Screening and immobilization of bacterial strains for the preparation of Forsythia suspensa from Forsythia suspensa glycosides in human gut microbiota
Forsythia suspensa is mainly found in traditional Chinese medicines such as Forsythia suspensa, Osmanthus fragrans, Chrysanthemum morifolium, and pomegranate seeds. In recent years, it has been found that forsythialipin has a variety of pharmacological activities, such as anti-inflammatory activity, inhibiting the proliferation of human lung cancer cell lines, inhibiting the proliferation and migration of pancreatic cancer cells, and regulating the systolic and diastolic blood pressure of spontaneously hypertensive rats. Forsythia suspensa is a glycoside derived from the deglycosylation of Forsythia suspensa (see Figure 1). Quan, Mao et al. found that Forsythia suspensa has better anti-inflammatory and anticancer effects than Forsythia suspensa. In addition, Ye et al. found that Forsythia suspensa has a higher bioavailability than Forsythia suspensa. At present, there are two main methods for preparing Forsythia suspensa: direct extraction from plants and conversion through Forsythia suspensa glycosides. Wang et al. prepared Forsythia suspensa extract by fermenting Forsythia suspensa leaves. Mei et al. screened a strain of mold from soil that can transform forsythin to produce forsythin, and also prepared forsythin by using cellulase or acid hydrolysis of forsythin. However, the above methods all have some drawbacks, such as the addition of enzymes easily causing emulsification, making it difficult to separate and prepare the products; Acid and mold can cause instability of Forsythia suspensa and produce by-products. Based on the above reasons, there is currently no simple, feasible, complete, and reliable conversion method that needs further improvement.

There are reports that co incubation of Forsythia suspensa glycoside with human gut bacteria can metabolize and produce Forsythia suspensa. Moreover, Forsythia suspensa can be further metabolized into small molecule substances. Therefore, this study aims to screen bacteria from human gut microbiota that can efficiently convert Forsythia suspensa glycosides into Forsythia suspensa lipids, in order to compensate for the shortcomings of existing methods and lay the foundation for the further development and utilization of Forsythia suspensa lipids.

Firstly, in the screening study, due to the antibacterial effect of Forsythia suspensa glycoside, we added a certain amount of Forsythia suspensa glycoside to both the liquid culture medium used for initial screening and the plate culture medium used for secondary screening. This can inhibit the growth of strains that are not friendly to Forsythia suspensa glycoside, narrowing down the scope of screening to a certain extent. Secondly, previous studies have shown that Forsythia suspensa glycosides, when converted into Forsythia suspensa lipids by intestinal bacteria, continue to be metabolized and produce by-products. The single strain screened in this study can maintain stability of the product within 20 hours after converting Forsythia suspensa glycoside to Forsythia suspensa glycoside, without further metabolism or the generation of by-products. This method overcomes the disadvantage of continued metabolism of Forsythia suspensa in existing microbial transformation methods, and does not introduce excess functional groups. Again, Escherichia coli is a Gram negative facultative anaerobic bacterium that has been genetically engineered as an engineered bacterium due to its excellent growth activity and ability to effectively utilize inexpensive carbon sources. The facultative anaerobic characteristics are of great significance for fermentation engineering, and the Escherichia coli screened in this study possesses both of these characteristics. Finally, we also used the method of immobilizing microorganisms to immobilize strain SXUXJ-1, and optimized the production process to ultimately produce a highly practical bacterial ball. Under optimal conversion conditions, Forsythia suspensa glycoside can be completely transformed in about 25 hours. The immobilized bacterial ball transformation method is a simple, efficient, and environmentally friendly method, and the transformed product Forsythia suspensa is easy to extract and separate, therefore it has a very good application prospect.