# Enzymatic synthesis of calcium lactobionate from cheese whey permeate as a value-added ingredient

## TL;DR

This study developed an efficient enzymatic process to produce calcium lactobionate from cheese whey permeate, achieving nearly 99% yield and suggesting potential for industrial application.

## Contribution

A high-yield enzymatic synthesis method for calcium lactobionate using cheese whey permeate at high substrate concentration is proposed.

## Key findings

- The enzymatic oxidation process achieved a molar conversion yield of nearly 99% in 7 hours.
- Critical parameters like pH, temperature, and dissolved oxygen were effectively controlled for scalable production.
- The produced calcium lactobionate showed potential antioxidant and metal chelating activities.

## Abstract

Summary: In this study, calcium lactobionate (Ca-LBN) was manufactured through an enzymatic oxidation process using cheese whey permeate as a substrate and Ca(OH)2 as a neutralizing base. The process was operated based on the pH-stat method using a benchtop bioreactor. The operational performance of the enzymatic conversion process was analyzed by estimating the real-time molar conversion rate (MCRRT) and molar conversion yield (MCY). The enzymatic oxidation process lasted for nearly 7 hours and achieved an MCY of nearly 99%. The critical operational parameters, such as dissolved oxygen (DO%), temperature, and pH, were monitored and controlled during the manufacturing process. The manufactured Ca-LBN showed possible 2,2-diphenyl-1-picrylhydrazyl-radical scavenging and ferrous ion chelating activities; future work is needed to establish feasible and reliable methods for quantifying their bioactivities. This work suggests that employed processing parameters were effective in achieving a high LBN yield and may be adopted by the manufacturing industry for scalable production of LBN.

Summary: In this study, calcium lactobionate (Ca-LBN) was manufactured through an enzymatic oxidation process using cheese whey permeate as a substrate and Ca(OH)2 as a neutralizing base. The process was operated based on the pH-stat method using a benchtop bioreactor. The operational performance of the enzymatic conversion process was analyzed by estimating the real-time molar conversion rate (MCRRT) and molar conversion yield (MCY). The enzymatic oxidation process lasted for nearly 7 hours and achieved an MCY of nearly 99%. The critical operational parameters, such as dissolved oxygen (DO%), temperature, and pH, were monitored and controlled during the manufacturing process. The manufactured Ca-LBN showed possible 2,2-diphenyl-1-picrylhydrazyl-radical scavenging and ferrous ion chelating activities; future work is needed to establish feasible and reliable methods for quantifying their bioactivities. This work suggests that employed processing parameters were effective in achieving a high LBN yield and may be adopted by the manufacturing industry for scalable production of LBN.

•Enzymatic synthesis of Ca-LBN at high substrate concentration was studied.•The enzymatic oxidation process lasted 7 hours and achieved an MCY of ~99%.•Process parameters (e.g., temperature, pH, and DO%) were monitored and maintained.•Ca-LBN exhibited potential 2,2-diphenyl-1-picrylhydrazyl-radical scavenging and metal chelation activities.•The knowledge obtained may be used for scale-up production of Ca-LBN.

Enzymatic synthesis of Ca-LBN at high substrate concentration was studied.

The enzymatic oxidation process lasted 7 hours and achieved an MCY of ~99%.

Process parameters (e.g., temperature, pH, and DO%) were monitored and maintained.

Ca-LBN exhibited potential 2,2-diphenyl-1-picrylhydrazyl-radical scavenging and metal chelation activities.

The knowledge obtained may be used for scale-up production of Ca-LBN.

Lactobionic acid (LBA) and its salt form (e.g., lactobionate [LBN]) have emerged as high-value-added functional ingredients in food and pharmaceutical applications, such as acidulant, antioxidant, metal chelator, and carrier in drug delivery systems. Enzymatic oxidation has been employed as a nontoxic, cost-effective, and environmentally friendly approach for the synthesis of LBA/LBN. The current study investigated, first, the feasibility of producing calcium lactobionate (Ca-LBN) via enzymatic oxidation using cheese whey permeate as a substrate at high concentration, and second, the bioactivity of the resulting Ca-LBN. The production experiment was performed using reconstituted cheese whey permeate solution (300 g·L−1 lactose) as a substrate, Ca(OH)2 as a titrant base, oxidase (dosage: 400 U·kg−1 lactose), and catalase (dosage: 168,000 U·kg−1 lactose) in a laboratory bioreactor. Target critical control parameters, such as pH 6.40; dissolved oxygen: 44%; and temperature: 38°C, were defined and monitored using an industrial human-machine interface (HMI) to ensure operational stability. The consumption of Ca(OH)2 was used to calculate real-time molar conversion rate (MCRRT) and accumulative molar conversion yield (MCY) according to the pH-stat method. Enzymatic oxidation reaction continued for 7 h, and MCY was observed at nearly 99%. The MCRRT rapidly reached a plateau value of ∼470 mmol·h−1 within 20 min of the process. The critical operational parameters remained controlled by the HMI cascade, suggesting that the process is scalable. The DPPH-radical scavenging and ferrous ion chelating activity of the obtained LBN could not be confirmed based on the colorimetric assays used in the present work; however, characterization processes need to be further optimized. The obtained knowledge may be applied to the scalable production of LBA/LBN, enabling higher yields and an efficient manufacturing process.