The mechanistic effect of bromelain and papain on tenderization in jumbo squid (Dosidicus gigas) muscle

Highlights

• Effects of bromelain and papain on tenderization in squid muscle were studied.

• Proteases decrease WHC and texture properties of muscle.

• Proteases hydrolyze myofibrillar and sarcoplasmic proteins.

• Proteases increase amino acids and generate small peptides and short fragments.

Abstract

Jumbo squid (Dosidicus gigas) muscle is rather hard and tough, which directly affects consumer acceptance. In this study, the tenderization effect of bromelain and papain on squid muscle during enzymolysis is examined and compared with an untreated control and water-treated sample. Squid mantle were incubated with different solutions (water, bromelain, and papain solution) for 40 min in a 30 °C water bath. Then, the mantle samples were subjected to water holding capacity (WHC) analysis, texture evaluation, biochemical determination, and histological observations. The results revealed that bromelain and papain disadvantageously decrease the water holding capacity when compared to the control and water-treated samples. Furthermore, following tenderization with bromelain or papain, muscle hardness, shear force, myofibrillar protein content, and Ca²⁺ ATPase activity were all significantly decreased. Additionally, some essential amino acids were released following tenderization. When examining the myofibrillar fragmentation index (MFI), bromelain and papain were shown to cause high levels of hydrolysis in myofibrillar and sarcoplasmic proteins. Moreover, microstructural imaging indicated that the tenderization treatments disrupted myofibrils and generated a larger number of small fragments in the muscle tissues, subsequently decreasing microstructure stability and integrity. SDS-PAGE analysis confirmed that bromelain and papain have a high proteolytic activity, with some small peptides and/or short fragments detected post-tenderization. The results presented herein demonstrated that bromelain and papain improved squid muscle tenderness and can be utilized to ensure a more desirable squid product.

Introduction

Jumbo squid (Dosidicus gigas) is one of the most important cephalopod species in the world, with 747,010 tons harvested in 2016 (FAO, 2018). In recent years, squid has captured an increasing economic interest in many countries, including China, Japan, Mexico, and Chile, mainly because of the increased land demands due to development and commercialization (Lemus-Mondaca, Zambra, Marín, Pérez-Won, & Tabilo-Munizaga, 2018). Importantly, squid muscle is a good source of nutrition and contains compounds such as protein, unsaturated fatty acids, vitamins A, B, and D, essential amino acids, and other important biomolecules (Torrinha, Gomes, Oliveira, Cruz, Mendes, Delerue-Matos, et al., 2014).

However, the squid muscle has a sticky, tough consistency and is difficult to get to soften, even following storage and heating, which negatively affects consumer acceptance. Squid muscle fibers structurally differ from those of other fish and mammals. They are present in a radial and circular orientation and tightly interposed with connective tissue fibers (Torres-Arreola, Ocaño-Higuera, Ezquerra-Brauer, López-Corona, Rodríguez-Felix, Castro-Longoria, et al., 2017). This particular tissue organization is directly associated with the observed tough muscle texture. Importantly, high content of insoluble myostromin (11%) in squid muscle makes industrial squid products (e.g. shredded squid, read-to-eat squid, dried squid) taste tough and hard, which significantly hinders the edibility of these products by the elderly and infants (Hu et al., 2014). Therefore, it is necessary to find a way to manage and increase values of the squid products.

To improve the textural muscle tissues properties, several approaches, such as exposure to hydrolytic enzymes, ultrasonic waves, electrical stimulation, chemical compounds, high hydrostatic pressure processing, and blade tenderization, have been explored (Bolumar et al., 2013, Vandenberghedescamps et al., 2018, Peña-Gonzalez et al., 2019). These studies have shown that the tenderization processes reduce the degree of sarcomere contraction and affect myofibrillar integrity/degradation and/or connective tissues cleavage to improve muscle tissue tenderness (Morton, Bhat, & Ahmed, 2019). Most of all, the use of exogenous enzymes has been shown to effectively improve muscle tenderness and enhance flavor, as well as increase the nutrient value. Commonly used exogenous proteases, including papain, bromelain, ficin, and actinidin, have been examined in beef (Barekat & Soltanizadeh, 2017), chicken muscle (Wang, Li, Xu, & Zhou, 2013), pork (Sun, Chen, Geng, Luo, Gong, & Jiang, et al., 2017), and lamb tenderization (Han, Morton, Bekhit, & Sedcole, 2009). Among the most promising, bromelain and papain (recognized as Generally Recognized as Safe by United States federal agencies) are plant-derived cysteine proteases and have broad spectrum activity, cleaving a wide variety of bonds, thus degrading myofibrillar and collagenous proteins in muscle. However, one of the major challenges of using these enzymes is countering the effect of overtenderizing, leading to the need for the processor to carefully monitor the process to avoid creating an overly soft texture (Bekhit, Hopkins, Geesink, Bekhit, & Franks, 2014).

While numerous studies investigating the effect of proteases on the textural properties of mammal muscle products have been performed, little is known regarding the actions of bromelain and papain on the squid mantle muscle during the tenderization process. The objective of this study was to explore the effects of bromelain and papain in regard to water holding capacity (WHC), texture, shear force, myofibrillar protein levels, myofibril morphology, and muscle protein degradation during tenderizing in squid mantle.