A visible light long wavelength activated photoswitching molecule and its synthesis method and application

Abstract

The application belongs to the technical field of optical switch molecular materials, and particularly relates to a visible light long wavelength activated optical switch molecule and a synthesis method and application thereof. The optical switch molecule is an optical switch molecule based on coumarin, rhodamine and cyanine mother nucleus; the synthesis method comprises the following steps: preparing the optical switch molecule through condensation reaction, coupling reaction, substitution reaction or Click reaction of an intermediate fluorophore and 1,4-oxathiacyclohexene; the photoactivated molecule designed and synthesized in the application will experience the process of gradually lighting and then photo bleaching in the process of excitation light irradiation, and thus is suitable for long time tracing and can be used for super-resolution imaging of cells, including living cells and dead cells. The optical switch molecule can be photoactivated through long wavelength irradiation, and can be widely applied to the field of photochemistry.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of photo-switching molecular materials, and particularly relates to a long-wavelength activated photo-switching molecule and a synthesis method and application thereof. BACKGROUND

[0002] Photoactivated fluorescent probes are also known as photo-luminescent probes, and have wide applications in the fields of optical devices and molecular imaging. Traditional photo-cages such as azide, o-nitrobenzyl, nitroso and tetrazine, etc. mostly need to be decomposed or removed by short-wavelength ultraviolet light. The ultraviolet light has low penetration in cell tissues and is easily absorbed, causing background fluorescence interference. Xiao's group converted the carbonyl group in the fluorophore into a thiocarbonyl group by using Lawson's reagent. In the visible light range, the thiocarbonyl group is oxidized to an oxycarbonyl group by intramolecularly generated singlet oxygen, realizing visible light activation. However, this method has particularity and requires the fluorophore itself to contain a carbonyl structure. Therefore, it is particularly important to develop a general photo-cage group and realize the photoactivation of fluorescent dyes by long-wavelength irradiation.

[0003] Literature Chem. Commun. 2003, 1756-1757 reports a 1,4-oxathiin compound which can combine singlet oxygen to make the double bond break.

[0004]

[0005] Patent CN 112457336 A combines 1,4-oxathiin with BODIPY, realizes fast photolysis speed and high photolysis efficiency by singlet oxygen generated by a photosensitizer, and is applied to long-afterglow luminescent materials in the field requiring fast photoactivation.

[0006]

[0007] However, so far, there is no compound combining 1,4-oxathiin with phycocyanin, rhodamine and coumarin dye molecules. And it is rarely reported to realize visible light photoactivation by oxidizing the photo-cage group 1,4-oxathiin using singlet oxygen generated after absorbing photons. SUMMARY

[0008] The purpose of the present application is to provide a visible light long-wavelength activated photo-switching molecule and a synthesis method and application thereof.

[0009] The visible light long-wavelength activated photo-switching molecule provided by the present application is a general photo-switching molecule based on the coumarin, rhodamine and phycocyanin mother nucleus, and a strategy for realizing photoactivation by long-wavelength irradiation and application thereof in the field of photochemistry. The molecular structure of the photo-switching molecule is shown in formula I, II or III as follows:

[0010]

[0011] wherein:

[0012] A is selected from oxygen or dimethylsilyl group;

[0013] n is selected from an integer 1, 3 or 5;

[0014] LK is a functional group or a chemical bond that links the upper moiety 1,4-cyclohexene structure and the lower moiety fluorophore structure;

[0015] said linking functional group is selected from any one of the following:

[0016]

[0017] R1, R2may be the same or different and are each selected from oxygen, sulfur, nitrogen or carbon;

[0018] R3, R4may be the same or different and are each selected from hydrogen, alkyl group having 1-50 carbon atoms, alkoxy group having 1-50 carbon atoms, alkylamino group having 1-50 carbon atoms;

[0019] R5, R6may be the same or different and are each selected from hydrogen, C1-C 12 alkyl group, C1-C 12 alkoxy group, C3-C 12 cycloalkyl group, C1-C 12 alkylamino group, C1-C 12 alkylthio group, C2-C 12 alkenyl group, C2-C 12 alkynyl group, C2-C 12 alkenyloxy group, C2-C 12 alkynyloxy group, C1-C 12 alkylsulfonyl group, C1-C 12 alkylcarboxy group;

