Coaxial Electrospraying for Black Seed Oil Nanoencapsulation: Improved Thymoquinone Stability and Bioactivity

Abstract

Thymoquinone is the primary bioactive compound in black seed oil (BSO), but it has limited stability under harsh conditions. Nanoencapsulation using zein via coaxial electrospraying is an effective approach to improve its stability and bioactivity during gastrointestinal transit. Therefore, this study aimed to develop BSO-loaded nanoparticles using coaxial electrospraying, evaluate their stability at different temperatures (4°C, 25°C and 60°C), and investigate their in vitro bioaccessibility, intestinal permeability using Caco-2 cell models, and immunomodulatory potential. Nanoparticles with high encapsulation efficiency (74.38% ± 2.6%) and loading capacity (12.99% ± 3.6%) retained thymoquinone content at 91.3%, 74.7%, and 52.0% after 55 days of storage at 4°C, 25°C, and 60°C, respectively. The bioaccessibility of thymoquinone in nanoparticles increased from 21.7% to 33.1% when orange juice was fortified with nanoparticles. The apparent permeability of thymoquinone in nonencapsulated oil, nanoparticles, and orange juice fortified with nonencapsulated oil and nanoparticles was 2.10 × 10⁻⁷, 5.76 × 10⁻⁷, 4.39 × 10⁻⁷, and 7.38 × 10⁻⁷ cm/s, respectively. When the intracellular signaling pathways were analyzed for its intracellular mechanism of action, thymoquinone nanoparticles affected the activation levels of the p38 MAPK and PI3K proteins and stimulated macrophages to generate substantial levels of pro-inflammatory cytokines, including TNFα, IL6, GMCSF, and IL12p40. BSO-loaded nanoparticles can be incorporated into functional food formulations to deliver health benefits.