Zinc and copper in human milk

Studies of zinc and copper contents of breast milk were performed in 34 mothers during their lactation period until 5 days postpartum. They experienced no problems during pregnancy and delivery. All women had full term healthy infants, 22 were multiparous and 12 primiparous. The highest zinc concentration was found in the colostrum, and subsequently the concentration declined. Copper content was stable during the 5 days of lactation. Significantly lower zinc concentration and higher copper concentration were found in the serum of lactating women 3 days post partum, when compared with non-lactating women. Factors including lactation history, age of lactating women, serum levels of zinc and copper did not affect the contents of these trace minerals in breast milk.     

Multielement determination in small samples of human milk by inductively coupled plasma atomic emission spectrometry

Inductively coupled plasma atomic emission spectroscopy (ICP-AES) was used for routine analysis of small samples of human milk. The concentrations of calcium (Ca), copper (Cu), iron (Fe), magnesium (Mg), manganese (Mn), phosphorus (P), and zinc (Zn) were determined in 203 milk samples from postpartum women at different stages of lactation after stepwise digestion in HNO₃, HCIO₄, and H₂O₂ under heat. Validation of the procedure was achieved using certified reference material of bovine liver (NBS 1577) with mean recoveries of 103.5%. The concentrations of the above elements in milk matrix were comparable with previously reported values. The analytical results from breast milk will provide reference information for mineral studies of Brazilian mothers and breast-fed infants.     

Copper, iron, and Zinc contents in human milk during the first three months of lactation

The aim of the study has been to analyze the evolution of copper, iron, and zinc contents in human milk, from colostrum to the third postpartum month, following a longitudinal design, under specific conditions of sample collection and to apply an analytical procedure previously optimized to reduce any variation outside physiological lactation. The copper, iron, and zinc concentrations in 144 milk samples from 39 healthy puerpera women, were analyzed in five stages by flame atomic absorption spectrometry, following a standardized protocol. Copper presented a gradual decrease from 0.38 mg/L to 0.19 mg/L by the 90th day; the particular analysis from colostrum to transitional milk manifested the following two tendencies. Whereas an increase from 0.19 to 0.42 mg/L was observed in some women, a decrease from 0.53 to 0.45 mg/L was detected in others; therefore, copper presented two significant behaviors in the evolution from colostrum to transitional milk. In both cases, the evaluated changes were significant. The iron content varied from 0.56 to 0.40 mg/L by the 30th day, remaining constant until the first trimester concluded. The average zinc concentration decreased sharply from 7.99 to 3.3 mg/L on d 15; the rate of decrease slowed down gradually until 1.05 mg/L.     

A study of factors that may influence the determination of copper, iron, and zinc in human milk during sampling and in sample individuals

The aim of this study was to establish the possible effects of the sampling protocol (between-breast, within-feed, and diurnal differences) and the mother’s personal factors (age, parity, iron supple-mentation, smoking habits, and lactation period) on the copper, iron, and zinc contents in human milk. One hundred thirty-six human milk samples identified by their origin and sampling conditions were analyzed. The samples were obtained from the 2nd to 15th d postpartum from 62 women. The data on the individuals required for the study were available. Mineral determinations were analyzed by flame atomic absorption spectrometry following a standarized protocol. The results showed that iron contents were higher in hind-milk samples and at the nighttime feeding and depended on the breast from which the sample was taken. The copper and zinc concentrations showed no significant variations. There was no significant relationship among the mothers’ age, parity, smoking habits, iron supplementation, and copper content. Milk from older women had lower zinc contents than that of younger women. Increased amounts of iron were found in multiparous women. Between colostrum and transitional milk, a sharp decrease in zinc content was observed, whereas copper and iron contents remained constant. All of these results make it clear that standardized sampling protocols are needed in order to obtain comparable values.     

Mineral- and trace element concentrations in human breast milk,placenta, maternal blood,and the blood of the newborn

The concentrations of the essential trace elements Cu, Fe, and Zn, and of the mineral elements Ca, K, Mg, and P during the perinatal period in human placenta and in the blood of the mother and the newborn (cord blood) were determined. Breast milk (colostrum and transitory milk) was also included to permit correlations between the different compartments. No correlations were found. The uptake by nutrition and the body-pools of the mother and their mobilization for these elements seem to be sufficiently high for an adequate supply of the fetus and the milk in the geographical region of Munich (Bavaria, FRG) under these investigations. Differences in the mineral- and trace element concentrations of colostrum and transitory milk for the elements P and Zn and to a lesser extent for Ca and Mg were observed. Additionally, breast milk samples from different geographical regions in Bavaria were investigated. Results for the heavy metals Cd, Hg, and Pb, and for the essential trace element Se are also presented for these samples, and can be seen as a reflection of the overall environmental and dietary influences during pregnancy in these geographical regions. ICP (Inductively Coupled Plasma)-emission spectrometry was used for the determination of the elements Cu, Fe, Zn, Ca, K, Mg, and P. For the additionally given elements in the milk-samples anodic stripping voltammetry (DPASV) (Cd, Pb), hydride atomic absorption spectrometry (AAS) (Se), and cold vapor AAS (Hg) were applied.     

