Abstract Background The rapid expansion of the functional mushroom, yeast, and algae market has increased the need for enzymatic quantification of 1,3-/1,6-β-glucans. The longstanding indirect approach, which calculates β-glucans by subtracting enzymatically measured α-glucans from total glucans obtained through acid hydrolysis, is increasingly challenged by modern supplement formulations containing resistant starches, trehalose, and sweeteners such as maltitol. Objective To evaluate the current indirect enzymatic method for β-glucan measurement and to refine the analytical workflow to support reliable β-glucan quantification across the expanding functional mushroom, yeast, and algae market and associated dietary supplement formats. Methods Key steps of the established indirect β-glucan method were systematically evaluated to identify matrix-dependent limitations related to acid solubilization, glucose stability during hydrolysis, α-glucan specificity, and formulation-derived interferences. Based on these findings, a refined analytical workflow was defined and applied for benchmarking and performance evaluation across mushrooms, yeast, algae, and multiple supplement delivery formats. Results Targeted refinements increased measured total glucan values, improved α-glucan determination in matrices containing resistant starches, trehalose, and maltitol, and reduced systematic glucose loss associated with acid hydrolysis. Comparative evaluation demonstrated matrix-dependent differences relative to the conventional method, while maintaining consistent analytical performance across complex formulations and reducing total assay time. Conclusion These advances in enzymatic β-glucan measurement address recognized limitations of the indirect approach and support more consistent β-glucan quantification across diverse natural product matrices and modern supplement formats. Highlights Advances in enzymatic β-glucan measurement were achieved through targeted refinements to total and α-glucan determination, improving measurement consistency across mushrooms, yeast, algae, and complex supplement formulations and supporting further collaborative evaluation.
Sweeney et al. (Wed,) studied this question.