| Lactoferrin is an important bioactive protein in mammalian milk and the core immune component in human milk.Adding lactoferrin to infant formula milk powder can promote the growth and development of infants and improve their immunity.Understanding the content of lactoferrin in breast milk and mastering the change law of lactoferrin content has important guiding significance for the design of formula of infant milk powder at various stages.At present,there are still many problems in the detection methods of lactoferrin in human milk.In this paper,the sample pretreatment process and detection conditions were optimized,and the ultra performance liquid chromatography method suitable for human milk lactoferrin was established,and the effects of lactation period,delivery mode and parity on lactoferrin were studied.The main contents and results are as follows:1.A method for the determination of human lactoferrin was established by optimizing the chromatographic conditions and the pretreatment process of human milk samples.Human lactoferrin was extracted with phosphate solution of 100 mm,p H 5.0±0.2.The extract was enriched by heparin affinity column,then washed with phosphate solution containing 100 m M sodium chloride,and finally eluted with phosphate solution containing 1 M sodium chloride.The elution was centrifuged and determined on the computer.The detection wavelength was set at 201nm and the injection volume was 3?L.2.The method was validated by methodology.The results showed that the linear range of human lactoferrin was 5?200mg/L,R~2>0.998,the standard equation y=16881x+161391.Both intra day and inter day relative standard deviations were less than 2.14%.When the scalar was 250-2000mg/L,the recovery was 96.52-107.43%.Twelve mature milk samples were used to verify the method.The lactoferrin content was 690-2830mg/kg,accounting for 10.04-27.10%of the total protein in human milk.3.Objective to study the changes of lactoferrin content in human milk and its influencing factors.The content of lactoferrin was the highest in human colostrum,and decreased in turn during transition milk and mature milk,and the contents were 3214.49±414.61,2046.74±539.70 and 907.50±421.10,respectively;The difference was statistically significant among the three groups;the content of lactoferrin was affected by parity,and the content of lactoferrin in multiple parity group was significantly higher than that in one parity group;there was no significant difference in lactoferrin content between different delivery modes,but the lactoferrin content in cesarean section group was slightly higher than that in natural birth group.4.The stability of lactoferrin in the production process of infant formula milk powder was studied,and the lactoferrin content in commercial infant formula milk powder was investigated.After being pasteurized,vacuum mixed and spray dried,lactoferrin decreased from 90.69 mg/kg to 13.86 mg/kg,with a loss rate of 87.10%.The content of lactoferrin in infant formula was 4.59-38.26 mg/kg.There were significant differences among samples of different brands,but there was no significant difference between domestic milk powder group and imported milk powder group.5.A formula of infant milk powder fortified with high concentration lactoferrin was designed.The theoretical concentration of lactoferrin was 68.69-77.71mg/100 m L,which was similar to that of lactoferrin in human milk.The theoretical calculation shows that the energy and nutrient contents in the formula meet the relevant national standards. |
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