Detection of Huperzine A and Huperzine B in fermentation broth of endophytic fungus Colletotrichum gloesporioides from Huperzia serrate by HPLC.

Liqin HU; Xincong Kang; Pengyuan Shen; Tian Chen; Jiayin Zhang; Dongbo Liu

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Detection of Huperzine A and Huperzine B in fermentation broth of endophytic fungus Colletotrichum gloesporioides from Huperzia serrate by HPLC.

Author: Liqin HU ¹ ; Xincong KANG ¹ ; Pengyuan SHEN ¹ ; Tian CHEN ¹ ; Jiayin ZHANG ² ; Dongbo LIU ¹
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1. Horticulture and Landscape College, Hunan Agricultural University, Changsha 410128, Hunan, China.
2. State Key Laboratory of Subhealth Intervention Technology, Changsha 410128, Hunan, China.
Publication Type:Journal Article
Keywords: HPLC; Huperzia serrate; Huperzine A; Huperzine B; endophytic fungi; secondary metabolites
From: Chinese Journal of Biotechnology 2018;34(5):777-784
CountryChina
Language:Chinese
Abstract: In this study, we established a rapid and efficient HPLC method to determine the accumulation of Huperzine A and Huperzine B in the fermentation broth of endophytic fungus Colletotrichum gloesporioides from Huperzia serrate. The chloroform extracts of fermentation broth were dissolved in methanol and filtered before injection for HPLC analysis. The analysis was performed on an Agilent Eclipse plus-C18 column (250 mm×4.6 mm, 5 μm) by isocratic elution. The mobile phase was 0.015 mol/L ammonium acetate-methanol (70:30, V/V), the flow rate was 1 mL/min and the detection wavelength was set at 308 nm. Huperzine A and Huperzine B could be well separated within 25 min. Good linearity of Huperzine A was found in the range of 1.50-48.00 μg/mL (r=0.999 5), and that of huperzine B was in 0.25-7.50 μg/mL (r=0.999 7). The average recoveries of Huperzine A and Huperzine B were 106.83% and 108.06%, respectively (RSD=3.34%, 3.60%). The results demonstrate that this method can detect the content of huperzine A and huperzine B in fermentation broth simply, rapidly, accurately and in good reproducibility. Under the optimized conditions, the accumulated content of huperzine A and huperzine B were measured from the sixth to the fifteenth day. Huperzine A and Huperzine B reached the highest (12.417 0 μg/mL and 4.660 3 μg/mL, respectively) at the fourteenth and eighth days. The analysis methodology could contribute to the future study of huperzine A and huperzine B biosynthesis in C. gloeosporioides, consequently facilitate the development of new drug resources.