CN1193793C - Diarylprophin photosensitizer and its preparing process and usage - Google Patents

Abstract

The invention belongs to the technical field of photosensitizers with photodynamic activity, and in particular relates to 5,15-diaryl porphine photosensitizers and their preparation methods and applications. The photosensitizer is: see the right formula, wherein R is selected from (1) hydrogen; (2) C _n H _2n+1 , n=1-8; (3) C _n H _2n+1 CO, n=1 -7; (4) Hydroxy; (5) C _n H _2n+1 O, n=1-8; (6) Halo X, X=F, Cl, Br or I; (7) C _n H _2n X , n=1-8, X=F, Cl, Br or I; (8) carboxylic acid; (9) COOC _n H _2n+1 , n=1-8; or (10) NR ¹ R ² , where R ¹ , R ² =H or C _n H _2n+1 , n=1-8. The photosensitizer of the invention has very good photodynamic properties, and can be used for preparing medicines for photodynamic therapy of cancer, blood diseases, blood purification and the like.

Description

Diarylprophin photosensitizer and its production and use

The invention belongs to photosensitizer technical field, especially relate to 5,15-Diarylprophin photosensitizer and its production and use with photodynamic activity.

Photodynamic therapy (photodynamic therapy is called for short PDT) is showed the human body photosensitizer administration, and illumination activates photosensitizer then, in the presence of oxygen, produces the purpose that active oxygen and other reactive intermediate (as the photosensitizer free radical) reach the treatment disease.Require the phototoxicity height of photosensitizer, dark toxicity is low, has strong absorption at phototherapy window (600-900 nanometer), and is easy to accumulate on the pathological tissues.Because phototherapy needs the participation of photosensitizer and light simultaneously, has double selection (drug selectivity enrichment and selective light are according to activating).Phototherapy just is easier to the drug effect point control in pathological tissues than general chemotherapy like this, reduces the damage to normal structure, thereby reduces the toxic and side effects of medicine greatly.

Though be used for clinical report relevant for optical dynamic therapy very early, optical dynamic therapy causes that really people pay attention to then starting from 1972 widely.The awns moral people such as (Diamond) of Dell has reported that at " scalpel " 1972 the 2nd volumes the 1175th page (Lancet1972,2,1175) phototherapy is used for the treatment of malignant tumor; Dao Erdi people such as (Dougherty) was at the 1333rd page of (J.Natl.Cancer Inst.1974 of " North Atlantic Treaty Organization's ICR magazine " 1974 the 51st volumes afterwards, 51,1333) and 1975 the 55th the volume the 115th page of (J.Natl.Cancer Inst.1975,55,115) reported phototherapy in shallow-layer cancers such as pulmonary carcinoma, neck cancer and cancer eye have been had obvious curative effects.After this, the scientific worker of various countries has carried out extensive, deep research to all many-sides such as the mechanism of photodynamic therapy, clinical characteristics.

Canadian healthy protect office on April 16th, 1 (The HealthProtection Bureau of Canada) announces that the commercialization of approval photofrin (Photofrin) is used for the treatment of bladder cancer.After this Japan, US and European have also been ratified to treat certain cancers with phototherapy in succession, as gastric cancer, pulmonary carcinoma etc.So far, photodynamic therapy has formally been established its status in clinical practice.

On the history of tumor phototherapy, hematoporphyrin derivative (Haematoporphyrinderivatives is called for short HpD) has critical role always.In Dao Erlin (Doiron) and height " porphyrin location and oncotherapy " book that (Gomer) do not compile, Dao Erdi people such as (Dougherty) is at " structure of hematoporphyrin derivative active component " (The structure of the active component ofhaematoporphyrin derivative.In Porphyrin Localizaion andTreatment of Tumors 1984 partly, 6,259-274) effective ingredient of having reported HpD is dihematoporphyrin ether or ester, i.e. DHE (Dihaematoporphyrin ethers andesters).The commodity preparation of HpD is exactly photofrin (Photofrin), the title of its commercially produced product has phototherapy element (Photosan) again, light spirit plain (Photogem) and light cancer element (Photocarcinorin) or the like, in fact they be exactly refining, the later product of purifying of HpD second phase, generally contain be lower than 20% non-activity monomer and be higher than 80% active porphyrin dimer and oligomer arranged.The phototherapy medicament that present China ratifies to use clinically has only HpD.

Although it is clinical with HpD to be that the mixing Porphyrin-Based Sensitizer of representative has been widely used in, these first generation photosensitizer have three very important shortcomings.At first, their selectivity is bad, easily causes the side effect of skin photosensitivity, and needs the lucifuge time long; The second, the absorption band of red light portion (band I, about 630 nanometers) is very weak, well absorptive red light; The 3rd, its component complexity, poor stability, this has all limited their application.

After the eighties, the research of second filial generation photosensitizer and screening have been stepped out a step phototherapy research again.The activity and the selectivity of these second filial generation photosensitizer are all improved.Bo Naite (Bonnett) is at " contemporary pharmacotherapy summary " 1999 the 10th phase page 1 (Rev.Contemp.Pharmacother.1999,10, delivered introduction 1-17) to some second filial generation photosensitizer just under development, these optical dynamic therapy medicines are the derivant of porphyrins mostly, comprise porphyrin, porphin, alizarinopurpurin, endogenous porphyrin etc.

