Effect of cold storage on human milk fat globule membrane: Microstructure and proteomic analysis

We investigated the changes in microstructure and composition of milk fat globule membrane (MFGM) in fresh human milk under different storage temperatures (-18, -3, -0.5, and 4 & DEG;C). Freezing at -18 & DEG;C resulted in the formation of ice crystals that ruptured the MFGM, leading to aggregation and increased particle size, which could potentially impact lipid digestion and absorption in infants. Conversely, storage at -3, -0.5, or 4 & DEG;C were more favorable for maintaining the stability of MFGM structure without ice crystals formation. Notably, -3 & DEG;C exhibited the highest preservation of MFGM integrity. Label-free proteomics analysis identified a total of 1525 different MFGM proteins in the initial sample (0 day) and samples preserved at varying temperatures (-18, -3, -0.5, and 4 & DEG;C) for a duration of 4 days. Principal component analysis and hierarchical clustering analysis demonstrated two distinct clusters of MFGM proteins across the different storage temperatures. Specifically, MFGM proteomic characteristics at -18 and -3 & DEG;C showed more similarities and clustered together with the 0-day samples, while -0.5 and 4 & DEG;C formed a separate cluster. Comparative analysis with the control sample (day 0) highlighted significant changes in 202 proteins, most of which played essential roles in binding, catalytic activities, and pathways related to protein, lipid, and fatty acid metabolism. Our findings highlight the superior ability of -3 & DEG;C to inhibit bacterial growth compared to 4 and -0.5 & DEG;C, and to preserve the microstructure of milk fat globules compared to -18 & DEG;C. Thus, for household storage up to 4 days, we recommend a temperature of -3 & PLUSMN;1 & DEG;C, considering temperature fluctuations in domestic refrigerators.