# Enhancing the Content of Hesperidin and Nobiletin in Citrus Wines through Multi-Strain Fermentation
## 4 Conclusions  
This study demonstrated that multi-strain combination fermentation methods can effectively enhance the content of hesperidin and nobiletin in citrus wine, with the (Hg.Ht)-10-Sc combination emerging as the optimal multi-strain combination for increasing the content of these flavonoids. Compared to commercial yeast monoculture fermentation, the content of nobiletin was increased by 16% and hesperidin by 5%. Additionally, the alcohol level in Nanfeng tangering wine created using a combination of multiple strains for fermentation was notably reduced compared to that made solely with Sc fermentation, rendering it better suited for today's popular consumer market. The fermentation process involving multiple strains showcased a diverse microbiome, active metabolic processes, fast sugar utilization, and rapid fermentation speed, despite a slightly reduced efficiency in converting sugar to ethanol. However, this method has the potential to enhance the taste and beneficial compounds in the wine. The aroma richness and complexity were better in Sc:(Ce.Hg.Ht)-1:1 and (Hg.Ht)-10-Sc. During fermentation, Sc, Hg, and Ht are known to produce \(\beta\)-glucosidase, with non-_Saccharomyces cerevisiae_ strains typically exhibiting higher enzymatic activity compared to _Saccharomyces cerevisiae_. Additionally, Ce displays tannase activity, facilitating the degradation of cell walls and promoting the release of compounds contained within. The utilization of a 1:1 and sequential multi-strain combination fermentation method may have the potential to enhance the co-fermentation process of multiple strains with minimal mutual influence compared to alternative fermentation approaches. This approach may effectively leverage the strengths of individual strains and ultimately enhance the flavor profile of Nanfeng tangering wine to a considerable extent. The 1:1 and 1:100 multi-strain combination fermentation methods facilitated malolactic fermentation, significantly reducing the content of malic acid and increasing the content of lactic and succinic acids, enhancing the smoothness of Nanfeng tangering wine and its health-promoting properties. Overall, the simulated complex microbial system in this experiment requires further exploration, as it did not achieve the complexity and richness of aroma similar to natural fermentation. This may be due to inappropriate inoculation ratios or the selection of strains not suited for multi-strain combination fermentation. The co-fermentation of different strains produces complex metabolites, but the interactions between strains may also hinder their development and adversely affect wine fermentation. Therefore, selecting appropriate strains and fermentation methods is crucial. Further investigation into the metabolic and molecular mechanisms underlying the promotion of bioactive substances by different strain combinations during fruit wine fermentation could be achieved by monitoring the biotransformation process. The following are available online at [[https://www.mdpi.com/article/10.3390/fermentation10050238/s1](https://www.mdpi.com/article/10.3390/fermentation10050238/s1)]([https://www.mdpi.com/article/10.3390/fermentation10050238/s1](https://www.mdpi.com/article/10.3390/fermentation10050238/s1)), Figure S1: The concentration of volatile aroma compounds identified in tangering wines. Table S1. Chemical composition of tangering juice and tangering wines (means \(\pm\) SD). Table S2: The standard curve, validation range, and coefficient of determination (r2) for the non-volatile aroma compounds in tangering wines. S.Z. : conceptualization, methodology, validation, investigation, and writing--original draft preparation. Y.O. : investigation, methodology, validation, and writing review and editing. L.X. : data curation and software. J.L. : conceptualization, writing--review and editing, visualization, project administration, and funding acquisition. Y.W. : investigation and methodology. B.G. : investigation. D.Z.