At present gone through to be applied to clinical second filial generation photosensitizer abroad, have only this medicine of benzoporphyrin derivative list acid ring A (to be called for short BPDMA, commodity are called Verteporfin), it was used for diseases such as clinical treatment tumour and macula retinae degeneration in 2000 by FDA (FDA) approval by the exploitation of Canadian QLT Phototherapeutics company.If sea (Rouhi) is at " chemical news " magazine second phase in November, 1,998 22 pages of (C﹠amp; EN 1998, and Nov.2 22-26) reported once that this photosensitizer of BPDMA compared with HpD, can be used for the treatment of the darker and bigger tumor tissues in position in the human body.It can be concentrated fast at the destination organization place, and the place removes fast in normal structure, can carry out phototherapy after medicine enters 5 minutes, and do not resemble will not arranged two day waiting time with HpD.Because it is also fast that absorption soon, is removed, and make the photosensitivity time to skin shorten to one day, and HpD will continue several weeks.

Yet as phototherapy medicament, the defective of BPDMA also is clearly, and main cause is its semi-synthetic product that is derived from natural product, and also more complicated of synthetic process, and overall productive rate is very low, so cost is very high.The preparation cost of its final drug Verteporfin is also very high, so cause a Verteporfin market price of containing 15 milligrams of effective ingredient to want about 1,000 dollars, this has just greatly limited its clinical practice.

In the second filial generation photosensitizer that all are being developed, porphin photosensitizer occupies critical role again.The framing structure of porphin is identical with porphyrin, and just the two keys of in its molecular structure are replaced by singly-bound.This just makes porphin compare with porphyrin, and the phototherapy window in the near infrared region has stronger absorption.Dao Erfen people such as (Dolphin) has delivered the structure of many porphin photosensitizers just under development and the exhaustive overview of characteristic at " tetrahedron " magazine 1998 the 54th volume the 4151st page (Tetrahedron, 1998,54,4151).The porphin photosensitizer that is carrying out at present clinical trial mainly contains the Npe6 of Japanese petro-chemical corporation (Nippon Petrochemicals) exploitation, and the m-THPC that develops of Britain Bo Naite (Bonnett) people of etc.ing (commodity are called Foscan ^TM).These two kinds of porphin photosensitizers all show good phototherapy characteristic in clinical trial.In " photodynamic therapy and biomedical laser " book that Si Binli people such as (Spinelli) writes, Alan people such as (Allen) reported once that Npe6 can treat skin carcinoma very effectively in " using the optical dynamic therapy of Npe6 to the shallow malignant disease of people's body surface " (Photodynamic therapy of superficial malignancieswith Npe6 in man.In Photodynamic therapy and biomedicallasers) part, and did not almost cause the secular photo sensitive reaction of application on human skin.Sai Worui people such as (Savary) reported once in 1998 the 30th phases of " endoscopy " magazine the 258th page (Endoscopy30,258,1998) that m-THPC was used for early stage bronchogenic carcinoma of optical dynamic therapy and esophageal carcinoma, and achieved satisfactory results.Particularly under the situation that absorbs identical light dosage, the phototoxicity of m-THPC is more much better than than HpD and Photofrin.And m-THPC's is simple in structure, synthetic easily, compares with BPDMA, has very large advantage in the medicine cost control.

Generally speaking, compare with first generation photosensitizer such as HpD, porphin photosensitizer has more superior characteristic, mainly shows: (1) chemical composition is single, and purity is very high; (2) about the extinction coefficient of the absworption peak of red light district are generally than the HpD and the big order of magnitude of Photofrin; (3) stronger phototoxicity; (4) littler photosensitive side effect.

Yet these two kinds of novel porphin photosensitizers of Npe6 and m-THPC are also intact no all roses, and Npe6 is owing to being the semi-synthetic photosensitizer that stems from natural product, so exist complex structure, separation difficulty, the higher shortcoming of cost.Though and m-THPC is simple in structure, easily synthetic, and good phototherapy effect is arranged, as phototherapy medicament, the m-THPC part that also comes with some shortcomings.Wei, Ge Nieer people such as (Wagnieres) was once at 382 pages of (Photochem.Photobiol.1998 of 1998 the 68th volumes of " photochemistry and photo bio " magazine, 68,382) though reported the photosensitive side effect of skin that m-THPC causes less than Photofrin under therapeutic dose, secular photosensitive side effect still exists for human body skin for it.

In sum, for optical dynamic therapy, develop brand-new porphin photosensitizer and just have very important significance.

5,15-diaryl porphyrin is a kind of novel porphyrins, and structurally, it possesses two kinds of most typical porphyrin models, i.e. characteristics of tetraphenylporphyrin and octaethylporphyrin simultaneously.But for the diphenyl porphyrin, it is synthetic method comparatively effectively, is reported at 1999 the 76th volumes of " organic synthesis " book series 287 pages (Organic Syntheses.1999,76,287) by Bao Li people such as (Boyle) recently.So the research and development based on the porphin photosensitizer of diaryl porphyrin is blank always.

The object of the present invention is to provide a series of simple in structurely, easily synthetic, and Diarylprophin photosensitizer of good phototherapy effect and its production and use is arranged; It is medicine that these porphin photosensitizers can be used to prepare aspects such as optical dynamic therapy cancer, hematopathy, blood purification, and human body is had no side effect.

