Chemical Compositions and Biological Activities of Chicken Meat Protein Hydrolysates

Abstract

The increasing consumer health awareness on food has accelerated the driving force of the functional food market. Chicken meat has long been considered as healthy meat because of high proteins and biological values. Some investigations have reported that peptides derived from chicken meat hydrolysates contained several biological activities such as antihypertensive, antioxidative, and antibacterial activities. In Thailand, different genotypes of chicken are available for consumption such as broiler, black-boned, Thai native, and spent hen chickens. Therefore, the main purpose of this study was to determine the effect of breast muscle from various chicken genotypes on nutritional components and biological activities. Physical, chemical and biological properties of the hydrolysate obtained from the most effective breed of chicken was evaluated. Furthermore, the structure of the bioactive peptides was further identified. In this study, ten chickens of broiler, black-boned, Thai native, and spent hen were slaughtered at a typical market age (2.1±0.2 kg for broiler, 1.2±0.1 kg for black-boned, 1.3±0.1 kg for Thai native, and 1.7±0.1 kg for spent hen) and breasts were separated from the carcass to determine chemical composition by proximate analysis. Protein pattern was investigated using SDS- polyacrylamine gel electrophoresis. The antioxidant capacities of chicken breasts were evaluated by DPPH, ABTS and FRAP assays. Carnosine and anserine, the antioxidant peptides in animal tissues, were also determined using high performance liquid chromatography (HPLC). It was shown that Thai indigenous chicken meat had the greatest protein composition and the highest antioxidant capacity (DPPH and FRAP assays) compared with the other genotypes (p<0.05). Protein pattern in breast meat of each genotype was not differ, however, Thai indigenous chicken presented the highest levels of myosin and actin (p<0.05). In addition, the highest amounts of total carnosine and anserine were found in Thai indigenous chickens breast meat (p<0.05). Therefore, it might be suggested that breast meat from Thai indigenous chicken is a source of functional health benefit for consumer. To produce antioxidant hydrolysate from breast meat of Thai indigenous chicken, commercial peptidases including alcalase and papain were employed in hydrolysis process and further quantified their antioxidant activities by standard in vitro antioxidant assays. The effects of enzyme concentration of each enzyme on degree of hydrolysis and antioxidant activities were investigated. It was found that incubation with 5% (v/w) alcalase for 3 hours and 0.125% (w/w) papain for 3 hours were the optimum conditions of each enzyme that produce the highest antioxidant activities. The digestion of breast meat by papain provided stronger antioxidant activities but lower degree of hydrolysis and peptide content than the digestion by alcalase. To enhance the production of antioxidant hydrolysate of breast meat, the combination of papain and alcalase was processed. We found that the hydrolysate obtained from the hydrolysis of alcalase before papain on breast meat showed higher antioxidant activities and peptide contents than the hydrolysate of papain prior to alcalase treatment. Next, the preparation of antioxidant hydrolysate powder of chicken breast meat was further investigated. The hydrolysate was microencapsulated with maltodextrin and dried with different two methods including spray drying and freeze drying. The result showed that the process by spray drying or freeze drying did not affect chemical and antioxidant properties of hydrolysate but spray drying gave a better value in physicochemical characteristics such as solubility and moisture. It might be summarized that the optimum condition for production of antioxidant hydrolysate powder of chicken breast meat was as followed; 1) digesting by combined enzymatic hydrolysis of 5% (v/w) alcalase for 3 hours and then 0.125% (w/w) papain for 3 hours, 2) using spray drying and microencapsulation with maltodextrin. Next, the antioxidant hydrolysate of Thai indigenous chicken breast meat was identified its primary protein structure. It was digested by exopeptidases including carboxypeptidase A and leucine aminopeptidase. Then, the resultant peptides were purified via size-exclusion chromatography (SEC). Primary structure of indigestible peptide was confirmed via liquid chromatography-electrospray ionization-tandem mass spectrometer (LC-MS/MS). It was observed that 21 peptides with molecular weight ranged from 75 to 1348 daltons were remained after exopeptidase digestion. The highest amount of detected peptides of chicken breast meat hydrolysate was anserine, beta-alanyl-3-methyl-L-histidine. The hydrolysate also contained 2 tripeptides, 17 dipeptides and 1 pyroglutamyl peptide. Furthermore, most indigestible peptides were composed of either proline or branch chain amino acid residue in their structure. It might be suggested that anserine was a major antioxidant peptide in hydrolysates derived from Thai indigenous chicken meat. Chronic inflammation is associated with various degenerative diseases including aging and cancer. We further investigated anti-inflammatory activities of some identified peptides in Thai indigenous chicken meat using RAW264.7 murine macrophages culture. Proline containing peptides including Gly-Pro, Pro-Pro and Leu-Pro as well al pyroglutamyl leucine (pEL) were evaluated. These synthesized peptides at 200 micromolar were not toxic to murine macrophage cells measured by MTT assay. Moreover, they did not produce nitric oxide which can cause inflammation to macrophage cells. Leu-Pro, Pro-Pro and pEL significantly inhibited the production of nitric oxide and suppressed the secretion of tumor necrosis factor-alpha (TNF-α) in lipopolysaccharide induced murine macrophage. Furthermore, Leu-Pro significantly inhibited the releasing of interleukin-6 (IL-6) in lipopolysaccharide induced murine macrophage. It might be suggested proline containing dipeptides and pyroglutamyl peptide obtained from Thai indigenous chicken meat exhibited anti-inflammatory activities at least partly due to suppression of pro-inflammatory cytokine production. In conclusion, breast meat of Thai indigenous chicken provided a great protein content and high antioxidant capacity compared with the other genotypes. The optimal production of antioxidant hydrolysate powder was prepared by the treatment with combined enzymes of alcalase and papain together with spray-drying method. Anserine was a major antioxidant peptide found in breast protein hydrolysate. Moreover, peptides containing either proline or leucine and pyroglutamyl group obtained from Thai indigenous chicken breast meat exhibited the anti-inflammatory activities in lipopolysaccharide-stimulated murine macrophage cells by suppression of pro-inflammatory cytokines secretion including TNF-α and IL-6. Thai indigenous chicken might be a potential source of bioactive peptides for antioxidant and anti-inflammatory activities in food supplement for health promotion.