The protein intake of breastfed term infants has been used as
the basis for estimating an infant’s protein requirements during
the first year. Daily protein gain is highest in the very young
infant and decreases rapidly in later infancy and in the second
year of life. The protein content of breast milk evolves depending
on the stage of lactation and time since delivery. Indeed,
protein concentration in breast milk is high during the first few
weeks of lactation and gradually subsides throughout the first
year. The quantity and quality of breast milk is critical to support
infant growth and long-term development.
Proteins are the third most abundant solids found in breast
milk. The variety of functions performed by the bioactive proteins
and peptides in breast milk shed light on why breastfed
infants have lower morbidity and fewer infections. Lactoferrin,
secretory IgA, osteopontin, and various cytokines modulate
the infant’s immune system alongside lysozyme, ĸ-casein, and
lactoperoxidase, which have antibacterial functions. Other proteins
regulate gut development and aid in the absorption of key
Based on our better understanding of protein evolution in
breastmilk across the stages of lactation, new infant formulas
with lower protein concentration but better protein quality
have been developed, tested, and made available in many
countries. Low-birth-weight infants have higher protein requirements
than term infants because of their higher daily
protein gain per unit body weight. The concentrations of protein
and amino acids in the breast milk of mothers who deliver
preterm are higher during the first weeks of lactation compared
to those of mothers who deliver at term. Supplementation of
breast milk is needed to meet the high protein requirements of
infants with very low and extremely low birth weight.
Lönnerdal B, Erdmann P, Thakkar SK, Sauser J, Destaillats F:
Longitudinal evolution of true protein, amino acids and bioactive
proteins in breast milk: a developmental perspective. J Nutr
Protein intake of breastfed term infants has been used
as a model to estimate protein requirements during
the first year. They are higher during the first months
when daily weight gain is fast and lower during later
infancy when daily weight gain slows down.
Breast milk contains a multitude of bioactive proteins
that are highly concentrated in early lactation and
decrease with progressing lactation.
Quantity and quality of protein in breast milk are
crucial for healthy growth and long-term development.
Protein ingested with breast milk provides indispensable
amino acids which are necessary for new protein synthesis
for growth and replacement of losses via urine, feces, and
the skin. Protein gain in the body of an infant is highest during
the first months when protein concentrations in breast
milk are higher than during later stages of lactation. Low-birth-weight
infants have higher protein needs than term infants
and need protein supplements during feeding with
breastmilk. Based on our better understanding of protein
evolution in breastmilk during the stages of lactation, new
infant formulas with lower protein concentration but better
protein quality have been created, successfully tested, and
are now available in many countries. Besides providing indispensable
amino acids, bioactive protein in breast milk can be
broadly classified into 4 major functions, that is, providing
protection from microbial insults and immune protection,
aiding in digestive functions, gut development, and being
carriers for other nutrients. Individual proteins and their proposed
bioactivities are summarized in this paper in brief. Indeed,
some proteins like lactoferrin and sIgA have been extensively
studied for their biological functions, whereas others
may require more data in support to further validate their
Breastfeeding is important for the healthy growth and
development of infants and young children. The WHO
recommends exclusive breastfeeding until 6 months and
continuation of breastfeeding until 2 years as part of a
mixed diet. However, recent DHS surveys indicate that
even in developing countries only about 32% of mothers
exclusively breastfeed their infants until 6 months ,
and the quality of complementary foods is very low.
Therefore, in many developing countries, stunting is still
prevalent in about 20% of children under 5 years of age
. In most developed countries, solids are introduced
between 4 and 6 months, and breastfeeding is often
stopped much earlier than recommended.
After carbohydrates and lipids, proteins are the third
abundant solids in breast milk (BM), not only providing
crucial amino acids indispensable for growth but also bioactive
proteins and peptides essential for many functions
Protein Needs for Growth
The protein intake of breastfed term infants has been
used as a model to estimate protein requirements during
the first year [3, 4]. The protein content in BM can be
quantified by directly assessing the true protein content
. True protein concentrations of 14–16, 8–10, and 7–8
g/L have been reported during early lactation, at 3–4
months, and at 6 months, respectively. A recent metaanalysis
of 43 studies  confirms that the protein concentration
in BM depends on the stage of lactation and
time since delivery. It also indicates a big variety in protein
concentration, in particular during the first few
months of lactation. However, the true protein intake
does not accurately reflect the amount of utilizable amino
acids to synthesize new body protein because some (bioactive)
BM proteins can be found intact in infant stools
Protein requirements for growth and daily turnover
strongly depend on rates of body protein gain. Daily protein
retention in the body can be calculated by measuring
absorption and excretion. Fomon et al.  published detailed
estimates of protein gain in children during the first
2 years and beyond. Based on total body water (estimating
the extra- and intracellular compartment), total body potassium
(estimating the intracellular compartment where
almost all of the protein is present), and total body calcium
(osseous minerals), all components of fat-free mass
were calculated. Because potassium is the main intracellular
cation, gains at different age ranges allow to estimate
gain of total body protein (Fig. 1). Daily protein gain is
highest in the very young infant and is rapidly decreasing
during later infancy and the second year of life: during the
first months, protein gain is 3 times higher than between
12 and 24 months. Indeed, protein concentration in BM
is high during the first few weeks of lactation and then
continues to decrease throughout the first year, but at
substantially lower rates than those observed in the first
weeks (Fig. 2). Casein and most whey proteins in BM are
utilized for growth. Their concentrations change profoundly
over the course of lactation: during the first 2
weeks of lactation, concentrations of whey proteins are
very high, while concentrations of caseins are low, which
results in a whey:casein ratio as high as 80:20. The ratio
drops to 65:35 by week 2 and stays constant at about
60:40 thereafter .
