Frozen storage of fresh human milk for up to 3 months and refrigerator storage for up to 72 hours is currently recommended. We examined the integrity of human milk that was subjected to refrigeration followed by prolonged freezing.
100 ml of freshly expressed milk was obtained from mothers (n=18) in the NICU and was divided into 5 aliquots. A time 0 sample was immediately stored at -800C. 2 samples were stored fresh in -200C freezer for 1 and 3 months (FF, Fresh-Frozen) and 2 samples were placed in refrigerator (+40C) for 72 hours and then stored in -200C freezer for 1 and 3 months (RF, Refrigerated-Frozen). Each sample was then maintained in -800C until analyzed. Milk integrity was described as the effect on pH, total protein content (TP), total bacterial (TBCC), Gram positive (GPCC) and Gram negative (GNCC) colony counts. Data were analyzed by two-way repeated measures ANOVA.
There was a significant (p < 0.001) decline in TP from a mean of 24 to 17 g/L from 0 time to 1 and 3 months FF. TP remained at 18 g/L for RF samples both at 1 and 3 months. Median pH declined significantly from a mean of 7.12 at time 0 to 6.4-6.5 range after frozen storage and did not differ significantly between 1 and 3 months. No significant differences were noted between the FF and RF samples at 1 or 3 months for TP and pH. There was a significant decline in TBCC with FF and RF (p < 0.001) in general, which was more pronounced after 1 month. TBCC did not change from 0 time (median 5.4 x 104 cfu/mL) to 1 month (5.6) with FF sample but was significantly reduced with RF (1.4). At 3 months TBCC for both FF and RF samples were significantly lower than baseline (0.20 and 0.25 respectively), p < 0.05. GPCC declined significantly from time 0 (4.2 x 104) to 1 month with RF (0.57) samples only. There was a pronounced decline for both FF and RF samples at 3 months, p< 0.05. Although only detectable in 4 samples, refrigeration and/or prolonged freezing did not affect GNCC.
These preliminary data suggest that refrigeration followed by prolonged frozen storage does not change the integrity of the human milk when compared to fresh-freezing alone. Freezer storage affects milk constituents initially but prolonged freezer storage does not change milk integrity adversely. The magnitude of these changes observed would not be expected to affect the use of stored human milk.