Center for Food Safety and Department of Food Science, University of Arkansas, Fayetteville, AR 72701, USA.
BMC Complement Altern Med. 2012 Aug 16;12:125. doi: 10.1186/1472-6882-12-125.
Staphylococcus aureus is the pathogen most often and prevalently involved in skin and soft tissue infections. In recent decades outbreaks of methicillin-resistant S. aureus (MRSA) have created major problems for skin therapy, and burn and wound care units. Topical antimicrobials are most important component of wound infection therapy. Alternative therapies are being sought for treatment of MRSA and one area of interest is the use of essential oils. With the increasing interest in the use and application of natural products, we screened the potential application of terpeneless cold pressed Valencia orange oil (CPV) for topical therapy against MRSA using an in vitro dressing model and skin keratinocyte cell culture model.
The inhibitory effect of CPV was determined by disc diffusion vapor assay for MRSA and vancomycin intermediate-resistant S. aureus (VISA) strains. Antistaphylococcal effect of CPV in an in vitro dressing model was tested on S. aureus inoculated tryptic soya agar plate. Bactericidal effect of CPV on MRSA and VISA infected keratinocyte cells was examined by enumeration of extra- and intra-cellular bacterial cells at different treatment time points. Cytotoxic effects on human skin cells was tested by adding CPV to the keratinocyte (HEK001) cells grown in serum free KSFM media, and observed by phase-contrast microscope.
CPV vapour effectively inhibited the MRSA and VISA strains in both disc diffusion vapour assay and in vitro dressing model. Compared to untreated control addition of 0.1% CPV to MRSA infected keratinocyte decreased the viable MRSA cells by 2 log CFU/mL in 1 h and in VISA strain 3 log CFU/mL reduction was observed in 1 h. After 3 h viable S. aureus cells were not detected in the 0.2% CPV treatment. Bactericidal concentration of CPV did not show any cytotoxic effect on the human skin keratinocyte cells in vitro.
At lower concentration addition of CPV to keratinocytes infected with MRSA and VISA rapidly killed the bacterial cells without causing any toxic effect to the keratinocytes. Therefore, the results of this study warrant further in vivo study to evaluate the potential of CPV as a topical antistaphylococcal agent.
金黄色葡萄球菌是引起皮肤和软组织感染最常见和最普遍的病原体。近几十年来，耐甲氧西林金黄色葡萄球菌（MRSA）的爆发给皮肤治疗、烧伤和伤口护理带来了重大问题。局部抗菌药物是治疗伤口感染的最重要组成部分。人们正在寻找治疗 MRSA 的替代疗法，其中一个关注领域是使用精油。随着人们对天然产品的使用和应用兴趣的增加，我们筛选了萜烯缺失的冷榨瓦伦西亚橙油（CPV）在 MRSA 局部治疗中的潜在应用，使用体外敷料模型和皮肤角质形成细胞培养模型。
通过对耐甲氧西林金黄色葡萄球菌（MRSA）和中间耐万古霉素金黄色葡萄球菌（VISA）菌株的圆盘扩散蒸汽测定法，确定 CPV 的抑制作用。在金黄色葡萄球菌接种胰蛋白酶大豆琼脂平板的体外敷料模型中，测试 CPV 的抗葡萄球菌作用。通过在不同治疗时间点计数细胞外和细胞内细菌细胞，检查 CPV 对感染角质形成细胞的 MRSA 和 VISA 的杀菌作用。通过将 CPV 添加到无血清 KSFM 培养基中生长的角质形成细胞（HEK001）中，测试对人皮肤细胞的细胞毒性作用，并通过相差显微镜观察。
CPV 蒸气在圆盘扩散蒸汽测定法和体外敷料模型中均能有效抑制 MRSA 和 VISA 株。与未处理的对照相比，在 1 小时内，向感染 MRSA 的角质形成细胞中添加 0.1%CPV 可使活的 MRSA 细胞减少 2 对数 CFU/mL，在 VISA 株中观察到 3 对数 CFU/mL 的减少。在 0.2%CPV 处理后 3 小时内，未检测到活的金黄色葡萄球菌细胞。CPV 的杀菌浓度对体外人皮肤角质形成细胞无任何细胞毒性作用。
在较低浓度下，CPV 与感染 MRSA 和 VISA 的角质形成细胞一起添加，可迅速杀死细菌细胞，而不会对角质形成细胞造成任何毒性作用。因此，这项研究的结果需要进一步的体内研究来评估 CPV 作为局部抗葡萄球菌剂的潜力。