The object of the present invention is achieved like this:

With existing various 5,15-diaryl porphyrin is a raw material, adopt unifor reducing process and Osmic acid. oxidizing process respectively, obtained three types corresponding Diarylprophin photosensitizer, that is: 5,15-diaryl porphin (I), 5,15-diaryl porphine of bacterium (II) and 5,15-diaryl-2,3 dihydroxy porphin (III).

The structure of Diarylprophin photosensitizer is suc as formula shown in (I):

Wherein R is selected from (1) hydrogen; (2) C _nH _2n+1, n=1-8; (3) C _nH _2n+1CO, n=1-7; (4) hydroxyl; (5) C _nH _2n+1O, n=1-8; (6) halogen X, X=F, Cl, Br or I; (7) C _nH _2nX, n=1-8, X=F, Cl, Br or I; (8) carboxylic acid; (9) COOC _nH _2n+1, n=1-8; Or (10) NR ¹R ², R wherein ¹, R ²=H or C _nH _2n+1, n=1-8 etc.

The structure of diaryl porphine of bacterium class photosensitizer is suc as formula shown in (II):

Wherein R is selected from (1) hydrogen; (2) C _nH _2n+1, n=1-8; (3) C _nH _2n+1CO, n=1-7; (4) hydroxyl; (5) C _nH _2n+1O, n=1-8; (6) halogen X, X=F, Cl, Br or I; (7) C _nH _2nX, n=1-8, X=F, Cl, Br or I; (8) carboxylic acid; (9) COOC _nH _2n+1, n=1-8; Or (10) NR ¹R ², R wherein ¹, R ²=H or C _nH _2n+1, n=1-8 etc.

Diaryl-2, the structure of 3-dihydroxy porphin photosensitizer is suc as formula shown in (III):

Wherein R is selected from (1) hydrogen; (2) C _nH _2n+1, n=1-8; (3) C _nH _2n+1CO, n=1-7; (4) hydroxyl; (5) C _nH _2n+1O, n=1-8; (6) halogen X, X=F, Cl, Br or I; (7) C _nH _2nX, n=1-8, X=F, Cl, Br or I; (8) carboxylic acid; (9) COOC _nH _2n+1, n=1-8; Or (10) NR ¹R ², R wherein ¹, R ²=H or C _nH _2n+1, n=1-8 etc.

Photosensitizer structural formula of the present invention is that the synthetic method of (I) and Diarylprophin photosensitizer (II) is as follows:

With 5,15-diaryl porphyrin, Anhydrous potassium carbonate and unifor join in exsiccant pyridine or the picoline, heating, and stirring reaction under the argon shield condition, in system, add the pyridine solution that contains unifor more in batches, heating, and under the argon shield condition, stir reaction; After having reacted, add organic solvent and water in mixed system, heated and stirred is told organic layer, use cold hydrochloric acid, saturated sodium bicarbonate and distilled water wash respectively, obtain the Diarylprophin photosensitizer of structural formula (I) and the diaryl porphine of bacterium photosensitizer of structural formula (II);

Under the stirring at normal temperature condition, use cold hydrochloric acid, phosphoric acid solution, saturated sodium bicarbonate and distilled water wash respectively, with anhydrous sodium sulfate or anhydrous magnesium sulfate drying, filter gained liquid evaporate to dryness, column chromatography is purified, and recrystallization obtains the diaryl porphine of bacterium class photosensitizer of structural formula for (II);

Or

Under the stirring at normal temperature condition, add tetrachloroquinone or DDQ, in this process with the ultraviolet spectra monitoring, stopped reaction during to the characteristic absorption peak disappearance of its corresponding by-product structural formula diaryl porphine of bacterium photosensitizer that be (II); System is used sodium sulfite, sodium hydroxide, phosphoric acid solution, saturated sodium bicarbonate and distilled water wash more respectively, with anhydrous sodium sulfate or anhydrous magnesium sulfate drying; After filtering gained liquid evaporate to dryness, separate, carry out recrystallization again, obtain the Diarylprophin photosensitizer of structural formula (I);

The synthetic method of the Diarylprophin photosensitizer that photosensitizer structural formula of the present invention is (III) is as follows:

With 5,15-diaryl porphyrin is dissolved in the organic solvent, adds exsiccant pyridine or picoline again; Add the organic solvent that contains Osmic acid. to this system, stirring reaction under room temperature, lucifuge and argon shield condition, in system, feed the stink damp precursor reactant then, solvent evaporated, gained solid are dissolved in the chloroform soln that contains methanol, and room temperature stirs down, filter then, gained filtrate evaporate to dryness, column chromatography, recrystallization; Obtain the Diarylprophin photosensitizer of structural formula for (III).

