CN1057834A - Be used for the treatment of the pyridine-benzoic acid derivative that is connected with the acid amides of leukotriene-related disease - Google Patents

Abstract

The invention relates to following formula: compound

T is that acid amides connects base in the formula, and each R base defines in specification sheets.These compounds are as leukotriene antagonist.

Description

Be used for the treatment of the pyridine-benzoic acid derivative that is connected with the acid amides of leukotriene-related disease

The present invention relates to be used for the treatment of the pyridine-benzoic acid derivative that is connected with the acid amides of leukotriene-related disease.These compounds are used in particular for treatment by hydroxyl leukotriene, especially LTB ₄And LTB ₄The disease that-gaonist active substance causes.

The biological activity lipid that is commonly referred to as leukotriene is for breathing, and cardiovascular and gastro-intestinal system produces pharmacological action.Leukotriene is divided into two subclass usually, i.e. peptide leukotriene (leukotriene C ₄, D ₄And E ₄) and hydroxyl leukotriene (leukotriene B ₄).The present invention relates generally to hydroxyl leukotriene (LTB ₄), but be not limited to this specific leukotriene.

The peptide leukotriene is relevant with biological response, and this biological response (SRS-A) interrelates with " the anaphylactoid material of slowly replying ".This replying shows as the prolongation bronchoconstriction in vivo, and the effect in cardiovascular is replied as coronary artery contraction and many other biologicals.The pharmacology of peptide leukotriene comprises that smooth muscle contraction, cardiac muscle prevent, increase vascular permeability and increase mucous generation.

By comparison, LTB ₄Be by stimulating white corpuscle and lymphocyte function to produce its biological action.It stimulates polymorphonuclear leukocyte (PMN _S) chemotaxis, chemical agonism and gathering.

In fact strict, they are relevant with the disease of the numerous species that transmits cardiovascular, lung, skin, kidney, hypersensitive and inflammatory, and these diseases comprise asthma, grownup's respiratory distress syndrome, the fibrosis of bladder, psoriasis and enteritis.

Leukotriene B ₄(LTB ₄) describe in 1979 by Borgeat and Samuelsson at first, turn out to be 5(S by Corey and co-worker thereof later on), 12(R)-dihydroxyl-(Z, E, E, Z)-6,8,10,14-eicosane tetraenoic acid.

By LTA ₄The product of enzymatic hydrolysis gained be exactly the arachidonic acid series connection.Have been found that it can be by the mast cell, polymorphonuclear leukocyte, monocyte and scavenger cell produce.Shown LTB ₄Be the leukocytic effective stimulator of PMN in vivo, thereby cause increasing migration, adhesion, gathering, the threshing of chemotactic and chemical agonism, the formation of super-oxide and cytotoxicity.LTB ₄Effect be by have on the leukocyte surface height stereospecificity special receptor site transmit.Show for the pharmaceutical research of people's blood PMN white corpuscle and to have two class LTB ₄Specific receptors, they are that specific receptors with the peptide chemokine separates.Every receptoroid can be connected on the PMN leukocyte function base that separates class.The calcium transporting action comprises two kinds of mechanism.

LTB ₄Be proved to be the transmission matter of inflammation in the body.Its concentration increased in being found in the lung lavage with serious cacergastic people of lung, it also excessively replied relevant with the air flue of dog.

By antagonism LTB ₄Effect, or antagonism other be positioned at the effect that the pharmacologically active on the organ end transmits matter such as airway smooth muscle, compound of the present invention and pharmaceutical composition comprise that to treatment human or animal curee's disease is valuable, and leukotriene is a factor in these diseases.Some this compound can also suppress the 5-lipoxygenase or they are LTD ₄Antagonist.

Compound of the present invention or its pharmaceutically acceptable salt or N-oxide compound represented by logical formula I,

T is that acid amides connects base in the formula

Carbonyl carbon wherein is connected with pyridine ring;

R is C ₁To C ₂₀Aliphatic group, the phenyl C that does not replace or replace ₁To C ₁₀Aliphatic group, wherein substituted-phenyl has and one or morely is selected from lower alkoxy, low alkyl group, trihalomethyl group, or the group of halogen, and perhaps R is C ₁To C ₂₀Aliphatic series-O-, or R is the phenyl C that does not replace or replace ₁To C ₁₀Aliphatic series-O-, wherein substituted-phenyl has one or more lower alkoxies, low alkyl group, trihalomethyl group, or halogen group;

R ₁Be R ₄,-(C ₁To C ₅Aliphatic group) R ₄,-(C ₁To C ₅Aliphatic group) CHO ,-(C ₁To C ₅Aliphatic group) CH ₂OR ₈,-CH ₂OH or-CHO;

R ₂Be hydrogen, COR ₅, R wherein ₅For-OH, the base-OR of pharmaceutically acceptable formation ester ₆, or OX, wherein X is pharmaceutically acceptable positively charged ion, perhaps R ₅For-N(R ₇) ₂, R wherein ₇Be H, or the aliphatic group of 1 to 10 carbon atom, the cycloalkyl-(CH of 4 to 10 carbon atoms ₂) the n-base, wherein n is 0-3, perhaps two R ₇Connect together and form the ring with 4 to 6 carbon atoms, perhaps R ₂Be NHSO ₂R ₉, R wherein ₉For-CF ₃, C ₁To C ₆Alkyl or phenyl;

R ₃Be hydrogen, lower alkoxy, halogen ,-CH, COR ₅, or OH;

R ₄Be COR ₅, R wherein ₅For-OH, the base-OR of pharmaceutically acceptable formation ester ₆, or OX, wherein X is pharmaceutically acceptable positively charged ion, or R ₅For-N(R ₇) ₂, R wherein ₇Be H, the aliphatic group of 1 to 10 carbon atom, the cycloalkyl-(CH of 4 to 10 carbon atoms ₂) the n-base, wherein n is 0-3, perhaps two R ₇Connect together and form ring with 4 to 6 carbon atoms;

R ₈Be hydrogen, C ₁To C ₆Alkyl, or C ₁To C ₆Acyl group.

On the other hand, the present invention includes the pharmaceutical composition that contains these compounds and pharmaceutically acceptable vehicle.

Treatment and leukotriene be LTB particularly ₄Relevant or by they caused diseases, or transmit the matter diseases associated with the pharmacologically active that is in the organ end and be included within the scope of the invention.This treatment can be independent or carry out with generalformula that pharmaceutically acceptable vehicle combines by taking one or more.

Another aspect the present invention relates to prepare the method for generalformula, is illustrated among reaction scheme that this method provides below and the embodiment that lists.

Following definition is used to describe the present invention and states that at this contriver thinks their invention.

" aliphatic series " is meant and comprises saturated and undersaturated group.It comprises straight chain and side chain, saturated single or a plurality of unsaturated chain, and wherein two keys and triple bond can any combination exist.Term " low alkyl group " is meant with any isomeric form but the alkyl of 1 to 6 carbon atom of particularly positive structure or linear form." lower alkoxy " is meant low alkyl group-O-group." acyl group " is meant the group with terminal carbonyl carbon." halogen " is meant fluorine, chlorine, bromine or iodine.Phenyl ring can replace with one or more these groups.Multi-substituent can be identical or different, as three cl radicals are wherein arranged, or the combination of chlorine and alkyl, and this being combined in chlorine/alkyl mode of the latter can have different alkyl.

At R ₂And R ₃In term " base of pharmaceutically acceptable formation ester " comprise all esters that form by the acid functional group who is present in these compounds.Resulting ester is used for pharmaceutical applications for acceptable at it.Therefore, this means that list or diester will keep the biological activity of parent compound, and in its application and be used for the treatment of in the disease and will do not have unfavorable or deleterious effects.These esters be for example with following by-OR ₆Those esters that one of group of representative forms, wherein R ₆Be: C ₁To C ₁₀Alkyl, phenyl-C ₁-C ₆Alkyl, cycloalkyl, aryl, aralkyl, alkaryl, alkylaryl alkyl, aminoalkyl group, 2,3-indanyl, pivalyl oxygen methyl, acetyl-o-methyl, propionyl oxygen methyl, glycyl oxygen methyl, phenyl glycyl oxygen methyl, or thiophene glycyl oxygen methyl.Aryl comprises phenyl and naphthyl, or heteroaryl such as furyl, thienyl, imidazolyl, triazolyl or tetrazyl.The base of most preferred formation ester is R wherein ₆Be alkyl, especially the alkyl of 1 to 10 carbon (is CH ₃-(CH ₂) n-, n wherein is 0-9) or phenyl-(CH ₂) n-, wherein n is those groups of 0-4.

The pharmaceutically acceptable salt of these compounds is also included within the scope of the invention.These salt are acceptable in its pharmacy is used.Therefore this means that salt will keep the biological activity of parent compound, and in its application and be used for the treatment of in the disease and will do not have unfavorable or deleterious effects.

Pharmaceutically acceptable salt can prepare by general method.Under the situation of acid salt, be in suitable solvent, to make parent compound and excessive organic or inorganic acid-respons, or work as R with alkali part ₄During for OH and excessive organic or inorganic alkali reaction.Representative acid has spirit of salt, Hydrogen bromide, sulfuric acid, phosphoric acid, acetate, toxilic acid, succsinic acid or methylsulfonic acid.Cationic salts can be by alkali metal base, easily prepares as the alkali and the ammonia of sodium, potassium, calcium, magnesium, zinc, copper etc.Organic bases comprises that list or two replaces amine, quadrol, amino acid, trimethyl-xanthine, trometamol, trometamol compound, triethylamine, piperazine or the like.

The pyridine ring nitrogen oxide can prepare by methods known in the art and the illustrated method of this paper.These are considered to a part of the present invention.

If by substituent some be combined in and can produce chiral centre or other forms of isomery center in the The compounds of this invention, then all these isomeric forms include within the scope of the invention.Have the form administration that the compound of chiral centre can racemic mixture, or racemoid separated and use the discrete enantiomorph separately.

As leukotriene antagonist, these compounds can be used for treating particularly LTB of various and leukotriene ₄Diseases associated, or its cause or invasion and attack are by leukotriene LTB particularly ₄The disease that causes.Therefore, expect that these compounds can be used for treating anaphylactic disease, comprise lung and disease non-lung kind.For example, these compounds can be used for the anaphylaxis of antigen induction.They are used for the treatment of asthma and allergic rhinitis.These compounds also can be used to treat the disease of eye, as uveitis and anaphylaxis conjunctivitis.

R in the preferred compound of the present invention is an alkoxyl group, the particularly alkoxyl group of 8 to 15 carbon, or replacement or unsubstituted phenyl-C ₁To C ₁₀-aliphatic group-O-; R ₁For-(C ₁To C ₅Aliphatic group) R ₄Or-(C ₁To C ₅Aliphatic group) CH ₂OR ₃; And R ₂Be COOH or its an alkali metal salt or NHSO ₂R ₉, R wherein ₉For-CF ₃, C ₁To C ₆Alkyl or phenyl.The present invention more preferably R in the compound is the alkoxyl group of 8 to 15 carbon atoms or phenyl-C that alkoxyl group replaces ₁To C ₈Alkenyloxy or C ₁To C ₈Alkoxyl group; R ₁For-COR ₅,-CH ₂-CH ₂COR ₅Or-CH=CH-COR ₅; R ₂For-COOH or NHSO ₂R ₉, R wherein particularly ₉Be CF ₃; R ₂Be hydrogen or chlorine.

Most preferred is listed in the table II.

The table II

^*Carbonyl carbon is replaced on the pyridine ring.

^*Transconfiguration.

Synthetic

These compounds can be prepared by intermediate and the reagent as shown in following reaction scheme.This one group of specific intermediate is used for illustrating general method.Reaction scheme 1 has illustrated that preparation is used to prepare the method for R group compound.In order to prepare the R group, the preparation of the raw material that other diagrams are used is described in diagram 1, or derives from the intermediate of commercial source, and explanation prepares the method for generalformula then.

The R group of formula I is maybe can being prepared by one of two kinds of methods described in the reaction scheme I of having bought from the chemical supplier shop.

