Characterization of Freeze-Dried Bitter Orange (Citrus aurantium L.) Peel and Recovery of Bioactive Compounds by Microwave- and Ultrasound-Assisted Extraction

Bitter orange peel (Citrus aurantium L.) is an abundant agro-industrial by-product and a promising source of natural bioactive compounds for high-value applications. This study comprehensively characterized freeze-dried bitter orange peel (BOP) and evaluated microwave-assisted extraction (MAE) and ultrasound-assisted extraction (UAE) for the recovery of antioxidant compounds. The freeze-dried powder exhibited low moisture content and suitable physicochemical, technological, and functional properties, supporting its use as a stable raw material for further valorization. Its chemical characterization revealed the presence of vitamin C, phenolic compounds, and carotenoids, including 0.40 mg/g DW of vitamin C and 4.31 mg/g DW of total carotenoids. Randomized full factorial designs were used to evaluate the effects of solvent type, extraction time, and microwave power or acoustic intensity on extraction yield, vitamin C recovery, total phenolic compounds (TPC), and antioxidant activity (TEAC). Solvent type was the most influential factor in both extraction processes, whereas extraction time and energy input exerted response-dependent effects. Among the experimentally evaluated MAE conditions, ethanol at 100 W for 30 s produced the highest TPC (514.38 ± 15.85 mg GAE/g extract) and the second-highest TEAC value (597.75 ± 2.40 µmol TE/g extract). For UAE, ethanol at 92 W/cm2 for 5 min provided the highest TPC (760.05 ± 81.06 mg GAE/g extract) and TEAC (513.11 ± 9.18 µmol TE/g extract), although aqueous extraction resulted in higher extraction yields. Confocal laser scanning microscopy (CLSM) revealed a greater reduction in residual green autofluorescence after UAE than after MAE, providing complementary qualitative evidence of a more extensive removal of naturally fluorescent constituents from the plant matrix. In summary, freeze-dried BOP represents a promising raw material for bioactive compound recovery, while MAE and UAE provide rapid extraction approaches whose environmental performance should be further assessed using quantitative sustainability metrics. The resulting extracts warrant further investigation for potential food, nutraceutical, cosmetic, and pharmaceutical applications.