(250e) The Use of ?-Galactosidase to Produce N-Acetyllactosamine As a High Value Molecule Directly from Dairy Whey.

Large volumes of whey are generated during the production of cheese, Greek yoghurt and other products from bovine milk. Much of the nutritional value of this whey is still under-utilised, being sent to wastewater or used as animal food. In this work, we focus on the use of this key resource to generate higher value ingredients. In particular, we consider the use of β-galactosidases in transgalactosylation reactions between lactose and N-acetylglucosamine (GlcNAc) to generate N-acetyllactosamine (LacNAc). This functionalised disaccharide is a prebiotic and can be used as a building block for a range of human milk oligosaccharides, which have very high value within infant formula mixtures.

We show that the cations present in dairy whey can have a positive impact on the transgalactosylation reaction. Specifically, concentrations of 100 mM of calcium or magnesium reduce the reaction rate constant for the undesirable hydrolysis reaction and formation of the undesired isomer, Allo-LacNAc at longer timeframes, due to a loss of enzyme activity. Addition of magnesium enhances the selectivity for LacNAc over Allo-LacNAc with no significant reduction in the LacNAc yield.

Further, we show that the β-galactosidase enzyme can be immobilised onto silica particles using a cross-linked layer-by-layer encapsulation method. Immobilization significantly improved the enzymatic LacNAc yield compared to the free enzyme. The immobilized enzyme was successfully reused in eight consecutive reaction cycles with no significant reduction in this yield.

In our laboratory, we have shown that we can generate LacNAc of over 92% purity directly from dairy whey using activated carbon followed by selective crystallisation with ethanol. This LacNAc was used directly in an enzymatic reaction with whey derived sialylated casein glycomacropeptide to form 3’-sialyl-N-acetyllactosamine (3’-SLN). This functionalised trisaccharide can be used to form antiviral drugs that inhibit the binding of sialic-acid-targeting viruses, including several human or avian influenza strains.

Our process simulations compare the economics of a range of different downstream separation designs once LacNAc is produced; Anion-exchange chromatography; activated carbon; GlcNAc crystallization and activated carbon; and selective crystallization. We show that selective crystallisation is the best approach, with a payback period of between 0.6 and 4.4 years for a production scale of 5 tonnes of LacNAc per year.

In conclusion, this body of work provides an exciting opportunity to take a low value dairy process stream and transform it into products of much higher value in an economic and sustainable manner.