DAVID V. M. ASHLEY AND G. HARVEY ANDERSON Department of Nutrition and Food Science, FitzGerald Building, University of Toronto, Toronto, Ontario, Canada, M5S 1A1 ABSTRACT The relationship between changes in blood plasma amino acids and the quantity of protein and energy self-selected by the weanling rat, simultaneously offered two diets varying only in protein concentration, was examined. The changes in the quantity of protein and energy consumed by the rats, which were brought about by the addition of the essential limiting amino acids and groups of essential amino acids to gluten, casein, and zein, were not linearly related to alterations in the total plasma amino acid concentrations or to the accumulation of the added amino acids in the blood plasma. However a consistent relationship between food intake and plasma acids was identified when the plasma tryptophan to neutral amino acid ratio was correlated with the protein intake selected. It is postulated that the ratio of tryptophan to neutral amino acids in the plasma is involved in, or reflects, a mechanism regulating protein intake. J. Nutr. 105: 1412-1421, 1975. INDEXING KEY WORDS acids •protein •energy
plasma amino acids •tryptophan •self-selection •weanling rats
Since the early observations of a relation ship between amino acid excess, decreased food intake, and the accumulation of amino acids in the blood of chicks ( 1 ), a number of studies have described correlations be tween the food intake depression that oc curs when high protein or amino acidimbalanced diets are fed, and altered blood amino acid patterns (2-4). Areas separate from the hypothalamus have been shown to mediate the food intake response caused by amino acid-imbalanced or high protein diets (5-9). Although the mecha nisms by which rats alter their food intake in response to these diets have not been identified, it has recently been shown, using a feeding system allowing weanling rats to select both protein and energy, that a mechanism operates to control protein intake (10, 11). Because protein is usually fed as a fixed proportion of the diet, the importance of this mechanism, which ap pears to interact with the mechanism con trolling energy intake, in the regulation of food intake has been disregarded. A precise relationship between plasma amino acid patterns and food intake has 1412
•neutral amino
been difficult to establish because many changes in plasma patterns lead to altera tions in food intake. However a specific as sociation between a neurotransmitter, serotonin, and food intake has been stated recently (12-15). The concentration of serotonin in the brain has been shown to be related to the brain concentration of tryptophan, from which it is synthesized ( 13, 14 ). Brain tryptophan concentration is dependent on the relative concentrations of plasma tryptophan to the five neutral amino acids with which it shares a com mon transport mechanism to the brain (i.e., tyrosine, phenylalanine, leucine, isoleucine, and valine) (16). Alterations in plasma amino acid concentrations brought about by alterations in the amino acid composition of the dietary protein have been shown to influence the synthesis of serotonin (14). The present experiments describe a relationship between protein intake and the ratio of plasma tryptophan Received for publication March 6, 1975. 1 Supported by grant MA-4392 from the Medical Research Council of Canada. * Address reprint requests to G. II. Anderson.
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Correlation between the Plasma Tryptophan to Neutral Amino Acid Ratio and Protein Intake in the Self-selecting Weanling Rat1'2
RAT PLASMA AMINO ACIDS AND PROTEIN SELECTION
1413
to the sum of the plasma concentration of the neutral amino acids in the self-select ing weanling rat.
