基于受体识别的白酒中塑化剂多残留荧光偏振检测方法研究

Phthalate esters (PAEs) have been widely used as plasticizers. As the PAEs are not chemically bound to plastic, they can migrate into the environment, cause pollution, and be hazardous to health. PAEs are mainly detected by chromatographies and immunoassays at present. The former can only be conducted in the laboratory and the latter focus on detecting one PAE or several PAEs, both of which have limitations. Nowadays, there is an urgent need to develop a fast, on-site and broad-spectrum method for monitoring multiple PAEs simultaneously to reduce the time and the cost of detection. In this work, a fluorescence polarization assay will be developed based on mPPARα-LBD* for the determination of PAEs in white wine by using C4-BODIPY-C9 as a probe. Furthermore, molecular docking approach will be performed to explore the probable binding mode between PAEs and mPPARα-LBD* for establishing the discriminate model to predict the PPARs binding efficiencies of novel ligands, which is helpful to new, biologically inert plasticizers screening. This work has important significance for control of PAEs residues and assurance of food safety.

邻苯二甲酸酯类塑化剂（PAEs）是广泛使用的塑料助剂，随着时间的推移，可迁移到环境中产生污染，进而危害人体健康。常用的塑化剂检测方法包括色谱分析法和免疫分析法。前者只能在实验室开展工作，后者只能识别一种或少数几种塑化剂，具有局限性。因此，开发塑化剂多残留的快速检测方法，对大量样品进行现场初筛，可降低检测费用，提高检测效率。本项目基于过氧化物酶体增殖物激活受体（PPARs）可特异性识别邻苯二甲酸酯类物质，以mPPARα-LBD*为识别元件，以C4-BODIPY-C9为荧光探针，建立白酒中塑化剂多残留荧光偏振检测方法。同时，利用分子对接技术，研究PAEs与mPPARα-LBD*的相互作用，建立PPARs配体活性预测模型，有助于筛选新的生物惰性塑化剂。本项目的实施，对于监控塑化剂残留、保障食品安全具有重要意义。

邻苯二甲酸酯类塑化剂（PAEs）是广泛使用的塑料助剂，随着时间的推移，可迁移到环境中产生污染，进而危害人体健康。开发塑化剂多残留的快速检测方法，对大量样品进行现场初筛，可降低检测费用，提高检测效率。.本项目基于过氧化物酶体增殖物激活受体（PPARs）可特异性识别邻苯二甲酸酯类物质，以mPPARα-LBD*为识别元件，以C4-BODIPY-C9为荧光探针，建立白酒中塑化剂多残留荧光偏振检测方法。优化后的荧光偏振分析条件为：探针浓度40nM，蛋白浓度80nM，孵育时间10分钟。竞争性结合反应表明，10种PAEs均可竞争受体的结合位点，以剂量依赖方式抑制C4-BODIPY-C9与mPPARα-LBD*的结合。对白酒产品重点监控的DEHP和DBP的检测限分别为8.9和0.7μM，低于国家卫计委评估的安全浓度。空白白酒样品中添加DEHP和DBP的平均回收率约为80%，变异系数低于10%，重复性较好，基本可满足白酒样品检测的要求。.本项目还利用分子对接技术，研究了PAEs与mPPARα-LBD*的相互作用，发现两者的结合力以氢键和疏水相互作用为主。建立了PPARs配体活性预测模型，基于结合位点和对接打分值，可在理论上快速判别PPAR未知配体的亲和力，尤其适用于指导筛选新的生物惰性塑化剂。.本项目的实施，对于监控塑化剂残留、保障食品安全具有重要意义。