Preformulation studies and in vitro cytotoxicity of naringin.

Objective: This study evaluates the chemical and enzymatic stability of naringin (NRG), identifies its degradation metabolites, assesses its in vitro cytotoxicity, and validates a high-performance liquid chromatography (HPLC) method for precise quantification.

Significance: NRG, a flavonoid with antioxidant, anti-inflammatory, and anticancer properties, faces clinical limitations due to poor solubility, rapid degradation, and low bioavailability. While research efforts on this promising compound have largely focused on overcoming these limitations through formulation strategies, it is equally necessary and complementary to focus on preformulation studies to enhance NRG's therapeutic potential. These studies represent a fundamental step in drug development, providing key insights into the physicochemical and biological properties of NRG and serving as the basis for the rational design of safe and effective formulations in future research.

Methods: NRG stability was analyzed under various temperature and pH conditions. Cytotoxicity was evaluated in 3T3 cells, and an HPLC method was developed and validated to quantify NRG and its primary metabolite, naringenin (NRGN).

Results: NRG remained stable up to 100 °C and under physiological pH (1.2, 5.8, and 7.4) but degraded at extreme pH, forming NRGN. Cytotoxicity assays showed low toxicity at ≤1 mM (viability >80%), whereas 5 mM significantly reduced viability. The validated HPLC method exhibited high precision, specificity, and accuracy in distinguishing NRG from NRGN.

Discussion: This study provides critical insights into NRG's stability, safety, and quantification, supporting its potential therapeutic development. These findings establish a foundation for future research aimed at enhancing NRG bioavailability and clinical applicability.