Macronutrient Content Of Milk Research Articles

Got (Optimal) Milk? Pooling Donations in Human Milk Banks with Machine Learning and Optimization

Problem definition: Human donor milk provides critical nutrition for millions of infants who are born preterm each year. Donor milk is collected, processed, and distributed by milk banks. The macronutrient content of donor milk is directly linked to infant brain development and can vary substantially across donations, which is why multiple donations are typically pooled together to create a final product. Approximately half of all milk banks in North America do not have the resources to measure the macronutrient content of donor milk, which means pooling is done heuristically. For these milk banks, an approach is needed to optimize pooling decisions. Methodology/results: We propose a data-driven framework combining machine learning and optimization to predict macronutrient content of donations and then optimally combine them in pools, respectively. In collaboration with our partner milk bank, we collect a data set of milk to train our predictive models. We rigorously simulate milk bank practices to fine-tune our optimization models and evaluate operational scenarios such as changes in donation habits during the COVID-19 pandemic. Finally, we conduct a year-long trial implementation, where we observe the current nurse-led pooling practices followed by our intervention. Pools created by our approach meet clinical macronutrient targets approximately 31% more often than the baseline, although taking 60% less recipe creation time. Managerial implications: This is the first paper in the broader blending literature that combines machine learning and optimization. We demonstrate that such pipelines are feasible to implement in a healthcare setting and can yield significant improvements over current practices. Our insights can guide practitioners in any application area seeking to implement machine learning and optimization-based decision support. History: This paper has been accepted as part of the 2023 Manufacturing & Service Operations Management Practice-Based Research Competition. Supplemental Material: The e-companion is available at https://doi.org/10.1287/msom.2022.0455 .

PTFS05-07-23 The Association of Human Milk Macronutrient Content With Early Childhood Growth and Neurodevelopment in Rural Bangladesh

PTFS05-07-23 The Association of Human Milk Macronutrient Content With Early Childhood Growth and Neurodevelopment in Rural Bangladesh

Breast-milk composition in women with cystic fibrosis: report of two cases and a review of the literature

Breast milk was collected from two women with cystic fibrosis (CF) and data from the literature was reviewed. The sodium concentration was within normal limits, 11-24 mmol/L in colostrum and 7-8 mmol/L in mature milk. Normal physiologic changes in milk composition after parturition (decreases in Na and increases in lactose) and during individual feeding periods (increases in fat and decreases in protein) were observed. Concentrations of milk protein, fat, and sugars were normal when the pulmonary disease of these patients was mild. During exacerbations of pulmonary disease, the concentrations of milk macronutrients were reduced. Milk secreted by women with CF appears to be physiologically normal, safe for the infant, and breast-feeding by mothers with CF should no longer be discouraged. Variations in the macronutrient content of CF milk warrants routine monitoring of the mother with CF and the breast-fed infant, especially during exacerbations in the pulmonary aspects of this disease.

Carbohydrate content of human milk is affected by seasonal variations: a retrospective observational study

The influence of seasonal variation upon human milk macronutrient content has not been elucidated. This study aimed to compare the macronutrient content of HM produced by lactating mothers during the winter and the summer seasons. Macronutrient content of colostrum milk samples collected from lactating mothers of healthy term infants between March 2012 and February 2016 was measured by mid-infrared spectroscopy and compared. The carbohydrate content of the colostrum was significantly higher in the summer season than in the winter season (6.2 ± 1.3 vs. 5.5 ± 1.4, p-value < 0.001). Protein, fat, and energy contents were similar in summer and winter in both groups (protein 2.7 ± 2.1 vs. 2.6 ± 2.2 g/100 ml, fat 2.6 ± 1.9 vs. 2.35 ± 1.9 g/100 ml, and energy 62 ± 19.1 vs. 60.5 ± 21 kcal/100 ml, respectively). The carbohydrate content in colostrum obtained from mothers of term infants was affected by seasonal variations.

Evaluation of Breast Milk Macronutrient Content in Preeclamptic Mothers.

