Studies On Chemical Constituents From Dodartia Orientalis L. And Their Antibacterial Activities And Action Mechanisms

Dodartia orientalis L.which is also called Niuhanshui,Daozhuacao and Duodecao belongs to the Dodartia of Scrophulariaceae family,and distributes in Xinjiang Uygur Autonomous Region and Inner Mongolia Autonomous Region of China,and other countries including People’s Republic of Mongolia and Kazakhstan also have its habitats.The whole herb of D.orientalis can be used medicinally,and it has the efficacy of clearing heat,detoxifying,dispersing wind,stopping itching,etc.D.orientalis has been as traditional Chinese medicine for the treatment of urticaria,allergic rhinitis and bronchitisin in our country for a long time.As an important medicinal plant resource,it is recorded in many kinds of pharmaceutical works such as "National Chinese Herbal Medicine Collection" and "Inner Mongolia Herbal Medicine".Up to now,there are few researches on the phytochemical composition and pharmacological activity of D.orientalis.On the other hand,foodborne pathogens have been threating people’s health which leads to the urgent need of new antibacterial drugs.Therefore,we reviewed the chemical constituents and pharmacological activities of D.orientalis,and separated the chemical constituents of D.orientalis systematically,and studied the antibacterial activity and antibacterial mechanism of its chemical constituents.1.The essential oil of D.orientalis was extracted by the hydro-distilled method,and the compositions in D.orientalis essential oil were identified by GC–MS analysis.The results show: Totally,34 compounds were identified,representing 94.65% of total compositions.And most of the identified compounds belonged to monoterpenes and sesquiterpenes,such as the highest contents of the two components were both monoterpenes,which were linalool(32.25%)and α-terpineol(13.83)respectively.2.The chemical compositions of 70% ethanol extracts from D.orientalis were studied systematically.50 compounds were isolated,and 40 compounds were identified,including eleven flavonoids: Apigenin(1),Luteolin(2),Quercetin(3),Diosmetin(4),Chrysoeriol(5)Tricin(6),5-hydroxy-4′,6,7-Trimechoxy flavone(7),Artemetin(8),Rutin(9),Hyperoside(10),Luteolin-7-O-β-D-glucoside(11);nine triterpenoids: Lupeol(12),Birch aldehyde(13),Betulin(14),Betulinic acid(15),Zizyberenalic acid(16),Oleanolic acid(17),Ursolic acid(18),Rubiarbonol A(19),Pomolic acid(20);four Steroids: β-sitosterol(21),Stigmasterol(22),Ergosterol(23),Daucosterol(24);four Phenylpropionic acids: Ferulic acid(25),Isoferulic acid(26),Caffeic acid(27),Chlorogenic acid(28);five Phenolic acids,phenolics and polyphenols: Gallic acid(29),Syringaldehyde(30),Syringic acid(31),Protocatechuic acid(32),Resveratrol(33);three iridoids and sesquiterpenoids: Specioside(34),Geniposide(35),Roxbughianin B(36);two lignans:(-)-5’-methoxyisolariciresinol(37),(-)-5’-methoxyisolariciresinol-3α-O-β-D-glucopyranoside(38);two other compounds:Mannitol(39),n-Eicosanol(40).There were 26 compounds obtained from D.orientalis for the first time and were also obtained from the genus of Dodartia for the first time.3.The antibacterial activities and action mechanisms against S.aureus,E.coli and S.enteritidis of D.orientalis essential oil were studied.The antibacterial activities of essential oil to the above three pathogens was determined by the DIZ test and broth microdilution method,the results showed that the MIC of D.orientalis essential oil were 0.5,1,and 2μL/m L while the MBC were 1,2 and 4 μL/m L against the strains of S.aureus,E.coli and S.enteritidis,respectively.AKP leak assay revealed that D.orientalis essential oil caused the destruction of cell wall,leading to the leak of AKP.Nucleic acids leak,potassium ions leak,and PI staining assays showed that D.orientalis essential oil aroused the destruction of cell membrane,leading to the leak of intracellular nucleic acids and potassium ions.SEM observation further certified that D.orientalis essential oil could suppress bactericidal activity via disrupting cell walls and membranes.On the other hand,the assays of activity of cellular metabolism of biofilm and biofilm biomass indicated that D.orientalis essential oil had the ability to resist biofilm through decreasing biofilm biomass and biofilm cellular activity.The bacterial surface hydrophobicity experiment showed that one of the anti-biofilm mechanisms of this essential oil is to reduce bacterial adhesion.4.The antibacterial activities of compounds from D.orientalis against MRSA were detected and the antibacterial mechanisms of oleanolic acid and ursolic acid against MRSA were studied.The anti-MRSA activity of oleanolic acid(17)and ursolic acid(18)was determined by broth microdilution method,the results showed that the MIC of oleanolic acid and ursolic acid were 64 and 32 μg/m L while the MBC were 128 and 64 μg/m L against MRSA,respectively;syringaldehyde(30)and resveratrol(33)also exhibited anti-MRSA activities and their MICs were both 512 μg/m L.NPN and nucleic acids leak assays revealed that oleanolic acid and ursolic acid caused the destruction of integrity of cell membrane.The assays of activity of cellular metabolism of biofilm and biofilm biomass indicated that oleanolic acid and ursolic acid both had the ability to resist biofilm.The assay of β-lactamase activity showed that ursolic acid had the ability to reduce the activity of β-lactamase produced by MRSA significantly,while oleanolic acid had no effect.