Inhibition of protease activity. Part 1. The effect on tenderness and indicators of proteolysis in ovine muscle

To examine the effect of particular enzyme groups on tenderness specific cysteine protease inhibitors were injected into muscle early post-mortem. The protease enzyme inhibitor E-64 was injected into the m. longissimus thoracis et lumborum (LTL) on the right side of 12 lamb carcasses within 15 min of death and in another 12 carcasses with the protease inhibitor Z-Phe-Ala-CHN(2). The left LTL (control) was injected with saline (0.25 M NaCl). To create variation in the rate of pH decline alternate carcasses were electrically stimulated (low voltage). The LTL was divided into cranial and caudal portions and aged for 1 or 2 days. Muscle samples at 1 day post-mortem were used for measurement of osmolality and sarcomere length (n=48), and others at 1 and 2 days post-mortem for shear force determination (n=96). The myofibrillar fragmentation index (MFI) was determined on samples taken at pH 6.2 and 1 and 2 days post-mortem (n=144). Other muscle samples were obtained at death, pH 6.2 and 6.0 and then at 1 and 2 days post-mortem (n=215). These samples were used for determination of protein solubility and the concentration of free amino acids. Stimulation caused a faster (P<0.05) decline in pH. There was no effect of stimulation (P>0.05) on shear force values, but injection of inhibitor and ageing both had effects (P<0.001). The inhibitor E-64 prevented any improvement in tenderness with ageing, whereas the inhibitor Z-Phe-Ala-CHN(2) and the control samples showed a similar ageing response. In the latter two treatments there was an average reduction of 1 kg in shear between 1 and 2 days post-mortem, whilst the inhibitor E-64 maintained shear force on average 2 kg higher than control samples. Injection and ageing had an effect on MFI (P<0.001) and there was an interaction (P<0.05) between stimulation and ageing for MFI, such that as stimulated muscle aged the rate of change of MFI was greater. There was an interaction between injection and ageing (P<0.05) for protein solubility such that samples treated with E-64 showed a minimal increase in protein solubility with ageing, whereas in samples treated with Z-Phe-Ala-CHN(2) and the control samples there was a significant increase. There was also an interaction between stimulation and ageing such that between sampling at pH 6.0 and 2 days post-mortem, stimulated muscle exhibited greater solubility (P<0.05). There were no effects (P>0.05) on the concentration of free amino acids. The evidence indicated that the cysteine proteases were responsible for post-mortem proteolysis and tenderisation, in particular the calpains, whereas the cathepsins (B and L) were unlikely to contribute to proteolysis and subsequent tenderisation in meat.