[关键词]
[摘要]
目的 研究大蓟Cirsium japonicum内生真菌Coniothyrium sp. DJ-1中抑制新型冠状病毒主蛋白酶(severe acute respiratory syndrome coronavirus 2 main protease,SARS-CoV-2 Mpro)活性的化学成分。方法 运用新型冠状病毒Mpro/3CLpro抑制剂筛选模型从30株菌株中筛选具有抑制活性的菌株和化学成分。采用正相硅胶、反相RP-18柱色谱和HPLC等技术对抑制率高的菌株的次生代谢产物进行分离纯化,通过NMR、质谱等谱学技术解析化合物结构,利用荧光共振能量转移法测定化合物对SARS-CoV-2 Mpro的抑制活性。结果 在30株菌株中发现抑制活性良好的菌株Coniothyrium sp. DJ-1,并从该菌的大米发酵提取物中分离到17个化合物,分别鉴定为德加隆内酯(1)、5'-表细链格孢烯(2)、细链格孢烯(3)、4'-表细链格孢烯(4)、3-表-二氢细链格孢烯A(5)、瓶霉酚(6)、1-脱氧红藻内酯(7)、6-羟基-8-甲氧基-3a-甲基-3a,9b-二氢-3H-呋喃并[3,2-c]异色烯-2,5-二酮(8)、黄色篮状菌酮(9)、链格孢毒素II(10)、二氢链格孢苝醇(11)、2-(2'-羟基丙基)-5-甲基-7-羟基色酮(12)、7-羟基-2-羟甲基-5-甲基-4H-色烯-4-酮(13)、三环链格孢霉烯6b(14)、三环链格孢霉烯D(15)、苯乙醇(16)和对羟基苯甲醛(17)。化合物5、9、11对SARS-CoV-2 Mpro的半数抑制浓度(median inhibition concentration,IC50)分别为42.77、25.85和24.26 μmol/L。结论 化合物1~15和17均为首次从Coniothyrium sp.属中分离得到,化合物5、9、11对SARS-CoV-2 Mpro有较好的抑制作用。通过筛选活性菌株的策略,快速从真菌天然产物中挖掘抗新冠病毒活性的化合物,为抗新冠病毒药物的研发提供了参考。
[Key word]
[Abstract]
Objective To study the chemical constituents with severe acute respiratory syndrome coronavirus 2 main protease (SARS-CoV-2 Mpro) inhibitory activity from Coniothyrium sp. DJ-1, an endophytic fungus isolated from Cirsium japonicum. Methods The strains and chemical constituents both with inhibitory activity were screened from thirty endophytic fungi by using 2019-nCoV Mpro/3CLpro inhibitor screening model. The secondary metabolites of the strain with high inhibition rate were separated and purified by normal phase silica gel column chromatography, reversed-phase RP-18 column chromatography, and HPLC. Their structures were identified by NMR, MS, and optical rotation data. And the compounds’ anti-SARS-CoV-2 Mpro activities were determined by fluorescence resonance energy transfer method. Results The Coniothyrium sp. DJ-1 strain with good inhibitory activity was screened from thirty endophytic fungi. Seventeen compounds were isolated from the rice fermentation extract of Coniothyrium sp. DJ-1, and identified as djalonensone (1), 5'-epialtenuene (2), altenuene (3), 4'-epialtenuene (4), 3-epi-dihydroaltenuene A (5), phialophoriol (6), 1-deoxyrubralactone (7), 6-hydroxy-8-methoxy-3a-methyl-3a,9b-dihydro-3H- furo[3,2-c]isochromene-2,5-dione (8), talaroflavone (9), altertoxin II (10), dihydroalterperylenol (11), 2-(2'-hydroxypropyl)-5- methyl-7-hydroxychromone (12), 7-hydroxy-2-hydroxymethyl-5-methyl-4H-chromen-4-one (13), tricycloalternarene 6b (14), tricycloalternarene D (15), 2-phenylethan-1-ol (16) and 4-hydroxybenzaldehyde (17). Compounds 5, 9 and 11 exhibited potent inhibitory effects on SARS-CoV-2 Mpro with IC50 values of 42.77, 25.85 and 24.26 μmol/L, respectively. Conclusion Compounds 1 —15 and 17 have been isolated from the extracts of Coniothyrium sp. genus for the first time. Compounds 5 , 9 and 11 exhibit potent inhibitory effects on SARS-CoV-2 Mpro. The research concludes that the method of screening active strains is helpful for us to quickly excavate active compounds from natural fungal products, moreover provides a basis for the research and development of drugs for COVID-19.
[中图分类号]
R284.1
[基金项目]
国家自然科学基金项目(31960095);国家自然科学基金项目(82160670);云南省应用基础项目(202101AS070022,202201AT070228);
云南省万人计划-青年拔尖人才(W.-G. Wang)