Downloaded from https://academic.oup.com/jn/article-abstract/105/11/1412/4768666 by East Carolina University user on 13 January 2019
TCA. Plasma tryptophan was determined fluorometrically on each sample by the method of Denckla and Dewey (17). One milliliter of the remaining plasma was deMETHODS proteinized with 3 ml of 10% sulfosalicylic Male weanling rats of the Wistar strain 3 acid, and 0.5 ml of the clear supernatant and of weight range 40-50 g were housed of the six samples in each group was pooled. A minimum of 0.5 ml of the pooled and maintained for 4 weeks under condi tions previously described (11). During sample was analyzed for plasma amino this time they were allowed to freely acids on an automatic amino acid ana balance their dietary protein and energy lyzer.4 Statistical analyses of the plasma tryptophan intakes from two food cups that were of using Tukey'sconcentrations multiple t testwere and performed Dunnett's fered simultaneously and that differed from procedures (18). each other only in protein concentration. Details of the experimental design have RESULTS been documented in the preceding paper (11), but the design is briefly restated Experiment 1. The relative changes in here. plasma amino acid concentrations of the Three experiments were conducted. In experimental groups are illustrated in fig experiment 1, 9 groups of 10 weanling rats ure 1 according to the format used by were offered a choice of 5 and 45%, 15 Peng et al. (19). Each section of the and 55%, or 30 and 60% gluten diets to figure represents a different dietary pro which either 0, 3, or 6 g of lysine had been tein combination. For each combination added to each 100 g of gluten. fed, the effect of increasing the level of In experiment 2, 8 groups of 10 wean lysine in gluten by 3 or 6% on plasma ling rats were offered a choice of 15 and amino acids is shown expressed as a per 55% protein diets formulated with gluten centage of the values obtained when the or with casein to which none, lysine, me- gluten containing no added lysine was thionine, or the four most limiting amino fed (table 1). acids were added to the protein. The increase in lysine level in gluten In experiment 3, 12 groups of 10 wean increased plasma lysine concentration re ling rats were fed 15 and 55% zein diets. gardless of the dietary combination (fig. Six groups were fed the zein diets to which 1). Arginine concentrations were in three levels of lysine (1.7, 3.4, or 6.8 g/100 creased with the increased dietary lysine g) were added to zein made adequate in concentration in gluten when the dietary tryptoohan (zein plus tryptophan) or zein, choices were 5 and 45% or 15 and 55%, partially corrected with other amino acids but not when the choice was 30 and 60% in addition to tryptophan (partially cor gluten. The addition of lysine to gluten rected zein plus tryptophan). The six re was consistently associated with a decrease maining groups were offered 15 and 55% in plasma essential amino acids, threonine, valine, leucine, isoleucine, and phenylzein diets to which three levels of trypto phan (0.3, 0.6, or 1.2 g/100 g) were added alanine. Of the dispensable amino acids, to zein made adequate in lysine (zein plus the most consistent changes were an in lysine) or to partially corrected zein that crease in glutamic acid and a decrease in was adequate in lysine (partially corrected glutamine. zein plus lysine). Plasma tryptophan concentrations (table At the end of each 4-week study, blood 2) were unaffected by the 3 or 6% addi samples were collected between 9 and tion of lysine except when 3% lysine was 11 AM from the abdominal vena cava of added and offered in the 15 and 55% six animals in each group. The animals dietary combination. On the other hand, were first anesthetized with ether. The the sum of the plasma concentrations of blood samples were stored in chilled valine, leucine, isoleucine, phenylalanine, heparinized tubes, centrifuged, and 1 ml 3 Woodlyn Farms, Guelph. Ontario. Cannila. of the plasma was aliquoted and deoro4 Beckman Spinco automatic analyzer, model 11G/ teinized with an equal volume of 20% 119, Beckman Instruments Inc., Palo Alto, Calif.
1414
DAVID V. M. ASHLEY AND G. HARVEY ANDERSON
un im IMO «D
no
UDO
in m 200 UO MO l« 120 HO
120 SUO
ino no
5n »to rit
i
B 20 230 200 I» l«
o«»
Sa §300
a «
£ MO MO 120
Fig. l Effects of lysine additions to gluten on relative plasma amino acid concentrations in weanling rats self-selecting from a, 5 and 45%; b, 15 and 55%; or c, 30 and 60% gluten diets (• •, 3% lysine added; •—-•, 6% lysine added). Single analysis of a pooled sample from six rats in each group.
and tyrosine (table 2) decreased when the lysine additions were made, and the ratio of tryptophan to neutral amino acids (Trp/2 NAA) increased, with the one ex
n ai a 40
a
Fig. 2 Effects of addition of lysine, methionine, and the first four limiting amino acids to a, gluten and b, casein on relative plasma amino acid concentrations in weanling self-selecting rats (• •, 6% lysine added; •——•, four limiting amino acids added; • •, 2.5% methionine added). Single analysis of a pooled sample from six rats in each group.
