WO2016029671A1 - 一种硅基罗丹明衍生物及其制备方法和应用 - Google Patents

一种硅基罗丹明衍生物及其制备方法和应用 Download PDF

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WO2016029671A1
WO2016029671A1 PCT/CN2015/073677 CN2015073677W WO2016029671A1 WO 2016029671 A1 WO2016029671 A1 WO 2016029671A1 CN 2015073677 W CN2015073677 W CN 2015073677W WO 2016029671 A1 WO2016029671 A1 WO 2016029671A1
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silicon
group
hydrogen
rhodamine derivative
butyl
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汪亭
王保刚
柴晓云
吴秋业
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中国人民解放军第二军医大学
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F7/00—Compounds containing elements of Groups 4 or 14 of the Periodic Table
    • C07F7/02—Silicon compounds
    • C07F7/08—Compounds having one or more C—Si linkages
    • C07F7/10—Compounds having one or more C—Si linkages containing nitrogen having a Si-N linkage
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    • C09K11/00—Luminescent, e.g. electroluminescent, chemiluminescent materials
    • C09K11/06—Luminescent, e.g. electroluminescent, chemiluminescent materials containing organic luminescent materials
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
    • G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
    • G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
    • G01N21/33—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry using ultraviolet light
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances

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  • the present invention relates to the field of chemistry, and in particular to a silicon-based rhodamine derivative, a process for its preparation and the use thereof in the preparation of fluorescent labels for cell dyes, biological stains, biomolecules or biological particles.
  • Rhodamine is one of the xanthene compounds, which has the advantages of good light stability, high fluorescence quantum yield and simple chemical modification. It is a good fluorescent small molecule dye probe widely used in biological samples. Dyeing with fluorescent markers.
  • rhodamine as a fluorescent probe has many shortcomings in the application process, especially when applied to biological samples. For example, the excitation wavelength of Rhodamine B dye is in the ultraviolet-visible region, and the biological sample will cause severe background signal interference in the ultraviolet-visible region. On the other hand, ultraviolet visible light has poor penetrating ability to tissues and is highly phototoxic to biological samples. These factors greatly affect the sensitivity and accuracy of rhodamine dye probes in fluorescence analysis of biological samples, which greatly limits their further application in fluorescence imaging of biological samples.
  • Silicon-based rhodamine is a new class of fluorescent dyes developed in recent years.
  • the advantage is that by replacing the oxygen atom in the rhodamine molecule with the silicon atom, while retaining the advantages of the rhodamine compound, the spectral range is red-shifted to reach the near-infrared region, thereby satisfying the requirements of biological sample analysis and imaging.
  • patents for silicon-based rhodamines such as the patent WO 2010/126077 A1, the patent WO 2012/111818 A1 and the patent WO 2013/029650 A1. These patents describe the properties, synthesis methods, and applications of silicon-based rhodamine.
  • silicon-based rhodamine derivatives are prepared by a nucleophilic addition method of a reactive metal reagent and a carbonyl group.
  • This synthesis method has many steps on one hand, harsh reaction conditions, low yield, and cannot be used for a large amount of Synthesis; on the other hand, the use of active metal reagents greatly limits the choice of substrate, some of which replace the silicon-based rhodamine to the inability to synthesize, affecting its application in the imaging of biological samples. Therefore, it is necessary to design a method for synthesizing a silicon-based rhodamine derivative by designing a method with few steps, simple reaction conditions, high yield, diverse substrate selection, and suitable for large-scale preparation.
  • the object of the present invention is to provide a silicon-based rhodamine derivative for the deficiencies in the prior art.
  • Still another object of the present invention is to provide a process for producing the above-described silicon-based rhodamine derivative.
  • Another object of the present invention is to provide the use of the above-described silicon-based rhodamine derivative.
  • R 2 and R 3 are each a C1-C6 linear or branched hydrocarbon group or a phenyl group, and the hydrocarbon group is selected from the group consisting of methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl. , sec-butyl, tert-butyl, n-pentyl, isopentyl, sec-pentyl, neopentyl, n-hexyl, preferably methyl, ethyl and phenyl, most preferably methyl.
  • R 4 , R 5 , R 6 , R 7 , R 8 and R 9 are each independently a hydrogen or a C1-C6 linear or branched hydrocarbon group selected from the group consisting of methyl, ethyl, n-propyl and iso-.
  • R 20 , R 21 , R 22 and R 23 are each a C1-C6 linear or branched saturated or unsaturated hydrocarbon group or a cycloalkyl group selected from the group consisting of methyl, ethyl, n-propyl and iso-.
  • R 20 or R 21 may be bonded to R 5 as a carbon chain to form a five- or six-membered ring structure with a parent benzene ring, preferably a five-membered ring.
  • R 22 or R 23 may be bonded to R 8 as a carbon chain to form a five- or six-membered ring structure with a parent benzene ring, preferably a five-membered ring.
  • the substituent -NR 20 R 21 may form a structure having the following formula (III) with -R 5 :
  • the substituent -NR 22 R 23 may form a structure having the following formula (IV) with -R 8 :
  • the substituents R 24 and R 25 are hydrogen, methyl, ethyl or allyl, more preferably ethyl.
  • R 11 , R 12 , R 13 , R 14 , R 15 , R 16 , R 17 , R 18 , R 19 may be the same or different any substituent selected from the group consisting of: a hydrocarbon group, a halogen group, a cyano group, Nitro, alkoxy, haloalkyl.
  • a preferred structure is one of the following:
  • R 11 or R 15 is hydrogen, and the other is -CN, -NO 2 , -SO 3 H, -SO 3 - , -OH, -OCH 3 , -NH 2 , -N 3 , -NCS, -CH 3
  • R 12 , R 13 , R 14 are all hydrogen.
  • R 11 or R 15 is hydrogen and the other is -CH 3 , R 12 , R 13 , and R 14 is -CN, -NO 2 , -SO 3 H, -SO 3 - , -OH, -OCH 3 , -NH 2 , -CH 3 , -N 3 , -NCS, -C ⁇ CH, -CONH 2 , -CH 2 NH 2 , -F, -Cl, -Br, -I, -COOH, -COO - Any one of them, the other two are hydrogen.
  • R 11 or R 15 is hydrogen, the other is -SO 3 H or -SO 3 - , and any one of R 12 , R 13 and R 14 is one of -SO 3 H, -SO 3 - group, and the rest Both are hydrogen.
  • R 11 or R 15 is hydrogen, the other is -COOH or -COO - , and any of R 12 , R 13 and R 14 is -CN, -NO 2 , -SO 3 H, -SO 3 - , -OH , -OCH 3 , -NH 2 , -CH 3 , -N 3 , -NCS, -C ⁇ CH, -CONH 2 , -CH 2 NH 2 , -F, -Cl, -Br, -I, -COOH, -COO - one of the groups, the remaining two being hydrogen.
  • any one of R 16 , R 17 , R 18 and R 19 is -CN, -NO 2 , -SO 3 H, -SO 3 - , -OH, -OCH 3 , -NH 2 , -CH 3 , -N 3 , -NCS, -C ⁇ CH, -CONH 2 , -CH 2 NH 2 , -F, -Cl , -Br, -I, -COOH, -COO - a group of the remaining three are hydrogen.
  • the N,N-dihydrocarbyl m-bromoaniline derivative is reacted with n-butyllithium to form the corresponding lithium reagent, which is further reacted with dihydrocarbyldichlorosilane to give a key silicon-based intermediate.
  • the reaction process can be expressed as:
  • R 2 , R 3 , R 4 to R 9 and R 20 to R 23 have the same meanings as defined above.
  • the specific method of operation is preferably: 1.0 molar amount of N,N-dihydrocarbyl m-bromoaniline derivative is reacted with 1.05 molar amount of n-butyl lithium in dry diethyl ether at 0 ° C for 2 hours, and then 0.6 molar amount of dihydrocarbyl group.
  • N,N-dihydrocarbyl m-bromoaniline derivative is reacted with n-butyllithium to form a corresponding lithium reagent, which is then reacted with a dihydrocarbyldichlorosilane to replace one of the chlorines to form a monochlorosilane product.
  • Another N,N-dihydrocarbyl m-bromoaniline derivative is reacted with butyl lithium to form the corresponding lithium reagent, which is further reacted with the monochlorosilane product to provide a key silicon-based intermediate.
  • the reaction process can be expressed as:
  • R 2 , R 3 , R 4 to R 9 and R 20 to R 23 have the same meanings as defined above;
  • the specific operation method is preferably: a 1.0 molar amount of N,N-dihydrocarbyl m-bromoaniline derivative is reacted with 1.05 molar amount of n-butyllithium in dry diethyl ether at 0 ° C for 2 hours, and added thereto to contain 5.0 molar amount.
  • the dihydrocarbyldichlorosilane is dried in diethyl ether at 0 ° C for 2 hours to obtain a monochlorosilane product; the other 0.5 molar amount of N,N-dihydrocarbyl m-bromoaniline derivative and 0.525 molar amount of positive
  • the butyl lithium was reacted in dry diethyl ether at 0 ° C for 2 hours.
