A concept of dietary dipeptides: a step to resolve the problem of amino acid availability in the early life of vertebrates
1 School of Natural Resources, The Ohio State University, Columbus, OH
43210, USA
2 Institute of Aquaculture Research, Protein and Amino Acid Section,
Sunndalsøra, N-6600, Norway
3 Aquaculture Protein Centre CoE, Protein and Amino Acid Section,
Sunndalsøra, N-6600, Norway
4 The Ohio State University Interdisciplinary Program in Nutrition,
Columbus, OH 43210, USA
5 Metabolism and Cancer Susceptibility Section, Laboratory of Comparative
Carcinogenesis, National Cancer Institute, Frederick, MD 21702, USA
6 Faculty of Applied Marine Science, Cheju National University, Jeju
690-756, South Korea
* Author for correspondence (e-mail: dabrowski.1{at}osu.edu)
Accepted 17 May 2005
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Summary |
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Key words: dipeptide, protein, amino acid, proline, teleost, rainbow trout, Oncorhynchus mykiss
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Introduction |
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Requirements of protein for maximum growth in teleost larval and juvenile
stages are nearly twice as high as in older fish
(Dabrowski, 1986). The reason
for this may be due to underestimated IDAA requirements because of suboptimal
growth rates in juvenile fish when purified diets were used
(Dabrowski, 1986
;
NRC, 1993
). There is
essentially no data for larval, pre-metamorphosed fish amino acid requirements
because of limitations in formulating acceptable diets.
Determination of tissue free amino acid (FAA) levels during requirement
studies or comparison with dietary amino acid composition has met with
variable success in fish (Schumacher et
al., 1997; Yamamoto et al.,
2000
). However, most of these measurements have been made for
plasma, while in fish the white musculature is quantitatively the most
important site for protein accretion
(Carter and Houlihan, 2001
).
In humans, muscle FAAs have been found to vary according to dietary levels and
pathological and catabolic conditions
(Fürst and Stehle, 2004
).
Ontogenetic, dietary and post-prandial effects on muscle FAAs are also
important in rainbow trout (Carter et al.,
1995
).
We documented previously that a synthetic dipeptide-based diet supported
growth of rainbow trout alevins (first-feeding) whereas a free amino acid
mixture diet did not (Dabrowski et al.,
2003). Hydrolysates as sources of peptides are insufficient, since
such peptides are impaired by their characteristics, i.e. the range of
molecular sizes and difficulties in controlling their amino acid composition
in requirement studies. There is evidence that tetra- and larger peptides, in
the absence of pancreatic enzymes and deficiencies of brush border peptidase
activity, become incapable of covering nitrogen requirements
(Grimble, 1994
;
Daniel, 2004
). Losses of IDAA
during hydrolysate preparation are frequently responsible for nutritional
inadequacies resulting in growth depression
(Langar et al., 1993
). This
may have been the case when the proportion of hydrolysate was higher than 20%
protein replacement in larval fish diets
(Cahu et al., 1999
).
Advantages of using specific di- or tripeptides as substrates for
particular peptide transporters have been determined based on the affinity of
PEPT1 transporters (Doring et al.,
1998). These transporters were found to be expressed in larval
teleosts prior to the first exogenous feeding
(Verri et al., 2003
). We
hypothesize that synthetic dipeptide diets can be instrumental in defining
amino acid requirements for pre-metamorphosed fish, which is largely unknown.
As a first step, we set out to determine whether physiological indices such as
muscle FAA and enzyme activities would respond to such diets, in comparison
with a dietary mixture of synthetic amino acids or other molecular forms. The
specific amino acid composition of the dipeptide diets was based on the
capacity of cytosolic peptidases in fish intestinal epithelial cells
(Aranishi et al., 1998
) and on
known IDAA requirements for rainbow trout juveniles
(NRC, 1993
). A series of
feeding trials was carried out with first-feeding alevins and larger, juvenile
rainbow trout (Dabrowski et al.,
2003
). We hypothesized that the concentrations of FAA in fish
muscle (constituting 6580% of body mass;
Weatherley and Gill, 1987
)
would be an integrated measure of availability of dietary amino acids for
protein synthesis and, at the same time, an indication of protein synthesis
and degradation rates. Although not anticipated, we discovered that free
proline concentration in muscle showed a strong dietary dependency and was
possibly a conditional indispensable amino acid, similar to in young mammals
(Kirchgessner et al., 1995
).
