Identification of different domains of calpain and calpastatin from chicken blood and their role in post-mortem aging of meat during holding at refrigeration temperatures

• Zymography and SDS–PAGE methods were standardized for simultaneous identification of calpains and calpastatin.
• DEAE Sephacel column proved to be more effective in separation of calpains and calpastatin.
• Biospectrometric method was employed for activity determination of calpains and calpastatin.

The aim of this study was to develop a simple, specific and rapid analytical method for accurate identification of calpain and calpastatin from chicken blood and muscle samples. The method is based on liquid–liquid extraction technique followed by casein Zymography detection. The target compounds were extracted from blood and meat samples by tris buffer, and purified and separated on anion exchange chromatography. It has been observed that buffer (pH 6.7) containing 50 mM tris-base appears to be excellent extractant as activity of analytes was maximum for all samples. The concentrations of μ-, m-calpain and calpastatin detected in the extracts of blood, breast and thigh samples were 0.28–0.55, 1.91–2.05 and 1.38–1.52 Unit/g, respectively. For robustness, the analytical method was applied to determine the activity of calpains (μ and m) in eighty postmortem muscle samples. It has been observed that μ-calpain activity in breast and thigh muscles declined very rapidly at 48 h and 24 h, respectively while activity of m-calpain remained stable. Shear force values were also declined with the increase of post-mortem aging showing the presence of ample tenderness of breast and thigh muscles. Finally, it is concluded that the method standardized for the detection of calpain and calpastatin has the potential to be applied to identify post-mortem aging of chicken meat samples.