Background: Preeclampsia remains among the leading causes of maternal and perinatal morbidity and mortality worldwide. This study aimed to evaluate the macronutrient content of colostrum in mothers with preeclampsia. Materials and Methods: Women who delivered by cesarean section at ≤37 weeks of gestation were included in the study. Fresh colostrum samples were collected by manual expression on postpartum days 2 and 4 and were analyzed using a mid-infrared human milk analyzer. Demographic characteristics of the patients were recorded. The protein, fat, carbohydrate, and energy composition of the milk samples were compared between mothers with and without preeclampsia. Results: A total of 58 colostrum samples (25 preeclampsia, 33 nonpreeclampsia) were analyzed. The median gestational age of the infants was similar in both groups (32 versus 33 weeks; p > 0.05). There were also no differences between the groups in terms of maternal age, maternal body mass index, maternal weight gain during pregnancy, and infant gender. Median protein, fat, and carbohydrate levels in the colostrum of the preeclamptic mothers were 2.2, 2.1, and 6.3 g/dL, respectively, compared with 1.8, 2.4, and 6.2 g/dL in the nonpreeclamptic mothers, respectively. Although the colostrum of preeclamptic mothers had higher protein content and lower fat content, the difference was not statistically significant. Carbohydrate and energy contents were similar in both groups. Conclusions: The presence of preeclampsia does not cause a significant change in the macronutrient composition of mother's milk. Analyses of micronutrients at different lactation stages in larger samples are needed.

Meeting Protein and Energy Requirements of Preterm Infants Receiving Human Milk.

Mother's own milk is universally recognized as the optimal source of nutrition for preterm infants, although most authorities agree a multi-nutrient fortifier must be added in order to support nutrient accretion at a rate comparable to in utero. Nevertheless, many preterm infants face a gap between achieved growth and what could have been achieved in utero. In this narrative review, we provide an overview on the macronutrient content in mother's own milk and donor milk and how this can be enhanced by the various available multi-nutrient fortifiers. We describe their general compositions and formulation, as well as several of their theoretical and practical advantages and drawbacks. In addition, differences between standardized fortification, or a more individualized approach like adjusted and targeted fortification are discussed. The optimal strategy however remains to be elucidated, and more experimental well-powered studies are therefore urgently needed. Until then, financial considerations and practical capabilities are likely to be the main drivers of local fortification strategies.

Macronutrient and Micronutrient Content in Breast Milk

Breast milk is the best source of nutrition for newborns. Breast milk content is composed of water, protein, carbohydrates, fats, vitamins, minerals, antibodies, and enzymes. Breast milk also contains white blood cells and substances that make up the baby`s immune system such as immunoglobulins and lysozyme. The composition of breast milk can change depending on the needs and ages of the baby. Exclusive breastfeeding is recommended by many doctors and other healthcare professionals. One of the factors that influence exclusive breastfeeding is the mother`s knowledge about the content of breast milk. Most breastfeeding mothers do not know the nutritional content of breast milk, both macronutrients, and micronutrients. Breast milk is also rich in immune cells, antibodies, and other bioactive ingredients that help protect the baby from infection until the baby has an effective antibody response in the first few months of life. Breast milk is an ideal nutrient for babies. The mixture of vitamins, proteins, and fats in breast milk can provide the nutrients and calories the baby needs to grow. Babies who have been breastfed for 6 months are more likely to reach their ideal weight.Therefore, the government recommends exclusive breastfeeding for infants up to the age of 6 months.

Effects of the circadian rhythm on milk composition in dairy cows: Does day milk differ from night milk?

Metabolism in most organisms can show variations between the day and night. These variations may also affect the composition of products derived from livestock. The aim of the present study was to investigate the difference in composition between the day milk and night milk of dairy cows. Ten multiparous Holstein cows (milk yield = 25.2 ± 5.00 kg/d) were randomly selected during mid lactation. Milk samples were collected at 0500 h ("night milk") and 1500 h ("day milk") and analyzed to determine their composition. Mid-infrared spectroscopy was used to analyze macronutrient content of milk. Metabolomics and lipidomics were used to detect and analyze small molecules and fatty acids, respectively. An automatic biochemical analyzer and ELISA kits were used to determine biochemical indicators, as well as antioxidant and immune parameters in the milk. Though milk fat, protein, lactose, and total milk solids were not different between day milk and night milk, small molecules, metabolites and lipids, and hormones and cytokines differed between day milk and night milk. Regarding biochemical and immune-related indicators, the concentrations of malondialdehyde, HSP70, and HSP90 in night milk were lower than that in day milk. However, interferon-γ levels were higher in night milk. Additionally, night milk was naturally rich in melatonin. Lipidomics analyses showed that the levels of some lipids in night milk were higher than those in day milk. Metabolomics analyses identified 36 different metabolites between day milk and night milk. Higher concentrations of N-acetyl-d-glucosamine, cis-aconitate, and d-sorbitol were observed in day milk. However, the other 33 metabolites analyzed, including carbohydrates, lipids, AA, and aromatic compounds, showed lower concentrations in day milk than in night milk. The present findings show that the composition of night milk differs considerably from that of day milk. Notable changes in the circadian rhythm also altered milk composition. These results provide evidence to support the strategic use and classification of day milk and night milk.

