乳脂肪球的组成结构、体外消化及抗菌特性

发布时间：2018-06-18 12:17

本文选题：乳脂肪球 + 加工处理　；参考：《江南大学》2017年博士论文

【摘要】：乳是哺乳动物新生儿的主要营养来源。其中的脂肪以脂肪球的形式存在,外围由独特机制分泌的三层膜包裹。本文围绕乳脂肪球(MFG)的脂质组成及结构特点,研究对比了三种哺乳动物(牛、羊和人)MFG的脂质组成和微观结构及不同加工处理对牛乳MFG脂质组成和微观结构的影响。在此基础上通过模拟体外消化进一步研究它们在婴儿及成人体内的消化特点及其对细菌与HT-29细胞的粘附及侵染作用,为开发合适的母乳替代品提供理论依据。主要研究内容包括:首先,应用色谱分析及显微检测手段全面分析了牛乳、羊乳和人乳MFG的脂质组成、理化性质和微观结构。结果表明:牛乳和羊乳含有较高比例的饱和脂肪酸(60%),而人乳中的饱和脂肪酸比例40%,且人乳中C18:2的含量大约是牛乳和羊乳的7倍以上。人乳中胆固醇的含量比牛乳和羊乳高约20%。牛乳与羊乳和人乳相比,含有更多的短、中链的甘油三酯。牛乳和羊乳中最主要的磷脂为磷脂酰胆碱(PC),分别占总磷脂组分的33.1%和31.6%。而人乳中最主要的磷脂则为鞘磷脂(SM)占29.28%。牛乳和羊乳中的磷脂酰乙醇胺(PE)、SM和磷脂酰肌醇(PI)含有的脂肪酸组成类似。人乳中的PE、磷脂酰丝氨酸(PS)和PC中含有的不饱和脂肪酸(40%)比牛乳和羊乳高。羊乳MFG的平均粒径最小,Z-电位的值也最低。从激光共聚焦显微图片可知,液态有序区(Lo)的形状和大小也与物种有关。牛乳MFG中主要为圆形,羊乳中大部分形状不规则,而人乳中则以圆形和不规则形共存。由拉曼共聚焦光谱图可知,人乳和牛乳MFG的不饱和度随着MFG粒径的增大而降低,而羊乳中则是中等大小的MFG的不饱和度最高。人乳中类胡萝卜素的峰强度比羊乳和牛乳高,且随着粒径的减小而增强。其次,对比研究了乳粉生产过程对MFG微观结构及组成的影响。结果表明:除总脂肪酸的组成没有明显改变外,磷脂、胆固醇和磷脂脂肪酸的组成都发生了变化。牛乳MFG经巴氏杀菌后,磷脂的含量和分布相对稳定。均质对PE、PS和SM的含量影响最大。喷雾干燥之后PE的含量由28%降低到22%。胆固醇的含量也随着巴氏杀菌和均质处理而降低。巴氏杀菌后的乳脂肪球膜(MFGM)依然呈圆环状包裹在MFG外围,但是荧光环结构较松散。均质和喷雾干燥之后MFGM结构完全破坏,荧光探针呈点状分布。经过真空冷冻干燥和喷雾干燥这两种不同的干燥处理后,MFG的粒径变大,但其MFGM结构依然保持完整。这两种不同的干燥处理对磷脂及胆固醇的含量影响较大,我们推测脂肪球的Lo区更容易受低温干燥的影响,而液态无序(Ld)区更容易受高温干燥的影响。然后,通过体外实验模拟研究不同动物MFG在婴儿及成人胃肠道的消化特点。结果表明:三种哺乳动物MFG在婴儿体外消化过程中的水解速率为:羊牛人。牛、羊和人乳MFG消化后的脂解产物中,游离脂肪酸(FFA)分别占总脂肪的76.06%、60.72%和68.89%。牛乳和羊乳比人乳产生更多的短、中链脂肪酸和饱和脂肪酸。牛乳和人乳消化产物中胆固醇的含量比未消化时增加了将近0.5倍,而羊乳则无明显差异。三种哺乳动物MFG中的磷脂主要是PE、PC和SM发生了水解,且PE的水解程度最高。由激光共聚焦显微图像可知,牛乳和羊乳经过30 min胃消化后,MFG的粒径减小,一部分MFG聚集在一起,之后相互融合形成大MFG。而人乳MFG的粒径则一直呈减小趋势。牛乳和羊乳MFG在成人体外肠消化前10 min内,没有出现之前文献报道的延滞期,水解速度很快,脂解产物的比例有明显区别。但是产生的FFA的组成及含量除了C10:0和C14:0并无显著差异。消化产物中胆固醇的含量比婴儿体外消化产物中高,分别为424mg/100g和387 mg/100g。成人体外消化过程中也主要是PE、PC和SM发生了水解,且PE的水解程度最高。激光共聚焦显微图像显示,在成人体外胃消化过程中MFG大量的聚集几乎不发生消化,MFGM的结构依然保持完整。经过肠液消化后,可以看到由消化产物与未消化的TAG及蛋白形成中间有空洞的脂肪颗粒。不同加工处理的MFG在婴儿体外胃消化时,生乳和巴氏杀菌乳的延滞期要比均质乳和喷雾干燥乳的长。喷雾干燥之后的乳,脂解速度明显快于其它三种乳。婴儿体外肠消化过程中,生乳、均质乳及喷雾干燥乳的脂解速率没有显著差异,而巴氏杀菌乳的消化速率最低。均质乳和喷雾干燥乳中未水解的TAG最高,分别占16%和21%。均质和喷雾干燥处理也影响了消化产物中胆固醇的含量,与生乳和巴氏杀菌乳相比约减少10%。巴氏杀菌、均质和喷雾干燥都增加了PC的水解度。四种不同加工处理的MFG经过婴儿体外胃消化1 h后,都有部分MFG发生聚集,尤其是均质乳和喷雾干燥乳。经过肠消化后进一步形成更小粒径的MFG,其中喷雾干燥乳消化后仍然有一部分大的脂肪液滴存在。随着消化的进行MFGM的环状荧光结构都消失了,整个颗粒都被磷脂探针Rh-DOPE染成了红色。生乳、巴氏杀菌乳、均质乳及喷雾干燥乳在成人体外肠消化的前10 min,脂解速率分别达到23、16.8、21.6和20.4μmol/m L乳。巴氏杀菌乳中未水解的TAG最高,占10.86%。加工处理对释放的FFA的种类和含量无影响,但是对水解产物MAG的脂肪酸组成却有明显的区别。成人体外胃肠道消化后与婴儿相比增加了消化产物中胆固醇的含量。不同加工处理的MFG在成人胃中容易发生聚集。牛乳和巴氏杀菌乳MFG在成人体外胃消化1 h后,依然能检测到MFGM的环状荧光。经过2 h肠消化后,环状的荧光环消失后膜磷脂被胆盐取代,MFG被胰脂肪酶进一步分解为小的颗粒。但是巴氏杀菌乳中依然有一些聚集的大微粒存在。最后,研究了不同MFG对细菌与HT-29细胞的粘附及侵染作用。结果表明:不同动物(牛、羊和人)的MFG都会促进益生菌(德氏乳杆菌保加利亚亚种、嗜酸乳杆菌、婴儿双岐杆菌和长双歧杆菌婴儿亚种)与HT-29细胞的粘附。羊乳MFG对长双歧杆菌有良好的促粘附的作用。人乳MFG对婴儿双歧杆菌表现出较高的促粘附作用。人乳MFG对于德氏乳杆菌保加利亚亚种的促粘附作用最小,不及牛乳MFG和羊乳MFG。并且羊乳MFG比牛乳和人乳MFG对四种益生菌都表现出较高的促粘附作用。这三种哺乳动物MFG都对致病菌(沙门氏菌、阪崎肠杆菌和大肠杆菌)与HT-29细胞的粘附有抑制作用,但是羊乳MFG对这三种致病菌的抑制效果明显优于牛MFG,人乳MFG对大肠杆菌的抑制作用最强。经过加工处理后的MFG对益生菌和致病菌粘附到HT-29细胞的数量有不同程度的影响。除巴氏杀菌外,加工处理的程度越大,益生菌与HT-29细胞粘附的数量越少,对致病菌的抑制程度越低。