Proteins are polymer chains made of amino acids
linked together by peptide bonds. During the digestion
process, most proteins are decomposed to simple amino
acids or small peptides which are absorbed. Amino acids
which are absorbed and not oxidized are the building
blocks of new protein which is synthesized in the body.
Despite the reduction in protein over time, the nutritional
value of protein in BM, as measured by the ratio of essential
amino acids to total amino acids, appears to be
consistent over time. These changes correlate well with
the evolving needs of the growing infant .
Because of their higher daily protein gain per unit of
body weight, low-birth-weight infants have higher protein
requirements than term infants [9, 10]. Concentrations
of protein and amino acids during the first weeks of
lactation are higher in BM of mothers who deliver preterm
than in BM of mothers who deliver at term .
However, feeding BM without supplements does not
meet protein requirements, in particular those of verylow-
and extremely-low-birth-weight infants. Supplements
which are on the market are based on protein fractions
of cow’s milk or human donor milk. Supplementation
of BM or the use of preterm formulas improves
growth rates of low-birth-weight infants, but it is questionable
if the right mix of indispensable amino acids for
synthesis of new body protein in low-birth-weight infants
has already been found.
Growth differences between breastfed infants and infants
fed high-protein formulas (i.e. >2.25 g/100 kcal)
have been shown more than 2 decades ago . Infants
fed a formula with high protein content grow faster during
the first 2 years and beyond , and they have higher
insulin and insulin-like growth factor 1 levels in blood
[13, 14]. Rapid growth during the first year is associated
with obesity during childhood. Three longitudinal studies
confirm a strong relationship between weight gain between
0 and 12 months and BMI at 12, 36, and 60 months
(Fig. 3) : 21% of the variance of BMI at 60 months was
explained by weight gain between birth and 12 months.
Narrowing the protein gap between BM and infant formula
requires a deep understanding of how protein quality
and quantity in BM changes over time. Composition
of infant formulas has evolved with increasing knowledge
of BM. Recently, a pooled analysis of individual growth
data (11 clinical trials; n = 1,882 ) of infants who received
a modified whey-based low-protein starter formula
(1.8 g protein/100 kcal)  with an amino acid profile
close to term BM  has become available. The weight
and length of formula-fed infants at 4 months correspond
to the 50th percentile of the WHO global growth standard
 . The CHOP study in Europe followed the growth of
children who had been fed low- or high-protein formulas
during the first year of life. At 6 years, BMI and the percentage
of children with obesity were significantly lower
in the low-protein formula group . Two randomized
controlled trials [19, 20] tested 2 low-protein follow-up
formulas (1.6 g protein/100 kcal) and followed children
until 5 years of age. BMI at 5 years of the children who
had been on the low-protein formula was similar to that
of the children who had been exclusively breastfed until
4–6 months. .
Therefore, it seems that the quantity and quality of
protein in BM are crucial for healthy growth and longterm
development. Some nutritive proteins which are
partly or well absorbed may also have biological functions.
In addition, there are bioactive proteins in BM
which are not absorbed. Specific functions of bioactive
proteins and peptides which have been studied in detail
can provide insights on why breastfed infants have lower
morbidity and shorter infection periods  as well as
different microbiota .
Bioactive (Functional) Proteins in BM
Immunomodulatory and Antimicrobial Activity
Lactoferrin, also known as lactotransferrin, was reported
to be present in bovine milk in the late 1930s and
quantified in BM in the early 1960s . Originally described
as “red protein from (bovine) milk,” it turned out
to be a multifunctional globular glycoprotein [24, 25].
Lactoferrin content of BM decreases with progressing
stages of lactation, found highest in colostrum at 5.5 g/L
and between 1.5 and 3.0 g/L in mature milk depending on
the stage of lactation . It is generally accepted that lactoferrin
resists digestion to some degree and therefore can
be found intact in infant feces. However, early in life, a
fraction of this protein may be taken up by the intestinal
mucosa and the rest is digested to yield bioactive peptides
. Due to its high affinity towards ferric iron, it not
only acts as a carrier of iron in BM, but also deprives
harmful microbes of iron that is key for their growth. Additionally,
due to its basic N-terminal domain that can
interact with various microbial and host targets, lactoferrin
not only has antimicrobial properties but also modulates
the innate and adaptive immune responses. This is
orchestrated by cytokines interleukin (IL)-4, IL-2, IL-12,
and interferon-γ and results in its ability to act as an antior
a proinflammatory agent. It has been demonstrated
that lactoferrin can compete with lipopolysaccharide for
binding to CD14 and thereby preventing lipopolysaccharide-
mediated proinflammatory series of events . Although
this molecule is resilient to digestion and can be
found intact in fecal material, it is digested to some extent
to form lactoferricin, a molecule that is able to inhibit
Escherichia coli attachment to intestinal cells [29, 30].
Lactoferricin may not be the only lactoferrin related peptide
to have antimicrobial activity, the role of Lf(1–11)
and lactoferrampin has also emerged in the recent past
. In one study, Lf(1–11) was demonstrated to be active
against gram-positive bacteria (Staphylococcus spp.
and Streptococcus mitis) as well as gram-negative bacteria
(Acinetobacter baumannii, Pseudomonas spp., Klebsiella
spp., and E. coli) .
