|Abstract:||Carotenoids are commonly consumed in the diet and are reported to have a wide range of bioactivities that can confer positive health benefits. The non-provitamin A carotenoid, lycopene, has shown to be one of the most abundant carotenoids in serum and is correlated with a wide range of health benefits. Lycopene has been thought to exhibit its bioactivity from its antioxidant structure or its metabolites. However, it remains unclear what enzymes cleave lycopene in vivo. The current literature shows that two enzymes, ß-carotene oxygenase 1 (BCO1) and ß-carotene oxygenase 2 (BCO2), cleave carotenoids in vivo. Several animal trials show that lycopene can be cleaved by BCO2, but metabolites have been detected in mice lacking BCO2. Additionally, in vitro studies in E. Coli have demonstrated that BCO1 preferentially cleaves lycopene. To confirm the primary cleaving enzyme on lycopene, we used four groups of mice; wild type (WT), Bco1 knock out (-/-), Bco2 knock out (-/-), and Bco1 -/- x Bco2 -/- double knock out (DKO) mice. All mice were orally gavaged with 1 mg of lycopene resuspended in cottonseed oil for 14 consecutive days. A WT reference group was orally gavaged with cottonseed oil for 14 consecutive days. The primary goal of this study was to elucidate what enzyme preferentially cleaves lycopene in vivo. This work analyzed tissue weights, hepatic lipid accumulation, gender differences in lycopene accumulation, and lycopene accumulation in the tissues. Our results indicate that tissue weights, when adjusted for body weights, were impacted by genotype. For example, DKO mice exhibited smaller hearts than WT and Bco1 -/- mice and larger gonadal adipose depots compared to WT and Bco2-/- mice. The lycopene accumulation observed in our study suggests that BCO2 primarily cleaved lycopene in the liver. Despite this, there were lower lycopene concentrations in serum and extrahepatic tissues in WT and Bco2 -/- mice compared to DKO and Bco1 -/- mice suggesting modified transportation of lycopene in certain genotypes or enhanced BCO1 enzymatic activity within the extrahepatic tissues. Hepatic lipid levels were higher in WT and Bco2 -/- mice than Bco1-/- mice, and there was a positive correlation (P> 0.051) between hepatic lipids and hepatic lycopene accumulation across groups. This correlation could partially explain the levels of lycopene observed in this study. Lycopene may be sequestered in lipid droplets, limiting the amount of lycopene available for further enzymatic activity within the liver. Lastly, female mice accumulated more lycopene in their tissues when compared to male mice. These results indicate that genetic knock-out of BCO1 and BCO2 impact tissue weights, tissue lycopene accumulation, and hepatic lipids.