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Milk | Human Milk
Human Milk A Darragh, Massey University, Palmerston North, New Zealand B Lo¨nnerdal, University of California, Davis, CA, USA ª 2011 Elsevier Ltd. All rights reserved. This article is a revision of the previous edition article by A. Darragh, Volume 3, pp 1350–1360, ª 2002, Elsevier Ltd.
Introduction The primary function of mammalian milk is to nourish the neonate in the most efficient manner possible from the mother’s perspective, while providing the neonate with additional benefits from enhanced immune protection, and a source of nonnutrient growth promoters, hormones, and other bioactive components that serve as biochemical messengers within the body. In addition, several components in human milk help to maintain the integrity of mammary tissue, optimizing milk output and composition. Human milk, a biologically complex fluid containing many hundreds of different constituents, is ideally suited to meet the term infant’s nutritional needs during the first 4–6 months of life, provided that the mother is also healthy and well nourished. For this reason, infant formula manufacturers often refer to the gross composition of human milk as the ‘gold standard’. However, it is important to remember that, unlike infant formula, the composition of human milk is not uniform. Significant changes in the composition of breast milk occur not only between individual women, but also within a single feed, between feeds, throughout the day, throughout the lactation period, with changing maternal diet, and as a result of other external factors such as exercise or metabolic illness. Implicit in these findings is the understanding that a single description of human milk is somewhat erroneous. Nevertheless, it is important to establish data on the composition of human milk at various key stages of lactation.
Gross Nutrient Composition There are three stages in human lactation that are clearly identified by changes in specific components in the milk, such as the whey proteins and lactose. These compositional changes appear uniquely to match the changing physiological needs of the infant, and can be defined based on the following time periods: 1–5 days post-partum, colostrum; 5–21 days postpartum, transitional milk; and >21 days post-partum, mature milk.
Nutrient information relating to human milk at the predominant stages of lactation is provided in Table 1. Human colostrum differs from mature milk in both the types and amounts of individual components. Colostrum has a much higher protein content and a lower lactose content than mature milk. The specific gravity of colostrum is 1.040 compared with 1.060 for mature milk, and the mean energy content of colostrum is also lower at 67 kcal 100 ml1 compared with mature milk, which supplies 75 kcal 100 ml1 of energy. Although the fat and ash contents of colostrum and mature milk do not differ significantly, variations in specific micronutrients are apparent, with the concentrations of sodium, chloride, and magnesium being higher, and that of potassium and calcium lower, in colostrum.
Component Composition Human milk contains hundreds of different components; Table 2 lists some of these grouped according to their physicochemical properties. The water-soluble constituents found in the aqueous phase account for 87% of milk volume, and include some salts, vitamins and trace elements, lactose, oligosaccharides, whey proteins, and nonprotein nitrogen (NPN) components. Complementing the aqueous phase is a colloidal dispersion (0.3% of volume) containing casein proteins held in micellar form, which contributes to the white color of milk. Calcium phosphate and some other proteins are also linked to the casein micelles. A further 6% of milk volume is made up of an emulsion containing fat globules, fat globule membranes, triglycerides, and other lipid components, including the fat-soluble vitamins. The remainder consists of cellular components, non-peptide hormones, and other minor fractions. Proteins Many of the functional properties of human milk are delivered via the milk proteins. Provision of essential amino acids for growth is the most obvious role of milk proteins. Determining the gross amino acid composition of human milk will accurately describe amino acid
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582 Milk | Human Milk Table 1 Gross nutrient content (g 100 ml1) of human milk at different stages in lactation Nutrient
Colostrum (1–5 days)
Transitional milk (5–21 days)
Mature milk (21 daysþ)
Protein Non-protein nitrogen Lactose Glucose Oligosaccharides Total lipid Ash
1.5–1.7 0.5 4.1–5.8 0.2–1.0 2.4 2–3 0.25
0.9
0.8–0.9 0.45 6.8 0.2–0.3 1.3 3–5 0.2
5.4
3–4
Table 2 Components of human milk grouped according to physicochemical properties Proteins
Non-protein nitrogen
-Casein -Lactalbumin Lactoferrin Lysozyme Serum albumin Secretory immunoglobulin A Immunoglobulin A Immunoglobulin G Immunoglobulin M Peptide hormones Enzymes Growth factors Binding proteins
Urea Glucosamine Creatine/creatinine Uric acid Nucleotides Nucleic acids Polyamines
Carbohydrates Lactose Oligosaccharides Glycoproteins Minerals Calcium Phosphate Potassium Sodium Sulfate Magnesium Chloride Trace minerals Chromium Cobalt Copper Iodine Iron Manganese Molybdenum Nickel Selenium Zinc
Lipids Fatty acids Phospholipids Triacylglycerols Fat-soluble vitamins (A, D, E, K) Sterols Other vitamins Biotin Choline Folic acid Inositol Nicotinic acid Pantothenic acid Riboflavin Thiamine Vitamin B1 Vitamin B2 Vitamin B6 Vitamin B12 Vitamin C Cells Epithelial cells Leukocytes Lymphocytes Macrophages Neutrophils Others Bicarbonate Citrate Taurine Glutamine Carnitine Nonpeptide hormones
content, and in the past this figure has been used to define the amount of protein available to the infant for growth and development. However, milk proteins also provide the infant with immune protection, increased digestive capacity, and enhanced micronutrient availability, and, as many bioactive components of human milk are of protein origin, implicit in their functionality is a resistance to digestion. Investigation of the nutritive value of human milk protein reveals that although most (95%) milk protein is available, a proportion does not contribute significantly to amino acid nutrition. To determine accurately the amounts of amino acids, and therefore protein, used for nutrition by the breast-fed infant, a correction needs to be made for bioavailability. A comparison of gross and available amino acid composition of mature human milk is made in Table 3. The total protein content of human milk consists of two major groups, caseins and whey proteins, as well as Table 3 The gross and available amino acida composition of mature human milk (mg 100 g1 proteinb) Amino acid
Gross
Available
Aspartic acid Threonine Serine Glutamic acid Proline Glycine Alanine Valine Isoleucine Leucine Tyrosine Phenylalanine Histidine Lysine Arginine Cysteine Methionine
9.2 4.7 5.0 18.4 8.5 2.3 3.8 6.4 5.9 9.7 4.2 3.9 2.3 6.8 4.0 2.2 1.6
9.1 4.6 4.2 16.8 8.4 2.1 3.4 5.1 5.1 9.3 3.8 3.8 2.3 6.2 3.1 2.6c 1.4
a
Includes free amino acids and peptides. Calculated from total g N 6.38. c Value for available is higher than gross due to correction for significant losses during the acid hydrolysis stage of amino acid determination. b
Milk | Human Milk Table 4 The types and amounts of protein (g 100 ml1) and non-protein nitrogen (mg 100 ml1) in human colostrum and mature milk
Casein proteins -Casein -Casein Whey proteins -Lactalbumin Serum albumin Lactoferrin Lysozyme Secretory immunoglobulin A/ immunoglobulin A Immunoglobulin G Immunoglobulin M Non-protein nitrogen (N) Urea N Creatine N Creatinine N Uric acid N Glucosamine -Amino N Ammonia N
Colostrum
Mature milk
0.26 0.12
0.3–0.5 0.1–0.3
0.36 0.04 0.35
0.2–0.3 0.03 0.1–0.3 0.05 0.05–0.1
0.2
0.034 0.012 48 12.1
0.5 14.2 4.5 0.2–0.8
0.001 0.002 50 15–25 3.7 3.5 0.5 4.7 13 0.2
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milk. -Lactoglobulin, prominent in bovine milk, does not occur in human milk. The predominant sources of amino acids for nutrition among the whey proteins are -lactalbumin and serum albumin. The other whey proteins, while contributing to nutrition to some extent, have a primary function in other roles. Lactoferrin, an iron-binding protein, has bacteriostatic properties that may impact on both the infant and the mammary gland. Lysozyme is capable of cleaving proteoglycans in the cell wall of certain bacteria, providing an antibacterial effect. The Igs in human milk are distinct from those found in blood serum, indicating selective synthesis in the mammary gland. sIgA, formed by the linking of two serum IgA molecules by disulfide bonds, is present at very high concentrations in colostrum, but the sIgA content declines rapidly to a much lower level within 14 days of lactation. sIgA protects the infant by blocking the attachment of pathogenic bacteria to the gut epithelium. Other Igs in human milk provide immune protection through activation of the complement system, or by promoting phagocytosis. Other proteins
some minor components (Table 1) and proteins present in the milk fat globule membrane. The ratio of whey proteins to casein changes during the initial stages of lactation, from 90:10 immediately post-partum, to 60:40 in mature milk and 50:50 in late lactation. These ratios directly reflect a decrease in the content of immune proteins (Table 4) in line with development of the infant’s own immune system. Casein
Human milk contains two ( and ) caseins, and is devoid of -casein, the predominant casein found in bovine milk. The casein pattern differs depending on the stage of lactation, with -casein detectable in human milk only from day 3–4 post-partum onward. A unique feature of human milk is the high degree of glycosylation of -casein (40–60% glycosylated) and other milk proteins (e.g., immunoglobulins (Igs) and lactoferrin). Glycosylated proteins exhibit anti-infective properties and enhance the absorption of some micronutrients. As part of the normal digestive process, the caseins in human milk release smaller peptides into the gut lumen. These casein fragments are thought to enhance the absorption of calcium by keeping it in solution in the gut lumen. Other casein-derived peptides have been linked to the regulation of intestinal motility and to the growth promotion of beneficial bacteria in the infant’s gut. Whey proteins
-Lactalbumin, lactoferrin, serum albumin, and secretory IgA (sIgA) are the main whey proteins found in human
In addition to casein and whey proteins, human milk contains other proteins, such as serum proteins, most of which are in the aqueous phase. The milk fat globule membrane proteins surround the lipid droplets, but are a minor fraction quantitatively (1–3% of total protein). However, these minor proteins can contribute in many ways to the health and well-being of the infant, for example, by facilitating transport of certain vitamins and minerals. Milk-borne peptide hormones and enzymes assist in the digestion of other milk components (e.g., bile salt-stimulated lipase (BSSL) and amylase). A detailed list of the hormones, both peptide and non-peptide, and enzymes found in human milk is given in Table 5.
