Development of a Homogenizer-Assisted Sulfuric Acid Hydrolysis Method for Quantifying Yeast Cell Wall Mannan

Accurate quantification of yeast cell wall mannan is critical for strain screening, metabolic engineering, and functional evaluation of yeast-derived biomaterials, yet existing methods often lack specificity or scalability. Here, we report a rapid and reproducible mannan quantification platform combining homogenizer-assisted cell disruption, sulfuric acid hydrolysis, and high-performance liquid chromatography with refractive index detection (HPLC–RID). High-pressure homogenization enabled efficient and uniform cell wall disruption, markedly improving analytical reproducibility compared with bead-beating pretreatment. Cell wall mannan was quantitatively converted to mannose using a sulfuric acid hydrolysis protocol, allowing selective monosaccharide-level detection with minimal interference. The method showed excellent linearity using commercial yeast mannan (0–2.5 g/L; R²=0.9994) and a strong correlation between released mannose and dry cell weight in <italic>Saccharomyces cerevisiae</italic> (R²=0.9927). The platform reliably distinguished metabolically engineered low- and high-mannan <italic>S. cerevisiae</italic> strains and was further validated in the probiotic yeast <italic>S. boulardii</italic>. Overall, this approach provides a rapid, precise, and accessible tool for high-throughput yeast mannan quantification.