Physiological, phytochemical, and antibacterial responses of Anoectochilus formosanus cuttings under in vitro culture to cold atmospheric plasma jet exposure

• Cold plasma jet enhanced growth and biomass of A. formosanus in vitro . • CPJ improved chlorophyll, carotenoid, and antioxidant enzyme activities. • CPJ enhanced isorhamnetin (1.27-fold) and kinsenoside (2.46-fold) levels analyzed by Liquid Chromatography-Mass Spectrometry. • CPJ extracts inhibited C. albicans, Escherichia coli , and S. aureus activities. This study aimed to investigate the effects of air cold plasma jet (CPJ) treatment with varying exposure times (30–120 s) on the physiological characteristics, bioactive compounds, antioxidant capacity, and antibacterial activity of jewel orchid ( Anoectochilus formosanus ) cultured in vitro over six weeks. The results demonstrated that CPJ treatment for 60 s produced the greatest improvements in plant growth, with plant height, fresh weight, and dry weight increasing by 57.87, 195, and 263%, respectively, compared with the control. These enhancements were attributed to multiple physiological and biochemical changes, including elevated chlorophyll and carotenoid contents, enhanced activities of antioxidant enzymes, as well as reduced membrane lipid peroxidation. Moreover, the total phenolic, flavonoid, DPPH radical scavenging, and FRAP assays were increased by 22.53, 7.40, 22.65, and 81.10%, respectively. CPJ treatment altered the phytochemical profile of A. formosanus, with isorhamnetin and kinsenoside levels increasing 1.27- and 2.46-fold after 120 s and 60 s exposures, respectively. CPJ-treated extracts (0–120 s) inhibited C. albicans, Escherichia coli , and S. aureus growth, with maximum inhibition of 37.55% against S. aureus at 313 µg/mL after 30 s treatment. Antimicrobial effects were strongest in the early weeks but diminished over time. Thus, these findings highlight that cold plasma technology is a promising method for improving growth, phytochemical profile, antioxidant defense, and antibacterial activity in A. formosanus in vitro culture.