Immobilization of β-galactosidase on chitosan-coated magnetic nanoparticles and its application for synthesis of lactulose-based galactooligosaccharides

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Process Biochemistry

journal homepage: www.elsevier.com/locate/procbio

Immobilization of β-galactosidase on chitosan-coated magnetic

nanoparticles and its application for synthesis of lactulose-based

galactooligosaccharides

Vuong D. Nguyena,b

, Gabriella Styevkóa

, Erzsébet Madarasa

, Gökce Haktanirlara

,

Anh T.M. Trana,b

, Erika Bujnaa

, Mai S. Damb

, Quang D. Nguyena,⁎

a Research Centre for Bioengineering and Process Engineering, Faculty of Food Science, Szent István University, H-1118 Budapest, Ménesi út 45, Hungary

b Institute of Food Technology and Biotechnology, Industrial University of Ho Chi Minh City, No. 12 Nguyen Van Bao, Ward 4, Go Vap District, Ho Chi Minh City, Viet Nam

ARTICLE INFO

Keywords:

Galactooligosaccharide

Lactulose

Immobilized β-galactosidase

Pectinex Ultra SP-L

Magnetic nanoparticles

ABSTRACT

β-Galactosidase was covalently immobilized on the chitosan-coated magnetic nanoparticles activated with

glutaraldehyde. The effect of three factors including protein content, pH and time on the yield of immobilization

was investigated, and the optimum values were 0.45 mg/ml, 4.9 and 4.9 h, respectively. The maximum yield of

immobilization reached 98.8% in term of enzyme activity. The stability of β-galactosidase significantly was

improved after immobilization from 2.5 days to over 6 days of half-life at 60 °C and was active in wider ranges of

temperature (45–60 °C), and pH (4.0–5.5) compared to soluble form. The activity of immobilized enzyme re￾mained 85% of origin after eight recycles (at 60 °C, pH 4.5 for 4 h). Different initial lactulose concentrations

(0.58–2.34 M) were applied for synthesis of galactooligosaccharides. The maximum yield of galactooligo￾saccharides was 17% mol/mol at the initial lactulose concentration of 2.34 M at 36 h of reaction. It showed that

both hydrolytic and trans-galactosylation activities of β-galactosidase remained well after immobilization. The

kinetic model of immobilized enzyme was determined. These results revealed that immobilized enzyme on

chitosan-coated magnetic nanoparticles has application potentials in the production of prebiotic lactulose-based

galactooligosaccharides.

1. Introduction

The concept to improve human health through modulation of the

microbiome should be an excellent strategy with high impacts and it is

part of a comprehensive, holistic approach to lifestyle wellness [1]. In

this concept, prebiotics play an essential role because of their benefitial

effects on health of host. Recently, two main prebiotic groups are in￾tensively studied and commonly used in industry, fructans (fructooli￾gosaccharides and inulins) and galactooligosaccharides that are pro￾duced (exception to inulin that are extracted from plant sources) either

from saccharose or from lactose by glycosyltransferase activity of β￾fructofuranosidase and β-galactosidase, respectively. Chemically, both

groups of oligosaccharides contain β-glycosidic bounds and one-one

glucose molecule at the end of chain that causes the limitation of their

use in food application. To overcome this problem, lactulose and lac￾tulose-based oligosaccharides (LuGOS) may provide very good

alternative resolution. Additionally, good prebiotic activity and index

of these sugars are reported by several authors [2–4]. While lactulose is

industrially produced by chemical isomerization of lactose [5], whereas

LuGOS can be synthetized enzymatically from lactulose [6–9]. Fur￾thermore, LuGOS were reported to have higher prebiotic index as well

as better anti-inflammatory profile than lactulose [10,11]. Theoreti￾cally, LuGOS can be classified into two types fructosyl or galactosyl

oligosaccharides depended on transferase activity of enzyme used. β￾Galactosidase (β-D-galactoside-galactohydrolase, EC 3.2.1.23) enzyme

was reported to be able to catalyze galactosyl transferation to synthetise

LuGOS from lactulose [12], thus, the application of β-galactosidase,

either free or immobilized forms, in the food industry has attracted the

attention of researchers. The Pectinex Ultra SP-L (from Aspergillus

aculeatus) is a commercial preparation including several enzymes such

as pectinase, glucosidases, invertase, β-galactosidase etc. [12–15], thus

it should be ideal source of biocatalyst for production of certain

https://doi.org/10.1016/j.procbio.2019.05.021

Received 17 August 2017; Received in revised form 21 April 2019; Accepted 19 May 2019

Abbreviations: GOS, galactooligosaccharides; Lu, lactulose; LuGOS, lactulose-based galactooligosaccharides; LaGOS, lactose-based galactooligosaccharide; CTS,

chitosan; ILC, initial lactulose concentration; MAG, magnetic Fe3O4 particles; PC, protein content; Ԏ, time of immobilization; YI, the yield of enzyme immobilization ⁎ Corresponding author.

E-mail address: [email protected] (Q.D. Nguyen).

Process Biochemistry 84 (2019) 30–38

Available online 21 May 2019

1359-5113/ © 2019 Elsevier Ltd. All rights reserved.

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