Role of Milk Whey in the Transmission of Human Cytomegalovirus Infection by Breast Milk

Breast-fed infants are susceptible to human cytomegalovirus (HCMV) infection via breast milk. In our previous study, HCMV was isolated more frequently from breast milk at later than one month after delivery than from colostrum or early breast milk. To clarify the role of milk cells and whey in vertical infection by breast feeding, we separated breast milk into milk cells and whey and examined each fraction for the presence of HCMV. We collected breast milk from mothers who breast-fed their infants (aged from 3 days to 2 months). The breast milk was centrifuged and separated into the middle layer (layer of milk whey) and the pellet (containing milk cells). We attempted to isolate HCMV from whey and to detect HCMV immediate early (IE) DNA in both milk whey and cells. HCMV was isolated from 7 out of 35 (20.0%) whey samples and HCMV IE DNA was detected from 15 out of 35 (42.9%) whey and/or milk cells. Detection rates of HCMV IE DNA in the whey layer and milk cells were 39.1% (25 out of 64) and 17.2% (11 out of 64), respectively. HCMV IE DNA was not detected in colostrum, but was detected in breast milk samples one month after delivery. Therefore, cell-free HCMV shed into milk whey may have a more important role in vertical infection by breast milk than cell-associated HCMV in the milk.     

Seasonal changes in the mineral composition of colostrum and milk whey of cows

It is shown that in summer the level of sodium, potassium and calcium in the colostrum and milk serum of cows for the first ten days of lactation is 1.5-2.0 times higher and the content of inorganic phosphorus lower than the analogous indices for cows in winter-spring. The concentration of iron, copper, zinc and chlorides in the colostrum and milk serum of cows in different seasons is unchanged.     

Environmental and injected cadmium are sequestered by two major isoforms of basal copper, zinc-metallothionein in gibel (Carassius auratus langsdorfi) liver

Gibels were exposed to cadmium in their aquarium at a concentration of 10 micrograms Cd/l for up to 39 weeks. Distributions of cadmium, copper and zinc in the liver soluble fraction were determined along with sulfur by high performance liquid chromatography-inductively coupled argon plasma-atomic emission spectrometry. Cadmium was sequestered by the two major isoforms of gibel metallothionein as in the case of cadmium injected intraperitoneally into gibel. Several peaks with cadmium, copper, zinc and sulfur were observed other than the two major isoforms and their relative ratios were different between the control and cadmium-exposed fishes.     

Bioavailability of zinc and its binding to casein in milks and formulas

Differences in zinc bioavailability among milk and formulas may be attributed to binding of zinc to various ligands. We determined the distribution of zinc and protein at different pHs and zinc and calcium concentrations. We used radiolabelled cow's milk, human milk, whey-predominant (WPF) and casein-predominant (CPF) infant formula. Lowering the pH changed zinc and protein distribution: zinc shifted from pellet (casein) to whey in cow's milk, from fat to whey in human milk and from fat and pellet to whey in formulas. Protein shifted from whey to pellet in human milk and from whey and pellet to fat in formulas. Increasing zinc and calcium concentrations shifted protein and zinc from pellet to whey for cow's milk and from whey and pellet to fat for the formulas. Protein distribution was not affected by calcium or zinc addition in human milk or CPF, while zinc shifted from whey to fat in human milk and from fat and pellet to whey in CPF. Zinc and calcium binding to isolated bovine or human casein increased with pH. At 500 mg/L of zinc, bovine casein bound 32.0 +/- 1.8 and human casein 10.0 +/- 0.9 mg zinc/g protein. At 500 mg/L of calcium, calcium was preferentially bound over zinc. Adding calcium and zinc resulted in 32.0 +/- 1.8 mg zinc/g bound to bovine casein and 17.0 +/- 0.8 mg zinc/g to human casein, while calcium binding was low. Suckling rat pups dosed with 65Zn labelled infant diets were killed and individual tissues were gamma counted. Lower zinc bioavailability was found for bovine milk at pH = 4.0 (%65Zn in liver = 18.7+1.4) when compared to WPF (22.8 +/- 1.6) or human milk (26.9 +/- 0.8). Lowering the pH further decreased zinc bioavailability from human milk, but not from cow's milk or WPF. Knowledge of the compounds binding minerals and trace elements in infant formulas is essential for optimizing zinc bioavailability.     

