Effects of concentration on in vivo absorption of a peptide containing protein hydrolysate - Gut
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Gut: first published as 10.1136/gut.23.4.304 on 1 April 1982. Downloaded from http://gut.bmj.com/ on November 1, 2021 by guest. Protected by copyright.
Gut, 1982, 23, 304-309
Effects of concentration on in vivo absorption of a
peptide containing protein hydrolysate
J E HEGARTY, P D FAIRCLOUGH, K J MORIARTY, M J KELLY, AND M L CLARK
From the Department of Gastroenterology, St. Bartholomew's Hospital, London
SUMMARY Amino acid absorption from a peptide-containing protein hydrolysate and an equiva-
lent amino acid mixture over a range of concentrations of the two-such as is thought to be found in
the normal intestine after a meal'-has been studied using a jejunal perfusion technique in man.
The relative rates of amino acid absorption from the protein hydrolysate and amino acid mixture
varied markedly with concentration, demonstrating that the global hypothesis that peptides confer
an advantage in amino acid absorption in vivo is too simple. There is a highly significant correlation
between amino acid absorption and the concentrations of amino acid in the perfusate, whether this
contained amino acid or protein hydrolysate, suggesting that, under these experimental conditions
at least, the presence of distinct amino acid and peptide transport systems is relatively unimpor-
tant. Doubt is thus cast upon suggestions derived from previous intestinal perfusion experiments
that intact peptide transport may be nutritionally significant in man.
The existence and independence of intestinal trans- as follows. Crude lactalbumin obtained from whey
port systems for amino acids and small peptides in during the commercial processing of milk was hydro-
animals and man has now been demonstrated.2 Intes- lysed with pancreatin. The resulting digest was separ-
tinal perfusion studies in man demonstrating greater ated into peptide fractions by ultrafiltration and the
and more rapid amino acid absorption from four approximate size of the peptides obtained was esti-
different protein hydrolysates containing small pep- mated by gel filtration on columns of Sephadex G10.
tides than from equivalent free amino acid mixtures As estimated by this technique, the preparation con-
have been interpreted as suggesting that the peptide tained 20% free amino acids, 30-35% di-, tri-, and
transport systems might be of nutritional impor- tetrapeptides, 20-30% larger oligopeptides up to
tance.3 4 5 However, each hydrolysate was studied MW 1500D and 10-20% peptides above MW 1500D.
only at a single concentration. As the relative rates It should be borne in mind that, because of the varia-
of absorption of amino acids from single pep- tion in the molecular weight of individual amino
tides and equivalent free amino acids are known to be acids, this distribution is an approximation. An ami-
critically dependent on concentration,6 7 8 the present no acid mixture simulating the amino acid composi-
study was designed to examine in vivo in man the tion of the hydrolysate was also prepared. Individual
effect of concentration on the relative rates of amino amino acid concentrations in solutions of protein
acid absorption from a protein hydrolysate contain- hydrolysate (both free and peptide bound) and ami-
ing a complex mixture of peptides of varying size and no acid are compared in Table 1. The amino acid
an equivalent free amino acid mixture. composition of the preparations was derived from ion
exchange chromatography of solutions of the hy-
Methods drolysate and amino acid mixture as described below.
LACTALBUMIN HYDROLYSATE AND EQUIVALENT PERFUSION TECHNIQUE
FREE AMINO ACID MIXTURE Jejunal absorption of amino acids from protein hy-
The hydrolysate of lactalbumin was prepared by drolysate and amino acids was studied in 18 normal
Nestle Technical Assistance (Lausanne, Switzerland) healthy adult volunteers using the intestinal perfu-
sion technique previously described.8 The study was
approved by the Ethical Committee of St. Bar-
tholomew's Hospital and informed consent was
Received for publication 17 August 1981 obtained from all subjects. Solutions containing pro-
304Gut: first published as 10.1136/gut.23.4.304 on 1 April 1982. Downloaded from http://gut.bmj.com/ on November 1, 2021 by guest. Protected by copyright.
