Spectral investigations of the solution properties of 5,10,15,20-tetrakis(4-N-benzyl-pyridyl)porphyrin (TBzPyP) and its interaction with human serum albumin (HSA)

Abstract

The association behavior of 5,10,15,20-tetrakis(4-N-benzyl-pyridyl)porphyrin ( TBzPyP ) was investigated in aqueous solution at 27 °C and various ionic strengths by optical absorption, fluorescence and resonance light scattering (RLS) spectroscopies. The results show that TBzPyP exists as monomer at low ionic strength and as ill-defined aggregates at high ionic strength and does not show concentration dependent aggregation over an extended concentration range between 5 × 10−7 to 1 × 10−4 M in water. The interaction of the TBzPyP with human serum albumin (HSA) in 5 mM phosphate buffer, pH = 7.00, and at 27 °C, has been also studied by optical absorption, fluorescence and RLS spectroscopies. The formation of two types of complexes (HSA: TBzPyP and (HSA)2: TBzPyP ) has been demonstrated by analysis of optical absorption spectral patterns of TBzPyP at various concentrations of HSA. Based on absorption data, a binding model has been proposed and used to analyze the binding process. The calculated binding constant for formation of HSA: TBzPyP and ( HSA )2: TBzPyP are 3.79 × 106 M−1 and 1.46 × 105 M−1, respectively. The RLS spectra of TBzPyP at various concentration of HSA do not show any aggregation of TBzPyP in the presence of HSA. Due to the fact that binding of TBzPyP quenches the HSA fluorescence, a step-by-step aggregation model, which has been developed by Borissevich, I. E. et al, was used to determine the average aggregation number of HSA, < J >, from the fluorescence quenching. The < J > value is 1.4, which further confirms the formation of the two mentioned complexes at this concentration range of HSA.