【作者】 王伟军；

【导师】 张兰威；

【作者基本信息】 哈尔滨工业大学 ， 化学工程与技术， 2013， 博士

【摘要】 牛乳加热处理是乳制品工业的重要环节。虽然出于感官、营养的考虑，低杀菌强度的使用已经成为人们的普遍愿望，而由于工艺和安全的原因也有使用较高杀菌强度的要求。同时，我国的原料乳供应在地域和季节上的不平衡使乳粉的生产和使用始终受到重视，然而乳粉加工中的热相关处理使其复原乳凝固特性受到很大影响。本研究的目的是分析加热处理以及乳粉加工中热相关工艺对乳凝固特性，包括酶诱凝固和酸诱凝固的影响；研究牛乳蛋白聚合和理化特性变化与乳凝固特性的关系以及相关机理；通过化学控制牛乳蛋白聚合和利用特定乳酸菌分别在乳粉加工环节和复原乳干酪生产环节初步解决复原乳酶凝胶及在干酪中的应用问题。不同强度加热乳的酶诱凝固和酸诱凝固特性研究表明：高强度热处理乳显示出更强的酸凝固促进作用，以及更大的酶凝固抑制效果。与同批次原料乳相比，70℃/15s加热乳的酶凝胶硬度降低了16.45%，而90℃/8min则降低了37.27%。采用分子排阻色谱（SEC）技术，本文首次分析了凝胶前的加热乳中乳浆蛋白聚合体（SPA）以及凝胶后的乳清中蛋白聚合体残留随不同加热强度的变化规律。发现SPA的变化与乳凝固质地参数之间存在高度相关性（|r|>0.75，P<0.001）；同时也发现乳清中蛋白聚合体残留与加热强度有高度依赖关系。采用SEC对SPA进行分离并对其性质和作用做进一步研究。结果表明SPA颗粒大小随着处理温度和时间的增加而增加，范围从低强度热处理的约30nm增加到高温处理的约100nm。在酶凝固添加试验中，发现SPA对酶凝固无抑制作用。随后考察了热诱酪蛋白胶束大小、表面疏水性以及活性钙离子浓度的变化，发现酪蛋白直径以及活性钙离子浓度变化是热诱酶凝固特性降低的主要因素。本文考察了乳粉加工中热相关工艺，包括预热杀菌、浓缩和喷雾干燥对乳凝固特性的影响。随着乳粉加工工艺的进行，乳样品酶凝胶的硬度逐步降低，同时，乳样品酸凝胶的硬度呈现逐步上升的趋势；发现经过加热、浓缩和干燥工艺后生产的乳粉，与同批次原料乳相比，其复原乳的酶凝胶硬度降低了73.99%-81.53%。对乳粉加工中不同乳样品的SPA变化、酪蛋白大小和钙离子浓度以及乳粉样品中蛋白二级结构进行了分析，结果表明这些属性均具有高度的热相关工艺及强度的相关性。为了考察热诱酪蛋白胶束与乳清蛋白，特别是与β-乳球蛋白之间的反应，以及其在乳凝固中的作用，本文采用蛋白免疫与电子显微镜结合技术研究了加热和乳粉工艺对乳蛋白聚合的影响。发现加热不会造成酪蛋白胶束的解离和聚集，然而加热会导致乳球蛋白在酪蛋白胶束表面的聚集反应；而在乳粉复原乳的微结构中，除上述发现外，也观察到酪蛋白胶束发生很大程度的解离和重排，因此认为酪蛋白胶束发生根本改变是造成乳粉复原乳酶凝胶特性降低的主要原因。研究了原料乳经碘酸钾（PI）或低聚异麦芽糖处理对随后热诱蛋白聚合分布的控制和修饰作用，SEC发现PI能明显增加SEC的乳清蛋白峰C的强度，降低SEC的蛋白聚合体峰B0和B1的强度。同时也发现这种修饰作用会提升加热乳和乳粉复原乳酶凝胶的质地特性。对PI处理乳的微结构观察也证明了热诱蛋白聚合是可控的，这为加热乳和乳粉复原乳酶凝固特性的改善提供了新的思路。在复原乳干酪（CRMP）的微生物改善研究中，发现直接酸化CRMP在成熟中会发生湿度的降低：从第1天的65.0%降低到第21天的62.5%再到45天的59.9%。而采用特定乳酸菌生产的CRMP能够在成熟期内保持湿度不变，同时使干酪质地得到改善，干酪产量轻微增加（4.6%）；并能极大改变CRMP的蛋白网络微结构，据此认为这是其改善干酪质地和技术特性的主要原因，该研究为CRMP的生产开发提供了理论支撑。更多还原