Absorption ofprotein hydrolysate 305
Table 1 Individual amino acid concentration (mmolll) of a to 290-300 MOsm/kg by addition of NaCl and the
solution of lactalbumin hydrolysate and its equivalentfree pH to 7 by addition of 1M NaOH. The sodium con-
amino acid mixture containing 100 mmol (1 32 g) total amino tent of perfusion solutions was measured by flame
acidsll photometry.
Amino Lactalbumin Amino acid
acid hydrolysate mixture ANALYTICAL METHODS AND CALCULATION OF
RESULTS
Asparagine 11-3 11-9
Threonine 8-8 8-4 Aliquots of the perfusion solutions containing amino
Serine 7-1 7-2 acid or protein hydrolysate and their respective intes-
Glutamine 16-5 16-1 tinal aspirates were hydrolysed under reflux condi-
Proline 7-9 7-2
Glycine 3-4 3-7 tions at 110°C for 24 hours with 6 M HCl in the
Alanine 7.5 7-4 presence of norleucine as internal standard to com-
Valine 7.5 7-3
Methionine 1-8 1-8 pensate for hydrolysis losses. After cooling, the ami-
Isoleucine 6-9 6.3 no acid content of the hydrolysed sample was esti-
Leucine 9-6 9-4
Tyrosine 1-9 1-9 mated by ion exchange chromatography using a
Phenylalanine 2-5 2-9 Locarte automatic loading amino acid analyser
Lysine 7-1 6-5 (Locarte Company, London, UK). The 14C PEG
Histidine 1-5 1-5
Arginine 1-9 1-9 content of the test solutions and intestinal aspirates
Individual amino acid concentration:
was measured by liquid scintillation counting.9 Ami-
no acid absorption was calculated using previously
mg/100 ml= Conc (mmol/l) x mol wt amino acid described formulae. 10 Luminal disappearance of ami-
10
no acid residues was taken to be equivalent to absorp-
tein hydrolysate and amino acids were infused at 20 tion. The significance of differences was assessed by
mi/min. After an equilibration time of 30 minutes, the paired t test."
three 10 minute samples were collected from the
distal collecting orifice by simple siphonage into a Results
vessel at 4°C. Samples were stored at -20°C until
required for analysis. Whereas five amino acids (isoleucine, leucine,
Three concentrations of protein hydrolysate and methionine, proline, and arginine) were absorbed to
amino acid mixture containing 40, 70, and 100 mmol/l a greater extent (pGut: first published as 10.1136/gut.23.4.304 on 1 April 1982. Downloaded from http://gut.bmj.com/ on November 1, 2021 by guest. Protected by copyright.
306 Hegarty, Fairclough, Moriarty, Kelly, and Cli'ark
so
a AMINO ACID MIXTURE
S0 = HYDROLYSATE
70
60
AMINO W0
ACID
ADSORPTION
1% of purfud
B )40
30
20
10f
0
MET LEU ILEU PRO ARG VAL PHE ALA TYR LYS GLY SER HIS THR GLU At5P
p NS NS NS NS NS NSGut: first published as 10.1136/gut.23.4.304 on 1 April 1982. Downloaded from http://gut.bmj.com/ on November 1, 2021 by guest. Protected by copyright.
Absorption ofprotein hydrolysate 307
8
7
M AMINO ACID MIXTURE
6- HYDROLYSATE
6~~
AMINO 5
ACID
ADSORPTION
(mMoilsbr/30cm)
4
3
2
MET LEU ILEU PRO ARG VAL PHE ALA TYR LYS GLY SER HIS THR GLU ASP
p NS NS NS NS NS NSGut: first published as 10.1136/gut.23.4.304 on 1 April 1982. Downloaded from http://gut.bmj.com/ on November 1, 2021 by guest. Protected by copyright.
