A new stachybotrin congener from a soil fungus Stachybotrys parvispora strain HS-FG-843
A new stachybotrin congener from a soil fungus Stachybotrys parvispora strain HS-FG-843
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来自土壤真菌 Stachybotrys parvispora 菌株 HS-FG-843 的新水苏霉素同系物
DOI：
10.1038/ja.2014.146
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
W. Xiang
中科院分区：
文献类型：
--
作者：
Dan Zhou;Lian;Huan Qi;Jun;Hui Zhang;Jidong Wang;W. Xiang
真菌已被认为是发现新的生物活性化合物的一个有吸引力的来源真菌的几种生物活性代谢物，如青霉素、头孢菌素、美伐他汀和洛伐他汀，已成为制药工业的重要产品为了从真菌中寻找生物活性代谢物，我们对各种生态类群的真菌缺陷进行了研究，发现了几种新的具有抗肿瘤活性的天然产物。4-6作为我们从土源真菌imperfeci中持续筛选更活跃的次级代谢物的一部分，我们从Stachybotrys parvispora菌株HS-FG-843中获得了一个新的stachybotrin同系物，命名为stachybotrin G(1)和一个已知的代谢物stachybotrin(2)。本文详细介绍了新化合物的发酵、分离、结构鉴定和生物活性。从湖南省竹林峰顶土壤样品中分离到小孢子s - fg -843菌株。由山东农业大学张天宇教授提供并鉴定为S. parvispora。菌株HS-FG-843保存于浙江海正集团股份有限公司医药科研培养库，入库号：hs - fg - 843。菌株在马铃薯葡萄糖琼脂斜面上培养，24°C孵育6-7 d。将原液移入含有250ml种子培养基的1 l Erlenmeyer烧瓶中，24℃孵育24 h， 150转/分摇匀。然后将1 l移入含有30 l生产培养基的50 l发酵罐，生产培养基由蛋白胨0.5%、马铃薯淀粉0.5%、酵母浸膏0.2%、NaCl 0.4%、KH2PO4 0.1%、MgSO4·7H2O 0.05%、CaCO3 0.2% （pH 6.2-6.4）组成。在24℃下发酵7天，以100转/分的速度搅拌，曝气率为每小时900升。从50 l发酵罐中过滤出最后30 l的肉汤，用水（3 l）洗涤所得饼，随后用甲醇（3 l）提取。上清液和洗涤水经HP-20树脂（Mitsubushi Chemical Co., Ltd, Tokyo, Japan）用95% EtOH （5 l）柱洗脱。将MeOH萃取物和EtOH洗脱液在50℃下减压蒸发至1l的体积，用等体积的EtOAc萃取三次所得浓缩液。结合的乙酸乙酯相在减压下浓缩，得到混合物（15g）。混合物经Sephadex LH-20凝胶（GE Healthcare, Glies， UK）柱，CHCl3/MeOH （1:1, v/v）洗脱，薄层色谱检测，得到四个组分（Fr.1至Fr.4）。fr2在硅胶柱（青岛海洋化工集团，山东青岛，中国；100-200目）上进行层析，以CHCl3/MeOH （100:0-50:50, v/v）梯度逐级洗脱，根据TLC谱图得到fr2 -1 ~ fr2 -3三个馏分。用CHCl3/MeOH （90:10-70:30, v/v）洗脱，得到三个馏分（Fr.2-2-1, Fr.2-2-2和Fr.2-2-3）。采用半制备型高效液相色谱法（Agilent 1100, Zorbax SB-C18, 5μm, 250× 9.4mm i.d; 1.5 mlmin 1; 220 nm; Agilent, Palo Alto， CA， USA）用甲醇/水（78:22,v/v）洗脱，进一步分离得到化合物1 （tR 23.4min, 3.6 mg）。用甲醇/水（74:26）半制备高效液相色谱（HPLC）对Fr.2-2-1进行纯化，得到stachybotrin (2) （tR 23.2min, 23mg）。1H和13C NMR波谱用Bruker DRX-400光谱仪（1H 400MHz, 13C 100MHz）测量（Bruker, rheinsteten, Germany）。采用Q-TOF Micro LC-MS-MS质谱仪（Milford， MA， USA）进行ESIMS和高分辨率电喷雾电离质谱分析。化合物1为白色无定形粉末，1 / 2a 25 D þ 50 （c 0.036, EtOH）和UV (EtOH) λmax nm （log ε）分别为338 nm (4.89), 285 nm (4.95), 228 nm（5.08）。分子式为C28H38N2O5（化学式m/z 483.2846 [m +H]+, calcd 483.2853）。1的红外光谱在官能团区3414 cm 1 （OH）、1650 （C=O）、1610 cm 1 （C=O）处有吸收带。1的1H NMR谱在δH 6.93 （1H, s）处有一个芳香质子单重态，δH 0.85 （3H, s）、0.96 （3H, s）和0.98 （3H, s）处有三个叔甲基信号，δH 0.69 （3H, d， J= 6.5 Hz）处有一个脂肪族甲基双重态。其13C NMR和DEPT谱显示28个碳共振，包括δ c169.4 （s, 2C）的两个酰胺或酯羰基碳，一个sp 2甲基，
Fungi have been regarded as an attractive source for discovering new bioactive compounds.1,2 Several bioactive metabolites from fungi, such as penicillins, cephalosporins, mevastatin and lovastatin, have been the important products of pharmaceutical industry.3 With the aim of searching for bioactive metabolites from fungi, our studies of fungi imperfeci from a variety of ecological groups have resulted in the discovery of several kinds of new natural products with antitumor activity.4–6 As part of our continuous screening for more active secondary metabolites from soil-derived fungi imperfeci, a new stachybotrin congener, named stachybotrin G (1) and a known metabolite, stachybotrin (2) were obtained from Stachybotrys parvispora strain HS-FG-843. Here, the report details the fermentation, isolation, structure elucidation and bioactivity of the new compound. The S. parvispora strain HS-FG-843 was isolated from a soil sample collected from the peak of a bamboo forest of Hunan province, China. It was provided and identified as S. parvispora by Professor Tianyu Zhang at the Shandong Agricultural