Non-protein Nitrogen Compared with the milk of other mammalian species, human milk is notably different with regard to the NPN fraction. In human milk, the NPN fraction represents a very high percentage (20–25%) of total nitrogen (N). Urea N accounts for almost 50% of the NPN in human milk, with more than 200 compounds, including free amino acids, carnitine, taurine, amino sugars, nucleic acids, nucleotides, and polyamines, making up the remainder. In general, the total NPN of human milk remains constant throughout the lactation period, although there may be some variation in individual NPN components (Table 4).
584 Milk | Human Milk Table 5 Hormones and enzymes found in mature human milk Hormones Peptide Growth factors
Gastrointestinal regulators
Hypothalamus/hypophyseal hormones
Thyroid–parathyroid group
Non-peptide Epidermal growth factor Insulin Insulin-like growth factor 1 Nerve growth factor Transforming growth factor- Transforming growth factor-
Thyroid
Thyroxine Triiodothyronine
Adrenal
Cortisol
Sexual
Progesterone Pregnane3( )20( )-diol Estrogen
Gastrin Gastric inhibitory polypeptide Gastric-releasing peptide Neurotensin Peptide histidine-methionine Peptide tyrosine–tyrosine Somatostatin Vasoactive intestinal peptide Gonadotropin-releasing hormone Growth-releasing factor Growth hormone Prolactin Thyrotropin-releasing hormone Thyroid-stimulating hormone Calcitonin-like hormone Parathyroid hormone Parathyroid hormone-related peptide
Enzymes Phosphoglucomutase Fatty acid synthase Galactosyltransferase Bile salt-stimulated lipase Lysozyme
-Glutamyltransferase Protease inhibitors Lipoprotein lipase Alkaline phosphatase
Given that the total protein content of human milk is low compared to other species, yet supports adequate growth of the breast-fed infant, the suggestion has been made that the urea found in the NPN fraction of human milk is used by the infant for protein synthesis. This would have major implications for infant formula manufacturers, necessitating a total reevaluation of infants’ protein requirements. Isotopic tracer experiments with infants have shown that as much as 40% of labeled urea N was retained within the body. However, this may not directly reflect the incorporation of urea N into body protein, as NH exchanges occurring during transamination may in part explain the appearance of urea 15N in body proteins. This remains an important area for future investigation of infant protein metabolism.
Lactose synthase Thioesterase Amylase Proteases Peroxidase Platelet-activating factor acetylhydrolase Sulfhydryl oxidase Glutathione peroxidase Xanthine oxidase
Taurine and glutamine, two predominant free amino acids in human milk, are considered conditionally essential, as the infant appears to have a requirement for these amino acids above what the body can synthesize. Taurine plays an integral part in fat digestion by conjugating with bile acids to form bile salts. Although no direct link has been made between a deficiency in taurine and abnormal development of retinal and brain tissue in infants, the addition of taurine to infant formulae is now routine. Glutamine is involved in the cellular metabolism of enterocytes and in the immune response associated with inflammation and sepsis. During periods of rapid growth or tissue repair, a dietary source of nucleotides may be beneficial, as de novo synthesis of nucleotides can be a limiting factor.
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Table 6 Nucleotide content of human milk, and the amounts of specific nucleotides approved for inclusion in infant formulae
Nucleotide
In human milk (mg 100 ml1)
Adenosine diphosphate (ADP) Adenosine monophosphate (AMP) Cytidine diphosphate (CDP) Cytidine monophosphate (CMP) Guanosine diphosphate (GDP) Guanosine monophosphate (GMP) Inosine monophosphate (IMP) Uridine diphosphate (UDP) Uridine monophosphate (UMP)
69 175 474 461 96 138 228 174 179
Maximum to be added to infant formula (mg 100 kcal1)
1.5 2.5 0.5 1.0 1.75
Adapted from Lawrence and Lawrence (1994); infant formula inclusion rates from Rudloff S and Kunz C (1997) Protein and nonprotein nitrogen components in human milk, bovine milk, and infant formula: Quantitative and qualitative aspects in infant nutrition. Journal of Pediatric Gastroenterology and Nutrition 24: 328–344.
The infant is in a state of rapid growth and cellular development, and human milk is rich in nucleotides (Table 6). To date, however, no link between nucleotides in human milk and enhanced cellular function in the infant has been made. Despite this lack of direct evidence, some regulatory authorities have approved the inclusion of nucleotides in infant formulae (Table 6).
Table 7 Quantities of some carbohydrates found in human milk
Carbohydrate Besides water, lactose constitutes the largest component of human milk (Table 1); lactose concentration decreases as lactation progresses, in line with the decline in milk volume. Oligosaccharides constitute the next greatest fraction of carbohydrate in human milk, representing more than 27% of the carbohydrate in colostrum. This gradually decreases to 15–16% of carbohydrate as oligosaccharides in mature human milk. Most oligosaccharides found in human milk contain lactose at the reducing end, and may contain fucose or sialic acid at the non-reducing end. Compared to other mammalian milks, human milk is unique due to its content of complex oligosaccharides. Milk oligosaccharides provide a low osmolar source of calories for the infant, and have been classed as ‘soluble’ fiber because they appear to remain largely undigested in the infant’s gut. Human milk oligosaccharides contain significant quantities of sialic acid, which is essential for brain development, and they are also thought to stimulate the growth of bifidus bacteria and to inhibit the adhesion of pathogenic bacteria to epithelial surfaces. Approximately 130 oligosaccharides have been identified in human milk, some of which are listed in Table 7. Glucose and galactose are present in much smaller quantities in human milk, as are other carbohydrates such as monosaccharides, and peptide-bound and protein-bound carbohydrates.
carbohydrate oligosaccharid
Concentration mg 100 ml1
Lactose Oligosaccharides Lacto-N-tetraose Lacto-N-fucopentaose I Lacto-N-fucopentaose II Lacto-N-fucopentaose III Lacto-N-difucohexaose I NeuAc( 2–6)lactose NeuAc( 2–3)lactose NeuAc-lacto-N-tetraose a NeuAc-lacto-N-tetraose b NeuAc2-lacto-tetraose
6800 1200–1300 5–15 12–17 3–10 0.1–2 1–2 3–5 1–3 0.3–2 1–6 2–6
Adapted from Kunz C, Rodriguez-Palmero M, Koletzko B, and Jensen R (1999) Nutritional and biochemical properties of human milk. 1. General aspects, proteins, and carbohydrates. Clinics in Perinatology 26: 307.
Lipids The lipids in human milk occur in globular form, emulsified in the aqueous phase, and provide the infant with a major proportion (45–55%) of the energy needed to support growth. They also contain bioactive components important to the infant’s retinal and neural tissue development. The triacylglycerols (TGs) account for 98% of total fat, and 90% of this is fatty acids (FAs). Phospholipids (0.8%), cholesterol (0.5%), and other minor lipid components make up the remainder of the fat, which includes fat-soluble vitamins. In the first few weeks of lactation, the total lipid content increases from around 2 to 3.5–5% (Table 8). As the fat content increases, so too does the average size of the milk fat globules, which in turn results in a reduction in the ratio of phospholipids and cholesterol in TGs.