  • the reaction liquid was slowly added to the monochlorosilane product at 0 ° C, slowly raised to room temperature, the reaction was continued for 12 hours, and the reaction was quenched by adding water, and extracted.
  • Step 2 Reaction of a key silicon-based intermediate with a different substituted benzaldehyde to form a silicon-based rhodamine derivative:
  • reaction process can also be expressed as:
  • R 2 , R 3 , R 4 to R 9 , R 11 - R 19 and R 20 to R 23 have the same meanings as defined above.
  • the molar ratio of the key silicon-based intermediate having the structure of the general formula (V) to the differently substituted benzaldehyde derivative is 1:5.
  • the catalyst is selected from CuBr 2 or p-toluenesulfonic acid monohydrate.
  • the molar ratio of the critical silicon-based intermediate having the structure of the general formula (V) to the catalyst is from 1:0.1 to 1.
  • the specific operation method is preferably: adding 1.0 molar amount of the key silicon-based intermediate, 5.0 molar amount of substituted benzaldehyde, and 0.1-1.0 molar amount of the catalyst to the thick-wall pressure-resistant reaction tube, sealing the tube, and placing it in a protective cover.
  • the reaction was carried out at 140 ° C for 2-24 hours. Naturally cooled, the organic solvent is dissolved. If the substituted benzaldehyde is not adjacent to the carboxyl group, the oxidizing agent is added for oxidation for 10 minutes. If the substituted benzaldehyde is added ortho to the carboxyl group, it is not oxidized by adding an oxidizing agent.
  • the solvent was distilled off and separated by silica gel column chromatography.
  • the substituted benzaldehyde is ortho-position is a carboxyl group
  • the obtained target compound of the silicon-based rhodamine derivative can also be further derivatized by a substituent group to obtain a new silicon-based rhodamine derivative, that is, the following third step can also be included.
  • the third step further derivatization of the silicon-based rhodamine derivative
  • the silicon-based rhodamine derivative undergoes partial hydrolysis such as cyano group, complete hydrolysis of cyano group, reduction of cyano group, reduction of nitro group, azide reaction of amino group, and the like.
  • the cell dye and biological stain can be used as a staining agent for experimental animals, living cells or cells after fixation.
  • the invention adopts a novel synthesis method, can conveniently synthesize a silicon-based rhodamine derivative, can improve the synthesis efficiency by the method, and can realize the mass preparation of the silicon-based rhodamine derivative.
  • This synthesis method greatly enriches the selection range of substrates, and can synthesize some silicon-based rhodamine derivatives which are difficult to synthesize by the former method, which is beneficial to the further modification and application of the silicon-based rhodamine derivatives.
  • the invention also studies and applies the spectroscopic properties of the synthesized silicon-based rhodamine derivatives, and is used for living cell imaging research.
  • the invention designs a brand-new synthetic route, and synthesizes a series of novel silicon-based rhodamine derivatives which are difficult to synthesize by a novel method by a novel method, and the structure thereof is shown in Table 1 below.
  • the method also has the advantages of high total yield, simple reaction conditions, convenient operation, large amount of preparation, wide selection of substrate and the like.
  • Figure 1 is a graph showing the absorption spectrum of the compound of the present invention.
  • Figure 2 is a graph showing the fluorescence emission spectrum of the compound of the present invention.
  • Figure 3 is a graph showing the effect of the compound 2-COOH SiR prepared in Example 13 on the laser confocal fluorescence imaging of mouse brain microvascular endothelial cells (bEND3).
  • Example 4 is a diagram showing the effect of the compound 2-COOH SiR prepared in Example 13 of the present invention on laser confocal fluorescence imaging of human hepatoma cells (HepG2).
  • Figure 5 is a graph showing the effect of the compound 2,4-DiSO 3 H SiR prepared in Example 13 of the present invention on laser confocal fluorescence imaging of mouse brain microvascular endothelial cells (bEND3).
  • Figure 6 is a graph showing the effect of the compound 2,4-DiSO 3 H SiR prepared in Example 13 of the present invention on laser confocal fluorescence imaging of human hepatoma cells (HepG2).
  • the method was the same as in Example 4 except that the benzaldehyde in the raw material was changed to o-methylbenzaldehyde.
  • the target compound 2-Me SiR yield was 34%.
  • the method was the same as in Example 4 except that the benzaldehyde in the raw material was changed to o-bromobenzaldehyde.
  • the target compound 2-Br SiR was obtained in a yield of 43%.
  • the method was the same as in Example 4 except that the benzaldehyde in the raw material was changed to o-nitrobenzaldehyde.
  • the target compound 2-NO 2 SiR was obtained in a yield of 47%.
  • the target compound 2-SO 3 H SiR was obtained in a yield of 16%.
  • the method was the same as in Example 4 except that the benzaldehyde in the raw material was changed to 2-methyl-5-nitrobenzaldehyde.
  • the target compound 2-Me-5-NO 2 SiR was obtained in a yield of 59%.
  • Example 13 The procedure was the same as in Example 13 except that the o-aldehyde benzoic acid in the starting material was changed to 2-aldehyde-4-bromobenzoic acid for 8 hours.
  • the target compound 2-COOH-5-Br SiR was obtained in a yield of 40%.
  • the method was the same as in Example 13 except that the starting aldehyde benzoic acid was changed to 2-aldehyde-4-cyanobenzoic acid in the starting material for 3 hours.
  • the target compound 2-COOH-5-CN SiR was obtained in a yield of 31%.
  • the method was the same as in Example 13 except that the starting aldehyde benzoic acid was replaced by 2-aldehyde-5-nitrobenzoic acid in a raw material for 3 hours.
  • the target compound 2-COOH-4-NO 2 SiR was obtained in a yield of 31%.
  • Example 13 The procedure was the same as in Example 13 except that the o-aldehyde benzoic acid in the starting material was changed to 2-aldehyde-5-bromobenzoic acid for 5 hours.
  • the target compound 2-COOH-4-Br SiR was obtained in a yield of 37%.
  • the prepared sample solution was measured by ultraviolet spectrophotometer (Agilent Cary 100) (quartz dish, optical path length 1 cm), the measurement range was 450 nm to 750 nm, the maximum absorption wavelength was measured, and the molar absorption coefficient of the sample was calculated.
  • the results are shown in Table 2, and the absorption spectrum is shown in Figure 1.
  • the fluorescence emission spectrum of the prepared sample solution was measured by fluorescence spectrometer.
  • the excitation wavelength was 610 nm
  • the measurement range was 620 nm to 750 nm
  • the maximum emission wavelength was measured.
  • the crystal violet was used as a reference material to calculate the fluorescence quantum yield. The results are shown in the following table. 2.
  • the fluorescence emission spectrum of the sample is shown in Figure 2.
  • Test cells mouse brain microvascular endothelial cells (bEND3) and human liver cancer cells (HepG2) were used;
  • DMEM medium containing 10% FBS, 0.1 mg/mL streptomycin and 100 U/mL penicillin was used in a cell culture incubator containing 5% CO 2 , 95% air, 37 ° C constant temperature, and saturated humidity. Cultured cells. When the degree of cell polymerization reached 90%, the medium was aspirated and the cells were washed twice with PBS solution, digested with 0.25% trypsin at 37 ° C for 1 minute, neutralized by adding 2 volumes of medium, centrifuged at 1000 rpm for 5 minutes, and the cells were 1:3. The ratio was passaged to a Petri dish and the medium was changed every 2 days.
  • 2-COOH SiR was formulated with DMSO as a 1.0 mM solution; 2,4-DiSO 3 H SiR was dispensed with 1.0 mM solution in PBS buffer.
  • the bEND3 cells were seeded in a laser confocal culture dish at a density of 5 ⁇ 10 4 , and the cells were cultured in a cell incubator containing 5% CO 2 , 95% air, 37° C., and saturated humidity for 24 hours, and then the medium was discarded. Wash twice with pre-cooled PBS. Add 10 ⁇ L of 2-COOH SiR solution to 1.0 mL of fresh medium, mix well, add to the culture dish, incubate for 40 minutes in the incubator, then add DAPI stain to the medium, continue incubating for 20 minutes in the incubator. The cells were washed three times with PBS buffer and observed under a laser confocal microscope. The excitation light sources were 488 nm and 633 nm.
  • the laser confocal fluorescence imaging is shown in Figure 3.
  • the 2-COOH SiR staining method for HepG2 cells is consistent with the bEND3 cell staining method.
  • the laser confocal fluorescence imaging is shown in Figure 4.
  • 2,4-DiSO 3 H SiR vs bEND3 cells The HepG2 cell staining method is consistent with 2-COOH SiR, and the laser confocal fluorescence imaging is shown in Fig. 5 and Fig. 6.

