Paraoxonase 1 (PON1), a high-density lipoprotein-associated enzyme known to protect against cellular damage from toxic agents, may also have antioxidant properties. PON1 activity levels have been reported to differ by sex in human and animal studies with females exhibiting higher basal levels. We measured PON1 activity frozen serum for 1,406 individuals in over 40 extended pedigrees from the San Antonio Family Heart Study (SAFHS). We used a maximum likelihood-based, variance decomposition approach implemented in SOLAR to test for genotype-by-sex (G × S) interaction on variation in PON1 activity and to determine if any of the four PON1 quantitative trait loci (QTL) previously reported by us for this population might account for sex differences in PON1 activity levels. The residual additive genetic correlation (ρG = 0.82) between males and females is significantly different from 1 (P = 0.009), suggesting that some of the genes that influence PON1 activity act differently in females and males or, possibly, that a different combination of genes influences this trait in each sex. In addition to the QTL at or near the PON structural locus on 7q21-22, three other potential QTLs were evaluated for sex-specific effects: one each on chromosomes 12, 17 and 19. The QTL on chromosome 17 (LOD = 2.32, P = 0.0003; flanked by microsatellite marker loci D17S974 and D17S969) shows a significant (P = 0.005) sex-specific effect on PON1 activity; accounting for 6% of the additive genetic variance in males and 20% in females. This study represents the first formal statistical genetic test for G × S interactions on normal quantitative variation in PON1 activity in humans. Genet. Epidemiol. © 2006 Wiley-Liss, Inc.