Continuous clarification of grape juice using a packed bed bioreactor including pectinase enzyme immobilized on glass beads

Highlights

• Covalent immobilization of pectinase on glass beads was carried out.

• The volumetric productivity of pectinase was decreased after immobilization.

• Grape juice clarification was done in a packed bed reactor using an immobilized enzyme.

• The maximum clarity of the juice was obtained with a flow rate of 0.5 mL min⁻¹.

• The physicochemical properties of the clarified juice were compared with the initial juice.

Abstract

A continuous clarification of grape juice was undertaken in a packed bed reactor (PBR) using pectinase enzymes immobilized on glass beads. The enzyme immobilization process decreased the specific activity and the volumetric productivity from 3 ± 0.2 to 2.7 ± 0.1 units mg⁻¹ and from ~0.085 to ~0.070 mg mL⁻¹ min⁻¹, respectively. Also, it increased the optimum pH from 5.0 to 5.5, but did not influence the optimum temperature (50 °C). The best conditions for clarification of the grape juice were obtained at a flow rate of 0.5 mL min⁻¹ and a treatment time of 42 min. After clarification, pH, total soluble solids, turbidity, total phenolic content, antiradical activity and viscosity of grape juice were decreased, and on the other hand, total titratable acidity and the color parameter L were increased. Generally, this study showed that the continuous operation in a PBR including glass bead immobilized enzyme can be a potential alternative for the batch system used in juice industries.

Introduction

Grapes (Vitis vinifera) are one of the most popular fruits in the world, and are usually consumed as fresh fruit, juice, wine or marmalade (Ghafoor et al., 2009). Grape juice is rich in flavonols, anthocyanins, procyanidins and phenolic acids and therefore has positive effects on human health as anti-cancer and anti-inflammatory agents (Capanoglu et al., 2013; da Silva Haas et al., 2019). However, the major problems in the industrial production of juice is its turbidity and the formation of sediment during storage (Tapre & Jain, 2014). The cause of these problems is mainly a hydrocolloid called pectin and so removing it is needed to produce a customer-friendly product.

Pectin is a plant polysaccharide composed of galacturonic acid chains and used as an emulsifier, thickener and stabilizer agent in the food and drug industries (Mosayebi et al., 2018; Pasandide et al., 2018). Despite these unique properties, the presence of this polysaccharide in many fruit juice samples has been problematic as stated earlier, and the best way to remove pectin during the clarification step remains an issue. (Sharma et al., 2014). Enzymatic clarification of different fruit juice samples has a high potential and could be a cost-effective method (Lettera et al., 2016; Salim et al., 2018). Pectinase is the most important enzyme used for fruit juice clarification. This enzyme reduces cloudiness and improves the quality of fruit juice by hydrolysis of pectic substances (Sojitra et al., 2017). However, the native form of the enzyme has limitations such as high cost, low stability and problems related to its recovery and reusability. Enzyme immobilization on a suitable carrier can overcome some of the limitations and defects (Chauhan et al., 2015; dos Santos et al., 2019; Wang et al., 2016; Yu et al., 2018) and lead to a continuous process. The enzyme immobilization has been done using different methods of which covalent immobilization, adsorption and entrapment on/in the supports are the main ones (Ko et al., 2018; Rehman, Nawaz, et al., 2014; Sheldon, 2007). For instance, Amin et al. (2017) immobilized the fungal exo-polygalacturonase on sodium alginate support by covalent (using glutaraldehyde as a cross-linker) and/or adsorption methods. Rehman, Aman, et al. (2014) applied the entrapment method for pectinase immobilization in an agar-agar matrix. Lei and Jiang (2011) used glutaraldehyde for covalent immobilization of pectinase onto macroporous polyacrylamide microspheres. The covalent immobilization method seemed to have better results with less enzyme release. The choice of a suitable support such as glass beads with high performance, easy availability, high stability and low cost are important benefits (Eskandarloo & Abbaspourrad, 2018).

Although the enzyme immobilization is a necessary condition for a continuous enzymatic process, it is impossible to do without the use of a bioreactor. A packed bed reactor (PBR), is a simple and inexpensive reactor, which includes one or more tubes filled with a catalyst, in which the reaction mixture flows continuously with controlled conditions (de Oliveira et al., 2018). Continuous reactors have the advantage of ease of operation, better control of the reaction conditions and product quality, high production performance and low operating costs (Nakkharat & Haltrich, 2007).

The aim of this study was the immobilization of a pectinase enzyme on glass beads using a covalent method and the development of a continuous process for clarifying grape juice in a PBR. Pectinase was immobilized on functionalized glass beads using glutaraldehyde and then the optimum conditions of enzyme activity were determined. In the next step, a bioreactor after obtaining optimum conditions was used to clarify grape juice.