Quality by design approach of apocynin loaded clove oil based nanostructured lipid carrier as a prophylactic regimen in hemorrhagic cystitis in vitro and in vivo comprehensive study

Abstract

Apocynin (APO) is a naturally occurring acetophenone with eminent anti-inflammatory and anti-oxidant peculiarities. It suffers from poor bioavailability due to low aqueous solubility. Herein, APO was loaded in a Clove oil (CO) based Nanostructured lipid carrier (NSLC) system using a simple method (ultrasonic emulsification) guided by a quality-by-design approach (23 full factorial design) to optimize the formulated NSLCs. The prepared NSLCs were evaluated regarding particle size (PS), polydispersity index (PDI), zeta potential (ZP), and entrapment efficiency (EE%). The optimal formula (F2) was extensively investigated through transmission electron microscope (TEM), Fourier transform infrared (FT-IR) spectroscopy, Differential scanning calorimetry (DSC), X-ray diffractometry (XRD), in vitro release, and stability studies. Cytotoxicity against human urinary bladder carcinoma (T24) cell line and in vivo activity studies in rats with induced cystitis were also assessed. The results disclosed that the optimal formula (F2) had PS of 214.8 ± 5.8 nm with EE% of 79.3 ± 0.9%. F2 also exhibited a strong cytotoxic effect toward the T24 cancer cells expressed by IC50 value of 5.8 ± 1.3 µg/mL. Pretreatment with the optimal formula (orally) hinted uroprotective effect against cyclophosphamide (CP)-induced hemorrhagic cystitis (HC) in rat models, emphasized by histopathological, immunohistochemical, and biochemical investigations. In consideration of the simple fabrication process, APO-loaded CO-based NSLCs can hold prospective potential in the prophylaxis of oncologic and urologic diseases.