Whey lactose was hydrolyzed via different biocatalytic and chemical methods in order to establish the optimum procedure to generate a carbon-rich substrate for haloarchaeal production of the biopolyester poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBHV). Biocatalytic hydrolysis was carried out using the commercially available bacterial β-galactosidase enzyme formulation Maxilact LG 2000 TM and solid fungal β-galactosidase from Aspergillus niger. Different enzyme concentrations, incubation times, temperatures and pH-ranges were investigated to assess the optimum hydrolysis conditions. As major outcome of the undertaken investigation, an addition of 0.25% (v/v) Maxilact LG 2000 TM to whey permeate at pH-value 6.5 and 38°C leads to almost complete (more than 90% w/w) lactose hydrolysis already after only 5 h of stirring, and performs beneficial in terms of hydrolysis kinetics compared to the solid enzyme formulation. As an inexpensive alternative, kinetics of hydrolysis of whey lactose was investigated using different amounts of HCl or H 2 SO 4 , respectively, at 90°C. By adjusting the pH-value to 0.7 or lower and stirring at 90°C for 5 h, a degree of hydrolysis of about 90% (w/w) was achieved. The hydrolysis matter was used as carbon sources for PHBHV bioproduction by the haloarchaeal species Haloferax mediterranei. Independent of the applied hydrolysis method, PHBHV biopolyesters of similar monomeric composition, molar mass and dispersity index Di were accumulated by the strain. The final decision of the most adequate method in future will depend upon the microbial production strain and production scale.