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Abstract

本发明涉及一种硅基罗丹明衍生物，所述硅基罗丹明衍生物的制备方法及其在制备细胞染料、生物染色剂和生物分子、粒子荧光标记中的应用。本发明提供的硅基罗丹明衍生物合成方法具有总产率高、反应条件简单、操作方便、能够大量制备、底物选择范围广等优点。

Description

一种硅基罗丹明衍生物及其制备方法和应用 技术领域
本发明涉及化学领域，具体地说，涉及一种硅基罗丹明衍生物及其制备方法和在制备细胞染料、生物染色剂、生物分子或生物粒子荧光标记中的应用。
背景技术
罗丹明是氧杂蒽类化合物中的一种，具有光稳定性好、荧光量子产率高、化学修饰简单等优点，是一种较好的荧光小分子染料探针，广泛应用于生物样品的染色与荧光标记中。但是罗丹明作为荧光探针在应用过程中，特别是应用于生物样品时，也存在着许多不足。比如罗丹明B染料的激发波长位于紫外可见光区，而生物样品在紫外可见光区会产生严重的背景信号干扰。另一方面，紫外可见光对组织的穿透能力较差，对生物样品的光毒性较大。这些因素大大影响了罗丹明染料探针在生物样品荧光分析中灵敏度和准确性，极大地限制了其在生物样品荧光成像中的进一步应用。
硅基罗丹明是近几年新发展起来的一类荧光染料。其优势在于通过将罗丹明分子中的氧原子替换为硅原子，在保留了罗丹明类化合物优点的同时，其光谱范围红移，达到近红外光区，从而满足了生物样品分析成像的要求。近几年来不断有硅基罗丹明方面的专利发表，例如专利WO 2010/126077 A1，专利WO 2012/111818 A1和专利WO 2013/029650 A1等。这些专利对硅基罗丹明的性质、合成方法和应用等进行了叙述。在这些专利中，都是采用活泼金属试剂与羰基亲核加成的方法制备硅基罗丹明衍生物，这种合成方法一方面步骤较多，反应条件苛刻，产率低，不能用于大量的合成；另一方面活性金属试剂的使用大大限制了底物的选择范围，其中一些取代硅基罗丹明甚至无法合成，影响了其在生物样品成像方面的应用。因此，设计一种步骤少、反应条件简单、产率高、底物选择多样、能适实现大量制备的方法用于合成硅基罗丹明衍生物是十分必要的。
发明内容
本发明的目的是针对现有技术中的不足，提供一种硅基罗丹明衍生物。
本发明的再一的目的是，提供上述硅基罗丹明衍生物的制备方法。
本发明的另一的目的是，提供上述硅基罗丹明衍生物的用途。
为实现上述目的，本发明采取的技术方案是：
一种硅基罗丹明衍生物，其结构通式如(I)所示：
Figure PCTCN2015073677-appb-000001
特别的，当R11或R15其中任意一个或两个为-COOH或-COO-时，可以形成通式(I)相应的内酯螺环形式，具有通式(II)结构的化合物：
Figure PCTCN2015073677-appb-000002
其中，R2、R3分别为C1-C6直链或支链的烃基或苯基，所述烃基选自：甲基、乙基、正丙基、异丙基、正丁基、异丁基、仲丁基、叔丁基、正戊基、异戊基、仲戊基、新戊基、正己基，优选的是甲基、乙基和苯基，最优选的是甲基。
R4、R5、R6、R7、R8、R9分别为氢或C1-C6的直链或支链的烃基，所述烃基选自：甲基、乙基、正丙基、异丙基、正丁基、异丁基、仲丁基、叔丁基、正戊基、异戊基、仲戊基、新戊基、正己基，优选的是甲基、乙基和苯基，最优选的是氢。
R20、R21、R22、R23分别为C1-C6直链或支链的饱和或不饱和的烃基或环烷基，所述烃基选自：甲基、乙基、正丙基、异丙基、烯丙基、正丁基、异丁基、仲丁基、叔丁基、正戊基、异戊基、仲戊基、新戊基、正己基，优选的是甲基、乙基、烯丙基，最优选乙基。
R20或R21可以与R5以碳链相连，与母体苯环形成五元或六元环状结构，优选五元环。
R22或R23可以与R8以碳链相连，与母体苯环形成五元或六元环状结构，优选五元环。
优选地，取代基-NR20R21可以与-R5形成具有以下通式(III)的结构：
Figure PCTCN2015073677-appb-000003
优选地，所述取代基-NR22R23可以与-R8形成具有以下通式(IV)的结构：
Figure PCTCN2015073677-appb-000004
优选地，所述取代基R24、R25为氢、甲基、乙基或烯丙基，更优选为乙基。
R11，R12，R13，R14，R15，R16，R17，R18，R19可以为相同或者不同的任何取代基，所述取代基选自：烃基、卤素、氰基、硝基、烷氧基、卤代烷基。优选的结构为下列中的一种：
R11或者R15一个为氢，另一个为-CN、-NO2、-SO3H、-SO3-、-OH、-OCH3、-NH2、-N3、-NCS、-CH3、-F、-Cl、-Br、-I基团中的一个，R12，R13，R14均为氢。
R11或者R15一个为氢，另一个为-CH3，R12，R13，R14中任意一个为-CN、-NO2、-SO3H、-SO3-、-OH、-OCH3、-NH2、-CH3、-N3、-NCS、-C≡CH、-CONH2、-CH2NH2、-F、-Cl、-Br、-I、-COOH、-COO-中的任一个，其余两个为氢。
R11或者R15一个为氢，另一个为-SO3H或-SO3-，R12，R13，R14中任意一个为-SO3H、-SO3-基团中的一个，其余两个为氢。
R11或者R15一个为氢，另一个为-COOH或者-COO-，R12，R13，R14中任意一个为-CN、-NO2、-SO3H、-SO3-、-OH、-OCH3、-NH2、-CH3、-N3、-NCS、-C≡CH、-CONH2、-CH2NH2、-F、-Cl、-Br、-I、-COOH、-COO-基团中的一个，其余两个为氢。
R11或者R15至少有一个为-COOH或者-COO-时，形成的内酯螺环结构中，R16，R17，R18，R19中任意一个为-CN、-NO2、-SO3H、-SO3-、-OH、-OCH3、-NH2、-CH3、-N3、-NCS、-C≡CH、-CONH2、-CH2NH2、-F、-Cl、-Br、-I、-COOH、-COO-基团中的一个，其余三个为氢。
为实现上述第二个目的，本发明采取的技术方案是.：
所述硅基罗丹明衍生物制备方法是按如下步骤进行的：
第一步：具有通式(V)结构的关键硅基中间体的合成：
Figure PCTCN2015073677-appb-000005
当-NR20R21与-NR22R23，R4与R7，R5与R8，R6与R9均具有相同结构，采用如下方法合成：
N,N-二烃基间溴苯胺衍生物与正丁基锂反应生成相应的锂试剂，再进一步与二烃基二氯硅烷反应得到关键硅基中间体。其反应过程可以表示为：
Figure PCTCN2015073677-appb-000006
其中R2，R3，R4～R9，R20～R23含义同前。
具体的操作方法优选是：1.0摩尔量N,N-二烃基间溴苯胺衍生物与1.05摩尔量的正丁基锂0℃下在干燥的乙醚中反应2小时，再将0.6摩尔量的二烃基二氯硅烷溶于干燥的乙醚中加入到反应液中，0℃慢慢升至室温，反应过夜。加入水淬灭反应，萃取，蒸除溶剂，硅胶柱层析分离，洗脱剂为石油醚和乙酸乙酯的混合溶剂，以体积计，乙酸乙酯:石油醚＝1:40～80，得所述的关键硅基中间体。
当-NR20R21与-NR22R23，R4与R7，R5与R8，R6与R9中有一对或多对具有不同结构时，采用如下方法合成：
一种N,N-二烃基间溴苯胺衍生物与正丁基锂反应生成相应的锂试剂，再与二烃基二氯硅烷反应，取代其中的一个氯，生成单氯硅烷的产物。另一种N,N-二烃基间溴苯胺衍生物与丁基锂反应生成相应的锂试剂，再进一步与单氯硅烷产物反应得到关键硅基中间体。其反应过程可以表示为：
Figure PCTCN2015073677-appb-000007
其中R2，R3，R4～R9，R20～R23含义同前；
具体操作方法优选是：一种1.0摩尔量N,N-二烃基间溴苯胺衍生物与1.05摩尔量的正丁基锂0℃下在干燥的乙醚中反应2小时，将其加入含有5.0摩尔量的二烃基二氯硅烷的干燥乙醚中，0℃下反应2小时，制得单氯代硅烷产物；另一种0.5摩尔量的N,N-二烃基间溴苯胺衍生物与0.525摩尔量的正丁基锂0℃下在干燥的乙醚中反应2小时，0℃下将该反应液缓慢加入到单氯代硅烷产物中，缓慢升至室温，继续反应12小时，加入水淬灭反应，萃取，蒸除溶剂，硅胶柱层析分离，洗脱剂为石油醚和乙酸乙酯的混合溶剂，以体积计，乙酸乙酯:石油醚＝1:60，得关键硅基中间体。
第二步：关键硅基中间体与不同取代苯甲醛生成硅基罗丹明衍生物的反应：
关键硅基中间体与不同取代的苯甲醛衍生物在催化剂条件下，封管后置于防护罩中，140℃下反应，再经过四氯苯醌氧化，或者不氧化，最后进行柱层析分离，以二氯甲烷 和甲醇的混合体系或者乙酸乙酯、石油醚和三乙胺的混合体系进行洗脱，得到硅基罗丹明衍生物。其反应过程可以表示为：
Figure PCTCN2015073677-appb-000008
当取代苯甲醛的邻位有羧基时，其反应过程还可以表示为：
Figure PCTCN2015073677-appb-000009
其中R2，R3，R4～R9，R11-R19，R20～R23含义同前。
所述具有通式(V)结构的关键硅基中间体与不同取代的苯甲醛衍生物的投料摩尔比为1:5。
所述催化剂选用CuBr2或者对甲苯磺酸一水合物。
所述具有通式(V)结构的关键硅基中间体与催化剂的摩尔比为1:0.1～1。
所述柱层析分离的洗脱剂，以体积计，甲醇和二氯甲烷的混合溶剂为甲醇:二氯甲烷＝1:5～40，乙酸乙酯和石油醚混合溶剂为乙酸乙酯:石油醚＝1:40～80，乙酸乙酯、石油醚和三乙胺的混合溶剂为乙酸乙酯:石油醚＝1:10～80，三乙胺的加入量为乙酸乙酯和石油醚混合物体积的1％。
具体操作方法优选是：将1.0摩尔量的关键硅基中间体、5.0摩尔量的取代苯甲醛、0.1～1.0摩尔量的催化剂加入厚壁耐压反应管中，封管后置于防护罩中，140℃下反应2-24小时。自然冷却，有机溶剂溶解，若加入的取代苯甲醛邻位不是羧基，则加入氧化剂氧化10分钟，若加入的取代苯甲醛邻位是羧基，不用加氧化剂氧化。蒸除溶剂，硅胶柱层析分离。若加入的取代苯甲醛邻位不是羧基，洗脱剂为二氯甲烷和甲醇的混合溶剂，以体积计，甲醇:二氯甲烷＝1:5～40，得目标化合物。若加入的取代苯甲醛邻位是羧基，洗脱剂为石油醚、乙酸乙酯和三乙胺的混合溶剂，以体积计，乙酸乙酯:石油醚＝1:10～80，三乙胺的加入量为乙酸乙酯和石油醚混合物体积的1％，得到目标化合物。
所得到的目标化合物硅基罗丹明衍生物还可以通过取代基团进一步的衍生化反应得到新的硅基罗丹明衍生物，即还可以包含以下第三步。
第三步硅基罗丹明衍生物进一步衍生化反应；
在常规条件下，所述的硅基罗丹明衍生物进行如氰基的部分水解、氰基的完全水解、氰基的还原、硝基的还原、氨基的叠氮化反应等。
为实现上述第三个目的，本发明采取的技术方案是：
所述的硅基罗丹明衍生物在制备细胞染料、生物染色剂、生物分子或生物粒子荧光标记中的应用。所述的细胞染料、生物染色剂可用于实验动物、活细胞或固定后细胞的染色剂。
