Optimisation of freeze drying conditions for purified serine protease from mango (Mangifera indica Cv. Chokanan) peel

Abstract

This study investigated the possible relationship between the encapsulation variables, namely serine protease content (9–50 mg/ml, X₁), Arabic gum (0.2–10% (w/w), X₂), maltodextrin (2–5% (w/w), X₃) and calcium chloride (1.3–5.5% (w/w), X₄) on the enzymatic properties of encapsulated serine protease. The study demonstrated that Arabic gum, maltodextrin and calcium chloride, as coating agents, protected serine protease from activity loss during freeze-drying. The overall optimum region resulted in a suitable freeze drying condition with a yield of 92% for the encapsulated serine protease, were obtained using 29.5 mg/ml serine protease content, 5.1% (w/w) Arabic gum, 3.5% (w/w) maltodextrin and 3.4% (w/w) calcium chloride. It was found that the interaction effect of Arabic gum and calcium chloride improved the serine protease activity, and Arabic gum was the most effective amongst the examined coating agents. Thus, Arabic gum should be considered as potential protection in freeze drying of serine protease.

Introduction

Protease is a commercially important enzyme and it has been reported that about 60% of the total worldwide market of enzymes is proteases (Rao, Tanksale, Ghatge, & Deshpande, 1998). Proteolytic enzyme (EC 3.4) is a class of proteins found in various sources, including animals, plants and microorganisms ubiquitous nature (Aehle, 2004). Proteases of plant origin perform vital functions, from the mobilisation of storage proteins during germination to the initiation of cell death and senescence (Schaller, 2004). Plant-derived proteases have been used in food and biotechnological industries for high stability in extreme conditions, substrate specificity, good solubility and activity over a wide range of temperatures and pH (Caffini, Lopez, Natalucci, & Priolo, 1988). Mangifera indica is one of the most popular tropical fruits in the world and currently ranked 5th in total world production among the major fruit crops (Villa-Corrales, Flores-Prieto, Xamán-Villaseñor, & García-Hernández, 2010). Mango peel contains different types of enzymes, with positive and negative effects and exhibiting a significant amount of protease, peroxidase, polyphenol oxidase, pectin lyase, xylanase, and amylase activities (Ajila, Naidu, Bhat, & Prasada Rao, 2007). Although mango peel is one of the major wastes of the food and beverage industries, it can be used as a valuable, economic and available media source for the commercial production of natural enzymes (Loelillet, 1994).

Freeze drying is an important drying process for products requiring the preservation of their flavour, bioactivities, and other properties (Liapis & Sadikoglu, 1997). This process produces a fine, homogeneous powder, with excellent control of impurity levels and decreases its environmental impact. However, this method of drying generates low temperature stress, stress of dehydration and ice crystal formation, which may deactivate and destabilize enzymes (Kawai & Suzuki, 2007). Thus, many kinds of stabilizers have been used to decrease deactivation and destabilization of freeze-dried enzymes (Arakawa, Prestrelski, Kenney, & Carpenter, 2001). There is no available information about the optimisation of freeze drying conditions of serine protease from mango peel. In this study therefore, response surface methodology (RSM) was employed for modelling the possible relationship between the enzyme encapsulation variables namely serine protease content (9–50 mg/ml, X₁), Arabic gum (0.2–10% (w/w), X₂), maltodextrin (2–5% (w/w), X₃) and calcium chloride (1.3–5.5% (w/w), X₄) on enzymatic properties of encapsulated serine protease using freeze drying. The main objective of this study was to determine the optimum concentration level of encapsulation variables resulting in maximum serine protease activity, yield, maximum pH stability and storage stability in terms of other response.