Predicting Protein and Fat Content in Human Donor Milk Using Machine Learning.

Donor milk is the standard of care for hospitalized very low birth weight (VLBW) infants when mother's milk is unavailable; however, growth of donor milk-fed infants is frequently suboptimal. Variability in nutrient composition of donated milk complicates the production of a uniform pooled product and, subsequently, the provision of adequate nutrition to promote optimal growth and development of VLBW infants. We reasoned a machine learning approach to construct batches using characteristics of the milk donation might be an effective strategy in reducing the variability in donor milk product composition. The objective of this study was to identify whether machine learning models can accurately predict donor milk macronutrient content. We focused on predicting fat and protein, given their well-established importance in VLBW infant growth outcomes. Samples of donor milk, consisting of 272 individual donations and 61 pool samples, were collected from the Rogers Hixon Ontario Human Milk Bank and analyzed for macronutrient content. Four different machine learning models were constructed using independent variable groups associated with donations, donors, and donor-pumping practices. A baseline model was established using lactation stage and infant gestational status. Predictions were made for individual donations and resultant pools. Machine learning models predicted protein of individual donations and pools with a mean absolute error (MAE) of 0.16g/dL and 0.10g/dL, respectively. Individual donation and pooled fat predictions had an MAE of 0.91g/dL and 0.42g/dL, respectively. At both the individual donation and pool levels, protein predictions were significantly more accurate than baseline, whereas fat predictions were competitive with baseline. Machine learning models can provide accurate predictions of macronutrient content in donor milk. The macronutrient content of pooled milk had a lower prediction error, reinforcing the value of pooling practices. Future research should examine how macronutrient content predictions can be used to facilitate milk bank pooling strategies.

The implications of routine milk fortification for the short and long-term health of preterm babies

Fortification refers to the practice of enriching human milk feeds for very preterm babies with macronutrients, minerals and vitamins. Though standard of care in some parts of the world, adoption of fortification is not universal. Fortification entered into use on the assumption that human milk macronutrient content, principally protein, is insufficient to support the growth and development of very preterm babies. However, because of the substantial variability in human milk composition, routine fortification risks exposing some babies to very high protein intakes, which may be dangerous. Some clinicians fear fortification with cow-milk derived products will increase the risk of necrotising enterocolitis, leading them to favour commercial fortifiers made from pooled human milk over cow milk based products, a practice that has additional ethical implications. Randomised controlled trials of multi-nutrient fortification to-date are inadequate. No trial has had power to detect important functional effects; the majority are methodologically weak and focus primarily upon short-term growth. Evidence to guide practice is inadequate. There is an urgent need for collaboration to conduct high-quality research to end these long-standing uncertainties.

Challenges in providing adequate and reliable nutrition for extremely low birth weight infants.

There is limited data on macronutrient content and its variability in mothers' milk of extremely low birth weight (ELBW) infants. The primary objectives were to determine the mean and the range of macronutrient content in milk from mothers of ELBW infants. A near-infrared milk analyzer was used to measure fat, protein and carbohydrate. Data is presented as mean±SD, median, and minimum/maximum values. Intraclass correlation coefficients (ICC) and linear regression were used to examine differences between repeat analyses. The mean GA and BW of infants were 25.6±1.9 weeks and 678.6±78.6g respectively. The mean caloric content of 212 milk samples (10 infants) was 20.1±5.4 cals/oz. The mean fat, protein and carbohydrate content were 3.2±1.8, 1.6±0.5g% and 8.0±0.8g% respectively. Large subject to subject and day to day variations were observed. The range of calories, fat, protein and carbohydrate content were 10.4-42.3 cals/oz, 0.2-14.1, 0.6-3.3 and 6.4-13.7g% respectively. Nearly half of all samples had 17 or less cals/oz and 10% had 15 or less cals/oz. There were no significant differences between repeat readings on a single sample. Standard fortification in presence of significant variation in macronutrient and caloric content of mother's milk can lead to large day to day variation in macronutrient and caloric intake of ELBW infants. Clinical significance of variable intake from one day to another in infants at high risk of NEC and growth failure is unclear and needs further study.