Detection of sulfur-containing compounds in control and cadmium-exposed rat organs by high-performance liquid chromatography--vacuum-ultraviolet inductively coupled plasma-atomic emission spectrometry (HPLC-ICP)

Sulfur-containing compounds in biological samples were separated by high-performance gel permeation chromatography and detected with a vacuum-ultraviolet inductively coupled plasma-atomic emission spectrometer. Distribution profiles of sulfur in the supernatants of liver, kidney, spleen, lung, and pancreas of control and cadmium-exposed rats were determined along with cadmium, copper, iron, phosphorus, and zinc profiles. Changes in sulfur distribution were induced by cadmium exposure not only in the metallothionein fraction, but also in the high-molecular-weight protein fraction, indicating the effect of cadmium exposure on diverse endogenous sulfur-containing compounds. Glutathione and taurine also were detected simultaneously as distinct peaks.     

Changes of insulin concentration in colostrum and milk of women, cows and sows

The method of whey extraction in order to determine insulin concentration in colostrum and milk was worked out, checked and applied. The observation of insulin changes showed high and similar insulin concentration in whey samples of women, cows and sows colostrum on the day before and in the day of parturition. One day after delivery hormone concentration maintained high in case of women and sows but in cow colostrum it decreased to 1/12 comparing to the level on the parturition day. During the postpartum following days it was decreasing gradually in the colostrum of all three examined species; the process was very fast in cows, slower in women and the slowest in sows.     

Trace elements in human milk: Correlation with blood levels, inter-element correlations and changes in concentration during the first month of lactation

Using inductively coupled plasma mass spectrometry (ICP-MS) based analytical procedures, the concentration of several trace elements (Mn, As, Pb, Co, Ni, Cu, Zn and Se) was determined in human milk samples collected from a group of healthy lactating Portuguese women (n=44), both on the 2nd day postpartum (i.e., colostrum; n=34) and at 1 month postpartum (i.e., mature milk; n=19). Blood samples (n=44), collected on the 2nd day after parturition, were also analyzed for the same trace elements. No major correlations were observed between the levels of the analyzed trace elements in blood and colostrum samples. All the studied elements, except for Co, Pb and Ni, showed a significant trend for a decrease in concentration in milk during the first month of lactation. This trend was more pronounced for Zn and Se, whose levels decreased to approximately 23% and 44% of their initial mean concentration, respectively. With the exception of Co (r=0.607) and Zn (r=0.487), no significant correlations were observed when comparing the levels of each trace element between samples of colostrum and mature milk. Several inter-element correlations were found within each type of milk sample. The most significant were: (i) Se vs Cu (r=0.828) and Se vs Co (r=0.605) in colostrum samples and (ii) Ni vs Pb (r=0.756), Ni vs Mn (r=0.743) and Se vs Co (r=0.714) in mature milk samples. An inverse correlation between Zn and Se was also found in both types of milk sample; however, it only reached statistical significance for mature milk (r=-0.624).     

Identification of lactoperoxidase in mature human milk

Myeloperoxidase (MPO) derived from milk leukocytes and lactoperoxidase (LPO) secreted from the mammary gland have been identified previously in human colostrum. These peroxidases are known to play host defensive roles through antimicrobial activity. The goals of this study were to measure the peroxidase activity in mature human milk and to characterize the enzyme responsible for the activity. As determined using 3,3',5,5'-tetramethylbenzidine as substrate, whey prepared from human milk samples obtained 1 and 5 months postpartum showed levels of peroxidase activity equivalent to 0.13 +/- 0.18 and 0.24 +/- 0.21 microg/mL bovine LPO (bLPO; n = 13), respectively. Whey from early milk was fractionated into two peaks of peroxidase activity by cation-exchange chromatography; the peroxidase in the first peak was sensitive to dapsone, which is an inhibitor of LPO, whereas the second peroxidase was not. Whey from mature milk showed only the first peak. Purified bLPO and MPO showed chromatographic behaviors that were similar to the first and second peaks, respectively. The dapsone-sensitive peroxidase from mature milk was further purified (952-fold from whey) by hydrophobic interaction chromatography. This preparation showed two bands with molecular masses of 80 and 90 kDa by polyacrylamide gel electrophoresis and immunoblotting using an antibody against bLPO. After deglycosylation, two distinct proteins with lower molecular weights were observed. Amino acid sequencing indicated that both of these proteins are LPO. These results provide evidence that LPO is present in mature human milk and that it is responsible for most of the peroxidase activity in mature milk.     