Secretory Immunoglobulin A
Mothers’ milk is rich in secretory immunoglobulin A
(sIgA) especially during early stages of lactation. Colostrum
shows a wide variability from mother to mother and
may contain on average 2.0 g/L of sIgA, which is reduced
to approximately 0.5 g/L in mature milk . The absorptive
fate of this protein is similar to that of lactoferrin in
that it is partly excreted intact and partly digested to bioactive
peptides  . This class of antibody has been documented
to be abundantly present in intestinal mucosa of
humans and other mammals and to protect the epithelium
from toxic assaults. As a first line of defense, in inhibiting
infectious incident, sIgA would block toxin adhering
to intestinal epithelium. In mouse models exposed to
Vibrio cholerae toxin, sIgA demonstrated a protective effect
. Another mechanism by which sIgA could block
pathogens is by direct recognition of receptor-binding
domains like reovirus type 1 Lang. When IgA knockout
mice were challenged with reovirus, the orally gavaged
IgA group of knockout mice was as effective as wild-type
mice in clearing the infection . sIgA may also have a
direct effect on the virulence of the bacteria. For example,
murine monoclonal IgA-binding Shigella flexneri suppressed
activity of the bacterial type 3 secretion (T3S),
necessary for S. flexneri to gain entry into intestinal epithelium
. Immune exclusion is often referred to as
sIgA’s ability to prevent pathogen access to intestinal epithelium
through a series of processes involving agglutination,
entrapment in mucus, and clearance via peristaltic
Osteopontin is a multifunctional, heavily glycosylated
and phosphorylated acidic protein with possible roles in
immune activation, inhibition of ectopic calcification,
cellular adhesion and migration, angiogenesis, and bone
remodeling . With high variability amongst populations
and stages of lactation, the average concentration of
osteopontin in BM is approximately 140 mg/L . When
compared to wild-type suckling mice, osteopontin knockout
suckling mice suffered from prolonged periods and
intense bouts of diarrhea upon rotavirus exposure . A
fine balance between T helper 1 (Th1) and T helper 2
(Th2) is required to alleviate an immune response. Osteopontin
has been demonstrated to induce Th1 expression
and inhibit Th2 along with IL-10 . Interestingly,
breastfed infants but not formula-fed infants showed induction
of Th1 response when immunized with measles,
mumps and rubella . This observation can presumably
be tied to presence of osteopontin in BM of the breast
fed group but not the formula fed group. Furthermore,
through electrostatic interactions, osteopontin can form
complexes with lactoferrin and thereby acts as a carrier
for other immunomodulatory proteins to further enhance
the immune competency of its consumers . Recently,
a randomized controlled trial was carried out
where 2 groups of formula-fed infants and a group of
breast-fed infants were recruited. The 2 formula-fed
groups were fed a standard formula with 65 mg/L bovinederived
osteopontin or an experimental formula containing
130 mg/L bovine-derived osteopontin. Apart from
comparable growth parameters, differences were observed
in lowered levels of proinflammatory cytokine
TNF-α and a significantly fewer number of days when the
infants had fever .
The effect of cytokines to regulate inflammatory processes
often associated with infection is usually like an
orchestra, operating in network and produces a cascading
effect. Cytokines are postulated to enhance proliferation
of thymocytes, inhibit IL-2 production from T-cells and
suppress IgE production [44–46]. The presence of several
cytokines in BM has been demonstrated over years. These
molecules include, but may not be limited to, IL-1β, IL-6,
IL-8, IL-10, TNF-α, interferon-γ, transforming growth
factor-β, and colony-stimulating factor [47–52]. Usually
they are present at very low concentrations (picograms)
and potentially may originate from epithelial mammary
gland cells, or activated macrophages and other cells in
BM . The biological function of these agents on infants
is to complement infants’ own source of cytokines
that are produced at lower quantities due to immaturity
of the immune system. Although cytokines are not as well
studied as other immunomodulatory agents described in
this section, it is postulated that these molecules balance
Th1 and Th2 to impart immunity-related benefits .
Lysozyme belongs to the whey class of protein fraction
in BM and possesses bactericidal properties by affecting
the cell wall of most of gram-positive and some gramnegative
bacteria. Higher amounts of lysozyme have been
observed in colostrum at approximately 0.36 g/L that is
reduced slightly in mature milk to 0.30 g/L . Attempts
have also been made to produce recombinant human lysozyme
and lactoferrin in dairy animals  . The mechanism
is yet unclear, but lysozyme of BM origin also contains
activity against HIV type 1 .
κ-Casein belongs to the casein family of phosphoproteins
that is involved in a number of physiological processes.
With an average concentration of approximately
1.25 g/L in colostrum and transition milk, it settles closer
to 1 g/L in mature milk . These glycosylated κ-casein
of human origin as opposed to bovine origin inhibit the
cell lineage-specific adhesion of Helicobacter pylori to human
gastric surface mucous cells .
A member of the heme peroxidase family, lactoperoxidase
is secreted by mammary glands and is persistently
present during lactation. In human BM, lactoperoxidase
is found at 1–1.5 units/mL during the first 6 months
of lactation . It is well accepted that this peroxidase
catalyzes oxidation of thiocyanate from the saliva of infants
to hypothiocyanate in the presence of small amounts
of hydrogen peroxides already in the mouth of the baby.
