Abstract: Multispectral techniques were employed to detect salt concentration (i.e. ionic strength)-modulated interaction of acid green 9 (AG9) and acid blue 9 (AB9) with bovine serum albumin (BSA). The Debye-Hückel limiting law was applied to quantitative analysis to calculate the true Gibbs free energy change (ΔG0I→0) and effective charge (ZB) in the anion receptor pocket of BSA. The sign of ZC value for AG9/AB9 was negative and the ZB values were positive. These results indicated the formation of ion-pair between opposite charges; for example, the negatively charged AG9/AB9 bound to the net positively charged binding pocket (site-I) of BSA, suggesting that the local charge rather than the overall or surface charge of BSA played a key role in dominating the interaction strength of BSA-AG9/AB9 system. Moreover, with the increase of salt concentration, the entropy gradually varied from positive to negative and the exothermic enthalpy term increased so that the ΔG0 was almost invariant, namely enthalpy-entropy compensation. Finally, the thermodynamic parameters changed from ΔH0 < 0, ΔS0 > 0 (electrostatic force) to ΔH0 < 0, ΔS0 < 0 (non-electrostatic force) at high salt concentration. These results showed that the mobility/local motion of AG9/AB9 complexed with protein was retarded due to the transformation of dominant force.

Key words: acid green 9, acid blue 9, bovine serum albumin, Debye-Hückel theory, enthalpy-entropy compensation