Macronutrient Composition of Donated Human Milk in a New Zealand Population.

A primary role of human donor milk banks is to provide pasteurized human milk for the sick and preterm infant populations and to support the mothers of these infants as they establish their own milk supply. The results of human milk pasteurization continue to be studied to provide information that enables optimal nutrition in this sick and preterm population. The aims of our study were to determine macronutrient characteristics (fat, protein, carbohydrate) and energy content of human milk donated to the Christchurch Women's Hospital Human Milk Bank in New Zealand, and the influence of Holder pasteurization on this macronutrient composition. This was a retrospective, pre/post pasteurization observational design to describe the macronutrient content within two groups of donors, mature preterm PDM (n = 13; 21%) and mature term PDM (n = 50; 79%). Sixty three samples of human milk donated to the human milk bank by 27 registered participants (mothers of preterm and term infants) were analyzed. This analysis took place July-September 2018 using a human milk analyzer before and after Holder pasteurization (62.5 °C for 30 min). Preterm milk contained on average 76 kcal/100 ml energy, 4.0 g/100 ml fat, 1.1 g/100 ml protein and 8.2 g/100 ml total carbohydrate; and mature term milk contained 68 kcal/100 ml energy, 3.5 g/100 ml fat, 0.8 g/100 ml protein and 7.9 g/100 ml total carbohydrate. Wide variation between single, donor-pooled samples was demonstrated and there was no major result of pasteurization. This research adds to the evidence regarding the macronutrient content of preterm and term milk and that these values are unaffected by Holder pasteurization. The variance in individual pooled donor human milk indicates the importance of determining the nutrient composition of donated milk to inform fortification procedures.

Effect of Freezing Time on Tissue Factor Activity and Macronutrients of Human Milk.

Human milk proteins are known as vital molecules for infant development and growth. Tissue factor is one of these human milk proteins that its role in human milk has not been cleared yet. Therefore, the first aim of this study was to detect the tissue factor activity of human milk and also was to investigate the effect of extended freezer storage on the milk tissue factor activity. The relationship between the tissue factor activity and macronutrient content and pH of milk was also investigated in this study. Under this aim, mature human milk samples were obtained from 8 healthy women. Collected human milk samples were pooled and divided into aliquots that were stored at -20°C until the day to be analyzed. Milk tissue factor activity, protein, fat, lactose, energy, water, density, and pH levels were determined for up to six months. By two months from the freezing, tissue factor activity did not significantly change but significantly decreased at the end of the six months. From the first month to six months from freezing, lactose, protein, fat, and energy levels showed a significant decline. Milk pH did not change with freezing at the end of 6months. In conclusion, TF activity maintained its first-day activity until the second month after being pumped. The increased interest in breast milk leads us to believe that the gap existing in the knowledge of breast milk bioactive components like TF will be complemented with new research data.

Rapid Analysis of Milk Using Low-Cost Pocket-Size NIR Spectrometers and Multivariate Analysis.

The miniaturisation of analytical devices, reduction of analytical data acquisition time, or the reduction of waste generation throughout the analytical process are important requirements of modern analytical chemistry, and in particular of green analytical chemistry. Green analytical chemistry has fostered the development of a new generation of miniaturized near-infrared spectroscopy (NIR) spectrometric systems. However, one of the drawbacks of these systems is the need for a compromise between the performance parameters (accuracy and sensitivity) and the aforementioned requirements of green analytical chemistry. In this paper, we evaluated the capabilities of two recently developed portable NIR instruments (SCiO and NeoSpectra) to achieve a rapid, simple and low-cost quantitative determination of commercial milk macronutrients. Commercial milk samples from Italy, Switzerland and Spain were chosen, covering the maximum range of variability in protein, carbohydrate and fat content, and multivariate calibration was used to correlate the recorded spectra with the macronutrient content of milk. Both SCiO and NeoSpectra can provide a fast and reliable analysis of fats in commercial milk, and they are able to correctly classify milk according to fat level. SCiO can also provide predictions of protein content and classification according to presence or absence of lactose.

The Effect of Physical Activity on Human Milk Macronutrient Content and Its Volume.

