源于稻曲病菌的黑粉菌素及其生物活性与检测分析

Rice false smut is one of the most destructive fungal diseases of rice (Oryza sativa L.) production. Ustilaginoidins are a group of mycotoxins produced by the rice false smut pathogen Villosiclava virens (anamorph: Ustilaginoidea virens). They exist in the rice false smut balls, false smut pathogen, as well as in the paddy, husked rice, straw and their products infected by the pathogen. They are phytotoxic to the host plant. Rice production as well as feed and food safety have been seriously threatened by these mycotoxins. Ten ustilaginoidins have been isolated from the rice false smut balls. Our previous results showed that many other ustilaginoidins have not been revealed in the rice false smut balls, false smut pathogen, and other infected rice materials collected from the different areas of China. The purposes of this project attempt to clarify the unknown ustilaginoidins in the rice materials from China including rice false smut balls and pathogen, to establish the standard sample system and HPLC profile for the ustilaginoidins, to evaluate their biological activities such as phytotoxicity and cytotoxicity, to establish the ELISA methods and LC-MS analysis for the main ustilaginoidins. Other aspects such as structure-activity relationships, mechanisms of action, biosynthesis pathway and metabolic regulation of the ustilaginoidins will also been studied. It aims to provide the basis for rice false smut disease control, feed and food safety, as well as the creation of new medicines and agrochemicals.

稻曲病是水稻生产中的一种毁灭性真菌病害。黑粉菌素是一类由稻曲病菌产生的真菌毒素，存在于稻曲球、稻曲病菌和由稻曲病菌污染的谷物、稻米、稻草及其产品中。黑粉菌素不但对水稻有毒害作用，而且影响水稻生产和食用安全性。前人已从稻曲球中分离鉴定出10个黑粉菌素。申请者课题组前期研究表明，源于我国的稻曲球、稻曲病菌，以及由稻曲病菌污染的谷物中尚存在多个未被揭示的黑粉菌素。本申请项目试图系统分离鉴定我国的代表性稻曲球、稻曲病菌等样品中未知的黑粉菌素，建立黑粉菌素的标样体系和液相色谱指纹图谱，评价黑粉菌素的植物毒、细胞毒等生物活性，建立主要黑粉菌素的酶联免疫检测方法和液相色谱-质谱分析方法，探讨构效关系和作用机制，同时对黑粉菌素的生物合成途径和代谢调控进行研究，为有效防治稻曲病、保障粮食安全以及药物创制提供依据。

本项目研究了稻曲球和稻曲病菌发酵培养物中黑粉菌素（即：稻绿核菌素）的种类和结构、生物活性、富集、制备、HPLC和ELISA检测分析、生物合成途径等方面。取得的主要成果如下：（1）共分离并鉴定出稻绿核菌素27个（其中新化合物17个：ustilaginoidins A~X、ustilaginoidin E1、2,3-dihydroustilaginoidin T和isochaetochromin B2）及其生物合成前体3个（hemiustilaginoidin F、epihemiustilaginoidin D和hemiustilaginoidin D），还分离鉴定了其他类型的化合物（包括1个新的稻曲菌素、10个新的山梨素、2个新生物碱）；（2）对代谢产物进行了植物毒活性、细胞毒活性、抗菌活性、抗氧化活性、植物细胞生理效应等评价；（3）建立了稻绿核菌素的HPLC分析方法和酶联免疫测定方法；（4）建立了稻绿核菌素的大孔树脂富集和逆流色谱制备工艺；（5）建立了稻绿核菌素的标样体系和指纹图谱；（6）测定了不同发育时期（早期、中期、晚期）稻曲球中稻绿核菌素的含量和种类的动态变化，发现2,3,2’,3’-位羟基化的稻绿核菌素在晚期稻曲球中的含量要明显高于早期和中期的稻曲球样品；（7）开展了稻曲病菌中稻绿核菌素的生物合成途径和调控的研究。研究结果为揭示稻曲病菌中稻绿核菌素的代谢网络和生物合成调控规律、稻绿核菌素的生物活性和生理功能、稻曲病菌与水稻之间的相互关系，同时为有效防治稻曲病、保障粮食和饲料安全，以及新农药和医药创制提供了依据。本项目已发表SCI刊源的论文7篇，培养研究生21名，其中6人获得硕士学位，6人获得博士学位。