Identification and Fermentation Conditions of Syringin-producing Endophytic Fungus CJ7

doi:10.11924/j.issn.1000-6850.casb2021-0888

Abstract

Abstract:

The paper aims to efficiently utilize endophytic fungi of Acanthopanax senticosus for the production of syringin. The fermentation broth of endophytic fugus CJ7 was analyzed by HPLC with syringin as reference. The antimicrobial activities of CJ7 were examined by agar diffusion. Meanwhile, the fermentation condition of CJ7 was screened by single factor combined with orthogonal experiments, and the endophytic fungus CJ7 was identified by morphological observation and ITS sequence analysis. The results showed that syringin was detected in the fermentation broth of CJ7, and good inhibitory effects of CJ7 on 10 common pathogens were recognized, especially on Staphylococcus aureus. After being optimized, the best culture condition was as follows: 2.5% glucose and 2.0% beef extract fix in PDA culture with initial pH value of 7.0, medium liquid volume of 40% and inoculation amount of 2%. Besides, the fermentation period was 8 d, and the table rotational speed was set at 120r/min. The strain of CJ7 was identified as Aspergillus fumigatus. As a conclusion, the endophytic fungus CJ7 is a syringin-producing one, and the study can lay a foundation for the production of syringin by microbial fermentation.

Key words: syringin, endophytic fungus, antimicrobial activity, fermentation condition, identification

CLC Number:

R931

ZHANG He, YOU Mengyao, WAN Lu, YAN Jiajia, LIU Songmei, ZHENG Chunying. Identification and Fermentation Conditions of Syringin-producing Endophytic Fungus CJ7[J]. Chinese Agricultural Science Bulletin, 2022, 38(25): 143-150.

Figures/Tables 15

References 29

[1]	国家药典委员会. 中华人民共和国药典[S]. 北京: 中国医药科技出版社, 2020:207.
[2]	BAYGILDIEVA D I, BRAUN A V, STAVRIANIDI A N, et al. Determination of eleutheroside B and eleutheroside E in extracts from Eleutherococcus senticosus by liquid chromatography/mass spectrometry[J]. Journal of analytical chemistry, 2020, 75(14):1832-1837.
[3]	SHI X L, YANG Y, REN H X, et al. Identification of multiple components in deep eutectic solvent extract of Acanthopanax senticosus root by ultra-high-performance liquid chromatography with quadrupole orbitrap masss pectrometry[J]. Phytochemistry letters, 2020, 35:175-185.
[4]	LI X H, ZHANG Z Q, GUO Z H, et al. Acanthopanax senticosus polysaccharide-loaded calcium carbonate nanoparticle as an adjuvant to enhance porcine parvovirus vaccine immune responses[J]. Medicine in drug discovery, 2021, 11:1-10.
[5]	JIA A L, ZHANG Y H, GAO H, et al. A review of Acanthopanax senticosus (Rupr and Maxim.) harms: From ethnopharmacological use to modern application[J]. Journal of ethnopharmacology, 2021, 286:1-17.
[6]	HUANG L Z, HUANG B, YE Q, et al. Bioactivity-guided fractionation for anti-fatigue property of Acanthopanax senticosus[J]. Journal of ethnopharmacology, 2011, 133:213-219.
[7]	ZHANG M L, SUN Y P, LIU Y, et al. Five new sesquiterpenoids from the fruits of Acanthopanax senticosus (Rupr. & Maxim.) Harms[J]. Fitoterapia, 2021, 149:1-6.
[8]	XIA Y G, HUANG Y X, LIANG J, et al. Comparable studies of two polysaccharides from leaves of Acanthopanax senticosus: Structure and antioxidation[J]. International journal of biological macromolecules, 2020, 147:350-362.
[9]	CHEN R H, DU W D, WANG Q, et al. Effects of Acanthopanax senticosus (Rupr. & Maxim.) Harms on cerebral ischemia-reperfusion injury revealed by metabolomics and transcriptomics[J]. Journal of ethnopharmacology, 2021, 264:1-12.
[10]	ZHANG X Q, LAI Y C, WANG L, et al. Phenylpropanoid constituents from Acanthopanax senticosus[J]. Biochemical systematics and ecology, 2011, 39:861-863.
[11]	LI N, SUN Y N, ZHANG L, et al. NF-kB inhibitory activities of triterpenoid glycosides from the stems of Acanthopanax senticosus (Rupr. & Maxim.) Harms[J]. Phytochemistry letters, 2016, 15:210-214.
[12]	ZHAO C J, CHENG J B, LI C Y, et al. Quality evaluation of Acanthopanax senticosus via quantitative analysis of multiple components by single marker and multivariate data analysis[J]. Journal of pharmaceutical and biomedical analysis, 2021, 201:1-6.
[13]	SUN H, HAN Y, ZHANG A, et al. UPLC-MS based metabolic profiling of the phenotypes of Acanthopanax senticosus reveals the changes in active metabolites distinguishing the diversities of the plant grown in northeast area of China[J]. Chinese journal of natural medicines, 2012, 10(3):196-206.
[14]	BRADER G, COMPANT S, MITTER B, et al. Metabolic potential of endophytic bacteria[J]. Current opinion in biotechnology, 2014, 27:30-37.
[15]	KAMBIZ B, NADER H, BEHROUZ H, et al. Isolation and identification of endophytic bacteria from potato tissues and their effects as biological control agents against bacterial wilt[J]. Physiological and molecular plant pathology, 2021, 116:1-9.
[16]	张萍, 吴丹, 矫佳陈琦, 等. 产原儿茶酸北五味子内生细菌WF17的鉴定及抑菌活性研究[J]. 中国食品学报, 2015, 15(7):198-202.
[17]	LIU Y T, YOU Y X, LI Y W, et al. Characterization of carboxymethylated polysaccharides from Catathelasma ventricosum and their antioxidant and antibacterial activities[J]. Journal of functional foods, 2017, 38:355-362.
[18]	邵力平, 沈瑞祥, 张素轩, 等. 真菌分类学[M]. 北京: 中国林业出版社, 1984:17.
[19]	魏景超. 真菌鉴定手册[M]. 上海: 上海科学技术出版社, 1982:84.
[20]	KATHAWUT S, WANPEN L, ANYALAK W, et al. Biological potential and chemical composition of bioactive compounds from endophytic fungi associated with thai mangrove plants[J]. South African journal of botany, 2021, 141:66-76.
[21]	YAN J H, TONG S Q, CHU J J, et al. Preparative isolation and purification of syringin and edgeworoside C from Edgeworthia chrysantha Lindl by high-speed counter-current chromatography[J]. Journal of chromatography A, 2004, 1043:329-332.
[22]	LEE C H, HUANG C W, CHANG P C, et al. Reactive oxygen species mediate the chemopreventive effects of syringin inbreast cancer cells[J]. Phytomedicine, 2019, 61:1-12.
[23]	TAN J Y, LUO J, MENG C C, et al. Syringin exerts neuroprotective effects in a rat model of cerebral ischemia through the FOXO3a/NF-κB pathway[J]. International immunopharmacology, 2021, 90:1-9.
[24]	郭晓璐, 卢玢宇, 郑春英. 刺五加内生细菌的分离及其抑菌活性研究[J]. 中国新药杂志, 2014, 23(22):128-135.
[25]	熊亚南, 邢朝斌, 吴鹏, 等. 刺五加内生真菌分离及分布研究[J]. 安徽农业科学, 2009, 37(24):11347-11348.
[26]	徐红艳, 刘爱华, 王彬, 等. 刺五加内生放线菌的分离与多样性研究[J]. 西南师范大学学报:自然科学版, 2016, 41(5):28-35.
[27]	郑春英, 崔宇, 纪鑫, 等. 药用植物刺五加内生真菌的分离及其抑菌活性研究[J]. 中国药学杂志, 2008, 43(20):1541-1544.
[28]	郑帅. 刺五加内生真菌抗氧化活性研究[D]. 哈尔滨: 黑龙江中医药大学, 2015.
[29]	邢朝斌, 熊亚南, 劳凤云, 等. 刺五加内生真菌对刺五加苷B和E含量的影响[J]. 安徽农业科学, 2009, 37(25):12010-12011.