本发明用一种全新的合成方法，可以方便的合成硅基罗丹明衍生物，通过这种方法可以提高合成效率，能够实现硅基罗丹明衍生物的大量制备。最主要的是这种合成方法大大丰富了底物的选择范围，可以合成一些采用前方法难以合成的硅基罗丹明衍生物，有利于硅基罗丹明衍生物的进一步修饰与应用。
通过本发明的方法合成若干新型的硅基罗丹明衍生物，以满足其在不同领域的应用。
本发明还对合成的硅基罗丹明衍生物进行光谱学性质的研究和应用，并用于活体细胞成像研究。
本发明优点在于：
本发明设计了一条全新的合成路线，通过一种全新的方法合成得到了一系列采用前方法难于合成的新型硅基罗丹明类衍生物，其结构如下表1所示。该方法还具有总产率高、反应条件简单、操作方便、能够大量制备、底物选择范围广等优点。
表1 本发明首次合成的硅基罗丹明类衍生物结构式
Figure PCTCN2015073677-appb-000010
Figure PCTCN2015073677-appb-000011
Figure PCTCN2015073677-appb-000012
附图说明
附图1为本发明中化合物的吸收光谱图。
附图2为本发明中化合物的荧光发射光谱图。
附图3为本发明中实施例13制备的化合物2-COOH SiR对小鼠脑微血管内皮细胞(bEND3)染色激光共聚焦荧光成像效果图。
附图4为本发明实施例13制备的化合物2-COOH SiR对人肝癌细胞(HepG2)染色激光共聚焦荧光成像效果图。
附图5为本发明实施例13制备的化合物2,4-DiSO3H SiR对小鼠脑微血管内皮细胞(bEND3)染色激光共聚焦荧光成像效果图。
附图6为本发明实施例13制备的化合物2,4-DiSO3H SiR对人肝癌细胞(HepG2)染色激光共聚焦荧光成像效果图。
图3～6中，(a)均表示激发光为633nm时激光共聚焦荧光成像效果图；(b)均表示激发光为488nm时激光共聚焦荧光成像效果图；(c)均表示细胞明视野效果图；(d)均表示(a)、(b)、(c)的叠加效果图。
具体实施方式
下面用具体实施例对本发明中的硅基罗丹明衍生物的合成方法、采用该方法所合成的具有全新结构的硅基罗丹明衍生物及将其应用于活细胞激光共聚焦显微镜成像做进一步详细说明，所述是对本发明的解释而不是限定。
实施例1
Figure PCTCN2015073677-appb-000013
在装有磁力搅拌子的500mL长颈反应瓶中，加入N,N-二乙基间溴苯胺27.376g(120.0mmol)。氩气保护条件下，加入无水乙醚120mL，0℃下磁力搅拌5min。将n-BuLi 52.5mL(2.4M in n-hexane,126.0mmol)缓慢滴加到反应液中，滴加完毕后，0℃下继续反应2h。将二甲基二氯硅烷8.8mL(72.0mmol)溶于30mL无水乙醚中，缓慢滴加到上 述反应液中。滴加完毕后，反应缓慢升至室温，搅拌过夜。反应液中加入水150mL，分液漏斗分取有机层，水层用乙醚萃取(50mL×3)，合并有机层，150mL水洗涤一次，150mL饱和NaCl水溶液洗涤一次，无水Na2SO4干燥，蒸除溶剂，硅胶柱层析纯化，洗脱剂以体积计为石油醚:乙酸乙酯＝80:1，得到黄色油状物SiN117.507g，产率82％。
其核磁1H NMR(300MHz,CDCl3):δ0.64(s,6H)，1.23(t,12H,J＝7.2Hz)，3.43(q,8H,J＝7.2Hz)，6.79-7.35(m,8H)；核磁13C NMR(75MHz,CDCl3):δ-2.13,12.67,44.45,112.79,117.75,121.47,128.76,139.11,147.10；质谱MS(ESI)calcd.for C22H34N2Si[M+H]+:355.26,found:355.32。
实施例2
Figure PCTCN2015073677-appb-000014
在装有磁力搅拌子的250mL长颈反应瓶中，加入1-乙基-6-溴吲哚啉4.070g(18.0mmol)。氩气保护条件下，加入无水乙醚50mL，0℃下磁力搅拌5min。将n-BuLi 11.8mL(1.6M in n-hexane,18.9mmol)缓慢滴加到反应液中，滴加完毕后，0℃下继续反应2h。将二甲基二氯硅烷1.3mL(10.8mmol)溶于10mL无水乙醚中，缓慢滴加到上述反应液中。滴加完毕后，反应缓慢升至室温，搅拌过夜。反应液中加入水50mL，分取有机层，水层用乙醚萃取(30mL×3)，合并有机层，50mL水洗涤一次，50mL饱和NaCl水溶液洗涤一次，无水Na2SO4干燥，旋蒸除溶剂，硅胶柱层析纯化，洗脱剂以体积计为石油醚:乙酸乙酯＝40:1，得到黄色油状物SiN2，2.423g，产率77％。
其核磁1HNMR(300MHz,CDCl3):δ0.49(s,6H),1.16(t,6H,J＝7.2Hz),2.94(t,4H,J＝8.1Hz),3.12(q,4H,J＝7.2Hz),3.30(t,4H,J＝8.4Hz),6.65-7.08(m,6H)；核磁13C NMR(75MHz,CDCl3):δ-1.80,12.09,28.57,43.17,52.20,112.39,124.00,124.07,131.64,137.17,151.80；质谱MS(ESI)calcd.for C22H30N2Si[M+H]+:351.23,found:351.26。
实施例3
Figure PCTCN2015073677-appb-000015
在装有磁力搅拌子的100mL长颈反应瓶中，加入N,N-二乙基间溴苯胺2.738g(12.0 mmol)。氩气保护条件下，加入无水乙醚10mL，0℃下磁力搅拌5min。将n-BuLi 7.9mL(1.6M in n-hexane,12.6mmol)缓慢滴加到反应液中，滴加完毕后，0℃下继续反应2h。将二甲基二氯硅烷7.3mL(60.0mmol)溶于30mL无水乙醚中，缓慢导入上述反应液中。滴加完毕后，反应缓慢升至室温，并室温下反应2小时。减压条件下除去溶剂和未反应的二甲基二氯硅烷，得到SiNCl粗品，氩气保护下加入无水乙醚5mL溶解SiNCl粗品，所得溶液备用。
在另一装有磁力搅拌子的50mL长颈反应瓶中，加入1-乙基-6-溴吲哚啉1.357g(6.0mmol)。氩气保护条件下，加入无水乙醚10mL，0℃下磁力搅拌5min。将n-BuLi 4.0mL(1.6M in n-hexane,6.3mmol)缓慢滴加到反应液中，滴加完毕后，0℃下继续反应2h。冰浴下，将该反应液缓慢滴加到之前制备的SiNCl溶液中，滴加完毕后，缓慢升至室温，搅拌过夜。反应液中加入水30mL，分液漏斗分取有机层，水层用乙醚萃取(20mL×3)，合并有机层，50mL水洗涤一次，50mL饱和NaCl水溶液洗涤一次，无水Na2SO4干燥，蒸除溶剂。硅胶柱层析纯化，洗脱剂以体积计为石油醚:乙酸乙酯＝60:1，得黄色油状物SiN3，1.116g，产率53％。
其核磁1HNMR(300MHz,CDCl3):δ0.50(s,6H),1.12(t,6H,J＝7.2Hz),1.16(t,3H,J＝7.2Hz),2.93(t,2H,J＝8.1Hz),3.11(q,2H,J＝7.2Hz),3.30(t,2H,J＝8.1Hz),3.32(q,4H,J＝7.2Hz),6.70-7.23(m,7H)；核磁13C NMR(75MHz,CDCl3):δ-2.01,12.13,12.60,28.56,43.17,44.38,52.22,112.34,112.73,117.72,121.45,123.99,124.02,128.68,131.64,137.00,139.20,147.05,151.84；质谱MS(ESI)calcd.for C22H32N2Si[M+H]+:353.60,found:353.49。
实施例4
Figure PCTCN2015073677-appb-000016
在装有磁力搅拌子的15mL厚壁耐压反应管中，加入原料SiN1 106mg(0.3mmol)，苯甲醛159mg(1.5mmol)，对甲苯磺酸一水合物57mg(0.3mmol)，封管后置于保护罩中，140℃油浴反应12小时，自然冷却至室温，加入二氯甲烷和甲醇各2mL溶解，加入足量四氯苯醌氧化，滤除不溶物，硅胶柱层析纯化，洗脱剂以体积计为二氯甲烷:甲醇＝40:1，得到目标化合物2-H SiR，产率40％。
其核磁1H NMR(300MHz,CD3OD)δ0.59(s,6H),1.29(t,12H,J＝6.9Hz),3.70(q,8H, J＝6.6Hz),6.74-7.55(m,10H)；核磁13C NMR(75MHz,CD3OD)δ-2.67,11.72,45.41,113.42,120.65,127.36,127.91,128.37,129.01,139.17,142.03,148.15,152.50,169.07；高分辨质谱HRMS(ESI)calcd.for C29H37N2Si+[M]+:441.2721,found:441.2765。
实施例5
Figure PCTCN2015073677-appb-000017
方法同实施例4，只是将原料中苯甲醛换为邻甲基苯甲醛。得到目标化合物2-Me SiR产率34％。
其核磁1H NMR(600MHz,CD3OD)δ0.51(s,3H),0.63(s,3H),1.31(t,12H,J＝7.2Hz),2.06(s,3H),3.73(q,8H,J＝7.2Hz),6.77-7.48(m,10H)；核磁13C NMR(150MHz,CD3OD)δ-2.90,-2.65,11.71,18.09,45.41,113.74,120.60,125.39,126.94,128.62,128.76,129.92,135.58,138.66,141.13,148.09,152.64,168.96；高分辨质谱HRMS(ESI)calcd.for C30H39N2Si+[M]+:455.2877,found:455.2888。
实施例6
Figure PCTCN2015073677-appb-000018
方法同实施例4，只是将原料中苯甲醛换为邻溴苯甲醛。得到目标化合物2-Br SiR，产率43％。
其核磁1H NMR(300MHz,CDCl3)δ0.56(s,3H),0.61(s,3H),1.16(t,12H,J＝7.2Hz),3.37(q,8H,J＝7.2Hz),6.59-7.74(m,10H)；核磁13C NMR(75MHz,CDCl3)δ-1.68,-0.92,12.84,46.08,113.91,120.70,122.69,126.99,127.48,130.48,130.74,133.04,139.55,141.45,148.39,152.51,166.83；高分辨质谱HRMS(ESI)calcd.for C29H36BrN2Si+[M]+:519.1826,found:519.1827,521.1810。
实施例7
Figure PCTCN2015073677-appb-000019
方法同实施例4，只是将原料中苯甲醛换为邻硝基苯甲醛。得到目标化合物2-NO2SiR，产率47％。
