Abstract:

To explore the non-covalent interaction between reduced tripeptide glutathione and L-aromatic amino acids, a stoichiometry of reduced γ-glutathione and three L-aromatic amino acids, including phenylalanine, tyrosine and tryptophan were mixed respectively, and then incubated at room temperature and physiological pH conditions for 1 h to reach the equilibrium. The electrospray ionization mass spectrometry(ESI-MS) results indicate that glutathione and three L-aromatic amino acids could form non-covalent complexes, respectively. The primary fragment ions of complexes obtained from MS² in a tandem mass spectrometer included aromatic amino acid, glutathione as well as its y₂, b₂, which confirmed the results of ESI-MS. Moreover, in UV spectroscopies, the discrepancy between complexes and reactants also confirmed the formation of complexes in aqueous solution. To avoid distinct ionization efficiency discrepancy and signal suppression in ESI-MS measurements, the interaction between glutathione and L-Tyrosine was evaluated quantitatively, which revealed that the reactant concentration should range from 5×10^-5 to 3.00×10^-4 mol/L. The dissociation constants of three complexes were determined using mass spectrometric titration method, the calculation results revealed that the stabilities of complexes formed by glutathione and three L-aromatic amino acids increased gradually according to the order of tyrosine, tryptophan and phenylalanine.

Key words: L-Aromatic amino acids, Glutathione, Non-covalent complexes, Mass spectrometric titration method, Dissociation constants