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Amino Acid Composition of Human Milk Is Not Unique1'2'3: Nutrient Requirements and Interactions
Amino Acid Composition of Human Milk Is Not Unique1'2'3: Nutrient Requirements and Interactions
of the milks secreted by different species. Human All samples were obtained from animals 10 or
milk is generally recognized as being distinct in its more days after parturition and therefore were con
amino acid composition (Heine et al. 1991); however, sidered "mature" milk samples. None of the samples
this conclusion has arisen principally from com was obtained during the late stage of lactation when
parison of human milk with bovine milk, its common offspring obtain a large proportion of their nutrients
substitute for human infant feeding. The most dis from foods other than milk. Each of the three pig
tinct difference between human and bovine milk milk samples was pooled from several pigs, but all
seems to be the greater concentrations of cystine and other milk samples were obtained from individual
tryptophan relative to the total amino acid concen animals. When possible, nipples of the animals were
tration and lower concentration of methionine cleaned prior to milking. Rats and most nonhuman
relative to the total amino acid concentration in primates were anesthetized prior to milk collection.
human milk. This has been ascribed to the greater a- Oxytocin was administered to some but not all non-
lactalbumin content. We do not know, however, if human primates and to pigs, rats and cats. The off
the amino acid composition of the human milk is spring had suckled just before the milk samples were
indeed unique or whether it is characteristic of the obtained from some animals. Complete evacuation of
mammals within the same phylogenetic group (i.e., the glands was not possible in some instances. In all
the great apes in particular and primates in general) or species except the human, single samples were ob
of slowly growing species, which would include not tained at one milking. Human milk samples were
only the great apes but also such species as the ele obtained from alternate breasts during a 24-h period
phant (McCullagh and Widdowson 1970). Therefore, while the infant suckled on the contralateral breast,
it was our objective to determine the amino acid and then the samples were pooled. All milk samples
(protein-bound plus free) composition of human milk
were frozen, shipped to the laboratory on dry ice, and
and to compare it with the amino acid compositions then frozen at -20°C until analyzed. Human milk
of milks of great apes, lower primates and nonprimate samples were obtained after approval by the Institu
species. tional Review Committees on Human Research of
Baylor College of Medicine and Texas Children's
Hospital. Animal care was in compliance with the
MATERIALS AND METHODS Guide for the Use and Care of Laboratory Animals
(NRC 1985).
Milk collection. Milk was obtained from five Milk analysis. Milk samples were warmed to 37°C
species of primate: human (Homo sapiens, n = 6), and inverted several times to mix. Duplicate aliquots
chimpanzee (Pan troglodytes, n = 5), gorilla (Gorilla (-0.20 mL) were weighed and an equal volume of
gorilla, n = 3), baboon (Papio cynocephalus anubis water was added. Samples were centrifuged at 3000 x
and Papio cynocephalus anubis/Papio cynocephalus, g for 15 min and frozen for 10 min at -70°C, and the
n = 5) and rhesus monkey (Macaca mulatta, n = 6).
upper fat layer was skimmed from the lower frozen
Milk was also obtained from the following ruminant aqueous layer. The skimmed milk was hydrolyzed in
and nonruminant nonprimate species: cow (Bos 4 mL of 6 mol/L HC1 under a blanket of nitrogen at
taurus, n = 4), goat (Capra hircus, n = 2), sheep (Ovis 110°Cfor 24 h. The protein hydrolysates were dried
aries, n = 6), llama (Lama glama, n = 3), pig (Sus
under vacuum (Speedvac, Savant Instruments, Far-
scrofa, n = 3), horse (Equus caballus, n = 8), elephant
(Elephas maximus, n = 3), cat (Felis catus, n = 4) and mingdale, NJ), 1 mL of water was added and evapo
rat (Rattus norvégiens, n = 3). Some of the primate rated two times, and 1 mL of 4.0 mmol/L methionine
sulfone was added as an internal standard. The
milks were purchased from Yerkes Regional Primate protein hydrolysates were filtered through a 0.2-/mi
Research Center, Emory University (Atlanta, GA).
Some of the sheep milk was donated by W. G. Pond filter, and the amino acid compositions were deter
(Children's Nutrition Research Center, Houston, TX). mined.
