Cumin Essential Oil Extraction Optimization and Inhibition Against Postharvest Pathogenic Fungi of Fruit and Vegetables

Author:Zhang Zhong

Supervisor:bi yang


Degree Year:2018





Cumin is an important spice consumed in the world,and essential oil(EO)is its most value added processing product.Cumin EO possesses good antioxidant,antimicrobial and insect repellent activity.In this paper,Dunyu No.1 cumin seeds were used to extract EO via the optimized extraction process,and the influence of the optimized process on the composition and biological activity of cumin EO was analyzed;the antifungal activity of EO and its main component cuminal against T.roseum were compared;the synergistic effect of EO and cuminal on inhibiting the T.roseum was investigated;microscopic observations and determination of key biochemical indexes of the fungal cells were performed;and RNA-seq comparative transcriptome and iTRAQ quantitative comparative proteome analyses were conducted to explore the antifungal mechanism of the cuminal.The results showed:1.The extent of acidolysis and pulverization significantly affects the distillation EO yield.PBD and BBD optimization determined: a pretreatment of 40 mesh pulverization and acidolysis in 3 mol/l HCl for 45 min confers an EO yield of 3.802%,and 48.4% higher than that of control sample(2.562%).GC-MS identified 53 components from EO of the optimized process.Among them,the components with relative content more than 10% were cuminal(25.14%),γ-terpineal(12.74%),γ-terpinene(11.88%),and α-terpinene(11.47%);while 47 oils were identified from the EO of conventional process,and the components more than 10% were cuminal(21.89%),β-pinene(20.49%),γ-terpinene(18.37%)and γ-terpineal(10.98%).The main component of the two EOs is cuminal,but the optimized process oil contains more phenols and alcohols.2.Compared with the EO of traditional process,the EO of optimized process possesses stronger activities of DPPH free radical scavenging,ABTS free radical scavenging,iron reducing and β-carotene bleaching capacity,and the potencies have reached 2.4,4.1,73.7 and 3.2 times of that of traditional process EO,respectively.The EO of optimized process has mightier antifungal activity against four post-harvest pathogens(Alternaria alternata,Penicillium expansum,T.roseum and Fusarium sulphureum)both in mycelium growth and spore germination.The T.roseum was most sensitive to the inhibitory activity.The MIC of the EO of the optimized process against the above four fungi is 1/2 of that of the traditional process;the MFCs of the optimized process against the four fungi are lower than that of the traditional process.3.The antifungal effect of cuminal on T.roseum was significantly stronger than that of EO,and they had different antifungal kinetic models with a dose-response relationship against T.roseum.When administered directly on PDA,the MIC of EO was 1 μl/ml,while that of cuminal was 0.5 μl/ml.The MIC values of them were 1/10 when in volatilized state compared with direct contact conditions;while the MIC of the cuminal was 1/2 of the EO under both administration modes.The cupric sulfate can significantly enhance the antifungal activity,and ascorbic acid can weaken the inhibitory effect.4.In PDB,the MIC and MFC of EO against T.roseum were 15.625 μl/ml and 31.250 μl/ml,respectively.The MIC and MFC of cuminal against T.roseum was 3.906 μl/ml and 7.813 μl/ml,respectively.Antifungal kinetic model showed the EO reached highest efficiency after 720 min,and cuminal reached that after 175 min.The optimal combination of them reached after 145 min.The antifungal capacity(k value)was optimal combination> cuminal> EO.Chessboard model and response surface model analysis showed that EO and cuminal had a significant synergistic effect in killing spores and inhibiting germination.5.Cumin treatment significantly decreased the vitality of T.roseum and after washing its infectivity to apple fruits was still weak;the treatment of cuminal decreased the ergosterol content of the plasma membrane;the ATPase and mitochondrial dehydrogenase activities of the fungus decreased.T.roseum spores and mycelium had reactive oxygen accumulation,and the fungus reacted to external oxidative stress through their intrinsic antioxidant system such as CAT and SOD.The treated fungal showed typical aging characteristics including cell swelling,rough surface,thin plasma membrane,and deformed intracellular organelles.6.After treatment with cuminal,a total of 1516 genes were up-regulated and 1309 genes were down-regulated in T.roseum.Up-regulated differential genes were involved in protein degradation,signal transduction system,oxoacid metabolism and oxi-reductive process;down-regulated genes mainly were involved in cell division,central metabolism and ribosomes.A total of 90 proteins was up-regulated and 135 proteins was down-regulated,and the up protein functions mainly involved oxoacid,amino acid and other small molecules catabolism;the down proteins were involved in biosynthesis,ribosome and translation.Integration of transcriptomic and proteomic information from the perspective of metabolic pathways revealed that treatment enhanced antibiotic anabolism and lipid catabolism,while the carbohydrate metabolism decreased and the fungus exhibited metabolic characteristics of aging.In summary,acidolysis and proper pulverization can significantly improve the hydrodistillation yield of cumin seeds EO.The EO obtained in this way has better antioxidant and antifungal activity than that of traditional hydrodistillation;cuminal has stronger antifungal activity against T.roseum than the EO itself,and they have synergistic effects on the antifungal activity against T.roseum.Cuminal can significantly reduce the pathogenicity of T.roseum and destroy its cell structure and energy production system,and exert oxidative stress to the fungus;transcriptomic and proteomic analysis revealed that the treatment speeded up the cell membrane lipid degradation and secondary metabolites synthesis,and reduced the carbohydrate metabolism.