Optimization of fermentation process and comparison of content determination methods of mannatide
10.13200/j.cnki.cjb.004523
- VernacularTitle:甘露聚糖肽发酵工艺的优化及其含量测定方法的比较
- Author:
ZHANG Xu
- Publication Type:Journal Article
- Keywords:
Mannatide;
Enterococcus faecalis;
Fermentation process;
Gel permeation chromatography(GPC);
Ethanol precipitation
- From:
Chinese Journal of Biologicals
2025;38(07):821-826+832
- CountryChina
- Language:Chinese
-
Abstract:
Objective To optimize the fermentation process of mannatide strain and screen a more accurate method for determination of mannatide content in order to lay a foundation for its large-scale production.Methods Molecular identification of α-hemolytic streptococcus strain 33# was performed by 16S rDNA sequencing and phylogenetic tree. Using the content of mannatide as the indicator, the activation times(1-4 times), inoculation time(2, 5, 7, 10 h), and fermentation mode(standing culture, shaking once every 6 h, shaking table 70 r/min, shaking table 200 r/min) were optimized by single factor test. The culture medium composition and inoculation amount were optimized by orthogonal test. In addition, the effect of pH on mannatide content was determined by dynamically controlling pH in 10 h before fermentation. The content of mannatide in the supernatant of fermentation broth prepared by five groups of formulations in orthogonal test was determined by phenolsulfuric acid method and gel permeation chromatography(GPC) method respectively, and the results were compared.Results The mannatide strain was identified as Enterococcus faecalis by molecular identification. The optimum conditions were 3 timesofactivation,5 hofinoculation and 70r/min shakingtableculture.Theoptimum culturemedium composition was0.2%glucose,1%yeastextract,0.3%beefextract,0.6%soybeanpeptoneand0.5%sodiumchloride,andtheinoculationamount was 10%. After dynamically adjusting the pH to 6. 8-7. 4, the mannatide content increased from(0. 451 ± 0. 018) mg/mL(without pH adjustment) to(0. 642 ± 0. 007) mg/mL. Compared with phenol-sulfuric acid method, GPC method could accurately separate the target product(after ethanol precipitation) and avoid the interference of small molecular sugars.Conclusion The fermentation process of mannatide strain was optimized successfully, and GPC was determined as a more reliable method for detecting mannatide content, laying a foundation for the large-scale production of mannatide.
