Effect of Holder Pasteurization and Frozen Storage on Macronutrients and Energy Content of Breast Milk

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The aim of this study was to explore the effect of Holder pasteurization and frozen storage at −20°C after pasteurization on fat, total nitrogen, lactose, and energy content of breast milk. Both procedures are routinely practiced in human milk banks.


A total of 34 samples of frozen breast milk, donated by 28 women, were collected. Once thawed, an aliquot of each sample was analyzed before pasteurization; the remaining milk was pasteurized (Holder method) and split into 8 aliquots. One aliquot was analyzed after pasteurization and the remainder frozen at −20°C and analyzed 30, 60, 90, 120, and 180 days later. For every aliquot, fat, total nitrogen, lactose, and energy content were determined using the device human Milk Analyzer.


We observed a significant reduction in fat (3.5%; −0.17 (−0.29; −0.04) g/dL) and energy content (2.8%; −2.03 (−3.60; −0.46) g/dL) after pasteurization. A significant decrease over time was observed for fat, lactose and energy content. No significant changes were observed for nitrogen content. Mean differences between day 0 postpasteurization and day 180 were −0.13 (−0.21; −0.06) g/dL for fat, −0.08 (−0.13; −0.03) g/dL for lactose, and −1.55 (−2.38; −0.71) kcal/dL for energy content. The relative decreases were 2.8%, 1.7%, and 2.2%, respectively. Overall (postpasteurization + frozen storage), a 6.2% and 5% decrease were observed for fat and energy, respectively.


Holder pasteurization decreased fat and energy content of human milk. Frozen storage at −20°C of pasteurized milk significantly reduced fat, lactose, and energy content of human milk.

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