586 Milk | Human Milk Table 8 Lipid content of human colostrum and milk Lipid component
Colostrum
Mature milk
Total lipid (percentage of milk volume) Phospholipid (percentage of total lipid) Triacylglycerol (percentage of total lipid) Cholesterol (percentage of total lipid)
2
3–5
1.1
0.8
97–98
97–98
1.3
0.5
Table 9 An example of the fatty acid composition of human milk (wt%) Fatty acid
Colostrum
Mature milk
Saturated 10:0 12:0 14:0 15:0 16:0 17:0 18:0 20:0 22:0 24:0
0.5 2.3 5.3 0.3 26.2 0.4 7.5 0.3 0.2 0.3
1.1 4.8 6.7 0.3 21.8 0.3 7.5 0.2 0.1 0.1
Monounsaturated 14:1n9 0.1 16:1n9 2.1 18:1n9 34.7 20:1n9 1.3 22:1n9 0.3 24:1 0.5
0.3 2.7 33.0 0.6 0.1 0.1
Polyunsaturated 18:3n3 0.8 20:3n3 0.2 20:5n3 0.04 22:5n3 0.2 22:6n3 0.6 18:2n6 8.8 18:3n6 0.1 20:2n6 0.6 20:3n6 0.6 20:4n6 0.8 22:4n6 0.2 22:5n6 0.4
1.0 0.1 0.04 0.1 0.2 10.7 0.2 0.3 0.4 0.5 0.1 0.2
The TG composition in human milk is determined by the types and quantities of FAs esterified to the glycerol molecule. Human milk contains eight major FAs in amounts greater than 1% (Table 9), and is an excellent source of the essential FAs C18:2n6 and C18:3n3 and their long-chain derivatives C20:4n6 (arachidonic acid) and C22:6n3 (docosahexaenoic acid). The latter FAs, although not classed as essential for adults, are the only FAs utilized by the brain and are important structural components of the membrane systems of all tissues.
Approximately 60% of the C16:0 FAs in human milk are situated at the sn-2 position on the TG, which is a unique feature of the FAs in human milk. Lipolytic hydrolysis cleaves FAs from the sn-1 and sn-3 positions, and a C16:0 remains as a monoacylglycerol. This facilitates the absorption of C16:0 across the infant’s gut epithelium, improving overall lipid utilization to around 90–95%. Lipid digestion and absorption are enhanced further by four enzyme systems. These are, in the order of utilization, the gastric phase with lingual lipase and gastric lipase, and the intestinal phase with pancreatic lipase and BSSL. The gastric phase is important because the milk fat globule would otherwise resist the action of pancreatic lipase and BSSL. Unlike the other lipases, which are endogenous, BSSL is secreted in human milk specifically to assist the infant with milk fat digestion. Of all the components in human milk, the lipid fraction is the most variable. Changes in fat content occur not only during the first few weeks of lactation (Table 8), but also in milk secretions throughout the day, and even within a single feed, with the hindmilk being significantly higher in fat content than the foremilk. Variations also exist in the fat content of milk from individual women, although by far the greatest variation occurs in the types and amounts of FAs.
Micronutrients Vitamins
Most vitamins found in human milk increase in concentration as lactation shifts from colostrum to a mature secretion (Table 10), with the exception of carotenoids ( -carotene, -carotene, lutein, cryptoxanthin, and lycopene), retinol, and vitamin E, which decrease. Reduction in the concentration of some vitamins may not alter the total amount available to the infant, however, as any reduction may be offset by an increase in the quantity of breast milk consumed. The milk from healthy, well-nourished women usually contains sufficient quantities of most of the vitamins required by the infant. There are a few exceptions, however, when the amount in milk may not be adequate. Deficiencies in vitamin D, with consequent development of rickets, have been reported in breast-fed infants, directly linked to the mother’s diet and the infant’s lack of exposure to sunlight. To meet physiological requirements, vitamin K is routinely given to newborn babies because transplacental transfer is limited, and the level in milk is low. Minerals
In general, the macromineral content of human milk remains stable during the lactation period after an initial decrease in the concentration of chloride, potassium,
Milk | Human Milk Table 10 Micronutrient content of human colostrum and milk Micronutrient
Units l
Water-soluble vitamins Ascorbic acid mg Biotin mg Folic acid mg Niacin mg Pantothenic acid mg Riboflavin mg Thiamine mg Vitamin B6 mg mg Vitamin B12
1
Colostrum
Mature milk
60 0.6 5 0.6 1.8 300 19.5
50–100 6 80–133 1.6–6.0 2.0–2.5 400–600 200 0.1–0.3 1.0
0.5
Fat-soluble vitamins Carotenoids Retinol Vitamin D Vitamin E Vitamin K
mg mg mg mg mg
2 1.8
Major minerals Calcium Chloride Citrate Magnesium Phosphorus Potassium Sodium Sulfur
mg mg mg mg mg mg mg mg
300 813
Trace minerals Chromium Cobalt Copper Fluoride Iodine Iron Manganese Molybdenum Nickel Selenium Zinc
ng ng mg mg mg mg mg mg mg mg mg
10–15 2–5
36 140 652 502 277
0.4–0.8
0.75 5–12
40 8–12
0.2–0.6 0.3–0.7 0.4 3–8 3–15 278 426 500a 35 140 530 180 142 200–400 100–200 0.2–0.4 4–15 146 0.72 3–6 1–2 0.5–2 10–20 1–3
a Citrate is not classed as a mineral, but is soluble in water and binds some minerals, so is included in this table.
sodium, and sulfur (Table 10). Most of the sulfur in human milk is found in the sulfur-containing amino acids methionine and cysteine, with only a small fraction (10%) existing in salt form. The sulfur amino acids predominate in whey proteins, so a reduction in whey protein content during the first few days of lactation is matched by a similar reduction in sulfur content. Human milk has a higher calcium-to-phosphorus ratio (2:1) than the milk from other mammalian species (e.g., in bovine milk, Ca:P ratio is 1.1:1), although the absolute amounts of calcium and phosphorus are lower. A low phosphorus content suits the infant’s limited renal capacity. In addition, the contribution of phosphate to the buffering capacity of human milk is such that a low pH results, promoting the growth of beneficial gut bacteria.
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Human milk contains what would initially appear to be an inadequate level of iron, yet breast-fed infants rarely deplete their iron stores until after 4–6 months of age. Maximum utilization of the iron in human milk is achieved with the assistance of lactoferrin, most likely via specific protein receptors in the gut that link with lactoferrin and therefore facilitate iron absorption.
Bioactive Factors Human milk contains an optimal balance of the nutrients required by the infant and, in addition, it contains an array of bioactive factors. In the first instance, these factors assist in the production and secretion of milk components and provide immune protection of mammary tissue. Once ingested by the infant, however, bioactive factors actively promote growth and development, and afford the infant immune protection until its own defense system has matured. Even the major components in milk may have specific bioactive functions either before being digested or as a result of releasing bioactive components during the digestive process. The main sites of action for these bioactive factors are mucosal surfaces, although some may enter the circulation and have a systemic effect elsewhere in the body. The various physiological roles of milk components and their derivatives, either within the mammary gland or in the infant, are listed in Table 11. Intact cellular components are found in human milk (Table 12), and although their exact role within either the mammary gland or the recipient infant is not clearly defined, there does appear to be a degree of biological activity associated with them. The total count of cells is high in colostrum, with the numbers decreasing as lactation becomes established. More than 90% of the cells in human milk are macrophages and neutrophils, while the remainder are lymphocytes (80% of which are T cells) and epithelial cells. The primary function of milk leukocytes is thought to be as a means of defense within the mammary gland, and not necessarily to transfer maternal immunocompetence to the infant. Macrophages,however, have been suggested as a potential vehicle for the storage and transport of milk-borne Igs, and may be involved in the formation of lactoperoxidase and other factors that enhance the growth of intestinal epithelium and digestive capacity. The lymphocytes present in human milk are part of the immunological system that synthesizes IgA, and considerable research data are available to support the suggestion that lymphocytes in colostrum and milk provide the infant with immunological benefits.
588 Milk | Human Milk Table 11 Physiological roles of components in human milk, either within the mammary gland or following ingestion by the infant Physiological function
Components in human milk
Within the mammary gland Component synthesis/secretion Immune effect/anti-infection/ anti-inflammatory
-Lactalbumin, lipoprotein lipase, xanthine oxidase Lactoferrin, lysozyme, -glutamyltransferase, cytokines
Within the infant Nutrition Enhancing nutrient availabilitya Gut colonization Gastrointestinal development Brain development Immunity Anti-infective Anti-inflammatory Immunomodulating
- and -casein, whey proteins, lactose, fats, minerals, and vitamins Bile salt-stimulated lipase, milk amylase, lactoferrin (Fe), glutamine (Zn), -lactalbumin (Ca), casein phosphopeptides (Ca), folate-binding protein Bifidus factor peptide, -casein oligosaccharides Epidermal growth factor, insulin-like growth factor, growth factor-binding proteins, hormones, enzymes, nucleotides, oligosaccharides, amino sugars Docosahexaenoic acid, other long-chain polyunsaturated fatty acids, taurine, carnitine Immunoglobulins, leukocyte-stimulating factors, T lymphocytes, macrophages, immunopeptides Lactoferrin, lysozyme, free fatty acids, cell adhesion molecules, cytokines, mucins, glycoproteins, glycopeptides, oligosaccharides Vitamins, prostaglandins, growth factors, cytokines (interleukin-10), transforming growth factor- Interleukins, tumor necrosis factor-
a
Nutrient that has increased availability due to a particular component in human milk is given in parentheses.
