title: Evaluation of antibacterial potential and toxicity of plant volatile compounds using new broth microdilution volatilization method and modified MTT assay.
abstract: With aim to develop effective proof-of-concept approach which can be used in a development of new preparations for the inhalation therapy, we designed a new screening method for simple and rapid simultaneous determination of antibacterial potential of plant volatiles in the liquid and the vapour phase at different concentrations. In addition, EVA (ethylene vinyl acetate) capmat™ as vapour barrier cover was used as reliable modification of thiazolyl blue tetrazolium bromide (MTT) assay for cytotoxicity testing of volatiles on microtiter plates. Antibacterial activity of carvacrol, cinnamaldehyde, eugenol, 8-hydroxyquinoline, thymol and thymoquinone was determined against Haemophilus influenzae, Staphylococcus aureus, and Streptococcus pneumoniae using new broth microdilution volatilization method. The cytotoxicity of these compounds was evaluated using MTT test in lung fibroblast cells MRC-5. The most effective antibacterial agents were 8-hydroxyquinoline and thymoquinone with the lowest minimum inhibitory concentrations (MICs) ranging from 2 to 128μg/mL, but they also possessed the highest toxicity in lung cell lines with half maximal inhibitory concentration (IC50) values 0.86-2.95μg/mL. The lowest cytotoxicity effect was identified for eugenol with IC50295.71μg/mL, however this compound produced only weak antibacterial potency with MICs 512-1024μg/mL. The results demonstrate validity of our novel broth microdilution volatilization method, which allows cost and labour effective high-throughput antimicrobial screening of volatile agents without need of special apparatus. In our opinion, this assay can also potentially be used for development of various medicinal, agricultural, and food applications that are based on volatile antimicrobials.
authors.affiliation: Department of Crop Sciences and Agroforestry, Faculty of Tropical AgriSciences, Czech University of Life Sciences Prague, Kamycka 129, 165 21 Praha 6 - Suchdol, Czech Republic.
authors.forename: Marketa
authors.initials: M
authors.lastname: Houdkova
authors.affiliation: Department of Crop Sciences and Agroforestry, Faculty of Tropical AgriSciences, Czech University of Life Sciences Prague, Kamycka 129, 165 21 Praha 6 - Suchdol, Czech Republic.
authors.forename: Johana
authors.initials: J
authors.lastname: Rondevaldova
authors.affiliation: Department of Microbiology, Nutrition and Dietetics, Faculty of Agrobiology, Food and Natural Resources, Czech University of Life Sciences Prague, Kamycka 129, 165 21 Praha 6 - Suchdol, Czech Republic.
authors.forename: Ivo
authors.initials: I
authors.lastname: Doskocil
authors.affiliation: Department of Crop Sciences and Agroforestry, Faculty of Tropical AgriSciences, Czech University of Life Sciences Prague, Kamycka 129, 165 21 Praha 6 - Suchdol, Czech Republic. Electronic address: kokoska@ftz.czu.cz.
authors.forename: Ladislav
authors.initials: L
authors.lastname: Kokoska
mesh_terms: D000171:Acrolein / Q000031:analogs & derivatives / Q000737:chemistry; D000900:Anti-Bacterial Agents / Q000737:chemistry*; D016227:Benzoquinones / Q000737:chemistry; D002460:Cell Line; D000081007:Cymenes; D005054:Eugenol / Q000737:chemistry; D006193:Haemophilus influenzae / Q000187:drug effects; D006801:Humans; D008826:Microbial Sensitivity Tests / Q000379:methods*; D039821:Monoterpenes / Q000737:chemistry; D015125:Oxyquinoline / Q000737:chemistry; D064209:Phytochemicals / Q000737:chemistry*; D013211:Staphylococcus aureus / Q000187:drug effects; D013296:Streptococcus pneumoniae / Q000187:drug effects; D013778:Tetrazolium Salts; D013844:Thiazoles; D013943:Thymol / Q000737:chemistry; D055549:Volatile Organic Compounds / Q000737:chemistry*; D014835:Volatilization
chemical_list: D000900:Anti-Bacterial Agents; D016227:Benzoquinones; D000081007:Cymenes; D039821:Monoterpenes; D064209:Phytochemicals; D013778:Tetrazolium Salts; D013844:Thiazoles; D055549:Volatile Organic Compounds; D013943:Thymol; D005054:Eugenol; D015125:Oxyquinoline; D000171:Acrolein; C073316:carvacrol; C022616:thiazolyl blue; C003466:thymoquinone; C012843:cinnamaldehyde
keywords: Antibacterial activity; Colorimetric assay; Cytotoxicity; Essential oil; Vapour; Volatilization method