Introduction: Annatto, a tropical shrub from Central and South America and parts of India, contains bixin, an apocarotenoid pigment. Bixin is conventionally used as a natural food colorant and is now gaining attention for its health-promoting nutraceutical properties, particularly in chronic diseases (Metabolic Syndrome—MetS—and cancers). This study investigates the dose-dependent anti-obesity and anti-steatotic effects of bixin in in vitro cell culture models.
Methods: The anti-adipogenic and anti-steatotic effects of bixin were examined in well-established in vitro models of obesity and non-alcoholic fatty liver disease (NAFLD/steatosis), using 3T3-L1 preadipocytes (according to a differentiation protocol) and HepG2 cells (steatosis-induced with oleic acid), respectively. Bixin was administered in the concentration range of 1ug/ml ─20ug/ml (obesity model) and 2.5ug/ml ─ 10ug/ml (NAFLD/steatosis model). The neutral lipid content was estimated using Oil Red O staining; ROS/RNS were quantified using 2',7'-dichlorofluorescein diacetate (DCFDA), nitrite assays, and malondialdehyde (MDA); and the biochemical marker of lipid peroxidation was assessed using the TBARS assay.
Results: At lower concentrations, 5 µg/ml in steatotic cells and 10 µg/ml in matured adipocytes, bixin significantly reduced lipid accumulation in both hepatocytes and adipocytes (P<0.05), demonstrating its potential as an anti-steatotic and anti-obesity agent. This beneficial effect was correlated with a reduction in oxidative stress levels (decreased MDA and ROS/RNS levels). Strikingly, bixin at higher concentrations (>10ug/ml) showed increased lipid accumulation and oxidative stress.
Conclusions: Bixin exhibits anti-obesity and anti-steatotic effects at lower doses, which correlates with its antioxidant properties. However, its bioactivity is dose-dependent, wherein at higher concentrations, it ceases to inhibit adipogenesis. This opposing response is accompanied by elevated oxidative stress levels, indicating a pro-oxidant effect at higher doses, which implicates its anti-cancer potential. The present study highlights the significance of dosage optimization of nutraceuticals and dietary ingredients with respect to their intended biological applications, such as MetS and cancer treatment.
