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Received: 22 December 2017 Revised: 30 March 2018 Accepted: 8 April 2018 DOI: 10.1002/fsn3.669
ORIGINAL RESEARCH
The combined effects of lactoperoxidase system and whey protein coating on microbial, chemical, textural, and sensory quality of shrimp (Penaeus merguiensis) during cold storage (4 ± 1°C) Maryam Farshidi
| Mohammad Yousefi
Faculty of Nutrition and Food Sciences, Department of Food Science and Technology, Tabriz University of Medical Sciences, Tabriz, Iran Correspondence Ali Ehsani, Faculty of Nutrition and Food Sciences, Department of Food Science and Technology, Tabriz University of Medical Sciences, Tabriz, Iran. Email:
[email protected] Funding information Tabriz University of Medical Sciences, Grant/Award Number: 5/97/580 and 09/05/2015
| Ali Ehsani
Abstract Growth and reproduction of gram-negative bacteria has a pivotal role in spoilage of seafood products. In order to identify the effect of lactoperoxidase system (LPOS), an antimicrobial activity was added to whey protein solution at the levels of 0 (control group), 1.25%, 2.5%, 5%, and 7.5%. Then, the shrimp samples were coated with immersion method and stored in the refrigerator for 16 days. In this period, the microbial tests of psychrotrophic bacteria, mesophilic bacteria, Pseudomonas fluorescens, Pseudomonas spp., and Shewanella putrefaciens, chemical tests of total volatile basic nitrogen, thiobarbituric acid, and pH, and sensory evaluation were carried out on the days of 0, 4, 8, 12, and 16. Adding LPOS in the coating resulted in a decrease in the total specific spoilage organisms and TVB-N. There was no significant effect for the TBA test. The levels of LPOS showed a positive correlation with the acceptable measurement in the sensory evaluation. KEYWORDS
antimicrobial coating, lactoperoxidase, shelf life, shrimp, whey protein
1 | I NTRO D U C TI O N
natural herbal extracts, particularly essential oils and bacteriocins such as nisin and LPOS have been found as appropriate additives for
The enzymatic and microbial activities of fresh shrimp have caused
the conservation of products (Lu, Ding, Ye, & Liu, 2010). Using edible
it to be more perishable than many other foods. The effect of the
coatings has expanded the shelf life of shrimp (Nowzari, Shábanpour,
microbial activity is usually the dominant agent limiting the shelf life
& Ojagh, 2013). In this procedure, a thin layer of edible and biocom-
and safety of shrimp during refrigeration storage and transporta-
patible coating is used to protect the shrimp from the physical and
tion. During the past several decades, certain methods have been
chemical damage and delay the microbial growth and increase the
developed in order to increase the shelf life of shrimp in an attempt
safety. Moreover, the additional benefits of the method are reduc-
to prevent the consumer health risks (Al-Dagal & Bazaraa, 1999).
ing waste packaging and producing new food (Valverde, Valero,
Furthermore, traditional methods such as cook- chill processes
Martínez-Romero, Guillén, Castillo, & Serrano, 2005).
and storage were commonly used to avoid the corruption of fresh
The growing consumer worries about the human health impacts
shrimp. Different natural antimicrobial and chemical compounds, an-
of chemical preservatives have encouraged several studies to focus
tioxidants, polymers, and biotechnology products have been devel-
on the edible coatings containing natural antioxidant and antimi-
oped as well and effectively used in foodstuff industry. In addition,
crobial properties. Chitosan, Chitin, and their derivatives have been
This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. © 2018 The Authors. Food Science & Nutrition published by Wiley Periodicals, Inc. Food Sci Nutr. 2018;1–9.
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FARSHIDI et al.
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utilized for years in the processing of marine products due to their
According to Min, Harris, and Krochta (2005), LPOS did not signifi-
antibacterial and antioxidant activities (Kamil, Jeon, & Shahidi, 2002).
cantly change the tensile features, oxygen permeability, and whey
Furthermore, shellfish produced ink of sepia has been found to re-
protein films color. Accordingly, the aim of this study was to develop
tard the corruption of shrimps as a reliable edible coating (Sadok,
a solution of whey protein as a protective antimicrobial coating or
Abdelmoulah, & El Abed, 2004).
low-cost food by blending, LPOS as an antimicrobial enzyme system
Bacterial contamination is one of the reasons of spoilage of sea-
with both the bactericidal and bacteriostatic impacts to enhance an-
food products. Recent researches have proved that usually in spe-
timicrobial quality, mainly on particular corruption of bacteria and
cial refrigeration conditions, the corruption occurs mainly due to the
extend shelf life fillets of shrimp in the refrigerator.
presence of specific microorganisms that are identified and introduced as organisms involved in corruption. According to the studies of researchers, some species of gram-negative organisms, particularly Pseudomonas fluorescens and Shewanella putrefacience, have been introduced as the organisms involved in the corruption of most
2 | M ATE R I A L S A N D M E TH O DS 2.1 | Materials
fish, mollusks, and crustaceans during the storage at cold tempera-
Lactoperoxidase system was composed of glucose oxidase (GO; Sigma-
tures (ice and refrigerator) (Xu, Lin, Sui, & Cao, 2012).
