In order to reveal a functional group required to form an analytically valuable complex such as chrome Azurol S (CAS)-human serum albumin (HSA) complex, the absorption and fluorescence spectra of ten dyes related to CAS were examined in the presence of HSA. A red shift similar to that caused by HSA in the case of CAS was observed in the absorption spectrum only for the dyes containing carboxyl group. HSA also caused increase of fluorescence of these dyes, whereas the dyes free from carboxyl group exhibited weak or no fluorescence even in the presence of HSA. In particular, the increase of fluorescence intensity of Eriochrome Gyanine R (ECR) was found to be larger than that of CAS, and ECR may be superior to CAS for the determination of HSA. The increase of fluorescence may be attributed to ionization of carboxyl group and/or wear of planarity of the dye by binding to HSA. The results indicate that carboxyl group is essential to form an analytically valuable dye-HSA complex in contrast with the case of Bromcresol Green-HSA complex.