Table 12 Cellular components (number per ml) in human colostrum and milk Cell type
Colostrum
Mature milk
Total cells Macrophages Neutrophils Lymphocytes
2840 1490 1375 250
51 52 8 1
Adapted from Jensen R (ed.) (1995) Handbook of Milk Composition. San Diego, CA: Academic Press.
Factors that Influence Milk Volume and Composition Factors Affecting Milk Volume In women, the principal determinant of milk volume is infant demand. Comprehensive review of milk volume from exclusively breast-feeding women throughout the world has revealed that milk volume, 6 months into lactation, is incredibly constant, with an average of 800 ml (range 500–1200 ml) of milk transferred from mother to infant each day. The potential for milk production far exceeds this amount, however, given that mothers of twins or triplets can adequately nourish their children with human milk. The energy density of human milk, in part, influences milk production, as infants consuming lower calorific milk will suckle more to obtain the energy required, in turn stimulating greater milk production. In one study, women with a low body fat content produced as much as 15% more milk as a result of increased infant demand. As soon as the weaning process begins, and human milk is
substituted by either supplementary formula or complementary solids, milk production declines, indicating yet again the dominant role the infant plays in controlling milk volume.
Major Factors Affecting Milk Composition Stage of lactation
Major compositional changes that occur from parturition through to day 21 of lactation have been discussed. The composition of human milk continues to change throughout lactation, however, with a gradual reduction in component content, although some components, for example, lysozyme, may increase. During weaning, changes in composition can occur depending on the time taken to wean. When weaning is rapid (several days/weeks), lactose and potassium contents decrease, while sodium, chloride, fat, and total protein (specifically the whey proteins) increase. Gradual weaning (over several months) results in an increase in sodium (220% from baseline), iron (172%), and protein (142%), no change in calcium, and a decrease in zinc (58% from plateau). Both rapid and gradual weaning produce a decrease in lactose content. Prematurity
Relative to normal human milk, the milk of mothers delivering prematurely contains higher concentrations of protein (1.8–2.4 g 100 ml1), short-, medium-, and long-chain FAs, calcium, phosphorus, sodium, chloride, magnesium, and iron, which is related to the low volume produced. Cellular components are also significantly
Milk | Human Milk
greater in premature colostrum (total cells 6800 per ml). Levels of the other components in premature milk do not differ markedly from those found in normal human milk. Changes during a feed and during the day
Changes that occur in composition during a feed relate more to the physicochemical nature of fractions in the milk, rather than a selective change in individual components. In general, most components remain relatively constant throughout a feed. The lipid content increases rapidly toward the end of a feed, however, reflecting a propensity of the mammary gland tissue to retain the lipid globules. Subjective evidence from breast-feeding women suggests that milk composition changes during the day, especially toward evening, when the infant appears to be less satisfied by milk consumed. An unsettled infant is more likely to be due to insufficient milk volume, however, as lipid content actually increases toward the end of the day (by two- to fivefold). No significant changes occur during the day in other components. Nationality/age/parity
The composition of milk from women from different geographic, ethnic, and socioeconomic backgrounds is remarkably similar, particularly in reference to the macronutrients. Any differences observed are more likely to be the result of dietary variation, rather than genetic modification of composition. Milk from teenagers has a significantly lower concentration of lactose and some macrominerals compared to adults, but diet may play an important part in this difference as the diet of teenagers is often suboptimal. Parity may have a significant effect on milk composition, with primiparous women having higher concentrations of protein and fat, and multiparous women experiencing a reduction in the content of major components in milk with each successive lactation. Maternal diet
Two of the most significant factors influencing milk composition are the nutritional status of the mother and her ongoing diet. An inadequate nutritional status will adversely affect milk volume and the content of milk minerals such as iron and selenium. Severe malnutrition and diminished nutritional status will cause a decrease in the protein and fat content of milk. The components most affected by maternal diet are FAs, vitamins, and some macro- and micro-minerals. The FA content of human milk can undergo pronounced changes that directly reflect the FA content of the diet. For example, the C18:2n6 content of milk from vegetarians is high compared to the milk from women consuming an omnivorous diet. Also, a low-fat diet will result in a
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marked increase in medium-chain FAs as the de novo synthesis of FAs in the mammary gland increases. Exercise
Recent evidence suggests that the lactic acid content of human milk can change following exercise, with a decline in infant acceptance of milk secreted immediately after exercise. Exercise regimens at 100, 75, or 50% of VO2 max resulted in increases in the lactic acid content of milk sampled immediately after exercise, although only 100% intensity exercise produced milk with a lactic acid content significantly different from baseline. Thus, breast-feeding women can complete a moderate exercise program without the fear that the infant will reject their milk. Illness and metabolic disorders Mastitis
Infection of the mammary tissue, with cellulitis and, more rarely, abscess formation, can alter milk composition to such an extent that the infant’s nutrition is compromised. During a bout of mastitis, milk volume and lactose content decrease, while the levels of sodium and chloride increase, giving the milk a much saltier taste. Immune factors such as sIgA, lysozyme, and lactoferrin also increase in milk from mastitic tissue, as part of the mammary gland immune response. Insulin-dependent diabetes mellitus
The delayed lactogenesis associated with mothers with insulin-dependent diabetes mellitus (IDDM) results in a lower volume of milk being consumed by the infant in the first few days following birth. Initially (<5 days’ lactation), the milk from an IDDM mother has a composition different from that of normal milk, with less lactose and fat, and more total nitrogen. Once lactation has been established, however, most of these differences diminish. The FA content of milk from IDDM women continues to differ throughout the lactation, with more C10, C12, and C14, and less C20 and C22, FAs. Drug use
The degree to which a drug taken by the mother will affect the breast-fed infant depends entirely on the class of drug. Detailed information on the transfer of pharmaceutical drugs in human milk is available. Recreational drugs, such as alcohol, caffeine, nicotine, and marijuana, also appear in human milk and can affect both milk volume and composition. Human milk from a mother who smokes not only contains many chemical by-products, including nitrates, nitrites, lead, and cadmium, but is also likely to have lower levels of certain vitamins. Exposure of an infant to drugs (both medical and recreational) in milk can be minimized by advising the mother to take any
590 Milk | Human Milk
medication immediately after a breast-feed, and/or just before the infant is due to have a lengthy sleep period.
Human Milk Banking Human milk is usually collected and stored in milk banks to assist in the nourishment of premature infants. Implicit in this action of ‘banking’ milk is the need to maintain functional integrity in the milk. Numerous steps in the process can affect the composition of human milk, including the suitability of the donor, and her dietary regimen; the method of milk collection and the type of container chosen to collect the milk; the length of time and temperature at which the milk is stored; and the methods used to thaw, mix, and reheat the milk prior to a feed. The Human Milk Banking Association of North America has compiled a set of guidelines for the establishment and operation of a donor milk bank. See also: Dehydrated Dairy Products: Infant Formulae. Mammals. Milk: Colostrum.
Further Reading Committee on Drugs (1994) The transfer of drugs and other chemicals into human milk. Pediatrics 93: 137–150. Hamosh M (2001) Bioactive factors in human milk. Pediatric Clinics of North America 48: 69–86. Jensen R (ed.) (1995) Handbook of Milk Composition. San Diego, CA: Academic Press. Jensen R (1999) Lipids in human milk. Lipids 34: 1243–1271. Kunz C, Rodriguez-Palmero M, Koletzko B, and Jensen R (1999) Nutritional and biochemical properties of human milk. 1. General aspects, proteins, and carbohydrates. Clinics in Perinatology 26: 307–333. Lawrence RM and Lawrence RA (1999) Biochemistry of human milk. Breastfeeding: A Guide for the Medical Profession, 5th edn., pp. 95–155. St Louis, MO: Mosby. McVeagh P and Brand Miller J (1997) Human milk oligosaccharides: Only the breast. Journal of Paediatrics and Child Health 33: 281–286. Picciano MF (2001) Representative values for constituents of human milk. Pediatric Clinics of North America 48: 53–67 (appendix 263–264). Rodriguez-Palmero M, Koletzko B, Kunz C, and Jensen R (1999) Nutritional and biochemical properties of human milk. 2. Lipids, micronutrients, and bioactive factors. Clinics in Perinatology 26: 335–359. Rudloff S and Kunz C (1997) Protein and nonprotein nitrogen components in human milk, bovine milk, and infant formula: Quantitative and qualitative aspects in infant nutrition. Journal of Pediatric Gastroenterology and Nutrition 24: 328–344.