[Abstract]:Milk is the main source of nutrition in mammal newborns. The fat in the form of fatty balls and three layers of membrane secreted by unique mechanisms. The lipid composition and microstructure of the three mammals (cattle, sheep and people) and the different processing places of the lipid composition and structure of the milk fat balls (MFG) are studied and compared in this paper. The lipid composition and structure characteristics of the milk fat balls (MFG) are compared with the three mammals. The effects of the lipid composition and microstructure of milk MFG were studied. On this basis, the digestive characteristics and the adhesion and infection of bacteria to HT-29 cells in infants and adults were further studied by simulated digestion in vitro, and the theoretical basis for the development of appropriate breast milk substitutes was provided. The lipid composition, physicochemical properties and microstructure of milk, goat milk and human milk MFG were analyzed by spectral analysis and microscopic examination. The results showed that milk and goat milk contain a high proportion of saturated fatty acids (60%), and the proportion of saturated fatty acids in human milk is 40%, and the content of C18:2 in human milk is about 7 times more than that of milk and sheep milk. The content of cholesterol is higher than milk and goat milk about 20%. milk and goat milk and human milk. It contains more short, middle chain triglycerides. The main phosphatidylcholine (PC) in milk and goat milk is 33.1% and 31.6%. of the total phosphatidylcholine, and the most important phosphatide in human milk is sphingomyelin (SM) in 29.28%. milk and goat milk. Phosphatidylethanolamine (PE), SM and phosphatidylinositol (PI) contain a similar composition of fatty acids. PE, phosphatidylserine (PS) and PC in human milk are higher than milk and goat milk. The average particle size of the goat milk MFG is the smallest and the Z- potential is the lowest. The shape of the liquid ordered region (Lo) is known from the laser confocal microscope picture. And the size is also related to species. The main shape of the milk MFG is round, most of the shape of the goat milk is irregular, and the human milk coexists with round and irregular shape. The Raman confocal spectrogram shows that the unsaturation of human milk and milk MFG decreases with the increase of the MFG size, while the middle size MFG is the highest unsaturation in the goat milk. The peak intensity of carotenoids in milk was higher than that of goat milk and milk, and increased with the decrease of particle size. Secondly, the effects of the milk powder production process on the microstructure and composition of MFG were compared. The results showed that the composition of the phospholipid, cholesterol and phospholipid fatty acids in Chengdu had changed in addition to the composition of the total fatty acid. After pasteurization, the content and distribution of phospholipids were relatively stable. Homogenization had the greatest influence on the content of PE, PS and SM. After the spray drying, the content of PE decreased from 28% to 22%. cholesterol and decreased with pasteurization and homogenization. After pasteurization, the milk fat globule membrane (MFGM) remained round and round in the periphery of MFG, but the fluorescent ring was in the ring. The structure was looser. After homogenization and spray drying, the structure