New insights into wine polysaccharides quantification: evaluation of acidic methanolysis challenges and different two-step sequential precipitation strategies

Abstract

Among the macromolecules present in wine, polysaccharides represent one of the most complex and abundant classes. Commonly classified as grape- or yeast-derived polymers, they play a crucial role in wine quality and colloidal stability1,2. Despite their importance, polysaccharide analysis remains challenging and requires a prior isolation step, most often achieved by ethanolic precipitation3. While several methodological improvements have been proposed to increase precipitation repeatability, only a limited number of studies have investigated how sample preparation and precipitation conditions influence the recovery of the different polysaccharide families4,5. As a result, a strong need for analytical approaches that combine efficient extraction with reliable qualitative and quantitative measurements still exist3. Recently, the influence of sample preparation and precipitation steps on wine polysaccharides recovery and detection by HRSEC-RI and GC–MS, following acidic methanolysis and derivatization, was re-evaluated by our group through the application of design of experiments (DoE) techniques6. This work highlighted the possibility of selectively recovering yeast-derived polysaccharides or both yeast- and most grape-derived polymers depending on precipitation conditions. However, the results also suggested that acidic methanolysis efficiency may depend on the nature of the precipitate obtained and that a single precipitation step may be insufficient to recover the full diversity of wine polysaccharides6. In the present study, the challenges and limitations of acidic methanolysis are investigated in detail, with particular attention to the water content of solvents and to the dehydration of polysaccharide extracts. Several strategies aimed at minimizing water content were evaluated, and a modified acidic methanolysis protocol is proposed to improve polysaccharide conversion into TMS methyl glycosyl derivatives while limiting their secondary hydrolysis into free monosaccharides. Moreover, a two-step sequential precipitation scheme was systematically applied under different conditions across various wines to identify conditions leading to maximal polysaccharides quantification and to examine how polysaccharides are distributed between the two precipitation steps. Overall, this work provides a refined methodological framework for wine polysaccharides quantification and offers new perspectives for studying their role in wine colloidal systems.

References

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