Introduction The primary function of mammalian milk is to nourish the neonate in the most efficient manner possible from the mother’s perspective, while providing the neonate with additional benefits from enhanced immune protection, and a source of nonnutrient growth promoters, hormones, and other bioactive components that serve as biochemical messengers within the body. In addition, several components in human milk help to maintain the integrity of mammary tissue, optimizing milk output and composition. Human milk, a biologically complex fluid containing many hundreds of different constituents, is ideally suited to meet the term infant’s nutritional needs during the first 4–6 months of life, provided that the mother is also healthy and well nourished. For this reason, infant formula manufacturers often refer to the gross composition of human milk as the ‘gold standard’. However, it is important to remember that, unlike infant formula, the composition of human milk is not uniform. Significant changes in the composition of breast milk occur not only between individual women, but also within a single feed, between feeds, throughout the day, throughout the lactation period, with changing maternal diet, and as a result of other external factors such as exercise or metabolic illness. Implicit in these findings is the understanding that a single description of human milk is somewhat erroneous. Nevertheless, it is important to establish data on the composition of human milk at various key stages of lactation.
Gross Nutrient Composition There are three stages in human lactation that are clearly identified by changes in specific components in the milk, such as the whey proteins and lactose. These compositional changes appear uniquely to match the changing physiological needs of the infant, and can be defined based on the following time periods: 1–5 days post-partum, colostrum; 5–21 days postpartum, transitional milk; and >21 days post-partum, mature milk.
Nutrient information relating to human milk at the predominant stages of lactation is provided in Table 1. Human colostrum differs from mature milk in both the types and amounts of individual components. Colostrum has a much higher protein content and a lower lactose content than mature milk. The specific gravity of colostrum is 1.040 compared with 1.060 for mature milk, and the mean energy content of colostrum is also lower at 67 kcal 100 ml1 compared with mature milk, which supplies 75 kcal 100 ml1 of energy. Although the fat and ash contents of colostrum and mature milk do not differ significantly, variations in specific micronutrients are apparent, with the concentrations of sodium, chloride, and magnesium being higher, and that of potassium and calcium lower, in colostrum.
Component Composition Human milk contains hundreds of different components; Table 2 lists some of these grouped according to their physicochemical properties. The water-soluble constituents found in the aqueous phase account for 87% of milk volume, and include some salts, vitamins and trace elements, lactose, oligosaccharides, whey proteins, and nonprotein nitrogen (NPN) components. Complementing the aqueous phase is a colloidal dispersion (0.3% of volume) containing casein proteins held in micellar form, which contributes to the white color of milk. Calcium phosphate and some other proteins are also linked to the casein micelles. A further 6% of milk volume is made up of an emulsion containing fat globules, fat globule membranes, triglycerides, and other lipid components, including the fat-soluble vitamins. The remainder consists of cellular components, non-peptide hormones, and other minor fractions. Proteins Many of the functional properties of human milk are delivered via the milk proteins. Provision of essential amino acids for growth is the most obvious role of milk proteins. Determining the gross amino acid composition of human milk will accurately describe amino acid
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582 Milk | Human Milk Table 1 Gross nutrient content (g 100 ml1) of human milk at different stages in lactation Nutrient
Colostrum (1–5 days)
Transitional milk (5–21 days)
Mature milk (21 daysþ)
Protein Non-protein nitrogen Lactose Glucose Oligosaccharides Total lipid Ash
1.5–1.7 0.5 4.1–5.8 0.2–1.0 2.4 2–3 0.25
0.9
0.8–0.9 0.45 6.8 0.2–0.3 1.3 3–5 0.2
5.4
3–4
Table 2 Components of human milk grouped according to physicochemical properties Proteins
Non-protein nitrogen
-Casein -Lactalbumin Lactoferrin Lysozyme Serum albumin Secretory immunoglobulin A Immunoglobulin A Immunoglobulin G Immunoglobulin M Peptide hormones Enzymes Growth factors Binding proteins
Urea Glucosamine Creatine/creatinine Uric acid Nucleotides Nucleic acids Polyamines
Carbohydrates Lactose Oligosaccharides Glycoproteins Minerals Calcium Phosphate Potassium Sodium Sulfate Magnesium Chloride Trace minerals Chromium Cobalt Copper Iodine Iron Manganese Molybdenum Nickel Selenium Zinc
Lipids Fatty acids Phospholipids Triacylglycerols Fat-soluble vitamins (A, D, E, K) Sterols Other vitamins Biotin Choline Folic acid Inositol Nicotinic acid Pantothenic acid Riboflavin Thiamine Vitamin B1 Vitamin B2 Vitamin B6 Vitamin B12 Vitamin C Cells Epithelial cells Leukocytes Lymphocytes Macrophages Neutrophils Others Bicarbonate Citrate Taurine Glutamine Carnitine Nonpeptide hormones
content, and in the past this figure has been used to define the amount of protein available to the infant for growth and development. However, milk proteins also provide the infant with immune protection, increased digestive capacity, and enhanced micronutrient availability, and, as many bioactive components of human milk are of protein origin, implicit in their functionality is a resistance to digestion. Investigation of the nutritive value of human milk protein reveals that although most (95%) milk protein is available, a proportion does not contribute significantly to amino acid nutrition. To determine accurately the amounts of amino acids, and therefore protein, used for nutrition by the breast-fed infant, a correction needs to be made for bioavailability. A comparison of gross and available amino acid composition of mature human milk is made in Table 3. The total protein content of human milk consists of two major groups, caseins and whey proteins, as well as Table 3 The gross and available amino acida composition of mature human milk (mg 100 g1 proteinb) Amino acid
Gross
Available
Aspartic acid Threonine Serine Glutamic acid Proline Glycine Alanine Valine Isoleucine Leucine Tyrosine Phenylalanine Histidine Lysine Arginine Cysteine Methionine
9.2 4.7 5.0 18.4 8.5 2.3 3.8 6.4 5.9 9.7 4.2 3.9 2.3 6.8 4.0 2.2 1.6
9.1 4.6 4.2 16.8 8.4 2.1 3.4 5.1 5.1 9.3 3.8 3.8 2.3 6.2 3.1 2.6c 1.4
a
Includes free amino acids and peptides. Calculated from total g N 6.38. c Value for available is higher than gross due to correction for significant losses during the acid hydrolysis stage of amino acid determination. b
Milk | Human Milk Table 4 The types and amounts of protein (g 100 ml1) and non-protein nitrogen (mg 100 ml1) in human colostrum and mature milk
Casein proteins -Casein -Casein Whey proteins -Lactalbumin Serum albumin Lactoferrin Lysozyme Secretory immunoglobulin A/ immunoglobulin A Immunoglobulin G Immunoglobulin M Non-protein nitrogen (N) Urea N Creatine N Creatinine N Uric acid N Glucosamine -Amino N Ammonia N
Colostrum
Mature milk
0.26 0.12
0.3–0.5 0.1–0.3
0.36 0.04 0.35
0.2–0.3 0.03 0.1–0.3 0.05 0.05–0.1
0.2
0.034 0.012 48 12.1
0.5 14.2 4.5 0.2–0.8
0.001 0.002 50 15–25 3.7 3.5 0.5 4.7 13 0.2
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milk. -Lactoglobulin, prominent in bovine milk, does not occur in human milk. The predominant sources of amino acids for nutrition among the whey proteins are -lactalbumin and serum albumin. The other whey proteins, while contributing to nutrition to some extent, have a primary function in other roles. Lactoferrin, an iron-binding protein, has bacteriostatic properties that may impact on both the infant and the mammary gland. Lysozyme is capable of cleaving proteoglycans in the cell wall of certain bacteria, providing an antibacterial effect. The Igs in human milk are distinct from those found in blood serum, indicating selective synthesis in the mammary gland. sIgA, formed by the linking of two serum IgA molecules by disulfide bonds, is present at very high concentrations in colostrum, but the sIgA content declines rapidly to a much lower level within 14 days of lactation. sIgA protects the infant by blocking the attachment of pathogenic bacteria to the gut epithelium. Other Igs in human milk provide immune protection through activation of the complement system, or by promoting phagocytosis. Other proteins
some minor components (Table 1) and proteins present in the milk fat globule membrane. The ratio of whey proteins to casein changes during the initial stages of lactation, from 90:10 immediately post-partum, to 60:40 in mature milk and 50:50 in late lactation. These ratios directly reflect a decrease in the content of immune proteins (Table 4) in line with development of the infant’s own immune system. Casein
Human milk contains two ( and ) caseins, and is devoid of -casein, the predominant casein found in bovine milk. The casein pattern differs depending on the stage of lactation, with -casein detectable in human milk only from day 3–4 post-partum onward. A unique feature of human milk is the high degree of glycosylation of -casein (40–60% glycosylated) and other milk proteins (e.g., immunoglobulins (Igs) and lactoferrin). Glycosylated proteins exhibit anti-infective properties and enhance the absorption of some micronutrients. As part of the normal digestive process, the caseins in human milk release smaller peptides into the gut lumen. These casein fragments are thought to enhance the absorption of calcium by keeping it in solution in the gut lumen. Other casein-derived peptides have been linked to the regulation of intestinal motility and to the growth promotion of beneficial bacteria in the infant’s gut. Whey proteins
-Lactalbumin, lactoferrin, serum albumin, and secretory IgA (sIgA) are the main whey proteins found in human
In addition to casein and whey proteins, human milk contains other proteins, such as serum proteins, most of which are in the aqueous phase. The milk fat globule membrane proteins surround the lipid droplets, but are a minor fraction quantitatively (1–3% of total protein). However, these minor proteins can contribute in many ways to the health and well-being of the infant, for example, by facilitating transport of certain vitamins and minerals. Milk-borne peptide hormones and enzymes assist in the digestion of other milk components (e.g., bile salt-stimulated lipase (BSSL) and amylase). A detailed list of the hormones, both peptide and non-peptide, and enzymes found in human milk is given in Table 5.