Reaction scheme I (a) has illustrated the method for preparing unsaturated phenyl-aliphatic R base.

Reaction scheme I (a)

Only pipe this explanation be the methoxyphenyl compound, but this series of steps and reagent can be used for preparing the phenyl-ω-aliphatic group by other represented replacements of R.Can buy on the starting raw material phenyl aldehyde market or available currently known methods easily is prepared.

For preparing acid (a), at first under inert atmosphere, alkyl sila trinitride is added in the inert solvent.Add microcosmic salt then.This doses under about room temperature and carries out.Through after the of short duration mixing, this mixture is generally suspension.Under about room temperature, slowly add phenyl aldehyde.Use the microcosmic salt of molar excess a little.Under about room temperature after additional of short duration stirring, water makes the reaction mixture quenching.Extract this acid with the solution acidifying and with appropriate organic solvent.Further use general separation and purification process as required.

By using the acid above the reductive agent reduction to make alcohol.Can use lithium aluminum hydride or similar reductive agent, reaction conditions can change according to carrying out the reductive needs.

In inert solvent, use Tosyl chloride and alkali such as pyridine to prepare p-toluenesulfonic esters.Appropriate condition is included in carries out this reaction 1 to 5 hour under about room temperature.Can prepare other suitable leavings groups that on function, are similar to p-toluenesulfonic esters, and as the method that R partly is added on the pyridine ring.

Reaction scheme I (b) has been summarized a kind of method of preparation alkoxyphenyl radical alkyl R group.This method can be used for preparing other R groups that phenyl is the ω group on the aliphatic chain, comprises the phenyl groups that contains of replacement.

Reaction scheme I (b)

ω-alkynes in these examples-1-alcohol is to can't buy on the market, and its preparation can be undertaken by handle this alcohol with highly basic by corresponding 3-alkynes-1-alcohol.Here use alkali metal amide.Then alcohol is protected so that required phenyl is added in and do not hold on the triple bond.In this example, form silyl ether; It has illustrated generalized case.The halogenophenyl adducts is used to phenyl is added on the triple bond.Here triple bond can be reduced, and optimum is to use catalytic method, as palladium/carbon and hydrogen.Another kind method is that triple bond is kept, and intermediate obtains p-toluenesulfonic esters according to illustrated reacting.The alcohol that silyl is removed and will be obtained is converted into p-toluenesulfonic esters or other group, and this group should have enough reactive behavioies, so that can form ether at an easy rate in these compounds synthetic afterwards.

In following reaction scheme, use the compound in the reaction scheme I, prepare and bought or with other compounds of currently known methods preparation, can prepare generalformula according to the reaction sequence of summarizing in the following reaction scheme.And these diagrams have illustrated the general method that how to prepare these compounds with concrete example.By changing or improving the chemical of explanation herein, these diagrams can be used for preparing other compound disclosed by the invention.This change or improvement will change reaction conditions, as temperature, pressure, reaction times, and reagent dosage, or the like.Reagent can be replaced by its Equivalent or similar agents, and this Equivalent or similar agents will produce product identical or of equal value.Similarly, also can change starting raw material and intermediate to adapt to the needs that prepare particular compound.

Listed a kind of method of preparation compound in reaction scheme 2, wherein the nitrogen of acid amides linking group is to be connected on the phenyl ring.

Reaction scheme 2

The above-mentioned a kind of synthetic route that illustrates the preparation generalformula, the carboxyl carbon in its formula I is to be connected on the pyridine ring.3-hydroxyl-2-(methylol) pyridine can have been bought on market maybe and can prepare with disclosed currently known methods.2-position methylol in this glycol is converted into aldehyde, is converted into the 3-alkoxy compound then, perhaps at first the 3-hydroxyl is converted into ether, then 2-position methylol is oxidized to aldehyde.Oxidation alcohol can use gentle oxygenant to carry out at an easy rate; Preferred Manganse Dioxide, but other oxygenants also can perform well in this step.Ether can be prepared at an easy rate by corresponding alpha-halogen-R group or the compound that resembles the p-toluenesulfonic esters class under alkaline condition.

Utilize suitable phosphoranyl inner salt ester being generally used under the condition of this reaction this 3-replacement-2-carboxyl aldehyde (c) being converted into 2-methoxycarbonyl vinyl form (d) then.Again the ester that obtains is prepared the N-oxide compound with peracid treatment, this N-oxide compound can be used for preparing pyridone (e).This step by-chloroperoxybenzoic acid is illustrated, but also can uses other similar oxygenants.Carry out the rearrangement of N-oxide compound with trifluoroacetic anhydride or similar agents afterwards, generate 2-pyridone (f).

Making the 2-pyridone be converted into acid amides is by acidylate 2-pyridone (g), in the presence of certain catalyzer and carbon monoxide, carries out with required Aminobenzoate (h) and this ester reaction then.With the trifluoromethanesulfanhydride anhydride is example explanation acidylate step.Amidate action is by at Pd(OAc) ₂, 1,1 '-two (diphenylphosphino) ferrocene exists and to be undertaken by the trifluoromethanesulfonic acid ester solution with the carbon monoxide bubbling down.Then the diester (ⅰ) that obtains is made hydrolysis of ester group with the mineral alkali saponification.In and the salt of gained so that obtain free acid.Can free acid be converted into another kind of ester or be prepared into corresponding amide with currently known methods.

Saturated 3-bit substituent can be prepared by catalytic hydrogenation at an easy rate by the alkene analogue.Reaction scheme 3 has illustrated this method.

Reaction scheme 3

In the catalytic reduction reaction of classics, utilize heavy metal catalyst and hydrogen that the diester catalytic reduction is (3a).After reduction reaction was finished, diacid (3b) can use alkali to make the diester hydrolysis if desired.By suitable oxidation, reduction, esterification, amidate action or by additive method, each compound all can be converted into other compounds of the present invention.

The carbon analogue of these compounds, promptly wherein connecting the R group is that the compound of methylene radical can being prepared according to the step of listing in the 4th schema in proper order to the atom on the 3-position.

Reaction scheme 4

With corresponding alkanol and acid catalyst the 3-hydroxy-picolinic acid is converted into alkyl ester.With trifluoromethanesulfanhydride anhydride and pyridine hydroxyl is converted into triflate (4a).Connect broadtail (4b) with suitable alkyl catechol boride then, used alkyl catechol boride is with palladium coupling condition ((Pd(OAc) ₂)) prepare by 1-tridecylene and catecholborane.With suitable hydride, alkyl ester is converted into corresponding aldehyde again as diisobutylaluminium hydride.Make this aldehyde carry out Wei Tixi (Wittig) olefination again, for example use (triphenylphosphine inner salt) methyl acetate.Afterwards, the identical series of steps with general introduction in the reaction scheme 2 in the above is converted into the purpose compound with the pyridyl acrylate that obtains.

Reverse acid amides can being prepared according to the step that provides in the reaction scheme 5 in proper order.

Reaction scheme 5

With acid catalyst and corresponding alkanol commercially available 3-hydroxy-picolinic acid is converted into alkyl ester.Then under general condition, carry out alkylated reaction with for example 1-iodo dodecane or similar 1-halogenated compound.Weak base such as the K that is in the dimethyl formamide preferably used in this reaction ₂CO ₃Carry out.Generate the 3-alkoxy derivative.The nitrogen of pyridine oxide is also reset the N-oxide compound of gained, generates the 2-pyridone.With peracid such as 3-chloroperoxybenzoic acid or similarly oxygenant can carry out oxidation at an easy rate.Can use the trifluoroacetic anhydride in suitable solvent such as dimethyl formamide to carry out the rearrangement of N-oxide compound (5a).

Forming triflate utilizes trifluoromethanesulfanhydride anhydride and alkali such as pyridine to carry out.The nucleophilic substitution reaction that carries out with sodiumazide generates 2-azido-pyridine derivate (5b).By catalytic hydrogenation trinitride is reduced to amine.With hydride such as diisobutylaluminium hydride reduction alkyl ester is aldehyde.Use Wei Tixi (Wittig) prepared in reaction 2-aminopyridine acrylate (5c) then.For example, can use (triphenylphosphine inner salt) methyl acetate.Acylated amine (methyl isophthalyl chloride) then uses alkali (LiOH, tetrahydrofuran (THF), methyl alcohol) with the ester hydrolysis, obtains the purpose acid amides.The common known method of method or this area with this paper explanation can further be converted into ester with these compounds, acid amides, salt or the similar compounds that is defined by formula I.

Preparation

Pharmaceutical composition of the present invention comprises pharmaceutical carrier or thinner and a certain amount of logical formula I compound.This compound exists carrying out the amount of physiological response, perhaps exists with less amount, makes the user need take two or the treatment expected of the composition of a plurality of units like this.These compositions can be formulated as solid, liquid or gaseous form.Perhaps when administration, can change one of these three kinds of forms into another kind of form, as when solid is provided by the aerosol mode, perhaps when liquid is provided with spraying or aerosol form.

The carrier of composition and medicine or the character of thinner depend on the route of administration of expection certainly, as method parenteral, partial, oral or by sucking.

For administered parenterally, the form of pharmaceutical composition is an aseptic parenteral solution, as the aqueous or anhydrous liquid suspension of ampoule.

For topical, the form of pharmaceutical composition is emulsifiable paste, ointment, liniment, lotion, paste and the drops that is suitable for eye, ear or nasal administration.

For oral administration, the form of pharmaceutical composition is tablet, capsule, pulvis, pill, atroche, lozenge, syrup, liquid or emulsion.

When pharmaceutical composition used with solution or form of suspension, the example of suitable pharmaceutical carrier or thinner comprises: for Aquo System was water; For non-aqueous system is the mixture of ethanol, glycerine, propylene glycol, Semen Maydis oil, oleum gossypii seminis, peanut oil, sesame oil, white oil and they and water; For solid system is lactose, kaolin and mannitol; For the aerosol system is Refrigerant 12, chloro-trifluoro-ethane and compressed carbon dioxide.Except pharmaceutical carrier or thinner, composition of the present invention can also comprise other components, as stablizer, antioxidant, sanitas, lubricant, suspension agent, viscosity modifier, or the like, as long as additional component does not have injurious effects for the therapeutic action of the present composition.

Described pharmaceutical preparation can be prepared according to the method that the pharmacist who is suitable for required purpose product uses always.

Usually, particularly for the prophylactic treatment of asthma, the suitable form of medication of composition is an inhalation.Therefore for utilizing general atomizer administration, composition comprises the aqeous suspension or the aqueous solution of active constituent.During by pressurization aerosol container administration, composition is included in the general liquefied propellant or the suspension or the solution of the active constituent in the pressurized gas with other method.For by the powder inhalator administration, composition also comprises the solid active component with the solid diluent dilution.In the superincumbent composition, the amount of carrier or thinner can change, but preferably it is the bigger part of the suspension or the solution of active constituent.When thinner is solid, the amount that its exists can less than, be equal to or greater than the amount of solid active component.

Usually, with generalformula with the form administration of composition in the curee, said composition comprises the generalformula of nontoxic amount, it is enough to produce inhibition is the symptom of the disease of the factor with the leukotriene.When using by this way, the dosage of composition is selected from the scope of each administration 50mg to 100mg active constituent.For convenience's sake, with the dosed administrations such as per daily dose every day of regulation 1 to 5 time, the per daily dose of defined is selected from extremely about 5000mg of about 50mg.

Comprise also within the scope of disclosure that treatment is by LTB ₄The method of the disease of transmitting, this method comprise the generalformula of taking the treatment significant quantity to the curee, preferably with the form administration of pharmaceutical composition.For example, suppress by the symptom that discharges the transmission anaphylaxis that matter produced by the generalformula of taking significant quantity.This is included within the scope of disclosure.Can under suitable interval, carry out administration as required with dose unit or with single dose.Usually, when needing mitigation symptoms especially, will implement this method.Yet this method also is used for carrying out continuously or prophylactic treatment.In the technical scope of this area,, list the effective dose that dosage range can be determined administration, will consider severity or the like the factor of the disease of being treated simultaneously by top by routine test.