Aldrich), LPO (150 U/mg, Sigma-Aldrich), D-(α)-glucose (Glu, Sigma-
The main purpose of adding the antimicrobial components for
Aldrich), H2O2 (Merck, Germany), and potassium thiocyanate (KSCN,
packaging films is enhancing the antimicrobial effect which leads
Bioserae, France). Glycerol, a plasticizer for improving coating flex-
to control the surface corruption growth agents and pathogenic
ibility, was obtained from Merck (Frankfurt, Germany). Whey protein
(Kuorwel, Cran, Sonneveld, Miltz, & Bigger, 2011; Ouattara, Simard,
(80% protein) was purchased from DMV Co. (Veghel, the Netherlands).
Piette, Bégin, & Holley, 2000). Moreover, instead of mixing antimicrobial components directly with the food, combining them into coating solutions lets the functional impact on the food surface be
2.2 | Preparation of LPOS
localized. So the use of such films in foods such as meat is very prac-
Lactoperoxidase system preparation was performed as described
tical. The advantage of this method is a slow and gradual release
by Cissé et al. (2012). The ratios of the LPOS components based on
of antimicrobial compounds into the food material (Coma, Sebti,
weight were 0.35, 1.00, 108.70, 2.17, and 1.09 for GO, LPO, Glu,
Pardon, Deschamps, & Pichavant, 2001). Lactoperoxidase (LPO) enzyme, a glycoprotein in milk, saliva,
H2O2, and KSCN, respectively. To prepare LPOS, the compounds were separately dissolved in 50 ml of phosphate buffer (pH 6.2,
and tears of mammals is introduced as one of the most import-
Sigma- Aldrich); and then, 15.5 mg lactoperoxidase enzyme was
ant enzymes used in food industries as a biological antimicrobial
added to it. To increase the antibacterial activity of LPOS, the final
agent with broad spectrum which can be utilized in food packaging.
solution was incubated at 23°C under shaking at 160 rpm for 24 hr
Investigations have shown that this enzyme has bactericidal impact
(Min, Krochta, & Rumsey, 2007).
on gram-negative bacteria along with inhibitory impact on gram- positive bacteria. In addition, antifungal and antiviral activity of this enzyme has been reported (Seifu, Buys, & Donkin, 2005; Yener,
2.3 | Shrimp fillet preparation
Korel, & Yemenicioğlu, 2009). LPOS has made of these three com-
Shrimps used for experiments were obtained from the Persian Gulf
pounds: lactoperoxidase, thiocyanate (SCN−), and hydrogen perox-
(Iran). They were gutted, beheaded, washed with tap water, and then
ide (H2O2) where the enzyme catalyzes oxidation of SCN− by H2O2
transferred immediately to the laboratory on ice.
and to produce antimicrobial compounds such as hypothiocyanite (OSCN−) and hypothiocyanous acid (HOSCN) That this substances are having the potential inhibition of microorganisms through oxidation of sulfhydryl (SH−) groups in their enzymes and proteins
2.4 | Preparation of the whey protein solution followed by treatment of shrimp fillets
systems. These substances have the potential inhibition of bacteria
Mixing 10 g Whey Protein with 100 ml distilled water and stirring in a
through oxidation of sulfhydryl (SH−) groups in their enzymes and
Controlled temperature of 90°C to result in a clear mixture prepared
protein systems (Cissé, Montet, Tapia, Loiseau, & Ducamp-Collin,
whey protein solution (WPS). Glycerol, as a plasticizer, was added
2012). The cytoplasmic microorganism membrane damage caused
to the solution (Min et al., 2005). LPOS concentrations prepared at
by the oxidation of sulfhydryl (SH−) groups has been reported as
1.25%, 2.5%, 5%, and 7.5% (v/v) were added to the WPS. In order to
the most important principle of the destruction of microbial cells
determine the optimum LPOS concentrations in WPS, a preliminary
(Mohamed, Clementine, Didier, Gérard, & Noëlle, 2013; Min, Harris,
disk diameter test was carried out with the concentrations of 0.31%,
& Krochta, 2005).
0.63%, 1.25%, 2.5%, 5%, 7.5%, and 10% LPOS (v/v) in a solution
Whey protein is the good source for biodegradable edible coating
of whey protein. The bacterial suspension was adjusted to 1 × 107
and is suggested in food industry. By adding plasticizer, clear, bland,
colony-forming Units (CFU)/ml and spread on the iron agar LYNGBY
and flexible edible films based on water will be produced which have
with the help of sterile cotton swab. After that, 6 mm in diameter fil-
excellent oxygen, oil, and aroma barrier features (Gennadios, 2002).
ter paper disks were impregnated with 20 μl of the each solution and
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FARSHIDI et al.
placed on the surface of insemination. After incubation at 4°C for
homogenized for 2 min with 9 ml of 0.25 molequi/L HCl solution con-
2 hr, the plates were placed in an incubator at 30°C for 3–4 days. The
taining 15 g/100 ml trichloroacetic acid (TCA) and 0.375 g/100 ml
antibacterial strength was investigated by measurement of the diam-
2-thiobarbituric acid (TBA). The mixture was heated in a boiling
eter of inhibitory zone. It was found that the concentration of LPOS
water bath for 10 min, then immediately cooled with running water.
