| Genus Dracocephalum is an important member of the Lamiaceae family.This genus comprises more than 60 species.Most plants distribute in temperate regions of the Northern Hemisphere.Some species have been documented as folk medicine for the treatment of a wide variety of symptom such as coronary heart diseases,icteritious hepatitis,gastritis,chronic bronchitis and ulcers.Dracocephalum moldavica L.is an annual plant belonging to this genus.In China,D.moldavica is mainly found in Xinjiang Autonomous Region and believed to have benefits for the treatment of coronary heart disorders,hypertension and headache.The same plant named as Moldavian balm in Iran,which was used for the treatment of tonic,stomachic,antiemetic and diaphoretic.In this paper,we first optimized the extraction process of D.moldavica.The most suitable extraction solvent(60%ethanol)was determined..Then we used different types of macroporous resins to accumulate the phenolic acid compounds in the D.moldavica,and the most suitable macroporous resins(DM130)was selected,in order to ensure the bioactive compounds were retained while the impurities such as pigments and sugars were removed at the same time.Using an ARE-dependent luciferase reporter assay,we identified the 40%,60%ethanol elution fraction were the most potent fraction,then these two parts were selected for the follow chemical component separation.In this study,50 compounds(including 13 new compounds and 37 known compounds)were isolated from the aerial parts of D.moldavica by MCI gel,silca gel,Sephadex LH-20,C18 chromatography and semi-preparative HPLC.The structure of the compounds was established by using spectroscopic data including NMR,MS,UV,IR and CD.The new compounds include three phenylpropionic acid dimers,dracomolphin A-C(1-3);a neoligan,dracomolphin D(4);five 3,4-seco-phenylpropionic acids,(R)-(+)-dracomolphin E-I(5-9);and two pairs of enantiomers of phenylpropionic acid dimers,(7'R,8'S)-(-)-dracomolphin J(10),(7'S,8'R)-(+)-dracomolphin J(11),(7'S,8'S)-(-)-dracomolphin J(12),(7'R,8'R)-(+)-dracomolphin J(13).Separation of 10-13 was achieved by using chiral HPLC and absolute configurations of 5,6,8-13 were assigned by using circular dichroism(CD)and optical rotation.37 known compounds were obtained,including 15 flavonoids:tilianin(14),cosmosiin(15),cymaroside(16),diosmetin-7-O-?-D-glucoside(17),apigenin-7-O-?-D-glucuronide methyl ester(18),acacetin-7-O-?-D-glucuronide methyl ester(19),diosmetin-7-O-?-D-glucuronide methyl ester(20),luteolin-7-O-?-D-glucuronide methyl ester(21),acacetin-7-O-?-D-glucuronide(22),acacetin(23),apigenin(24),circimaritin(25),kumatakenin(26),santaflavone(27),xanthomicrol(28);13 phenylpropanoid:methyl(Z)-3-(4-hydroxy-3-methoxyphenyl)-2-{2-methoxy-4[(E)-3-methoxy-3-oxoprop-1-enyl]phenoxy}-prop-2-enoate(29),3-benzofurancarboxylic acid-2-(3,4-dihydroxyphenyl)-7-hydroxy-5-(3-methoxy-3-oxo-l-propenyl)-methyl ester(30),methyl(E)-3-[2-(3,4-dihydroxyhenyl)-7-hydroxy-3-methoxycarbonyl-2,3-dihydro-l-benzofuran-5-yl]prop-2-enoate(31),(R)-(+)-Oresbiusin B(32),(R)-(+)-rosmarinic acid methyl ester(33),(R)-(+)-rosmarinic acid ethyl ester(34),d-pinoresinol(35),epi-pinoresinol(36),eugenol(37),caffeic acid methyl ester(38),ferulic acid methyl ester(39),(R)-(+)-Oresbiusin A(40),scopoletin(41);3 triterpenoids:ursolic acid(42),2a-hydroxy-ursolic acid(43),oleanic acid(44);1 steroid:?-sitosterol(45);5 aromatic compounds:3-methoxy-4-hydroxyanisole(46),syringaldehyde(47),methyl protocatechuate(48),4-hydroxybenzoic acid methyl ester(49),3,4-dihydroxyphenylacetic acid methyl ester(50).Nrf2 is a transcription factor which mainly regulate the hemostasis of cells.Nrf2 regulate the expression of phase ? and antioxidant enzymes,which could protect cells from the damage of free radicals and exogenous toxicants.It is reported that induction of Nrf2 expression was helpful for the treatment of a number of chronic diseases such as cardiovascular disease,diabetes and neurodegenerative disease.Some of the compounds obtained were evaluated the potential Nrf2 inducing activity using HEK 293 cells which stable transfected with luciferase.Results showed that compounds 5,6,8,9,14,15,16,32,38 significantly induced expression of luciferase and could be potential Nrf2 activitors.Immunoblot analysis and immunofluorescence were also achieved among compounds 5,6,8,9 to confirm the activation of Nrf2 pathway.The inhibition of lung cancer cells proliferation was also tested,it was found that the IC50 of compound 30 was 4.33 ?M in NCI-H292 cells,which indicated that this compound might be a lung cancer cell proliferation inhibitor. |
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