibility of muscle homogenates to iron-induced lipid oxidation was determined by a modification of the method of Kornbrust and Mavis (1980). Homogenates (approximately 1 mg/ml buffer) were incubated at 37 C in 80 mm Tris-maleate buffer (ph 7.4) in the presence of.2 M ascorbic acid in a total volume of 10 ml. Aliquots were obtained at fixed time intervals for measurement of thiobarbituric acid-reactive substances (TBARS), which were expressed as nanomoles of malonaldehyde (MDA) per milligram of protein. The protein content of individual samples was determined using the method of Bradford (1976). For the enzyme assays, liver samples were homogenized in three volumes of ice-cold buffer (.02 M Tris- HCl,.25 M sucrose, 2 mm EDTA,.1 M sodium fluoride,.5 mm phenylmethylsulfonylfluoride,.01 M β-mercaptoethanol, ph 7.4), and homogenates were centrifuged at 48,000 g at 4 C for 40 min. The soluble protein content of liver homogenates was determined with the method of Bradford (1976) using BSA as the standard. Selected lipogenic enzyme activities of the supernatant were quantified with spectrophotometric procedures: glucose-6-phosphate dehydrogenase (G6PD, EC ) according to Bautista et al. (1988) and malic enzyme (MEz, EC ) according to Ochoa (1955). The enzymatic activity units (IU), defined as micromoles of substrate converted to product per minute at the assay temperature (30 C) were expressed per milligram of hepatic soluble protein (specific activity) or per gram of liver tissue (wet weight). Statistical Analysis An analysis of variance for a completely random design was used to compare differences between treatments. In addition, a repeated measures test was used

3 3324 ALVAREZ ET AL. Table 1. Main ingredients, chemical composition, gross energy, digestible energy, and protein of rainbow trout diets containing high (H) or low (L) digestible protein (P) levels and gelatinized (G) or crude (NG) cornstarch LP HP Item G NG G NG Ingredient, g/kg Fish meal (herring, CP > 70%) Fish protein soluble concentrate Fish oil Gelatinized cornstarch Crude cornstarch Corn gluten Vitamin premix a Mineral premix b Binder (Na alginate) Chemical composition Dry matter, g/kg Protein, g/kg DM Crude fat, g/kg DM Ash, g/kg DM Gross energy, kj/g DM Digestible energy, kj/g DM Digestible protein, g/kg DM a Per kilogram of diet: 60 mg DL-α-tocopheryl acetate, 5 mg Na menadione bisulfate, 15,000 IU retinyl acetate, 3,000 IU DL-cholecalciferol, 15 mg thiamine, 30 mg riboflavin, 15 mg pyridoxin,.05 mg B 12, 175 mg nicotinic acid, 5 mg folic acid, 500 mg ascorbic acid, 1,000 mg inositol, 2.5 mg biotin, 50 mg calcium pantothenate, and 2,000 mg choline chloride. b Per kilogram of diet: 2.15 g calcium carbonate (40% Ca), 1.24 g magnesium oxide (60% Mg),.2 g ferric citrate,.4 mg potassium iodide (75% I),.4 g zinc sulfate (36% Zn),.3 g copper sulfate (25% Cu),.3 g manganese sulfate (33% Mn), 5 g dibasic calcium phosphate (20% Ca, 18% P), 2 mg cobalt sulfate, 3 mg sodium selenite (30% Se),.9 g KCl, and.4 g NaCl. to compare the oxidation rates of groups. The experimental units for analysis of data were individual fish except for growth and feed intake, which were determined by group-weighing. Data were analyzed using SAS procedures (1988) and are presented as group means and pooled standard deviations with the corresponding significance levels of the main effects and interactions. Results and Discussion The growth performance and dorsal muscle composition of rainbow trout and European sea bass fed the experimental diets are shown in Tables 3 and 4. Gelatinized starch showed a significant effect on final body weight in rainbow trout (P <.001). Moreover, the effect of gelatinization on final body weight was more pronounced when the concentration of dietary protein was lower (P =.037). This interaction indicates that gelatinized starch may partially replace protein as the energy source without having a negative effect on growth. Hillestad and Johnson (1994) reported that optimal growth of Atlantic salmon may be obtained with only 39% dietary