The formed hypothiocyanate may be responsible to exterminate
gram-positive and gram-negative bacteria
Haptocorrin is a vitamin B 12 -binding protein found in
many body fluids including BM with a concentration
range of approximately 5 μg/
mL in colostrum to 3 μg/mL
in mature milk . Structurally,
haptocorrin did not show
much alternations after exposure
to digestive enzymes and
was able to inhibit the growth
of E. coli in an in vitro system
. Further systematic study
of exposure of haptocorrin to
34 commensal and pathogenic bacteria indicated suppression
of only Bifidobacterium breve implicating its role
that may be limited to certain strains and a blanket antimicrobial
label might not be relevant for this protein warranting
further studies .
A well-characterized and primary protein in BM, α-
lactalbumin is made up of 123 amino acids and 4 disulfide
bridges and accounts for 20–25% of total BM proteins
[66, 67]. Since it is also a rich source of many indispensable
amino acids, a fraction of the protein is digested well
and the rest yields polypeptides that exert antimicrobial
activities mostly against gram-positive bacteria and not
gram-negative bacteria [68, 69]. Additionally, a folding
variant of α-lactalbumin was also found to be bactericidal
against an antibiotic resistant strain of Streptococcus
pneumonia . Not only for its antimicrobial benefits
but also to mimic the BM closer for additional benefits,
all efforts have been made to enrich BM substitute with
Bile Salt-Stimulated Lipase
The major source of energy for breast-fed infants is the
predominant form of lipid in BM, the triacylglycerols.
Milk triacylglycerols are efficiently digested by complementary
actions of gastric lipase, colipase-dependent
pancreatic lipase, and bile salt-stimulated lipase (BSSL).
While there are 2 sources of these enzymes, infants’ exocrine
pancreas, the major source is maternal milk BSSL.
In the early 1950s, it was first demonstrated by Freudenberg
that mothers’ milk contains an inactive lipase that is
stimulated when the chyme reaches duodenum and
comes in contact with bile salts [72, 73]. BSSL was purified
and characterized in the early 1980s , is demonstrated
to have a broad substrate specificity [75–77] and
to inactivate by pasteurization of BM . Therefore, digestion
and absorption of lipids is significantly lower
when pasteurized donor milk is fed to preterm infants
. Recently in a randomized
phase 3 study, recombinant
human BSSL was added
to infant formula to assess if
it had any impact on growth
velocity, presumably due to
better lipid digestion and absorption.
benefits on growth were not
observed in appropriate-forgestational-
age preterm infants but were present in smallfor-
gestational-age preterm infants .
In absence of pancreatic amylase, BM amylase may
catalyze hydrolysis of starch, glycogen and other related
saccharides by cleavage of α-1,4 linkages to produce maltose,
dextrins, and glucose. Activity of amylase varies from
1,000 to 5,000 units per liter of BM . Colostrum is
known to contain higher activities compared to transition
or mature milk . There is a further decrease of approximately
35% of the activity beyond the first trimester
of breastfeeding . Additionally, higher parity may
also reduce amylase activity by half . Preterm milk
contains equal amounts of amylase activity as term milk
. Apart from its obvious digestion-aiding role, amylase
may also act as antibacterial by attacking the polysaccharides
of the bacterial cell wall .
Thy physiological role of protease inhibitors like α1-
antitrypsin (A1AT) in BM is not completely understood.
However, as generally accepted for other mammals, protease
inhibitors may play a role in digestion and/or absorption
of bioactive proteins present at relatively higher
concentrations in colostrum. Indeed, McGilligan et al.
 showed the highest presence of A1AT in colostrum
(1.4–5.2 g/L) compared to the first 26 weeks (0.07 g/L) or
26–52 weeks of lactation (0.05 g/L). A1AT resists digestion
in the enteral tract and can be found intact in feces
of infants in significant quantities . Efforts have been
made to express human A1AT in transgenic sheep for
potential human applications [87, 88].
Growth factors, their concentrations in BM, and biological
sources of growth factors have been described in the literature
. Potentially originating from epithelial and stromal
cells as well as from macrophages of mammary glands,
the growth factors are present at microgram-per-liter quantities
in BM. Growth factors that are present in the intestinal
lumen, such as epidermal growth factor and insulin-like
growth factors 1 and 2, originate either from salivary glands
of the infants or from mothers’ milk . It still remains unclear
how they are able to exert effects upon their receptors
that are located on the basolateral side of the absorptive intestinal
epithelial cells. Indeed, one suggestion is that the immature
gut of the infant provides access of the ligands to the
basolateral compartment. Preterm infants, whose gut is relatively
underdeveloped compared to that of term infants,
may have significantly higher concentrations of epidermal
growth factor in milk secreted by their mothers .
Lactoferrin exposure to intestinal cell culture models
shows increased proliferation and differentiation in a
dose-dependent manner  . Additionally, it also has
greater proliferation of intestinal crypt cells in a piglet
model  . Indeed, it is plausible that rapid maturation
of absorptive intestinal epithelia in presence of lactoferrin
may contribute to the higher weight gain in infants fed
BM substitute with lactoferrin compared to a control
group without lactoferrin .
Carriers for Other Nutrients
Iron absorption in breastfed infant was reported to be
more efficient than cow’s milk-based infant formula .
Certainly, this was later attributed to thepresence of relatively
high levels of lactoferrin in BM compared to bovine
milk (approx. 1 mg/mL and 10 μg/mL, respectively) and
the majority of the iron in milk is bound to lactoferrin [25,
26]. Furthermore, a receptor of lactoferrin was later identified
that had greater affinity for human lactoferrin in
contrast to bovine lactoferrin for iron absorption .
Efforts are underway to express recombinant human lactoferrin
in rice, and a comparison of the stability and bioactivity
has shown promising results for its potential use
in BM substitutes .
