Mathematical Modelling of Galactooligosaccharides Synthesis: Insights into Aspergillus oryzae Derived β-Galactosidase Kinetics

Öz

The objective of this study was to adapt experimental data with a mathematical model based on transgalactosylation mechanisms identified in the literature for the synthesis of galactooligosaccharides and lactose hydrolysis. Galactooligosaccharides, synthesized from lactose through β-galactosidase-catalyzed transgalactosylation reactions, are acknowledged as specific bifidogenic factors among prebiotic carbohydrates. Modelling galactooligosaccharide synthesis is challenging due to simultaneous hydrolysis and transferase reactions with identical nucleophiles and substrates. Therefore, researchers tried to simplify the mechanism by ignoring intermediate reactions or inhibitory effects of glucose and galactose. This study explored various kinetic models, considering the complexity of the reaction mechanism and balancing accuracy with feasibility. Experimental data from galactooligosaccharide synthesis using a continuous stirred tank reactor was adapted to kinetic models (Models A and B) with modifications, incorporating glucose and galactose inhibitions. The COPASI software was used for testing fitness of reaction models. Eight models were evaluated for their suitability, utilizing six replicated experimental datasets under specific reaction conditions, including a temperature of 45°C, lactose concentration of 18.25°Brix, and enzyme concentration of 10-unit g lactose solution-1. The analysis was conducted using the "random research" and "particle swarm" algorithms. Model evaluations revealed that Model B, augmented with glucose and galactose inhibitions, provided the closest alignment between experimental and simulated data. The study underscored the significance of glucose and galactose impacts on galactooligosaccharide synthesis efficiency. The Model B, with addition of glucose and galactose inhibitions, emerged as a valuable tool for predicting galactooligosaccharide yield, contributing to a deeper understanding of A.oryzae-derived β-galactosidase kinetics due to its significant influence on parameters such as productivity, conversion of lactose, and composition of product. The findings will guide optimization strategies for enhanced galactooligosaccharide production efficiency, advancing knowledge in enzymatic galactooligosaccharide synthesis.

Anahtar Kelimeler

• Kinetics
• Transgalactosylation
• Hydrolysis
• Galactooligosaccharides
• A. oryzae

Destekleyen Kurum

This research received partial support from the Scientific Research Projects Fund of Istanbul Technical University (ITUBAP, Project ID: MDK-2017-40556).

Proje Numarası

ITUBAP, Project ID: MDK-2017-40556

Etik Beyan

There is no need to obtain permission from the ethics committee for this study.

Teşekkür

The β-galactosidase from Aspergillus oryzae was generously provided by Enzyme Development Corporation (EDC), New York, USA.

Galaktooligosakkarit Sentezinin Matematiksel Modellenmesi: Aspergillus oryzae kaynaklı β-Galaktosidaz Kinetiğine İlişkin Bilgiler

Öz

Bu çalışmanın amacı, elde edilen deneysel verileri literatürde tanımlanmış laktoz hidrolizi ve galaktooligosakkaritlerin sentezi için belirlenmiş transgalaktosilasyon mekanizmalarına dayalı matematiksel bir modele uyumlu hale getirmektir. Galaktooligosakkaritler, laktozun β-galaktosidaz katalizli transgalaktosilasyon reaksiyonları aracılığıyla sentezlenen özel bifidojenik faktörler olarak bilinirler. Galaktooligosakkarit sentez mekanizmasının, aynı nükleofillere ve substratlara sahip hidroliz ve transferaz reaksiyonlarının eş zamanlı olarak gerçekleşmesi nedeniyle matematiksel modellemesi zorluklar içermektedir. Bu nedenle araştırmacılar, meydana gelen ara reaksiyonları veya glukoz ve galaktozun inhibe edici etkilerini göz ardı ederek mekanizmayı basitleştirmeye çalıştılar. Bu çalışma, reaksiyon mekanizmasının karmaşıklığını göz önünde bulundurarak doğruluk ile uygulanabilirlik arasında denge sağlamak amacıyla çeşitli kinetik modellerin uygunluğunu değerlendirmektedir. Sürekli karıştırmalı tank reaktörü kullanılarak elde edilen deneysel veriler, glukoz ve galaktoz inhibitörlerini içeren modifikasyonlarla kinetik modellere (Model A ve B) adapte edilmiştir. COPASI yazılımı, model denklemleri test etmek amacıyla kullanılmıştır. Toplam 8 adet model, 6 tekrarlı deneysel veri setinde (sıcaklık:45°C, laktoz konsantrasyonu:18.25 Briks ve enzim konsantrasyonu:10 ünite g laktoz çözeltisi-1) uygunluğu test edilmiştir. Çözümleme sırasıyla “random research” ve “particle swarm” algoritmaları kullanılarak gerçekleştirilmiştir. Model değerlendirmeleri, glukoz ve galaktoz inhibitörlerini içeren Model B'nin deneysel ve simüle veriler arasında en yakın uyumu sağladığını göstermiştir. Çalışma, glukoz ve galaktozun galaktooligosakkarit sentezi verimliliği üzerindeki inhibitör etkilerinin önemini ortaya koymuştur. Glukoz ve galaktoz inhibitörlerini içeren Model B, galaktooligosakkarit veriminin öngörülmesi ve laktoz dönüşümü, üretkenlik, ürün bileşimi gibi parametreler üzerindeki önemli etkisi nedeniyle A. oryzae kaynaklı β-galaktosidaz kinetiğinin daha iyi anlaşılmasına katkı sağlayacak önemli bir araçtır. Çalışma sonucunda elde edilen bulgular, galaktooligosakkarit üretim verimliliğini artırmak için optimizasyon stratejilerine rehberlik edecek ve enzimatik galaktooligosakkarit sentezi konusunda literatüre katkı sağlayacaktır.

Anahtar Kelimeler

• Kinetik
• Transgalaktosilasyon
• Hidroliz
• Galaktooligosakkaritler
• A. oryzae

Destekleyen Kurum

This research received partial support from the Scientific Research Projects Fund of Istanbul Technical University (ITUBAP, Project ID: MDK-2017-40556).

Proje Numarası

ITUBAP, Project ID: MDK-2017-40556

Etik Beyan

Bu çalışma için etik kuruldan izin alınmasına gerek yoktur.

Teşekkür

The β-galactosidase from Aspergillus oryzae was generously provided by Enzyme Development Corporation (EDC), New York, USA.

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