Inhibition of tyrosinase activity by Maillard reaction products derived from cysteine and glucose (Cys-MRPs) was studied. Pre-incubation of mushroom tyrosinase with Cys-MRPs decreased enzyme activity with increasing reaction time. We show that Cys-MRPs irreversibly block the active site of mushroom tyrosinase and that the competitive inhibitors dithiothreitol and kojic acid protect the enzyme from Cys-MRPs inactivation. Correlation of tyrosinase inhibition ability, volatile compounds, non-volatile compounds (HMF, DDMP and maltol), and Maillard reaction conditions of Cys-MRPs was analyzed by partial least squares regression (PLSR). 3-Ethyl-2-formylthiophene, α-dimethylformylthiophene, 2,6-dimethylpyrazine, ethylpyrazine, 2-ethyl-6-methylpyrazine, 2-methyl-3-(2-thienyldithio) thiophene, and furfural showed a significant and positive contribution to inhibition ability, while 2-propionylfuran and α-dimethyl-2-formylfuran showed a significant but negative correlation with inhibition ability. Of the three non-volatile compounds analyzed, only 2,3-dihydro-3,5-dihydroxy-6-methyl-4(H)-pyran-4-one (DDMP) showed a significant and positive correlation with inhibition ability, while HMF and maltol showed a weak negative correlation. The reaction temperature and time showed a significant and positive correlation with inhibition rate, whereas the ratio of sugar to amino acid showed a negative effect within the experimental range.