| Plants have evolved multiple disease defense mechanisms to selectively suppress pathogens by production of secondary metabolites with antimicrobial activities.Therefore,compounds with potent antiviral activity but not toxic to hosts can be directly isolated from plants and used to control plant virus diseases which are known as "plant cancer".Ailanthus altissima(Mill.)Swingle(family Simaroubaceae),belonging to genus Ailanthus,is a fast-growing medicinal plant.The fruit of Ailanthus altissima is a samara with emmenagogue,antidiarrhea,astringent and antimicrobial activity.Previous research showed that the extract of Ailanthus altissima fruit exhibited potent inhibitory effects on Tobacco mosaic virus(TMV),but there was still no systematic research on the chemical constituents with antiviral activity from Ailanthus altissima fruit.Herein,comprehensive research on the chemical constituents from Ailanthus altissima fruit and their antiviral activity against TMV replication has been conducted.The main results showed below:1)80 pure compounds were isolated from the CHC13-and n-BuOH-soluble fraction which was extracted from the MeOH extract of Ailanthus altissima fruit,including 21 quassinoids and quassinoid glycosides(1-21):chuglycoside A(1),chuglycoside B(2),chuglycoside C(3),chuglycoside D(4),chuglycoside E(5),chuglycoside F(6),shinjuglycoside A(7),chuglycoside G(8),chuglycoside H(9),chuglycoside I(10),shinjuglycoside B(11),chaparrinone(12),glaucarubolone(13),ailanthinone(14),glaucarubinone(15),ailanthone(16),?13(18)-dehydroglaucarubolone(17),dehydroailanthinone(18),chuglycoside J(19),chuglycoside K(20),shinjuglycoside C(21);14 lignans and lignan glycosides(22-35):(+)-isolariciresinol(22),(+)-isolariciresinol 3a-O-P-D-glucopyranoside(23),burselignan(24),densispicoside(25),secroisolariciresinol(26),erythro-guaiacylglycerol-?-O-4'-coniferyl ether(27),7R,8R-threo-4,7,9,9'-tetrahydroxy-3,3'?dimethoxy-8-O-4'-neolignan(28),dihydrodehydrodiconiferyl alcohol(29),curcasinlignan B(30),(+)-pinoresinol(31),(+)-(1R,2S,5R,6S)-2,6-di(4'-hydroxyphenyl)-3,7-dioxabicyclo[3.3.0]octane(32),(-)-lariciresinol(33),tetrahydro-2-(4-hydroxy-3-methoxyphenyl)-4-[(4-hydroxyphenyl)-methyl]-3-furanmethanol(34),thero-2,3-bis-(4-hydroxy-3-methoxy-phenyl)-3-methoxy-propanol(35);3 N-phenylpropionamides and N-phenylpropionamide glycosides(36-38):2-hydroxy-N-[(2-O-?-D-glucopyranosyl)phenyl]propionamide(36),2-hydroxy-N-[(2-0-?-D-glucopyranosyl-(1?6)-?-D-glucopyranosyl)phenyl]propionamide(37),2-hydroxy-N-(2-hydroxy-phenyl)propionamide(38);1 piperidine derivative(39):2?-carboxyl-piperidine-4?-acetic acid methyl ester(39);14 phenols and phenol glycosides(40-53):4-hydroxyphenyl-1-O-[6-(hydrogen 3-hydroxy-3-methylpentanedioate)]-?-D-glucopyranoside(40),arbutin(41),P-D-glucopyranosyl-(1?6)-arbutin(42),hydroquinone(43),2-(4-hydroxyphenyl)propane-1,3-diol(44),4-methoxyphenylacetic acid(45),4-hydroxybenzoic(46),protocatechuic acid(47),vanillic acid(48),gallic acid(49),methyl gallate(50),1-O-galloyl-?-D-glucose(51),3,4,8,9,10-pentahydroxydibenzo[b,d]-pyran-6-one(52),corilagin(53);10 flavonoids and flavonoid glycosides(54-63):astragalin(54),kaempferol 3-O-rutinoside(55),kaempferol 3-0-(2"-O-galloyl)-rutinoside(56),quercetin(57),isoquercitrin(58),quercitrin(59),quercetin 3-0-(2"-O-galloyl)-p-D-glucopyranoside(60),quercetin 3-O-(6"-O-galloyl)-?-D-glucopyranoside(61),rutin(62),quercetin 3-0-(2"-O-galloyl)-rutinoside(63);3 coumarins(64-66):altissimacoumarin H(64),esculetin(65),scopoletin(66);9 phenylpropanoids and phenylpropanoid glycosides(67-75):4-hydroxyphenol-1-O-[6-0-(E)-feruloyl-?-D-glucopyranosyl]-(1 ?6)-?-D-glucopyranoside(67),methyl chlorogenate(68),5-O-caffeoyl quinic acid butyl ester(69),syringin(70),p-coumaric acid(71),caffeic acid(72),3-hydroxy-1-(4-hydroxyphenyl)-1-propanone(73),?