[0020] R9, R 10 may be the same or different and are each selected from hydrogen, C1-C 12 alkyl group, C1-C 12 alkoxy group, C3-C 12 cycloalkyl group, C1-C 12 alkylamino group, C1-C 12 alkylthio group, C2-C 12 alkenyl group, C2-C 12 alkynyl group, C2-C 12 alkenyloxy group, C2-C 12 alkynyloxy group, C1-C 12 alkylsulfonyl group, C1-C 12 alkylcarboxy group;

[0021] R 11 , R 12 may be the same or different, and are each selected from hydrogen, alkyl, alkoxy, alkylamino, alkylcyano, aryl or benzylaryl groups having 1-16 carbon atoms;

[0022] wherein the aryl group is free of substituent groups or is substituted with one or more groups L;

[0023] L is selected from alkoxy, hydroxy, carboxy, amino, ester, nitro or sulfonic acid groups.

[0024] According to an embodiment of the present application, the photo-switching molecule is any one of the following compounds:

[0025]

[0026]

[0027]

[0028] The present application also provides a synthesis method of the photo-switching molecule, comprising: preparing the photo-switching molecule from an intermediate fluorophore and 1,4-oxathie, through condensation reaction, coupling reaction, substitution reaction or Click reaction; and a synthesis route thereof is as follows:

[0029]

[0030] wherein:

[0031] The condensation reaction has the following specific steps:

[0032] The fluorophore and 1,4-oxathie are added into an organic solvent, a condensing agent is added, the molar ratio of the fluorophore, 1,4-oxathie and the condensing agent is 1:1:1, the reaction is carried out for 12-24 hours to obtain a target solution, and the photo-switching molecule is obtained through post-treatment.

[0033] The condensing agent is selected from one of dicyclohexyl carbodiimide (DCC), 1-ethyl-3(3-dimethylpropylamine) carbodiimide (EDCI), 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyl urea hexafluorophosphate (HATU), benzotriazole (HOBt) and 1-ethyl-3(3-dimethylpropylamine) carbodiimide (EDCI).

[0034] The coupling reaction (including Suzuki coupling, Heck coupling, Sonogoshira coupling) has the following specific steps:

[0035] The fluorophore and 1,4-oxathiine are added to an organic solvent, a coupling reaction catalyst and a base are added, the molar ratio of the fluorophore, 1,4-oxathiine, catalyst and base is 1:1:0.01:3-1:1:0.1:8 (1:1:(0.01-0.1):(3-8)), preferably 1:1:0.05:5, and the reaction is carried out for 8-24 hours to obtain a target solution, and the photo-switching molecule is obtained after post-treatment.

[0036] The catalyst is selected from one of palladium chloride, palladium acetate or tetrakis(triphenylphosphine)palladium.

[0037] The substitution reaction, in particular, comprises the following steps:

[0038] The fluorophore and 1,4-oxathiine are added to an organic solvent, a base is added, the molar ratio of the fluorophore, 1,4-oxathiine and base is 1:1:3-1:1:10 (1:1:(3-10)), preferably 1:1:5, and the reaction is carried out for 12-24 hours to obtain a target solution, and the photo-switching molecule is obtained after post-treatment.

[0039] The Click reaction, in particular, comprises the following steps:

[0040] The fluorophore and 1,4-oxathiine are added to an organic solvent, a Click reaction catalyst is added, the molar ratio of the fluorophore, 1,4-oxathiine and Click reaction catalyst is 1:1:0.01-1:1:0.09 (1:1:(0.01-0.09)), preferably 1:1:0.05, and the reaction is carried out for 12-24 hours to obtain a target solution, and the photo-switching molecule is obtained after post-treatment.

[0041] The catalyst is selected from one of copper sulfate or cuprous iodide.

[0042] In the above reactions, the organic solvent is one or a mixture of several of toluene, tetrahydrofuran, 1,4-dioxane, ethanol, methanol, N,N-dimethylformamide or dimethyl sulfoxide.

[0043] In the above synthesis method, the intermediate fluorophore is selected from any one of the following compounds:

[0044]

[0045]

[0046] The 1,4-oxathiine is selected from any one of the following compounds:

[0047]

[0048] The traditional fluorescent molecules will have serious photobleaching when applied to cell imaging, so long-term tracking cannot be realized, and the light-activated molecules designed and synthesized in the application will experience the process of gradually lighting up and then photobleaching in the process of excitation light irradiation, so they are suitable for long-term tracking.