CP. In contrast, neither gelatinization nor protein concentration in the diet produced an effect on final body weight in the European sea bass (Table 4). This lack of response to dietary protein concentration over the range presently tested may be of economic and environmental interest. The intramuscular neutral lipid concentration in the European sea bass was reduced with the gelatinization of the starch (P =.034) and by feeding high dietary crude protein levels (P <.001) (Table 4), but no such effect was observed in rainbow trout (Table 3), although the trend was similar to that observed in the sea bass. Greater dietary protein levels, however, did lead to a higher concentration of intramuscular neutral lipids in the dorsal muscle (P =.0139) (Table 3). The fatty acid types of neutral and polar lipids from the dorsal muscle of rainbow trout and European sea bass are shown, respectively, in Tables 5 and 6. In rainbow trout, high dietary protein levels led to a reduced concentration of total (n-3) (P =.0457) and (n-6) (P =.0522) fatty acids and to an increased concentration of total monounsaturated fatty acids (P =.0006) in neutral lipid. Lower concentrations of (n-3) fatty acids (P =.0034) and higher concentrations of saturated fatty acids (P =.0007) were also observed in the neutral lipid fraction of the muscle of fish fed diets containing gelatinized starch. The concentration of fatty acids in the polar fraction of intramuscular lipids was scarcely affected by the dietary treatment (Table 5). The phospholipid fraction is considered to be particularly stable due to its role in supporting cell membrane structures (Farkas and Csengeri, 1976). A similar trend also was observed in the types of fatty acids found in the European sea bass neutral lipids (Table 6), although gelatinized starch only significantly

4 GELATINIZED STARCH AND OXIDATION IN FISH 3325 Table 2. Main ingredients, chemical composition, gross energy, digestible energy, and protein of European sea bass diets containing high (H) or low (L) digestible protein (P) levels and gelatinized (G) or crude (NG) cornstarch LP HP Item G NG G NG Ingredient, g/kg Fish meal (herring, CP > 70%) Fish protein soluble concentrate Fish oil Gelatinized cornstarch Crude cornstarch Corn gluten Vitamin premix a Mineral premix b Binder (guaranate) Chemical composition Dry matter, g/kg Protein, g/kg DM Crude fat, g/kg DM Ash, g/kg DM Gross energy, kj/g DM Digestible energy, kj/g DM Digestible protein, g/kg DM a Per kilogram of diet: 60 mg DL-α-tocopheryl acetate, 5 mg Na menadione bisulfate, 15,000 IU retinyl acetate, 3,000 IU DL-cholecalciferol, 15 mg thiamine, 30 mg riboflavin, 15 mg pyridoxin,.05 mg B 12, 175 mg nicotinic acid, 5 mg folic acid, 500 mg ascorbic acid, 1,000 mg inositol, 2.5 mg biotin, 50 mg calcium pantothenate, and 2,000 mg choline chloride. b Per kilogram of diet: 2.15 g calcium carbonate (40% Ca), 1.24 g magnesium oxide (60% Mg),.2 g ferric citrate,.4 mg potassium iodide (75% I),.4 g zinc sulfate (36% Zn),.3 g copper sulfate (25% Cu),.3 g manganese sulfate (33% Mn), 5 g dibasic calcium phosphate (20% Ca, 18% P), 2 mg cobalt sulfate, 3 mg sodium selenite (30% Se),.9 g KCl, and.4 g NaCl. Table 3. Average daily intake and gain, final weight, protein retention, and intramuscular lipid content of rainbow trout fed diets containing a high (HP) or low (LP) level of digestible protein and a gelatinized (G) or crude (NG) source of cornstarch LP HP Probability of contrasts d Item G NG G NG SD Protein Gelatinization Interaction Intake ab Digestible protein, g kg ABW 1 d Digestible fat, g kg ABW 1 d Digestible carbohydrate, g kg ABW 1 d Digestible energy, kj kg ABW 1 d Dorsal muscle composition, g/kg c Neutral lipids Polar lipids Growth b Final body weight, g N, mg Fat, g Carcass composition, % of fresh c Moisture Protein Fat Ash a ABW = average body weight. b n = 3. Triplicate groups of 10 fish per group were used. Group was considered the experimental unit. c n = 10 fish per dietary group. Fish was the experimental unit. d Not significant (P >.2).