-hydroxypropi-oguaiacone(74),threo-1-(4-hydroxyphenyl)-1-methoxy-2,3-propanediol(75);3 norsesquiterpenes and norsesquiterpene glycosides(76-78):(3S,5R,6R,7E,9S)-3,5,6,9-tetrahydroxy-7-megastigmene(76),3?-gluco-pyranosyloxy-?-ionone(77),corchoionoside C(78);1 monoterpene glycoside(79):betulalbuside A(79);1 sterol glycoside(80):daucosterol(80).Among them,there were 17 new compounds(1-6,8-10,19-20,36-37,39-40,64,67)and 32 compounds(13-14,18,24-26,28,31-35,41,43-45,47,51,55-56,60,63,68-70,73,75-79)were isolated from this plant for the first time.2)Anti-TMV replication bioassay for the 80 pure compounds has been conducted to indicate that quassinoid aglycones(IC50 0.19-10.38?M)and quassinoid glycosides(IC50 18.37-137.74 ?M)exhibited significant anti-TMV replication activity.In addition,the antiviral activity of all quassinoids was more potent than the antiviral agents ningnanmycin(IC50 183.31 ?M)and ribavirin(IC50 255.19 ?M).It was worth mentioning that ailanthone(16)and chaparrinone(12)possessed the most significant antiviral activity with IC50 values at 0.19 ?M and 0.93?M,respectively.Further structure-activity relationships were also determined to indicate that:i)Glycosylation at C-2 weakens activity;the order of the antiviral activity:quassinoid aglycones>quassinoid monoglycosides>quassinoid diglycosides,ii)Carbonylation at C-12 weakens activity.iii)Dehydrogenation at C-13 and C-21 to form a exocyclic double bond enhances activity.iv)Hydroxylation at C-15 weakens activity,while esterification of the OH group may enhance activity.v)Hydrogenation of the double bond at C-3 and C-4 and the loss of an oxygen bridge from C-8 to C-11 may weaken activity.Apart from quassinoids,some lignans and phenols such as 7R,8R-threo-4,7,9,9'-tetrahydroxy-3,3'-dimethoxy-8-0-4'-neolignan(28),dihydrodehydrodiconiferyl alcohol(29),arbutin(41),4-methoxyphenylacetic acid(45)and corilagin(53)possessed moderate activity against TMV replication with IC50 values at 287.70,367.80,488.19,266.03,and 452.21 ?M,respectively.3)Using Western blot analysis,bioassay of inhibitory effects of 3 quassinoids ailanthone(16),chaparrinone(12),and glaucarubinone(15)with significant antiviral activity on accumulation of TMV CP in Nicotiana tabacum cv.K326 has been conducted to show that these 3 quassinoids exhibited strong inhibitory effects on the expression of TMV CP in tobacco plants,with a positive correlation between the degree of inhibition and the concentration of the compounds.And the expression of TMV CP can be obviously inhibited by pretreated with ailanthone,chaparrinone,and glaucarubinone at concentrations of 0.625,2.5 and 20?M,respectively.4)Bioassay of inhibitory effects of ailanthone,the best antiviral compound among 80 compounds,on the replication and spread of TMV in Nicotiana benthamiana has been conducted by using the green fluorescent protein(GFP)gene as reporter gene combined with the agroinfiltration method.The results showed that ailanthone possessed potent inhibitory effects on the replication and spread of TMV in N.benthamiana,with a positive correlation between the degree of inhibition and the concentration of the compounds.And the replication of TMV can be strongly inhibited by pretreated with ailanthone,at a concentration of 0.625 ?M.In summary,we've isolated 80 compounds from MeOH extract of Ailanthus altissima fruit.Among them,there were 17 novel compounds and 32 compounds were isolated from this plant for the first time.A series of components with potent antiviral activity has been afforded.In addition,quassinoids were comfirmed to be the main active components with anti-TMV replication activity and the antiviral structure-activity relationships of quassinoids against TMV were also discussed.The results will provide a theoretical basis for further development and utilization of plant Ailanthus altissima and new antiviral agents of plant source with high efficiency and safety. |
PDF Full Text Request