40 6 Vol 40 No 6 2016 11 Journal of Jiangxi Normal UniversityNatural Science Nov 2016 1000-5862201606-0569-05 SF4 1 2 2 2 2 2 3 3 4* 1 330025 2 330022 3 330013 4 336000 ITS-rDNA AChEI SF4 Acremonium sp Acremonium sp SF4 SF4 AChEI 28 150 r min - 1 14 d AChEI 0 331 g L - 1 60 27% AChEI R 284 2 A DOI10 16357 /j cnki issn1000-5862 2016 06 05 0 ITS-rDNA SF4 Acetylcholinesterase Inhibiters AChEI Alzhe- AChEI imer s disease AD AChEI AChEI AChEI AD 1-2 1 AChEI 1 1 AChEI SF4 2-6 AChEI ATCI 5 5- -2- DTNB 7-8 AChE Sigma HupA Sabouraud's dextrose broth SDB 12 Cazpek broth CB 13 Potato dextrose broth PDB 12 9-11 1 2 Huperzia serrata 1 2 1 14 AChEI SF4 2016-06-01 81260617 31300051 20142BAB214020 1986-
570 2016 160 r min - 1 10 d 1 2 2 ITS-rDNA CTAB 1 2 5 1 2 6 DNA 15 PCR 7 AChE 1 2 3 ITS-rDNA 3 7Neighbour-joining 80 150 200 r min - 1 10 d 1 2 5 1 2 4 205 ml 1 2 6 60 ml PDB 3 d AChE 4 25 150 r min - 1 3 d 500 ml 120 ml 10 14 18 d 1 2 5 8% 1 2 6 AChE 1 2 5 1 2 8 1 2 7 AChE 3 95% 2 d 3 2 1 mg 2 1 1 ml PBS ph 7 7 SF4 PDA 1 mg ml -1 25 7 d 20 μg ml - 1 25 ~ 35 mm AChE 1 2 6 AChE G L Ellman 16 17 I I = OD - OD 1 100% OD 1 2 7 1 2 8% 14 18 PDB SDB CB 25 150 r min - 1 SF4 10 d 1 2 5 Hyphomycetes Moniliales 1 2 6 Moniliaceae Amerosporoideae of Moniliaceae Botrytideae AChE 2 SDB Acremonium 25 28 30 1 SF4 PDA 2 SF4
6 SF4 571 2 2 SF4ITS-rDNA sp P42E5 99% 3 ITS-rDNA ITS1 ITS4 ITS-rDNA SF4 549 bprdna GU985205Blast Acremonium sp Acremonium Acremonium sp SF4 3 SF4 ITS-rDNA 2 3 SF4 AChE AChEI 4 SF4 PDB 2 4 1 1 AChE PDB SBD 11 23% AChE CB SBD 46 94% SF4 AChEI AChE PDB SF4 AChE SBD SF4 AChE 1 SF4 SBD PDB CB / g L - 1 0 191 ± 0 006 0 185 ± 0 009 0 089 ± 0 012 AChE /% 57 25 ± 3 18 46 94 ± 2 19 45 36 ± 1 63 2 4 2 2 SBD 30 28 25 28 4 SF4 AChE AChE 2 4 65 13% 30 25 AChE AChE 28 SF4 AChEI SF4 25 AChE AChE AChEI
572 2016 28 2 19% SBD 2 SF4 / 25 28 30 / g L - 1 0 211 ± 0 026 0 289 ± 0 014 0 315 ± 0 016 AChE /% 51 95 ± 2 56 65 13 ± 3 21 37 89 ± 2 42 2 4 3 3 3 SBD 7 19-20 28 150 r min - 1 200 r 7-8 min - 1 1 SF4 AChEI 80 r min - 1 AChEI 3 AChEI AChE SF4 150 r min - 1 3 SF4 SF4 AChEI SBD / r min - 1 80 150 200 / g L - 1 0 276 ± 0 031 0 319 ± 0 023 0 257 ± 0 008 AChE /% 62 05 ± 2 56 60 18 ± 3 17 63 33 ± 1 26 ITS-rDNA SF4 2 4 4 Acremonium sp Acremonium sp SF4 4 14 d SF4 0 327 g L - 1 18 d 4 1Blusztajn J K Berse B The cholinergic neuronal phenotype in Alzheimer s disease J Metabolic Brain Dis- 3 AChE ease 2000 15145-64 14 d 2Zhao Qin Tang Xican Effects of huperzine A on acetylcholinesterase isoforms in vitrocomparison with 4 SF4 tacrine /d 10 14 18 / g L - 1 0 251 ± 0 021 0 327 ± 0 013 0 237 ± 0 006 AChE /% 65 05 ± 1 71 63 18 ± 2 39 61 38 ± 1 18 2 5 4 J 2008 2721-5 5 PDB 25 150 r J min - 1 10 d 0 185 ± 2014 385489-495 0 009g L - 1 AChE 46 94 ± 28 150 r min - 1 14 d 0 331 ± 0 017g L - 1 AChE 60 27 ± 3 01% SF4 AChEI 8% 28 150 r min - 1 14 d 0 331 ± 0 017g L - 1 AChE 60 27 ± 3 01% donepezil rivastigmine and physostigmine J European Journal of Pharmacology 2002 4552 /3101-107 3Ma Xiaoqiang Gang D R The Lycopodium alkaloids J Natural Product Reports 2004 216 752-772 6Ma Xiaoqiang Tan Changheng Zhu Dayuan et al Hu-
