Investigation of Non‐ <i>Saccharomyces</i> Yeasts for Developing Unique Flavor Profiles in Nonalcoholic Mulberry Fermented Beverage

This study presents an innovative exploration of non-<i>Saccharomyces</i> yeast strains isolated from fragrant flowers for use in mulberry juice fermentation, aiming to develop nonalcohol fermented beverages with distinctive and complex aroma profiles. Seventy yeast isolates were screened for sugar utilization efficiency and ethanol production. Thirty-six strains that produced more than 20-fold less ethanol than <i>Saccharomyces cerevisiae</i> were selected for volatile compound analysis using headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS). Among these, nine isolates-identified as <i>Hanseniaspora thailandica</i>, <i>Pichia kudriavzevii</i>, and <i>Pichia pijperi</i>-produced a wide range of esters and higher alcohols associated with desirable wine aromas. Notably, <i>H. thailandica</i> strain S64-2 demonstrated the highest aromatic potential, producing elevated levels of isoamyl acetate, phenethyl alcohol, linalool, ethyl acetate, and methyl salicylate-compounds associated with floral and fruity notes. Although <i>H. thailandica</i> S64-2 exhibited ethanol productivity comparable to <i>S. cerevisiae</i> under high-sugar conditions, ethanol concentration was successfully limited to below 0.5% v/v (nonalcoholic beverage standard) through fourfold dilution of the juice and omission of exogenous sugar. A key novelty of this work is to report the fermentation potential of <i>H. thailandica</i> S64-2 in mulberry juice and to provide detailed insights into its volatile profile. Furthermore, the study introduces a new strategy for leveraging indigenous floral yeasts with aroma-enhancing capabilities to develop nonalcoholic fermented beverages with enriched sensory quality. These findings align with emerging consumer demands and offer a sustainable alternative to traditional wine fermentation practices.