Free zinc concentration in bovine milk measured by analytical affinity chromatography with immobilized metallothionein

A new analytical affinity chromatography method was developed for measuring the free [Zn²⁺] concentration in bovine milk. The column was generated by immobilizing avidin and attaching biotinylated metallothionein (MT) on controlled-pore glass beads. Zinc bound to the MT column at physiological free [Zn²⁺] concentration and was dissociated again in an elution buffer of pH 2. The distributions of extrinsically added⁶⁵Zn and native zinc in different fractions of milk were virtually identical, validating the use of extrinsic labeling in studies of the free [Zn²⁺] concentration in milk. Extrinsically labeled whey fractions were mixed with standard solutions whose free [Zn²⁺] concentrations were calculated by computer model.⁶⁵Zn retained by the column provided an indication of free [Zn²⁺] concentration in the mixture, and by interpolation, in the original milk. The free [Zn²⁺] concentration measured by the affinity chromatography method in the milk of a group of six cows was 90.4±29.7 pM. This value is similar to estimates of free [Zn²⁺] concentrations in other biological fluids by entirely different methods. Measurement of free [Zn²⁺] may be helpful in understanding the physiology and biochemistry of zinc metabolism.     

Role of zinc in regulating the levels of hepatic elements following nickel toxicity in rats

This study was designed to determine the protective effects of zinc on the hepatotoxicity induced by nickel in rats. Female Sprague-Dawley (SD) rats received either nickel sulfate alone in the dose of 800 mg/L nickel in drinking water, zinc sulfate alone in the dose of 227 mg/L zinc in drinking water, and nickel plus zinc or drinking water alone for a total duration of 8 wk. The effects of different treatments were studied on activities of rat liver marker enzymes like alkaline phosphatase (ALP), alanine aminotransferase (ALT), and aspartate aminotransferases (AST) and on the status of essential elements in rat liver. The study revealed a significant increase in the activities of enzymes ALP and ALT in rats subjected to nickel treatment. Interestingly, zinc supplementation to rats treated with nickel brought back the raised activities of these enzymes to within normal limits. Further, the levels of elements in liver that include zinc, copper, selenium, and potassium were found to be significantly suppressed following nickel treatment, whereas the levels of iron and sulfur were elevated. However, zinc treatment alone did not cause any appreciable change in the concentration of these elements. To the contrary, when zinc was given to nickel-treated rats, the concentrations of zinc, copper, potassium, and phosphorus were not significantly different from that of normal controls, whereas the levels of iron, selenium, and sulfur were improved in comparison to nickel-treated rats but were not within the normal limits. The present study concludes that zinc has the ability to maintain the levels of hepatic elements and has bearing in regulating the liver functions by maintaining the activities of marker enzymes in conditions of nickel toxicity.     

Daily dietary intake of copper,zinc, and selenium of exclusively breast-fed infants of middle-class women in Burundi,Africa

Copper (Cu), zinc (Zn), and selenium (Se) in human milk of middle-class Burundian women during the first 10 mo of lactation have been determined. Wet acid digestion, using nitric and perchloric acids, and atomic absorption spectrometric analysis have been used. Daily intakes have been calculated and proven to decrease from 0.39±0.05 (colostrum) to 0.16±0.02 (mature milk), 2.3±0.3 (colostrum), to 1.2±0.2 mg (mature milk) and 10.9±1.5 (colostrum) to 5.3±0.8 μg (mature milk) for Cu, Zn, and Se, respectively. Since values for this African country are nonexistent, intake levels are compared with literature data and found to be somewhat higher than those observed in other poorly nourished countries. The recommended safe and adequate daily intake for infants of 0–6 mo of age, as proposed by the National Research Council of the USA, is only met for Burundian infants <1 mo of age. The function of copper (Cu) and zinc (Zn) as essential trace elements has been known for quite a number of years (1). Also, selenium (Se) is a trace element essential for the activity of glutathione peroxidase (2) and type I iodothyronine 5-deiodinase (3). For all three elements, an adequate intake is necessary for satisfactory infant growth and development (4). In view of the almost total lack of relevant data on Burundi (Africa), we have determined Cu, Zn, and Se in human milk of middle-class Burundian women during the first 10 mo of lactation (5). The aim of this study is to assess infants' elemental intake for this country and compare this with literature data on trace elemental intake of exclusively breast-fed infants.     

