The Effect Of Cathepsins L, B, H On Yak Meat Quality And Microstructure During Postmortem Ageing

It is well known that tenderness is one of the meat quality attributes consideredto be a determining factor in the acceptability and repurchasing intent of consumersand producers. Tenderness can be improved through postmortem storage atrefrigerated temperatures, a process called “conditioning”. The increase in tendernessduring postmortem ageing is due to the action of endogenous proteolytic enzymes.Three proteolytic systems have been implicated in the tenderization of meat:calcium-dependent proteases (calpains), the lysosomal proteases (cathepsins), andproteasomes. As yet, little information of cathepsins L, B and H was available in yakmeat. Accordingly, the research taking ten Datong yak Longissimus dorsi muscle(obtained randomly from Haiyan County, Qinghai province, China) as raw materials,the objectives of this work were to purify and characterize the endogenous cathepsinsL, B and H, observed the cathepsins L, B and H activity, meat quality andmicrostructure changes during postmortem ageing, through the correlation analysis toclarify the role of cathepsins L, B and H during postmortem ageing. This studyprovides further insights into the tenderization process of yak meat, which mayultimately be used for the advantageous manipulation of the process.Cathepsins L, B and H from yak muscle were purified to6.05,13.25,15.67foldrespectively by buffer extract, ammonium sulfate fractionation and a series ofchromatographic separations. The specific activity of purified enzyme were1.15,0.53,0.47units/mg, yield coefficient were5.51,11.51,9.82%, total activity were20.19,7.91,5.58U and the molecular mass were36,35,27kDa as determined bySDS-PAGE.The optimum temperature of cathepsins L, B, H were50℃,45℃and50℃respectively, optimum pH were5.5,6.0,6.5. Through observed the effect of inhibitors,activator and metal ions, suggested that the purified cathepsins belongs to cysteineproteinase family and the activity centers containing-SH. The optimum substrate ofcathepsins L, B and H were Z-Phe-Arg-AMC, Z-Arg-Arg-AMC, and L-Arg-AMCrespectively. The optimum pH of cathepsins L and B degradation myofibril were5.0~5.7, the optimum pH of cathepsin H was4.0.Cathepsins L, B and H activities were at a minimum at the initial postmortemand increased significantly with time postmortem (P <0.05), and cathepsin L was detected in more important amounts than cathepsins B and H. pH, glycogen changedsignificantly (P <0.05), and these changes was mainly concentrated in the0~3dduring postmortem ageing. Ageing have impact on pressuring losses, cooking losses,shear force, color and texture parameters, and the most direct impact on meat is toimprove its tenderness. Correlation analysis showed, the cathepsins L and pH havesignificant correlation (P <0.05), cathepsin B and pH have highly significantcorrelation (P <0.01), and cathepsins L, B, H and glycogen was a significant negativecorrelation (P <0.01).The MFI increased with time postmortem and relative to at-death values, anincrease (P <0.05) in MFI was detected during the first3d postmortem. Observed bytransmission electron microscope, we found that after8d ageing the muscle fibrilsstructure was completely disrupted, Z line rupture, M line disappear completely. Themuscle protein electrophoresis showed that ageing can lead to protein degradation ofthe different degree, the most obvious was Troponin. Ageing can make the yak musclefiber diameter decreases, thus improving the yak meat tenderness. Accordingcorrelation analysis with MFI, muscle fiber diameter and cathepsins L, B, H enzymeactivity, we concluded that cathepsins L, B, H and MFI have very significant positivecorrelated (P <0.01), cathepsins L, B, H and muscle fiber diameter have verynegatively significantly correlated (P <0.01), the muscle fiber diameter and MFIhave significant negative correlation (P <0.01).In conclusion, meat quality and microstructure indexes also changed duringageing, these changes lead to the most direct impact was improved the tenderness ofyak meat. Accordance with the cathepsins L, B, H activity, myofibril degradationcharacteristic and the correlations between cathepsins L, B, H activity and meatquality, microstructure index, we found that cathepsin H had little effect on ageingprocess, the role of cathepsin L is greater than B.