Study On The Chemical Constituents And Antimicrobial Effect And Mechanism Of Alpinia Coriandriodora D. Fang

Alpinia coriandriodora D.Fang belongs to the Alpinia.The rhizome has the effect of dispelling rheumatism and influenza.Folk use whole herbs to treat stomach pain and wound ulceration.As edible plants,rhizome and stem of A.coriandriodora can be used as condiment,and the leaves can be used to pack rice dumplings.At present,there is a lack of study on chemical constituents and related pharmacological activities of A.coriandriodora.Due to the main medicinal and edible parts was the rhizome,the chemical constituents and related pharmacological activities of rhizome were studied.The paper mainly studies chemical constituents and pharmacological activities of essential oil（EO）,water extract（WE）and ethanol extract（EE）were extracted from rhizome of A.coriandriodora.（1）EO was extracted by steam distillation method,and the extraction rate was0.56%.WE and EE were extracted by condensation reflux method,and the extraction rates were 2.04%and 1.37%,respectively.The chemical constituents of EO were analyzed by GC-MS,33 compounds were identified,and it was rich of（E）-2-decennal and trans-2-decene-1-acetate.The contents of total phenolic acids and total flavonoids of WE and EE were determined by Folin-Ciocalteu method and Na NO₂-Al（NO₃）₃-Na OH colorimetric method.The results showed that WE（60.49±0.24 mg GAE/g）and EE（52.99±0.16 mg GAE/g）owned high total phenolic acids contents.UPLC-Q-Orbitriap MS was used to identify the chemical constituents of WE and EE.It was total of 64compounds from WE（38 compounds）and EE（55 compounds）,which including 37phenolic acids and 13 flavonoids compounds.WE and EE showed high total phenolic acids content and abundant variety.（2）DPPH and ABTS assay were used to study the antioxidant activity of the rhizome of A.coriandriodora.The results showed that WE and EE had strong antioxidant activities.The IC₅₀ values of WE（6.59±0.88μg/m L）and EE（17.70±1.15μg/m L）showed stronger than BHT（21.83±0.89μg/m L）agianst DPPH radical scavenging.And the IC₅₀ values of WE（20.48±0.44μg/m L）and EE（18.27±0.55μg/m L）showed good ABTS radical scavenging activity.However,EO showed poor antioxidant activity.（3）The inhibitory activities of tyrosinase,α-glucosidase,acetylcholinesterase and butyrylcholinesterase of A.coriandriodora were measured.The results showed that the inhibitory activity of WE(IC₅₀ values:17.43±1.69μg/m L)and EE(IC₅₀ values:13.41±0.72μg/m L)onα-glucosidase was stronger than that of acarbose(IC₅₀ values:236.68±9.78μg/m L).EO,WE and EE had good activities for tyrosinase(IC₅₀ values:1010.91-19849.64μg/m L),acetylcholinesterase(IC₅₀ values:1587.89-6668.89μg/m L)and butyratecholinesterase(IC₅₀ values:379.51-5730.65μg/m L).（4）The antimicrobial activity of rhizome of A.coriandriodora was preliminarily evaluated by inhibition zone,MIC and MBC experiments and Aspergillus flavus experiments.The results showed that EO had the strong antibacterial effect against Staphylococcus aureus,Pseudomonas aeruginosa and Proteus vulgaris.The diameter of the inhibition zone range from15.59 mm to 22.17 mm,the MIC value range from 781.25μg/m L to 6250μg/m L,and the MBC range from 3125μg/m L to 6250μg/m L.However,WE and EE showed weak antibacterial activity.Particularly,EO showed strong antifungal activity,the minimum inhibitory concentration was 362.5μg/m L,the minimum fungicidal concentration was 625μg/m L.WE and EE have poorly resisted against A.flavus.Hence,EO had good antimicrobial activity.（5）The fractionation inhibitory concentration index（FICI）of EO combined with antimicrobial agents（streptomycin,tetracycline and gentamicin）on S.aureus,B.subtilis,P.aeruginosa,E.coli,P.vulgaris and E.faecalis was measured by chessboard method to determine the synergistic effect.The results showed that the FICI of EO combined with streptomycin against the six strains ranging from 0.375 to 2.5,and the FICI against S.aureus was 0.375 with the better synergistic effect.The FICI of EO combined with tetracycline against the six strains ranging from 0.5 to 2.5,and that against P.vulgaris was 0.5,EO showed good synergistic effect.The FICI of EO combined with gentamicin against the six strains ranging from 0.75 to 7,and that against S.aureus was 0.75.Hence,EO showed a general synergistic effect.（6）The antibacterial mechanism of A.coriandriodora was studied by using S.aureus exposed to EO.AKP experiment showed that the cell wall was destroyed.Rhodamine 123 experiment and relative conductivity experiment showed that the permeability of cell membrane was damaged.The extracellular protein excretion experiment showed that the integrity of cell membrane was destroyed.Intracellular protein experiment showed that intracellular protein synthesis was inhibited.According to ROS experiment,EO activated oxidative stress in bacterial cells.Hence,experiments preliminarily showed that the antimicrobial mechanism of EO may be through activating the level of oxidative stress,reducing the synthesis of proteins,thereby affecting（the conformation of cell wall,the permeability of cell membrane,and membrane potential）,finally inhibiting the proliferation of bacteria or leading to bacterial death.（7）Total phenolic acids and flavonoids,antioxidant and enzyme inhibitory activities of WE and EE of stems and leaves were studied.The results showed that the contents of total phenolic acids were the highest in stems EE（positive equivalents:84.88±0.68mg GAE/g）and leaves WE（positive equivalents:83.70±0.62 mg GAE/g）.The highest content of total flavonoids was EE of leaves（positive equivalent:79.19±4.84mg RE/g）.The results of antioxidant experiments showed that WE and EE of all parts from A.coriandriodora showed good antioxidant effects.The IC₅₀of DPPH radical scavenging rate was 6.59-22.93μg/m L,and that of ABTS radical scavenging rate was8.84-32.58μg/m L.EO,WE and EE of A.coriandriodora had good inhibitory effects against the four selected enzymes,especially forα-glucosidase,all of part of A.coriandriodora(IC₅₀ values:2.34-54.81μg/m L)showed stronger than acarbose(IC₅₀values:236.68±9.78μg/m L).In summary,the chemical constituents of rhizomes of A.coriandriodora were analyzed.EO was rich of（E）-2-Decennal and trans-2-decene-1-acetate,WE and EE owned high total phenolic acids content and abundant variety.WE and EE have good antioxidant activity and enzyme inhibition activity,especially for DPPH andα-glucosidase,those were stronger than positive（BHT,acarbose）.EO had good antimicrobial activity and strong inhibitory effect on the growth of A.flavus,and EO combined with streptomycin had a good synergistic effect against S.aureus.According to the preliminary mechanism study of EO showed anti-microbial effect may be by activating the level of oxidative stress,reducing protein synthesis,thereby affecting（the conformation of cell wall,membrane permeability and membrane potential）,finally inhibiting the proliferation of bacteria or leading to bacterial death.Hence,the above research shows that A.coriandriodora has great development value in food preservation,medical industry,health care and other aspects.