Described structural formula for the synthetic method of (I) and Diarylprophin photosensitizer (II) is:

Below related amount be relative quantity, with 5 of 0.2-2 mM, the Anhydrous potassium carbonate of 15-diaryl porphyrin, 500 milligrams-10 grams and the unifor of 0.6-10 mM, join in exsiccant pyridine of 25-200 milliliter or the picoline, be 100-110 ℃ and under the argon shield condition, stir in temperature, reacted 2-4 hour; In system, add total amount more in batches and be the pyridine solution of 1-30 milliliter of the unifor of 0.6-80 mM, be 100-110 ℃ and under the argon shield condition, stir that in temperature total reaction time is 2-30 hour; After having reacted, in mixed system, add the water that organic solvent that 0.3-2 rises and 0.15-1 rise, on steam bath heated and stirred 1-1.5 hour; Tell organic layer, use cold hydrochloric acid, saturated sodium bicarbonate and the distilled water wash of 2 mol respectively, obtain the Diarylprophin photosensitizer of structural formula (I) and the diaryl porphine of bacterium photosensitizer of structural formula (II);

Under the stirring at normal temperature condition, respectively with the cold hydrochloric acid of 2 mol, phosphoric acid solution, saturated sodium bicarbonate and the distilled water wash that percentage by weight is 60-80%, with anhydrous sodium sulfate or anhydrous magnesium sulfate drying; Filter gained liquid evaporate to dryness, column chromatography is purified, and recrystallization obtains the diaryl porphine of bacterium class photosensitizer 0.04-0.5 milligram of structural formula for (II), productive rate 10%-25%;

Or

Under the stirring at normal temperature condition, add the tetrachloroquinone or the DDQ of 0.4-4 mM in batches, in this process with ultraviolet spectra monitoring, stopped reaction during to the characteristic absorption peak disappearance of its corresponding by-product structural formula diaryl porphine of bacterium photosensitizer that be (II); System is the sodium sulfite of 2-10%, the sodium hydroxide of 2-10%, phosphoric acid solution, saturated sodium bicarbonate and the distilled water wash of 40-60% with percentage by weight respectively again, with anhydrous sodium sulfate or anhydrous magnesium sulfate drying; After filtering gained liquid evaporate to dryness, use column chromatography to separate, carry out recrystallization again; Get corresponding Diarylprophin photosensitizer 0.1-1 mM, productive rate is 30%-51%;

Described structural formula is as follows for the synthetic method of the Diarylprophin photosensitizer of (III): following related amount is a relative quantity, with 5 of 1-5 mM, 15-diaryl porphyrin is dissolved in the organic solvent of 0.2-1 liter, adds exsiccant pyridine of 5-20 milliliter or picoline again; The 5-20 milliliter organic solvent that adds the Osmic acid. that contains the 1.1-5.5 mM to this system; Reaction system stirred 3-8 hour under room temperature, lucifuge and argon shield condition; In system, fed hydrogen sulfide gas 15-30 minute then, solvent evaporated, the gained solid is dissolved in 400 milliliters-2 liters and contains in the chloroform soln that percentage by weight is a 10%-30% methanol, and room temperature stirred 10-20 minute down, filtered then; Gained filtrate evaporate to dryness, column chromatography, recrystallization; Obtain the Diarylprophin photosensitizer 0.3-2 mM of structural formula for (III), productive rate is 30%-45%.

Described structural formula is that (I) and organic solvent (II) are benzene, toluene, dichloromethane, chloroform.

Described structural formula is benzene, toluene, dichloromethane, chloroform or acetone for the organic solvent of (III).

The present invention's three class Diarylprophin photosensitizers can be used to prepare the medicine of aspects such as optical dynamic therapy cancer, hematopathy, blood purification.

The synthetic three class Diarylprophin photosensitizers of the present invention all have strong absorption at phototherapy window (600-900 nanometer), and wherein photosensitizer II is very strong in the absorption of 734 nanometers, is very beneficial for treating tumor and cancer.Photosensitizer III has certain fat water amphipathic, help medicine in vivo with the combination of target cell.The photodynamic activity of these photosensitizer is strong, and dark toxicity is little, and being expected to becomes the of new generation optical dynamic therapy medicine better, more with better function than existing photosensitizer performance, is used for the phototherapy of tumor and other relevant disease.

Three class Diarylprophin photosensitizers involved in the present invention synthetic has that reactions steps is few, the comparatively easy and overall productive rate advantage of higher of operation.Optical physics and Photochemistry Study for this three classes Diarylprophin photosensitizer show that this three classes Diarylprophin photosensitizer all has very high photodynamic activity.They can both be under aerobic conditions photosensitive generation creating singlet oxygen by using ( ¹O ₂) and superoxide anion (O ₂ ^-).Photosensitizer I (during R=H) has very high creating singlet oxygen by using quantum yield (Φ _Δ=0.72); Photosensitizer III (during R=H) also has higher creating singlet oxygen by using quantum yield (Φ _Δ=0.65) photosensitization produces hydroxyl radical free radical (OH) very efficiently, and in aqueous solution.Simultaneously, under oxygen free condition, these photosensitizer photosensitization efficiently produce self anion free radical (Sen ^-).The cell in vitro experiment shows that this three classes Diarylprophin photosensitizer all has very strong lethal effect for tumor cell under illumination condition, and its phototoxicity all is better than hematoporphyrin derivative (HpD), and dark toxicity is lower.Wherein photosensitizer III (during R=H) phototoxicity is more than 200 times of hematoporphyrin derivative (HpD), and dark toxicity characteristic is suitable with HpD basically.

Below in conjunction with embodiment and accompanying drawing technical scheme of the present invention is further described.

Fig. 1. the survival rate of the embodiment of the invention 6 cancerous cell is mapped to light application time.

Embodiment 1.