Downloaded from https://academic.oup.com/jn/article-abstract/105/11/1412/4768666 by East Carolina University user on 13 January 2019
ino isoo
ception, that of the 3% lysine, 15 and 55% group. Experiment 2. The plasma amino acid profiles of the groups self-selecting from the gluten and casein diets are shown in figure 2a and b, respectively. The indi vidual concentrations of plasma amino acids of the control groups are shown in table 3. Increasing the lysine level of gluten by 6% caused an increase in the plasma lysine concentration and a decrease in plasma ornithine but had little effect on the con centrations of the other plasma amino acids (fig. 2a). With the exception of the added amino acids, lysine, threonine, and
1410
RAT PLASMA AMINO ACIDS AND PROTEIN SELECTION TABLE 1
for the groups fed casein to which no amino acid additions were made and those self-selecting from gluten diets fed casein to which lysine or the four most with no added lysine1 limiting amino acids were added, except for the plasma concentrations of lysine, Dietary choice leucine, and methionine. Plasma lysine con Amino acid 5 + 45 15 + 55 30 + 60 centrations were elevated when lysine alone was added to casein but were un mlAspart limóles/too affected by the addition of the four most acidThreonineSerineAsparagineGlu ic limiting amino acids to casein. Leucine and methionine concentrations were in creased in the plasma of all treatment neGlutamic tam i groups. When methionine additions were acidGlycineAlanineValineMethionineIsoleucineLeucineTv made to casein, the plasma concentrations of all amino acids with the exception of leucine and methionine were lower than the plasma amino acid concentrations of the animals fed the casein diet containing rosine no added amino acids. PhenylalanineOrnithineLysineHistidineArginine3.430.033.39.075.914.824.854.833.35.815.928.213.2 14.46.54.98.69.32.329.435.28.373.715.720.561.143.96.820.037.113.5 10.55.05.011.013.4 11.77.44.39.16.93.430.039.35.461.221.822.059.946.17.919.238.712.4 There were no significant differences in plasma tryptophan concentrations of the animals fed gluten or casein diets (table 4). However, the animals fed casein diets had higher plasma tryptophan concentra 1Single analysis of a pooled sample from six rats. tions than those fed gluten diets. The sum of the five neutral amino acids (table 4) methionine, plasma amino acid concen was lowered by all amino acid additions to trations were lower in the plasma of rats gluten, and as a result the Trp/2 NAA fed gluten to which the four limiting amino ratio was increased. The most marked ef acids, rather than lysine alone or no amino fects on the neutral amino acids and Trp/2 NAA ratio were observed when acids, had been added. When methionine was added to gluten, plasma amino acid methionine additions were made to gluten. On the other hand, the sum of the neutral concentrations, with the exception of me amino acids was less severely affected by thionine, were depressed. When the casein diets were fed, the the amino acid additions to casein, and plasma amino acid profiles were similar plasma Trp/2 NAA ratio was higher when I'lasnia untino acid concentrations of weanling rats
Protein choice5
addeda/100
4515 +
5530 +
+ 60Lvsine
g0
ml6.4
3 60
±0.7'«' 6.2±0.5"6 27.3 25.5 14.9 13.5 9.6 6.2±0.4°»7.224.243.9 30.837.1 14.220.0 13.213.5 8.811.7
0.068 90.8 91.3126.2 0.0380.057
3 60
±0.5» 5.4±0.56 40.8 35.5 18.4 11.3 9.9 6.2 ±0.5° 67.0±0.3°» 29.146.1 27.738.7 16.419.2 12.912.4 9.110.5
115.8 0.046 95.3126.9 0.0640.055
5.7 ±0.3°» 30.3 27.2 15.0 14.0 7.8 94.3 0.060 3 6.2±0.4°»33.32 6Ti-pValLeuHeTyrPheZNAATrp/SNAAltmolee/100 35.328.22 26.315.92 14.913.22 11.214.42 8.5105.0 96.10.0610.063
1 Mean ±SEMof six samples. Means with different superscripts are significantly different from each other (P < 0.05). from single analysis of a pooled sample of six rats.
