Extraction of betel leaves (Piper betle L.) essential oil and its bio-actives identification: Process optimization, GC-MS analysis and anti-microbial activity

Betel leaf (Piper belle L.) var. Bangla is an essential oil (EO) rich plant belongs to Piperaceace family used as traditional herbal medicine. The extraction of EO from fresh and cured betel leaves is of high interest for industrial application. Thus, the present study was aimed to optimize the EO extraction and its bio-chemical characterisation. Box-Behnken Design coupled with response surface methodology was employed to optimize the independent variables such as liquid to solid ratio (30:1-50:1 mL/g), extraction time (5-7 h), and particle size (0-20 mesh). Quadratic polynomial models were found to be highly significant (p < 0.05) obtaining fresh and cured betel leaf EO with a yield of 0.18% +/- 0.01% and 0.22% +/- 0.02% at optimal condition: 40:1 mL/g, 6 h, and 10 mesh. Gas chromatography mass spectrometry (GC-MS) analysis revealed thirty-three and thirty volatile compounds, representing 98.41% and 97.34% of the optimized fresh and cured leaf EO, respectively which have a varied range of biological activities and industrial applications. Eugenol, estragole, linalool, a-copaene, anethole, chavicol, and caryophyllene were found to be highly abundant in both EO with different percentages. Antibacterial activity was evaluated against Mycobacterium smegmatis, Staphylococcus aureus and Pseudomonas aeruginosa and the results demonstrated that cured leaf EO showed significantly higher antimicrobial activity against M. smegmatis than fresh leaf EO. Moreover, morphological study was carried out to investigate the extraction mechanism of fresh and cured betel leaves with the help of scanning electron microscopy (SEM).