Horse and cat milks were donated by S. C. Zicker and Amino acid chromatography. Amino acids in the
milk protein hydrolysates were pre-column deriva-
Q. R. Rogers, respectively (University of California,
Davis, CA). Horse, sheep and goat milks were also tized with phenylisothiocyanate and separated on a
PICOTAG reverse-phase column (Waters, Milford,
donated by G. S. Smith (New Mexico State
University, Las Cruces, NM). Elephant milk was do MA). Derivatized amino acids were detected on-line
nated by E. Miller (St. Louis Zoological Park, St. spectrophotometrically and quantified by comparing
Louis, MO), C. L. Wallace (Burnet Park Zoo, Syracuse, the area under the sample peak against that of an
NY) and J. Glazier (Dickerson Park Zoo, Springfield, amino acid standard solution (Pierce H standard,
MO). Llama milk was donated by E. Domatti (Sun Sigma Chemical, St. Louis, MO) of known concen
shine Acres Llamas, Simonton, TX). Pig and bovine tration.
milk was purchased from Texas A&M University Tryptophan is destroyed by acid hydrolysis
(College Station, TX). Rats were purchased from (McKenzie 1970); therefore tryptophan values are not
Charles River Laboratories (Wilmington, MA). reported. Because glutamine was converted to
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1128 DAVIS ET AL.
acids of andnonprimate
greatest abundance in primate branched-chainamino
essential amino acids (EAA) and total
milks1-2Species milks1'2Species
acids (BCAA) in primate and nonprimate
n GlutamateProlinemg
Leucine n EAA
BCAAmg
acid/gtotal
amino acid/gtotal
amino
acidPrimateHuman amino acidPrimateHuman amino
inhorse, milks (P < 0.001). They were lowest forinfant milk is the most common milk source
milks.Although
pig and rat widelyestablished
formula feeding even though it has been
notdiffer total sulfur amino acids in milk did humanand that the amino acid patterns of
milk,which
among species (except for rat milk and cat Renner1983).
bovine milk differ (Heine et al. 1991,
therelative
were rich in total sulfur amino acids), milkprotein
Although discussions in the literature on
tothe contributions of methionine and cystine ofhuman pattern are dominated by a comparison
widelyamong
total sulfur amino acids in milk varied humanmilkand bovine milk and it is implied that
lowermethionine
species (Table 4). Primate milks had hasnot is unique, the uniqueness of human milk
nonprimatemilks
and higher cystine than Thus,our
been established in any systematic way.
ofthe (P < 0.001). Among the primates, the milks thegeneral
principal objective was to determine whether
methioninebut
great apes and humans were lower in fromthe conclusion that human milk is different
oflower
higher in cystine (P < 0.001) than the milks isindeed
milk of other species in its amino acid pattern
themilksprimates, which in turn were similar to besimilarities
valid. We hypothesized that there would
milkwas of the nonprimates. In addition, human aphylogenetic in milk amino acid patterns within
ofthe significantly higher in cystine than the milks comparehuman order. Therefore, we chose to
thehighest
great apes (P < 0.002). However, rat milk had thatwere and bovine milk to milk from species
sampled.Table
cystine content of all the milks human,chimpanzee,
close in the phylogenetic order (i.e.,
uniqueproportions
5 shows the amino acids that were in onone gorilla, baboon and rhesus monkey
(althoughthere in the milks of some species theother).
hand, and cow, goat, sheep and llama on
were no statistically significant differences be acidpattern
We further questioned whether the amino
individualamino
tween primates and nonprimates for these slowgrowthof human milk might be related to the
milk.Serine
acids). Glycine was uniquely high in pig rate of the species; for this reason, we com
inrat and cystine were high and proline was low pared
slow-growingspecies,
human milk with that of another
catbut
milk. Arginine was highest in the milk of the Widdowson1970).
the elephant (McCullagh and
was also high in horse milk.TABLE Other species were chosen because of specific
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1130 DAVIS ET AL.
