The temperature at which collagen denatures from a triple helix to a random coil structure is a useful measure of the degree of crosslinking. A new multi-sample denaturation temperature tester (DTT) has been constructed for rapid determination of the collagen denaturation temperature of natural tissues and collagenous biomaterials. To validate the system, the denaturation temperatures measured for the DTT are compared with results from differential scanning calorimetry (DSC). Data are presented for bovine pericardium in three states with denaturation temperatures ranging from 68 to 85 degrees C: fresh, or crosslinked with glutaraldehyde or the epoxide reagent Denacol EX-512 poly (glycidyl ether). Denaturation temperatures measured by DTT were not significantly different from those measured by differential scanning calorimetry (DSC); however, DSC onset systematically occurred at a slightly lower temperature than that measured by DTT. This result, seen only for fresh tissue is in agreement with earlier experiments using hydrothermal isometric tension (HIT) testing. By contrast, DTT and DSC onset were identical for the exogenously crosslinked materials. Since the measured transition temperature was independent of initial load, this variable may be chosen to yield sharper force-temperature transitions with a given sample geometry. This instrument allows accurate assessment of collagen denaturation temperatures for multiple samples in a fraction of the time required by other methods.

Although the pericardium is recognized as having important contributions to ventricular function, the relationship between its functional role and structural composition remains poorly understood. Conflicting evidence from low strain rate experiments has shown that differences exist between the mechanical properties of canine, human, and bovine pericardium but with no structural explanation for these differences. This paper examines the pericardia of calves, dogs, pigs, and sheep using a structural/mechanical approach with techniques novel to the pericardial literature. High strain rate mechanical testing for stress-strain response, stress relaxation, and forced vibration response has shown the pericardium to be much more viscoelastic than previously believed under large deformations, but to be quite elastic in small vibrations. The thinner canine and porcine pericardia were found to be significantly stiffer than the thicker bovine and ovine tissues, but equivalently viscoelastic. Biochemical analysis shows these thinner tissues to have significantly higher levels of type III collagen combined with a higher degree of cross-linking. This is the first structural explanation for differences in mechanical properties between the pericardia of different species.

The need for alternative crosslinking techniques in the processing of bioprosthetic materials is widely recognized. While glutaraldehyde remains the most commonly used crosslinking agent in biomaterial applications there is increasing concern as to its biocompatibility--principally due to its association with enhanced calcification, cytotoxicity, and undesirable changes in the mechanical properties of bioprosthetic materials. Hexamethylene diisocyanate (HMDC), like glutaraldehyde, is a bifunctional molecule which covalently bonds with amino groups of lysine residues to form covalent crosslinks. Evidence within the literature indicates HMDC-treated materials are less cytotoxic than glutaraldehyde-treated materials; however, there is limited characterization of the material properties of HMDC-treated tissue. This study uses a multi-disciplined approach to characterize the mechanical, thermal, and biochemical properties of HMDC-treated bovine pericardial tissue. Further, to facilitate stabilization of the HMDC reagent, non-aqueous solvent environments were investigated. HMDC treatment produced changes in mechanical properties, denaturation temperature, and enzymatic resistance consistent with crosslinking similar to that seen in glutaraldehyde treated tissue. The significantly lower extensibility and stiffness observed under low stresses may be attributed to the effect of the 2-propanol solvent environment during crosslinking. While the overall acceptability of HMDC as a crosslinking agent for biomaterial applications remains unclear, it appears to be an interesting alternative to glutaraldehyde with many similar features.

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Characterization of the viscoelastic behavior of pencardial biomatenals for heart valve or patching applications demands testing under loading times or frequencies typical of those that occur in physiological function or in a bioprosthetic device We have used a servo-hydraulic testing system to evaluate the behavior of these materials under loading times as low as 0.05 s, frequencies up to 10 Hz and strain rates exceeding 24 000 %/min Mechanical tests included large deformation cyclic loading, stress relaxation experiments, and small deformation forced vibration This paper reviews our expenence with these tests, interpreted using results from collagen denaturation temperature testing and biochemical analysis, in three distinct studies: (1) examination of the effects of glutaraldehyde, poly (glycidyl ether) (a diepoxide compound), and cyanamide on bovine pericardium, (2) comparison of bovine and porcine pericardia, and (3) evaluation of the effects of extraction of cellular components from bovine pericardium These tools provide better means toward understanding the viscoelastic properties of these materials and the structural/functional relationships that determine those properties.