6 SF4 573 perzine A from Huperzia speciesan ethnopharmacolgical 2007 review J Journal of Ethnopharmacology 2007 1131 13 M 5 15-34 1997 7Wang Ya Zeng Qinggui Zhang Zhibin et al Isolation and 14 M 1979 characterization of endophytic Huperzine A-producing fun- 15Guo L D Hyde K D Liew E C Y Identification of endophytic fungi from Livistona chinensis based on morphology gi from Huperzia serrata J Journal of Industrial Microbiology & Biotechnology 2011 389 1267-1278 and rdna sequences J New Phytologist 2000 147 8Yan Riming Zhang Zhibin Wang Ya et al Efficient strategy for maintaining and enhancing the huperzine A pro- 16Ellman G L Courtney K D Andre V A new and rapidcolo- 3617-630 duction of Shiraia sp Slf14 through inducer elicitation rimetric determination of acetylcholinesterase activity J J Journal of Industrial Microbiology & Biotechnology Biochemical Pharmocology 1961 72 88-95 2014 417 1175-1179 17 g5 9Stierle A Strobel G Stierle D Taxol and taxane production J 2008 35 by Taxomyces andreanae endophytic fungus of Pacific 111764-1768 Yew J Science 1993 2609 214-216 18 M 10Stone J K Bacon C WWhite J F An overview of endophytic microbesendophytism defined M Bacon C W 19Rivera-Chaávez J Figueroa M Gonzaález MDC et al α- 2009 White JFJr Microbial Endophytes New YorkMarcel Dekker 2000 with Hintonia latiflora J Journal of Natural Products 11Wang Ya Gao Boliang Li Xixi et al Phylogenetic diversi- 2014 784730-735 Glucosidase inhibitors from a Xylaria feejeensis associated ty of culturable endophytic fungi in Dongxiang wild rice 20Liu Ling Gao Hao Chen Xulin et al Brasilamides A-D Oryza rufipogon Griffdetection of polyketide synthase sesquiterpenoids from the plant endophytic fungus Paraconiothyrium brasiliense J European Journal of Organic gene and their antagonistic activity analysis J Fungal Biology 2015 11911 1032-1045 Chemistry 2010 173302-3306 12 M The Identification and Optimization of Fermentation Conditions of Acetylcholinesterase Inhibiters-Producing Endophytic Fungi Strain SF4 LI Xixi 1 2 YAN Riming 2 ZHANG Zhibin 2 YANG Huilin 2 ZHU Du 2 3 WANG Ya 3 4* 1 Science and Technology College Jiangxi University of Traditional Chinese Medicine Nanchang Jiangxi 330025 China 2 Key Laboratory of Protection and Utilization of Subtropic Plant Resources of Jiangxi Province Jiangxi Normal University Nanchang Jiangxi 330022 China 3 Key Lab of Bioprocess Engineering of Jiangxi Province College of Life Sciences Jiangxi Science and Technology Normal University Nanchang Jiangxi 330013 China 4 Key Laboratory for Research on Active Ingredients in Natural Medicine of Jiangxi Province Yichun University Yichun Jiangxi 336000 China AbstractThe acetylcholinesterase inhibitersachei-producing endophytic fungi strain SF4 was identified as Acremonium sp SF4 based on its morphological characteristics and ITS-rDNA sequence analysis The effects of fermentation medium temperature rotation speed and cultural time on acetylcholinesterase AChEinhibitory activity and AChEI metabolites amount were analyzed It was indicated that the appropriate fermentation condition wassabouraud dextrose brothsdbmedium 28 150 r min - 1 for 14 d Under the optimized conditions of fermentation the amount of AChEI metabolites and AChE inhibitory activitycytotoxic activity is 0 331 g L - 1 and 60 27% respectively higher than initial conditions These results can lay the foundation for the further separation and purification of the AChEI metabolites Key wordsendophytic fungiacetylcholinesterase inhibitersacetylcholinesteraseidentificationfermentation