TABLE 4
Sulfur amino acids in primate and nonprimate milks1'2
sulfur to
SpeciesPrimateHumanChimpanzeeGorillaBaboonRhesusNonprimateCowGoatSheepLlamaPigHorseElephantCatRatn65356426338343Methioninemg16.1
acidsacid36.3
amino cystine
ratio0.81
acid/g
amino20.2
total
0.917.0
± 2.616.2
± 3.333.2
± 0.091.06
±
2.219.8
± 1.715.5
± 3.635.3
± 0.111.28
±
1.721.2
± 1.210.1
± 0.731.2
± 0.202.13
±
1.824.8
± 1.711.7
± 3.236.5
± 0.292.18
±
2.026.3
± 2.58.9
± 4.135.2
± 0.392.97
±
0.925.5
± 0.88.6
± 0.934.1
± 0.302.97
±
2.228.7
± 1.17.5
± 3.336.3
± 0.143.82
±
0.731.1
± 0.57.3
± 0.838.4
± 0.284.29
±
1.021.7
± 0.915.6
± 0.337.3
± 0.661.40
±
0.422.0
± 1.311.3
± 1.433.4
± 0.112.03
±
0.721.8
± 2.310.6
± 2.532.4
± 0.462.36
±
2.732.0
± 3.912.1
± 2.644.0
± 1.282.65
±
0.625.0
± 0.825.7
± 1.250.7
± ±0.170.97
±0.5Cystineamino ±0.3Total ±0.6Methionine ±0.02
'Values for methionine and cystine are means ±so calculated from the sum of each amino acid (in mg| divided by the total amino acids (in
g, excluding tryptophan). Total sulfur amino acids are means ±so of the sum of methionine and cystine values.
^Methionine differed in primates vs. nonprimates, humans and great apes vs. lower primates, and humans vs. elephants |P < 0.001 ).
Cystine differed in primates vs. nonprimates (P < 0.01), humans and great apes vs. lower primates (P < 0.001], humans vs. great apes (P < 0.01),
and humans vs. elephants (P < 0.01). The methionine to cystine ratio differed in primates vs. nonprimates, humans and great apes vs. lower
primates, humans vs. great apes, and humans vs. elephants (P < 0.001).
taxonomic relationships of interest (Jenness 1986, concentration tended to be higher in the milk of those
lenness and Sloan 1970). We chose species born at allowed to suckle their young before sampling (13.9 ±
similar stages of maturity (such as the cow, llama, 0.4 vs. 10.0 ±1.6 g/L), the amino acid pattern of the
horse and pig), species that share similar ecological milk was the same for all baboons (data not shown).
niches (such as the cow and horse), species that nurse The lack of effect of sampling technique on amino
on demand (such as primates and the horse), species acid pattern is also reflected in the small CV for each
that vary in litter size (human and cow vs. rat and species (<10% for all amino acids).
pig), species that as adults have radically different The results of the present study show a remarkable
dietary amino acid patterns (omnivores, herbivores commonality in the general amino acid pattern of
and carnivores), and species that differ in site and milk despite the 10-fold difference in the total amino
extent of digestion (ruminants, nonruminant herbi acid concentrations of the milks that we analyzed.
vores and nonruminants). We further questioned The amino acids in greatest abundance in the milks of
whether the unique amino acid needs of a species all species were glutamate (plus glutamine), leucine
might be reflected in the amino acid pattern of the and proline, which together were 40% of the total
milk of that species. These unique needs include the amino acids in milk. Essential amino acids together
cat's essential requirement for arginine and the were -40%, branched-chain amino acids -20%, and
sheep's, llama's and rat's need to synthesize large sulfur amino acids -4% of the total amino acids in
amounts of a tissue (i.e., hair or wool) with a radically the milks of all species. If one accepts the idea of
different amino acid pattern from that of other coevolution of milk and neonate, these results
tissues. suggest a commonality in the pattern of amino acid
To determine the amino acid pattern of milk requirements of the young of the species that we
samples from a large number of species, different surveyed.