[0049] The light-activated fluorescent molecules involved in the application can be used for super-resolution imaging of cells, including living cells and dead cells. BRIEF DESCRIPTION OF DRAWINGS

[0050] Figure 1 NMR hydrogen spectrum of Cy5-SO in deuterated methanol for Example 2.

[0051] Figure 2 NMR carbon spectrum of Cy5-SO in deuterated methanol for Example 2.

[0052] Figure 3 NMR hydrogen spectrum of Coumarin-SO in deuterated chloroform for Example 3.

[0053] Figure 4 NMR carbon spectrum of Coumarin-SO in deuterated chloroform for Example 3.

[0054] Figure 5 NMR hydrogen spectrum of Rhodamine-SO in deuterated chloroform for Example 1.

[0055] Figure 6 NMR carbon spectrum of Rhodamine-SO in deuterated chloroform for Example 1.

[0056] Figure 7 Light activation process of Cy5-SO (10 μM) in pH 7.4 PBS phosphate buffer for Example 2, 405 nm (a), 473 nm (b), 532 nm (c), 635 nm (d) laser irradiation for 2 hours (power density 35 W / cm 2 ), and the emission spectrum was tested after the time shown in the figure.

[0057] Figure 8 Light activation process of Rhodamine-SO (50 μM) in pH 7.4 PBS phosphate buffer for Example 1, 405 nm (a), 473 nm (b), 532 nm (c), 635 nm (d) laser irradiation for 2 hours (power density 35 W / cm 2 ), and the emission spectrum was tested after the time shown in the figure.

[0058] Figure 9For the photoactivation process of Coumarin-SO (10 μM) in DMSO, the 405 nm (a), 473 nm (b), 532 nm (c), 635 nm (d) laser irradiation for 2 hours (power density 35 W / cm 2 The emission spectra were tested after the time indicated in the figure.

[0059] Figure 10 For the photoactivation process of Cy3-SO (10 μM) in DMSO, the 405 nm (a), 473 nm (b), 532 nm (c), 635 nm (d) laser irradiation for 2 hours (power density 35 W / cm 2 The emission spectra were tested after the time indicated in the figure.

[0060] Figure 11 For the photoactivation process of Cy5-SO in Hela cells under 635 nm laser irradiation and its fluorescence intensity change curve, the cell imaging of photoactivation of Example 6.

[0061] Figure 12 For the photoactivation process of Rhodamine-SO in Hela cells under 532 nm laser irradiation and its fluorescence intensity change curve, the cell imaging of photoactivation of Example 6.

[0062] Figure 13 For the photoactivation process of Coumarin-SO in Hela cells under 405 nm laser irradiation and its fluorescence intensity change curve, the cell imaging of photoactivation of Example 6.

[0063] Figure 14 For the super-resolution imaging of Rhodamine-SO-PA for fixing DRG neurons, (a) is the comparison of traditional wide field and single molecule localization super-resolution images, (b) is the FWHM analysis of microtubules in the yellow frame in (a), (c) is the periodic analysis of DRG neurons. DETAILED DESCRIPTION

[0064] The experimental methods used in the following examples are conventional methods unless otherwise specified.

[0065] Example 1: Preparation of rhodamine-SO

[0066]

[0067] Into a 25 mL three-necked flask, rhodamine-Br (47 mg, 0.1 mmol, 1 eq), boronate (64 mg, 0.15 mmol, 1.5 eq), tetrakis(triphenylphosphine)palladium (5.8 mg, 0.005 mmol, 0.05 eq) and sodium carbonate (42 mg, 0.4 mmol, 4 eq) were added successively. Then 1,4-dioxane / ethanol / water (10 mL, 4:1:1) was added to the reaction mixture, which was stirred at 100 °C under nitrogen protection overnight. The reaction was cooled to room temperature and the solvent was removed under reduced pressure. Purification was performed by silica gel column chromatography (eluent: dichloromethane / methanol = 20:1) to give 41 mg of brown solid with a yield of 60%. The product was confirmed by1H NMR,13C NMR and MS. 1 H NMR, 13 C NMR and MS: 1 H NMR (400 MHz, CDC13) δ 7.97 (d, J = 7.8 Hz, 1H), 7.72 (d, J = 7.8 Hz, 1H), 7.25-7.23 (m, 4H), 7.16 (s, 1H), 7.04 (d, J = 8.3 Hz, 2H), 6.64 (d, J = 8.3 Hz, 2H), 6.55-6.45 (m, 4H), 6.37 (d, J = 7.3 Hz, 2H), 4.46 (s, 2H), 3.19 (s, 2H), 2.95 (s, 12H), 2.87 (s, 6H). 13 C NMR (101 MHz, CDC13) δ 169.8, 154.4, 152.9, 152.0, 149.7, 147.5, 143.9, 137.5, 137.1, 130.9, 129.2, 128.9, 128.2, 126.5, 126.1, 125.9, 125.0, 122.0, 112.2, 109.4, 108.7, 106.9, 98.6, 65.6, 40.3, 40.3, 28.5; HR-ESI-MS m / z: [M+H] + calcd. for C 42 H 40 N3O4S + , 682.2734; found, 682.2733.

