电子束辐照场对花生粕中黄曲霉毒素 B1 消减机制研究

Aflatoxins (AFT) are highly toxic, mutagenic, teratogenic and carcinogenic compounds. Although 4 million ton peanut meals are produced every year in our country, it could not be utilized sufficiently because of the high quantity of AFT. Thus, a safe and effective detoxification method in peanut meal is needed to find at present situation, which are the short of present detoxification method, the need to scale applications, and the heterogeneous nature of peanut meal. It is found that 67.96% AFB1 can be deleted by 5 kGy electron beam irradiation (EBI), and at the same time the quantity of the peanut meal is not affected by it. The most important is that the detoxification method can be used in industrial. But the mechanism of EBI to remove AFB1 remains unknown, which become the development bottleneck of EBI in food safety..AFB1 in peanut meal is the target in this study, and the degradation behavior & degradation kinetics will be studied firstly. Secondly, the degradation products will be screened and identified by UPLC-Q-TOF/MS and HPLC-NMR. And then, the kinetic variation of the degradation products under different condition will be tracked and the degradation pathway will be proposed at the same time. At last, the degradation mechanism of AFB1 in peanut meal using EBI will be elucidated from the perspective of quantum chemistry, and the cytotoxicity of the degradation products will be evaluated by in vitro evaluating model. Theoretical direction and basis information will be supplied for EBI to be applied in food widespread.

我国花生粕年产400万吨，因黄曲霉毒素（AFT）超标严重影响其充分利用，针对现有消减方法缺陷、规模化应用需求及花生粕非均质固态特性等现状，寻找安全有效又不影响花生粕品质的消减AFT方法亟待解决。申请人前期研究已发现5 kGy电子束辐照场（EBI）可穿透花生粕消减其中67.96%AFB1，同时证实EBI在剂量范围内对花生粕品质无影响且有规模化应用潜质，但降解机理尚不明确，这成为该法消减花生粕中AFT应用的瓶颈。本研究以AFB1为目标物，研究EBI作用下AFB1降解行为及动力学；采用超高效液相色谱-四级杆飞行时间质谱与高效液相色谱-核磁共振联用技术，筛查AFB1各级降解产物，追踪降解物动态变化，阐明AFB1的降解路径，结合量子化学从能量角度诠释电子束辐照场消减AFB1机理；采用体外AFB1细胞毒理模型评估降解产物的安全性，为EBI在食品行业的应用提供理论指导和基础数据。

花生粕营养丰富，但花生极易感染真菌，产生黄曲霉毒素，严重限制了花生粕的应用。电子束（EB）加速辐照技术作为一种去除黄曲霉毒素的新方法，具有高效、安全、等优点，广泛应用于食品的杀菌、脱毒等。该技术在去除简单体系中黄曲霉毒素的有效性已得到证实，但应用到花生粕中黄曲霉毒素去除的有效性、降解机理、安全性及对花生粕品质的影响等方面仍有待研究。本课题以黄曲霉毒素B1（简称AFB1）为研究对象，研究其在水以及花生粕中的降解行为、降解产物、路径等机理，并通过鼠伤寒沙门氏菌回复突变试验和细胞毒理实验对降解产物的致突变性和细胞毒性进行了研究，同时评估了EB去毒工艺对花生粕品质的影响。. 首先，研究花生粕中辐照剂量、初始浓度、物料厚度、水分含量等因素对AFB1电子束降解行为的影响。结果表明，花生粕中AFB1的降解效果与辐照剂量有关，受初始浓度、固体粕厚度和水分含量影响。在实验范围内，随着辐照剂量、初始浓度和固体粕中水分含量的增加，花生粕中AFB1的降解效果增强；固体粕厚度增加降解率降低。. 然后，运用UPLC-Q-TOF/MS技术鉴定出水中五种降解产物分别为P1（C16H12O7）、P2（C16H16O6）、P3（C17H14O8）、P4（C15H12O6）和P5（C17H12O7）。五种EB降解产物结构与AFB1类似，只在部分基团稍有不同。花生粕中AFB1电子束降解反应剧烈，AFB1经过电子束辐照以后，迅速发生氧化反应，断裂为小分子物质。. 最后，应用Ames和细胞毒理实验研究水、花生粕两种体系中AFB1降解产物的致突变性和细胞毒性。结果表明：阳性对照AFB1对鼠伤寒沙门氏菌组氨酸缺陷型菌株和HepG2细胞毒性强烈，导致细胞活性降低72.44%。经过EB去除两种体系中AFB1后，辐照降解产物致突变性和细胞毒性已大幅度降低（p＜0.001），其中水中EB降解产物致突变性降低50%以上，细胞活性降低18.61%，花生粕中降解产物已无毒性（p＜0.001）。