Objective Formulation and evaluation of Paraoxonase1 (PON1) microparticles (MPs) efficacy on chlorpyrifos (CPF) induced hepatotoxicity. Methods PON1 micro particles are prepared by using solvent evaporation method. The prepared MPs were characterised for entrapment efficacy (EE), drug content, and drug release. The surface morphology was identified using scanning electron microscopy (SEM). In-vivo studies were performed biochemical markers such as acetylcholinesterase (AchE), nitric oxide (NO), thiobarbituric acid reactive substances (TBARS), superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx), and malondialdehyde (MDA) using animal model by CPF induced hepatotoxicity method. Results Obtained results revealed that formulated micro particles had 84.37% EE and the drug content was 89.71. The scanning electron microscopy (SEM) images confirm the spherical structure and smooth texture of particles. The maximum % drug was released in 30 days. PON1-MPs supplementation significantly reduces liver weight, liver index, and alanine transaminase (ALT), aspartate transaminase (AST) , alkaline phosphatase (ALP) and AchE, NO, TBARS and ameliorates hepatic toxicity with increased anti oxidant levels. Conclusion In the present investigation principal reduction in AchE, elevated levels of anti oxidant enzymes in hepatic tissue by the PON1 and improved efficacy by PON1-MPs contributed hepato protection against CPF induced hepatotoxicity. Further studies have to be emphasizing molecular mechanisms involved in hepato protection.