Objective: There are multiple health benefits associated with both breastfeeding and practicing physical activity (PA). Therefore, it is likely that many women might want to engage in both. We designed the current randomized clinical trial to examine the effect of moderate- to high-intensity PA on human milk (HM) volume and macronutrient contents. Methods and Study Design: In this prospective, randomized, crossover clinical trial, we recruited 31 healthy mothers who had been exclusively breastfeeding their infants. Mothers expressed HM twice each day on 2 consecutive days-a day with PA (1-hour before and 1-hour after PA) and a control day without PA (at the exact same hours of the day). The order of days (with/without PA) was determined randomly. Macronutrients and energy contents of HM were analyzed using the Human Milk Analyzer (Miris AB, Uppsala, Sweden). PA was graded according to the Borg Rating of Perceived Exertion scale (RPE scale). Results: A total 124 HM samples from 31 mothers were analyzed. Moderate- to high-intensity PA affected neither macronutrients (fat, carbohydrates, protein) nor energy content. Milk volume remained unaffected by PA as well. Conclusions: Maternal PA does not affect HM volume or its macronutrient contents. Lactating mothers can be reassured regarding their breast milk volume and composition while practicing PA of moderate to high intensity.

Maternal Undernutrition and Breast Milk Macronutrient Content Are Not Associated with Weight in Breastfed Infants at 1 and 3 Months after Delivery.

This study examined whether maternal nutritional intake and breast milk macronutrient content influence the weight of breastfed infants. We investigated 129 healthy mothers with singleton babies born from July 2016 to December 2017 in a university hospital in Tokyo, Japan. Information was obtained by a self-administered food frequency questionnaire at 1 (valid response n = 92; mean age, 34 years) and 3 (n = 57) months after delivery. Breast milk was sampled at 1 and 3 months and the macronutrient contents were analyzed. The average pre-pregnancy body mass index and weight gain during pregnancy were 20.7 ± 2.6 kg/m2 and 9.6 ± 3.7 kg, respectively. At 1 month, average maternal calorie intake was 1993 ± 417 kcal/day, which was lower than the intake recommended by Japanese Dietary Reference Intakes for breastfeeding mothers. There were no significant differences with regard to maternal calorie and protein intake, and breast milk macronutrient content between breastfed infants with weight above and below the 25th percentile of its distribution at both 1 and 3 months. This study suggests that suboptimal calorie intake by breastfeeding mothers and breast milk macronutrient content were not associated with weight of their infants at 1 and 3 months after delivery.

Maternal body mass index, parity and smoking are associated with human milk macronutrient content after preterm delivery

BackgroundMaternal characteristics may be associated with human milk macronutrients but no definite conclusions have been made to date. AimThis study aimed to determine the relationship of maternal-associated factors on the content of macronutrients in human milk for the first six weeks after preterm delivery. Study designProspective observational cohort study. SubjectsMilk samples were collected from mothers after premature birth between 24 + 0–35 + 6 weeks. Outcome measuresMacronutrients and energy content were analyzed by mid-infrared transmission spectroscopy. Demographic and anthropometric data from mothers were systematically recorded. ResultsA total 1.558 human milk samples from 192 mothers were analyzed.Colostrum: higher protein (p = 0.001) and lower carbohydrate content (p = 0.003) were present in primiparous compared to multiparous milk. Vaginal birth was associated with increased carbohydrate content (p = 0.021). Fat and energy content in colostrum was not related to any maternal characteristics.Mature human milk: similarly to colostrum, higher protein content (p = 0.001) and lower carbohydrates content (p = 0.022) were observed in primiparous compared to multiparous milk. The mode of delivery was found to be another factor possibly influencing protein and carbohydrate levels (p = 0.036, p = 0.003, respectively). Pre-pregnancy obesity was associated with increased fat (p = 0.030) and energy content (p = 0.020) in human milk. On the contrary, smoking had a negative relationship to fat and energy content (p = 0.026, p = 0.007, respectively). ConclusionHuman milk macronutrient concentration after preterm delivery is associated with pre-pregnancy obesity, parity, mode of delivery and smoking. The impact of maternal factors on human milk composition should be taken into account in a strategy of feeding in premature infants.

Macronutrient composition of longitudinal milk samples from captive aardvarks (Orycteropus afer).