We followed this line of inquiry and, for the first time, analyzed
pyrroline-5-carboxylate reductase (P5CR) activity in fish, the final step in
proline biosynthesis.
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Materials and methods |
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Experiment 1
Rainbow trout Oncorhynchus mykiss Walbaum of a local strain
(London, OH, USA) were hatched and, at the first-feeding stage (114±16
mg wet mass individual1), distributed randomly into
triplicate tanks for each dietary treatment. After 5 weeks of feeding, fish
were collected from each tank and transferred into an ice water bath and then
individually frozen on dry ice and stored at 80°C for FAA
analyses.
Experiment 2
Juvenile rainbow trout of 0.78±0.08 g initial mass
individual1 and approximately 5 weeks old were used. Three
diets were tested: Free AA, dipeptide and casein(). At the completion
of the 2-week experiment, sampling tissues for FAA and P5CR activity analyses
commenced in the morning, 24 h after the last feeding, and were conducted
randomly across tanks and treatments. Fish were killed and samples stored in
the same manner as in Experiment 1.
Experiment 3
This experiment aimed to determine changes in P5CR activity with time
duration of the assay, i.e. to optimize kinetics. Furthermore, it aimed to
analyze tissue distribution of P5CR activity between week 2 and 6 after the
first feeding. Thus, it would include the time-periods of rainbow trout
ontogeny studied in Experiments 1 and 2. Rainbow trout alevins were reared as
described earlier and fed a casein-based diet (30% casein, 6% gelatin, 6%
casein-hydrolysate; K. Dabrowski and M. Penn, personal communication) until
sampling at weeks 2 and 6. Fish were collected from each tank at sampling and
individually frozen in liquid nitrogen and stored at 80°C. To
establish the assay conditions and presence of P5CR activity in fish, we used
fresh tissues and individuals of larger size to separate organs of interest:
liver and intestine. Tissue samples of 12-month-old rainbow trout were
therefore assayed. These fish were fed a standard commercial diet
(Zeigler-Bros., Gardners, PA, USA). After 48 h fasting, four fish
(53±12 g individual1, 17.6±1.5 cm length) were
killed by a sharp blow to the head, and the intestine (including pyloric
caeca) and the liver dissected out, rinsed in 0.9% NaCl and assayed
immediately.
FAA analysis
Individual fish from Experiments 1 and 2 were rapidly dissected while still
frozen on ice-cooled boards. The head and tail (beyond the abdominal cavity)
were removed, and the dorsal body region, anterior and posterior to the dorsal
fin, was dissected out (28±8% of body mass; N=76). Care was
taken to avoid remains of kidney tissue by scraping any blood containing
tissue from the upper part of the body cavity. The tissue samples were weighed
and re-frozen at 80°C. Thus, these muscle tissues also contained
skin, cartilage and bone, but were largely composed of white muscle, and are,
for simplicity, termed `muscle'. Within two days, muscle samples were
extracted in 0.1 mol l1 HCl containing 160 µmol
l1 norleucine, using a tissue:extraction medium ratio of 1:4
(juveniles) or 1:10 (alevins) (w/v), according to Cohen et al.
(1989). The norleucine
recovery was 104±28% (N=52). Tissue extracts were spun at 12
000 g (4°C, 15 min), and supernatants filtered (Millipore,
Billerica, MA, USA; 10 kDa cut-off at 2000 g, 4°C, 90
min). Samples of blanks and external standards (Sigma acid/neutral and basic
amino acids), supplemented with glutamine, were prepared at the same time as
sample extraction by adjusting appropriately with distilled and deionized
H2O, and 0.1 mol l1 HCl containing 160 µmol
l1 norleucine. Tissue extracts, standards and blanks were
then stored at 80°C, for later analysis using the Waters (Milford,
MA, USA) PicoTag method with pre-column derivatization and reverse-phase
high-performance liquid chromatography (RP-HPLC;
Cohen et al., 1989
).