308 Hegarty, Fairclough, Moriarty, Kelly, and Clark
of protein hydrolysates as the concentration of pep- sumably the more important for protein synthesis. In
tides increases and saturation of the free amino acid Fig. 1 amino acid absorption is expressed, firstly, as a
transport system by amino acids released at the brush percentage of the perfused load and in Fig. 2 as actual
border occurs. This explanation must of necessity be amounts of amino acid absorbed. Quite clearly the
tentative as confirmation would require considerably 'pattern' of presentation of amino acids to the tissue
more detailed analysis of amino acid and peptide bears no relationship to the pattern of absorption
transport kinetics than is allowed by the results of the obtained when the results are expressed as a percen-
present studies. tage of the perfused load, but is, in fact, related to the
The importance of the pattern of amino acid composition of the solution perfused (Fig. 3), as pre-
absorption from amino acid mixtures and protein viously demonstrated in vitro by Gardner,16 17 and in
hydrolysates has previously been emphasised.45 Dur- vivo in man by Marrs et al. 18 A similar result was also
ing jejunal perfusion of a number of different protein obtained at 40 and 70 mmol/l total amino acid (results
hydrolysates and their equivalent amino acid not shown). Previous studies quoted in support of the
mixtures3 45 it has been observed that the variation in suggestion that protein hydrolysates may be superior
percentage amino acid absorption from the protein to amino acid mixtures in terms of protein synthesis
hydrolysates was considerably less than that seen emphasised the importance of simultaneity of pre-
with the equivalent amino acid mixtures. It was sug- sentation of amino acids to the tissues for protein
gested that this observation might be of nutritional synhesis'9 rather than that the actual amounts of
importance because of more 'even' presentation of amino acids were importnt in this respect. It is pos-
the amino acids to the tissues for protein synthesis. sible, but unproved, that rapid absorption of some
While expression of the absorption rates of individual amino acids from protein hydrolysates compared
amino acids as a percentage of perfused load is valid with equivalent amino acid mixtures shown in perfu-
and useful for comparing amino acid absorption from sion experiments will confer an advantage in protein
different nitrogen sources, the 'pattern' of absorption synthesis in normal man fed orally with a hy-
so obtained is quite different from the pattern one drolysate. This possibility is the subject of further
sees when absolute amounts of amino acid absorbed investigation.
are considered, because percentage absorption takes Our results illustrate the fundamental importance
no account of the wide variation in concentration of of studying kinetic phenomena over a range of con-
the amino acids in the perfusate-for example, com- centration since results at a single concentration are
pare histidine and glutamine (Table 1). The pattern frequently misleading.20 Although we have shown no
of absolute amounts of amino acid absorbed is pre- significant absorptive advantage due to either prepa-
8-
E
U
0
m
LEU *LEU
PRO*
E
6- 6-
0
E r=081
E I LEU@ pGut: first published as 10.1136/gut.23.4.304 on 1 April 1982. Downloaded from http://gut.bmj.com/ on November 1, 2021 by guest. Protected by copyright.
Absorption ofprotein hydrolysate 309
ration the recommendation that chemically defined electrolytes in the normal human jejunum. Clin Sci Mol
enteral diets should include protein hydrolysates con- Med 1975; 49:401-8.
taining small peptides rather than free amino acids as 8 Sladen GE, Dawson AM. Further studies on the perfu-
the nitrogen source is probably reasonable on the sion method for measuring intestinal absorption in man.
The effects of proximal occlusive balloon and a mixing
grounds of palatability, lower osmostic pressure, and segment. Gut 1970; 11:947-54.
lower cost. Further studies of absorption of amino 9 Wingate DL, Sandberg RJ, Phillips SF. A comparison of
acids, peptides, and whole protein nitrogen sources stable and 14C labelled polyethylene glycol as volume
after meals and estimation of their effects on protein marcators in the human jejunum. Gut 1977; 13:812-5.
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tional case for or against the use of peptides in enteral the absorption of glucose, sodium and water by the
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11 Snedecor GW, Cochrane WG. Statistical methods 6th
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