Haptocorrin, also known as transcobalamin 1, perhaps
a name derived from “transporter of cobalamin,” an alternative
name for vitamin B 12, is a binding protein found
in BM . In adults, vitamin B 12 absorption is dependent
on digestive juices, enzymes, binding protein secreted
in stomach, intrinsic factors, and their receptors in the
small intestine . However, in infants, very low
amounts of intrinsic factors have been found in fecal material,
perhaps indicating that haptocorrin may have a
larger role to play in the transport of vitamin B 12 .
Identified in the late 1960s, folate-binding protein
(FBP) almost entirely binds all naturally occurring folate
in BM as well as bovine milk [101, 102]. Since its discovery,
it has been thought that FBP sequesters various forms
of folate and ensures adequate supply to the neonate by
also preventing the oxidation in the digestive tract [103,
104]. Solids of pooled human and bovine milk contained
approximately 2,000 nmol/kg, whereas goat milk contains
twice as much and freeze-drying or spray-drying of
milk to powder retains practically all FBP [105, 106].
Since FBP is able to withstand digestion, it is plausible
that permeable intestinal lining of the infant gut is able to
take up the folate-FBP complex at least for weeks or even
months postpartum until the tight junctions are formed
Originally proposed to carry divalent cations like calcium
and zinc , it did show higher absorption in infant
rhesus monkeys . However, research work in
human infants did not show any signs of altered absorption
of iron α-lactalbumin-enriched infant formula ,
warranting further studies for this biological benefit.
The total casein concentration increases during lactation
and represents approximately 10–20% during earlier
stages and 40–50% when lactation matures [109, 110]. In
mature milk, β-casein may represent up to 25% or approximately
2.7 g/L in BM . This protein is highly
phosphorylated, which at least in preclinical model has
shown to solubilize calcium and uptake by intestinal cells
at least in part by forming casein phosphopeptides that
may act as calcium ionophores or calcium carriers across
the membrane [111, 112]. More research remains to be
done to elucidate the role of casein phosphopeptides in
enhanced uptake of other divalent cations like zinc and
Recent findings on nutritive and bioactive proteins in
breastmilk support the WHO recommendations that
breastfeeding should be continued during the first year
and beyond. Infant formula manufacturers should eliminate
all high-protein formulas from the market. New formulas
for infants should be low in protein, in particular
follow-up formulas and growing-up milks. Protein quality
in formulas (amino acid profiles) should be closer to
that in BM. Before bioactive proteins are added to infant
formulas, safety and efficacy tests must be provided by
F.H. is a board member of the Nestlé Nutrition Institute, a nonprofit-
making Swiss association which receives educational grants
from Nestec S.A., Switzerland, and other sources. S.K.T. is an employee
of Nestlé Research Center, Lausanne, Switzerland.
Haschke F, et al: Feeding patterns during the
first 2 years and health outcome. Ann Nutr
Metab 2013; 62(suppl 3):16–25.
Global Burden of Disease Study 2013 Collaborators:
Global, regional, and national incidence,
prevalence, and years lived with disability
for 301 acute and chronic diseases and
injuries in 188 countries, 1990–2013: a systematic
analysis for the Global Burden of
Disease Study 2013. Lancet 2015; 386: 743–
Fomon SJ: Requirements and recommended
dietary intakes of protein during infancy. Pediatr
Res 1991; 30: 391–395.
Heinig MJ, et al: Energy and protein intakes
of breast-fed and formula-fed infants during
the first year of life and their association with
growth velocity: the DARLING Study. Am J
Clin Nutr 1993; 58: 152–161.
Lönnerdal B, Erdmann P, Thakkar SK, Sauser
J, Destaillats F: Longitudinal evolution of
true protein, amino acids and bioactive proteins
in breast milk: a developmental perspective.
J Nutr Biochem 2016; 41: 1–11.
Donovan SM, et al: Partition of nitrogen intake
and excretion in low-birth-weight infants.
Am J Dis Child 1989; 143: 1485–1491.
Fomon SJ, et al: Body composition of reference
children from birth to age 10 years. Am
J Clin Nutr 1982; 35(suppl):1169–1175.
Zhang Z, et al: Amino acid profiles in term
and preterm human milk through lactation:
a systematic review. Nutrients 2013; 5: 4800–
Ziegler EE, et al: Body composition of the reference
fetus. Growth 1976; 40: 329–341.
Haschke F, Grathwohl D, Haiden N: Metabolic
programming: effects of early nutrition
on growth, metabolism and body composition.
Nestle Nutr Inst Workshop Ser 2016; 86:
Dewey KG, et al: Breast-fed infants are leaner
than formula-fed infants at 1 y of age: the
DARLING study. Am J Clin Nutr 1993; 57:
Weber M, et al: Lower protein content in infant
formula reduces BMI and obesity risk at
school age: follow-up of a randomized trial.
Am J Clin Nutr 2014; 99: 1041–1051.
Axelsson IE, Ivarsson SA, Raiha NC: Protein
intake in early infancy: effects on plasma
amino acid concentrations, insulin metabolism,
and growth. Pediatr Res 1989; 26: 614–
Socha P, et al: Milk protein intake, the metabolic-
endocrine response, and growth in infancy:
data from a randomized clinical trial.
Am J Clin Nutr 2011; 94(suppl):1776S–1784S.
Alexander DD, et al: Growth of infants consuming
whey-predominant term infant formulas
with a protein content of 1.8 g/100
kcal: a multicenter pooled analysis of individual
participant data. Am J Clin Nutr 2016;
Raiha NC, et al: Whey predominant, whey
modified infant formula with protein/energy
ratio of 1.8 g/100 kcal: adequate and safe
for term infants from birth to four months. J
Pediatr Gastroenterol Nutr 2002; 35: 275–
Haschke F, et al: Postnatal high protein intake
can contribute to accelerated weight
gain of infants and increased obesity risk.
