Abstract
We have developed a method to detect UV-dependent formation of carbonylated proteins in human hair using fluorescein-5-thiosemicarbazide (5-FTSC) and fluorescent microscopy. In this study, hair keratin films consisting of α-keratin and keratin-associated proteins (KAPs) were prepared and utilized as a substitute device for a hair sample. When the fluorescence intensity was observed in the hair keratin films after irradiation with UV rays, the formation of carbonylated proteins in the hair keratin film prepared by the pre-cast method was higher than that of keratin films prepared by other methods. The fluorescent intensity was increased in proportion to the irradiation time with a high coefficient of correlation (R2=0.97) and this became evident within 10min of irradiating the film. The sensitivity of 5-FTSC in the keratin film was approximately 6-fold higher compared with that of hair samples. The keratin film was sensitive to both UVA and UVB exposures. Immunoblot analysis also showed that the increase of oxidative proteins from the keratin films was observed in the film irradiated over 120min. The sensitivity, however, was low compared with the fluorescent microscopic observation. Fourier-transform infrared measurement showed that the formation of cysteic acid was detected in the film irradiated for 240min, though that of carbonyl compounds was not.