Pharmaceutical composition and using method thereof also comprise generalformula and H ₁The combination of blocker, the amount of two kinds of compounds that comprise in this combination are enough to treat the respiratory hypersensitivity reaction or the similarly anaphylaxis of antigen induction.At this useful representative H ₁Blocker comprises: Sodium Cromoglicate, alcohol amine compound (diphenhydramine), ethylenediamines (pyrilamine), alkyl amine (Chlophenamin), piperazines (chloreyclizine), and phenothiazines (promethazine).Useful especially in the present invention H ₁Blocker is 2-(4-(5-bromo-3-picoline-2-yl) fourth amino)-5-((6-picoline-3-yl) methyl)-4-pyrimidone.

Biological test

The specificity of the antagonistic activity of chemical compound lot of the present invention is for resembling Repone K, carbachol, histamine and PGF by them ₂The antagonistic action than low degree of this class gaonist is confirmed.

The receptors bind avidity that is used for the compound of the inventive method be on the U937 cytolemma that is attached to the people by these compounds ( ³H)-LTB ₄Ability on the combining site is measured.The LTB that is used for the compound of the inventive method ₄Antagonistic activity is with the relevant mode antagonism LTB of dosage by them ₄Can bring out that the ability of calcium transition state measures, be that the calcium sample of fluorescence is measured the calcium transition state with fura-2.Used method is as follows:

The U937 cell culture condition

The U937 cell is from Dr.John Bomalaski(Medical College of PA) and Dr.John Lee(Smithkline Beecham, Dept.of Immunology) that obtains, adding 10%(V/V) in the RPMI-1640 medium of foetal calf serum of the deactivation of heat, containing 5% CO ₂, 95% air is cultivated above-mentioned cell in 37 ℃ the moist environment.Cell not only can but also can have been cultivated in the turner culture in T type flask.In order to distinguish U937 cell and mononuclear type cell with DMSO, in containing the above-mentioned medium of 1.3%DMSO with 1 * 10 ⁵The concentration of individual cell/ml is cell inoculation, and cultured continuously 4 days.Cell density is generally 0.75-1.25 * 10 ⁶Individual cell/ml, by under 800Xg centrifugal 10 minutes with its collection.

The preparation of U937 cytolemma enrichment composition

With the U937 cell collected 25 ℃ with 50mM Tris-HCl washing, this washings pH is 7.4 and contains 1mM EDTA(buffer A).With cell with 5 * 10 ⁷The concentration resuspending of individual cell/ml and makes cell cracked by the nitrogen air pocket with 750psi with the Pa Er bomb under 0 ℃ in buffer A, continues 10 minutes.Broken cell preparation under 1000Xg centrifugal 10 minutes.With supernatant liquor 50, under the 000Xg centrifugal 30 minutes.Gained particle buffer A washed twice.Concentration with about 3mg membranin/ml is 7.4 at 25 ℃ with 50mM Tris HCl(pH) with the particle resuspending, and make the aliquots containig quick freezing, in-70 ℃ of storages down.

( ³H)-LTB ₄Be incorporated into the U937 membrane receptor

( ³H)-LTB ₄In conjunction with test is at 25 ℃, is containing 10mMCaCl ₂, 10mM MgCl ₂, ( ³H)-LTB ₄, the 50mM Tris-HCl(pH 7.5 of U937 epicyte protein) and in the damping fluid (standard conditions), be with or without the LTB of different concns ₄Or SK ﹠amp; Carry out under the existence of F compound.The mean value of the mensuration of triplicate is represented in each test point.Not or the unlabelled LTB of 2 μ M arranged ₄Exist down, measured respectively ( ³H)-LTB ₄Total and non-specific binding.Calculate specificity with difference non-specific binding and combine according to total.What radioligand was striven test unexpectedly is under standard conditions, with about 0.2 μ M ( ³H)-LTB ₄, the U937 epicyte protein of 20-40 μ g increases LTB in the 0.2ml reaction volume ₄Concentration (0.1nM to 10nM) or other concentration of striving property part (0.1 μ M to 30 μ M) unexpectedly, and cultivated 30 minutes down in 25 ℃.By the vacuum filtration method unconjugated radioligand and competitive drug are separated with the film binding partner.Measure on the filter film in conjunction with radioactivity with the liquid-scintillation spectrometry method.

The saturated of U937 cell is under standard conditions in conjunction with test, with about 15-50 μ g U937 membranin, and in the 0.2ml reaction volume, increase ( ³H)-LTB ₄Concentration (0.02 to 2.0mM), and cultivate at 22 ℃ and to carry out in 30 minutes.LTB ₄(2 μ M) is placed in the independent cover culture test tube so that measure non-specific binding.By the saturated computer assisted nonlinear least square method curve fitting analysis of data process, and further use the Scatchard methods analyst in conjunction with the test gained.

Carry out the picked-up of Fura-2 with the U937 cell that is broken up

The cell of collecting is with 2 * 10 ⁶The concentration resuspending of individual cell/ml is in Krebs Ringer Hensilet damping fluid, and this damping fluid contains the 0.1%BSA(RIA level), 1.1mM MgSO ₄, 1.0mM CaCl ₂With 5mM HEPES(pH7.4, buffer B).With fura-2(fura-2/AM) diacetyl methoxyl group ester to add to ultimate density be 2nM, in the dark under 37 ℃ with cell cultures 30 minutes.Under 800Xg with cell centrifugation 10 minutes, and with 2 * 10 ⁶The concentration of individual cell/ml in fresh buffer B, is cultivated its resuspending 20 minutes at 37 ℃, makes remaining ester complete hydrolysis.Under 800Xg with cell centrifugation 10 minutes, and with 5 * 10 ⁶The concentration of individual cell/ml with its resuspending in cold fresh buffer B.Cell is held in the dark place on ice, until the mensuration that is used for fluorescence.

The mensuration calcium transporting action of fluorescence

Use the fluorescence of measuring the U937 cell that contains fura-2 by the photofluorometer of Johnson Foundation Biomedical Instrumentation Group design.Photofluorometer is equipped with temperature controller and magnetic stirrer under the cuvette supporting apparatus.Excitation wavelength is positioned at 339nm, and emission wavelength is positioned at 499nm.All have a try to test all and under 37 ℃, carry out with constant agitation.

It is 1 * 10 that the U937 cell is diluted to concentration with fresh damping fluid ⁶Individual cell/ml remains on the dark place on ice.The aliquots containig (2ml) of cell suspending liquid is put into the 4ml cuvette, and temperature rises to 37 ℃, (remaining on 37 ℃, water-bath 10 minutes).Cuvette is moved in the photofluorometer, measured about 1 minute of fluorescence before adding stimulant or antagonist, then measured fluorescence about 2 minutes behind the adding stimulant, gaonist and antagonistic all add with 2 μ l aliquots containigs.

Earlier antagonist is added in the cell in the photofluorometer, so that measure the potential stimulant activity.Add 10nM LTB after about then 1 minute ₄(near maximum effective concentration), maximum Ca ²⁺Migration concentration (Ca ²⁺) _i; Calculate with following formula:

F is that the relative fluorescence of the maximum of sample is measured.Fmax is to be 0.02% by the 10%Triton X-100(ultimate density with 10 μ l) dissolved cell measures.Measured after the Fmax, 67 μ l 100mM EDTA solution (pH10) have been added to the Ca of complete chelating ²⁺In, make the fura-2 blackout, and obtain Fmin.Not having under the situation of antagonist, for 10nM LTB ₄(Ca ²+) _iConcentration is 100%, minimum (Ca ²⁺) _iBe 0%.IC ₅₀Concentration is the 10nM LTB of blocking-up 50% ₄Inductive (Ca ²⁺) _iThe antagonist concentration of transporting action.For LTB ₄Inductive (Ca ²⁺) _iThe EC of the increase of transporting action ₅₀Concentration partly for the maximum increase.Ki for the calcium transporting action determines with following formula:

$K_i=\frac{{\mathrm{IC}}_{50}}{1+\frac{\left[{\mathrm{LTB}}_4\right]}{\left[{\mathrm{EC}}_{50}\right]}}$

For described test, LTB ₄Concentration is 10nM, EC ₅₀Be 2nM.

The result of compound that will be by these method tests lists in the table III.

The table III

In conjunction with IC ₅₀, (Ki), μ M Ca-transporting action

U-937 PMN U-937 PMN

Complete complete IC ₅₀The stimulant stimulant

Constitute theca cell cell μ M % %

Ex1 4.0（1.4） 2.0 2.4 3.7 0 0

Ex2 23（8.0） 4.7 - 3.0 0 0

Ex3 47（17） 5.8 0.65 0.58 0 0

Ex4 6.5（2.2） 3.4 2.2 8.5 0 0

Ex5 41（14） 1.1 2.2 0.72 0 0

Ex6 ^＊6.1（2.0） 0.68 0.14 0.74 0 0

^*Title compound

Embodiment

How embodiment given below explanation prepares and uses compound of the present invention.These embodiment are example, and do not limit or limit the scope of the invention.Made the foundation that keeps the claim of giving the contriver in order to limit by this document.

Embodiment A

The 8-(4-p-methoxy-phenyl) 7-octyne-1-alcohol A(1 octane-1-(4-tosylate))

The mineral oil of 35%KH (27g, 240mmol) solution is used hexane wash under argon atmospher, and with 1, the 3-diaminopropanes is drop by drop handled.Mixture at room temperature stirs until evenly.Flask is cooled to 0 ℃, adds 3-octyne-1-alcohol (10g, 79mmol, Lancaster Synthesis) lentamente.Then reaction mixture was at room temperature stirred 18 hours.Reaction mixture water (50ml) quenching, the product extracted with diethyl ether.Organic layer is with 10%HCl(3 * 15ml) and salt water washing, and dry (MgSO ₄).Obtain title product through evaporation, no longer purify and just can use. ¹H NMR(90MHz, CDCl ₃) δ 3.65(t, J=5Hz, 2H, OCH ₂), 2.23(m, 2H, CH ₂), 2.0(m, 1H, alkynyl), 1.7-1.2(m, 8H, (CH ₂) ₄); The pure sample of 1R() ν max 3350,2930,2125cm ^-1

A(2) 7-octyne-1-t-butyldiphenylsilyl ether

With 7-octyne-1-alcohol (3.8g 30mmol) is dissolved in the dimethyl formamide (10ml), 0 ℃ with the tertiary butyl chloride diphenyl silane (10.2ml, 33mmol) and imidazoles (3.65g 45mmol) handles.Reaction mixture stirred 10 minutes and stirring at room 3 hours at 0 ℃.Add entry, and use the ethyl acetate extraction product.Acetic acid ethyl acetate extract water and salt water washing, and dry (Ma ₂SO ₄).Evaporate solvent, residuum is purified with flash column chromatography (silicon-dioxide, hexane), obtains yellow oil.

¹H NMR(250MHz, CDCl ₃) δ 7.7(d, 4H, aryl), 7.4(m, 6H, aryl), 3.63(t, 2H, OCH ₂), 2.23(m, 2H, CH ₂), 1.97(t, 1H, alkynyl), 1.6-1.3(m, 8H, (CH ₂) ₄), 1.05(s, 9H, the tertiary butyl); The 1R(film) ν max 3321,2940,2125 cm ^-1

A(3) 8-(4-p-methoxy-phenyl)-7-octyne-1-t-butyldiphenylsilyl ether

Under argon atmospher, in the dry flask of crossing of flame, add 4-iodo phenylmethylether (5.34g, triethylamine 22mmol) (50ml) solution, then add 7-octyne-1-t-butyldiphenylsilyl ether (9.84g, 27mmol), (Ph ₃P) ₂PdCl ₂(350mg, 0.44mmol), and CuI(200mg, 0.88mmol).The mixture that obtains was 50 ℃ of heating 4 hours.When cool to room temperature, filter reaction mixture, and evaporating solvent.Residuum distributes between ethyl acetate and water, and collected organic layer is used the salt water washing, and dry (Na ₂SO ₄).Evaporating solvent, residuum is purified with flash column chromatography (silicon-dioxide, the hexane solution of 1% ethyl acetate), obtains oily matter.