水平	因素
水平	A.氮源浓度/%	B.碳源浓度/%	C.接种量(v/v)/%	D.发酵时间/d
1	1.5	1.5	1	6
2	2.0	2.0	2	8
3	2.5	2.5	3	10

水平	因素
水平	A.氮源浓度/%	B.碳源浓度/%	C.接种量(v/v)/%	D.发酵时间/d
1	1.5	1.5	1	6
2	2.0	2.0	2	8
3	2.5	2.5	3	10

样品	大肠杆菌	枯草芽孢杆菌	金黄色葡萄球菌	短小芽孢杆菌	单增李斯特菌	鲍曼不动杆菌	肺炎克雷伯菌	铜绿假单孢菌	屎肠球菌	白假丝酵母
CJ7	15.02±0.32	14.79±0.80	18.05±0.17	14.93±0.13	14.02±0.14	18.55±0.09	15.91±0.18	16.53±0.28	17.10±0.24	12.57±0.19
甲醇	-	-	-	-	-	-	-	-	-	-
链霉素	26.71±0.31	27.14±0.30	28.52±0.07	19.13±0.21	24.20±0.35	28.07±0.05	26.50±0.01	28.72±0.30	25.00±0.08	-
制霉素	-	-	-	-	-	-	-	-	-	19.90±0.05

样品	大肠杆菌	枯草芽孢杆菌	金黄色葡萄球菌	短小芽孢杆菌	单增李斯特菌	鲍曼不动杆菌	肺炎克雷伯菌	铜绿假单孢菌	屎肠球菌	白假丝酵母
CJ7	15.02±0.32	14.79±0.80	18.05±0.17	14.93±0.13	14.02±0.14	18.55±0.09	15.91±0.18	16.53±0.28	17.10±0.24	12.57±0.19
甲醇	-	-	-	-	-	-	-	-	-	-
链霉素	26.71±0.31	27.14±0.30	28.52±0.07	19.13±0.21	24.20±0.35	28.07±0.05	26.50±0.01	28.72±0.30	25.00±0.08	-
制霉素	-	-	-	-	-	-	-	-	-	19.90±0.05

组别	因素				抑菌圈直径/mm
组别	A	B	C	D	抑菌圈直径/mm
1	1	1	1	1	25.83±0.30
2	1	2	2	2	28.33±0.30
3	1	3	3	3	29.33±0.30
4	2	1	2	3	30.50±0.00
5	2	2	3	1	29.17±0.70
6	2	3	1	2	30.67±0.70
7	3	1	3	2	23.50±0.00
8	3	2	1	3	21.67±0.40
9	3	3	2	1	26.67±0.20
K1	83.49	81.83	80.17	81.67	ΣY=249.67 CT= 6926.12
K2	94.34	79.17	87.50	84.50
K3	71.84	88.67	82.00	83.50
R	7.50	3.17	2.45	0.95
S	84.41	16.02	9.71	1.38