University, China. The strain HS-FG-843 has been deposited in the Pharmaceutical Research Culture Collection, Zhejiang Hisun Group Co., Ltd with accession No. HS-FG-843. The strain was grown and maintained on potato dextrose agar slant and incubated for 6–7 days at 24 °C. The stock culture was transferred into 1 l Erlenmeyer flasks containing 250ml of the seed medium and incubated at 24 °C for 24 h, shaken at 150 r.p.m. Then, 1 l of the culture was transferred into a 50-l fermentor containing 30 l of producing medium consisting of peptone 0.5%, potato starch 0.5%, yeast extract 0.2%, NaCl 0.4%, KH2PO4 0.1%, MgSO4·7H2O 0.05%, CaCO3 0.2% (pH 6.2–6.4). The fermentation was carried out at 24 °C for 7 days stirred at 100 r.p.m. with an aeration rate of 900 l of air per hour. The final 30 l of broth from 50 l fermentor was filtered and the resulting cake was washed with water (3 l) and subsequently extracted with MeOH (3 l). The supernatant and the wash water were subjected to a Diaion HP-20 resin (Mitsubushi Chemical Co., Ltd., Tokyo, Japan) column eluting with 95% EtOH (5 l). The MeOH extract and the EtOH eluents were evaporated under reduced pressure to a volume of 1 l at 50 °C and the resulting concentrate was extracted three times using an equal volume of EtOAc. The combined EtOAc phase was concentrated under reduced pressure to yield a mixture (15 g). The mixture was subjected to a Sephadex LH-20 gel (GE Healthcare, Glies, UK) column eluted with CHCl3/MeOH (1 : 1, v/v) and detected by TLC to give four fractions (Fr.1 to Fr.4). The Fr.2 was chromatographed on a silica gel (Qingdao Haiyang Chemical Group, Qingdao, Shandong, China; 100–200 mesh) column and successively eluted with a stepwise gradient of CHCl3/MeOH (100:0–50:50, v/v) to obtain three fractions Fr.2-1 to Fr.2-3 based on the TLC profiles. The Fr.2-2 was subjected to another silica gel column eluted with CHCl3/MeOH (90:10–70:30, v/v) to give three fractions (Fr.2-2-1, Fr.2-2-2 and Fr.2-2-3). Fr.2-2-3 was further isolated by semi-preparative HPLC (Agilent 1100, Zorbax SB-C18, 5μm, 250× 9.4mm i.d; 1.5 mlmin 1; 220 nm; Agilent, Palo Alto, CA, USA) eluting with MeOH/H2O (78:22, v/v) to obtain compound 1 (tR 23.4min, 3.6 mg). The Fr.2-2-1 was purified by semi-preparative HPLC eluting with MeOH/H2O (74:26) to yield stachybotrin (2) (tR 23.2min, 23mg). 1H and 13C NMR spectra were measured with a Bruker DRX-400 (400MHz for 1H and 100MHz for 13C) spectrometer (Bruker, Rheinstetten, Germany). The ESIMS and high-resolution electrospray ionization MS (HRESIMS) spectra were taken on a Q-TOF Micro LC-MS-MS mass spectrometer (Milford, MA, USA). Compound 1 was isolated as a white amorphous powder with 1⁄2a 25 D þ 50 (c 0.036, EtOH) and UV (EtOH) λmax nm (log ε): 338 nm (4.89), 285 nm (4.95), 228 nm (5.08). Its molecular formula was determined as C28H38N2O5 according to HRESIMS (found m/z 483.2846 [M+H]+, calcd 483.2853). The IR spectrum of 1 showed absorption bands at 3414 cm 1 (OH), 1650 (C=O), 1610 cm 1 (C=O) in the functional group region. 1H NMR spectrum of 1 showed an aromatic proton singlet at δH 6.93 (1H, s), three tertiary methyl signals at δH 0.85 (3H, s), 0.96 (3H, s) and 0.98 (3H, s), one aliphatic methyl doublet at δH 0.69 (3H, d, J= 6.5 Hz). Its 13C NMR and DEPT spectra displayed 28 carbon resonances, including two amide or ester carbonyl carbons at δC 169.4 (s, 2C), one sp 2 methine,
影响因子：
5.1
作者：
Wang X;Sena Filho JG;Hoover AR;King JB;Ellis TK;Powell DR;Cichewicz RH
通讯作者：
Cichewicz RH