¹H NMR(250MHz, CDCl ₃) δ 7.7(d, 4H, aryl), 7.4(m, 6H, aryl), 7.35(d, 2H, aryl) and, 6.8(d, 2H, aryl), 3.8(s, 3H, OCH ₃), 3.7(t, 2H, OCH ₂), 2.4(t, 2H, CH ₂), 1.7-1.3(m, 8H, (CH ₂) ₄), 1.05(s, 9H, the tertiary butyl).

A(4) octane-1-t-butyldiphenylsilyl ether 8-(4-p-methoxy-phenyl)

With 5% Pd/C(100mg) adding 8-(4-p-methoxy-phenyl)-(2.2g is in ethanol 4.6mmol) (10ml) and ethyl acetate (10ml) solution for 7-octyne-1-t-butyldiphenylsilyl ether.Mixture hydrogenation 4 hours under 75PSi.Reaction mixture filters by diatomite (Celite), and evaporating solvent obtains oily matter.

¹H NMR(250MHz, CDCl ₃) δ 7.7(d, 4H, aryl, 7.4(m, 6H, aryl), 7.05(d, 2H, aryl), 6.8(d, 2H, aryl) and, 3.8(s, 3H, OCH ₃), 3.6(t, 2H, OCH ₂), 2.5(t, 2H, benzyl), 1.75-1.3(m, 12H, (CH ₂) ₆), 1.0(s, 9H, the tertiary butyl).

A(5) octane-1-alcohol 8-(4-p-methoxy-phenyl)

With the 8(4-p-methoxy-phenyl) (2.2g, tetrahydrofuran (THF) 4.6mmol) (20ml) solution is cooled to 0 ℃ to octane-1-t-butyldiphenylsilyl ether, handles with tetrabutylammonium (14ml, 14mmol, the tetrahydrofuran solution of 1M).Remove cooling bath, reaction mixture at room temperature stirred 24 hours.Reaction mixture dilutes with ethyl acetate, water and salt water washing, and dry (Na ₂SO ₄).Evaporating solvent, residuum is purified with flash column chromatography (silicon-dioxide, the hexane solution of 0-20% ethyl acetate), obtains white solid.Fusing point 47-49 ℃; ¹H NMR(250MHz, CDCl ₃) δ 7.15(d, 2H, aryl), 6.86(d, 2H, aryl), 3.85(s, 3H, OCH ₃), 3.68(t, 2H, OCH ₂), 2.62(t, 2H, benzyl), 1.75-1.3(m, 12H, (CH ₂) ₆).

A(6) octane-1-(4-tosylate 8-(4-p-methoxy-phenyl))

Under argon atmospher with the 6-(4-p-methoxy-phenyl) (5.9g 25mmol) is dissolved in anhydrous CH to octane-1-alcohol ₂Cl ₂(100ml), be cooled to 0 ℃.To wherein add pyridine (2.5ml, 30mmol) and the 4-toluene sulfonyl chloride (5.4g, 28mmol).Reaction mixture stirred 20 minutes at 0 ℃, stirring at room 24 hours, and reaction soln water and salt water washing, and dry (Na ₂SO ₄).Evaporating solvent, residuum is purified with flash column chromatography (silicon-dioxide, the hexane solution of 0-10% ethyl acetate), obtains white solid. ¹H NMR(250MHz, CDCl ₃) δ 7.79(d, 2H, aryl), 7.35(d, 2H, aryl), 7.09(d, 2H, aryl) and, 6.82(d, 2H, aryl), 4.04(s, 2H, OCH ₂), 3.8(s, 3H, OCH ₃), 2.55(t, 2H, benzyl), 2.46(s, 3H, CH ₃), 1.75-1.15(m, 12H, (CH ₂) ₆).

Embodiment B

The 6-(4-p-methoxy-phenyl)-and hexane-1-(4-tosylate) B(1) 5-hexin-1-t-butyldiphenylsilyl ether

With 5-hexin-1-alcohol (3g, 30mmol Aldrich) are dissolved in the dimethyl formamide (10ml), 0 ℃ with the tertiary butyl chloride diphenyl silane (10.2ml, 33mmol) and imidazoles (3.65g 45mmol) handles.Reaction mixture stirred 10 minutes at 0 ℃, stirring at room 3 hours.Add entry, the product ethyl acetate extraction.Acetic acid ethyl acetate extract water and salt water washing, and dry (Na ₂SO ₄).Evaporating solvent, residuum is purified with flash column chromatography (silicon-dioxide, hexane), obtains yellow oil. ¹H NMR(250MHz, CDCl ₃) δ 7.7(d, 4H, aryl), 7.4(m, 6H, aryl), 3.65(t, 2H, OCH ₂), 2.2(m, 2H, CH ₂), 1.9(t, 1H, alkynyl), 1.7(m, 4H, CH ₂-CH ₂), 1.05(s, 9H, the tertiary butyl).

B(2) 6-(4-p-methoxy-phenyl)-5-hexin-1-t-butyldiphenylsilyl ether

Under argon atmospher, in the dry flask of crossing of flame, add 4-iodo phenylmethylether (5.34g, triethylamine 22mmol) (50ml) solution, then add 5-hexin-1-t-butyldiphenylsilyl ether (8.83g, 27mmol), (Ph ₃P) ₂PdCl ₂(350mg, 0.44mmol), and CuI(200mg, 0.88mmol).The mixture that obtains was 50 ℃ of heating 4 hours.Filter reaction mixture when being cooled to room temperature, evaporating solvent.Residuum distributes between ethyl acetate and water, and collected organic layer is used the salt water washing, and dry (Na ₂SO ₄).Evaporating solvent, residuum is purified with flash column chromatography (silicon-dioxide, the hexane solution of 1% ethyl acetate), obtains oily matter. ¹H NMR(250MHz, CDCl ₃) δ 7.7(d, 4H, aryl), 7.4(m, 6H, aryl), 7.35(d, 2H, aryl) and, 6.8(d, 2H, aryl), 3.8(s, 3H, OCH ₃), 3.7(t, 2H, OCH ₂), 2.4(t, 2H, CH ₂), 1.7(m, 4H, CH ₂-CH ₂), 1.05(s, 9H, the tertiary butyl).

B(3) hexane-1-t-butyldiphenylsilyl ether 6-(4-p-methoxy-phenyl)

With 5%Pd/C(100mg) adding 6-(4-p-methoxy-phenyl)-(2.0g is in ethanol 4.6mmol) (10ml) and ethyl acetate (10ml) solution for 5-hexin-1-t-butyldiphenylsilyl ether.Mixture hydrogenation 4 hours under 75psi.Reaction mixture filters by diatomite, and evaporating solvent obtains oily matter. ¹H NMR(250MHz, CDCl ₃) δ 7.7(d, 4H, aryl), 7.4(m, 6H, aryl), 7.05(d, 2H, aryl) and, 6.8(d, 2H, aryl), 3.8(s, 3H, OCH ₃), 3.6(t, 2H, OCH ₂), 2.5(t, 2H, benzyl), 1.55(m, 4H, CH ₂-CH ₂), 1.3(m, 4H, CH ₂-CH ₂), 1.0(s, 9H, the tertiary butyl).

B(4) hexane-1-alcohol 6-(4-p-methoxy-phenyl)

With the 6-(4-p-methoxy-phenyl) (2.0g, tetrahydrofuran (THF) 4.6mmol) (20ml) solution is cooled to 0 ℃ to hexane-1-t-butyldiphenylsilyl ether, handles with tetrabutylammonium (14ml, 14mmol, the tetrahydrofuran solution of 1M).Remove cooling bath, reaction mixture was stirring at room 24 hours.Reaction mixture dilutes with ethyl acetate, uses H ₂O and salt water washing, and dry (Na ₂SO ₄).Evaporating solvent, residuum is purified with flash column chromatography (silicon-dioxide, the hexane solution of 0-20% ethyl acetate), obtains white solid. ¹H NMR(250MHz, CDCl ₃) δ 7.05(d, 2H, aryl), 6.8(d, 2H, aryl), 3.8(s, 3H, OCH ₃), 3.65(t, 2H, OCH ₂), 2.55(t, 2H, benzyl), 1.6(m, 4H, CH ₂-CH ₂), 1.4(m, 4H, CH ₂-CH ₂).

B(5) hexane-1-(4-tosylate 6-(4-p-methoxy-phenyl))

Under argon atmospher, with the 6-(4-p-methoxy-phenyl) (5.36g 25mmol) is dissolved in anhydrous CH to hexane-1-alcohol ₂Cl ₂(100ml), be cooled to 0 ℃.To wherein add pyridine (2.5ml, 30mmol) and the 4-toluene sulfonyl chloride (5.4g, 28mmol).Reaction mixture stirred 20 minutes at 0 ℃, stirring at room 24 hours.Reaction soln H ₂O and salt water washing, and dry (Na ₂SO ₄).Evaporating solvent, residuum is purified with flash column chromatography (silicon-dioxide, the hexane solution of 0-10% ethyl acetate), obtains white solid. ¹H NMR(250MHz, CDCl ₃) δ 1.6-1.3(m, 8H, (CH ₂) ₄), 2.4(s, 3H, CH ₃), 2.5(t, 2H, benzyl), 3.8(s, 3H, OCH ₃), 4.0(t, 2H, OCH ₂), 6.80(d, 2H, aryl), 7.0(d, 2H, aryl), 7.3(d, 2H, aryl) and, 7.8(d, 2H, aryl).

Embodiment C

The E-6-(4-p-methoxy-phenyl)-the 1-(4-tosylate)-the 5-hexene

C(1) E-6-(4-p-methoxy-phenyl)-the 5-hexenoic acid

Under room temperature and argon atmospher, (17.6g, 30mmol) suspension in tetrahydrofuran (THF) (45ml) adds in freshly prepd hexamethyl two silicon Lithium Azides (Iithium hexamethyldisilazide) tetrahydrofuran (THF) (30ml) solution (64mmol) with bromination (4-carboxybutyl) triphenyl phosphonium.Reaction mixture stirred 15 minutes, formed the orange inner salt during this period.(4.5g, tetrahydrofuran (THF) 30mmol) (30ml) solution continue to stir 20 minutes more dropwise to add the 4-aubepine.Reaction mixture H ₂O(50ml) quenching is with ether (30ml) dilution.Water layer is acidified to pH1.0 with 3N HCl.(3 * 50ml) extract product with ethyl acetate.The organic layer that merges is carried out drying (MgSO ₄), with flash column chromatography (silicon-dioxide, the CH of 1% methyl alcohol ₂Cl ₂Solution) purify, obtain the E-alkene, solid. ¹H NMR(200MHz, CDCl ₃) δ 7.3(d, 2H, aryl), 6.8(d, 2H, aryl), 6.3(d, 1H, alkene) and, 6.0(m, 1H, alkene), 3.8(s, 3H, OCH ₃), 2.3(m, 4H, allylic CH ₂And CH ₂CO ₂), 1.8(q, 2H, CH ₂).

C(2) E-6-(4-p-methoxy-phenyl)-the 5-hexen-1-ol

Under argon atmospher, with the E-6-(4-p-methoxy-phenyl)-(1.1g, anhydrous diethyl ether 5.0mmol) (10ml) solution adds LiAlH to the 5-hexenoic acid lentamente ₄(240mg is 6.0mmol) in the suspension in ether (10ml).Reaction mixture refluxed 45 minutes.When cool to room temperature, use H ₂O(10ml) 6N H is then used in quenching ₂SO ₄(7ml) quenching.Add ethyl acetate (20ml), separate organic layer, and dry (MgSO ₄).Obtain white crystalline solid after the evaporation.Fusing point 65-66 ℃; ¹H NMR(200MHz, CDCl ₃) δ 7.2(d, 2H, aryl), 6.8(d, 2H, aryl), 6.3(d, 1H, alkene) and, 6.1(m, 1H, alkene), 3.8(s, 3H, OCH ₃), 3.6(t, 2H, OCH ₂), 2.2(q, 2H, allyl group), 1.5(m, 4H, CH ₂-CH ₂); Ultimate analysis, calculated value (C ₁₃H ₁₈O ₂): C, 75.65; H, 8.80, experimental value: C, 75.45; H, 8.95; MS(CI): 207(M+H).