[0068] Example 2, Preparation of Cy5-SO

[0069]

[0070] Into a 25 mL three-necked flask, Cy5-Br (64 mg, 0.1 mmol, 1 eq), boronate (64 mg, 0.15 mmol, 1.5 eq), tetrakis(triphenylphosphine)palladium (5.8 mg, 0.005 mmol, 0.05 eq) and sodium carbonate (42 mg, 0.4 mmol, 4 eq) were added successively. Then 1,4-dioxane / ethanol / water (10 mL, 4:1:1) was added to the reaction mixture, which was stirred at 100 °C under nitrogen protection for 3 h. The reaction was cooled to room temperature and the solvent was removed under reduced pressure. Purification was performed by silica gel column chromatography (eluent: dichloromethane / methanol = 15:1) to give 58 mg of brown solid with a yield of 68%. The product was confirmed by1H NMR,13C NMR and MS. 1 H NMR, 13 C NMR and MS: 1 H NMR (400 MHz, CDC13) δ 8.22 - 8.17 (m, 2H), 7.45 - 7.31 (m, 6H), 7.24 - 7.10 (m, 6H), 7.04 (d, J = 7.7 Hz, 2H), 6.58 (d, J = 8.2 Hz, 2H), 5.71 - 5.58 (m, 2H), 4.55 (s, 2H), 3.77 (s, 4H), 3.25 (s, 2H), 2.92 (s, 6H), 2.37 (s, 2H), 1.93 - 1.73 (m, 12H), 1.66 - 1.56 (m, 4H), 1.22 (s, 3H). 13 C NMR (101 MHz, CDC13) δ 175.8, 173.2, 172.7, 153.0, 149.7, 144.1, 141.8, 141.4, 141.4, 141.2, 136.7, 135.0, 133.7, 131.0, 129.6, 129.3, 128.7, 128.7, 126.5, 125.5, 122.6, 122.4, 112.3, 110.8, 110.4, 109.4, 101.6, 101.2, 65.7, 49.6, 44.2, 40.6, 39.2, 33.9, 28.6, 28.2, 28.1, 26.6, 22.3, 12.1; HR-ESI-MS m / z: [M+H] + calcd. for C 50 H 56 N3O3S + , 778.4037; found, 778.4034.

[0071] Example 3, preparation of coumarin-SO

[0072]

[0073] Into a 25 mL three-necked flask, coumarin-Br (42 mg, 0.1 mmol, 1 eq), boronate (64 mg, 0.15 mmol, 1.5 eq), tetrakis(triphenylphosphine)palladium (5.8 mg, 0.005 mmol, 0.05 eq) and sodium carbonate (42 mg, 0.4 mmol, 4 eq) were added successively. Then 1,4-dioxane / ethanol / water (10 mL, 4:1:1) was added to the reaction mixture, which was stirred at 90 °C for 12 h under nitrogen protection. The reaction was cooled to room temperature and the solvent was removed under reduced pressure. Purification was performed by silica gel column chromatography (eluent: dichloromethane / methanol / tetrahydrofuran = 100:1:1) to give 28 mg of yellow solid with a yield of 45%. The product was confirmed by1H NMR,13C NMR and MS. 1 H NMR, 13 C NMR and MS: 1 H NMR (400 MHz, CDC13) δ 8.04 (d, J = 7.9 Hz, 2H), 7.38 (d, J = 8.9 Hz, 1H), 7.27 (d, J = 7.9 Hz, 2H), 7.23 (d, J = 8.1 Hz, 2H), 7.12 (d, J = 8.9 Hz, 2H), 7.04 (d, J = 8.1 Hz, 2H), 6.70 - 6.41 (m, 4H), 4.63 (s, 2H), 4.49 (s, 2H), 3.55 (d, J = 6.9 Hz, 2H), 3.29 - 3.11 (m, 2H), 2.89 (s, 6H), 2.12 (s, 3H), 1.26 (t, J = 6.9 Hz, 3H). 13 C NMR (101 MHz, CDC13) δ 171.4, 161.7, 154.8, 150.6, 149.7, 148.2, 144.5, 144.2, 136.1, 133.6, 130.9, 130.8, 129.8, 128.5, 126.4, 126.2, 121.7, 112.4, 110.6, 109.1, 108.6, 98.3, 65.6, 53.9, 46.1, 40.5, 28.5, 16.3, 12.2; HR-ESI-MS m / z: [M+H] + calcd. for C 38 H 37 N2O5S + , 633.2418; found, 633.2393.

