【摘要】 基于传统热处理方式的蛋白质糖基化反应存在反应时间长、易产生有害糖基化反应末期产物等缺点。以卵类粘蛋白（OVM）-还原糖（1∶0.03,W/W）体系为对象，研究新型高温热处理技术-过热蒸汽对OVM糖基化反应及蛋白质结构的影响。结果表明：短时间（1～3 min）的过热蒸汽（110和120℃）处理即可促使OVM发生糖基化反应，其中不同糖的糖基化反应活性顺序为核糖（五碳糖）>葡萄糖（六碳糖）>乳糖（二糖）,糖基化反应后蛋白质的自由氨基含量由19.97 mg·mL^-1分别最低降为2.93, 5.04和6.69 mg·mL^-1。不同条件处理下OVM糖基化产物的紫外光谱未见显著的波峰位移，但最大吸收峰峰值发生一定的变化，其中大部分为降低的，说明OVM的球蛋白分子结构发生变化，还原糖的修饰掩盖了OVM上的发色基团（苯丙氨酸和酪氨酸）,其中核糖糖基化反应下效果最为显著。糖基化反应后，荧光光谱峰强度显著降低，其中核糖糖基化反应降低率最大，其次为葡萄糖和乳糖，表明蛋白质内部暴露的荧光基团去折叠导致荧光猝灭。此外，还原糖作为猝灭剂渗透入蛋白质框架中，与荧光基团反应，抑制其荧光发射。红外光谱分析发现经过热蒸汽处理后OVM糖基化产物3 300 cm^-1处的峰变窄，表明N—H的含量减少或者发生反应，从而使N—H伸缩振动频率降低。2 000～2 500和500 cm^-1附近出现了较多杂峰，表明蛋白质发生降解或者有糖基化反应中间产物生成。MALDI TOF MS分析表明热蒸汽处理下，OVM-核糖体系、 OVM-葡萄糖体系各有约6个糖基化反应位点，OVM-乳糖体系有约2个糖基化反应位点。糖基化反应后有OVM二聚体、三聚体和多聚体形成，其中核糖和葡萄糖与OVM反应产生的聚合物最为显著，乳糖与OVM反应产生的聚合物较少。该研究可为短时微糖糖基化反应及过热蒸汽在食品加工中的应用提供一定的技术和理论指导。更多还原

【Abstract】 Protein glycation reaction based on traditional heating methods has disadvantages, such as long reaction time consumption and easy to produce harmful advanced glycation end products. In this study, the ovomucoid（OVM）-reducing sugar（1∶0.03, W/W） system served as the research object, effects of superheated steam, a new high-temperature treatment technology, on the OVM glycation and protein structure were studied. The results showed that the glycation of OVM was induced by a short time（1～3 min） of superheated steam treatment（110 and 120 ℃）, and the sequence of glycation reaction activity of different sugars was ribose（pentose） > glucose（hexose） > lactose（disaccharide）. After glycation, the free amino content of protein decreased from 19.97 mg·mL^-1 to the minimum of 2.93, 5.04 and 6.69 mg·mL^-1, respectively. The ultraviolet spectra of OVM glycation products under different processing conditions showed no significant changes in wave displacement. However, the maximum absorption had some changes, with most of them were reduced, indicating OVM globulin molecular structure was altered, and the chromophore groups（phenylalanine and tyrosine） of OVM were masked by the reducing sugars. The effect was most significant under ribose glycation. After glycation, the fluorescence intensities were significantly decreased, in which the ribose glycation decreased the most, followed by glucose and lactose, indicating that the fluorescence quenching occurred, which resulted from the folding of exposed fluorophore in protein. In addition, reducing sugar as a quenching agent penetrated the protein framework and reacted with fluorophore, inhibiting the fluorescence emission. Fourier transform infrared spectroscopy showed that, after superheated steam treatment, the protein peak at 3 300 cm^-1 became narrow, indicating that the content of N—H decreased or a reaction occurred, thus reducing the stretching vibration frequency of N—H. Some fingerprint peaks in the vicinity of 2 000～2 500 and 500 cm^-1appeared, indicating that the protein degradation or glycation middle product formation occurred. MALDI TOF MS analysis showed that there were about 6 glycation reaction sites in each the OVM-ribose system and OVM-glucose system, and about 2 glycation reaction sites in the OVM-lactose system. Moreover, after glycation, OVM dimer, trimer and multimer were formed. Ribose and glucose produced the most significant polymers with OVM, while lactose produced fewer polymers with OVM. This study can provide technical and theoretical guidance for the glycation reaction featured by trace amounts of sugar participation, short time consumption, and application of superheated steam in food processing.更多还原