Characterization and synergistic antimicrobial activities of food-grade dilution-stable microemulsions against Bacillus subtilis Hui Zhang, Yan Shen, Yin Bao, Yibo He, Fengqin Feng * , Xiaodong Zheng * Department of Food Science and Nutrition, Huajiachi Campus, Zhejiang University, Hangzhou 310029, China article info Article history: Received 29 October 2007 Accepted 24 February 2008 Keywords: Microemulsion Glycerol monolaurate Sodium lactate Full dilution Antimicrobial activity abstract The physicochemical characterization of a food-grade glycerol monolaurate (GML)/ethanol/Tween 20/ sodium lactate (SL)/water microemulsion system and the antimicrobial activities against Bacillus subtilis have been studied in this paper. The influence of ethanol and SL on oil solubilization capability was clearly reflected in the phase behavior of these systems. The addition of ethanol in GML/Tween 20/water system greatly improved the oil solubilization and induced the formation of a U-type microemulsion sys- tem, while a transition from U-type to S-type microemulsion system was observed with the increasing concentration of sodium lactate. Two microemulsion formulations were obtained and remained stable by physical stability studies. Antimicrobial results showed that the microemulsions were effective in inhibiting the viable bacteria cells and the bacteria growth. Later it was found that the microemulsions had synergistic antimicrobial activities with a major contribution of GML and the solubilization of SL enhanced the antimicrobial effects. This work indicated that microemulsions are promising delivery vec- tors for antimicrobial applications in food industry. Ó 2008 Elsevier Ltd. All rights reserved. 1. Introduction Microemulsions are thermodynamically stable, transparent, low viscosity, and isotropic dispersions consisting of oil and water stabilized by an interfacial film of surfactant molecules, typically in conjunction with a cosurfactant, and have potential use as delivery systems for substances which are normally of limited use due to their hydrophobicity, toxicity or inability to access the site of ac- tion (Flanagan & Singh, 2006). There are few papers relevant to microemulsions for antimicrobial purpose. Al-Adham, Khalil, Al- Hmoud, Kierans, and Collier (2000) found that microemulsions as pharmaceutical preparations were stable and self-preserving sys- tems by the results of the challenge test and kinetics of killing experiments. Their later study (Al-Adham, Al-Hmoud, Khalil, Kier- ans, & Collier, 2003) reported that microemulsions were highly effective antibiofilm agents that resulted in rapid losses of viability in planktonic populations of tested bacteria. However, food-grade microemulsions are difficult to formulate because the complexity of food systems and the need for all of the components to be approved as being of food grade limit the choice of components, leading to difficulties in formulations (Garti, Yagh- mur, Leser, Clement, & Watzke, 2001). This study was conducted to prepare a food-grade microemulsion system with glycerol monola- urate (GML) as oil, Tween 20 as surfactant, ethanol as cosurfactant, sodium lactate (SL) and water, and evaluate the antimicrobial activities against Bacillus subtilis, which is a typical Gram-positive foodborne bacterial pathogen (Kramer & Gilbert, 1989). Glycerol monolaurate, the glycerol monoester of lauric acid, is a derivative extracted from coconut oil and has been recognized as safe by the US FDA. In addition, it is well-known as surfactant like lipid with the greatest overall antimicrobial activity of all lipids evaluated as antimicrobials (Shibasaki, 1982). These years, many research papers about its antimicrobial effects have been pub- lished (Bergsson, Arnfinnsson, Steingrimsson, & Thormar, 2001; Wang & Johnson, 1997). There are very few surfactants that can be used in food formu- lations (Dungan, 1997). In this respect, Tweens, (ethoxylated deriv- atives of sorbitan esters), are an interesting family of surfactants. It was observed, in some investigations, the goal of formulating iso- tropic water dilution lines was easily achieved in the Tween-based system (Garti et al., 2001). The liquid crystalline (LC) phase was not detected, and the two-phase region was drastically reduced. The microemulsion formulations, by the addition of short-chain alcohols, have been recently discussed (Yaghmur, Aserin, & Garti, 2002). It was reported that the addition of short-chain alcohol (such as ethanol), which increased the penetration of the surfac- tant film and decreased the polarity of water, seemed to favor the formation of microemulsions. There have been many investigations about the effects of added salts on the phase behavior (Iwanaga, Suzuki, & Kunieda, 1998; Kabalnov, Olsson, & Wennerstrgm, 1995). Sodium lactate (SL), Food Research International 41 (2008) 495–499 Contents lists available at ScienceDirect Food Research International journal homepage: www.elsevier.com/locate/foodres 0963-9969/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.foodres.2008.02.006 * Corresponding authors. Tel.: +86 571 86971163; fax +86 571 86971139. E-mail addresses: Fengfq@zju.edu.cn (F. Feng), xdzheng@zju.edu.cn (X. Zheng).