Nestle Nutr Inst Workshop Ser 2016; 85: 101–
WHO: WHO Child Growth Standards:
Weight-for-Length, Weight-for-Height and
Body Mass Index-for-Age, Methods and Development.
Geneva, WHO, 2016.
Inostroza J, et al: Low-protein formula slows
weight gain in infants of overweight mothers.
J Pediatr Gastroenterol Nutr 2014; 59: 70–
Ziegler EE, et al: Adequacy of infant formula
with protein content of 1.6 g/100 kcal for infants
between 3 and 12 months. J Pediatr
Gastroenterol Nutr 2015; 61: 596–603.
Dewey KG, Heinig MJ, Nommsen-Rivers
LA: Differences in morbidity between
breast-fed and formula-fed infants. J Pediatr
1995; 126: 696–702.
Isolauri E: Development of healthy gut microbiota
early in life. J Paediatr Child Health
2012; 48(suppl 3):1–6.
Blanc B, Bujard E, Mauron J: The amino acid
composition of human and bovine lactotransferrins.
Experientia 1963; 19: 299–301.
Siqueiros-Cendon T, et al: Immunomodulatory
effects of lactoferrin. Acta Pharmacol
Sin 2014; 35: 557–566.
Lonnerdal B, Iyer S: Lactoferrin: molecular
structure and biological function. Annu Rev
Nutr 1995; 15: 93–110.
Rai D, et al: Longitudinal changes in lactoferrin
concentrations in human milk: a global
systematic review. Crit Rev Food Sci Nutr
2014; 54: 1539–1547.
Lonnerdal B: Bioactive proteins in breast milk.
J Paediatr Child Health 2013; 49(suppl 1):1–7.
Actor JK, Hwang SA, Kruzel ML: Lactoferrin
as a natural immune modulator. Curr Pharm
Des 2009; 15: 1956–1973.
Tomita M, et al: Potent antibacterial peptides
generated by pepsin digestion of bovine lactoferrin.
J Dairy Sci 1991; 74: 4137–4142.
Edde L, et al: Lactoferrin protects neonatal
rats from gut-related systemic infection. Am
J Physiol Gastrointest Liver Physiol 2001;
Bruni N, et al: Antimicrobial activity of lactoferrin-
related peptides and applications in
human and veterinary medicine. Molecules
Brouwer CP, Rahman M, Welling MM: Discovery
and development of a synthetic peptide
derived from lactoferrin for clinical use.
Peptides 2011; 32: 1953–1963.
Goldman AS, et al: Immunologic factors in
human milk during the first year of lactation.
J Pediatr 1982; 100: 563–567.
Lycke N, et al: Lack of J chain inhibits the
transport of gut IgA and abrogates the development
of intestinal antitoxic protection. J
Immunol 1999; 163: 913–919.
Silvey KJ, et al: Role of immunoglobulin A in
protection against reovirus entry into Murine
Peyer’s patches. J Virol 2001; 75: 10870–
Forbes SJ, et al: Transient suppression of Shigella
flexneri type 3 secretion by a protective
O-antigen-specific monoclonal IgA. MBio
Stokes CR, Soothill JF, Turner MW: Immune
exclusion is a function of IgA. Nature 1975;
Ashkar S, et al: Eta-1 (osteopontin): an early
component of type-1 (cell-mediated) immunity.
Science 2000; 287: 860–864.
Schack L, et al: Considerable variation in the
concentration of osteopontin in human
milk, bovine milk, and infant formulas. J
Dairy Sci 2009; 92: 5378–5385.
Maeno Y, et al: Effect of osteopontin on diarrhea
duration and innate immunity in suckling
mice infected with a murine rotavirus.
Viral Immunol 2009; 22: 139–144.
Pabst HF, et al: Differential modulation of
the immune response by breast- or formulafeeding
of infants. Acta Paediatr 1997; 86:
Azuma N, Maeta A, Fukuchi K, Kanno C: A
rapid method for purifying osteopontin
from bovine milk and interaction between
osteopontin and other milk proteins. Int
Dairy J 2006; 16: 370–378.
Lonnerdal B, et al: Growth, nutrition, and
cytokine response of breast-fed infants and
infants fed formula with added bovine osteopontin.
J Pediatr Gastroenterol Nutr 2016; 62:
Soder O: Isolation of interleukin-1 from human
milk. Int Arch Allergy Appl Immunol
1987; 83: 19–23.
Hooton JW, et al: Human colostrum contains
an activity that inhibits the production
of IL-2. Clin Exp Immunol 1991; 86: 520–524.
Sarfati M, et al: Presence of IgE suppressor
factors in human colostrum. Eur J Immunol
1986; 16: 1005–1008.
Munoz C, et al: Interleukin-1 beta in human
colostrum. Res Immunol 1990; 141: 505–513.
Rudloff HE, et al: Tumor necrosis factor-alpha
in human milk. Pediatr Res 1992; 31: 29–
Saito S, et al: Detection of IL-6 in human
milk and its involvement in IgA production.
J Reprod Immunol 1991; 20: 267–276.
Garofalo R, et al: Interleukin-10 in human
milk. Pediatr Res 1995; 37: 444–449.
Eglinton BA, Roberton DM, Cummins AG:
Phenotype of T cells, their soluble receptor
levels, and cytokine profile of human breast
milk. Immunol Cell Biol 1994; 72: 306–313.