5 milliliters (49.1 mMs) of benzaldehyde and new distilled pyrroles 125 milliliters (1.8 moles) mix, and stirring at normal temperature was also led to argon 15 minutes, added 0.38 milliliter trifluoroacetic acid (4.9 mM) then, and restir is 15 minutes under the room temperature argon shield.Room temperature decompression down steams the pyrroles, adds 50 milliliters of dichloromethane in the residue dark oil thing, washs with the aqueous sodium hydroxide washes of 0.1 mol then, and anhydrous sodium sulfate drying is used in reuse distillation washing twice, filters.Column chromatography (silica gel, 200 orders) separates, and dichloromethane uses thin layer chromatography (developing solvent: dichloromethane/normal hexane=1/1) monitor as eluent in this process.Collect suitable component, the evaporate to dryness dichloromethane, the residue yellow oil places the distillation device, fine vacuum (0.01 millimetres of mercury) is slowly intensification (1 ℃/5 minutes) down, up to being warming up to 130 ℃, distillation is spent the night, and collects the white crystal on the cold finger, can get 5-phenyl two pyrroles's methane (a) 5.7 grams, productive rate 50%.

The evaluation of compound a:

Fusing point: 101-102 ℃

Nuclear magnetic resonance, NMR (δ ¹H NMR): 7.90 (bs, 2H), 7.33-7.19 (m, 5H), 6.67 (q, 2H), 6.13 (q, 2H), 5.90 (m, 2H), 5.45 (s, 1H)

Mass spectral analysis: m/z 222 (M ⁺)

Elementary analysis: theoretical value, C ₁₅H ₁₄N ₂, C, 81.05; H, 6.3 5; N, 12.60; Actual measurement, C, 81.26; H, 6.32; N, 12.49.

Embodiment 2

5-phenyl two pyrroles's methane (a), 1.2 grams (5.4 mM) and trimethyl orthoformate 44 milliliters (0.4 moles) are dissolved in 1.6 liters of dichloromethane, stir under lucifuge, the room temperature argon shield condition.In this mixture, slowly drip 600 milliliters the dichloromethane solution that contains 21.2 gram (130 mM) trichloroacetic acids, add about half an hour.This mixture restir 4 hours under room temperature, lucifuge condition adds 40 milliliters of pyridines then, and restir is 17 hours under room temperature, the lucifuge condition.The logical oxygen of reaction system 30 minutes stirred 4 hours seeing under the optical condition again.On Rotary Evaporators, steam dichloromethane, in this process, after the dichloromethane when 90 percent steams, in system, add 150 milliliters water, continue on Rotary Evaporators, to steam solvent, become clarification up to system, and have a large amount of solids to occur.Filter, the gained solid is positioned over air drying, on silicagel column (200 order), carry out chromatography (eluant: dichloromethane/normal hexane=7/3) collect suitable component then, the evaporate to dryness eluant, gained purple solid is with after washed with methanol, the filtration, with the toluene recrystallization that contains one of percentage pyridine.Get 5,220 milligrams of 15-diphenyl porphyrins (b), productive rate 18%.

The evaluation of compound b:

Fusing point:＞300 ℃

Uv absorption λ max (log ε): 406nm (5.61), 500nm (4.25), 536nm (3.71), 574nm (3.75), 630nm (3.16)

Nuclear magnetic resonance, NMR (δ ¹H NMR) :-3.01 (brs, 2H), 7.85-7.90 (m, 6H), 8.35 (dd, 4H), 8.97 (d, 4H), 9.38 (d, 4H), 10.31 (s, 2H)

Mass spectral analysis (FAB): m/z 463 (M+1 ⁺).

Elementary analysis: theoretical value, C ₃₂H ₂₂N ₄, C, 83.09; H, 4.79; N, 12.11; Actual measurement, C, 82.81; H, 4.89; N, 11.90.

Embodiment 3

5,200 milligrams of 15-diphenyl porphyrins (b) (0.43 mM), unifor 0.2 gram (1.1 mM) and 500 milligrams of mixing of Anhydrous potassium carbonate place 50 milliliters of exsiccant pyridines, stir 2 hours under 105 ℃ of argon shield conditions.In this mixture, add 1 milliliter of pyridine solution containing 0.2 gram unifor then again, system restir 2 hours under 105 ℃ of argon shield conditions.Add 300 milliliters of benzene and 150 ml waters in the mixed system, heated and stirred is 1 hour on steam bath.Tell organic facies, use cold hydrochloric acid, saturated sodium bicarbonate and the distilled water wash of 2 mol respectively.Under the stirring at normal temperature condition, slowly add total amount is the tetrachloroquinone of 220 milligrams (0.89 mMs) in batches then, in this process with ultraviolet spectra monitoring, stopped reaction when the absworption peak of 734 nanometers disappears.System is respectively 50% phosphoric acid solution, saturated sodium bicarbonate and distilled water wash again with 5% sodium sulfite, 5% sodium hydroxide, weight ratio, uses anhydrous sodium sulfate drying.Filter gained liquid evaporate to dryness on Rotary Evaporators.Use silicagel column (200 order) to carry out chromatography (eluant: dichloromethane/normal hexane=1/1) collect suitable component, evaporate to dryness eluant, gained blue solid toluene recrystallization.Get 5,15-diphenyl porphin (I, R=H) 102 milligrams, productive rate 51%.