of MFGM was completely destroyed and the fluorescent probe was distributed in a spot. After two different drying processes, vacuum freeze drying and spray drying, the particle size of MFG became larger, but the structure of MFGM remained intact. These two different drying treatments had great influence on the content of phospholipid and cholesterol. We speculate that the Lo region of the fat ball is more susceptible to low temperature drying, and the liquid disorder (Ld) region is more susceptible to the effect of high temperature drying. Then, the digestive characteristics of different animals MFG in the gastrointestinal tract of infants and adults are studied by experiments in vitro. The results show that the hydrolysis rate of three kinds of mammalian MFG in the process of infant digestion is: sheep The free fatty acid (FFA) accounted for 76.06% of the total fat, 60.72% and 68.89%. milk and goat milk produced more short, medium chain fatty acid and saturated fatty acid than human milk. The content of cholesterol in cow milk and human milk digestion products increased by nearly 0.5 times than that of undigested, and the sheep milk was less obvious than that of the MFG. The phospholipids in the three mammalian MFG were mainly PE, PC and SM, and the hydrolysis of PE was the highest. By laser confocal microscopy, the milk and goat milk were digested by 30 min, the particle size of MFG decreased and some MFG gathered together and then formed a large MFG. and the size of the human milk MFG decreased. The trend. Milk and goat milk MFG in 10 min before the digestion of human intestines, there is no prior period of delay reported in the literature, the rate of hydrolysis is very fast, and the proportion of the hydrolysate products is distinct. But the composition and content of the produced FFA have no significant difference between C10:0 and C14:0. The content of cholesterol in the digestive products is more than in the digestive products of the baby's in vitro. High, PE, PC and SM were hydrolyzed mainly in the process of 424mg/100g and 387 mg/100g. in vitro digestion, and the hydrolysis degree of PE was the highest. The laser confocal microscopic image showed that in the digestive process of the adult stomach, a large number of MFG gathered almost without digestion, and the structure of MFGM remained intact. After digestion of the intestine, it could be obtained. It is found that there are hollow fat particles between the digestive products and the undigested TAG and protein formation. The delay period of the raw milk and pasteurized milk is longer than that of the homogenized milk and the spray dry milk during the digestion of the baby's stomach in vitro. The milk after spray drying is faster than the other three kinds of milk. During the process, there was no significant difference in the lipo rate of raw milk, homogeneous milk and spray drying milk, but the digestion rate of pasteurized milk was the lowest. The highest TAG in homogeneous milk and spray dried milk, 16% and 21%. homogenization and spray drying, were also affected by the content of cholesterol in the