Grosvenor CE, Picciano MF, Baumrucker
CR: Hormones and growth factors in milk.
Endocr Rev 1993; 14: 710–728.
Lonnerdal B: Human milk proteins: key
components for the biological activity of human
milk. Adv Exp Med Biol 2004; 554: 11–
Lonnerdal B: Nutritional and physiologic
significance of human milk proteins. Am J
Clin Nutr 2003; 77: 1537S–1543S.
Golinelli LP, Del Aguila EM, Flosi Paschoalin
VM, Silva JT, Conte-Junior CA: Functional
aspect of colostrum and whey proteins
in human milk. J Hum Nutr Food Sci 2014; 2:
Cooper CA, Maga EA, Murray JD: Production
of human lactoferrin and lysozyme in
the milk of transgenic dairy animals: past,
present, and future. Transgenic Res 2015; 24:
Lee-Huang S, et al: Lysozyme and RNases as
anti-HIV components in beta-core preparations
of human chorionic gonadotropin.
Proc Natl Acad Sci USA 1999; 96: 2678–2681.
Cuilliere ML, et al: Changes in the kappacasein
and beta-casein concentrations in human
milk during lactation. J Clin Lab Anal
1999; 13: 213–218.
Stromqvist M, et al: Human milk kappa-casein
and inhibition of Helicobacter pylori adhesion
to human gastric mucosa. J Pediatr
Gastroenterol Nutr 1995; 21: 288–296.
Shin K, Tomita M, Lonnerdal B: Identification
of lactoperoxidase in mature human
milk. J Nutr Biochem 2000; 11: 94–102.
Bjorck L, et al: Antibacterial activity of the
lactoperoxidase system in milk against pseudomonads
and other gram-negative bacteria.
Appl Microbiol 1975; 30: 199–204.
Aoki,Y, Kobayashi K, Kajii T: Enzyme-linked
immunoassay of haptocorrin: analysis of
milk and granulocytes. Biochem Med Metab
Biol 1992; 47: 189–194.
Adkins Y, Lonnerdal B: Potential host-defense
role of a human milk vitamin B-
12-binding protein, haptocorrin, in the gastrointestinal
tract of breastfed infants, as assessed
with porcine haptocorrin in vitro. Am
J Clin Nutr 2003; 77: 1234–1240.
Jensen HR, et al: Effect of the vitamin B12-
binding protein haptocorrin present in human
milk on a panel of commensal and
pathogenic bacteria. BMC Res Notes 2011; 4:
Lonnerdal B, Lien EL: Nutritional and physiologic
significance of alpha-lactalbumin in
infants. Nutr Rev 2003; 61: 295–305.
Permyakov EA, Berliner LJ: Alpha-lactalbumin:
structure and function. FEBS Lett
2000; 473: 269–274.
Pellegrini A, et al: Isolation and identification
of three bactericidal domains in the bovine
alpha-lactalbumin molecule. Biochim
Biophys Acta 1999; 1426: 439–448.
Wada Y, Lonnerdal B: Bioactive peptides derived
from human milk proteins – mechanisms
of action. J Nutr Biochem 2014; 25:
Hakansson A, et al: A folding variant of alpha-
lactalbumin with bactericidal activity
against Streptococcus pneumoniae . Mol Microbiol
2000; 35: 589–600.
Lien EL: Infant formulas with increased concentrations
of alpha-lactalbumin. Am J Clin
Nutr 2003; 77: 1555S–1558S.
Blackberg L, et al: The bile salt-stimulated lipase
in human milk is an evolutionary newcomer
derived from a non-milk protein.
FEBS Lett 1980; 112: 51–54.
Freudenberg E: Lipase of human milk; studies
on its enzymological and nutritional significance.
Bibl Paediatr 1953; 9: 1–68.
Blackberg L, Hernell O: The bile-salt-stimulated
lipase in human milk. Purification and
characterization. Eur J Biochem 1981; 116:
Blackberg L, et al: Bile salt-stimulated lipase
in human milk and carboxyl ester hydrolase
in pancreatic juice: are they identical enzymes?
FEBS Lett 1981; 136: 284–288.
Hernell O, Blackberg L: Digestion of human
milk lipids: physiologic significance of sn-2
monoacylglycerol hydrolysis by bile saltstimulated
lipase. Pediatr Res 1982; 16: 882–
Lindquist S, Hernell O: Lipid digestion and
absorption in early life: an update. Curr
Opin Clin Nutr Metab Care 2010; 13: 314–
Fredrikzon B, et al: Bile salt-stimulated lipase
in human milk: evidence of activity in
vivo and of a role in the digestion of milk
retinol esters. Pediatr Res 1978; 12: 1048–
Andersson Y, et al: Pasteurization of mother’s
own milk reduces fat absorption and
growth in preterm infants. Acta Paediatr
2007; 96: 1445–1449.
Casper C, et al: Recombinant bile salt-stimulated
lipase in preterm infant feeding: a
randomized phase 3 study. PLoS One 2016;
Heitlinger LA, et al: Mammary amylase: a
possible alternate pathway of carbohydrate
digestion in infancy. Pediatr Res 1983; 17: 15–
Lindberg T, Skude G: Amylase in human
milk. Pediatrics 1982; 70: 235–238.
Dewit O, Dibba B, Prentice A: Breast-milk
amylase activity in English and Gambian
mothers: effects of prolonged lactation, maternal
parity, and individual variations. Pediatr
Res 1990; 28: 502–506.