Enzyme activity
P5CR (EC 1.5.1.2) activity was measured using modifications of mammalian
protocols (Herzfeld et al.,
1977; Dekaney et al.,
2003
). DL-P5C was produced from its
2,4-dinitrophenylhydrazone (Sigma), purified by cation-exchange
chromatography, analyzed with o-aminobenzaldehyde and stored at
4°C in 0.5 mol l1 HCl
(Mezl and Knox, 1977
). Liver
and intestine from juvenile trout (assays in fresh tissues), or liver and
intestine from alevins or juveniles from Experiment 3 (N=3 tanks),
for P5CR activity analysis were obtained using dissection techniques described
above. The whole intestine, including pyloric caeca and pancreas, was used.
Liver constituted 12%, and intestine 89%, of total body mass,
and no changes in hepatic or intestinal indices were found when comparing
Experiment 3 alevins fed for 2 weeks (0.27±0.03 g body mass) with those
fed for 6 weeks (1.08±0.09 g body mass).
To assess total P5CR activity in juvenile trout fed the different
experimental diets (Experiment 2), assays were run on the whole fish body
(Terjesen et al., 2001).
Tissues to be assayed for P5CR activity were homogenized on ice
(Potter-Elvehjem at 40 rev min1, 60 s, two strokes) in 250
mmol l1 sucrose, 50 mmol l1 phosphate
buffer (pH 7.2), 1 mmol l1 EDTA, 2.6 mmol
l1 dithiothreitol, and 2.5 µg ml1 each
of aprotinin, pepstatin A, chymostatin and phenyl methyl sulphonyl fluoride.
Whole-body samples were gently disrupted prior to the Potter-Elvehjem by an
Omni GLH homogenizer (4000 rev min1, 15 s). All extracts
were spun at 600 g (10 min, 4°C), the supernatant further
centrifuged at 15 000 g (10 min, 4°C), and the final
supernatant used for measurements. This approach
(Dekaney et al., 2003
) was
followed assuming that P5CR has a cytosolic location in fishes, as in mammals.
Proline oxidase, which could utilize the P5CR reaction product, has a
mitochondrial location in mammals (Wu and
Morris, 1998
). Immediately before assay, the P5C substrate was
neutralized on ice, using 1 mol l1 Hepes (pH 7.0) and 1 mol
l1 NaOH. Duplicate reactions were run in 100 µl total
volume (10 µl extract), containing 4.5 mmol l1
DL-P5C, 2 mmol l1 NADH and 45 mmol
l1 phosphate buffer (pH 6.8). After 015 min at
26°C (Terjesen et al.,
2001
), reactions were terminated by 25 µl of 1.5 mol
l1 HClO4 and neutralized with 12.5 µl of 2 mol
l1 K2CO3. After centrifugation (600
g), proline was determined by PicoTag RP-HPLC
(Cohen et al., 1989
), and
protein determined by the Bradford method
(Bradford, 1976
). The amount of
proline formed was linear with time and protein concentrations, and control
assays without P5C or enzyme extract did not result in detectable activity
(chromatograms not shown).
Data analysis
Data are presented as means ± S.D. Tank means
were used as the statistical unit, and data were analyzed in SPSS version
12.0.1. (SPSS Inc., Chicago, IL, USA) using one-way analysis of variance
(ANOVA) for all tests except for Experiment 3 regarding P5CR tissue
distribution and age since first-feeding (two-way ANOVA). If significant
(P<0.05), Duncan's multiple range tests were employed.
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Results |
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Several FAAs in muscle, such as histidine, threonine, valine, arginine and alanine, corresponded to dietary amino acid profiles in Experiments 1 and 2. The casein(+/) diets had a higher IDAA level than the Free AA and dipeptide diets, and this was mirrored in both alevin and juvenile muscle FAA concentrations (Figs 1B, 2B; Tables 2, 3), and resulted in the highest growth rates (Figs 1, 2, top diagram). However, these differences in muscle free IDAA, e.g. histidine (38-fold), were in many cases several folds larger than the dietary level differences (Figs 1B, 2B; Tables 2, 3). Among free DAA, alanine, which was high in the Free AA diet, reached 18 mmol kg1 wet mass in muscle of juveniles (Fig. 2A), and alanine concentrations were significantly higher in fish of both life stages fed the Free AA diet compared with trout fed other diets. Free aspartic or glutamic acids in muscle did not indicate any impact of dietary levels (Figs 1A, 2A).