【Abstract】 The heat treatment of bovine milk is an important process in the dairy industry.For the reasons of sensory and nutritional requirements, the low intensity ofsterilization has become the universal desire of people. However, the high intensityof heat treatment was also used for the purpose of technological demands and safetyreasons. In China, the production of raw milk is imbalance in geography and season.Therefore, the use of milk powder has always received much attention. However, thethermal processes during the production of milk powder changed not only thenutritional characteristics of milk but also its technological applications. The aims ofthis study were to analyze the effects and mechanisms of the heat treatments and thethermal-related processes during the manufacturing of milk powder on thecoagulation properties, including the rennet-induced coagulation and acid-inducedcoagulation, to investigate the relationship of changes in the protein aggregation andphysiochemical characteristic with subsequent difference in the coagulationproperties of milk, and to solve the problems in rennet coagulation of reconstitutedmilk powder and its application in the production of cheese by chemicallycontrolling the aggregation of milk proteins during the production of milk powderand by use of certain lactic acid bacteria in the production of cheese, respectively.The research on the properties of rennet-induced gel and acid-induced gel fromdifferent heated milk showed that the high heat treatment had the increasinginhibitory effect on rennet coagulation and promoting effect on acid coagulation.Compared with raw milk in the same batch, heat treatment of70°C/15s decreasedthe hardness of texture by16.45%, while90°C/8min decreased by37.27%. Sizeexclusion chromatography (SEC) was used to analyze the protein distribution inmilk serum. The changes in serum protein aggregates (SPA) of heated milk beforecoagulation and in the residual SPA of whey after coagulation as affected bydifferent heat treatments were investigated for the first time. High correlations(|r|>0.75, P<0.001) were found between the changes of SPA and the variables ofcoagulation properties. The content of residual SPA depended on the heatingintensity of milk before coagulation.SPA was separated by SEC, and its characteristics and role in rennet-inducedcoagulation were studied. The results showed that the size of SPA increased with theincrease of heating temperature and time, ranging from~30nm at low intensity ofheat treatment to~100nm at high intensity of heat treatment. The rennet coagulationof raw milk with the addition of SPA was not inhibited, suggesting that the inhibition of heating on rennet coagulation did not derive from SPA. Subsequently,the heat-induced changes in casein micelle size, surface hydrophobicity and calciumion activity were investigated. In conclusion, the properties of casein micelles andcalcium ion activity were the main factors that caused the decrease of therennet-induced coagulation properties of milk.In this paper, the effects of thermal-related processes during the production ofmilk powder, including pre-heat treatment, concentration and spray drying on thecoagulation properties of milk were investigated. During the manufacture of milkpowder, the hardness of rennet gel gradually decreased, while that of acid gelshowed a gradual upward trend. The milk powder samples what experienced theprocesses of heat, concentration and drying decreased the hardness of their rennetgel by73.99-81.53%compared with raw milk in the same batch. In addition, thestudies on SPA, the sizes of casein micelles, the calcium ion activity and thecontents of protein secondary structures of milk powder showed that highdependence of these variables was found on the changes of the heat-relatedprocesses and their intensity.In order to investigate the heat-induced aggregation of the casein micelles withwhey proteins, especially with β-lactoglobulin and its role in milk coagulation, theelectron microscopy was used to study the microstructure of heated milk andreconstituted milk powder when combined with the technology of proteinimmunology. Using IPP software the differences of protein microstructures wereanalyzed. In results, the heat treatment did not cause dissociation and aggregation ofcasein micelles, but lead to the aggregation of β-lactoglobulin on the surface ofcasein micelles. It was also found that the casein micelles in reconstituted milkpowder displayed a large degree of dissociation and rearrangement as well as theaggregation of β-lactoglobulin with casein micelles. Therefore the qualitativechange in the microstructure of casein micelles was considered as the main reason ofdecrease of rennet coagulation properties in heated milk and reconstituted milkpowder.By addition of potassium iodate (PI) or isomaltooligosaccharide in raw milk，thecontrol and modification of SPA in subsequent heated milk and reconstituted milkpowder were studied. In results, PI significantly increased the concentration of wheyproteins and decreased the intensity of SEC protein aggregates peak B0and B1.Meanwhile, PI significantly improved the textural properties of rennet gel producedfrom heated milk or reconstituted milk powder. Furthermore, the microstructureresearch on the aggregation of casein micelle with in PI-processed milk alsocorroborated the modification of protein aggregation. Therefore, this finding provided a new idea for the improvement of rennet coagulation properties of heatedmilk and reconstituted milk powder.In addition, an improvements study of microbiology was performed on thecheese from reconstituted milk powder (CRMP). In results, the direct acidifiedCRMP resulted in a decrease of humidity: from65.0%in the first day to62.5%in21days and then to59.9%in45days. While the use of exopolysaccharide(EPS)-producing lactic acid bacteria kept the constant humidity of the cheese duringthe storage, improved the texture and increased slightly (4.6%) the yield of thecheese. Moreover, EPS-producing strain greatly changed the network of cheesemicrostructure, which was the main reason for the improvement of cheese textureand technical characteristics. In conclusion, the related theory and understanding ofthe study could provide the technical guidance for the development and productionof CRMP.更多还原