Development of superoxide dismutase based visual and spectrophotometric method for rapid differentiation of fresh and frozen-thawed buffalo meat.

Study aimed to develop biomarker-based assay for rapid detection of fresh and frozen-thawed buffalo meat in the supply chain. The method is based on development of a solvent system and identification of suitable substrate and developer for screening of biomarkers. For the confirmation column chromatography, gel electrophoresis and Western Blotting were carried out. Validation was done by intra- and inter-day validation, storability study, and determination of thermal history. Best results were shown with pH 8.0 Tris-HCl; extraction buffer, 205 µM nicotinamide adenine dinucleotide hydrogen; substrate, 184 µM Nitroblue tetrazolium, and 1.9 µM phenazine methosulfate; developer. The thermal history ranged from 0.14 to 0.17 during storage at -20 °C. The intra- and inter-day assay precision (CV %) ranged from 5.3 to 6.5 %; in chilled and 14.1 - 9.2 % in frozen-thawed samples. The study confirmed SOD as a viable biomarker. Developed method using SOD has significant potential for rapidly differentiating chilled or frozen-thawed meat.

In-gel and OFFGEL-based proteomic approach for authentication of meat species from minced meat and meat products.

BACKGROUND: Fraudulent mislabelling of processed meat products on a global scale that cannot be detected using conventional techniques necessitates sensitive, robust and accurate methods of meat authentication to ensure food safety and public health. In the present study, we developed an in-gel (two-dimensional gel electrophoresis, 2DE) and OFFGEL-based proteomic method for authenticating raw and cooked water buffalo (Bubalus bubalis), sheep (Ovis aries) and goat (Caprus hircus) meat and their mixes. RESULTS: The matrix-assisted liquid desorption/ionization time-of-flight mass spectrometric analysis of proteins separated using 2DE or OFFGEL electrophoresis delineated species-specific peptide biomarkers derived from myosin light chain 1 and 2 (MLC1 and MLC2) of buffalo-sheep-goat meat mix in definite proportions at 98:1:1, 99:0.5:0.5 and 99.8:0.1:0.1 that were found stable to resist thermal processing. In-gel and OFFGEL-based proteomic approaches are efficient in authenticating meat mixes spiked at minimum 1.0% and 0.1% levels, respectively, in triple meat mix for both raw and cooked samples. CONCLUSIONS: The study demonstrated that authentication of meat from a complex mix of three closely related species requires identification of more than one species-specific peptide due to close similarity between their amino acid sequences. © 2017 Society of Chemical Industry.

OFFGEL electrophoresis and tandem mass spectrometry approach compared with DNA-based PCR method for authentication of meat species from raw and cooked ground meat mixtures containing cattle meat, water buffalo meat and sheep meat.

The present study compared the accuracy of an OFFGEL electrophoresis and tandem mass spectrometry-based proteomic approach with a DNA-based method for meat species identification from raw and cooked ground meat mixes containing cattle, water buffalo and sheep meat. The proteomic approach involved the separation of myofibrillar proteins using OFFGEL electrophoresis, SDS-PAGE and protein identification by MALDI-TOF MS. Species-specific peptides derived from myosin light chain-1 and 2 were identified for authenticating buffalo meat spiked at a minimum 0.5% level in sheep meat with high confidence. Relative quantification of buffalo meat mixed with sheep meat was done by quantitative label-free mass spectrometry using UPLC-QTOF and PLGS search engine to substantiate the confidence level of the data. In the DNA-based method, PCR amplification of mitochondrial D loop gene using species specific primers found 226bp and 126bp product amplicons for buffalo and cattle meat, respectively. The method was efficient in detecting a minimum of 0.5% and 1.0% when buffalo meat was spiked with cattle meat in raw and cooked meat mixes.