Enzymatic Synthesis of Galacto-oligosaccharides in an Organic–Aqueous Biphasic System by a Novel β-Galactosidase from a Metagenomic Library

Abstract

Prebiotic galacto-oligosaccharides (GOS) were effectively synthesized from lactose in organic-aqueous biphasic media by a novel metagenome-derived β-galactosidase BgaP412. A maximum GOS yield of 46.6% (w/w) was achieved with 75.4% lactose conversion rate in the cyclohexane/buffer system [95:5 (v/v) cyclohexane/buffer] under the optimum reaction conditions (initial lactose concentration = 30% (w/v), T = 50 °C, pH 7.0, and t = 8 h). The corresponding productivity of GOS was approximately 17.5 g L(-1) h(-1). The GOS mixture consisted of tri-, tetra-, and pentasaccharides. Trisaccharides were the chief component of reaction products. These experimental results showed that a low water content, a high initial lactose concentration, and an elevated reaction temperature could significantly promote the transgalactosylation activity of β-galactosidase BgaP412; at the same time, the enhanced GOS yield in an organic-aqueous biphasic system is because of the fact that thermodynamic equilibrium can be shifted to the synthetic direction by reversing the normal hydrolysis.

Transgalactosylation of lactose for synthesis of galacto-oligosaccharides using Kluyveromyces marxianus NCIM 3551

“…In contrast to this, several authors have reported maximum GOS at high (30-40%) lactose concentration [7,23,24,32]. This was attributed to high availability of both the donor and the acceptor molecules for transgalactosylation, suppression of water activity thereby reducing hydrolysis [24]. In the present study, the b-galactosidase produced by the selected yeast performed well at lower lactose concentration producing significant amount of GOS as compared to other studies.…”

“…These results were supported by the b-galactosidase of B. longum where maximum GOS yield was observed at 6.8 [7]. Similar observations of high GOS yield at pH 7.0 were observed for a b-galactosidase derived from a metagenomic library [24]. In general, b-galactosidase from yeast or bacterial origin have more neutral pH optima between 6.0 and 7.5, while b-galactosidases from filamentous fungus, such as A. oryzae, show maximum GOS formation at low pH [24].…”

“…The results were in conformity with that of b-galactosidase BgaP412 isolated and screened from a metagenomic library, expressed in Pichia pastoris indicating that an increase in temperature leads to higher solubility of lactose, and a reduction in viscosity, which in turn, increased the rate of reaction. Transgalactosylation was thus favored over hydrolytic activity [24] as also seen with Bifidobacterium infantis RW-8120 [5] for GOS synthesis. Other report revealed the best GOS production at 508C using purified b-galactosidase of Penicillium simplicissimum, whereas, at high temperature of 558C inactivation of the enzyme occurred leading to lowered production of GOS [33].…”

“…A 41% (w/w) yield with initial lactose concentration of 20.5% (w/v) was reported with recombinant b-galactosidase from Lactobacillus plantarum [31]. In contrast to this, several authors have reported maximum GOS at high (30-40%) lactose concentration [7,23,24,32]. This was attributed to high availability of both the donor and the acceptor molecules for transgalactosylation, suppression of water activity thereby reducing hydrolysis [24].…”

“…[24]. Maximum GOS concentration of 7.6 g/L (16% w/w of total sugar) has been reported with commercial b-galactosidase (Biolactase) from Bacillus circulans after 1.5 h with lactose conversion of 50% (w/v).…”