植物乳杆菌抑制黄曲霉活性代谢物的初步研究

植物乳杆菌（Lactobacillus plantarum）能够抑制黄曲霉生长，但起主要抑菌作用的物质尚未明确。该研究采用非靶向代谢组学技术比较分析了8株抑菌活性较好（S组）和8株抑菌活性较差（W组）的L. plantarum发酵上清液。结果显示，两组L. plantarum发酵上清液的代谢组存在显著差异（P<0.05）。通过数据库比对鉴定得到咪唑乙酸、酪氨酸等30个显著差异代谢物（P<0.05），其中有机酸、脂肪酸等酸性物质较多为22个。通过与已报道的乳酸菌产生的抗真菌物质相比较，找到十八烷酸、吲哚乙酸等结构一致或结构类似物，表明上清液中酸性物质起主要的抑菌作用，且其抑菌活性依赖于低 pH 值的酸性环境。在L. plantarum产生的主要有机酸中，乳酸、乙酸、丙酸的抑菌活性良好，其抑制黄曲霉活性从大到小依次为丙酸、乙酸、乳酸。当乙酸浓度为2.64g/L、丙酸浓度为1.76 g/L时，可完全抑制浓度为106个/mL的黄曲霉孢子生长。综合表明，植物乳杆菌代谢物中有机酸和脂肪酸为主要抑菌物质，且抑菌活性随酸性物质浓度增大而增强。

Preliminary study on active metabolites of Lactobacillus plantarum against Aspergillus flavus

Abstract: Lactic acid bacteria have a long history of safe use in food. Among them, the antifungal properties of Lactobacillus plantarum are particularly interesting. Numerous studies have pointed out that the natural compounds produced by L. plantarum can significantly inhibit the growth of fungi and Aspergillus flavus, spores, thereby degrading aflatoxin, especially for the longer shelf life of a variety of food. Moreover, the acid is attributed to the antifungal activity of L. plantarum, where organic acid is the most important acidic metabolite of L. plantarum. In addition, acetic acid is the most effective antifungal metabolite among the many organic acids produced by L. plantarum. At the same time, there is the highest content of lactic acid, acetic acid, and propionic acid in the L. plantarum fermentation supernatant organic acid. However, the inhibitory effect of L. plantarum on A.flavus is mostly focused on the antifungal substances produced by a single strain in recent years. Only a few studies were on the differential metabolites of multiple strains with different antifungal activities. Moreover, standard products are still lacking to verify the activity of inhibiting A.flavus on the quantitative amount of lactic acid, acetic acid, and propionic acid in the supernatant of L. plantarum. Therefore, the purpose of this research is to find the small molecular metabolites that inhibit A.flavus through the comparison between multiple strains, with emphasis on the practical application of L. plantarum. 16 strains of L. plantarum with different antifungal activities were selected, 8 strains of which presented strong antifungal activity (Strong group) and the rest was weak (Weak group). Ultra-high performance liquid chromatography-quadrupole flight Time mass spectrometry combined with PCA (Principal Components Analysis) and OPLS-DA (Orthogonal Partial Least Square-Discriminate Analysis) was utilized to explore the different metabolites between strains with different antifungal activities, further to determine the substances with main antifungal effects. At the same time, standard products of 1, 2, and 3 times the concentration were used to verify the inhibit activity of A.flavus, according to the quantitative determination for the content of lactic acid, acetic acid, and propionic acid in the fermentation supernatant of L. plantarum in the early stage of the laboratory. The results showed that there were significantly different metabolites in the two groups of L. plantarum fermentation supernatants (P<0.05). Database comparison demonstrated 30 significantly different metabolites, including imidazoleacetic acid, tyrosine were identified (P<0.05). Among them, acidic substances were relatively different, such as organic and fatty acids. Correspondingly, the acidic substance in the supernatant was attributed to the main antifungal effect, whereas the antifungal activity depended on the acidic environment of low pH value. Lactic acid, acetic acid, and propionic acid presented excellent antifungal activities among organic acids produced by L. plantarum. Specifically, the antifungal activity was ranked in order of propionic acid>acetic acid>lactic acid. Comprehensively, organic and fatty acids are widely expected to be the main antifungal substances, where the L. plantarum antifungal activity increased with the concentration of the acidic substances.