The microstructure and stability of collagen hydrogel cross-linked by glutaraldehyde

Collagen hydrogels, which are comprised of fibrils and possess three-dimensional network structure, were prepared via self-assembly of collagen molecules and sequentially incubated in glutaraldehyde (GTA) solutions with different [CHO]/[NH2] ratios (0-9). The cross-linking degree of outer and inner parts of hydrogels was similar, demonstrating the homogeneous reaction. Based on the results of atomic force microscopy, differential scanning calorimetry and dynamic rheological measurements, it was conjectured that the stability of hydrogels were closely associated with the structural changes of collagen fibrils. When the [CHO]/[NH2] ratios ≤3, cross-linking preferentially occurred between adjacent fibrils; therefore, the fibrils presented in pairs and then densely agglomerated. As a result, the thermal denaturation temperature (from 47.1 to 73.6 °C) and elastic modulus (from 108.32 to 1618.55 Pa) increased drastically, accompanied by a distinct decrease in enzymatic degradation degree (from 93.69 to 26.91%). The effective binding ratio (EB) of aldehyde groups reduced from 72.66 to 43.92%. Moreover, hydrogels turned into yellow and yellowness (Δb*) increased from 0 to 1.68. When the [CHO]/[NH2] ratio reached 3, the arrangement of fibrils became very compact; therefore, although the GTA dosage was multiplicative ([CHO]/[NH2] = 9), there was only a relatively small improvement in thermal stability and anti-enzymolysis. Furthermore, the EB value was only 16.40%, indicating that intensive self-polymerization of GTA molecules caused a large consumption of aldehyde groups, accompanied by a remarkable yellow-stain (Δb* = 2.73). These data on the stability of cross-linked hydrogels could be helpful for the design and fabrication of materials based on collagen hydrogels.