The evaluation of photosensitizer I (during R=H):

Fusing point:＞300 ℃

Uv absorption λ max (log ε): 363nm (4.53), 409nm (5.21), 505nm (4.11), 532nm (3.72), 593nm (3.62), 645nm (4.63)

Infrared spectrum cm ^-1: 3380,2923,2849,1606,1440,1417

Nuclear magnetic resonance, NMR (δ ¹H NMR) :-1.8 (brs, 2H), 4.38 (t, 2H), 4.69 (t, 2H), 7.77 (m, 6H), 7.96 (m, 2H), 8.21 (d, 1H), 8.33 (d, 1H), 8,51 (m, 2H), 8.89 (m, 2H), 9.11 (d, 1H), 9.35 (d, 1H), 9.97 (s, 1H), 10.78 (s, 1H)

Mass spectral analysis (FAB): m/z 465 (M+1 ⁺).

Elementary analysis: theoretical value, C ₃₂H ₂₄N ₄, C, 82.73; H, 5.21; N, 12.06; Actual measurement, C, 82.80; H, 5.37; N, 11.94.

Embodiment 4

5,100 milligrams of 15-diphenyl porphyrins (b) (0.22 mM), unifor 0.2 gram (1.1 mM) and Anhydrous potassium carbonate 2 gram mixing are dissolved in 30 milliliters of exsiccant picolines, stir 20 hours under 110 ℃ of argon shield conditions.In this process, add total amount in batches and be 12 milliliters the picoline solution that contains 1.8 gram (3.96 mM) unifor (wherein system can be under the argon shield condition room temperature place spent the night in 8 hours).Add 200 milliliters of toluene and 100 ml waters then in the mixed system, bathe under the heating condition at water vapour and stirred 1 hour.Telling organic facies, is respectively 68% phosphoric acid solution, saturated sodium bicarbonate and distilled water wash with cold hydrochloric acid, the weight ratio of 2 mol, uses anhydrous magnesium sulfate drying.Filter gained liquid evaporate to dryness on Rotary Evaporators.Use silicagel column (200 order) to carry out chromatography and (carry out eluting with normal hexane earlier, in eluant, progressively sneak into dichloromethane again, up to dichloromethane: normal hexane=1: 1), collect suitable component, the evaporate to dryness eluant, gained green solid dichloromethane/normal hexane recrystallization.Get 5,15-diphenyl porphine of bacterium (II, R=H) 20 milligrams, productive rate 19%.

The evaluation of photosensitizer II (during R=H):

Fusing point:＞300 ℃

Uv absorption λ max (log ε): 349nm (5.07), 373nm (5.16), 506nm (4.66), 734nm (5.11)

Infrared spectrum cm ^-1: 3372,2925,2851,1606,1440,1419

Nuclear magnetic resonance, NMR (δ ¹H NMR) :-1.53 (brs, 2H), 4.39 (t, 4H), 4.67 (t, 4H), 7.67 (m, 6H), 7.86 (m, 4H), 8.19 (d, 2H), 8.81 (d, 2H), 9.97 (s, 2H)

Mass spectral analysis (FAB): m/z 467 (M+1 ⁺)

Elementary analysis: theoretical value, C ₃₂H ₂₆N ₄, C, 82.38; H, 5.62; N, 12.01; Actual measurement, C, 82.19; H, 5.61; N, 11.97.

Embodiment 5

5,15-diphenyl porphyrin (b) 800 milligrams (1.76 mM) is dissolved in the mixed solvent of 300 milliliters of chloroform and 5 milliliters of exsiccant pyridines, adds 5 milliliters of chloroform solns that contain 500 milligrams of Osmic acid .s (1.97 mM) in the system.Reaction system stirred 3 hours under room temperature lucifuge argon shield condition.In mixed system, led to hydrogen sulfide gas 15 minutes then, solvent evaporated on Rotary Evaporators, the gained solid is dissolved in the mixed solvent that 400 milliliters of chloroform and 50 ml methanol form, room temperature stirred ten minutes down, filter then, gained filtrate evaporate to dryness on Rotary Evaporators, use silicagel column (200 order) to carry out chromatography (eluant: methylene chloride=20/1), collect suitable component, the evaporate to dryness eluant, gained blue solid dichloromethane/normal hexane recrystallization, obtain 5,15-diphenyl-2,3-dihydroxy porphin (III, R=H) 372 milligrams, productive rate 43%.

The evaluation of photosensitizer III (during R=H):

Fusing point: 260-262 ℃

Uv absorption λ max (log ε): 364nm (4.54), 402nm (5.19), 504nm (4.13), 531nm (3.89), 587nm (3.67), 638nm (4.55)

Infrared spectrum cm ^-1: 3423,2925,1610,1419,1034,962

Nuclear magnetic resonance, NMR (δ ¹H NMR) :-2.0 (brs, 2H), 5.90 (d, 1H), 6.45 (d, 1H), 7.75 (m, 6H), 8.12 (d, 1H), 8.24 (d, 2H), 8,48 (d, 1H), 8.55 (d, 1H), 8.65 (d, 1H), 8.89 (d, 1H), 8.95 (d, 2H), 9.10 (d, 1H), 9.28 (s, 1H), 9.96 (s, 1H)

Mass spectral analysis (FAB): m/z 496 (M ⁺)

Elementary analysis: theoretical value, C ₃₂H ₂₄N ₄O ₂, C, 77.40; H, 4.87; N, 12.06; Actual measurement: C, 77.18; H, 4.73; N, 12.11

Embodiment 6

The MDAMB543 breast cancer cell is incubated in the RPM1640 culture medium that contains 10% hyclone, places to contain 5%CO ₂, 95% air incubator in 37 ℃ of cultivations.Select for use the cell that is in exponential phase to experimentize.

