Human paraoxonase 1 (h‐PON1) hydrolyzes variety of substrates and the hydrolytic activities of enzyme can be broadly grouped into three categories; arylesterase, phosphotriesterase, and lactonase. Current models of the catalytic mechanism of h‐PON1 suggest that catalytic residues H115 and H134 mediate the lactonase and arylesterase activities of the enzyme. H‐PON1 is a strong candidate for the development of catalytic bioscavenger for organophosphate poisoning in humans. Recently, Gupta et al. (Nat. Chem. Biol. 2011. 7, 120) identified amino acid substitutions that significantly increased the activity of chimeric‐PON1 variant (4E9) against some organophosphate nerve agents. In this study we have examined the effect of these (L69G/S111T/H115W/H134R/R192K/F222S/T332S) and other substitutions (H115W/H134R and H115W/H134R/R192K) on the hydrolytic activities of recombinant h‐PON1 (rh‐PON1) variants. Our results show that the substitutions resulted in a significant increase in the organophosphatase activity of all the three variants of rh‐PON1 enzyme while had a variable effect on the lactonase/arylesterase activities. The results suggest that H residues at positions 115 and 134 are not always needed for the lactonase/arylesterase activities of h‐PON1 and force a reconsideration of the current model(s) of the catalytic mechanism of h‐PON1.