Abstract A glucose–gluconic acid biotransformation system was suggested for the experimental study of oxygen transfer in bioreactors. This biosystem was used for the investigation of the effect of the flow rate and biomass concentration on the volumetric oxygen transfer coefficient k L a in a 10 dm 3 internal-loop airlift bioreactor. For this purpose, the fermentation broth of the mycelial strain Aspergillus niger was employed, representing a three-phase system, where bubbles come into contact with dense rigid pellets. The results showed that the presented biotransformation system can be successfully utilised for the determination of the oxygen transfer rate in airlift bioreactors. The experiments showed a strong positive influence of the air flow rate on the rate ( r Glu), specific rate of gluconic acid production ( k Glu/X) as well as on the volumetric oxygen transfer coefficient ( k L a). This confirmed an expected limitation of production rate by the oxygen transport from the gas to the liquid phase in the whole range of air flow rates applied. Moreover, consistent curves of the production rate r Glu and k L a values vs. biomass concentration c X (amount of enzymes) were observed. These exhibited a local maximum for c X equal to 6.68 g dm −3. On the other hand, the specific production rate monotonously decreased with increasing biomass concentration. A decline of k L a values at higher c X values was attributed to a bubble coalescence promoting effect of mycelial pellets.