Photosensitizer HpD, 5,15-diphenyl porphin (I, R=H), 5,15-diphenyl porphine of bacterium (II, R=H) and 5, (III R=H) respectively gets 1 milligram and is dissolved in respectively in 1 milliliter the dimethyl sulfoxide (DMSO) 15-diphenyl-2,3 dihydroxy porphin, being diluted to the required concentration of experiment with the RPM1640 culture medium that contains 10% hyclone then uses for phototoxicity and dark toxicity test, in this process, the ultimate density of DMSO in culture fluid is no more than 2 ‰, and this concentration can not influence the survival of cell.Wherein, HpD, photosensitizer I and II concentration are respectively 2 mcg/ml, and photosensitizer III then is diluted to 0.01 mcg/ml, 0.05 mcg/ml, 0.2 mcg/ml, four concentration of 1 mcg/ml.

Be taken on the flat culture plate in 96 holes and cultivate well, and the MDAMB543 breast cancer cell of adherent growth (concentration is 100,000 units per ml) mixes with the RPM1640 culture medium that contains various concentration photosensitizer respectively.On a plate, nine holes of each concentration of each photosensitizer plantation are planted nine in addition again and are not contained photosensitizer and only contain the cell hole of culture medium and 2 ‰ DMSO as matched group.Plant 6 blocks of plates altogether, will plant 96 good orifice plates then and place incubator, 37 ℃, contain 5%CO ₂, 95% air environment in lucifuge cultivated 4 hours.Taking out 5 blocks of plates wherein, is respectively the copper-vapor laser illumination 20,40,80,160,320 seconds of 510 nanometers (energy density 20 milliwatt/square centimeters) with wavelength.Cell in these 96 orifice plates was cultivated 24 hours in incubator more then, for measuring cell survival rate.

The survival rate of cell is measured with mtt assay.A plantation that does not have illumination is had 96 orifice plates of cell and various concentration photosensitizer and 96 orifice plates of all the other 5 illumination, and every hole adds 20 microlitre MTT solution (10 mg/ml are dissolved in the PBS buffer solution) respectively, and then cultivates 4 hours.Liquid in the careful sucking-off hole, every hole add 200 microlitre DMSO with the reduction of prevention MTT and dissolve blue crystallization.Jolt 10 minutes under the room temperature on dull and stereotyped oscillator, Bio-Rad 3550 type microplate reader are measured the absorbance value of 595 nanometers.Record the survival rate that data can calculate tumor cell as follows:

The survival rate of tumor cell (the %)=average OD value of the average OD value of administration group ÷ matched group * 100

The present embodiment result as shown in Figure 1, concentration be the photosensitizer I (during R=H) of 2 mcg/ml when irradiation time is more than 40 seconds, its fragmentation effect to tumor cell has substantially exceeded the HpD of same concentration; And photosensitizer II (during R=H) is under 2 mcg/ml concentration, and irradiation time is more than 160 seconds the time, and fragmentation effect has also surpassed the HpD of same concentration; For photosensitizer III (during R=H), under the 0.01 mcg/ml concentration, the fragmentation effect that illumination was shown in 20 seconds has surpassed the HpD of 2 mcg/ml.

Claims (6)

1. Diarylprophin photosensitizer, it is characterized in that: described photosensitizer is:

Or

Or

Wherein R is:

(1) hydrogen,

(2)C _nH _2n+1，n＝1-8、

(3)C _nH _2n+1CO，n＝1-7、

(4) hydroxyl,

(5)C _nH _2n+1O，n＝1-8、

(6) halogen X, X=F, Cl, Br or I,

(7) C _nH _2nX, n=1-8, X=F, Cl, Br or I,

(8) carboxylic acid,

(9)COOC _nH _2n+1，n＝1-8、

Or (10) NR ¹R ², R wherein ¹, R ²=H or C _nH _2n+1, n=1-8.

2. the preparation method of photosensitizer as claimed in claim 1 is characterized in that:

Described photosensitizer structural formula is that the synthetic method of (I) and Diarylprophin photosensitizer (II) is as follows:

With 5,15-diaryl porphyrin, Anhydrous potassium carbonate and unifor join in exsiccant pyridine or the picoline, heating, and stirring reaction under the argon shield condition, in system, add the pyridine solution that contains unifor more in batches, heating, and under the argon shield condition, stir reaction; After having reacted, add organic solvent and water in mixed system, heated and stirred is told organic layer, use cold hydrochloric acid, saturated sodium bicarbonate and distilled water wash respectively, obtain the Diarylprophin photosensitizer of structural formula (I) and the diaryl porphine of bacterium photosensitizer of structural formula (II);

Under the stirring at normal temperature condition, use cold hydrochloric acid, phosphoric acid solution, saturated sodium bicarbonate and distilled water wash respectively, with anhydrous sodium sulfate or anhydrous magnesium sulfate drying, filter gained liquid evaporate to dryness, column chromatography is purified, and recrystallization obtains the diaryl porphine of bacterium class photosensitizer of structural formula for (II);

