Antibacterial peptides generated by Alcalase hydrolysis of goat whey
Introduction
The safety and shelf life of foods are of significant concern in the food industry. These are affected by the incidence of pathogenic and spoilage bacteria which can contaminate food through several routes. A broad spectrum of methods have been employed to prevent the growth of such bacteria in food including the use of synthetic and natural antimicrobial agents. However, the use of synthetic agents is precluded owing to the possible negative impact of such chemicals on the environment and human health. So, novel antimicrobial agents from natural sources are highly required. A number of strategies have been suggested to improve the antimicrobial activities of proteins, including chemical modification such as esterification (Sitohy, Mahgoub, Osman, El-Masry, & Al-Gaby, 2013) and enzymatic hydrolysis (using digestive, microbial, or plant proteolytic enzymes; Arruda et al., 2012, Korhonen and Pihlanto, 2006, Kotlar et al., 2013, Théolier et al., 2013).
Whey is a major by-product from cheese manufacture and contains 6–7% total solids. Whey proteins represent nearly 20% of total milk proteins (Fox, 1989). The major proteins in whey are beta-lactoglobulin (β-Lg), α-lactalbumin (α-La), immunoglobulins (Ig), and bovine serum albumin (BSA). In addition, whey contains a range of minor proteins including lactoferrin (LF), lactoperoxidase and various growth factors (Park, Juárez, Ramosc, & Haenlein, 2007).
By enzymatic hydrolysis of caprine whey proteins peptides with important biological activities, such as antimicrobial, immunomodulatory, antioxidant, and antihypertensive activities, have been released (De Gobba et al., 2014, Eriksen et al., 2008, Murata et al., 2013). Several studies have investigated the formation of antibacterial peptides by digestion of goat whey proteins using various proteolytic enzymes (reviewed in Atanasova and Ivanova, 2010, Hernández-Ledesma et al., 2011). Digestion of caprine whey proteins by human gastric and duodenal juice has produced various antibacterial components that exhibited a strong inhibition of the growth of Listeria monocytogenes and other pathogenic bacteria (Almaasa, Berner, Holm, Langsrud, & Vegarud, 2008). Hydrolysis of goat whey proteins with pepsin released peptides with antibacterial activity against some food pathogens (El-Zahar et al., 2004, Théolier et al., 2013). One particular peptide, lactoferricin C (fragment 14–42 of the LF) produced by pepsin and trypsin showed strong antimicrobial activity against various types of bacteria (Kimura et al., 2000, Recio & Visser, 2000).
Alcalase® is an enzyme extract from Bacillus licheniformis (Sukan & Andrews, 1982) containing several proteinases with different specificities. Alcalase has been used extensively to prepare soluble hydrolysates of soy protein (Fox, 1989) and fish protein (Rebeca, Peña-Vera, & Díaz-Castañeda, 1991) as well as to produce bioactive peptides (Corrêa et al., 2011, Espejo-Carpio et al., 2013, Sila et al., 2014, Zhang et al., 2013). However, to date the antibacterial activity of goat whey hydrolysates produced with Alcalase has not been investigated.
Therefore, the aim of this work was to i) evaluate the antibacterial activity of goat whey hydrolysate (GWH) generated by Alcalase and fractions thereof obtained by size-exclusion chromatography, and to ii) characterize the active fractions. This information is relevant in assessing the potential use of GWH or fractions as bio-preservatives in food.
Section snippets
Materials and chemicals
Goat milk was obtained from the Food Science Department, Faculty of Agriculture, Zagazig University, Zagazig, Egypt. Sweet whey was prepared from this milk by rennet coagulation as described by Ha, Bekhit, McConnell, Mason, and Carne (2014). The resulting sweet whey was pasteurized at 73 °C for 15 s, cooled, lyophilized, and kept frozen at −20 °C until use. Alcalase 2.4 L (FG; EC 3.4.21.6) from B. licheniformis was purchased from Sigma (St. Louis, MO, USA).