sampling techniques had to be used. However, it Nevertheless, differences in the pattern of in
seems unlikely that the use of different sampling dividual amino acids were observed among species,
techniques influenced the results. For example, and these differences were principally among species
within one species (baboon), two of the five individual of different phylogenetic orders. Thus, the amino acid
animals suckled their young just prior to sampling pattern of human milk did not seem to be unique but
and three did not. Although the total amino acid was similar to those of the milks of the great apes
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AMINO ACID PATTERN OF HUMAN MILK IS NOT UNIQUE 1131
TABLE 5
Unique differences in milk amino acid patterns among species^
SpeciesPrimateHumanChimpanzeeGorillaBaboonRhesusNonprimateCowGoatSheepLlamaPigHorseElephantCatRatn6S356426338343Glycine22
acid/g
total20 acid95
and, to a lesser degree, similar to those of the milks of other individual amino acids, as well as the obser
lower primates. The milk of the cow was most vation that milk from the rat, a rapidly growing
similar to those of the goat and sheep (which are of species, had the highest cystine content in its milk of
the same phylogenetic suborder) and, to a lesser all species surveyed.
degree, was similar to the milk of the llama, which is Although comparison of milk amino acid patterns
of the same order (Artiodactyla) but different sub among species by specific classifications such as stage
order. of maturity at birth, litter size and nursing schedule
The relationship between phylogenetic order and revealed little relationship, there did seem to be some
amino acid pattern of milk is most apparent for the relationship between milk amino acid content and
amino acids cystine and methionine. The primates as the unique amino acid needs of some species. The
a whole had lower methionine and higher cystine high content of arginine that we found in cat milk
contents in their milk compared with the artiodac and that others have found in the milk of the tiger
tyles. The human and the great apes had lower (Bock 1984) may be related to the high arginine re
methionine and higher cystine in their milks than did quirement of felines (Morris 1985). However, milk
the lower primates, and indeed the lower primates from the horse was also relatively high in arginine.
had methionine and cystine contents in their milks Because we (Davis et al. 1993) had previously found
the serine content of rat milk to be higher than
that were more similar to those of the nonprimates
available literature values for the milks of other
than to those of the great apes. Human milk had the
species except the sheep (USDA 1976), we speculated
highest cystine content of all the primate milks. Be
that a high serine content in milk might be related to
cause the requirement for cystine, as a proportion of
the need to synthesize large quantities of hair or
total amino acids, is higher for maintenance than for wool; serine is required for the synthesis of cystine,
growth (Fuller et al. 1989), and because maintenance and there is a proportionally high abundance of
contributes a greater proportion of the nutrient re cystine in hair and wool proteins. However, in the
quirements of slow-growing species, we questioned
current study we found that sheep and llama milks
whether the high cystine content in the milks of the were not serine rich, and thus the rat was unique
human and the great apes might be related to the among species sampled in its high serine content in
slow growth of these species. However, comparison of milk. This high serine and cystine but low proline
the cystine content of milks from the human and the content in rat milk is in agreement with a previously
elephant, another slow-growing species (McCullagh published description of the amino acid composition
and Widdowson 1970), suggests that the amino acid of rat casein (Woodward and Messer 1976). Addi
pattern of human milk is unrelated to growth rate. tionally, pig milk was unique in its high glycine
This conclusion is supported by the lack of close content, consistent with previously reported values
similarity between human and elephant milk for for sow milk (Elliott et al. 1971).
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1132 DAVIS ET AL.
+1§-H -H -H +1 -H +1 -Hl -H +1 +1 -H +1
ACKNOWLEDGMENTS "°°^~^-H
— 22
LITERATURE CITED
-H'uOg +1 +1 -H -H +1 +1 +1 -H -H -H +1 -H -H
National Research Council (1985) Guide for the Care and Use of oo...m.
oo oo oo Is-- Is-- r*-- i^- f^- Is"- ON \Q oo
Laboratory Animals. Publication no. 85-23 (rev.), National Insti
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Renner, E. (1983) Milk and Dairy Products in Human Nutrition. ^c,«««««^«(UW
Volkswirtschaftlicher Verlag, München, Germany.
Snedecor, G. W. & Cochran, W. G. (1967) Statistical Methods. Iowa
State University Press, Ames, IA. 4Jallaga
U.S. Department of Agriculture (1976) Composition of Foods: Dairy
and Egg Products, Raw, Processed, Prepared. Agricultural
Handbook no. 8-1, USD A, Washington, DC.
«1«E. 1 a«
«,§&«*g3j=0-^X
S 'Hoïï I-ISSI
§OO^J3.£fo^««a.
Woodward, D. R. & Messer, M. (1976) Chemical composition of rat —.'•-» 2
casein. Comp. Biochem. Physiol. 55B: 141-143.