Or

Under the stirring at normal temperature condition, add tetrachloroquinone or DDQ, in this process with the ultraviolet spectra monitoring, stopped reaction during to the characteristic absorption peak disappearance of its corresponding by-product structural formula diaryl porphine of bacterium photosensitizer that be (II); System is used sodium sulfite, sodium hydroxide, phosphoric acid solution, saturated sodium bicarbonate and distilled water wash more respectively, with anhydrous sodium sulfate or anhydrous magnesium sulfate drying; After filtering gained liquid evaporate to dryness, separate, carry out recrystallization again, obtain the Diarylprophin photosensitizer of structural formula (I);

The synthetic method of the Diarylprophin photosensitizer that described photosensitizer structural formula is (III) is as follows:

With 5,15-diaryl porphyrin is dissolved in the organic solvent, adds exsiccant pyridine or picoline again; Add the organic solvent that contains Osmic acid. to this system, stirring reaction under room temperature, lucifuge and argon shield condition, in system, feed the stink damp precursor reactant then, solvent evaporated, gained solid are dissolved in the chloroform soln that contains methanol, and room temperature stirs down, filter then, gained filtrate evaporate to dryness, column chromatography, recrystallization; Obtain the Diarylprophin photosensitizer of structural formula for (III).

3. the preparation method of photosensitizer as claimed in claim 2 is characterized in that: described structural formula for the synthetic method of (I) and Diarylprophin photosensitizer (II) is:

Below related amount be relative quantity, with 5 of 0.2-2 mM, the Anhydrous potassium carbonate of 15-diaryl porphyrin, 500 milligrams-10 grams and the unifor of 0.6-10 mM, join in exsiccant pyridine of 25-200 milliliter or the picoline, be 100-110 ℃ and under the argon shield condition, stir in temperature, reacted 2-4 hour; In system, add total amount more in batches and be the pyridine solution of 1-30 milliliter of the unifor of 0.6-80 mM, be 100-110 ℃ and under the argon shield condition, stir that in temperature total reaction time is 2-30 hour; After having reacted, in mixed system, add the water that organic solvent that 0.3-2 rises and 0.15-1 rise, on steam bath heated and stirred 1-1.5 hour; Tell organic layer, use cold hydrochloric acid, saturated sodium bicarbonate and the distilled water wash of 2 mol respectively, obtain the Diarylprophin photosensitizer of structural formula (I) and the diaryl porphine of bacterium photosensitizer of structural formula (II);

Under the stirring at normal temperature condition, respectively with the cold hydrochloric acid of 2 mol, phosphoric acid solution, saturated sodium bicarbonate and the distilled water wash that percentage by weight is 60-80%, with anhydrous sodium sulfate or anhydrous magnesium sulfate drying; Filter gained liquid evaporate to dryness, column chromatography is purified, and recrystallization obtains the diaryl porphine of bacterium class photosensitizer 0.04-0.5 mM of structural formula for (II), productive rate 10%-25%;

Or

Under the stirring at normal temperature condition, add the tetrachloroquinone or the DDQ of 0.4-4 mM in batches, in this process with ultraviolet spectra monitoring, stopped reaction during to the characteristic absorption peak disappearance of its corresponding by-product structural formula diaryl porphine of bacterium photosensitizer that be (II); System is the sodium sulfite of 2-10%, the sodium hydroxide of 2-10%, phosphoric acid solution, saturated sodium bicarbonate and the distilled water wash of 40-60% with percentage by weight respectively again, with anhydrous sodium sulfate or anhydrous magnesium sulfate drying; After filtering gained liquid evaporate to dryness, use column chromatography to separate, carry out recrystallization again; Get corresponding Diarylprophin photosensitizer 0.1-1 mM, productive rate is 30%-51%;

Described structural formula is as follows for the synthetic method of the Diarylprophin photosensitizer of (III): following related amount is a relative quantity, with 5 of 1-5 mM, 15-diaryl porphyrin is dissolved in the organic solvent of 0.2-1 liter, adds exsiccant pyridine of 5-20 milliliter or picoline again; The 5-20 milliliter organic solvent that adds the Osmic acid. that contains the 1.1-5.5 mM to this system; Reaction system stirred 3-8 hour under room temperature, lucifuge and argon shield condition; In system, fed hydrogen sulfide gas 15-30 minute then, solvent evaporated, the gained solid is dissolved in 400 milliliters-2 liters and contains in the chloroform soln that percentage by weight is a 10%-30% methanol, and room temperature stirred 10-20 minute down, filtered then; Gained filtrate evaporate to dryness, column chromatography, recrystallization; Obtain the Diarylprophin photosensitizer 0.3-2 mM of structural formula for (III), productive rate is 30%-45%.

4. as the preparation method of claim 2 or 3 described photosensitizer, it is characterized in that: described structural formula is that (I) and organic solvent (II) are benzene, toluene, dichloromethane, chloroform.

5. as the preparation method of claim 2 or 3 described photosensitizer, it is characterized in that:

Described structural formula is benzene, toluene, dichloromethane, chloroform or acetone for the organic solvent of (III).

6. the purposes of the described photosensitizer of claim 1, it is characterized in that: described photosensitizer is used to prepare the medicine of optical dynamic therapy cancer.