接第二篇：

Discussion

After optimizing the medium formula, the amount of polysaccharide of 8.09 g/L produced by C. cicadae liquid fermentation was 642.20% higher than that without optimisation (1.09 g/L). This increase in polysaccharide yield was much higher than that obtained by Yang and Xu (21.70 and 50%) (39). Additionally, the optimized result was 1.41 times that obtained by Wang using shaking flask fermentation (5.71g/L) (40), and 16 times that obtained byWang(486.16 ± 19.60 mg/L) (41). In the present study, after optimizing the components of the liquid medium, not only was the improvement in polysaccharide yield greater than those reported in other publications, but the yield was also relatively high. Therefore, this preliminary work provided an excellent approach to improving the polysaccharide yield. The FT-IR absorption peaks near 605 cm−1 (Se=O), 760 cm−1 (C–O–Se), and 1610 cm−1 (O–Se–O) indicated that the selenite group underwent substitution on the polysaccharide (40). The FT-IR spectrum of polysaccharide component PACI-1 contained an absorption peak at 762 cm−1, which corresponded to Se-O-C stretching vibrations (42, 43). These changes indicated that PACI-1 had been selenised.

The chromatographic analysis revealed PACI-1 to be a homopolysaccharide composed of fructose with an average molecular weight of 9.945 × 103 Da. As a comparison, the molecular weights of polysaccharides JCH-1 and JCH-2, purified from Isaria cicadae Miquel, were 3.09 × 104 and 5.55 × 105 Da (21), respectively, while the molecular weight of heteropolysaccharide (IPS1) from Paecilomyces cicadae was 2.40 × 106 Da (44). Chen determined that the molecular weight of the extracellular polysaccharide W-CBP50 II in fermentation broth was 9.97 × 104 Da (45). PACI-1, with its lower molecular weight of <1 × 104 Da, would be classified as a low-molecular weight polysaccharide (33). The differences in the molecular weights of these polysaccharides may be related to the different sources, extraction methods, and purification steps for polysaccharides obtained fromthe fermentation liquids of different strains. Alternatively, the lower molecular weight of PACI-1 may have been caused by the addition of inorganic selenium (6, 46).

PACI-1 was determined to be a homopolysaccharide composed of fructose, which was inconsistent with previous experimental results (47). It was previously reported that the polysaccharide of C. cicadae mycelium was a heteropolysaccharide, with glucose, mannose, and galactose as the main components. Table 2 shows that the polysaccharides extracted from C. cicadae have been mainly heteropolysaccharides, while there are few reports in which the extracted polysaccharide was a homopolysaccharide (51). Therefore, this separation and purification study provides the first report on selenium-enriched syn-fructose polysaccharide.

The antioxidant experiment conducted in this study showed that the selenium-enriched polysaccharide PACI-1 had a certain antioxidant capacity, which was higher than that of natural polysaccharides but lower than that of other seleniumenriched polysaccharides (52). It has been reported that low-molecular weight polysaccharides have high antioxidant activity. Therefore, the lower antioxidant activity of natural polysaccharide components FVRP-1 and PKP-E-1-1 compared to PACI-1 may have been due to their higher average molecular weights of 29,930 and 1.09 × 104 Da, respectively. The antioxidant activity of polysaccharides is not related to just one factor but several (45), including the degree of branching, connections, conformation, and other factors (31). Therefore, the complex relationship between the polysaccharide structure and antioxidant activity needs further exploration and research to elucidate the reasons underpinning the activity of PACI-1.

Conclusion

In this study, the components of the liquid fermentation medium of strain S1 were optimized. The response surface results identified the optimal medium ratio at which the greatest yield of extracellular polysaccharide was obtained (8.09 g/L). A selenium source was added to the fermentation medium of strain S1 to prepare the selenium polysaccharide and the extracellular polysaccharide in the fermentation broth was studied. The structure of component PACI-1 was analyzed by HPLC, FT-IR, CD, AFM, and NMR. The results showed that PACI-1 had an average molecular weight of 9.945 × 103 Da in selenium- and fructose-enriched medium, and a three-strand helical-spherical structure in aqueous solution. Additionally, the antioxidant activity of PACI-1 was between those of the natural polysaccharides and selenium polysaccharides. However, the complete structureactivity relationship of PACI-1 remains unclear. Future studies will further explore the relationship between the bioactivity and structure of extracellular polysaccharides from C. cicadae.

Data availability statement

The original contributions presented in the study are included in the article/Supplementary material, further inquiries can be directed to the corresponding author.

Author contributions

WZ: conceptualization, methodology, software, investigation, formal analysis, and writing—original draft. JX: data curation and writing—original draft. HL: visualization and investigation. YZ: resources. LC: Supervision. SSh: software. SSo: validation. HZh: visualization. TD: data analysis. HZe and QX: conceptualization, funding acquisition, resources, and supervision. All authors contributed to the article and approved the submitted version.

Funding

This work was financially supported by the Open Project Program from Key Laboratory of Se-enriched Products Development and Quality Control, Ministry of Agriculture/National-Local Joint Engineering Laboratory of Se-enriched Food Development (Grant No. Se-2021C08), Major science and technology projects in Huaibei (Grant No. HK2021016), and Industry university research cooperation project of Anhui Kouzi Distillery Co., Ltd. andHuaibei Normal University (Grant No. KZJY-001), National University Student Innovation and Entrepreneurship Project (Grant No. 202210373018).

Conflict of interest

Authors JX, HL, YZ, LC, SS, and QX were employed by Anhui Kouzi Distillery Co., Ltd.

The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Publisher’s note

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.

Supplementary material

The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fnut.2022.1032289/full#supplementary-material

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