其核磁1H NMR(300MHz,CD3OD)δ0.58(s,3H),0.63(s,3H),1.28(t,12H,J＝7.2Hz),3.70(q,8H,J＝6.9Hz),6.73-8.41(m,10H)；核磁13C NMR(75MHz,CD3OD)δ-3.31,-2.45,11.70,45.46,113.92,120.64,124.89,126.68,130.39,131.79,133.65,134.42,140.04,147.87,148.37,152.62,165.10；高分辨质谱HRMS(ESI)calcd.for C29H36N3O2Si+[M]+:486.2571,found:486.2588。
实施例8
Figure PCTCN2015073677-appb-000020
方法同实施例4，只是将原料中苯甲醛换为苯甲醛-2-磺酸钠，洗脱剂以体积计为二氯甲烷:甲醇＝10:1。得到目标化合物2-SO3H SiR，产率16％。
其核磁1H NMR(600MHz,CD3OD)δ0.60(s,6H),1.30(t,12H,J＝7.2Hz),3.70(q,8H,J＝7.2Hz),6.70-8.17(m,10H)；核磁13C NMR(150MHz,CD3OD)δ-2.93,-2.62,11.72,45.23,113.01,119.78,127.84,128.15,128.52,129.27,130.11,136.40,142.58,144.09,148.02,152.39,170.25；高分辨质谱HRMS(ESI)calcd.for C29H37N2O3SSi+[M+H]+:521.2289,found:521.2301；C39H36N2O3SSiNa+[M+Na]+:543.2108,found:543.2116。
实施例9
Figure PCTCN2015073677-appb-000021
方法同实施例4，只是将原料中苯甲醛换为2-甲基-4溴苯甲醛。得到目标化合物2-Me-4-Br SiR，产率43％。
其核磁1H NMR(300MHz,CD3OD)δ0.59(s,3H),0.60(s,3H),1.29(t,12H,J＝7.2 Hz),2.04(s,3H),3.71(q,8H,J＝7.2Hz),6.77-7.61(m,9H)；核磁13C NMR(75MHz,CD3OD)δ-2.91,-2.70,11.70,17.92,45.44,113.91,120.78,122.25,126.63,128.59,130.53,132.74,137.76,138.30,140.79,148.01,152.65,166.93；高分辨质谱HRMS(ESI)calcd.for C30H38BrN2O2Si+[M]+:533.1982,found:533.1978,535.1966。
实施例10
Figure PCTCN2015073677-appb-000022
方法同实施例4，只是将原料中苯甲醛换为2-甲基-5-硝基苯甲醛。得到目标化合物2-Me-5-NO2SiR，产率59％。
其核磁1H NMR(300MHz,CD3OD)δ0.61(s,3H),0.64(s,3H),1.30(t,12H,J＝7.2Hz),2.18(s,3H),3.73(q,8H,J＝7.2Hz),6.77-8.36(m,9H)；核磁13C NMR(75MHz,CD3OD)δ-2.91,-2.70,11.70,18.25,45.51,114.17,121.13,123.29,123.63,126.39,131.35,139.97,140.40,143.89,146.24,148.10,152.71,164.49；高分辨质谱HRMS(ESI)calcd.for C30H38N3O2Si+[M]+:500.2728,found:500.2729。
实施例11
Figure PCTCN2015073677-appb-000023
方法同实施例4，只是将原料中苯甲醛换为3-甲基-4醛基苯甲酸。得到目标化合物2-Me-4-COOH SiR，产率33％。
其核磁1H NMR(300MHz,CD3OD)δ0.60(s,3H),0.62(s,3H),1.29(t,12H,J＝7.2Hz),2.12(s,3H),3.72(q,8H,J＝7.2Hz),6.76-8.07(m,9H)；核磁13C NMR(75MHz,CD3OD)δ-2.92,-2.69,11.69,18.04,45.44,113.94,120.82,126.34,126.72,129.16,131.08,131.15,136.29,140.70,143.43,148.00,152.68,167.18,167.84；高分辨质谱HRMS(ESI)calcd.for C31H39N2O2Si+[M]+:499.2775,found:499.2759。
实施例12
Figure PCTCN2015073677-appb-000024
方法同实施例4，只是将原料中苯甲醛换为苯甲醛-2,4-二磺酸钠，洗脱剂以体积计为二氯甲烷:甲醇＝5:1，得到目标化合物2,4-DiSO3H SiR，产率10％。
其核磁1H NMR(300MHz,CD3OD)δ0.57(s,3H),0.58(s,3H),1.25(t,12H,J＝7.2Hz),2.02(s,1H),3.67(q,8H,J＝6.9Hz),6.71-8.65(m,9H)；核磁13C NMR(75MHz,CD3OD)δ-2.98,-2.60,11.70,45.75,113.65,120.49,125.70,126.87,127.88,130.63,135.62,137.01,138.25,141.47,142.42,147.89,152.00高分辨质谱HRMS(ESI)calcd.for C29H36N2O6S2Si[M+H]+:601.1857,found:601.1867；[M+Na]+:623.1676,found:623.1681。
实施例13
Figure PCTCN2015073677-appb-000025
在装有磁力搅拌子的15mL厚壁耐压反应管中，加入原料SiN1106mg(0.3mmol)，邻醛基苯甲酸225mg(1.5mmol)，溴化铜7mg(0.03mmol)，封管后置于防护罩中，140℃油浴反应5小时，自然冷却至室温，加入二氯甲烷和甲醇各2mL溶解，硅胶柱层析纯化，洗脱剂以体积计为石油醚:乙酸乙酯:三乙胺＝80:1:1，得到目标化合物2-COOH SiR，产率52％。
其核磁1H NMR(300MHz,CDCl3)δ0.64(s,3H),0.66(s,3H),1.18(t,12H,J＝7.2Hz),3.39(q,8H,J＝7.2Hz),6.50-8.01(m,10H)；核磁13C NMR(75MHz,CDCl3)δ-1.78,0.44,12.55,44.28,92.30,112.45,115.94,124.86,125.58,127.48,128.46,128.65,130.79,133.50,137.30,146.58,154,15,170.63；高分辨质谱HRMS(ESI)calcd.for C30H36N2O2Si[M+H]+:485.2619,found:485.2623。
实施例14
Figure PCTCN2015073677-appb-000026
方法同实施例13，只是将原料中SiN1换为SiN2，洗脱剂以体积计为石油醚:乙酸乙酯:三乙胺＝70:1:1，得到目标化合物2-COOH SiR2，产率16％。
其核磁1H NMR(300MHz,CDCl3):δ0.54(s,3H),0.58(s,3H),1.18(t,12H,J＝7.2Hz),2.70-2.88(m,4H),3.18(q,4H,J＝7.2Hz),3.29(t,4H,J＝8.4Hz),6.63-7.96(m,8H)；13C NMR(75MHz,CDCl3):δ-0.39,0.06,12.04,28.37,42.67,51.70,92.06,109.97,123.47,123.83,125.49,125.77,128.37,132.54,133.03,133.95,134.07,151.37,156.39,171.46；高分辨质谱HRMS(ESI)calcd.for C30H32N2O2Si[M+H]+:481.2306,found:481.2307。
实施例15
Figure PCTCN2015073677-appb-000027
方法同实施例13，只是将原料中SiN1换为SiN3，洗脱剂以体积计为石油醚:乙酸乙酯:三乙胺＝70:1:1，得到目标化合物2-COOH SiR3，产率31％。
其核磁1H NMR(300MHz,CDCl3):δ0.54(s,3H),0.58(s,3H),1.18(t,12H,J＝7.2Hz),2.70-2.88(m,4H),3.18(q,4H,J＝7.2Hz),3.29(t,4H,J＝8.4Hz),6.63-7.96(m,8H)；核磁13C NMR(75MHz,CDCl3):δ-0.39,0.06,12.04,28.37,42.67,51.70,92.06,109.97,123.47,123.83,125.49,125.77,128.37,132.54,133.03,133.95,134.07,151.37,156.39,171.46；高分辨质谱HRMS(ESI)calcd.for C30H32N2O2Si[M+H]+:481.2306,found:481.2307。
实施例16
Figure PCTCN2015073677-appb-000028
方法同实施例13，只是将原料中邻醛基苯甲酸换为2-醛基-4-溴苯甲酸，反应时间8小时。得到目标化合物2-COOH-5-Br SiR，产率40％。
其核磁1H NMR(300MHz,CDCl3)δ0.59(s,3H),0.63(s,3H),1.17(t,12H,J＝7.2Hz), 3.37(q,8H,J＝7.2Hz),6.51-7.83(m,9H)；核磁13C NMR(75MHz,CDCl3)δ-1.36,0.26,12.58,44.30,91.75,112.78,115.82,125.93,126.80,127.79,128.49,128.94,129.87,132.20,136.84,146.67,156.71,169.95；高分辨质谱HRMS(ESI)calcd.for C30H35BrN2O4Si[M+H]+:563.1724,found:563.1740,565.1725。
实施例17
Figure PCTCN2015073677-appb-000029
方法同实施例13，只是将原料中邻醛基苯甲酸换为2-醛基-4-氰基苯甲酸，反应时间3小时，洗脱剂以体积计为石油醚:乙酸乙酯:三乙胺＝60:1:1。得到目标化合物2-COOH-5-CN SiR，产率31％。