digestive products, compared with the milk and pasteurized milk. Reducing 10%. pasteurization, homogenization and spray drying increased the degree of hydrolysis of PC. Four different processed MFG had a partial aggregation of MFG after the digestion of the stomach for 1 h in the baby's stomach, especially the homogenized milk and spray dry milk. After digestion, the MFG was further formed with a smaller particle size, and there was still one after the spray drying milk was digested. The large fat droplets exist. As the cyclic fluorescent structure of MFGM disappears, the whole particle is dyed red by the phospholipid probe Rh-DOPE. Raw milk, pasteurized milk, homogenized milk and spray dry milk are in the first 10 min of the digestive tract of human intestines, the rate of lipid is 23,16.8,21.6 and 20.4 mu mol/m L milk. Pasteurized milk The unhydrolyzed TAG was the highest, which accounted for no effect on the type and content of FFA released by 10.86%. processing, but there was a distinct difference in the fatty acid composition of the hydrolysate MAG. The content of cholesterol in the digestive products was increased after the digestive tract digestion in adults. The MFG in the adult stomach was easy to accumulate in the adult stomach. Milk and pasteurized milk MFG can still detect the ring fluorescence of MFGM after digestion of the stomach of adults in vitro for 1 h. After digestion of 2 h intestines, the ring phospholipid is replaced by the bile salt and MFG is further decomposed into small particles by the pancreatic lipase. However, there are some large particles in the pasteurized milk. Finally, the study is done. The adhesion and infection effect of different MFG on bacteria and HT-29 cells showed that the MFG of different animals (cattle, sheep and human) would promote the adhesion of probiotics (Lactobacillus bulgaricus subspecies, Lactobacillus acidophilus, Bifidobacterium infantile and Bifidobacterium infantile subspecies) with HT-29 cells. Goat milk MFG has good adhesion to bifidobacteria. Human milk MFG showed higher adhesion to Bifidobacterium infantile. Human milk MFG had the smallest adhesion to the Bulgarian subspecies of Lactobacillus Deshi, less milk MFG and goat milk MFG., and the goat milk MFG showed higher adhesion to four probiotics than milk and MFG of human milk. These three mammalian MFG were both induced by MFG. The bacteria (Salmonella, sakazakibacillus sakazakybacilli and Escherichia coli) inhibited the adhesion of HT-29 cells, but the inhibitory effect of goat milk MFG on these three pathogenic bacteria was obviously better than that of bovine MFG, and the inhibitory effect of human milk MFG on Escherichia coli was the strongest. The number of probiotics and pathogenic bacteria adhered to HT-29 cells after processing MFG was different. In addition to pasteurization, the greater the degree of processing, the less the probiotics adhered to HT-29 cells, and the lower the degree of inhibition to pathogenic bacteria.
【学位授予单位】：江南大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：TS252.1

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