The muscle concentrations of several free IDAA were affected by dietary
molecular form. In particular, isoleucine and methionine were present in
muscle at higher levels when given a Free AA diet, in both alevin (Experiment
1) and juvenile trout (Experiment 2), compared with the dipeptide diet fed
fish, despite similar dietary levels (Figs
1B,
2B; Tables
2,
3). This observation may imply
that an excess of amino acids was not utilized for protein synthesis in the
Free AA diet fed alevins (Experiment 1), since negligible weight gains
(Fig. 1) and elevated ammonia
excretion were observed (Dabrowski et al.,
2003). To the contrary, there was no significant difference in
ammonia excretion rate in juveniles fed Free AA and casein() diets, and
improved, although low, weight gain was observed in juveniles fed a Free AA
diet (Fig. 2;
Dabrowski et al., 2003
).
The assays of fresh rainbow trout juvenile tissues showed that P5CR
activity was present in liver and intestine (data not shown). P5CR activity
was present in the intestine and liver of rainbow trout alevins fed for 2
weeks (Fig. 3A). Liver had
significantly higher P5CR activity than intestine (P<0.01), but no
significant effects of age, or age x tissue, were noted (two-way ANOVA).
The effects of different dietary sources of amino acids on P5CR activity were
investigated in Experiment 2 (Fig.
3B). The juvenile trout fed the dipeptide diet showed numerically
higher P5CR activity than juveniles fed Free AA- and casein-based diets,
although differences were not significant (P=0.25; one-way ANOVA).
When the total P5CR activity in the whole body was estimated in the
casein-diet-fed trout (0.7 U g1 wet mass) and compared with
the activity of intestine (1.0 U g1 wet mass) and liver (3.1
U g1 wet mass), it was calculated that these two tissues
(11% of body mass) accounted for 20% of whole-body P5CR activity. In
conclusion, rainbow trout alevins and juveniles expressed P5CR activity in
liver and intestine, but fish could not maintain muscle free proline when fed
proline-deficient dipeptide diets (free proline muscle levels were 10-fold
lower) in comparison with fish fed proline-containing casein()
(juveniles) or both casein(+) and () diets (alevins) (Figs
1C,
2C).
|
Rainbow trout alevins fed Free AA- or dipeptide-based diets (Experiment 1)
showed different responses in muscle levels of ornithine and proline and
interactions with growth. Ornithine, which may be synthesized from P5C, was
higher in alevins fed Free AA diet than in alevins fed other diets
(Fig. 1C;
Table 2), concurrent with a
negligible growth rate (Fig. 1)
and very low proline muscle concentrations
(Fig. 1C), despite being given
a diet high in proline. By contrast, dipeptide-fed alevins showed
significantly higher growth rates than the FAA-fed groups
(Fig. 1;
Dabrowski et al., 2003), and
the dipeptide-fed fish had comparable muscle ornithine levels to that of the
casein(+/)-fed fish (Fig.
1C). Despite being given a dipeptide diet, devoid of peptide
proline, these fish produced significantly higher [Orn]/[Pro] ratios
(Table 2). In regard to
hydroxyproline, in juveniles (Experiment 2) the free muscle levels were
comparable in the Free AA- and dipeptide-fed groups, but 7-fold lower than in
fish fed the casein()-based diet
(Fig. 2C). In alevins
(Experiment 1), the dipeptide-fed fish (126±20 µmol
kg1 wet mass) had a numerically higher free hydroxyproline
concentration in muscle than did the Free AA-fed fish (37±2 µmol
kg1 wet mass); however, this was 27-fold lower
(3394±430 µmol kg1 wet mass) than in the
fast-growing (4.5% day1) rainbow trout alevins fed the
casein() diet (Fig.
1C).
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Discussion |
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In fed animals, weight gains usually arise as a result of increased protein
synthesis and decreased protein degradation in comparison with fasted animals
(Waterlow, 1999). Several
factors are pivotal in determining the effectiveness of these processes, such
as concentrations and correct proportions, i.e. balanced amino acid
composition to achieve protein accretion, i.e. growth. We submit that the
differences in muscle FAA shown here (1) are due to differences in amino acid
absorption rates from free, dipeptide or protein dietary sources, (2) result
in uneven accumulation rates and post-prandial peak times for muscular FAA and
(3) consequently result in different metabolic handling of the amino acids and
availability for protein synthesis. Superimposed on these are differences in
growth rate, protein synthesis rate and developmental stages, which demand
varying amounts of amino acids at the quantitatively most important site for
protein accretion, white muscle. This set of factors, together with enzyme
expression data, provides more insight into amino acid nutrition than nitrogen
balance or growth rate alone. As an example, methionine was provided in
comparable amounts in all three diets, but methionine given in dipeptide form
was present in muscle of alevins in significantly lower amounts than in fish
with essentially no growth (Free AA group) and higher growth rates
[casein(+/) groups], suggesting limitations of the GlyMet
dipeptide availability (Fig.