其核磁1H NMR(300MHz,CDCl3):δ0.63(s,3H),0.68(s,3H),1.21(t,12H,J＝6.9Hz),3.41(q,8H,J＝6.9Hz),6.50-8.01(m,9H)；核磁13C NMR(75MHz,CDCl3):δ-1.46,0.27,12.54,44.31,92.59,112.68,115.95,117.16,117.77,126.43,128.32,128.65,129.01,130.54,132.22,137.03,146.80,155.29,168.96；高分辨质谱HRMS(ESI)calcd.for C31H35N3O2Si[M+H]+:510.2571,found:510.2581。
实施例18
Figure PCTCN2015073677-appb-000030
方法同实施例13，只是将原料中邻醛基苯甲酸换为2-醛基-5-硝基苯甲酸，反应时间3小时，洗脱剂以体积计为石油醚:乙酸乙酯:三乙胺＝70:1:1。得到目标化合物2-COOH-4-NO2SiR，产率31％。
其核磁1H NMR(300MHz,CDCl3):δ0.59(s,3H),0.63(s,3H),1.16(t,12H,J＝7.2Hz),3.37(q,8H,J＝7.2Hz),6.48-8.80(m,9H)；核磁13C NMR(75MHz,CDCl3):δ-1.36,0.18,12.52,44.27,92.56,112.73,115.88,121.31,125.70,128.38,128.52,128.73,136.82,146.84,148.57,160.51,168.49；高分辨质谱HRMS(ESI)calcd.for C30H35N3O4Si[M+H]+:530.2470,found:530.2481。
实施例19
Figure PCTCN2015073677-appb-000031
方法同实施例13，只是将原料中邻醛基苯甲酸换为2-醛基-4-炔基苯甲酸，反应时间2小时。得到目标化合物2-COOH-5-CCH SiR，产率16％。
其核磁1H NMR(300MHz,CDCl3)δ0.59(s,3H),0.62(s,3H),1.15(t,12H,J＝7.2Hz),3.22(s,1H),3.36(q,8H,J＝7.2Hz),6.48-7.92(m,9H)；核磁13C NMR(75MHz,CDCl3)δ-1.55,0.30,12.55,44.25,80.57,82.69,92.10,112.60,115.82,125.42,127.14,127.50,128.28,128.48,130.12,132.47,136.98,146.62,154.60,169.94；高分辨质谱HRMS(ESI)calcd.for C32H36N2O2Si[M+H]+:509.2619,found:509.2614。
实施例20
Figure PCTCN2015073677-appb-000032
方法同实施例13，只是将原料中邻醛基苯甲酸换为2-醛基-5-溴苯甲酸，反应时间5小时。得到目标化合物2-COOH-4-Br SiR，产率37％。
其核磁1H NMR(300MHz,CDCl3)δ-0.59(s,3H),0.62(s,3H),1.14(t,12H,J＝6.9Hz),3.35(q,8H,J＝7.2Hz),6.46-8.09(m,9H)；核磁13C NMR(75MHz,CDCl3)δ-1.82,0.42,12.56,44.30,92.65,112.46,115.99,122.58,126.45,128.42,128.45,129.61,130.01,136.57,137.38,146.68,152.95,169.09。
实施例21
Figure PCTCN2015073677-appb-000033
方法同实施例13，只是将原料中邻醛基苯甲酸换为2-醛基-5-碘苯甲酸，反应时间5小时。得到目标化合物2-COOH-4-I SiR，产率36％。
其核磁1H NMR(300MHz,CDCl3)δ-0.60(s,3H),0.62(s,3H),1.16(t,12H,J＝6.9Hz),3.37(q,8H,J＝6.9Hz),6.47-8.31(m,9H)；核磁13C NMR(75MHz,CDCl3)δ-1.83,0.39,12.55,44.27,92.63,93.50,112.42,115.93,126.61,128.42,129.71,129.95,134.54,137.32,142.19,146.67,153.65,168.91。
实施例22
Figure PCTCN2015073677-appb-000034
在装有磁力搅拌子的10mL茄形瓶中加入2-COOH-5-CN SiR 51mg(0.10mmol)，碳酸钾17mg(0.12mmol)，DMSO 5mL。磁力搅拌，再加入双氧水(30％水溶液)200μL。60℃反应1小时，将反应液倒入25mL冰水中，二氯甲烷萃取三次，合并有机层，有机层水洗三次，无水硫酸钠干燥，蒸除溶剂，硅胶柱层析，洗脱剂以体积计为石油醚:乙酸乙酯:三乙胺＝40:1:1，得到目标化合物2-COOH-5-CONH2SiR，产率67％。
其核磁1H NMR(300MHz,CDCl3)δ0.58(s,3H),0.63(s,3H),1.15(t,12H,J＝6.9Hz),3.35(q,8H,J＝6.9Hz),6.46-8.01(m,9H)；核磁13C NMR(75MHz,CDCl3)δ-1.52,0.30,12.54,44.25,92.55,112.67,115.83,123.55,125.85,127.93,128.55,129.83,130.01,137.06,138.40,146.68,155.15,168.09,169.81；高分辨质谱HRMS(ESI)calcd.for C31H37N3O3Si[M+H]+:528.2677,found:528.2676。
实施例23
Figure PCTCN2015073677-appb-000035
在装有搅拌子的50mL反应瓶中，加入2-COOH-4-NO2SiR 106mg(0.20mmol)，甲醇15mL，二氯甲烷10mL，10％Pd/C 50mg。在H2气氛中室温反应4小时，减压过滤除去不溶物，加入足量的四氯苯醌室温下反应10分钟，过滤，蒸除溶剂，硅胶柱层析纯化，洗脱剂以体积计为石油醚:乙酸乙酯:三乙胺＝100:10:1，得到目标化合物 2-COOH-4-NH2SiR，产率79％。
其核磁1H NMR(300MHz,CDCl3)δ0.60(s,6H),1.14(t,12H,J＝6.9Hz),3.35(q,8H,J＝6.9Hz),3.97(s,2H),6.44-7.25(m,9H)；核磁13C NMR(75MHz,CDCl3)δ-2.18,0.58,12.56,44.26,92.46,109.53,112.22,116.06,121.11,125.72,128.46,129.24,131.62,137.75,143.44,146.52,147.21,170.70；高分辨质谱仪HRMS(ESI)calcd.for C30H35N3O2Si[M+H]+:500.2728,found:500.2712。
实施例24  化合物光学性质测定
1.溶液配制
分别精确称取2-H SiR，2-Me SiR，2-Br SiR，2-NO2SiR，2-SO3H SiR，2-Me-4-Br SiR，2-Me-5-NO2SiR,2-Me-4-COOH SiR，2,4-DiSO3H SiR 1～3mg，用乙醇溶解并定容至10mL得样品的高标溶液，用微量移液器量取一定体积的高标溶液，用乙醇定容至10mL，控制其浓度在2.4μM～3.2μM之间，备用。
分别精确称取2-COOH SiR，2-COOH SiR2，2-COOH SiR3，2-COOH-4-Br SiR，2-COOH-4-CN SiR，2-COOH-4-NO2 SiR，2-COOH-5-CCH SiR，2-COOH-5-CONH2 SiR，2-COOH-4-NH2 SiR 1～3mg，用乙醇溶解并定容至10mL得样品的高标溶液，用微量移液器量取一定体积的高标溶液，加入含1.0‰盐酸的乙醇溶液80μL，用乙醇定容至10mL，控制其浓度在2.4μM～3.2μM之间，备用。
2.样品吸收光谱的测定
将配好的样品溶液用紫外分光光度计(Agilent Cary 100)(石英皿，光路长度1cm)测定其紫外吸收光谱，测定范围450nm～750nm，测定其最大吸收波长，计算其样品的摩尔吸光系数，结果见表2，吸收光谱图见附图1。
3.样品发射光谱的测定
将配好的样品溶液用荧光光谱仪测定其荧光发射光谱，激发波长610nm，测定范围620nm～750nm，测定其最大发射波长，以结晶紫为参比物，计算其荧光量子产率，结果见下表2，样品的荧光发射光谱见附图2。
表2 化合物光谱数据
Figure PCTCN2015073677-appb-000036
Figure PCTCN2015073677-appb-000037
实施例25  化合物的细胞染色及激光共聚焦荧光成像实验
1.细胞培养方法：
试验细胞：选用小鼠脑微血管内皮细胞(bEND3)和人肝癌细胞(HepG2)；
细胞培养条件：使用含10％FBS、0.1mg/mL链霉素和100U/mL的青霉素的DMEM培养基，在含5％CO2、95％空气，37℃恒温，饱和湿度的细胞培养箱中培养细胞。细胞聚合度达到90％时，吸出培养基并用PBS溶液清洗细胞2次，使用0.25％胰蛋白酶37℃消化1分钟，加入2倍体积培养基中和，1000rpm离心5分钟，将细胞以1:3比例传代到培养皿，每2天更换一次培养基。
2.试验用荧光染料的配置
将2-COOH SiR用DMSO配置成1.0mM溶液备用；2,4-DiSO3H SiR用PBS缓冲液配置成1.0mM溶液备用。
3.细胞染色方法
将bEND3细胞以5×104密度接种于激光共聚焦专用培养皿中，在含5％CO2、95％空气，37℃恒温，饱和湿度的细胞培养箱中培养细胞24h后，弃去培养基并用预冷PBS清洗2遍。在1.0mL新鲜培养基中加入2-COOH SiR溶液10μL，混合均匀后加入培养皿中，培养箱中孵育40分钟，之后在该培养基中加入DAPI染色剂，培养箱中继续孵育20分钟，用PBS缓冲液洗涤三遍，激光共聚焦显微镜下观察，激发光源488nm和633nm。激光共聚焦荧光成像图见附图3。2-COOH SiR对HepG2细胞染色方法同对bEND3细胞染色方法一致，激光共聚焦荧光成像图见附图4。2,4-DiSO3H SiR对bEND3细胞和HepG2细胞染色方法同2-COOH SiR一致，激光共聚焦荧光成像图见附图5、附图6。
4.细胞染色实验结果
通过激光共聚焦荧光成像对细胞染色效果观察，发现2-COOH SiR和2,4-DiSO3H SiR均可透过活细胞的细胞膜进入细胞中，并均匀分布于胞质中，但不能对细胞核进行染色。如附图3～6激光共聚焦荧光成像效果图所示(照片转化为灰度模式)。
以上所述仅是本发明的优选实施方式，应当指出，对于本技术领域的普通技术人员，在不脱离本发明方法的前提下，还可以做出若干改进和补充，这些改进和补充也应视为本发明的保护范围。

