Nikkhah Mehdi, Hashemi Maryam, Habibi Najafi Mohammad B, Farhoosh Reza
Ferdowsi University of Mashhad, Faculty of Agriculture, Department of Food Science and Technology, P.O. Box 91775-1163, Mashhad, Iran.
Microbial Biotechnology Department, Agricultural Biotechnology Research Institute of Iran (ABRII), Agricultural Research, Education and Extension Organization (AREEO), 3135933151 Karaj, Iran.
Int J Food Microbiol. 2017 Sep 18;257:285-294. doi: 10.1016/j.ijfoodmicro.2017.06.021. Epub 2017 Jun 24.
The development of natural protective agents as alternatives to chemical fungicides is currently in the spotlight. In the present investigation, chemical composition and antifungal activities of thyme, cinnamon, rosemary and marjoram essential oils (EO), as well as synergism of their possible double and triple combinations were investigated. The compositions of the oils were determined by GC/MS. For determination of antifungal activity against Penicillium expansum and Botrytis cinerea, a broth microdilution method was used. The possible interactions of some essential oil combinations were performed by the two and three-dimensional checkerboard assay and isobologram construction. An in vivo antifungal assay was performed by artificial wounding of pear fruits. The maximum antifungal activity was demonstrated by thyme and cinnamon oils which displayed lower MIC values whereas rosemary and marjoram oils with MIC range between 2500 and 10,000μg/mL exhibited weak antifungal activities against tested fungi. In synergy testing, some double combinations (thyme/cinnamon, thyme/rosemary, cinnamon/rosemary) were found to be synergistic (FICi≤0.5). The triple combination of thyme, cinnamon and rosemary was synergistic for B. cinerea and P. expansum (FICi values of 0.5 and 0.375, respectively); while combination of cinnamon, marjoram and thyme exhibited additive and synergistic effect against P. expansum (FIC=0.625) and B. cinerea (FIC=0.375) respectively. The usage of a mathematical Gompertz model in relation to fungal kinetics, showed that the model could be used to predict growth curves (R=0.993±0.05). For B. cinerea, Gompertz parameters for double and triple combination treatments showed significant increase in lag phase (1.92 and 2.92days, respectively) compared to single treatments. Increase lag time up to 2.82days (P<0.05) also observed in P. expansum treated by triple combination of EOs. Base on the results, the lowest maximum growth rate (0.37mm/day) was observed in B. cinerea treated by triple combination of thyme, cinnamon and rosemary. The in vivo test also demonstrated considerable inhibitory effects of EO combination treatments. Average lesion diameter of pears treated with triple combination of cinnamon/rosemary/thyme (78, 1250, 39μg/mL) was 6mm and 8mm against B. cinerea and P. expansum respectively, in 10days at 25°C. Results also showed that double combination of thyme/cinnamon (78, 156μg/mL) has more inhibitory effect than single EO treatments.
开发天然保护剂作为化学杀菌剂的替代品目前备受关注。在本研究中，对百里香、肉桂、迷迭香和马郁兰精油（EO）的化学成分和抗真菌活性及其可能的二元和三元组合的协同作用进行了研究。通过气相色谱/质谱联用仪（GC/MS）测定了这些精油的成分。采用肉汤微量稀释法测定对扩展青霉和灰葡萄孢的抗真菌活性。通过二维和三维棋盘法及等效线图构建来研究某些精油组合的可能相互作用。通过对梨果实进行人工创伤来进行体内抗真菌试验。百里香和肉桂精油表现出最大的抗真菌活性，其最低抑菌浓度（MIC）值较低，而MIC范围在2500至10000μg/mL之间的迷迭香和马郁兰精油对受试真菌的抗真菌活性较弱。在协同试验中，发现一些二元组合（百里香/肉桂、百里香/迷迭香、肉桂/迷迭香）具有协同作用（FICi≤0.5）。百里香、肉桂和迷迭香的三元组合对灰葡萄孢和扩展青霉具有协同作用（FICi值分别为0.5和0.375）；而肉桂、马郁兰和百里香的组合对扩展青霉（FIC = 0.625）和灰葡萄孢（FIC = 0.375）分别表现出相加和协同作用。使用与真菌动力学相关的数学Gompertz模型表明，该模型可用于预测生长曲线（R = 0.993±0.05）。对于灰葡萄孢，二元和三元组合处理的Gompertz参数显示与单一处理相比，延迟期显著增加（分别为1.92天和2.92天）。在经精油三元组合处理的扩展青霉中也观察到延迟时间增加至2.82天（P<0.05）。基于这些结果，在经百里香、肉桂和迷迭香三元组合处理的灰葡萄孢中观察到最低的最大生长速率（0.37mm/天）。体内试验也证明了精油组合处理具有相当大的抑制作用。在25°C下10天时，用肉桂/迷迭香/百里香（78、1250、39μg/mL）三元组合处理的梨对灰葡萄孢和扩展青霉的平均病斑直径分别为6mm和8mm。结果还表明，百里香/肉桂二元组合（78、156μg/mL）比单一精油处理具有更强的抑制作用。
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