C(3) E-6-(4-p-methoxy-phenyl)-the 1-(4-tosylate)-the 5-hexene

Under argon atmospher, with the E-6-(4-p-methoxy-phenyl)-(1.6g 7.0mmol) is dissolved in anhydrous CH to the 5-hexen-1-ol ₂Cl ₂(50ml), with the 4-toluene sulfonyl chloride (7.0g, 36mmol) and pyridine (3ml) processing.Reaction soln at room temperature stirred 3.5 hours.In reaction mixture, add entry (40ml), separate organic layer, and dry (MgSO ₄).Product is purified with flash column chromatography (silicon-dioxide, the hexane solution of 10% ethyl acetate), obtains oily matter. ¹H NMR(200MHz, CDCl ₃) δ 7.8(d, 2H, aryl), 7.3(d, 2H, aryl), 7.2(d, 2H, aryl) and, 6.8(d, 2H, aryl), 6.2(d, 1H, alkene) and, 6.0(m, 1H, alkene), 4.1(t, 2H, OCH ₂), 3.8(s, 3H, OCH ₃), 2.4(s, 3H, CH ₃), 2.1(q, 2H, allyl group), 1.6(m, 4H, CH ₂-CH ₂); MS(CI): 361(M+H).

Embodiment 1

The N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide disodium salt

1(a) 3-oxygen in last of the ten Heavenly stems base-2-(methylol) pyridine

With 3-hydroxyl-2-(methylol) pyridine hydrochloride (500mg, 3.09mmol, Aldrich, 85%) is dissolved in the anhydrous dimethyl formamide (10ml), sequentially uses anhydrous K ₂CO ₃(1.30g, 9.27mmol) (0.80ml 3.71mmol) handles with 1-iodo decane.Under argon atmospher in 90 ℃ with reaction mixture high degree of agitation 1.5 hours.When cool to room temperature, reaction mixture is used H with ethyl acetate (100ml) dilution ₂O(5 * 20ml) and salt water washing, and dry (MgSO ₄).The compound that obtains is purified with flash column chromatography (silicon-dioxide, the petroleum ether solution of 20% ethyl acetate), obtains title product. ¹H NMR(250MHz, CDCl ₃) δ 8.17(m, 1H, 6-pyridyl), 7.2(m, 2H, 4-pyridyl, 5-pyridyl), 4.78(s, 2H, CH ₂), 4.48(is wide unimodal, 1H, OH), 4.05(t, J=6.6Hz, 2H, OCH ₂), 1.9-0.90(m, 19H, aliphatic series).

1(b) 3-oxygen in last of the ten Heavenly stems base-2-pyridylaldehyde

Will be by 1(a) 3-oxygen in last of the ten Heavenly stems base-2-(methylol of obtaining) pyridine (560mg, anhydrous CH 2.11mmol) ₂Cl ₂(7ml) solution MnO ₂(1.80g 20.7mmol) handles, and at room temperature stirs 24 hours.Reaction mixture filters by diatomaceous earth filler, uses solvent removed in vacuo, obtains aldehyde, is light yellow oil.This aldehyde is directly used in next step without further purifying.

1(c) 2-(E-2-carboxymethyl vinyl)-3-oxygen in last of the ten Heavenly stems yl pyridines

Under argon atmospher, (429mg 1.63mmol) is dissolved in the dry toluene (3.5ml) 3-oxygen in last of the ten Heavenly stems base-2-pyridylaldehyde that previous step is obtained, and (820mg 2.45mmol) handles with triphenylphosphine inner salt methyl acetate.Reaction mixture was 45 ℃ of heating 30 minutes, and reaction mixture becomes evenly under this temperature.When cool to room temperature, reaction mixture is used H with ethyl acetate (100ml) dilution ₂O(2 * 20ml) and salt water washing, and dry (MgSO ₄).Product flash column chromatography (silicon-dioxide, ethyl acetate: CH ₂Cl ₂: sherwood oil=10: 5: 85) purify, obtain product, be light yellow solid. ¹H NMR(200MHz, CDCl ₃) δ 8.25(m, 1H, 6-pyridyl), 8.1(d, J=16.2Hz, 1H, alkene), 7.25(m, 2H, 4-pyridyl, 5-pyridyl) and, 7.05(d, J=16.2Hz, 1H, alkene), 4.05(t, J=6.6Hz, 2H, OCH ₂), 3.85(s, 3H, CO ₂CH ₃), 1.95-0.90(m, 19H, aliphatic series).

1(d) 2-(E-2-carboxymethyl vinyl)-3-oxygen in last of the ten Heavenly stems yl pyridines N-oxide compound

Under argon atmospher, (390mg 1.22mmol) is dissolved in anhydrous CH with 2-(E-2-carboxymethyl vinyl)-3-oxygen in last of the ten Heavenly stems yl pyridines ₂Cl ₂(6mlml), be cooled to 0 ℃, (278mg 1.34mmol) handles with 85%3-chlorine peroxybenzoic acid.Then remove cooling bath.Reaction mixture at room temperature stirred 24 hours.Reaction soln CH ₂Cl ₂(50ml) saturated NaHCO is poured in dilution into ₃In the aqueous solution (50ml).Water CH ₂Cl ₂(3 * 50ml) extractions, the CH of merging ₂Cl ₂Extraction liquid salt water washing, and dry (MgSO ₄).With flash column chromatography (silicon-dioxide, 10%CH ₂Cl ₂Ethyl acetate solution) purify.Obtain the N-oxide compound, be light yellow solid. ¹H NMR(250MHz, CDCl ₃) δ 8.18(d, J=16.2Hz, 1H, alkene), 7.97(d, J=6.5Hz, 1H, 6-pyridyl), 7.58(d, J=16.2Hz, 1H, alkene), 7.11(dd, J=8.6,6.5Hz, 1H, 5-pyridyl), 6.82(d, J=8.6Hz, 1H, 4-pyridyl), 4.08(t, J=6.6Hz, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.93-0.88(m, 19H, aliphatic series).

1(e) 6-(E-2-carboxymethyl vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridone

Under argon atmospher, with 2-(E-2-carboxymethyl vinyl)-(180mg 0.537mmol) is dissolved in the anhydrous dimethyl formamide (2.2ml) 3-oxygen in last of the ten Heavenly stems yl pyridines N-oxide compound, is cooled to 0 ℃.(0.76ml 5.38mmol), then removes cooling bath to wherein adding trifluoacetic anhydride lentamente.Reaction mixture at room temperature stirred 18 hours.Reaction soln is poured saturated NaHCO into lentamente with ethyl acetate (75ml) dilution ₃(30ml) in the aqueous solution.Organic layer Na ₂HCO ₃(20ml) with the salt water washing, and dry (MgSO ₄).Obtain product, be yellow solid.Further do not purify and to use. ¹H NMR(250MHz, CDCl ₃) δ 7.75(d, J=16.3Hz, 1H, alkene), 7.40(d, J=9.8Hz, 1H, 3-pyridyl), 7.01(d, J=16.3Hz, 1H, alkene) and, 6.73(d, J=9.8Hz, 1H, 4-pyridyl), 3.95(t, J=6.6Hz, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.82-0.88(m, 19H, aliphatic series); MS(CI): 336(M+H).

1(f) 6-(E-2-carboxymethyl vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine trifluoromethane sulfonic acid ester

Under argon atmospher, to refrigerative (0 ℃) 6-(E-2-carboxymethyl vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridone (200mg, anhydrous CH 0.596mmol) ₂Cl ₂(3.0ml) add in the solution anhydrous pyridine (0.48ml, 5.96mmol) and trifluoromethanesulfanhydride anhydride (0.30ml, 1.78mmol).Reaction mixture stirred 15 minutes at 0 ℃.Reaction mixture is used H respectively with ethyl acetate (50ml) dilution ₂O(20ml), 2%HCl(10ml), saturated NaHCO ₃(20ml) with the salt water washing, and dry (MgSO ₄).Purify with flash column chromatography (silicon-dioxide, the petroleum ether solution of 5% ethyl acetate), obtain sulphonate, be colorless oil. ¹H NMR(250MHz, CDCl ₃) δ 7.97(d, J=15.8Hz, 1H, alkene), 7.36(d, J=8.8Hz, 1H, 3-pyridyl), 7.11(d, J=8.8Hz, 1H, 4-pyridyl) and, 6.96(d, J=15.8Hz, 1H, alkene), 4.05(t, J=6.5Hz, 2H, OCH ₂), 3.83(s, 3H, CO ₂CH ₃), 1.92-0.88(m, 19H, aliphatic series).

1(g) N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide

With 6-(E-2-carboxymethyl vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine trifluoromethane sulfonic acid ester (160mg, 0.342mmol) be dissolved in the anhydrous dimethyl formamide (1.25ml), sequentially use the 3-Methyl anthranilate (775mg, 5.13mmol, Lancaster), Pd(OAc) ₂(4.5mg, 0.020mmol) and 1,1 '-(22mg 0.040mmol) handles two (diphenylphosphino) ferrocene.Lentamente the carbon monoxide bubbling was passed through this solution 5 minutes.Then, (balloon pressure) and 90 ℃ of reacting by heating mixtures 4 hours under CO-atmosphere.When being cooled to room temperature, with ethyl acetate (75ml) diluted reaction mixture, with 2%HCl(5 * 10ml), H ₂O(15ml), saturated NaHCO ₃(15ml) with the salt water washing, and dry (MgSO ₄).With flash column chromatography (silicon-dioxide, ethyl acetate: CH ₂Cl ₂: sherwood oil=10: 20: 70) purify.Obtain acid amides, be colorless solid. ¹H NMR(250MHz, CDCl ₃) δ 9.85(s, 1H, NH), and 8.29(s, 1H, 2-phenyl), 8.27(d, J=8.7Hz, 1H, 3-pyridyl), 8.14(d, J=7.9Hz, 1H, 4-phenyl), 8.10(d, J=15.8Hz, 1H, alkene), 7.84(d, J=7.9Hz, 1H, the 6-phenyl), 7.48(dd, J=7.9Hz, 1H, 5-phenyl), 7.38(d, J=8.7Hz, 1H, 4-pyridyl), 7.08(d, J=15.8Hz, 1H, alkene), 4.12(t, J=6.6Hz, 2H, OCH ₂), 3.95(s, 3H, CO ₂CH ₃), 3.88(s, 3H, CO ₂CH ₃), 1.96-0.88(m, 19H, aliphatic series); Ultimate analysis, calculated value (C ₂₈H ₃₆O ₆N ₂): C, 67.72; H, 7.31; N, 5.64, experimental value: C, 67.50; H, 7.27; N, 5.57; MS(CI): 497.5(M+H).

1(h) N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide disodium salt

With N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide (60mg, 0.121mmol) be dissolved in tetrahydrofuran (THF) (1.25ml) and MeOH(0.50ml) in, use 1M LiOH(0.50ml) handle.Reaction mixture at room temperature stirred 6 hours.By adding 2%HCl(0.75ml) make reaction mixture acidifying lenitively, reaction mixture is with ethyl acetate (50ml) dilution, water (3 * 10ml) and the salt water washing, and dry (MgSO ₄).Solvent removed in vacuo.Diacid is dissolved in saturated Na ₂CO ₃In the aqueous solution (3-5ml), purify by anti-phase MPLC method (RP-18 silicon-dioxide, the 10-65%MeOH aqueous solution), and the lyophilize separation, obtain product, be white amorphous solid. ¹H NMR(250MHz, CD ₃OD) δ 8.22(s, 1H, 2-phenyl), 8.13(d, J=8.7Hz, 1H, the 3-pyridyl), 7.90-7.70(m, 2H, 4-phenyl, 6-phenyl), 7.73(d, J=15.8Hz, 1H, alkene), 7.65(d, J=8.7Hz, 1H, 4-pyridyl), 7.48(dd, J=7.9Hz, 1H, the 5-phenyl), 7.17(d, J=15.8Hz, 1H, alkene), 4.26(t, J=6.6Hz, 2H, OCH ₂), 1.98-0.82(m, 19H, aliphatic series); FAB-MS:(+ve), 513.1(M+H); (-ve), 489.0(M-Na).

