A multi-sample denaturation temperature tester for collagenous biomaterials

Abstract

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.

Mechanical overload decreases the thermal stability of collagen in an in vitro tensile overload tendon model

“…Prior to testing, the pans were carefully cleaned to avoid baseline drift due to wicking and superheating effects. 17 Specimens were trimmed with a razor blade to fill the pan and ensure contact with the pan for optimal heat flux. The tissue was tested fully hydrated with Hank's solution but not immersed in a volume of liquid (gently patted dry superficially), as liquid was found to cause a highly variable baseline in pilot experiments.…”

Crosslinking of biological tissues using genipin and/or carbodiimide

“…This technique was widely used in studying the thermal transitions of collagenous tissues. 19,20 Details of the methodology used in the measurement of denaturation temperature were previously described.…”

The role of collagen in the declining mechanical properties of aging human cortical bone

“…14 This thermally induced structural transition shows itself as the so-called collagen shrinkage phenomenon in which, if collagen is heated slowly, it contracts 15,16 at a certain temperature or, if it is held at a constant length, it develops a contractile force. 17,18 The shrinkage behavior is indicative of the overall condition of the crosslinks in the collagen network and the stability they induce. 19 The technique has proved valuable in assessing the efficiency and density of artificially induced intermolecular crosslinks, produced by various chemical tanning agents, on a number of tissues.…”