Solid- and Vapor-Phase Antimicrobial Activities of Six Essential Oils: Susceptibility of Selected Foodborne Bacterial and Fungal Strains P. LO Ä PEZ, † C. SA Ä NCHEZ, †,§ R. BATLLE,* ,† AND C. NERI ÄN † Department of Analytical Chemistry, Arago ´n Institute of Engineering Research, i 3 A, CPS-University of Zaragoza, Marı ´a de Luna st. 3, E-50018 Zaragoza, Spain, and Department of I+D+i, ARTIBAL S.A., Can ˜ada Real st. 12, E-22600 Sabin ˜a ´nigo, Spain The antimicrobial activity of essential oils (EOs) of cinnamon (Cinnamon zeylanicum), clove (Syzygium aromaticum), basil (Ocimum basillicum), rosemary (Rosmarinus officinalis), dill (Anethum graveolens), and ginger (Zingiber officinalis) was evaluated over a range of concentrations in two types of contact tests (solid and vapor diffusion). The EOs were tested against an array of four Gram-positive bacteria (Staphylococcus aureus, Bacillus cereus, Enterococcus faecalis, and Listeria monocytogenes), four Gram-negative bacteria (Escherichia coli, Yersinia enterocolitica, Salmonella choleraesuis, and Pseudomonas aeruginosa), and three fungi (a yeast, Candida albicans, and two molds, Penicillium islandicum and Aspergillus flavus). The rationale for this work was to test the possibility of creating a protective atmosphere by using natural compounds that could extend the shelf life of packaged foodstuffs while minimizing organoleptic alterations. In the solid diffusion tests, cinnamon and clove gave the strongest (and very similar) inhibition, followed by basil and rosemary, with dill and ginger giving the weakest inhibition. The fungi were the most sensitive microorganisms, followed by the Gram-positive bacterial strains. The Gram-negative strain P. aeruginosa was the least inhibited. The composition of the atmosphere generated by the EOs, and their minimum inhibitory concentrations (MICs), were determined using a disk volatilization method, in which no inhibition from rosemary or basil was observed. Cinnamon and clove, once again, gave similar results for every microorganism. As a general rule, MIC (fungi) , MIC (bacteria) with no clear differences between Gram-positive or -negative strains except for P. aeruginosa, which was not inhibited by any of the EOs in the vapor phase. The atmosphere generated from the EOs was analyzed by means of solid-phase microex- traction combined with gas chromatography-ion trap mass spectrometry. Differences among the volatiles in the EOs, which may be responsible for the differences in their antimicrobial performances, were found. KEYWORDS: Essential oils; comparative evaluation; vapor-phase antimicrobial activity; minimum inhibitory concentration; atmospheric composition INTRODUCTION Many food products are perishable and require protection from microbial spoilage during their shelf life. Traditional food preservation techniques or the use of refrigeration alone cannot ensure the quality and safety of all foods, and alternative preservation techniques, such as modified atmosphere packaging (MAP), the use of pulsed light, electric, or magnetic fields, high pressure, irradiation, or a wide range of food-grade chemicals, are being applied or investigated for their preservative potential (1-5). Nevertheless, food processors and consumers have expressed a desire to reduce the use of both aggressive techniques and synthetic chemicals in food, mainly because the presence of chemical residues in foods and labeling of preserva- tives in food packages are major concerns nowadays. Therefore, there is a clear need for new methods of preserving food using natural additives, and a very interesting option is the use of essential oils (EOs) as antimicrobial additives, because they are rich sources of biologically active compounds. EOs are mainly obtained by steam distillation from various plant sources. The antimicrobial activity of EOs has been extensively studied and demonstrated against a number of microorganisms, mostly in vitro rather than in tests with food, and usually using a direct-contact antimicrobial assay. In these tests EOs are brought into contact with the selected microorgan- isms, and their inhibition is monitored by means of direct inspection or by measuring a physical property that is directly * Author to whom correspondence should be addressed (telephone 34- 976761000, ext. 5296; fax 34-9762388; e-mail r13bat@unizar.es). † Arago ´n Institute of Engineering Research. § ARTIBAL S.A. J. Agric. Food Chem. 2005, 53, 6939-6946 6939 10.1021/jf050709v CCC: $30.25 © 2005 American Chemical Society Published on Web 08/02/2005