Claims (10)

  1. 一种硅基罗丹明衍生物，其特征在于，所述硅基罗丹明衍生物是具有通式(I)结构的化合物：
    Figure PCTCN2015073677-appb-100001
    R11，R15其中任意一个为或两个均为-COOH或者-COO-时，可以形成通式(I)相应的内酯螺环形式，具有通式(II)结构的化合物：
    Figure PCTCN2015073677-appb-100002
    其中：
    R2、R3分别为C1-C6直链或支链的烃基或苯基，所述烃基选自：甲基、乙基、正丙基、异丙基、正丁基、异丁基、仲丁基、叔丁基、正戊基、异戊基、仲戊基、新戊基、正己基；
    R4、R5、R6、R7、R8、R9分别为氢或C1-C6的直链或支链的烃基，所述烃基选自：甲基、乙基、正丙基、异丙基、正丁基、异丁基、仲丁基、叔丁基、正戊基、异戊基、仲戊基、新戊基、正己基；
    R20、R21、R22、R23分别为C1-C6直链或支链的饱和或不饱和的烃基或环烷基，所述烃基选自：甲基、乙基、正丙基、异丙基、烯丙基、正丁基、异丁基、仲丁基、叔丁基、正戊基、异戊基、仲戊基、新戊基、正己基；
    R11、R12、R13、R14、R15，R16，R17，R18，R19为相同或者不同的取代基，所述取代基选自：烃基、卤素、氰基、硝基、烷氧基、卤代烷基。
  2. 根据权利要求书1所述的硅基罗丹明衍生物，其特征在于，所述硅基罗丹明衍生物取代情形选自下列中的一种、两种或两种以上：
    (a)所述取代基R2、R3为甲基、乙基或苯基；
    (b)所述取代基R4、R5、R6、R7、R8、R9为氢；
    (c)所述取代基R20、R21、R22、R23为甲基、乙基或烯丙基。
  3. 根据权利要求书1所述的硅基罗丹明衍生物，其特征在于，所述硅基罗丹明衍生物具备下列情形中的一种或两种：
    a)所述取代基R20或R21与R5以碳链相连，与母体苯环形成五元或六元环状结构；
    b)所述取代基R22或R23与R8以碳链相连，与母体苯环形成五元或六元环状结构。
  4. 根据权利要求书3所述的硅基罗丹明衍生物，其特征在于，所述硅基罗丹明衍生物具备下列情形中的一种或两种：
    所述取代基-NR20R21与-R5形成具有以下通式(III)的结构：
    Figure PCTCN2015073677-appb-100003
    所述取代基-NR22R23与-R8形成具有以下通式(IV)的结构：
    Figure PCTCN2015073677-appb-100004
  5. 根据权利要求书4所述的硅基罗丹明衍生物，其特征在于，所述硅基罗丹明衍生物具备下列情形中的一种或两种：
    a)所述的取代基R24为氢、甲基、乙基或烯丙基；
    b)所述的取代基R25为氢、甲基、乙基或烯丙基。
  6. 根据权利要求书1-4任一所述的硅基罗丹明衍生物，其特征在于，所述硅基罗丹明衍生物取代情形选自下列中的一种：
    (a)R12、R13、R14均为氢，R11或R15其中任一个为-CN、-NO2、-SO3H、-SO3 -、-OH、-OCH3、-NH2、-N3、-NCS、-CH3、-F、-Cl、-Br、-I中的任意一个基团，另一个为氢；
    (b)R11、R15其中任意一个为甲基，另一个为氢，R12、R13、R14中任意一个为-CN、-NO2、-SO3H、-SO3 -、-OH、-OCH3、-NH2、-CH3、-N3、-NCS、-C≡CH、-CONH2、-CH2NH2、-F、-Cl、-Br、-I、-COOH、-COO-中的任意一个基团，其余两个均为氢；
    (c)R11、R15其中任意一个为-SO3H或-SO3 -，另一个为氢，R12、R13、R14中任意一个为-SO3H或-SO3 -，另外两个为氢；
    (d)R11或者R15一个为-H，另一个为-COOH或者-COO-，R12，R13，R14中任意一个为-CN、-NO2、-SO3H、-SO3-、-OH、-OCH3、-NH2、-CH3、-N3、-NCS、-C≡CH、-CONH2、-CH2NH2、-F、-Cl、-Br、-I、-COOH、-COO-基团中的一个，其余两个为氢；
    (e)R16，R17，R18，R19中任意一个为-CN、-NO2、-SO3H、-SO3 -、-OH、-OCH3、-NH2、-CH3、-N3、-NCS、-C≡CH、-CONH2、-CH2NH2、-F、-Cl、-Br、-I、-COOH、-COO-基团中的一个，其余三个为氢。
  7. 权利要求书1-6任一所述硅基罗丹明衍生物的制备方法，其特征在于，所述制备方法包括下述步骤：
    第一步：具有通式(V)结构的关键硅基中间体的合成
    Figure PCTCN2015073677-appb-100005
    当-NR20R21与-NR22R23，R4与R7，R5与R8，R6与R9两两均具有相同结构时，采用如下方法合成；
    N,N-二烃基间溴苯胺衍生物与正丁基锂反应生成相应的锂试剂，再进一步与二烃基二氯硅烷反应，硅胶柱层析，以乙酸乙酯和石油醚混合体系进行洗脱，生成具有通式(V)结构的关键硅基中间体；
    当-NR20R21与-NR22R23，R4与R7，R5与R8，R6与R9两两之间有一对或者一对以上具有不同结构时，采用如下方法合成；
    一种N,N-二烃基间溴苯胺衍生物与正丁基锂生成相应的锂试剂，再与二烃基二氯硅烷反应，取代其中的一个氯，生成单氯硅烷的产物；另一种N,N-二烃基间溴苯胺衍生物与正丁基锂反应生成相应的锂试剂，再进一步与单氯硅烷产物反应，硅胶柱层析，以乙酸乙酯和石油醚混合体系进行洗脱，生成具有通式(V)结构的关键硅基中间体；
    第二步：硅基罗明丹衍生物的合成
    具有通式(V)结构的关键硅基中间体与不同取代的苯甲醛衍生物在催化条件下，封管后置于防护罩中140℃反应，再经四氯苯醌氧化或者不氧化，最后进行柱层析分离，以二氯甲烷和甲醇的混合体系或乙酸乙酯、石油醚和三乙胺的混合体系进行洗脱，得到硅基罗丹明衍生物。
  8. 根据权利要求书7所述的制备方法，其特征在于，所述制备具有通式(V)结构的关键硅基中间体时所用的溶剂为乙醚；所述制备具有通式(V)结构的关键硅基中间体时反应温度为0℃；
    所述制备具有通式(V)结构的关键硅基中间体时，若-NR20R21与-NR22R23，R4与R7，R5与R8，R6与R9两两均具有相同结构时，各反应物的投料比为N,N-二烃基间溴苯胺:丁基锂:二烃基二氯硅烷＝1:1.05:0.6；若-NR20R21与-NR22R23，R4与R7，R5与R8，R6 与R9两两之间有一对或者一对以上具有不同结构时，各反应物的投料比为N,N-二烃基间溴苯胺:丁基锂:二烃基二氯硅烷＝1:1.05:5；
    所述具有通式(V)结构的关键硅基中间体与不同取代的苯甲醛衍生物的投料摩尔比为1:5；
    所述催化剂选用CuBr2或者对甲苯磺酸一水合物；
    所述具有通式(V)结构的关键硅基中间体与催化剂的摩尔比为1:0.1～1；
    所述柱层析分离的洗脱剂，以体积计，甲醇和二氯甲烷的混合溶剂为甲醇:二氯甲烷＝1:5～40，乙酸乙酯和石油醚混合溶剂为乙酸乙酯:石油醚＝1:40～80，乙酸乙酯、石油醚和三乙胺的混合溶剂为乙酸乙酯:石油醚＝1:10～80，三乙胺的加入量为乙酸乙酯和石油醚混合物体积的1％。
  9. 根据权利要求7所述的制备方法，其特征在于，所述制备方法还包括第三步硅基罗丹明衍生物进一步衍生化反应；所述的衍生化反应为：氰基的部分水解反应、氰基的完全水解反应、氰基的还原反应、硝基的还原反应或氨基的叠氮化反应。
  10. 权利要求书1-6任一所述的硅基罗丹明衍生物在制备细胞染料、生物染色剂、生物分子或生物粒子荧光标记中的应用。
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020033681A3 (en) * 2018-08-10 2020-04-09 Life Technologies Corporation Silicon-substituted rhodamine dyes and dye conjugates