Embodiment 2

The N-(3-carboxyl phenyl)-the 6-(2-carboxy ethyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide

2(a) N-(3-carboxymethyl phenyl)-6-(2-carboxymethyl ethyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide

With N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide (70mg 0.141mmol) is dissolved in the ethyl acetate (1ml), uses 5%Pd/C(10mg) processing, at H ₂Atmosphere (balloon pressure) stirred 4 hours down.Reaction mixture does not carry out TLC, and product is insoluble to ethyl acetate.By adding CH ₂Cl ₂(5ml) make sedimentary product dissolving, solution filters by diatomaceous earth filler.Product flash column chromatography (silicon-dioxide, the CH of 5% ethyl acetate ₂Cl ₂Solution) purify, obtain the pyridine carboxamide of title, be white solid. ¹H NMR(250MHz, CDCl ₃) δ 10.02(s, 1H, NH), 8.48(s, 1H, the 2-phenyl), 8.18(d, J=7.9Hz, 1H, 4-phenyl), 8.11(d, J=8.5Hz, 1H, 3-pyridyl), 7.81(d, J=7.9Hz, 1H, 6-phenyl), 7.46(dd, J=7.9Hz, 1H, the 5-phenyl), 7.20(d, J=8.5Hz, 1H, the 4-pyridyl), 4.05(t, J=6.4Hz, 2H, OCH ₂), 3.94(s, 3H, CO ₂CH ₃), 3.68(s, 3H, CO ₂CH ₃), 3.24(t, J=6.9Hz, 2H, CH ₂), 2.88(t, J=6.9Hz, 2H, CH ₂), 1.88-0.86(m, 19H, aliphatic series); Ultimate analysis, calculated value (C ₂₈H ₃₈O ₆N ₂): C, 67.45; H, 7.68; N, 5.62, experimental value: C, 67.26; H, 7.76; N, 5.54; MS(CI): 499(M+H).

2(b) N-(3-carboxyl phenyl)-the 6-(2-carboxy ethyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide di-potassium

With N-(3-carboxymethyl phenyl)-6-(2-carboxymethyl ethyl)-(54mg 0.108mmol) is suspended in tetrahydrofuran (THF) (1.1ml) and the methyl alcohol (0.70ml) 5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide, uses 1M LiOH(0.45ml, 0.45mmol) handles.Reaction mixture at room temperature stirred 30 hours.Reaction mixture is poured 2%HCl(15ml into ethyl acetate (50ml) dilution) in.Ethyl acetate layer H ₂O(3 * 20ml) and salt water washing, and dry (MgSO ₄).Solvent removed in vacuo is dissolved in KHCO with the solid diacid ₃In the aqueous solution (3-5ml).Purify with anti-phase MPLC method (RP-18 silicon-dioxide, 10-65% methanol aqueous solution), and the lyophilize separation, obtain salt, be white amorphous solid. ¹H NMR(250MHz, CD ₃OD) δ 8.49(s, 1H, 2-phenyl), 8.00(d, J=8.5Hz, 1H, 3-pyridyl) and, 7.88(d, J=7.9Hz, 1H, 4-phenyl), 7.72(d, J=7.9Hz, 1H, 6-phenyl) and, 7.36(m, 2H, 4-pyridyl, 5-phenyl), 4.11(t, J=6.4Hz, 2H, OCH ₂), 3.19(t, J=6.9Hz, 2H, CH ₂), 2.66(t, J=6.9Hz, 2H, CH ₂), 1.92-0.87(m, 19H, aliphatic series); FAB-MS:(+ve), 547.4(M+H); (-ve), 507.3(M-K).

Embodiment 3

The N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-tetradecyloxyaniline-2-pyridine carboxamide dilithium salt

3(a) 3-hydroxyl-2-pyridylaldehyde

With 3-hydroxyl-2-(methylol) pyridine hydrochloride (1.32g, 6.9mmol, Aldrich, 85%) is dissolved in anhydrous CH ₂Cl ₂(35ml), with triethylamine (1.1ml, 7.89mmol) and MnO ₂(6.0g 69mmol) handles.Reaction mixture at room temperature stirred 18 hours.Filter vacuum concentration by diatomaceous earth filler.Thick aldehyde is not further purified and is directly used in next step.

3(b) 3-tetradecyloxyaniline-2-pyridylaldehyde

3-hydroxyl-2-pyridylaldehyde (about 6.9mmol) that previous step is obtained is dissolved in the anhydrous dimethyl formamide (10ml), sequentially uses anhydrous K ₂CO ₃(2.86g, 20.7mmol) (2.00ml 7.59mmol) handles above-mentioned solution with the 1-iododecane.Reaction mixture stirred 4.5 hours intensely with 90 ℃ under argon atmospher.When cool to room temperature, reaction mixture is used H with ethyl acetate (100ml) dilution ₂O(5 * 20ml) and salt water washing, and dry (MgSO ₄).Purify with flash column chromatography (silicon-dioxide, the petroleum ether solution of 30% ethyl acetate), obtain aldehyde, be light yellow oil. ¹H NMR(250MHz, CDCl ₃) δ 10.43(s, 1H, CHO), and 8.38(dd, J=4.1,1.5Hz, 1H, 6-pyridyl), 7.42(m, 2H, 4-pyridyl, 5-pyridyl), 4.10(t, J=6.5Hz, 2H, OCH ₂), 1.91-0.88(m, 27H, aliphatic series).

3(c) 2(E-2-carboxymethyl vinyl)-3-tetradecyloxyaniline pyridine

According to preparation 2-(E-2-carboxymethyl vinyl)-the described method preparation of 3-oxygen in last of the ten Heavenly stems yl pyridines. ¹H NMR(250MHz, CDCl ₃) δ 8.22(dd, J=4.0,1.8Hz, 1H, 6-pyridyl), 8.10(d, J=15.8Hz, 1H, alkene), 7.21(m, 2H, 4-pyridyl, 5-pyridyl) and, 7.02(d, J=15.8Hz, 1H, alkene), 4.02(t, J=6.5Hz, 2H, OCH ₂), 3.81(s, 3H, CO ₂CH ₃), 1.88-0.88(m, 27H, aliphatic series).

3(d) 2-(E-2-carboxymethyl vinyl)-3-tetradecyloxyaniline pyridine N-oxides

This compound is according to preparation 2-(E-2-carboxymethyl vinyl)-the described method preparation of 3-oxygen in last of the ten Heavenly stems yl pyridines N-oxide compound. ¹H NMR(250MHz, CDCl ₃) δ 8.18(d, J=16.2Hz, 1H, alkene), 7.95(d, J=6.5Hz, 1H, 6-pyridyl), 7.58(d, J=16.2Hz, 1H, alkene), 7.10(dd, J=8.5,6.5Hz, 1H, 5-pyridyl), 6.80(d, J=8.5Hz, 1H, 4-pyridyl), 4.08(t, J=6.6Hz, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.88-0.88(m, 27H, aliphatic series).

3(e) 6-(E-2-carboxymethyl vinyl)-5-tetradecyloxyaniline-2-pyridone

This compound is according to preparation 6-(E-2-carboxymethyl vinyl)-the described method preparation of 5-oxygen in last of the ten Heavenly stems base-2-pyridone. ¹H NMR(250MHz, CDCl ₃) δ 7.75(d, J=16.3Hz, 1H, alkene), 7.40(d, J=9.8Hz, 1H, 3-pyridyl), 7.01(d, J=16.3Hz, 1H, alkene) and, 6.73(d, J=9.8Hz, 1H, 4-pyridyl), 3.95(t, J=6.6Hz, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.82-0.88(m, 27H, aliphatic series).

3(f) 6-(E-2-carboxymethyl vinyl)-5-tetradecyloxyaniline-2-pyridine trifluoromethane sulfonic acid ester

This compound is according to the above-mentioned 6-(E-2-carboxymethyl vinyl for preparing)-preparation of the method for 5-oxygen in last of the ten Heavenly stems base-2-pyridine trifluoromethane sulfonic acid ester. ¹H NMR(250MHz, CDCl ₃) δ 7.96(d, J=15.7Hz, 1H, alkene), 7.35(d, J=8.8Hz, 1H, 3-pyridyl), 7.10(d, J=8.8Hz, 1H, 4-pyridyl) and, 6.96(d, J=15.7Hz, 1H, alkene), 4.04(t, J=6.5Hz, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.85-0.88(m, 27H, aliphatic series).

3(g) N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-tetradecyloxyaniline-2-pyridine carboxamide

Embodiment 1(g) method is used to prepare N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-tetradecyloxyaniline-2-pyridine carboxamide. ¹H NMR(250MHz, CDCl ₃) δ 9.86(s, 1H, NH) 8.29(s, 1H, 2-phenyl), 8.27(d, J=8.7Hz, 1H, the 3-pyridyl), 8.13(d, J=7.9Hz, 1H, 4-phenyl), 8.09(d, J=15.8Hz, 1H, alkene), 7.84(d, J=7.9Hz, 1H, the 6-phenyl), 7.48(dd, J=7.9Hz, 1H, 5-phenyl), 7.38(d, J=8.7Hz, 1H, 4-pyridyl), 7.08(d, J=15.8Hz, 1H, alkene), 4.12(t, J=6.6Hz, 2H, OCH ₂), 3.95(s, 3H, CO ₂CH ₃), 3.88(s, 3H, CO ₂CH ₃), 1.94-0.88(m, 27H, aliphatic series); MS(CI): 553.4(M+H).

3(h) N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-tetradecyloxyaniline-2-pyridine carboxamide dilithium salt

With N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-tetradecyloxyaniline-2-pyridine carboxamide (173mg 0.313mmol) is dissolved in tetrahydrofuran (THF) (4.0ml) and the methyl alcohol (1.0ml), uses 1M LiOH(1.0ml) processing.Reaction mixture at room temperature stirred 48 hours.The gel that obtains is dissolved in H ₂O(3ml) in, vacuum is removed tetrahydrofuran (THF) and methyl alcohol.Product is purified with anti-phase MPLC method (RP-18 silicon-dioxide, 10-65% methanol aqueous solution), and the lyophilize separation, obtains salt, is colourless amorphous solid.

¹H NMR(250MHz, CD ₃OD) δ 8.32(s, 1H, 2-phenyl), 8.12(d, J=8.7Hz, 1H, the 3-pyridyl), 7.85(d, J=15.7Hz, 1H, alkene), 7.83(d, J=7.9Hz, 1H, 4-phenyl), 7.76(d, J=7.9Hz, 1H, the 6-phenyl), 7.52(d, J=8.7Hz, 1H, 4-pyridyl), 7.38(dd, J=7.9Hz, 1H, 5-phenyl), 7.26(d, J=15.7Hz, 1H, alkene), 4.16(t, J=6.6Hz, 2H, OCH ₂), 1.94-0.89(m, 27H, aliphatic series); FAB-MS:(+ve), 537(M+H); (-ve), 529(M-Li).

Embodiment 4

The N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-dodecyloxy-2-pyridine carboxamide (dilithium salt)

According to preparation N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-tetradecyloxyaniline-described method of 2-pyridine carboxamide dilithium salt is by replacing 1-iododecane (referring to embodiment 3) with 1-iodo dodecane, preparation N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-dodecyloxy-2-pyridine carboxamide dilithium salt.

4(a) 3-dodecyloxy-2-pyridylaldehyde ¹H NMR(250MHz, CDCl ₃) δ 10.43(s, 1H, CHO), and 8.38(dd, 1H, 6-pyridyl), 7.42(m, 2H, 4-pyridyl, 5-pyridyl), 4.1(t, 2H, OCH ₂), 1.91-0.88(m, 23H, aliphatic series).