Non-protein Nitrogen Compared with the milk of other mammalian species, human milk is notably different with regard to the NPN fraction. In human milk, the NPN fraction represents a very high percentage (20–25%) of total nitrogen (N). Urea N accounts for almost 50% of the NPN in human milk, with more than 200 compounds, including free amino acids, carnitine, taurine, amino sugars, nucleic acids, nucleotides, and polyamines, making up the remainder. In general, the total NPN of human milk remains constant throughout the lactation period, although there may be some variation in individual NPN components (Table 4).
584 Milk | Human Milk Table 5 Hormones and enzymes found in mature human milk Hormones Peptide Growth factors
Gastrointestinal regulators
Hypothalamus/hypophyseal hormones
Thyroid–parathyroid group
Non-peptide Epidermal growth factor Insulin Insulin-like growth factor 1 Nerve growth factor Transforming growth factor- Transforming growth factor-
Thyroid
Thyroxine Triiodothyronine
Adrenal
Cortisol
Sexual
Progesterone Pregnane3( )20( )-diol Estrogen
Gastrin Gastric inhibitory polypeptide Gastric-releasing peptide Neurotensin Peptide histidine-methionine Peptide tyrosine–tyrosine Somatostatin Vasoactive intestinal peptide Gonadotropin-releasing hormone Growth-releasing factor Growth hormone Prolactin Thyrotropin-releasing hormone Thyroid-stimulating hormone Calcitonin-like hormone Parathyroid hormone Parathyroid hormone-related peptide
Enzymes Phosphoglucomutase Fatty acid synthase Galactosyltransferase Bile salt-stimulated lipase Lysozyme
-Glutamyltransferase Protease inhibitors Lipoprotein lipase Alkaline phosphatase
Given that the total protein content of human milk is low compared to other species, yet supports adequate growth of the breast-fed infant, the suggestion has been made that the urea found in the NPN fraction of human milk is used by the infant for protein synthesis. This would have major implications for infant formula manufacturers, necessitating a total reevaluation of infants’ protein requirements. Isotopic tracer experiments with infants have shown that as much as 40% of labeled urea N was retained within the body. However, this may not directly reflect the incorporation of urea N into body protein, as NH exchanges occurring during transamination may in part explain the appearance of urea 15N in body proteins. This remains an important area for future investigation of infant protein metabolism.
Lactose synthase Thioesterase Amylase Proteases Peroxidase Platelet-activating factor acetylhydrolase Sulfhydryl oxidase Glutathione peroxidase Xanthine oxidase
Taurine and glutamine, two predominant free amino acids in human milk, are considered conditionally essential, as the infant appears to have a requirement for these amino acids above what the body can synthesize. Taurine plays an integral part in fat digestion by conjugating with bile acids to form bile salts. Although no direct link has been made between a deficiency in taurine and abnormal development of retinal and brain tissue in infants, the addition of taurine to infant formulae is now routine. Glutamine is involved in the cellular metabolism of enterocytes and in the immune response associated with inflammation and sepsis. During periods of rapid growth or tissue repair, a dietary source of nucleotides may be beneficial, as de novo synthesis of nucleotides can be a limiting factor.
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Table 6 Nucleotide content of human milk, and the amounts of specific nucleotides approved for inclusion in infant formulae
Nucleotide
In human milk (mg 100 ml1)
Adenosine diphosphate (ADP) Adenosine monophosphate (AMP) Cytidine diphosphate (CDP) Cytidine monophosphate (CMP) Guanosine diphosphate (GDP) Guanosine monophosphate (GMP) Inosine monophosphate (IMP) Uridine diphosphate (UDP) Uridine monophosphate (UMP)
69 175 474 461 96 138 228 174 179
Maximum to be added to infant formula (mg 100 kcal1)
1.5 2.5 0.5 1.0 1.75
Adapted from Lawrence and Lawrence (1994); infant formula inclusion rates from Rudloff S and Kunz C (1997) Protein and nonprotein nitrogen components in human milk, bovine milk, and infant formula: Quantitative and qualitative aspects in infant nutrition. Journal of Pediatric Gastroenterology and Nutrition 24: 328–344.
The infant is in a state of rapid growth and cellular development, and human milk is rich in nucleotides (Table 6). To date, however, no link between nucleotides in human milk and enhanced cellular function in the infant has been made. Despite this lack of direct evidence, some regulatory authorities have approved the inclusion of nucleotides in infant formulae (Table 6).
Table 7 Quantities of some carbohydrates found in human milk
Carbohydrate Besides water, lactose constitutes the largest component of human milk (Table 1); lactose concentration decreases as lactation progresses, in line with the decline in milk volume. Oligosaccharides constitute the next greatest fraction of carbohydrate in human milk, representing more than 27% of the carbohydrate in colostrum. This gradually decreases to 15–16% of carbohydrate as oligosaccharides in mature human milk. Most oligosaccharides found in human milk contain lactose at the reducing end, and may contain fucose or sialic acid at the non-reducing end. Compared to other mammalian milks, human milk is unique due to its content of complex oligosaccharides. Milk oligosaccharides provide a low osmolar source of calories for the infant, and have been classed as ‘soluble’ fiber because they appear to remain largely undigested in the infant’s gut. Human milk oligosaccharides contain significant quantities of sialic acid, which is essential for brain development, and they are also thought to stimulate the growth of bifidus bacteria and to inhibit the adhesion of pathogenic bacteria to epithelial surfaces. Approximately 130 oligosaccharides have been identified in human milk, some of which are listed in Table 7. Glucose and galactose are present in much smaller quantities in human milk, as are other carbohydrates such as monosaccharides, and peptide-bound and protein-bound carbohydrates.
carbohydrate oligosaccharid
Concentration mg 100 ml1
Lactose Oligosaccharides Lacto-N-tetraose Lacto-N-fucopentaose I Lacto-N-fucopentaose II Lacto-N-fucopentaose III Lacto-N-difucohexaose I NeuAc( 2–6)lactose NeuAc( 2–3)lactose NeuAc-lacto-N-tetraose a NeuAc-lacto-N-tetraose b NeuAc2-lacto-tetraose
6800 1200–1300 5–15 12–17 3–10 0.1–2 1–2 3–5 1–3 0.3–2 1–6 2–6
Adapted from Kunz C, Rodriguez-Palmero M, Koletzko B, and Jensen R (1999) Nutritional and biochemical properties of human milk. 1. General aspects, proteins, and carbohydrates. Clinics in Perinatology 26: 307.
Lipids The lipids in human milk occur in globular form, emulsified in the aqueous phase, and provide the infant with a major proportion (45–55%) of the energy needed to support growth. They also contain bioactive components important to the infant’s retinal and neural tissue development. The triacylglycerols (TGs) account for 98% of total fat, and 90% of this is fatty acids (FAs). Phospholipids (0.8%), cholesterol (0.5%), and other minor lipid components make up the remainder of the fat, which includes fat-soluble vitamins. In the first few weeks of lactation, the total lipid content increases from around 2 to 3.5–5% (Table 8). As the fat content increases, so too does the average size of the milk fat globules, which in turn results in a reduction in the ratio of phospholipids and cholesterol in TGs.