[0074] Example 4: Preparation of Cy3-SO

[0075]

[0076] Into a 25 mL three-necked flask, Cy3-Br (59 mg, 0.1 mmol, 1 eq), boronate (64 mg, 0.15 mmol, 1.5 eq), tetrakis(triphenylphosphine)palladium (5.8 mg, 0.005 mmol, 0.05 eq) and sodium carbonate (42 mg, 0.4 mmol, 4 eq) were added successively. Then 1,4-dioxane / ethanol / water (10 mL, 4:1:1) was added to the reaction mixture, which was stirred at 100 °C for 12 h under nitrogen protection. The reaction was cooled to room temperature and the solvent was removed under reduced pressure. Purification was performed by silica gel column chromatography (eluent: dichloromethane / methanol = 20:1) to give 53 mg of yellow solid with a yield of 65%. The product was confirmed by1H NMR,13C NMR and MS. 1 H NMR, 13 C NMR and MS: 1 H NMR (400 MHz, CDC13) δ 8.47 - 8.41 (m, 1H), 7.57 (d, J = 7.8 Hz, 1H), 7.51 (s, 1H), 7.43 - 7.35 (m, 6H), 7.29 (t, J = 8.2 Hz, 3H), 7.15 (d, J = 7.8 Hz, 4H), 6.61 (d, J = 8.2 Hz, 2H), 4.54 (s, 2H), 4.34 (q, J = 6.6 Hz, 4H), 3.25 (s, 2H), 2.94 (s, 6H), 1.73 (s, 12H), 1.52 (t, J = 6.6 Hz, 6H). 13 C NMR (101 MHz, CDC13) δ 173.1, 172.8, 150.4, 149.6, 144.1, 141.7, 141.3, 140.9, 140.7, 138.3, 138.1, 136.2, 130.9, 129.9, 129.3, 128.9, 127.6, 126.1, 125.9, 125.3, 125.1, 122.2, 120.6, 112.2, 110.9, 110.8, 108.9, 104.5, 65.6, 48.9, 40.4, 40.2, 40.1, 28.5, 28.1, 28.1, 13.0; HR-ESI-MS m / z: [M] calcd for C 45 H 50 N3OS + , 680.3669; found, 680.3665.

[0077] Example 5, Compound Light Switching Activity Test

[0078] Cy5-SO, Rhodamine-SO, Coumarin-SO and Cy3-SO fluorescent dyes were irradiated with 405 nm, 473 nm, 532 nm and 635 nm lasers, respectively, and the fluorescence spectra at different irradiation time points were tested Figures 7-10 ). The experimental results showed that 1,4-oxathiacyclohexenyl (SO) combined with cyanine, rhodamine and coumarin dyes, and visible light activation could be achieved by long wavelength irradiation. Cy5-SO, Rhodamine-SO and Coumarin-SO all observed significant fluorescence enhancement under light irradiation within their absorption spectra Figures 7-9 ). The fluorescence intensity of Cy5-SO increased 8-fold, 5-fold, 7-fold and 6-fold after 2 hours of irradiation with 405 nm, 473 nm, 532 nm and 635 nm lasers, respectively Figure 7 ). The fluorescence intensity of Rhodamine-SO increased 81-fold, 37-fold and 61-fold after 2 hours of irradiation with 405 nm, 473 nm and 532 nm lasers, respectively, and only a weak fluorescence enhancement factor was observed under 635 nm laser irradiation outside the absorption spectrum Figure 8 ). The fluorescence intensity of Coumarin-SO increased 34-fold after 2 hours of irradiation with 405 nm laser, and no significant fluorescence enhancement was observed under 473 nm, 532 nm and 635 nm laser irradiation outside the absorption spectrum Figure 9 ). In addition, due to the poor light stability of Cy3-SO, no significant fluorescence enhancement was observed under light irradiation within its absorption spectrum, and only a weak fluorescence enhancement was observed under 405 nm light irradiation Figure 10 ).

