Application of response surface methodology for optimizing flavonoid extraction and antimicrobial activity from Pistacia terebinthus male flowers

Pistacia terebinthus is a Mediterranean dioecious shrub rich in flavonoids and exhibits various biological activities, including antimicrobial effects. While the leaves and fruits of this species have been studied extensively, its flowers remain underexplored. This study aims to optimize the extraction conditions for total flavonoid content from P. terebinthus male flowers and assess their antimicrobial activity against selected bacterial and fungal strains. The optimization was carried out using response surface methodology with a Box-Behnken design, evaluating four parameters: extraction temperature, time, methanol concentration, and solid-to-liquid ratio. The optimal extraction conditions were: 60% methanol, a solid-to-liquid ratio of 1:19.5, and extraction at 32.6 °C for 63 min, yielding a total flavonoid content of 953.27 mg quercetin equivalents 100 g–1 dry mass. The extract showed significant antimicrobial activity, particularly against Aspergillus niger (19.12 mm inhibition zone), followed by Bacillus cereus (16.53 mm). Fungal strains were generally more sensitive than bacterial strains. ANOVA confirmed the significance and predictive power of the quadratic models used for optimization. The findings support the potential use of male flower extracts in the development of natural antimicrobial agents, and underscore the effectiveness of response surface methodology with a Box-Behnken design in optimizing extraction conditions.