4(b) 2-(E-2-carboxymethyl vinyl)-3-dodecyloxy pyridine ¹H NMR(250MHz, CDCl ₃) δ 8.22(dd, 1H, 6-pyridyl), 8.1(d, 1H, J=15.8Hz, alkene), 7.21(m, 2H, 4-pyridyl, 5-pyridyl) and, 7.02(d, 1H, J=15.8Hz, alkene), 4.02(t, 2H, OCH ₂), 3.81(s, 3H, CO ₂CH ₃), 1.88-0.88(m, 23H, aliphatic series).

4(c) 2-(E-2-carboxymethyl vinyl)-3-dodecyloxy pyridine N-oxides ¹H NMR(250MHz, CDCl ₃) δ 8.15(d, 1H, J=16.2Hz, alkene), 7.9(d, 1H, 6-pyridyl), 7.58(d, 1H, J=16.2Hz, alkene) and, 7.1(dd, 1H, 5-pyridyl), 6.8(d, 1H, 4-pyridyl) and, 4.08(t, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.88-0.88(m, 23H, aliphatic series).

4(e) 6-(E-2-carboxymethyl vinyl)-5-dodecyloxy-2-pyridone ¹H NMR(250MHz, CDCl ₃) δ 8.0(s, 1H, OH), and 7.75(d, 1H, J=16Hz, alkene), 7.4(d, 1H, 3-pyridyl), 7.0(d, 1H, J=16Hz, alkene) and, 6.7(d, 1H, 4-pyridyl), 4.0(t, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.85-0.88(m, 23H, aliphatic series).

4(f) 6-(E-2-carboxymethyl vinyl)-5-dodecyloxy-2-pyridine trifluoromethane sulfonic acid ester ¹H NMR(250MHz, CDCl ₃) δ 7.95(d, 1H, J=15.9Hz, alkene), 7.37(d, 1H, 3-pyridyl), 7.1(d, 1H, 4-pyridyl) and, 6.95(d, 1H, J=15.9Hz, alkene), 4.1(t, 2H, OCH ₂), 3.8(s, 3H, CO ₂CH ₃), 1.89-0.88(m, 23H, aliphatic series).

4(g) N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-dodecyloxy-2-pyridine carboxamide ¹H NMR(250MHz, CDCl ₃) δ 9.86(s, 1H, NH), 8.29(s, 1H, aryl), 8.27(d, 1H, 3-pyridyl), 8.13(d, 1H, aryl), 8.09(d, 1H, J=15.8Hz, alkene), 7.84(d, 1H, aryl), 7.5(t, 1H, aryl), 7.38(d, 1H, 4-pyridyl), 7.08(d, 1H, J=15.8Hz, alkene), 4.15(t, 2H, OCH ₂), 3.98(s, 3H, CO ₂CH ₃), 3.88(s, 3H, CO ₂CH ₃), 1.94-0.88(m, 23H, aliphatic series); Ultimate analysis, calculated value (C ₃₀H ₄₀N ₂O ₆): C, 68.68; H, 7.69; N, 5.34, experimental value: C, 68.43; H, 7.54; N, 5.21; MS(CI): 525(M+H).

4(h) N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-dodecyloxy-2-pyridine carboxamide dilithium salt ¹H NMR(250MHz, CD ₃OD) δ 8.37(s, 1H, aryl), 8.12(d, 1H, the 3-pyridyl), 7.85(d, 1H, J=15.7Hz, alkene), 7.83(d, 1H, aryl), 7.77(d, 1H, aryl), 7.55(d, 1H, 4-pyridyl), 7.38(t, 1H, aryl), 7.26(d, 1H, J=15.7Hz, alkene), 4.16(t, 2H, OCH ₂), 1.90-0.88(m, 23H, aliphatic series); FAB-MS:(+ve), 509(M+H); (-ve), 501(M-Li).

Embodiment 5

The N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-octyloxy-2-pyridine carboxamide dilithium salt

According to preparation N-(3-carboxyl phenyl)-6-(E-2-carboxy vinyl-5-tetradecyloxyaniline-2-pyridine carboxamide dilithium salt (embodiment 3) described method and replace the 1-iododecane with 1-iodo octane, prepare the N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-octyloxy-2-pyridine carboxamide dilithium salt.

5(a) 3-octyloxy-2-pyridylaldehyde ¹H NMR(250MHz, CDCl ₃) δ 10.43(s, 1H, CHO), and 8.38(dd, 1H, 6-pyridyl), 7.42(m, 2H, 4-pyridyl, 5-pyridyl), 4.1(t, 2H, OCH ₂), 1.91-0.88(m, 15H, aliphatic series).

5(b) 2-(E-2-carboxymethyl vinyl)-3-octyloxy pyridine ¹H NMR(250MHz, CDCl ₃) δ 8.22(dd, 1H, 6-pyridyl), 8.1(d, 1H, J=15.8Hz, alkene), 7.21(m, 2H, 4-pyridyl, 5-pyridyl) and, 7.02(d, 1H, J=15.8Hz, alkene), 4.02(t, 2H, OCH ₂), 3.81(s, 3H, CO ₂CH ₃), 1.88-0.88(m, 15H, aliphatic series).

5(c) 2-(E-2-carboxymethyl vinyl)-3-octyloxy pyridine N-oxides ¹H NMR(250MHz, CDCl ₃) δ 8.15(d, 1H, J=16.2Hz, alkene), 7.9(d, 1H, 6-pyridyl), 7.58(d, 1H, J=16.2Hz, alkene) and, 7.1(dd, 1H, 5-pyridyl), 6.8(d, 1H, 4-pyridyl) and, 4.08(t, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.88-0.88(m, 15H, aliphatic series).

5(d) 6-(E-2-carboxymethyl vinyl)-5-octyloxy-2-pyridone ¹H NMR(250MHz, CDCl ₃) δ 8.0(s, 1H, OH), and 7.75(d, 1H, J=16Hz, alkene), 7.4(d, 1H, 3-pyridyl), 7.0(d, 1H, J=16Hz, alkene) and, 6.7(d, 1H, 4-pyridyl), 4.0(t, 2H, OCH ₂), 3.82(s, 3H, CO ₂CH ₃), 1.85-0.88(m, 15H, aliphatic series).

5(e) 6-(E-2-carboxymethyl vinyl)-5-octyloxy-2-pyridine trifluoromethane sulfonic acid ester ¹H NMR(250MHz, CDCl ₃) δ 7.95(d, 1H, J=15.9Hz, alkene), 7.37(d, 1H, 3-pyridyl), 7.1(d, 1H, 4-pyridyl) and, 6.95(d, 1H, J=15.9Hz, alkene), 4.1(t, 2H, OCH ₂), 3.8(s, 3H, CO ₂CH ₃), 1.89-0.88(m, 15H, aliphatic series).

5(f) N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-octyloxy-2-pyridine carboxamide ¹H NMR(250MHz, CDCl ₃) δ 9.86(s, 1H, NH), 8.29(s, 1H, aryl), 8.27(d, 1H, 3-pyridyl), 8.13(d, 1H, aryl), 8.09(d, 1H, J=15.8Hz, alkene), 7.84(d, 1H, aryl), 7.5(t, 1H, aryl), 7.38(d, 1H, 4-pyridyl), 7.08(d, 1H, J=15.8Hz, alkene), 4.15(t, 2H, OCH ₂), 3.98(s, 3H, CO ₂CH ₃), 3.88(s, 3H, CO ₂CH ₃), 1.94-0.88(m, 15H, aliphatic series).

5(g) N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-octyloxy-2-pyridine carboxamide dilithium salt ¹H NMR(250MHz, CD ₃OD) δ 8.37(s, 1H, aryl), 8.12(d, 1H, the 3-pyridyl), 7.85(d, 1H, J=15.7Hz, alkene), 7.83(d, 1H, aryl), 7.77(d, 1H, aryl), 7.55(d, 1H, 4-pyridyl), 7.38(t, 1H, aryl), 7.26(d, 1H, J=15.7Hz, alkene), 4.16(t, 2H, OCH ₂), 1.90-0.88(m, 15H, aliphatic series); FAB-MS:(+ve), 601.3(M+H); (-ve), 598.9(M-H).

Embodiment 6

The N-(3-carbonyl phenyl)-the 6-(E-2-carboxy vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-2-pyridine carboxamide dilithium salt

According to preparation N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-tetradecyloxyaniline-described method of 2-pyridine carboxamide dilithium salt (embodiment 4), and use the 8-(4-p-methoxy-phenyl) octane-1-(4-tosylate) replace 1-iododecane, preparation N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-2-pyridine carboxamide dilithium salt (referring to embodiment 3).

According to embodiment 3(d) and following method prepare following compounds.

6(a) 2-(E-2-carboxymethyl vinyl)-and 3-(8-(4-p-methoxy-phenyl) octyloxy) pyridine

The tosylate of embodiment A is used to prepare this compound. ¹H NMR(250MHz, CDCl ₃) δ 8.28(dd, J=4.0,1.8Hz, 1H, 6-pyridyl), 8.17(d, J=15.8Hz, 1H, alkene), 7.28(m, 2H, the 4-pyridyl, the 5-pyridyl), 7.12(d, J=8.6Hz, 2H, aryl), 7.02(d, J=15.8Hz, 1H, alkene), 6.89(d, J=8.6Hz, 2H, aryl), 4.08(t, J=6.5Hz, 2H, OCH ₂), 3.87(s, 3H, CO ₂CH ₃), 3.85(s, 3H, OCH ₃), 2.61(t, J=7.5Hz, 2H, benzyl), 1.94-1.38(m, 12H, aliphatic series).

6(b) 2-(E-2-carboxymethyl vinyl)-and 3-(8-(4-p-methoxy-phenyl)-octyloxy) pyridine N-oxides ¹H NMR(250MHz, CDCl ₃) δ 8.02(d, J=16.2Hz, 1H, alkene), 7.80(d, J=6.5Hz, 1H, 6-pyridyl), 7.39(d, J=16.2Hz, 1H, alkene), 7.00(m, 2H, 5-pyridyl, the 4-pyridyl), 6.85(d, J=8.6Hz, 2H, aryl), 6.65(d, J=8.6Hz, 2H, aryl), 3.91(t, J=6.5Hz, 2H, OCH ₂), 3.68(s, 3H, CO ₂CH ₃), 3.62(s, 3H, OCH ₃), 2.37(t, J=7.5Hz, 2H, benzyl), 1.82-1.10(m, 12H, aliphatic series).

6(c) 6-(E-2-carboxymethyl vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-the 2-pyridone ¹H NMR(250MHz, CDCl ₃) δ 7.75(d, J=16.2Hz, 1H, alkene), 7.40(d, J=9.8Hz, 1H, 3-pyridyl), 7.10(d, J=8.6Hz, 2H, aryl), 7.00(d, J=16.2Hz, 1H, alkene), 6.82(d, J=8.6Hz, 2H, aryl), 6.70(d, J=9.8Hz, 1H, 4-pyridyl), 3.95(t, J=6.5Hz, 2H, OCH ₂), 3.85(s, 3H, CO ₂CH ₃), 3.82(s, 3H, OCH ₃), 2.57(t, J=7.5Hz, 2H, benzyl), 1.85-1.22(m, 12H, aliphatic series).

6(d) N-(3-carboxymethyl phenyl)-6-(E-2-carboxymethyl vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-70-73 ℃ of 2-pyridine carboxamide fusing point; ¹H NMR(250MHz, CDCl ₃) δ 9.87(s, 1H, NH), 8.31(s, 1H, the 2-phenyl), 8.28(d, J=8.7Hz, 1H, 3-pyridyl), 8.15(d, J=7.9Hz, 1H, 4-phenyl), 8.08(d, J=15.8Hz, 1H, alkene), 7.85(d, J=7.9Hz, 1H, the 6-phenyl), 7.48(dd, J=7.9Hz, 1H, the 5-phenyl), 7.36(d, J=8.7Hz, 1H, 4-pyridyl), 7.10(d, J=8.6Hz, 2H, aryl), 7.08(d, J=15.8Hz, 1H, alkene), 6.85(d, J=8.6Hz, 2H, aryl), 4.12(t, J=6.5Hz, 2H, OCH ₂), 3.95(s, 3H, CO ₂CH ₃), 3.88(s, 3H, CO ₂CH ₃), 3.79(s, 3H, OCH ₃), 2.56(t, J=7.5Hz, 2H, benzyl), 1.99-1.28(m, 12H, aliphatic series); Ultimate analysis, calculated value (C ₃₃H ₃₃N ₂O ₇): C, 68.97; H, 6.67; N, 4.88, experimental value: C, 69.21; H, 6.88; N, 4.46; MS(CI): 575(M+H).