5 3326 ALVAREZ ET AL. Table 4. Average daily intake and gain, final weight, protein retention, and intramuscular lipid content of European sea bass fed diets containing a high (HP) or low (LP) level of digestible protein (P) and a gelatinized (G) or crude (NG) source of cornstarch LP HP Probability of contrasts d Item G NG G NG SD Protein Gelatinization Interaction Intake ab Digestible protein, g kg ABW 1 d Digestible fat, g kg ABW 1 d Digestible carbohydrate, g kg ABW 1 d Digestible energy, kj kg ABW 1 d Dorsal muscle composition, g/kg c Neutral lipids Polar lipids Growth b Final body weight, g N, mg Fat, g Carcass composition, % fresh c Moisture Protein Fat Ash a ABW = average body weight. b n = 3. Triplicate groups of 10 fish per group were used. Group was considered the experimental unit. c n = 10 fish per dietary group. Fish was the experimental unit. d Not significant (P >.2). affected the concentration of monunsaturated fatty acids in the neutral lipid fraction (P =.0107). High dietary protein concentration led to a higher proportion of saturated fatty acids in the neutral (P =.0742) and polar (P =.0033) lipid fractions. Saturated and monounsaturated fatty acids are the final end products of lipid synthesis in animals (Enser, 1984). The present findings indicate that rainbow trout and European sea bass fed gelatinized starch or high-protein diets have an intake of energy above their physiological needs and, because part of this energy is used for fat synthesis, this leads to a greater accumulation of nonessential fatty acids. In order to identify the possible modifications to the fatty acid synthesis pathway as a consequence of the increased amount of protein in the diet, the activities of the two main NADPH-generating enzymes in the cytoplasm, G6PD and MEz were assessed (Table 7). In both species, the activity of G6PD was 10 to 20 times that of MEz, in agreement with previous reports (Alvarez et al., 1998; Dias et al., 1998), which suggests that cytoplasmic reduction equivalents (NADPH) are Table 5. Fatty acid composition (g/kg of total fatty acids) of neutral and polar fractions of intramuscular lipids of rainbow trout fed diets containing a high (HP) or low (LP) level of digestible protein and a gelatinized (G) or crude (NG) source of cornstarch (n = 10) LP HP Probability of contrasts a Lipids G NG G NG SD Protein Gelatinization Interaction Neutral lipids (n-3) (n-6) (n-3)/ (n-6) Saturated Monounsaturated Polar lipids (n-3) (n-6) (n-3)/ (n-6) Saturated Monounsaturated a Not significant (P >.2).

6 GELATINIZED STARCH AND OXIDATION IN FISH 3327 Table 6. Fatty acid composition (g/kg total fatty acids) of neutral and polar fractions of intramuscular lipids of European sea bass fed diets containing a high (HP) or low (LP) level of digestible protein and a gelatinized (G) or crude (NG) source of cornstarch (n = 10) LP HP Probability of contrasts a Lipids G NG G NG SD Protein Gelatinization Interaction Neutral lipids (n-3) (n-6) (n-3)/ (n-6) Saturated Monounsaturated Polar lipids (n-3) (n-6) (n-3)/ (n-6) Saturated Monounsaturated a Not significant (P >.2). mainly provided by the pentose phosphate pathway. Thus, the changes observed between diets in terms of net NADPH reflect differences in G6PD activity rather than that of MEz. The enzyme G6PD is a regulatory, branching protein that produces reducing equivalents for fatty acid biosynthesis and recycling of pentose sugars in energy metabolism. As seen in Table 7, G6PD activity was significantly higher in groups of fish fed gelatinized starch than in those fed native starch in both rainbow trout and European sea bass (P <.0001 and P =.0136, respectively). This indicates that the carbohydrate, and thus carbon units, available to the fish should stimulate the consumption of NADPH, which generates NADP and in turn enhances the pentose phosphate pathway, in agreement with previous reports (Hilton and Atkinson, 1982). It would seem that the availability of carbohydrate in the diet regulates the generation of reducing equivalents through G6PD. With regard to the effect of protein in the diet, a net decrease in the production of NADPH was observed at high dietary protein concentrations regardless of the amount of gelatinized starch in the diet in the European sea bass. In rainbow trout, there was a dietary protein gelatinized starch interaction, and the dietary protein level was seen to affect G6PD activity in the groups that were fed gelatinized starch only. Such data indicate that part of the available carbohydrate and protein is utilized for lipogenesis. Hence, increased amounts of protein and available carbohydrate favor lipogenesis and consequently lead to an increased proportion of saturated fat compared to the higher deposition of polyunsaturated fatty acids from fat observed in lower-protein diets. The effects of the experimental diets on the susceptibility of rainbow trout and European sea bass dorsal muscle to oxidation, assessed as induced peroxidation, was subsequently investigated (Figures 1 and 2, respectively). Significantly lower oxidation rates were recorded in the groups of European sea bass fed gelatinized starch (P <.01). A similar trend was observed in rainbow trout, although the differences were not significant. This species variability was expected and may be explained by the different fatty acid concentrations in each species. Saturated and monounsaturated fatty Table 7. Glucose-6-phosphate dehydrogenase (G6PD) and malic enzyme (MEz) activities in the liver of rainbow trout and European sea bas fed diets containing a high (HP) or low (LP) level of digestible protein and a gelatinized (G) or crude (NG) source of cornstarch (values expressed as IU/g liver) (n = 6) LP HP Probability of contrasts a Species G NG G NG SD Protein Gelatinization Interaction Rainbow trout G6PD MEz European sea bass G6PD MEz a Not significant (P >.2).