a Value
Downloaded from https://academic.oup.com/jn/article-abstract/105/11/1412/4768666 by East Carolina University user on 13 January 2019
TABLE 2 Plasma tryptophan and neutral amino acid concentrations of weanling rats self-selecting from gluten diets
1416
DAVID V. M. ASHLEY AND G. HARVEY ANDERSON
TABLE 3 Plasma amino acid concentrations of control animais self-selecting from gluten or casein diets1
TABLE 4 Plasma tryptophan and neutral amino acid concentrations of weanling rats self-selecting from gluten or casein diets Dietary additionsTrpValLeuHeTyrPheZNAATrp/SNAAlimóles/100 mlGlutenNone
(control) Lysine Methionine Lysine, methionine, tryptophanCaseinNone threonine, (control) Lysine Methionine Methionine, arginine, tryptophan1 phenylalanine,
12
6.8±0.8° 34.4 24.4 13.4 12.4 93.1 0.073 8.5 5.4±0.5°6.7±0.9«7.8±1.3° 13.112.015.8 0.0950.0890.086 22.429.640.5 7.813.814.3 6.411.812.7 7.18.27.256.875.390.5
7.8±0.6° 46.7 28.9 17.0 13.2 0.069 6.3 112.076.8102.9(P ±1.0°9.8±0.9«31.32 8.3 0.1080.0951 4.17.2vith 28.943.128. 22.124.616.02 11.413.712.22 10.314.313.02
Mean - - s Mof six samples. Means
plasma amino acids •tryptophan •self-selection •weanling rats
Since the early observations of a relation ship between amino acid excess, decreased food intake, and the accumulation of amino acids in the blood of chicks ( 1 ), a number of studies have described correlations be tween the food intake depression that oc curs when high protein or amino acidimbalanced diets are fed, and altered blood amino acid patterns (2-4). Areas separate from the hypothalamus have been shown to mediate the food intake response caused by amino acid-imbalanced or high protein diets (5-9). Although the mecha nisms by which rats alter their food intake in response to these diets have not been identified, it has recently been shown, using a feeding system allowing weanling rats to select both protein and energy, that a mechanism operates to control protein intake (10, 11). Because protein is usually fed as a fixed proportion of the diet, the importance of this mechanism, which ap pears to interact with the mechanism con trolling energy intake, in the regulation of food intake has been disregarded. A precise relationship between plasma amino acid patterns and food intake has 1412
•neutral amino
been difficult to establish because many changes in plasma patterns lead to altera tions in food intake. However a specific as sociation between a neurotransmitter, serotonin, and food intake has been stated recently (12-15). The concentration of serotonin in the brain has been shown to be related to the brain concentration of tryptophan, from which it is synthesized ( 13, 14 ). Brain tryptophan concentration is dependent on the relative concentrations of plasma tryptophan to the five neutral amino acids with which it shares a com mon transport mechanism to the brain (i.e., tyrosine, phenylalanine, leucine, isoleucine, and valine) (16). Alterations in plasma amino acid concentrations brought about by alterations in the amino acid composition of the dietary protein have been shown to influence the synthesis of serotonin (14). The present experiments describe a relationship between protein intake and the ratio of plasma tryptophan Received for publication March 6, 1975. 1 Supported by grant MA-4392 from the Medical Research Council of Canada. * Address reprint requests to G. II. Anderson.
Downloaded from https://academic.oup.com/jn/article-abstract/105/11/1412/4768666 by East Carolina University user on 13 January 2019
Correlation between the Plasma Tryptophan to Neutral Amino Acid Ratio and Protein Intake in the Self-selecting Weanling Rat1'2
RAT PLASMA AMINO ACIDS AND PROTEIN SELECTION
1413
to the sum of the plasma concentration of the neutral amino acids in the self-select ing weanling rat.
Downloaded from https://academic.oup.com/jn/article-abstract/105/11/1412/4768666 by East Carolina University user on 13 January 2019
