Degradation potential of biological tissues fixed with various fixatives: an in vitro study

The purpose of this study was to investigate the in vitro degradation potential of porcine pericardia fixed with various aldehyde or epoxy compound (EC) fixatives, using bacterial collagenase and pronase. The fixatives investigated were formaldehyde (FA), glutaraldehyde (GA), monofunctional EC (EX-131), and multifunctional ECs (EX-810, EX-313, and EX-512). Fresh porcine pericardium was used as a control. The test samples were well immersed in a 20-U/mL collagenase solution or a 10-U/mL pronase solution and incubated at 37 degrees C at pH 7.5 for 24 h. The extent of degradation of each test sample was determined by measuring its increment in free amino group content and changes in collagen structure, denaturation temperature, and tensile stress after degradation. In general, the extent of tissue degradation with pronase was more notable than with collagenase. As observed with fresh tissue, the EX-131 EC fixed tissue radically disintegrated after either collagenase or pronase degradation, whereas the other test samples remained intact. The reason for this may reside in the more random molecular packing of the EX-131 EC-fixed tissue, which led to some loss in its helical integrity. This made penetration of enzymes into biological tissue easier. Of the multifunctional EC test groups, tissues fixed with tetrafunctional EC (EX-521) or trifunctional EC (EX-313) had relatively better resistance to degradation than those fixed with bifunctional EC (EX-810). The extent of degradation for the EX-313 or EX-512 EC fixed tissues was similar to that observed for the FA- or GA-fixed tissues. The results of this study indicated that the biological tissue fixed with monofunctional EC (EX-131) cannot resist bacterial collagenase or pronase degradation. However, resistance to degradation of the multifunctional EC (EX-313 or EX-152)-fixed tissues was comparable to that of the aldehyde (FA or GA)-fixed tissues. Therefore, of various EC fixatives, the EC with a greater number of functional groups should be chosen for tissue fixation to increase its resistance to enzymatic degradation.