Families Citing this family (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP6898301B2 (ja) * 2015-07-28 2021-07-07 ザ リージェンツ オブ ザ ユニバーシティ オブ カリフォルニアＴｈｅ Ｒｅｇｅｎｔｓ Ｏｆ Ｔｈｅ Ｕｎｉｖｅｒｓｉｔｙ Ｏｆ Ｃａｌｉｆｏｒｎｉａ 長波長電圧感受性色素
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CN110922783A (zh) * 2018-09-19 2020-03-27 深圳华大生命科学研究院 硅基罗丹明衍生物及其制备方法
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CN110872314A (zh) * 2019-11-21 2020-03-10 华东师范大学 一种不对称硅基取代罗丹明衍生物及制备方法和应用
CN112266388B (zh) * 2020-10-22 2021-12-28 大连理工大学 一类具有标记功能用于超分辨成像的亚硝基笼化罗丹明衍生物及应用
CN112538089B (zh) * 2020-12-04 2023-04-07 河北大学 近红外硅基罗丹明荧光染料、制备方法及其在线粒体脊膜原位免洗成像中的应用
CN112574246B (zh) * 2020-12-14 2021-09-07 山西大学 Zn2+比率荧光探针、制备及应用

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012083064A1 (en) * 2010-12-16 2012-06-21 Southern Illinois University Carbondale Silaanthracene as a red and near infrared sensor and a method to manufacture such a sensor
WO2012111818A1 (ja) * 2011-02-18 2012-08-23 国立大学法人 東京大学 蛍光プローブ
WO2014106957A1 (ja) * 2013-01-07 2014-07-10 国立大学法人 東京大学 非対称Ｓｉローダミン及びロドールの合成

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN100361999C (zh) * 2006-03-03 2008-01-16 大连理工大学 含硅元素的阳离子发色团及其衍生物
JP5526124B2 (ja) * 2009-04-30 2014-06-18 国立大学法人 東京大学 近赤外蛍光化合物
CN103113380B (zh) * 2013-03-08 2014-11-05 苏州大学 一种罗丹明衍生物、其制备方法及应用

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012083064A1 (en) * 2010-12-16 2012-06-21 Southern Illinois University Carbondale Silaanthracene as a red and near infrared sensor and a method to manufacture such a sensor
WO2012111818A1 (ja) * 2011-02-18 2012-08-23 国立大学法人 東京大学 蛍光プローブ
WO2014106957A1 (ja) * 2013-01-07 2014-07-10 国立大学法人 東京大学 非対称Ｓｉローダミン及びロドールの合成

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2020033681A3 (en) * 2018-08-10 2020-04-09 Life Technologies Corporation Silicon-substituted rhodamine dyes and dye conjugates
CN112566986A (zh) * 2018-08-10 2021-03-26 生命技术公司 经硅取代的罗丹明染料和染料缀合物
AU2019319901B2 (en) * 2018-08-10 2022-07-07 Life Technologies Corporation Silicon-substituted rhodamine dyes and dye conjugates

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