TCA. Plasma tryptophan was determined fluorometrically on each sample by the method of Denckla and Dewey (17). One milliliter of the remaining plasma was deMETHODS proteinized with 3 ml of 10% sulfosalicylic Male weanling rats of the Wistar strain 3 acid, and 0.5 ml of the clear supernatant and of weight range 40-50 g were housed of the six samples in each group was pooled. A minimum of 0.5 ml of the pooled and maintained for 4 weeks under condi tions previously described (11). During sample was analyzed for plasma amino this time they were allowed to freely acids on an automatic amino acid ana balance their dietary protein and energy lyzer.4 Statistical analyses of the plasma tryptophan intakes from two food cups that were of using Tukey'sconcentrations multiple t testwere and performed Dunnett's fered simultaneously and that differed from procedures (18). each other only in protein concentration. Details of the experimental design have RESULTS been documented in the preceding paper (11), but the design is briefly restated Experiment 1. The relative changes in here. plasma amino acid concentrations of the Three experiments were conducted. In experimental groups are illustrated in fig experiment 1, 9 groups of 10 weanling rats ure 1 according to the format used by were offered a choice of 5 and 45%, 15 Peng et al. (19). Each section of the and 55%, or 30 and 60% gluten diets to figure represents a different dietary pro which either 0, 3, or 6 g of lysine had been tein combination. For each combination added to each 100 g of gluten. fed, the effect of increasing the level of In experiment 2, 8 groups of 10 wean lysine in gluten by 3 or 6% on plasma ling rats were offered a choice of 15 and amino acids is shown expressed as a per 55% protein diets formulated with gluten centage of the values obtained when the or with casein to which none, lysine, me- gluten containing no added lysine was thionine, or the four most limiting amino fed (table 1). acids were added to the protein. The increase in lysine level in gluten In experiment 3, 12 groups of 10 wean increased plasma lysine concentration re ling rats were fed 15 and 55% zein diets. gardless of the dietary combination (fig. Six groups were fed the zein diets to which 1). Arginine concentrations were in three levels of lysine (1.7, 3.4, or 6.8 g/100 creased with the increased dietary lysine g) were added to zein made adequate in concentration in gluten when the dietary tryptoohan (zein plus tryptophan) or zein, choices were 5 and 45% or 15 and 55%, partially corrected with other amino acids but not when the choice was 30 and 60% in addition to tryptophan (partially cor gluten. The addition of lysine to gluten rected zein plus tryptophan). The six re was consistently associated with a decrease maining groups were offered 15 and 55% in plasma essential amino acids, threonine, valine, leucine, isoleucine, and phenylzein diets to which three levels of trypto phan (0.3, 0.6, or 1.2 g/100 g) were added alanine. Of the dispensable amino acids, to zein made adequate in lysine (zein plus the most consistent changes were an in lysine) or to partially corrected zein that crease in glutamic acid and a decrease in was adequate in lysine (partially corrected glutamine. zein plus lysine). Plasma tryptophan concentrations (table At the end of each 4-week study, blood 2) were unaffected by the 3 or 6% addi samples were collected between 9 and tion of lysine except when 3% lysine was 11 AM from the abdominal vena cava of added and offered in the 15 and 55% six animals in each group. The animals dietary combination. On the other hand, were first anesthetized with ether. The the sum of the plasma concentrations of blood samples were stored in chilled valine, leucine, isoleucine, phenylalanine, heparinized tubes, centrifuged, and 1 ml 3 Woodlyn Farms, Guelph. Ontario. Cannila. of the plasma was aliquoted and deoro4 Beckman Spinco automatic analyzer, model 11G/ teinized with an equal volume of 20% 119, Beckman Instruments Inc., Palo Alto, Calif.
1414
DAVID V. M. ASHLEY AND G. HARVEY ANDERSON
un im IMO «D
no
UDO
in m 200 UO MO l« 120 HO
120 SUO
ino no
5n »to rit
i
B 20 230 200 I» l«
o«»
Sa §300
a «
£ MO MO 120
Fig. l Effects of lysine additions to gluten on relative plasma amino acid concentrations in weanling rats self-selecting from a, 5 and 45%; b, 15 and 55%; or c, 30 and 60% gluten diets (• •, 3% lysine added; •—-•, 6% lysine added). Single analysis of a pooled sample from six rats in each group.
and tyrosine (table 2) decreased when the lysine additions were made, and the ratio of tryptophan to neutral amino acids (Trp/2 NAA) increased, with the one ex
n ai a 40
a
Fig. 2 Effects of addition of lysine, methionine, and the first four limiting amino acids to a, gluten and b, casein on relative plasma amino acid concentrations in weanling self-selecting rats (• •, 6% lysine added; •——•, four limiting amino acids added; • •, 2.5% methionine added). Single analysis of a pooled sample from six rats in each group.
