De. Stevenson et al., OPTIMIZATION OF ALKYL BETA-D-GALACTOPYRANOSIDE SYNTHESIS FROM LACTOSEUSING COMMERCIALLY AVAILABLE BETA-GALACTOSIDASES, Biotechnology and bioengineering, 42(5), 1993, pp. 657-666
Commercially available lactase (beta-D-galactoside galactohydrolase, E
C 3.2.1.23) enzymes produced from Kluyveromyces fragilis and Kluyverom
yces lactis were accessed as catalysts for use in the production of be
ta-galactopyranosides of various alcohols using lactose as galactosyl
donor. The yield of galactoside was enhanced by using the highest prac
tical concentrations of both lactose and alcohol acceptor. The concent
rations and thus yield, were limited by the solubility of the substrat
es. The increase in galactoside yield with increasing lactose concentr
ation appeared to be specific to the lactose substrate and not due to
water activity alterations, because addition of maltose to a fixed con
centration of lactose had no effect. During the course of the reaction
, the yield of galactoside peaked after around 70% to 80% of the lacto
se was consumed, due to hydrolysis of the product by the enzyme. A wid
e variety of compounds with primary or secondary hydroxyl groups could
act as acceptors, the essential requirement being at least some water
solubility. Addition of organic cosolvents had little effect on galac
toside yield except when it increased the water solubility of sparingl
y soluble alcohols. Some galactosides were synthesized on a gram scale
to determine practical product recoveries and improve purification me
thods for large-scale synthesis. Initial purification by hydrophobic c
hromatography (for galactosides of hydrophobic alcohols) or strong ani
on-exchange chromatography (for galactosides of hydrophilic alcohols)
separated galactosides, galactobiosides, and higher oligomers from red
ucing sugars. A facile separation of the galactoside and galactobiosid
e could then be effected by flash chromatography on silica gel. (C) 19
93 John Wiley & Sons, Inc.