Presence of CTAK/CCL27, MCP-3/CCL7 and LIF in human colostrum and breast milk

Human colostrum and breast milk are known to contain high levels of cytokines and chemokines, which are thought to contribute to the development of the newborn. The aim of this study was to investigate the difference in the presence and levels of 21 soluble cytokines and chemokines in paired samples of human colostrum (day 2 after delivery) and breast milk (day 4–5 after delivery) by using the multiplex technology. Of the 21 cytokine investigated in 10 pairs of samples, only β-NGF was absent in both colostrum and milk, while INF-α2, SCF and TNF-β were present in colostrum but not in human milk. As a general rule, colostrum contained higher concentrations of cytokines and chemokines with respect to breast milk. The majority of cytokines, detected in colostrum alone or in colostrum and human milk (IL-1α, IL-2Rα, IL-3, IL-16, IL-18, GRO-α, HGF, IFN-α2, M-CSF, MIF, MIG, TNF-β, SDF-1α, TRAIL) have been described in previous studies, while for the first time we describe the presence of additional cytokines either in colostrum alone (SCF) or in both colostrum and breast milk (CTAK/CCL27, MCP-3/CCL7, LIF). Our data confirm and expand previous studies showing that some cytokines/chemokines, which might contribute to the development of the gastro-intestinal and nervous systems, are overexpressed in human colostrum and breast milk, and might contribute to the development of these systems.     

Gel electrophoretic analysis of proteins in human milk and colostrum

Protein patterns of skim milk and fat globule phases were relatively unchanging throughout 9 months lactation, the only notable differences being a rise in lysozyme and a decline up to 6 months with increase thereafter in lactoferrin. Profound changes in protein patterns of the breast secretion were observed during the first week postpartum. True colostrum, in which peptides of sIgA dominate the protein patterns, exists for at most the 2 initial days of secretion. Proteins of mature milk are not coordinated to appear in the colostral secretion simultaneously. A band corresponding to alpha-lactalbumin is present from the initial secretion; that for beta-casein emerges approximately 2 days layer.     

Comparison of the levels of the growth factors in umbilical cord serum and human milk and its clinical significance

The process of the growth of the fetus begins in the uterus and gets further accelerated following the birth, especially during initial few months. The role of the growth factors in the physiology of the cellular growth is already well established. Vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) seem to be imperative for angiogenesis, cell development and proliferation as well as maintenance of the tissues. The levels of these factors in the maternal serum during pregnancy as well as during postpartum period are insignificant. Consequently, we hypothesized that the fetus receives moderate supply of these growth factors from the placenta during its stay in the uterus. This supply gets further augmented during the postpartum period through the different source, i.e. mother's milk. To study this physiological transition of the source of the growth factors from the placenta to the breast milk, the concentrations of VEGF and HGF in the cord serum of full term neonates and that in the breast milk of the corresponding mothers were analyzed during ELISA. The human milk, especially the colostrum revealed significantly higher levels of VEGF and HGF (1541.759 ± 119.349 pg/ml and 7129.249 ± 273.472 pg/ml) than cord serum (16.632 ± 0.773 pg/ml and 2581.6 ± 108.275 pg/ml) respectively. The multifold higher levels of VEGF observed in colostrum probably correlates with its high neonatal requirement for the maturation of the gastrointestinal epithelium following birth. The higher levels of both the growth factors in the breast milk than those observed in the cord serum probably explain their higher needs by the neonates for immunological protection, protein synthesis and neurocognitive development. The observations of the present study strengthen the policy of the colostrum feeding, which is promoted by organizations like World Health Organization (WHO). This study further documents the fact that the commercial milk formulae cannot replace the human milk.     

Heroin dependence effects on some major and trace elements

In this study, 56 (14 control and 42 addicts) adult human subjects of both sexes of different periods of heroin dependence were subjected to the measurement of whole blood, serum, and red blood cell levels of some trace elements (zinc, managanese, iron copper, and bromine), as well as some major elements (phosphorus, sulfur, calcium, potassium, and chlorine). This was done by the energy-dispersive X-ray fluorescence (EDXRF) technique, in which copper and bromine showed a significant rise in whole blood (male) (22 and 32%, respectively), while zinc, iron, manganese, calcium, sulfur phosphorus, potassium, and chlorine showed a significant drop (49, 8, 25, 34, 21, 51, 61 and 72%, respectively) in proportion to the period of heroin intake (6 yr) and in comparison with the control group. No significant sexual variation has been reported.