Fluorescence spectroscopic analysis of the interaction of papain and bromelain with l-ascorbic acid, α-tocopherol, β-carotene and astaxanthin

In this study, the interaction between four classic dietary antioxidants (including l-ascorbic acid, α-tocopherol, β-carotene and astaxanthin) and papain/bromelain was investigated by fluorescence spectroscopy. The results show that the quenching mechanisms are all static quenching at lower concentrations of antioxidants, but at higher concentrations of antioxidants, predominantly by the “sphere of action” quenching mechanisms. The binding processes of the four antioxidants to papain/bromelain are all synergistically driven by enthalpy and entropy, and the major driving forces are electrostatic effect and hydrophobic interactions. The binding constants of papain/bromelain with the four antioxidants are in the following order as: astaxanthin-papain > β-carotene-papain > astaxanthin-bromelain > l-ascorbic acid-papain > l-ascorbic acid-bromelain > β-carotene-bromelain >α-tocopherol-papain >α-tocopherol-bromelain. Synchronous fluorescence spectroscopy shows the interaction between l-ascorbic acid/β-carotene/astaxanthin and papain/bromelain decreases the hydrophobicity of the microenvironment of tryptophan (Trp) and tyrosine (Tyr) residues. The hydrophobicity of Trp is increased while the hydrophility of Tyr is increased in the presence of α-tocopherol.