1B).
A major finding in the present study is the strong dependence of free
proline in muscle tissue on dietary level of proline in dipeptide- and
casein-based diets (Figs 1C,
2C), since it can be assumed
that any excess of proline would be catabolized for energy via P5C
and ornithine or glutamate. In rainbow trout juveniles fed a dipeptide-based
diet devoid of proline, endogenous synthesis did not maintain proline levels
comparable with fish fed diets with proline [casein(+/)]. The free
proline levels in muscle of the dipeptide-fed group are at the lower end of
the range reported in fish muscle
(Torrissen et al., 1994;
Yamamoto et al., 2000
;
Ogata, 2002
). In large rainbow
trout fed a low-protein, 5% casein-containing diet, only `trace' amounts of
free proline were found in muscle, whereas in fish on a 50% casein diet, the
proline concentration was 3400 µmol kg1
(Yokoyama and Nakazoe, 1991
).
In mammals, although proline can be synthesized from ornithine or glutamate
via P5C, young mammals, rats and pigs still require a dietary source
of proline for maximum growth and protein retention
(Ball et al., 1986
).
Furthermore, increased dietary proline levels in young rats and pigs correlate
with plasma proline concentrations
(Samuels et al., 1989
;
Kirchgessner et al., 1995
),
and no significant response was found in P5CR activity when comparing pigs fed
control and proline-deficient diets. The findings in young mammals correspond
to the present study on rainbow trout. In fish, a 10-fold drop in the free
proline in muscle was associated with a 48% increase (ANOVA not significant)
in P5CR activity in the dipeptide-fed rainbow trout, a diet devoid of proline
(Fig. 3B). In other fish
(Pacific salmon), juveniles fed AA-based diets were reported for the first
time by Halver et al. (1957
)
to grow at the rate of 0.45% day1. That is similar to
rainbow trout juveniles in the present study (0.48% day1;
Fig. 2). The growth data for
fish fed diets with no proline in the Halver et al. study, however, ended much
earlier than with other treatments where AA were omitted from the diets.
Therefore, results cannot be compared directly. Similarly, growth of young
tilapia on an AA-based diet devoid of proline was shown to be inferior
(Aoe et al., 1970
). However,
these authors discontinued the study. Since larger fish were used in most
amino acid requirement studies, a possible proline conditional
indispensability for optimal growth of larval fish was not demonstrated. The
classical study (Halver and Shanks,
1960
) indicates that the final mass of sockeye salmon fed a
proline-deficient diet was less than that of fish fed a control diet, and the
experiment was shortened to only 5 weeks. In addition, in this previous study
(Halver and Shanks, 1960
) and
earlier experiments with Chinook salmon
(Halver et al., 1957
), a
proline-deficient diet was supplemented with more than a generous amount of
arginine (3.65%). Interconversion of arginine to ornithine and then
ornithine to proline in neonatal mammals can reach 25 and 57%, respectively
(Bertolo et al., 2003
).
Regarding fish, early life stages are characterized by high protein synthesis
rates (170300%), and protein deposition rates of 50% body protein per
day have been observed in larval catfish
(Terjesen et al., 1997
) and
other species (Fauconneau et al.,
1986
). Furthermore, during the saltatory development of larval
fish, muscle fibres and skeleton undergo a considerable change
(Blaxter, 1988
), demanding
collagen synthesis. Since proline and hydroxyproline constitute more than 20%
of the amino acid residues in collagen in mammals
(Smith and Phang, 1978
), and
in fish hydroxyproline alone is estimated at 7%
(Sato et al., 1989
), a
requirement for dietary proline supplementation should be investigated in
exogenous feeding during the early life stages. We submit that dipeptide-based
diets can be instrumental in determining dietary requirements for IDAA,
including conditional indispensability for proline, in larval and juvenile
fish.