Downloaded from https://academic.oup.com/jn/article-abstract/105/11/1412/4768666 by East Carolina University user on 13 January 2019
ino isoo
ception, that of the 3% lysine, 15 and 55% group. Experiment 2. The plasma amino acid profiles of the groups self-selecting from the gluten and casein diets are shown in figure 2a and b, respectively. The indi vidual concentrations of plasma amino acids of the control groups are shown in table 3. Increasing the lysine level of gluten by 6% caused an increase in the plasma lysine concentration and a decrease in plasma ornithine but had little effect on the con centrations of the other plasma amino acids (fig. 2a). With the exception of the added amino acids, lysine, threonine, and
1410
RAT PLASMA AMINO ACIDS AND PROTEIN SELECTION TABLE 1
for the groups fed casein to which no amino acid additions were made and those self-selecting from gluten diets fed casein to which lysine or the four most with no added lysine1 limiting amino acids were added, except for the plasma concentrations of lysine, Dietary choice leucine, and methionine. Plasma lysine con Amino acid 5 + 45 15 + 55 30 + 60 centrations were elevated when lysine alone was added to casein but were un mlAspart limóles/too affected by the addition of the four most acidThreonineSerineAsparagineGlu ic limiting amino acids to casein. Leucine and methionine concentrations were in creased in the plasma of all treatment neGlutamic tam i groups. When methionine additions were acidGlycineAlanineValineMethionineIsoleucineLeucineTv made to casein, the plasma concentrations of all amino acids with the exception of leucine and methionine were lower than the plasma amino acid concentrations of the animals fed the casein diet containing rosine no added amino acids. PhenylalanineOrnithineLysineHistidineArginine3.430.033.39.075.914.824.854.833.35.815.928.213.2 14.46.54.98.69.32.329.435.28.373.715.720.561.143.96.820.037.113.5 10.55.05.011.013.4 11.77.44.39.16.93.430.039.35.461.221.822.059.946.17.919.238.712.4 There were no significant differences in plasma tryptophan concentrations of the animals fed gluten or casein diets (table 4). However, the animals fed casein diets had higher plasma tryptophan concentra 1Single analysis of a pooled sample from six rats. tions than those fed gluten diets. The sum of the five neutral amino acids (table 4) methionine, plasma amino acid concen was lowered by all amino acid additions to trations were lower in the plasma of rats gluten, and as a result the Trp/2 NAA fed gluten to which the four limiting amino ratio was increased. The most marked ef acids, rather than lysine alone or no amino fects on the neutral amino acids and Trp/2 NAA ratio were observed when acids, had been added. When methionine was added to gluten, plasma amino acid methionine additions were made to gluten. On the other hand, the sum of the neutral concentrations, with the exception of me amino acids was less severely affected by thionine, were depressed. When the casein diets were fed, the the amino acid additions to casein, and plasma amino acid profiles were similar plasma Trp/2 NAA ratio was higher when I'lasnia untino acid concentrations of weanling rats
Protein choice5
addeda/100
4515 +
5530 +
+ 60Lvsine
g0
ml6.4
3 60
±0.7'«' 6.2±0.5"6 27.3 25.5 14.9 13.5 9.6 6.2±0.4°»7.224.243.9 30.837.1 14.220.0 13.213.5 8.811.7
0.068 90.8 91.3126.2 0.0380.057
3 60
±0.5» 5.4±0.56 40.8 35.5 18.4 11.3 9.9 6.2 ±0.5° 67.0±0.3°» 29.146.1 27.738.7 16.419.2 12.912.4 9.110.5
115.8 0.046 95.3126.9 0.0640.055
5.7 ±0.3°» 30.3 27.2 15.0 14.0 7.8 94.3 0.060 3 6.2±0.4°»33.32 6Ti-pValLeuHeTyrPheZNAATrp/SNAAltmolee/100 35.328.22 26.315.92 14.913.22 11.214.42 8.5105.0 96.10.0610.063
1 Mean ±SEMof six samples. Means with different superscripts are significantly different from each other (P < 0.05). from single analysis of a pooled sample of six rats.
a Value
Downloaded from https://academic.oup.com/jn/article-abstract/105/11/1412/4768666 by East Carolina University user on 13 January 2019
TABLE 2 Plasma tryptophan and neutral amino acid concentrations of weanling rats self-selecting from gluten diets
1416
DAVID V. M. ASHLEY AND G. HARVEY ANDERSON
TABLE 3 Plasma amino acid concentrations of control animais self-selecting from gluten or casein diets1
TABLE 4 Plasma tryptophan and neutral amino acid concentrations of weanling rats self-selecting from gluten or casein diets Dietary additionsTrpValLeuHeTyrPheZNAATrp/SNAAlimóles/100 mlGlutenNone
(control) Lysine Methionine Lysine, methionine, tryptophanCaseinNone threonine, (control) Lysine Methionine Methionine, arginine, tryptophan1 phenylalanine,
12
6.8±0.8° 34.4 24.4 13.4 12.4 93.1 0.073 8.5 5.4±0.5°6.7±0.9«7.8±1.3° 13.112.015.8 0.0950.0890.086 22.429.640.5 7.813.814.3 6.411.812.7 7.18.27.256.875.390.5
7.8±0.6° 46.7 28.9 17.0 13.2 0.069 6.3 112.076.8102.9(P ±1.0°9.8±0.9«31.32 8.3 0.1080.0951 4.17.2vith 28.943.128. 22.124.616.02 11.413.712.22 10.314.313.02
Mean - - s Mof six samples. Means
