Monitoring glycation using the intrinsic fluorescence of biological fluorophores

Abstract

The high blood glucose levels associated with diabetes affect various cells and proteins in the body. In response to high blood glucose collagen and keratin proteins experience glycation. This work aims to establish if the intrinsic fluorescence of collagen and keratin could be used to monitor the glycation of said compounds, and thus offer an alternative method to monitoring long term glycaemic control. We have studied the evolution of the intrinsic fluorescence of both compounds in response to glucose in vitro, using steady state and time-resolved fluorescence spectroscopy techniques. Changes in the intrinsic fluorescence of both collagen and keratin were observed. For collagen, contrary to the traditional fluorescence intensity decay measurement at arbitrarily selected excitation and detection wavelengths, we conducted systematic wavelength- and time-resolved measurements to achieve time-resolved emission spectra (TRES). These showed changes in the intrinsic fluorescence kinetics, caused by both collagen aggregation and glycation. In keratin, the addition of glucose caused an increase in the fluorescence intensity at the characteristic wavelength of 460 nm, due to faster formation of new cross-links. The results also suggest that glucose may cause the formation of two new fluorescent complexes with peak fluorescence at ~525 nm and ~575 nm. In conclusion, monitoring the intrinsic fluorescence of collagen or keratin could be used as a method to monitor long term glycaemic control in patients with diabetes.