Formulation, Optimization & evaluation of Azadirachta indica Methanolic extract & allicin-loaded niosomal dry powder inhaler for the treatment of mucormycosis

ABSTRACTBackground: Mucormycosis is a rapidly progressive, angioinvasive fungal infection with mortality ratesexceeding 40–80%, particularly devastating immunocompromised individuals. Conventional antifungaltherapy, primarily liposomal amphotericin B, is limited by severe nephrotoxicity, poor tissue penetrationinto necrotic lesions, and inadequate pulmonary targeting. The COVID-19 pandemic triggered anunprecedented surge of coronavirus-associated mucormycosis (CAM), with India reporting over 50,000cases, underscoring the urgent need for novel therapeutic strategies.Objective: This study aimed to develop, optimize, and comprehensively evaluate a niosomal dry powderinhaler (DPI) co-loaded with Azadirachta indica methanolic extract (AIME) and allicin for targetedpulmonary delivery against mucormycosis, integrating synergistic phytotherapeutic antifungal activitywith advanced nanocarrier technology.Methods: AIME and allicin were extracted, standardized, and characterized. Dual-drug loaded niosomeswere prepared by thin-film hydration and optimized via Box-Behnken design (surfactant:cholesterolratio, hydration temperature, drug:lipid ratio). The optimized niosomal dispersion was spray-dried withmannitol:leucine carrier to produce respirable DPI powder, characterized for aerodynamic performanceusing Next-Generation Impactor (NGI).Results: The optimized niosomal formulation (F-Opt) exhibited vesicle size 192.4 ± 9.6 nm, PDI 0.232 ±0.022, zeta potential −29.8 ± 2.6 mV, with entrapment efficiencies of 77.4 ± 4.2% (allicin) and 80.6 ± 4.6%(AIME). Niosomal encapsulation dramatically extended allicin half-life from 18.4 minutes (free) to >72hours. Spray-dried DPI powder (SD-F-Opt) achieved fine particle fraction (FPF) of 62.8 ± 4.4%, massmedian aerodynamic diameter (MMAD) 2.86 ± 0.32 μm, and 48.4% alveolar deposition (Stage 3–5)—significantly superior to free drug DPI (FPF 42.6%, p < 0.001). Sustained release over 24 hours insimulated lung fluid confirmed diffusion-controlled kinetics. Accelerated stability (40°C/75% RH, 6months) demonstrated acceptable parameter preservation with projected 18-month shelf life.Conclusion: The developed AIME-allicin niosomal DPI represents a transformative phytomedicine-basedapproach for pulmonary mucormycosis therapy, offering synergistic antifungal action, enhanced lungtargeting, protection of labile allicin, and reduced systemic toxicity compared to conventional intravenousantifungals.