586 Milk | Human Milk Table 8 Lipid content of human colostrum and milk Lipid component
Colostrum
Mature milk
Total lipid (percentage of milk volume) Phospholipid (percentage of total lipid) Triacylglycerol (percentage of total lipid) Cholesterol (percentage of total lipid)
2
3–5
1.1
0.8
97–98
97–98
1.3
0.5
Table 9 An example of the fatty acid composition of human milk (wt%) Fatty acid
Colostrum
Mature milk
Saturated 10:0 12:0 14:0 15:0 16:0 17:0 18:0 20:0 22:0 24:0
0.5 2.3 5.3 0.3 26.2 0.4 7.5 0.3 0.2 0.3
1.1 4.8 6.7 0.3 21.8 0.3 7.5 0.2 0.1 0.1
Monounsaturated 14:1n9 0.1 16:1n9 2.1 18:1n9 34.7 20:1n9 1.3 22:1n9 0.3 24:1 0.5
0.3 2.7 33.0 0.6 0.1 0.1
Polyunsaturated 18:3n3 0.8 20:3n3 0.2 20:5n3 0.04 22:5n3 0.2 22:6n3 0.6 18:2n6 8.8 18:3n6 0.1 20:2n6 0.6 20:3n6 0.6 20:4n6 0.8 22:4n6 0.2 22:5n6 0.4
1.0 0.1 0.04 0.1 0.2 10.7 0.2 0.3 0.4 0.5 0.1 0.2
The TG composition in human milk is determined by the types and quantities of FAs esterified to the glycerol molecule. Human milk contains eight major FAs in amounts greater than 1% (Table 9), and is an excellent source of the essential FAs C18:2n6 and C18:3n3 and their long-chain derivatives C20:4n6 (arachidonic acid) and C22:6n3 (docosahexaenoic acid). The latter FAs, although not classed as essential for adults, are the only FAs utilized by the brain and are important structural components of the membrane systems of all tissues.
Approximately 60% of the C16:0 FAs in human milk are situated at the sn-2 position on the TG, which is a unique feature of the FAs in human milk. Lipolytic hydrolysis cleaves FAs from the sn-1 and sn-3 positions, and a C16:0 remains as a monoacylglycerol. This facilitates the absorption of C16:0 across the infant’s gut epithelium, improving overall lipid utilization to around 90–95%. Lipid digestion and absorption are enhanced further by four enzyme systems. These are, in the order of utilization, the gastric phase with lingual lipase and gastric lipase, and the intestinal phase with pancreatic lipase and BSSL. The gastric phase is important because the milk fat globule would otherwise resist the action of pancreatic lipase and BSSL. Unlike the other lipases, which are endogenous, BSSL is secreted in human milk specifically to assist the infant with milk fat digestion. Of all the components in human milk, the lipid fraction is the most variable. Changes in fat content occur not only during the first few weeks of lactation (Table 8), but also in milk secretions throughout the day, and even within a single feed, with the hindmilk being significantly higher in fat content than the foremilk. Variations also exist in the fat content of milk from individual women, although by far the greatest variation occurs in the types and amounts of FAs.
Micronutrients Vitamins
Most vitamins found in human milk increase in concentration as lactation shifts from colostrum to a mature secretion (Table 10), with the exception of carotenoids ( -carotene, -carotene, lutein, cryptoxanthin, and lycopene), retinol, and vitamin E, which decrease. Reduction in the concentration of some vitamins may not alter the total amount available to the infant, however, as any reduction may be offset by an increase in the quantity of breast milk consumed. The milk from healthy, well-nourished women usually contains sufficient quantities of most of the vitamins required by the infant. There are a few exceptions, however, when the amount in milk may not be adequate. Deficiencies in vitamin D, with consequent development of rickets, have been reported in breast-fed infants, directly linked to the mother’s diet and the infant’s lack of exposure to sunlight. To meet physiological requirements, vitamin K is routinely given to newborn babies because transplacental transfer is limited, and the level in milk is low. Minerals
In general, the macromineral content of human milk remains stable during the lactation period after an initial decrease in the concentration of chloride, potassium,
Milk | Human Milk Table 10 Micronutrient content of human colostrum and milk Micronutrient
Units l
Water-soluble vitamins Ascorbic acid mg Biotin mg Folic acid mg Niacin mg Pantothenic acid mg Riboflavin mg Thiamine mg Vitamin B6 mg mg Vitamin B12
1
Colostrum
Mature milk
60 0.6 5 0.6 1.8 300 19.5
50–100 6 80–133 1.6–6.0 2.0–2.5 400–600 200 0.1–0.3 1.0
0.5
Fat-soluble vitamins Carotenoids Retinol Vitamin D Vitamin E Vitamin K
mg mg mg mg mg
2 1.8
Major minerals Calcium Chloride Citrate Magnesium Phosphorus Potassium Sodium Sulfur
mg mg mg mg mg mg mg mg
300 813
Trace minerals Chromium Cobalt Copper Fluoride Iodine Iron Manganese Molybdenum Nickel Selenium Zinc
ng ng mg mg mg mg mg mg mg mg mg
10–15 2–5
36 140 652 502 277
0.4–0.8
0.75 5–12
40 8–12
0.2–0.6 0.3–0.7 0.4 3–8 3–15 278 426 500a 35 140 530 180 142 200–400 100–200 0.2–0.4 4–15 146 0.72 3–6 1–2 0.5–2 10–20 1–3
a Citrate is not classed as a mineral, but is soluble in water and binds some minerals, so is included in this table.
sodium, and sulfur (Table 10). Most of the sulfur in human milk is found in the sulfur-containing amino acids methionine and cysteine, with only a small fraction (10%) existing in salt form. The sulfur amino acids predominate in whey proteins, so a reduction in whey protein content during the first few days of lactation is matched by a similar reduction in sulfur content. Human milk has a higher calcium-to-phosphorus ratio (2:1) than the milk from other mammalian species (e.g., in bovine milk, Ca:P ratio is 1.1:1), although the absolute amounts of calcium and phosphorus are lower. A low phosphorus content suits the infant’s limited renal capacity. In addition, the contribution of phosphate to the buffering capacity of human milk is such that a low pH results, promoting the growth of beneficial gut bacteria.
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Human milk contains what would initially appear to be an inadequate level of iron, yet breast-fed infants rarely deplete their iron stores until after 4–6 months of age. Maximum utilization of the iron in human milk is achieved with the assistance of lactoferrin, most likely via specific protein receptors in the gut that link with lactoferrin and therefore facilitate iron absorption.
Bioactive Factors Human milk contains an optimal balance of the nutrients required by the infant and, in addition, it contains an array of bioactive factors. In the first instance, these factors assist in the production and secretion of milk components and provide immune protection of mammary tissue. Once ingested by the infant, however, bioactive factors actively promote growth and development, and afford the infant immune protection until its own defense system has matured. Even the major components in milk may have specific bioactive functions either before being digested or as a result of releasing bioactive components during the digestive process. The main sites of action for these bioactive factors are mucosal surfaces, although some may enter the circulation and have a systemic effect elsewhere in the body. The various physiological roles of milk components and their derivatives, either within the mammary gland or in the infant, are listed in Table 11. Intact cellular components are found in human milk (Table 12), and although their exact role within either the mammary gland or the recipient infant is not clearly defined, there does appear to be a degree of biological activity associated with them. The total count of cells is high in colostrum, with the numbers decreasing as lactation becomes established. More than 90% of the cells in human milk are macrophages and neutrophils, while the remainder are lymphocytes (80% of which are T cells) and epithelial cells. The primary function of milk leukocytes is thought to be as a means of defense within the mammary gland, and not necessarily to transfer maternal immunocompetence to the infant. Macrophages,however, have been suggested as a potential vehicle for the storage and transport of milk-borne Igs, and may be involved in the formation of lactoperoxidase and other factors that enhance the growth of intestinal epithelium and digestive capacity. The lymphocytes present in human milk are part of the immunological system that synthesizes IgA, and considerable research data are available to support the suggestion that lymphocytes in colostrum and milk provide the infant with immunological benefits.
588 Milk | Human Milk Table 11 Physiological roles of components in human milk, either within the mammary gland or following ingestion by the infant Physiological function
Components in human milk
Within the mammary gland Component synthesis/secretion Immune effect/anti-infection/ anti-inflammatory
-Lactalbumin, lipoprotein lipase, xanthine oxidase Lactoferrin, lysozyme, -glutamyltransferase, cytokines
Within the infant Nutrition Enhancing nutrient availabilitya Gut colonization Gastrointestinal development Brain development Immunity Anti-infective Anti-inflammatory Immunomodulating
- and -casein, whey proteins, lactose, fats, minerals, and vitamins Bile salt-stimulated lipase, milk amylase, lactoferrin (Fe), glutamine (Zn), -lactalbumin (Ca), casein phosphopeptides (Ca), folate-binding protein Bifidus factor peptide, -casein oligosaccharides Epidermal growth factor, insulin-like growth factor, growth factor-binding proteins, hormones, enzymes, nucleotides, oligosaccharides, amino sugars Docosahexaenoic acid, other long-chain polyunsaturated fatty acids, taurine, carnitine Immunoglobulins, leukocyte-stimulating factors, T lymphocytes, macrophages, immunopeptides Lactoferrin, lysozyme, free fatty acids, cell adhesion molecules, cytokines, mucins, glycoproteins, glycopeptides, oligosaccharides Vitamins, prostaglandins, growth factors, cytokines (interleukin-10), transforming growth factor- Interleukins, tumor necrosis factor-
a
Nutrient that has increased availability due to a particular component in human milk is given in parentheses.
