杏鲍菇副产物免疫活性物质追踪与结构鉴定  

Trace and Structural Identification of Immunomodulate Activity Substances from Pleurotus eryngii's Byproducts

作　　者：樊铭聪[1,2] 李文香 胡欣蕾[1,2] 孙亚男 于戈 张欣[1,2] 李铭 Fan Mingcong;Li Wenxiang;Hu Xinlei;Sun Yanan;Yu Ge;Zhang Xin;Li Ming(Food Science and Engineering College,Qingdao Agricuitural University,Qingdao 266109,Shandong;Shandong Province Laboratory of Applied Mycology,Qingdao 266109,Shandong)

机构地区：[1]青岛农业大学食品科学与工程学院,山东青岛266109 [2]山东省应用真菌重点实验室,山东青岛266109

出　　处：《中国食品学报》2018年第9期254-264,共11页Journal of Chinese Institute Of Food Science and Technology

基　　金：山东省科技发展计划项目(2014GNC113007);山东省现代农业产业技术体系建设经费资助项目(SDAIT-07-07)

摘　　要：以杏鲍菇副产物为试验材料,采用不同极性部位分段萃取、硅胶柱层析2个步骤对甲醇提取物中的活性物质进行分离纯化,并对每步中得到的提取物进行免疫活性研究,观察其对小鼠巨噬细胞的免疫刺激作用,以此对其中的免疫活性物质进行筛选、活性追踪。利用液相串联四级杆飞行时间质谱对主要活性成分进行结构表征。结论显示:对杏鲍菇甲醇提取物进行系统溶剂法分段萃取,其中正丁醇部位表现出最强的促进细胞增殖活性、吞噬活性和细胞因子分泌活性。当正丁醇部位提取物质量浓度为25μg/m L时,促进Ana-1细胞的增殖和吞噬活性作用最强,细胞因子TNF-α和IL-6均出现分泌量高峰。通过硅胶柱层析对正丁醇部位萃取物进行分离,得到7个洗脱组分,其中Fr.2表现出最强的促进细胞增殖活性、吞噬活性和细胞因子分泌活性。当Fr.2质量浓度为12.5μg/m L时,促进Ana-1细胞增殖作用,吞噬活性最强,细胞因子TNF-α和IL-6出现分泌量高峰。通过液相质谱对Fr.2中主要成分进行结构表征,确定Fr.2中主要成分为腺苷,化学分子式为C10H13N5O4。In order to extract and purify some bioactive substances from the byproducts of Pleurotus eryngii, the methanol extracts of Pleurotus eryngii byproducts was used as raw material and two-step studies were carried out on different polar extracts preparation, slica column chromatography. The immune activity were investigated on those extracts which obtained from two extraction steps though evaluate effect of the immunomodulate activity on mice macrophage cell.With the activity-guided isolation method, the immunomodulate active substances were screened and purified based on the in vitro assays of immunomodulate activity, and the immune functional components were characterized by HPLC-QTOF-MS/MS. The results showed that： four extracts were obtained by different polar organic solvent in order to purify bioactive substances from methanol extracts, butyl alcohol extracts showed the highest promoting Ana-1 cell proliferative activity and cytokines secretion. In concetration of 25 μg/m L, the cell survival rate, phagocytosis ability and cytokines secretion were reached peak value. Followed by silica-gel column chromatography to purify butyl alcohol extracts, 7 elution fractions were obtained and Fr.2 showed the highest promoting Ana-1 cell proliferative activity and cytokines secretion（TNF-α and IL-6）. In concetration of 12.5 μg/m L, the cell survival rate and cytokines secretion were reached peak value. Followed by HPLC-QTOF-MS/MS to elucidate the structure of active substances in Fr.2, the result showed that the main active compound in Fr.2 is adenosine and the chemical formula is C10 H13 N5 O4.

关 键 词：杏鲍菇副产物 免疫活性 活性追踪 结构鉴定 腺苷 

分 类 号：TS209[轻工技术与工程—食品科学]