7 3328 ALVAREZ ET AL. Figure 1. Iron-induced lipid peroxidation (mean ± SE; nanomoles of malonaldehyde/milligram of protein) in dorsal muscle homogenates incubated at 37 C for up to 240 min in rainbow trout fed diets containing a high (HP) or low (LP) level of digestible protein and a gelatinized (G) or crude (NG) source of cornstarch (n = 10). acids are less susceptible to lipid oxidation than highly unsaturated fatty acids, particularly (n-3) fatty acids (Austreng and Krogdahl, 1987; Lopez-Bote et al., 1997). Hammer and Wills (1978) suggested enhanced susceptibility to lipid peroxidation of (n-3) fatty acids either as pure lipid or in tissues of rats fed fish oil compared to rats fed corn oil. Previous investigations indicate that the susceptibility to oxidation of (n-3) fatty acids seems to particularly involve highly unsaturated fatty acid residues. This is also in agreement with the results of L Abbe et al. (1991), who found a relationship between tissue incorporation of long-chain (n-3) fatty acids and urinary TBARS. A significantly higher TBARS concentration was also found in the muscle homogenates of rainbow trout fed the higher DP diet (P <.05) (Figure 1). The prooxidant effect of high DP was somewhat unexpected and needs further investigation. According to the composition of the fatty acids (Tables 5 and 6), high DP would be expected not to affect, or even reduce, lipid oxidation. It is suggested that additional factors may be involved in this greater tendency toward oxidation. Indeed, the rate of lipid oxidation in muscle systems depends on a number of factors other than the unsaturated nature of the fatty acids, including the presence of free fatty acids, the presence of antioxidants (Frigg et al., 1990), and the concentration of prooxidants such as iron and enzyme mediators (Tichivangana and Morrissey, 1985). The significant effect of the different diets on liver enzyme activity may partly explain the difference in the susceptibility to oxidation shown by the diet groups. The G6PD is the main enzyme that generates reducing power in the form of NADPH, which has been related to intracellular antioxidant capacity through a cascade of other enzyme activities (Bautista et al., 1989; Barroso et al., 1994). Data corresponding to G6PD activity (i.e., the production of reducing equivalents) may be correlated with the susceptibility to peroxidation; reduced G6PD activity is detected in animals with a higher susceptibility to oxidation. Because no correlation between total fatty acids and membrane lipids that could account for the higher susceptibility to oxidation in animals fed protein-enriched diets was observed, the data for G6PD activity could indicate a possible involvement of pro-antioxidant environments linked to enzyme activity and the availability of metabolic NADPH. Another possible explanation might involve the differences in the total amount of intramuscular fat observed in the different diet groups (Table 3). Several reports indicate that, as long as the degree of unsaturation falls within a reasonable range, the amount of intramuscular fat has a great influence on susceptibility to lipid oxidation (Shewfelt, 1981; Ingemansson et al., 1995). However, the effect of dietary protein on the susceptibility to muscle lipid oxidation has not been previously described and requires further investigation. The findings reported here indicate that protein can be partially replaced with gelatinized starch in rainbow trout and European sea bass diets with no negative effect on growth. The inclusion of digestible carbohydrate in the diet is known to improve nitrogen utilization and reduce nitrogen losses (Médale et al., 1995). A reduced loss of nitrogen into the environment and a greater stability of fish flesh to oxidative deterioration can thus be achieved, particularly in rainbow trout, Figure 2. Iron-induced lipid peroxidation (mean ± SE; nanomoles of malonaldehyde/milligram of protein) in dorsal muscle homogenates incubated at 37 C for up to 360 min in European sea bass fed diets containing a high (HP) or low (LP) level of digestible protein and a gelatinized (G) or crude (NG) source of cornstarch (n = 10).

8 GELATINIZED STARCH AND OXIDATION IN FISH 3329 when digestible carbohydrates are supplied at the expense of proteins in the diet. Implications The inclusion of gelatinized starch and high levels of protein in the diets of rainbow trout and sea bass seems to reduce the susceptibility of fish flesh to oxidative deterioration. This may be partly explained by differences in intramuscular fat content, fatty acid composition, and the activity of certain metabolic enzymes. Reduction in the digestible protein concentration of nutrient-dense diets may induce beneficial effects on tissue lipid oxidation and reduce environmental nitrogen loading without showing appreciable negative effects on growth rate and muscle composition. Literature Cited Alvarez, M. J., C. J. Lopez-Bote, A. Díez, G. Corraze, J. Arzel, J. Dias, S. J. Kaushik, and J. M. 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