Abstract: The aim of this study was to screen the endophytic fungal strains of Sophora alopecuroides that can produce Quinolizidine alkaloid, to explore the fermentation conditions and improve alkaloid production. Fifty endophytic fungal strains isolated from healthy seeds of S. alopecuroides in the previous period were used as materials. The alkaloid-producing strains were screened by Thin-Layer Chromatography, High-Performance Liquid Chromatography and Acid-dye colorimetry. The alkaloid-producing strains obtained were analyzed by morphology and multi-gene sequence comparison to determine the taxonomic status. Subsequently, the effects of the addition of precursors such as L-pipericolic acid, L-lysine and α-ketoglutaric acid on the alkaloid yields of alkaloid producing strains were explored by one-way and response surface methodology. The results showed that a total of three highly active Sophoridine-producing strains, DSD103, DSD201 and DSD308, all belonging to Alternaria alternata, were screened, and the alkaloid yields of Sophoridine in their mycelia were 0.212, 0.544 and 0.276 mg·g^-1. The highest alkaloid yielding strain, DSD201, was obtained with the addition of the precursors L-pipericolic acid at a concentration of 1.079 μmol·L^-1, L-lysine at a concentration of 6.251 μmol·L^-1, and α-ketoglutaric acid at a concentration of 0.153 μmol·L^-1. Under this conditions, the alkaloid yield in mycelium reached the maximum of 0.949 mg·g^-1, which was 74.12% higher than that under the control conditions. The screened alkaloid-producing strains provide a new method for the production of Quinolizidine alkaloid by microbial fermentation.

Key words: Endophytic fungi of Sophora alopecuroides, Alkaloid-producing strains, Screening, Optimization of fermentation conditions, Quinolizidine alkaloid

摘要： 本研究旨在筛选出能够产喹诺里西啶类生物碱的苦豆子（Sophora alopecuroides）内生真菌菌株,探究其发酵条件,提高碱产率。以前期从苦豆子健康种子中分离的50株内生真菌为材料,采用薄层色谱法、高效液相色谱法和酸性染料比色法进行产生物碱菌株的筛选,对获得的菌株进行形态学和多基因序列比对分析,确定分类地位。随后,通过单因素和响应面法探究添加L-哌啶酸、L-赖氨酸和α-酮戊二酸对产生物碱菌株碱产率的影响。结果表明,共筛选出3株高活性产槐定碱菌株DSD103,DSD201和DSD308,均属于互隔链格孢（Alternaria alternata）,其菌丝中槐定碱的产率分别为0.212,0.544和0.276 mg·g^-1。碱产率最高的菌株DSD201在L-哌啶酸浓度为1.079 μmol·L^-1、L-赖氨酸浓度为6.251 μmol·L^-1和α-酮戊二酸浓度为0.153 μmol·L^-1时,菌丝中碱产率达到最大,为0.949 mg·g^-1,比对照提高了74.12%。所筛选的产生物碱菌株为通过微生物发酵生产喹诺里西啶类生物碱提供了一种新方法。

关键词: 苦豆子内生真菌, 产生物碱菌株, 筛选, 发酵条件优化, 喹诺里西啶类生物碱