Abstract
Background: Human milk is the preferred nutrition for neonates due to the multiple nutritional and immunologic benefits over formula. Preterm neonates are often fed human milk that has been stored frozen for up to 3 months. Human milk is sometimes pasteurized prior to feeding to preterm neonates to reduce the transmission of pathogens.
Goal: To assess changes in the concentrations and activity of host defense proteins in human milk following prolonged storage or pasteurization.
Methodology: (A) Samples obtained through Cincinnati Children's Hospital Human Milk Bank were frozen at -20 °C for 2 days, 4, 8, or 12 weeks prior to analysis. Whey fraction was isolated and protein analyzed by dot blot protein analysis followed by immunoreaction with antibodies to human lysozyme, lactoferrin, lactoperoxidase, secretory leukoprotease inhibitor, HNP 3, or human beta defensin 2. Protein levels were determined by computer-assisted densitometric analysis. Concentration of sIgA was measured by ELISA. (B) Freshly-collected human milk samples were pasteurized at 65 °C for 30 minutes, then stored at -20 °C and analyzed by dot blot for host defense proteins as described in A. (C) Muramidase activity was assessed by incubating 10 μg total protein with a suspension of Micrococcus luteus and calculated from the rate of decrease in optic density. Recombinant human lysozyme was used to generate a standard curve.
Results: Levels of lysozyme, lactoferrin, and lactoperoxidase were not significantly changed for up to 8 weeks of storage at −20 °C. In contrast, levels of sIgA were significantly reduced within 1 week at −20 °C (40% at 1 week, 10% at 2 weeks, and <5% at 8 weeks p <0.05 relative to fresh milk). Muramidase also activity decreased sharply with storage (120 units/μg vs. 260 units, p <0.02). In comparison with age-matched control samples, concentrations of lysozyme, lactoferrin, lactoperoxidase, sIgA, and muramidase activity were unaltered following pasteurization. HNP-3 and human beta defensin 2 were not detected.
Discussion: Although the nutritional value of human milk may be preserved following prolonged storage at −20 °C, protein activity may be significantly compromised thereby reducing the immunologic benefits from human milk. Pasteurization of milk to reduce transmission of viruses appears safe with respect to levels and activity of host defense proteins.
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Zimmerly, K., Clark, R., Ma, Y. et al. 17 Effects of Prolonged Storage or Pasteurization on the Levels And Activity of Host Defense Proteins in Human Milk.. Pediatr Res 60, 493 (2006). https://doi.org/10.1203/00006450-200610000-00039
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DOI: https://doi.org/10.1203/00006450-200610000-00039