| Milk has been recognised as an excellent source of nutrition for mammal newborns. Lipids in milk are present as milk fat globules(MFGs) which are surrounded by a tri-layer membrane outside that is derived from a unique secretion mechanism. The lipid composition and microstructure of bovine, caprine and human MFG and the changes in MFG during milk powder manufacture were studied. Besides, the changes of the structure and lipid composition of MFG during in vitro digestion of infant and adult gastrointestional were compared. And the effect of MFG on adherence and internalization of bacterial to HT-29 cell were also studied. These results would provide theoretical basis for the improvement and development of powdered milk formulas.First of all, the lipid composition, physicochemical properties and microstructure of bovine, goat and human MFGs were analyzed by means of chromatographic techniques and confocal microscopic studies. We showed that saturated fatty acid content was higher in bovine and caprine MFGs(>60%) than in human MFGs(<40%), but the content of the unsaturated fatty acid C18:2 in human MFGs was >7 times higher than in bovine and caprine MFGs.The cholesterol content of human milk was ~20% higher than that of bovine and caprine milk. The TAG composition of bovine milk fat contained a wide range of short and medium-chain TAG species compared with that of the caprine and human MFGs. Among the different polar lipid species, SM(29.28%) was more abundant in human milk, and PC was less abundant relative to that in bovine and caprine milk(33.1% and 31.6%, respectively). The distribution of the fatty acids of PE, SM and PI in caprine milk was similar to the fatty acid profile of polar lipids in bovine milk. The unsaturated fatty acids represented in PE, PS and PC in human milk(>40%) were higher than those in bovine and caprine milk. Contrastingly, more saturated fatty acids were found in PI and SM in human milk. The average diameter and the Z-potential of the fat globules in caprine milk was smaller than that in bovine milk and human milk. Confocal laser scanning microscopy images of MFGs revealed that the shape of the liquid-ordered domains varied by species. Confocal Raman spectroscopy showed that the unsaturated ratios of MFGs in human milk and bovine milk were decreased with increasing size of the globules, but the ratio of the medium caprine MFG was the highest in caprine milk. The carotenoid peaks was appeared high intensities in the human MFGs than bovine and caprine MFGs, and an increase in the concentration in carotenoids was observed with increasing size of the MFGs.Then, the effects of milk powder processing conditions on the microstructure and composition of MFGs were studied. The phospholipid, sterol, and the fatty acid composition of the phospholipids, except for the fatty acids of total lipids, were significantly changed under different processing conditions compared with the untreated MFGs. The contents and distribution of the phospholipids in pasteurised milk indicated a high level of stability, but the amounts of phosphatidylethanolamine, phosphatidylserine, and sphingomyelin were significantly changed by homogenisation. However, the proportion of PE decreased from 28% to 22% after spray-drying. A reduction in the cholesterol content was observed after pasteurisation and homogenisation. The MFGs of the PM looked similar to those of the RM. However, the red emission fluorescence rings were much looser and the circular areas were incomplete. The fluorescence probe in homogenised and spray-drying MFGs was spotted. The globule sizes increased after freeze drying and spray drying, but the integrity of their microstructures was still maintained. The main phospholipids and cholesterol showed some differences between the freeze-dried milk and spray-dried milk. The results indicated that freeze drying had a significant effect on the liquid-ordered domains in the MFG membrane, which was formed by sphingolipids and cholesterols. However, the liquid-disordered domains were more influenced by spray-drying, which were formed by glycerophospholipids.And then, the digestive characteristics of different MFGs in vitro infant and adult gastrointestinal were studied. It was found that the amount of free fatty acids released per unit volume of the milk samples decreased in the following order: caprine milk > bovine milk > human milk. The contents of the free fatty acids in the lipolysis products were accounted for 76.06%, 60.72% and 68.89%, respectively. Moreover, short and medium fatty acids and saturated fatty acids released from bovine and caprine milk were more than human milk, whereas more unsaturated fatty acids were released from human milk. The cholesterol content in the lipolysis products of bovine and human milk was about 0.5 times than the undigested milk, while there was no significant difference in caprine milk. The phospholipids of PE, PC and SM were hydrolyzed, and PE was mostly hydrolyzed in these three milks. Confocal micrographs showed that some small particales were detected after 30 min digestion of the three milks. While a few large particles appeared in bovine and caprine milk. The sizes of the MFGs were increased as some MFGs aggregated together and then formed large