High concentrations of hydroxyproline in muscle of fast-growing animals is
an indicator of a high rate of collagen turnover
(Adams and Frank, 1980). Since
hydroxyproline released by collagen breakdown and subsequent hydrolysis of
Hyp-containing peptides is destined for catabolism and excretion, present
results in rainbow trout are the best indication thus far of the muscle
limiting degradation of collagen when there is no dietary supply of proline
(Figs 1C,
2C). In juvenile salmon, muscle
hydroxyproline was the only FAA that correlated with growth rates
(Sunde et al., 2001
). Although
we indicate the presence of hydroxyproline in the casein(+/)-based
diets, we must assume that transport of dietary hydroxyproline from intestine
to muscle is limited (Adams and Frank,
1980
), and thus high concentrations in muscle of a
casein/gelatin-based diet-fed fish would reflect high protein turnover rates.
This would suggest that low proline and hydroxyproline concentrations are
indicative of either fasting (the Free AA diet fed alevins) or dietary
limitation (the dipeptide diet). We submit that this paradox is related to the
only cursory understanding of the prolineornithinearginine
pathway in fish, conditional indispensability and interconversion of these
amino acids.
In conclusion, present observations and data collected with the use of dipeptideprotein (1:1 ratio)-based diets (B.F.T. and K.D., unpublished data) suggest that dietary peptide transport and hydrolysis in rainbow trout early life stages can be improved by the inclusion of protein supplement and more efficiently hydrolysable dipeptides. Consequently, monitoring disproportions of FAA in muscle and expression of glutamateprolineornithine pathway enzymes, in concert with growth indices, is the right approach to address basic mechanisms of amino acid utilization in larval fishes. The modest success of nutrient administration in the form of dipeptides points to the potential for evaluation of amino acid requirements in early life stages of fishes that has not been possible thus far.
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Acknowledgments |
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References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Adams, E. and Frank, L. (1980). Metabolism of proline and the hydroxyprolines. Annu. Rev. Biochem. 49,1005 -1061.[CrossRef][Medline]
Aoe, H., Masuda, I., Abe, I., Saito, T., Toyoda, T. and Kitamura, S. (1970). Nutrition in young carp. 1. Nutritive value of free amino acids. Bull. Jap. Soc. Sci. Fish. 36,407 -413.
Aranishi, F., Watanabe, T., Osatomi, K., Cao, M., Hara, K. and Ishihara, T. (1998). Purification and characterization of thermostable dipeptidase from carp intestine. J. Mar. Biotechnol. 6,116 -123.
Ball, R., Atkinson, J. and Bayley, H. (1986). Proline as an essential amino acid for the young pig. Br. J. Nutr. 55,659 -668.[Medline]
Berge, G. M. and Storebakken, T. (1996). Fish protein hydrolyzate in starter diets for Atlantic salmon (Salmo salar) fry. Aquaculture 145,205 -212.[CrossRef]
Bertolo, R. F. P., Brunton, J. A., Pencharz, P. B. and Ball, R.
O. (2003). Arginine, ornithine, and proline interconversion
is dependent on small intestinal metabolism in neonatal pigs. Am.
J. Physiol. Endocrinol. Metab. 284,E915
-E922.
Blaxter, J. H. S. (1988). Pattern and variety in development. In Fish Physiology, vol.XIA (ed. W. S. Hoar and D. J. Randall), pp.1 -58. London, New York: Academic Press.
Boge, G., Roche, H. and Balocco, C. (2002). Amino acid transport by intestinal brush border vesicles of a marine fish, Boops salpa. Comp. Biochem. Physiol. B 131, 19-26.[CrossRef][Medline]
Bradford, M. M. (1976). A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of the protein-dye binding. Anal. Biochem. 72,248 -254.[CrossRef][Medline]
Cahu, C., Zambonino-Infante, J. L., Quazuguel, P. and Le Gall, M. M. (1999). Protein hydrolyzate vs. fish meal in compound diets for 10-day old sea bass (Dicentrarchus labrax) larvae. Aquaculture 171,109 -119.[CrossRef]
Carter, C. G. and Houlihan, D. F. (2001). Protein synthesis. In Fish Physiology. Vol.20 (ed. P. A. Wright and P. M. Anderson), pp.31 -75. San Diego, CA: Academic Press.
