Optimizing Ultrasound-Assisted Extraction of Calicotome spinosa (L.) Link leaves using a Full-Factorial Design: linking Polyphenol Recovery to Biological Activities

Introduction: Calicotome spinosa (L.) Link., is a Mediterranean medicinal plant traditionally used for the management of several health disorders. The present study aimed to optimize the extraction of total phenolic compounds from its leaves and to investigate the biological properties of the optimized extract. Methods: Phenolic compounds were extracted by ultrasound-assisted extraction, and a three-level complete factorial design was applied to evaluate the influence of the ethanol/water ratio and sonication time on extraction efficiency. The optimized extract was characterized by determining its total flavonoid content and phenolic profile using HPLC-UV/VIS. Its antioxidant, acute oral toxicity, and anti-inflammatory activities were subsequently evaluated. Results: The ethanol/water ratio had the greatest effect on extraction efficiency, while both extraction parameters significantly influenced the recovery of phenolic compounds. The highest total phenolic content was obtained using 60% ethanol and 20 min of sonication. The optimized extract contained 36.94 ± 0.96 mg QE/g of flavonoids, and six phenolic compounds were identified by HPLC-UV/VIS. It also showed moderate antioxidant activity (DPPH IC₅₀ = 75.48 ± 5.04 μg/ mL), no signs of acute oral toxicity, and inhibited carrageenan-induced paw edema by 31.72% at 200 mg/ kg. Discussion: The results indicate that optimizing the extraction conditions improved the recovery of phenolic compounds, which may have contributed to the observed biological activities. These findings support the value of ultrasound-assisted extraction for obtaining phenolic-rich extracts from C. spinosa leaves. Conclusion: The optimized extract of C. spinosa leaves constitutes a promising source of bioactive phenolic compounds with antioxidant and anti-inflammatory potential, supporting its further investigation for pharmaceutical and nutraceutical applications.