Galacto-oligosaccharides (GOS) are formed from lactose in discontinuous mode of conversion using beta-galactosidase from Lactobacillus sp. (beta-gal). The discontinuous process was optimized for technical application with regard to GOS yield, enzyme preparation, reaction temperature and substrate source. It proved to be advantageous to directly apply the crude cell-free enzyme extract for the conversion, since similar GOS yields and composition were obtained as when using the pure enzyme preparation, but expensive purification could be avoided. Reaction temperature was lowered to 17 degrees C to limit microbial contamination when using technical substrates. Thereby GOS yield decreased from 30% to 28% of total sugars and enzyme demand increased 2.7-fold. Whey permeate was compared to buffered lactose solution as a substrate source. The initial reaction rate was found to be 1.8 times higher for the whey permeate substrate; however, GOS yield was slightly lower (approximately 25% of total sugar at 17 degrees C) mainly due to smaller amounts of allolactose[beta-D-Galp-(1-->6)-D-Glc] and the trisaccharide beta-D-Galp-(1-->6)-D-Lac formed.

We developed a novel analytical method for concentration determination of tandem single‐chain antibody diphtheria toxin (immunotoxin). The method is based on polymethacrylate monoliths with Protein L ligands as the binding moiety. Different buffers were tested for elution of the Protein L‐bound immunotoxin and 4.5 M guanidinium hydrochloride performed best. We optimized the elution conditions and the method sequence resulting in a fast and robust method with a runtime <10 min. Fast determination of immunotoxin is critical if any process decisions rely on this data. We determined method performance and a lower limit of detection of 27 μg/mL and a lower limit of quantification of 90 μg/mL was achieved. The validity of the method in terms of residual analysis, precision, and repeatability was proven in a range from 100 to 375 μg/mL. The short runtime and ease of use of a high‐performance liquid chromatography method is especially useful for a process analytical tool approach. Bioprocesses related to immunotoxin where fermentation or other process parameters can be adjusted in accordance to the immunotoxin levels will be benefited from this method to achieve the highest possible purity and productivity.