Carter, C., He, Z.-Y., Houlihan, D., McCarthy, I. and Davidson, I. (1995). Effect of feeding on the tissue free amino acid concentrations in rainbow trout (Oncorhynchus mykiss Walbaum). Fish Physiol. Biochem. 14,153 -164.[CrossRef]
Cohen, S., Meys, M. and Tarvin, T. (1989). The Pico-Tag® Method: A Manual of Advanced Techniques for Amino Acid Analysis.124 pp. Milford, USA: Millipore Corporation.
Dabrowski, K. (1986). Ontogenetical aspects of nutritional requirements in fish. Comp. Biochem Physiol. A 85,639 -655.[CrossRef][Medline]
Dabrowski, K., Lee, K.-J. and Rinchard, J.
(2003). The smallest vertebrate, teleost fish, can utilize
synthetic dipeptide-based diets. J. Nutr.
133,4225
-4229.
Daniel, H. (2004). Molecular and integrative physiology of intestinal peptide transport. Annu. Rev. Physiol. 66,361 -384.[CrossRef][Medline]
Dekaney, C. M., Wu, G. and Jaeger, L. A.
(2003). Gene expression and activity of enzymes in the arginine
biosynthetic pathway in porcine fetal small intestine. Pediatr.
Res. 53,274
-280.
Doring, F., Walter, J., Will, J., Focking, M., Boll, M.,
Amasheh, S., Clauss, W. and Daniel, H. (1998).
Delta-aminolevulinic acid transport by intestinal and renal peptide
transporters and its physiological and clinical implications. J.
Clin. Invest. 101,2761
-2767.
Fauconneau, B., Aguirre, P. and Bergot, P. (1986). Protein synthesis in early life of coregonids: Influence of temperature and feeding. Arch. Hydrobiol. Beih. Ergebn. Limnol. 22,171 -188.
Fürst, P. and Kuhn, K. S. (2000). Amino-acid substrates in new bottles: Implications for clinical nutrition in the 21st century. Nutrition 16,603 -606.[CrossRef][Medline]
Fürst, P. and Stehle, P. (2004). What are
the essential elements needed for the determination of amino acid requirements
in humans? J. Nutr. 134,1558S
-1565S.
Grimble, G. (1994). The significance of peptides in clinical nutrition. Annu. Rev. Nutr. 14,419 -447.[CrossRef][Medline]
Halver, J. (2002). Fish Nutrition. San Diego, CA: Academic Press.
Halver, J. E. and Shanks, W. (1960). Nutrition of salmonoid fishes. VIII. Indispensable amino acids for sockeye salmon. J. Nutr. 72,340 -346.[Medline]
Halver, J. E., Delong, D. and Mertz, E. (1957). Nutrition of salmonoid fishes. V. Classification of essential amino acids for Chinook salmon. J. Nutr. 63, 95-105.[Medline]
Herzfeld, A., Mezl, V. and Knox, W. (1977). Enzymes metabolizing 1-pyrroline-5-carboxylate in rat tissues. Biochem. J. 166,95 -103.[Medline]
Kim, K. I., Grimshaw, T. W., Kayes, T. B. and Amundson, C. H. (1992). Effect of fasting or feeding diets containing different levels of protein or amino acids on the activities of the liver amino acid-degrading enzymes and amino acid oxidation in rainbow trout (Oncorhynchus mykiss). Aquaculture 107,89 -106.[CrossRef]
Kirchgessner, v. M., Fickler, J. and Roth, F. (1995). Effect of dietary proline supply on N-balance of piglets. 3. Communication on the importance of non-essential amino acids for protein retention (in German). J. Anim. Physiol. A Anim. Nutr. 73, 57-65.
Langar, H., Guillaume, J., Metailler, R. and Fauconneau, B. (1993). Augmentation of protein synthesis and degradation by poor dietary amino acid balance in European sea bass (Dicentrarchus labrax). J. Nutr. 123,1754 -1761.[Medline]
Lee, K.-J., Dabrowski, K., Rinchard, J., Gomez, C., Guz, L. and Vilchez, C. (2004). Supplementation of maca (Lepidium meyenii) tuber meal in diets improves growth rate and survival of rainbowtrout Oncorhynchus mykiss (Walbaum) alevins and juveniles. Aquac. Res. 35,215 -223.[CrossRef]
Matthews, D. (1991). Protein absorption. New York: Wiley-Liss.