Hegardt P, et al: Amylase in human milk
from mothers of preterm and term infants. J
Pediatr Gastroenterol Nutr 1984; 3: 563–566.
McGilligan KM, Thomas DW, Eckhert CD:
Alpha-1-antitrypsin concentration in human
milk. Pediatr Res 1987; 22: 268–270.
Davidson LA, Lonnerdal B: Fecal alpha 1-antitrypsin
in breast-fed infants is derived from
human milk and is not indicative of enteric
protein loss. Acta Paediatr Scand 1990; 79:
Wright G, et al: High level expression of active
human alpha-1-antitrypsin in the milk
of transgenic sheep. Biotechnology (NY)
1991; 9: 830–834.
Carver A, et al: Expression of human alpha 1
antitrypsin in transgenic sheep. Cytotechnology
1992; 9: 77–84.
Donovan SM, Odle J: Growth factors in milk
as mediators of infant development. Annu
Rev Nutr 1994; 14: 147–167.
Playford RJ, Macdonald CE, Johnson WS:
Colostrum and milk-derived peptide growth
factors for the treatment of gastrointestinal
disorders. Am J Clin Nutr 2000; 72: 5–14.
Dvorak B, et al: Increased epidermal growth
factor levels in human milk of mothers with
extremely premature infants. Pediatr Res
2003; 54: 15–19.
Buccigrossi V, et al: Lactoferrin induces
concentration-dependent functional modulation
of intestinal proliferation and differentiation.
Pediatr Res 2007; 61: 410–414.
Reznikov EA, et al: Dietary bovine lactoferrin
increases intestinal cell proliferation in
neonatal piglets. J Nutr 2014; 144: 1401–
Hernell O, Lonnerdal B: Iron status of infants
fed low-iron formula: no effect of added
bovine lactoferrin or nucleotides. Am J
Clin Nutr 2002; 76: 858–864.
Saarinen UM, Siimes MA, Dallman PR:
Iron absorption in infants: high bioavailability
of breast milk iron as indicated by
the extrinsic tag method of iron absorption
and by the concentration of serum ferritin.
J Pediatr 1977; 91: 36–39.
Kawakami H, Lonnerdal B: Isolation and
function of a receptor for human lactoferrin
in human fetal intestinal brush-border
membranes. Am J Physiol 1991; 261:G841–
Suzuki YA, et al: Expression, characterization,
and biologic activity of recombinant
human lactoferrin in rice. J Pediatr Gastroenterol
Nutr 2003; 36: 190–199.
Burger RL, Allen RH: Characterization of
vitamin B12-binding proteins isolated
from human milk and saliva by affinity
chromatography. J Biol Chem 1974; 249:
Seetharam B, Alpers DH: Absorption and
transport of cobalamin (vitamin B12).
Annu Rev Nutr 1982; 2: 343–369.
Adkins Y, Lonnerdal B: Mechanisms of vitamin
B(12) absorption in breast-fed infants.
J Pediatr Gastroenterol Nutr 2002; 35:
Nygren-Babol L, Jagerstad M: Folate-binding
protein in milk: a review of biochemistry,
physiology, and analytical methods.
Crit Rev Food Sci Nutr 2012; 52: 410–425.
Ford JE, Salter DN, Scott KJ: A folate-protein
complex in cow’s milk. Proc Nutr Soc
1969; 28: 39A–40A.
Ford JE: Observations on the possible nutritional
significance of vitamin-binding proteins
in milk. Proc Nutr Soc 1974; 33: 15A.
Ford JE: Some observations on the possible
nutritional significance of vitamin B12-and
folate-binding proteins in milk. Br J Nutr
1974; 31: 243–257.
Wigertz K, Svensson UK, Jagerstad M: Folate
and folate-binding protein content in
dairy products. J Dairy Res 1997; 64: 239–
Indyk HE, Filonzi EL, Gapper LW: Determination
of minor proteins of bovine milk
and colostrum by optical biosensor analysis.
J AOAC Int 2006; 89: 898–902.
Kelleher SL, et al: Glycomacropeptide and
alpha-lactalbumin supplementation of infant
formula affects growth and nutritional
status in infant rhesus monkeys. Am J Clin
Nutr 2003; 77: 1261–1268.
Szymlek-Gay EA, et al: α-Lactalbumin and
casein-glycomacropeptide do not affect
iron absorption from formula in healthy
term infants. J Nutr 2012; 142: 1226–1231.
Kunz C, Lonnerdal B: Re-evaluation of the
whey protein/casein ratio of human milk.
Acta Paediatr 1992; 81: 107–112.
Kunz C, Lonnerdal B: Human-milk proteins:
analysis of casein and casein subunits
by anion-exchange chromatography, gel
electrophoresis, and specific staining
methods. Am J Clin Nutr 1990; 51: 37–46.
Sato R, Noguchi T, Naito H: Casein phosphopeptide
(CPP) enhances calcium absorption
from the ligated segment of rat
small intestine. J Nutr Sci Vitaminol (Tokyo)
1986; 32: 67–76.
Ferraretto A, et al: Casein phosphopeptides
influence calcium uptake by cultured human
intestinal HT-29 tumor cells. J Nutr
2001; 131: 1655–1661.
Liu KY, et al: Natural killer cell populations
and cytotoxic activity in pigs fed mother’s
milk, formula, or formula supplemented
with bovine lactoferrin. Pediatr Res 2013;
Hurley WL, Theil PK: Perspectives on immunoglobulins
in colostrum and milk. Nutrients
2011; 3: 442–474.
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