Table 12 Cellular components (number per ml) in human colostrum and milk Cell type
Colostrum
Mature milk
Total cells Macrophages Neutrophils Lymphocytes
2840 1490 1375 250
51 52 8 1
Adapted from Jensen R (ed.) (1995) Handbook of Milk Composition. San Diego, CA: Academic Press.
Factors that Influence Milk Volume and Composition Factors Affecting Milk Volume In women, the principal determinant of milk volume is infant demand. Comprehensive review of milk volume from exclusively breast-feeding women throughout the world has revealed that milk volume, 6 months into lactation, is incredibly constant, with an average of 800 ml (range 500–1200 ml) of milk transferred from mother to infant each day. The potential for milk production far exceeds this amount, however, given that mothers of twins or triplets can adequately nourish their children with human milk. The energy density of human milk, in part, influences milk production, as infants consuming lower calorific milk will suckle more to obtain the energy required, in turn stimulating greater milk production. In one study, women with a low body fat content produced as much as 15% more milk as a result of increased infant demand. As soon as the weaning process begins, and human milk is
substituted by either supplementary formula or complementary solids, milk production declines, indicating yet again the dominant role the infant plays in controlling milk volume.
Major Factors Affecting Milk Composition Stage of lactation
Major compositional changes that occur from parturition through to day 21 of lactation have been discussed. The composition of human milk continues to change throughout lactation, however, with a gradual reduction in component content, although some components, for example, lysozyme, may increase. During weaning, changes in composition can occur depending on the time taken to wean. When weaning is rapid (several days/weeks), lactose and potassium contents decrease, while sodium, chloride, fat, and total protein (specifically the whey proteins) increase. Gradual weaning (over several months) results in an increase in sodium (220% from baseline), iron (172%), and protein (142%), no change in calcium, and a decrease in zinc (58% from plateau). Both rapid and gradual weaning produce a decrease in lactose content. Prematurity
Relative to normal human milk, the milk of mothers delivering prematurely contains higher concentrations of protein (1.8–2.4 g 100 ml1), short-, medium-, and long-chain FAs, calcium, phosphorus, sodium, chloride, magnesium, and iron, which is related to the low volume produced. Cellular components are also significantly
Milk | Human Milk
greater in premature colostrum (total cells 6800 per ml). Levels of the other components in premature milk do not differ markedly from those found in normal human milk. Changes during a feed and during the day
Changes that occur in composition during a feed relate more to the physicochemical nature of fractions in the milk, rather than a selective change in individual components. In general, most components remain relatively constant throughout a feed. The lipid content increases rapidly toward the end of a feed, however, reflecting a propensity of the mammary gland tissue to retain the lipid globules. Subjective evidence from breast-feeding women suggests that milk composition changes during the day, especially toward evening, when the infant appears to be less satisfied by milk consumed. An unsettled infant is more likely to be due to insufficient milk volume, however, as lipid content actually increases toward the end of the day (by two- to fivefold). No significant changes occur during the day in other components. Nationality/age/parity
The composition of milk from women from different geographic, ethnic, and socioeconomic backgrounds is remarkably similar, particularly in reference to the macronutrients. Any differences observed are more likely to be the result of dietary variation, rather than genetic modification of composition. Milk from teenagers has a significantly lower concentration of lactose and some macrominerals compared to adults, but diet may play an important part in this difference as the diet of teenagers is often suboptimal. Parity may have a significant effect on milk composition, with primiparous women having higher concentrations of protein and fat, and multiparous women experiencing a reduction in the content of major components in milk with each successive lactation. Maternal diet
Two of the most significant factors influencing milk composition are the nutritional status of the mother and her ongoing diet. An inadequate nutritional status will adversely affect milk volume and the content of milk minerals such as iron and selenium. Severe malnutrition and diminished nutritional status will cause a decrease in the protein and fat content of milk. The components most affected by maternal diet are FAs, vitamins, and some macro- and micro-minerals. The FA content of human milk can undergo pronounced changes that directly reflect the FA content of the diet. For example, the C18:2n6 content of milk from vegetarians is high compared to the milk from women consuming an omnivorous diet. Also, a low-fat diet will result in a
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marked increase in medium-chain FAs as the de novo synthesis of FAs in the mammary gland increases. Exercise
Recent evidence suggests that the lactic acid content of human milk can change following exercise, with a decline in infant acceptance of milk secreted immediately after exercise. Exercise regimens at 100, 75, or 50% of VO2 max resulted in increases in the lactic acid content of milk sampled immediately after exercise, although only 100% intensity exercise produced milk with a lactic acid content significantly different from baseline. Thus, breast-feeding women can complete a moderate exercise program without the fear that the infant will reject their milk. Illness and metabolic disorders Mastitis
Infection of the mammary tissue, with cellulitis and, more rarely, abscess formation, can alter milk composition to such an extent that the infant’s nutrition is compromised. During a bout of mastitis, milk volume and lactose content decrease, while the levels of sodium and chloride increase, giving the milk a much saltier taste. Immune factors such as sIgA, lysozyme, and lactoferrin also increase in milk from mastitic tissue, as part of the mammary gland immune response. Insulin-dependent diabetes mellitus
The delayed lactogenesis associated with mothers with insulin-dependent diabetes mellitus (IDDM) results in a lower volume of milk being consumed by the infant in the first few days following birth. Initially (<5 days’ lactation), the milk from an IDDM mother has a composition different from that of normal milk, with less lactose and fat, and more total nitrogen. Once lactation has been established, however, most of these differences diminish. The FA content of milk from IDDM women continues to differ throughout the lactation, with more C10, C12, and C14, and less C20 and C22, FAs. Drug use
The degree to which a drug taken by the mother will affect the breast-fed infant depends entirely on the class of drug. Detailed information on the transfer of pharmaceutical drugs in human milk is available. Recreational drugs, such as alcohol, caffeine, nicotine, and marijuana, also appear in human milk and can affect both milk volume and composition. Human milk from a mother who smokes not only contains many chemical by-products, including nitrates, nitrites, lead, and cadmium, but is also likely to have lower levels of certain vitamins. Exposure of an infant to drugs (both medical and recreational) in milk can be minimized by advising the mother to take any
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medication immediately after a breast-feed, and/or just before the infant is due to have a lengthy sleep period.
Human Milk Banking Human milk is usually collected and stored in milk banks to assist in the nourishment of premature infants. Implicit in this action of ‘banking’ milk is the need to maintain functional integrity in the milk. Numerous steps in the process can affect the composition of human milk, including the suitability of the donor, and her dietary regimen; the method of milk collection and the type of container chosen to collect the milk; the length of time and temperature at which the milk is stored; and the methods used to thaw, mix, and reheat the milk prior to a feed. The Human Milk Banking Association of North America has compiled a set of guidelines for the establishment and operation of a donor milk bank. See also: Dehydrated Dairy Products: Infant Formulae. Mammals. Milk: Colostrum.
Further Reading Committee on Drugs (1994) The transfer of drugs and other chemicals into human milk. Pediatrics 93: 137–150. Hamosh M (2001) Bioactive factors in human milk. Pediatric Clinics of North America 48: 69–86. Jensen R (ed.) (1995) Handbook of Milk Composition. San Diego, CA: Academic Press. Jensen R (1999) Lipids in human milk. Lipids 34: 1243–1271. Kunz C, Rodriguez-Palmero M, Koletzko B, and Jensen R (1999) Nutritional and biochemical properties of human milk. 1. General aspects, proteins, and carbohydrates. Clinics in Perinatology 26: 307–333. Lawrence RM and Lawrence RA (1999) Biochemistry of human milk. Breastfeeding: A Guide for the Medical Profession, 5th edn., pp. 95–155. St Louis, MO: Mosby. McVeagh P and Brand Miller J (1997) Human milk oligosaccharides: Only the breast. Journal of Paediatrics and Child Health 33: 281–286. Picciano MF (2001) Representative values for constituents of human milk. Pediatric Clinics of North America 48: 53–67 (appendix 263–264). Rodriguez-Palmero M, Koletzko B, Kunz C, and Jensen R (1999) Nutritional and biochemical properties of human milk. 2. Lipids, micronutrients, and bioactive factors. Clinics in Perinatology 26: 335–359. Rudloff S and Kunz C (1997) Protein and nonprotein nitrogen components in human milk, bovine milk, and infant formula: Quantitative and qualitative aspects in infant nutrition. Journal of Pediatric Gastroenterology and Nutrition 24: 328–344.