6(e) N-(3-carboxyl phenyl)-the 6-(E-2-carboxy vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-315 ℃ of fusing points of 2-pyridine carboxamide dilithium salt (decomposition): ¹H NMR(250MHz, CD ₃OD) δ 8.31(s, 1H, 2-phenyl), 8.12(d, J=8.7Hz, 1H, the 3-pyridyl), 7.86(d, J=7.9Hz, 1H, the 4-phenyl), 7.85(d, J=15.8Hz, 1H, alkene), 7.76(d, J=7.9Hz, 1H, 6-phenyl), 7.52(d, J=8.7Hz, 1H, 4-pyridyl), 7.39(dd, J=7.9Hz, 1H, 5-phenyl), 7.26(d, J=15.8Hz, 1H, alkene), 7.07(d, J=8.6Hz, 2H, aryl), 6.80(d, J=8.6Hz, 2H, aryl), 4.15(t, J=6.5Hz, 2H, OCH ₂), 3.74(s, 3H, OCH ₃), 2.53(t, J=7.5Hz, 2H, benzyl), 1.93-1.37(m, 12H, aliphatic series); Ultimate analysis, experimental value: C, 61.69; H, 5.91; N, 4.60; FAB-MS:(+ve), 559.4(M+H); (-ve), 551.4(M-Li).

According to identical method, but replace the 3-Methyl anthranilate, the preparation following compounds with the 3-Methyl anthranilate that suitable chlorine replaces:

N-(3-carboxyl-6-chloro-phenyl-)-the 6-(E-2-carboxy vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-2-pyridine carboxamide dilithium salt; With

N-(3-carboxyl-4-chloro-phenyl-)-6-(E-2-carboxy vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-2-pyridine carboxamide dilithium salt.

Embodiment 7

By with in the water-soluble solution of salt and add enough acid, can make salt be converted into free acid so that pH is transferred to approximately about neutral (pH7.0).Though preferably use mineral acid such as HCl etc., can use any acid.Preferred use diluted acid and without concentrated acid, for example 1 to 6 normal solution is the most useful.Acid can add under about room temperature; Do not need special conditions.When solution-when reaching neutral pH or becoming acidity, acid just is precipitated out from solution, can reclaim by crystallization method or to any other useful method of given acid.

Embodiment 8

The medicinal preparations that comprises The compounds of this invention can prepare with various forms with various vehicle.The embodiment of these preparations is as follows:

Inhalation

Formula I compound is dissolved in the isotonic saline solution, is made into the solution that concentration is 1-10mg/ml.The each use under the airflow of regulating by the atomizer atomisation so that the dose that needs to be provided.

Tablet

Every per 10,000 of components

1. active ingredient (formula I compound) 40mg 400g

2. W-Gum 20mg 200g

3. alginic acid 20mg 200g

4. sodiun alginate 20mg 200g

5. Magnesium Stearate 1.3mg 13g

101.3mg 1013g

The method for preparing tablet:

The 1st step mixed component 1,2,3 and 4 in suitable mixing tank/blending machine.

The 2nd step added enough water in batches in the mixture that the first step obtains, carefully mix each back that adds.Add entry like this and mix and have certain consistence until material and make it can be transformed into wet particle.

The 3rd step made wet stock be converted into particle by vibrating nodulizer with No. 8 (2.38mm) sieve.

The 4th step, wet granular was dried in baking oven at 60 ℃, until drying then.

The 5th step dry granules is lubricated with component 5.

The 6th step, lubricated particle was suppressed on suitable tabletting machine in flakes.

Suppository

Per 1000 bolts of each bolt of component

1. active ingredient, formula I compound 40.0mg 40g

2. cetomacrogol 1000 1350.0mg 1,350g

3. Macrogol 4000 450.0mg 450g

1840.0mg 1，840g

The preparation method

The 1st step fuse component 2 and 3 together, and stirring is until evenly.

The 2nd step was dissolved in component 1 in the 1st fused material that obtains of step, and stirred until evenly.

The molten materials that the 3rd step obtained for the 2nd step is poured in the suppository mould, and suppository is taken out in cooling from mould, packing.

Claims (14)

1, the method for preparation or its pharmaceutically acceptable salt or N-oxide compound,

In the formula:

T is that acid amides connects base, and wherein carbonyl carbon is connected on the pyridine ring;

R is C ₁-C ₂₀Aliphatic group, the unsubstituted or phenyl C that replaces ₁-C ₁₀Aliphatic group, wherein the phenyl of Qu Daiing has one or more bases that are selected from the following groups: lower alkoxy, low alkyl group, trihalogenmethyl or halogen, perhaps R is C ₁-C ₂₀Aliphatic series oxygen base, perhaps R is phenyl C unsubstituted or that replace ₁-C ₁₀Aliphatic series oxygen base, wherein the phenyl of Qu Daiing has one or more lower alkoxies, low alkyl group, trihalogenmethyl or halogen;

R ₁Be R ₄,-(C ₁-C ₅Aliphatic group) R ₄,-(C ₁-C ₅Aliphatic group) CHO ,-(C ₁-C ₅Aliphatic group) CH ₂OR ₈,-CH ₂OH or-CHO;

R ₂Be hydrogen ,-COR ₅, R wherein ₅Be-OH, the group-OR of pharmaceutically acceptable formation ester ₆Or OX, wherein X is pharmaceutically acceptable positively charged ion, perhaps R ₅Be-N (R ₇) ₂, R wherein ₇Be the aliphatic group of a H or 1-10 carbon atom, the cycloalkyl-(CH of a 4-10 carbon atom ₂) _n-Ji, wherein n is 0-3, perhaps two R ₇Base is in conjunction with the ring that is formed with 4-6 carbon atom, perhaps R ₂Be NHSO ₂R ₉, R wherein ₉Be-CF ₃, C ₁-C ₆Alkyl or phenyl;

R ₃Be hydrogen, lower alkoxy, halogen ,-CN, COR ₅Or OH;

R ₄Be COR ₅, R wherein ₅Be-OH, the group-OR of pharmaceutically acceptable formation ester ₆Or OX, wherein X is pharmaceutically acceptable positively charged ion, perhaps R ₅Be-N (R ₇) ₂, R wherein ₇Be the aliphatic group of a H or 1-10 carbon atom, the cycloalkyl-(CH of a 4-10 carbon atom ₂) _n-Ji, wherein n is 0-3, perhaps two R ₇In conjunction with the ring that is formed with 4-6 carbon atom;

R ₈Be hydrogen, C ₁-C ₆Alkyl or C ₁-C ₆Acyl group;

This method comprises:

A) with formula I compound

L is the group as trifyl in the formula, uses carbon monoxide, 1, and the amine of two (diphenylphosphino) ferrocene of 1-, suitable palladium catalyst and formula 2 is handled,

R in the formula ₂Not acid amides or acid; Or

B) compound of hydrogenation of formula 3

R in the formula ₁Be-(C ₂-C ₅Alkenyl) R ₄,-(C ₂-C ₅Alkenyl) CHO or-(C ₂-C ₅Alkenyl) CH ₂OR ₈, form the compound of formula I, wherein R ₁Be-(C ₂-C ₅Alkyl) R ₄,-(C ₂-C ₅Alkyl) CHO or-(C ₂-C ₅Alkyl) CH ₂OR ₈Or

C) hydrolyzing type I compound, wherein R ₁, R ₂Or R ₃Be ester, form salt or acid; Or

D) esterification formula I compound, wherein R ₁, R ₂Or R ₃Be acid; Or

E) form the acid amides of formula I compound, wherein R ₁R ₂Or R ₃Be acid; Or

F) salt is changed into acid.

2, according to the method for claim 1, preparing wherein, R is C ₁-C ₂₀Aliphatic series oxygen base or C ₁-C ₂₀Aliphatic group, R ₁Be-(C ₁-C ₅Aliphatic group) R ₄Or R ₄, and R ₂Be-compound or its pharmaceutically acceptable salt of COOH.

3, according to the method for claim 2, preparing wherein, R is C ₈-C ₁₅Alkoxyl group and R ₁Be-CH=CHR ₄The compound of (wherein two keys are cis or trans).

4, according to the method for claim 3, preparing wherein, R is H ₁₇C ₈-O-, R ₁Be trans-CH=CHCOOH, and R ₂The position replaces-compound of COOH between being, i.e. compound N-(3-carboxyl phenyl)-6-(E-2-carboxy vinyl)-5-octyloxy-2-pyridine carboxamide, its dilithium salt or its pharmaceutically acceptable other salt or ester.

5, according to the method for claim 3, preparing wherein, R is H ₂₁C ₁₀-O-, R ₁Be trans-CH=CHCOOH, and R ₂The position replaces-compound of COOH between being, i.e. compound N-(3-carboxyl phenyl)-6-(E-2-carboxy vinyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide, its disodium salt or its pharmaceutically acceptable other salt or ester.

6, according to the method for claim 3, preparing wherein, R is H ₂₅C ₁₂-O-, R ₁Be trans-CH=CHCOOH and R ₂The position replaces-compound of COOH between being, i.e. compound N-(3-carboxyl phenyl)-6-(E-2-carboxy vinyl)-5-dodecyloxy-2-pyridine carboxamide, its dilithium salt or its pharmaceutically acceptable other salt or ester.

7, according to the method for claim 3, preparing wherein, R is H ₂₉C ₁₄-O-, R ₁Be trans-CH=CHCOOH, and R ₂The position replaces-compound of COOH between being, i.e. compound N-(3-carboxyl phenyl)-6-(E-2-carboxy vinyl)-5-tetradecyloxyaniline-2-pyridine carboxamide, its dilithium salt or its pharmaceutically acceptable other salt or ester.

8, according to the method for claim 2, preparing wherein, R is that replace or unsubstituted phenyl-C ₁-C ₁0 aliphatic group, R ₁Be-(C ₁-C ₅Aliphatic group) R ₄Compound.

9, method according to Claim 8, preparing wherein, R is phenyl-C that lower alkoxy replaces ₁-C ₈The compound of alkoxyl group.

10, according to the method for claim 9, preparing wherein, R is contraposition H ₃CO-phenyl-(CH ₂) ₈-O-, R ₁Be HO ₂CCH=CH-, and R ₂The position replaces-compound of COOH between being, be compound N-(3-carboxyl phenyl)-6-(E-2-carboxy vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-the 2-pyridine carboxamide, N-(3-carboxyl-6-chloro-phenyl-)-the 6-(E-2-carboxy vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-2-pyridine carboxamide dilithium salt, or N-(3-carboxyl-4-chloro-phenyl-)-the 6-(E-2-carboxy vinyl)-5-(8-(4-p-methoxy-phenyl) octyloxy)-2-pyridine carboxamide dilithium salt or its pharmaceutically acceptable other salt or ester.

11,, prepare wherein R according to the method for claim 2 ₁Be R ₄CH ₂CH ₂-compound.

12, according to the method for claim 11, preparing wherein, R is H ₂₁C ₁₀-O-, R ₁Be HO ₂CCH ₂CH ₂-and R ₂The position replaces-compound of COOH between being, i.e. compound N-(3-carboxyl phenyl)-6-(2-carboxy ethyl)-5-oxygen in last of the ten Heavenly stems base-2-pyridine carboxamide, its di-potassium or its pharmaceutically acceptable other salt or ester.

13, the method for pharmaceutical compositions comprises formula I compound or its pharmaceutically acceptable salt and pharmaceutically acceptable carrier combination with definition in the claim 1.

14, according to the method for claim 13 pharmaceutical compositions, make be suitable for sucking, form parenteral, oral or topical.