Hand-rearing and assisted-rearing aardvarks in captivity has become commonplace and has led to success in breeding the species. However, the macronutrient content of aardvark milk past 1 month of age is unknown. A better understanding of aardvark milk composition would enhance captive management efforts. Here, we assayed milk samples from two captive individuals from 2 to 114 days postpartum (N = 21) for dry matter, fat, crude protein, total sugar, ash (total minerals), calcium (Ca), phosphorus (P), and gross energy. The body weight of one calf was measured from birth to weaning. Milk macronutrient composition was compared to that of other Afrotherian species and Xenarthra species with similar diets. Average protein, fat, and sugar concentrations of aardvark milk across lactation were 12.3%, 13.6%, and 2.5%, respectively. Ash averaged 1.9%, with Ca (0.50%) and P (0.35%) accounting for about 45% of total minerals. All measured nutrients increased over lactation except sugar, which decreased. Aardvark milk is high in energy (2.12 kcal/g) mostly derived from fat and protein and little energy from sugar. Calf growth was linear (r2 = 0.995) with a mean gain of 159 g/day, achieving almost 30% of adult weight at weaning. Within Afrotheria, aardvark milk is higher in fat and protein and lower in sugar than elephant milk and more closely resembles the milk of its fellow insectivore, the elephant shrew. Aardvark milk is also similar in composition to milk of insectivorous Xenarthra species (nine-banded armadillo and giant anteater). Aardvark milk composition is consistent with the species' high-protein diet, fast growth, and nursing pattern.

Buffered or impaired: Maternal anemia, inflammation and breast milk macronutrients in northern Kenya.

Maternal anemia has adverse consequences for the mother-infant dyad. To evaluate whether and how milk nutrient content may change in ways that could "buffer" infants against the conditions underlying maternal anemia, this study assessed associations between milk macronutrients and maternal iron-deficiency anemia (IDA), non-iron-deficiency anemia (NIDA), and inflammation. A secondary analysis of cross-sectional data and milk from northern Kenya was conducted (n = 204). The combination of hemoglobin and transferrin receptor defined IDA/NIDA. Elevated serum C-reactive protein defined acute inflammation. The effects of IDA, NIDA, and inflammation on milk macronutrients were evaluated in regression models. IDA (β = 0.077, p = .022) and NIDA (β = 0.083, p = .100) predicted higher total protein (ln). IDA (β = -0.293, p = .002), NIDA (β = -0.313, p = .047), and inflammation (β = -0.269, p = .007) each predicted lower fat (ln); however, anemia accompanying inflammation predicted higher fat (β = 0.655, p = .007 for IDA and β = 0.468, p = .092 for NIDA). NIDA predicted higher lactose (β = 1.020, p = .003). Milk macronutrient content both increases and decreases in the presence of maternal anemia and inflammation, suggesting a more complicated and dynamic change than simple impairment of nutrient delivery during maternal stress. Maternal fat delivery to milk may be impaired under anemia. Mothers may buffer infant nutrition against adverse conditions or poor maternal health by elevating milk protein (mothers with IDA/NIDA), lactose (mothers with NIDA), or fat (mothers with anemia and inflammation). This study demonstrates the foundational importance of maternal micronutrient health and inflammation or infection for advancing the ecological understanding of human milk nutrient variation.

Breast Milk Macronutrient Components in Prolonged Lactation.

Human milk (HM) is the first dietary exposure in infancy and the best nutritional option for growth and healthy development of the newborn and infant. The concentrations of macronutrients, namely proteins, carbohydrates and lipids, change during lactation stages. HM has been studied extensively in the first 6 months of lactation, but there are limited data regarding milk composition beyond 1 or 2 years postpartum. The aim of our study was to describe longitudinal changes in HM macronutrient concentrations during the prolonged lactation of healthy mothers from the 1st to the 48th month. For the macronutrient content of milk of mothers breastfeeding for longer than 18 months, fat and protein increased and carbohydrates decreased significantly, compared with milk expressed by women breastfeeding up to 12 months. Moreover, the concentration of fat, protein and carbohydrates in HM over 2 years of lactation from the 24th to the 48th month remained at a stable level. However, analyzed macronutrients were positively (carbohydrate, r = 0.51) or negatively (fat, r = −0.36 and protein, r = −0.58) correlated with the amount of feeding. Our results create a greater potential for understanding the nutritional contribution of HM over 2 years of lactation and showed that the source of calories in breast milk for older children is mainly fat, while carbohydrates play a greater role in infant nutrition in the early stage. The observed changes of macronutrient concentrations from the 1st to the 48th month of lactation are probably related to the adaptation of milk composition to the increased energy demand of the intensively growing child.