银杏内生真菌Aspergillus oryzae YX-5抗肿瘤活性物质的分离与鉴定

Isolation and identification of antitumor substance from endophytic fungus Aspergillus oryzae YX-5 isolated from Ginkgo biloba

摘要:

【背景】植物内生真菌是天然活性物质的重要来源。【目的】对一株具有抗肿瘤活性的银杏内生真菌米曲霉Aspergillus oryzae YX-5进行活性物质的分离与鉴定。【方法】将该菌株发酵培养后，发酵液经乙酸乙酯萃取，采用减压柱层析、葡聚糖凝胶柱层析和高效液相色谱分析，从其代谢产物中分离活性化合物，在分离过程中以MTT法跟踪检测分离到的各组分及纯化合物的抗肿瘤活性。【结果】从菌株YX-5的发酵产物中分离纯化得到4个化合物。经核磁共振和高分辨质谱分析，将其分别鉴定为羟基曲霉酸(1)、环(4-羟脯氨酸-苯丙氨酸) (2)、环(亮氨酸-苯丙氨酸) (3)和环(丙氨酸-苯丙氨酸) (4)。其中羟基曲霉酸对人宫颈癌HeLa细胞具有显著的细胞毒活性，其IC50为1.07 μg/mL。【结论】报道羟基曲霉酸在抗肿瘤方面的活性，表明米曲霉及羟基曲霉酸在抗肿瘤天然产物开发中具有一定的应用潜力。

Abstract:

[Background] Plant endophytic fungi are an important source of natural bioactive substances. [Objective] The active substance of an antitumor endophytic fungus Aspergillus oryzae YX-5 isolated from Ginkgo biloba was investigated. [Methods] First, strain YX-5 was fermented. Then, the culture broth was extracted with ethyl acetate. By using vacuum liquid chromatography, Sephadex gel chromatography and high performance liquid chromatography, the active compound was isolated from the crude extract and the antitumor activity was measured by MTT assay. [Results] Four pure compounds were isolated from the crude extract of strain YX-5. Based on the analysis of NMR and HR-ESI-MS, these compounds were identified as hydroxyaspergillic acid (1), cyclo(4-Hydroxypro-Phe) (2), Cyclo(Leu-Phe) (3) and Cyclo(Ala-Phe) (4). Hydroxyaspergillic acid exhibited significant cytotoxic activity against HeLa cell with IC50 of 1.07 μg/mL. [Conclusion] Hydroxyaspergillic acid exhibited antitumor activity, indicating that Aspergillus oryzae and hydroxyaspergillic acid might have application potential in antitumor natural products development.