Development and optimisation of a synbiotic low-lactose cow milk beverage via enzymatic hydrolysis using commercial β-galactosidase preparations

Background and Scope: Lactose intolerance (LI) affects approximately 65–70% of the global adult population, yet comparative data on the performance of commercially available β-galactosidase (β-gal) preparations under standardised conditions remain limited. This study developed an innovative functional low-lactose cow milk beverage by evaluating four commercial β-gal preparations Lacta-Free (Aspergillus oryzae), Maxi-Lact-2000 (Kluyveromyces lactis), HA-Lactase-2100 (K. lactis) and NOLA-Fit-2800 (Kluyveromyces fragilis) at concentrations of 0.1, 0.15 and 0.2% (w/v), systematically optimising enzyme concentration, incubation temperature (10–70°C), hydrolysis time (1–6 h), pH (4.0–7.5), starter culture ratios and fermentation conditions using one-way ANOVA and Duncan's multiple range test (P ≤ 0.05). Key Findings and Conclusions: Lactose was reduced from 4.71 ± 0.13% to 0.16 ± 0.01% (96.50% hydrolysis). NOLA-Fit-2800 achieved the highest hydrolysis rate (94 ± 3%) in the shortest time (2.5 ± 0.1 h) at 0.2%, pH 6.0, and 45°C, confirming it as the most efficient preparation for neutral milk processing. Lacta-Free (fungal) performed optimally at pH 5.5, while Maxi-Lact and HA-Lactase (yeast-derived) were most effective at pH 6.5. The optimal starter culture ratio of L. lactis:L. cremoris:L. diacetylactis at 2:2:1, fermented at 30 ± 2°C for 8 ± 0.1 h with a 5% (v/v) inoculum and Bifidobacterium bifidum BGN4 supplementation, yielded the highest viable cell count [(1.73 ± 0.05) × 10⁹ cfu/g] and conferred synbiotic functionality to the final product. Significance and Novelty: This study provides the first systematic side-by-side comparison of four commercial β-gal preparations under identical standardised conditions in the same milk substrate, delivering a validated, industrially applicable process framework to produce functional synbiotic low-lactose dairy beverages. The findings offer actionable guidance for matching enzyme source to dairy application pH and for designing LAB starter systems that maximise probiotic viability in low-lactose matrices.