Evaluation by high-performance liquid chromatography of the hydrolysis of human milk triacylglycerides during storage at low temperatures.

The effect of various storage methods on the stability of the triacylglyceride fraction of human milk was evaluated. Samples were treated as follows: Group I--stored at -20 degrees C for 4 months, group II--heated for 1.5 min at 80 degrees C and stored at -20 degrees C for 4 months, group III--stored at -80 degrees C for 4 months and thawed rapidly at room temperature (25 degrees C) just before analysis and group IV--stored at -80 degrees C for 2 months, thawed rapidly at room temperature (25 degrees C), then stored at -80 degrees C for a further 2 months and finally thawed rapidly at 25 degrees C just before analysis. The absence of hydrolysis products in group II and group III indicated that these storage procedures were satisfactory even when samples were rapidly thawed for a short time (group IV). Only storage at -20 degrees C without previous heat treatment led to the hydrolysis of triacylglycerides and the appearance of free fatty acids (group I). On the other hand, the effect that freezing and thawing had over the lipolysis grade was studied. Samples were treated as follows: group V--stored at -20 degrees C for 2 months, thawed slowly at refrigerator temperature (5 degrees C), held at this temperature for one week and stored for a further month at -20 degrees C. Freezing and thawing activated lipolysis and increased the production of free fatty acids, monoacylglycerides and diacylglycerides. Milk samples were analyzed by reversed-phase HPLC with a ternary gradient of acetonitrile-dichloromethane-acetone and an evaporative light-scattering detector.