particles.Lipolysis was very fast in the first 10 min of bovine and caprine milk during in vitro adult intestinal digestion and no lag phase was observed. There were significantly differences of the lipolysis products from bovine and caprine milk. The profiles of free fatty acids released from bovine and caprine milk were not different except for C10:0 and C14:0. Chloesterol content in the adult digestived product was higher than in the infant digestive products, which was 424 mg/100 g and 387 mg/100 g, respectively. The main phospholipids of PE, PC and SM were hydrolyzed, and PE was mostly hydrolyzed in the adult digestion of these three milks. Confocal micrographs showed that large coalesced fat globules appeared, but the structure of MFGM was not destroyed under the adult gastric condition. Intact fat particles along with the empty/hollow particles were observed after intestinal digestion.It was found that the lag phase of raw and pasteurized milk was longer than that of homogenized milk and spray dried milk by comparison with the hydrolysis kinetics performed on the different processed milk during in vitro infant gastric digestion. The lipolysis speed of the spray-dried milk was significantly faster than the other three milks. There was no significant difference in the lipolysis rate of raw milk, homogenized milk and spray-dried milk during in vitro infant gastric digestion, while the digestion rate of pasteurized milk was the lowest. After the whole digestion, there was no significant difference of the percentage of lipolysis products of raw milk and pasteurized milk, but the free fatty acid composition of the hydrolyzate was obviously different. The highest undigested TAG in homogenized and spray dried milk was 16% and 21%, respectively. Homogenization and spray-drying also affected the cholesterol content of digested products, which was about 10% less than that of raw milk and pasteurized milk. These processes all increased the degree of hydrolysis of PC and decreased the degree of hydrolysis of SM. After the in vitro infant gastric dgestion of four processed milks, large coalesced fat globules appeared and the homogenized and spray-dried milk had the greatest degree of aggregation. After intestinal digestion, much smaller particles were formed. However, there were still some large fat droplets in spray-dried milk. The circular fluorescent structure of MFGM disappeared and the whole particle was colored red by the phospholipid probe Rh-DOPE during the digestion progresses.In the adult gastrointestinal digestion of the first 10 min, the levels of free fatty acids released from raw milk, pasteurized milk, homogenized milk and spray-dried milk were 23, 16.8, 21.6 and 20.4 ?mol/mL, respectively. There was no significant difference between raw milk, homogenized milk and spray-dried milk in the whole gastrointestinal digestion, but fat digestion of pasteurized milk was significantly lower than the others. The highes undigested TAG in pasteurized milk was the highest among others and the content was about 10.86%. Processing had no effect on the type and content of the released FFA, but there is a significant difference in fatty acid composition of the MAG was observed. The cholesterol levels in the digestive products during in vitro adult gastrointestinal digestion were increased than infant gastrointestinal digestion. Large aggreated and flocculated fat globules were observed during in vitro adult gastric digestion of different processed milks. After gastric digestion of 1 h, the fluorescence rings were still detected on the surface of the globules. After digestion of 2 h in the small intestine, the circular fluorescence was disappeared and the membrane phospholipid was replaced by bile salt. MFGs were decomposed into small particles by pancreatic lipase. Some aggregated large particles still existed in pasteurized milk after digestion.At last, the effects of different MFGs on adhesion and invasion of bacteria to HT-29 cells were studied. The bovine, caprine and human MFGs all promoted the adherence of probiotics(Lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus acidophilus, Bifidobacterium infants and Bifidobacterium longum) to HT-29 cells. The MFGs from caprine had a good promotion effect on Bifidobacterium longum. The effect of human MFGs on the adhesion of Bifidobacterium infantum was better than the other bacteria. The promotion effect of human MFGs on Lactobacillus delbrueckii subsp. bulgaricus was less than that of bovine MFGs and caprine MFGs. The results showed that MFGs showed higher adhesions to four kinds of probiotics. The three type of MFGs had significantly inhibited the binding adherence of pathogenic bacterias(Salmonella, Enterobacter sakazakii and Escherichia coli) to HT-29 cells, and the effect of caprine MFGs on these three pathogens was the best. The MFGs from human had the strongest inhibitory effect of Escherichia coli. The different processed MFGs had different adherences of the probiotics and pathogens to HT-29 cells. The greater of the process degree, the less number of probiotics will be adhere to HT-29 cells, and the lower inhibited effect of pathogens will be presented apart from the pasteurization. |
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