Preparation of goat whey hydrolysate (GWH)
Lyophilized goat whey (11% protein)
Production of goat whey hydrolysates and fractionation by SEC
Alcalase has been used to produce bioactive peptides from whey proteins having diverse activities (Corrêa et al., 2014, De Gobba et al., 2014; Espejo-Carpio et al., 2013). In the present study, goat whey proteins were used as substrates for hydrolysis with Alcalase. Hydrolysates obtained after 60, 120, 180 and 240 min had DH values of 12%, 19%, 25% and 28%, respectively. The high DH values are consistent with the broad specificity of Alcalase. Similar DH values have been reported from Alcalase
Conclusions
In this study goat whey was hydrolyzed by Alcalase at pH 7.8 and 55 °C for 240 min to release peptides possessing potent antimicrobial activity. The results showed that enzymatic hydrolysis enhanced the antibacterial activity of goat whey against selected pathogenic and spoilage bacteria (St. aureus, B. cereus, S. typhimurium and E. coli). The GWH exhibited bactericidal activity against S. typhimurium, E. coli and B. cereus and bacteriostatic activity against St. aureus under our experimental
Acknowledgements
The financial support from the Faculty of Agriculture, Cairo University, Egypt, and from the Danish Strategic Research Council through the NOVENIA project (file no. 09-514 067076) is gratefully acknowledged. The authors wish to thank Kirsten Sjøstrøm, University of Copenhagen, for her technical assistance with the LC-MS analysis.
References (47)
- et al.
Degradation of whey from caprine milk by human proteolytic enzymes, and the resulting antibacterial effect against Listeria monocytogenes
Small Ruminant Research
(2008) - et al.
New peptides obtained by hydrolysis of caseins from bovine milk by protease extracted from the latex Jacaratia corumbensis
LWT Food Science and Technology
(2012) - et al.
Hydrolysates of sheep cheese whey as a source of bioactive peptides with antioxidant and angiotensin-converting enzyme inhibitory activities
Peptide
(2014) - et al.
Antioxidant peptides from goat milk protein fractions hydrolyzed by two commercial proteases
International Dairy Journal
(2014) - et al.
Inhibition of Listeria monocytogenes growth in Cheddar cheese by an anionic peptide-enriched extract from whey proteins
International Dairy Journal
(2013) - et al.
Effect of milk proteins their hydrolysates on in vitro immune responses
Small Ruminant Research
(2008) - et al.
Angiotensin I-converting enzyme inhibitory activity of enzymatic hydrolysates of goat milk protein fractions
International Dairy Journal
(2013) - et al.
Antibacterial surfaces developed from bio-inspired approaches
Acta Biomaterialia
(2012) - et al.
Fractionation of whey proteins from red deer (Cervus elaphus) milk and comparison with whey proteins from cow, sheep and goat milks
Small Ruminant Research
(2014) - et al.
Bioactive components of ovine and caprine cheese whey
Small Ruminant Research
(2011)
Antimicrobial peptide of Korean native goat lactoferrin and identification of the part essential for this activity
Biochemical Biophysical Research. Communications
Bioactive peptides: production and functionality
International Dairy Journal
Improvement of functional and antimicrobial properties of brewery byproduct hydrolyzed enzymatically
LWT Food Science and Technology
Antibacterial activity of peptides and folding variants from milk proteins
International Dairy Journal
Identification of milk proteins enhancing the antimicrobial activity of lactoferrin and lactoferricin
Journal of Dairy Science
Fractionation and identification of ACE-inhibitory peptides from α-lactalbumin and b-casein produced by thermolysin-catalysed hydrolysis
International Dairy Journal
The molecular characterization and antimicrobial properties of amidated bovine β-lactoglobulin
International Dairy Journal
Physico-chemical characteristics of goat and sheep milk
Small Ruminant Research
Antibacterial and binding characteristics of bovine, ovine and caprine lactoferrins: a comparative study
International Dairy Journal
Antibacterial peptides from barbel muscle protein hydrolysates: activity against some pathogenic bacteria
LWT Food Science and Technology
Isolation and identification of antimicrobial peptides derived by peptic cleavage of whey protein isolate
Journal of Functional Foods
Potent antibacterial peptides generated by pepsin digestion of bovine lactoferrin
Journal Dairy Science
Contributions of the lipopolysaccharide outer core oligosaccharide region on the cell surface properties of Pseudomonas aeruginosa
Comparative Immunology, Microbiology & Infectious. Diseases
Cited by (96)
Peptides with biological and technofunctional properties produced by bromelain hydrolysis of proteins from different sources: A review
2023, International Journal of Biological MacromoleculesCricket protein hydrolysates pre-processed with ultrasonication and microwave improved storage stability of goat meat emulsion
2023, Innovative Food Science and Emerging TechnologiesGoat milk as a natural source of bioactive compounds and strategies to enhance the amount of these beneficial components
2023, International Dairy Journal