Background Controversy exists regarding the role of the subfractions of high-density lipoproteins (HDL 2 and HDL 3 ) in cardiovascular disease. The functionality of these particles, and their protective role, is due in part to the paraoxonase 1 (PON1) presence in them. The polymorphisms rs662 (Q192R, A/G), rs854560 (L55 M, T/A), and rs705379 (C-108T) of the PON1 gene have been related to enzyme activity and, with the anti-oxidative capacity of the HDL. The objective was to determine the arylesterase PON1 activity in HDL 3 and HDL 2 and its relationship with the polymorphisms mentioned, in a young population. Methods The polymorphisms were determined through mini-sequencing (SnaPshot). The HDL subpopulations were separated via ionic precipitation, cholesterol was measured with enzymatic methods, and PON1 activity was measured through spectrophotometry. Results The results show that the PON1 polymorphisms do not influence the cholesterol in the HDL. A variation between 40.02 and 43.9 mg/dL was in all the polymorphisms without significant differences. Additionally, PON1 activity in the HDL 3 subfractions was greater (62.83 ± 20 kU/L) than with HDL 2 (35.8 ± 20.8 kU/L) in the whole population and in all the polymorphisms (p < 0.001), and it was independent of the polymorphism and differential arylesterase activity in the Q192R polymorphism (QQ > QR > RR). Thus, 115.90 ± 30.7, 88.78 ± 21.3, 65.29 ± 10.2, respectively, for total HDL, with identical behavior for HDL 3 and HDL 2 . Conclusions PON1 polymorphisms do not influence the HDL -c , and the PON activity is greater in the HDL 3 than in the HDL 2 , independent of the polymorphism, but it is necessary to delve into the functionality of these findings in different populations.