Mezl, V. A. and Knox, W. E. (1977). Properties and analysis of a stable derivative of pyrroline-5-carboxylic acid for use in metabolic studies. Anal. Biochem. 74,430 -440.
NRC (1993). Nutrient requirements of fishes. In National Research Council, Board of Agriculture (ed. C. Cowey, Y. C. Cho, K. Dabrowski, S. Hughes, S. Lall, R. Lovell, T. Murai and R. Wilson). Washington, DC: National Academy Press.
Ogata, H. (2002). Muscle buffering capacity of yellowtail fed diets supplemented with crystalline histidine. J. Fish Biol. 61,1504 -1512.[CrossRef]
Reshkin, S. and Ahearn, G. (1991). Intestinal glycyl-L-phenylalanine and L-phenylalanine trasnport in a euryhaline teleost. Am. J. Physiol. 260,R563 -R569.[Medline]
Samuels, S., Aarts, H. and Ball, R. O. (1989). Effect of dietary proline on proline metabolism in the neonatal pig. J. Nutr. 119,1900 -1906.[Medline]
Sato, K., Yoshinaka, R. and Sato, M. (1989). Hydroxyproline content in the acid-soluble collagen from muscle of several fishes. Nippon Suisan Gakkaishi 55, 1467.
Schumacher, A., Wax, C. and Gropp, J. (1997). Plasma amino acids in rainbow trout (Oncorhynchus mykiss) fed intact protein or a crystalline amino acid diet. Aquaculture 151, 15-28.[CrossRef]
Smith, R. and Phang, J. (1978). Proline metabolism in cartilage: the importance of proline biosynthesis. Metab. Clin. Exp. 27,685 -694.[Medline]
Sunde, J., Taranger, G. and Rungruangsak-Torissen, K. (2001). Digestive protease activities and free amino acids in white muscle as indicators for feed conversion efficiency and growth rate in Atlantic salmon (Salmo salar L.). Fish Physiol. Biochem. 25,335 -345.[CrossRef]
Terjesen, B. F., Verreth, J. and Fyhn, H. J. (1997). Urea and ammonia excretion by embryos and larvae of the African Catfish Clarias gariepinus (Burchell 1822). Fish Physiol. Biochem. 16,311 -321.[CrossRef]
Terjesen, B. F., Chadwick, T. D., Verreth, J. A. J., Rønnestad, I. and Wright, P. A. (2001). Pathways for urea production during early life of an air-breathing teleost, the African catfish Clarias gariepinus Burchell. J. Exp. Biol. 204,2155 -2165.[Medline]
Torrissen, K., Lied, E. and Espe, M. (1994). Differences in digestion and absorption of dietary protein in Atlantic salmon (Salmo salar) with genetically different trypsin isozymes. J. Fish Biol. 45,1087 -1104.[CrossRef]
Verri, T., Kottra, G., Romano, A., Tiso, N., Peric, M., Maffia, M., Boll, M., Argenton, F., Daniel, H. and Storelli, C. (2003). Molecular and functional characterisation of the zebrafish (Danio rerio) PEPT1-type peptide transporter. FEBS Lett. 549,115 -122.[CrossRef][Medline]
Waterlow, J. (1999). The nature and significance of nutritional adaptation. Eur. J. Clin. Nutr. 53,S2 -S5.
Weatherley, A. H. and Gill, H. S. (1987). The Biology of Fish Growth. London: Academic Press.
Wu, G. and Morris, S. M. (1998). Arginine metabolism: nitric oxide and beyond. Biochem. J. 336, 1-17.[Medline]
Yamamoto, T., Unuma, T. and Akiyama, T. (2000). The influence of dietary protein and fat levels on tissue free amino acid levels of fingerling rainbow trout (Oncorhynchus mykiss). Aquaculture 182,353 -372.[CrossRef]
Yokoyama, M. and Nakazoe, J.-I. (1991). Effects of dietary protein levels on free amino acid and glutathione contents in the tissues of rainbwo trout. Comp. Biochem. Physiol. 99,203 -206.[CrossRef]