[0079] Example 6, Light-activated cell imaging

[0080] To study the light switch activity of the photoactivated fluorescent probe in cells, confocal laser scanning microscopy was used to perform photoactivation imaging experiments on HeLa cells Figures 11-13Hela cells were incubated with 10 micromolar of fluorescent probes (Cy5-SO, Rhodamine-SO and Coumarin-SO dissolved in pH 7.4 PBS buffer solution respectively) in dark at 37℃ for 30 min, and the intracellular fluorescent probes were photoactivated by confocal laser scanning microscope, and the relationship between the intracellular fluorescence intensity and the irradiation time of excitation light was explored. The excitation wavelengths of Cy5-SO, Rhodamine-SO and Coumarin-SO were 637 nm, 561 nm and 405 nm respectively, and the excitation power was 31 μW, 70 μW and 15 μW respectively. Under the continuous scanning of laser, continuous fluorescence enhancement was observed, and since the probe would undergo photobleaching after photoactivation, the fluorescence intensity would continuously decrease after reaching the peak, and the fluorescence intensity showed the trend of first increasing and then decreasing, thereby confirming that the SO type photoactivated fluorescent probe could realize visible light long wavelength photoactivation in living cells.

[0081] Example 7, photoactivated super-resolution imaging

[0082] In order to further explore the excellent photoactivation performance of the photoactivated fluorescent probe and its application prospect, the molecular structure of Rhodamine-SO is modified in the present application, and the polyene paclitaxel group (Rhodamine-SO-PA) targeting microtubules is connected to realize microtubule super-resolution imaging in fixed DRG neurons. Figure 14 The DRG neuron sample cultured in vitro for 9 days was incubated with 2 micromolar of Rhodamine-SO-PA at room temperature in dark for 60 minutes after being fixed by glutaraldehyde, and then the sample was excited by 532 nm laser with a power density of 2.5 kW / cm 2 The data set collected was processed by ThounderSTORM software, and the super-resolution image beyond the diffraction limit was reconstructed, and the more detailed structure of the microtubule was displayed. Compared with the 347 nanometer full width at half maximum of the microtubule obtained by the traditional wide field imaging, the reconstructed super-resolution image showed a microtubule full width at half maximum of 66.9 nanometers. The reconstructed super-resolution image also showed that the microtubule protein was distributed periodically along the axon about 150 nanometers.

Claims

1. A visible light long wavelength activated photoswitching molecule, characterized in that, The cation of the photo-switching molecule is any one of the following:

2. A method for synthesis of a visible light long wavelength activated photoswitching molecule as claimed in claim 1, wherein the cation of the photochromic molecule is Cy5-SO-1, characterized in that, comprising: intermediate fluorophore and 1,4-oxathiine are prepared into photo-switching molecules by coupling reaction; The specific steps are: The intermediate fluorophore and 1,4-oxathiine are added into an organic solvent, a coupling reaction catalyst and a base are added, the molar ratio of the intermediate fluorophore, 1,4-oxathiine, catalyst and base is 1:1:0.01:3-1:1:0.1:8, the reaction is carried out for 8-24 hours to obtain a target solution, and the photo-switching molecule is obtained after post-treatment; the catalyst is selected from one of palladium chloride, palladium acetate or tetrakis(triphenylphosphine)palladium; The intermediate fluorophore is The 1,4-oxathiin is 3. The method of synthesis of claim 2, wherein, The organic solvent is one of toluene, tetrahydrofuran, 1,4-dioxane, ethanol, methanol, N,N-dimethylformamide or dimethyl sulfoxide or a mixture of several thereof.

4. Use of the photo-switching molecule of claim 1 activated by visible light long wavelength in the preparation of a cell imaging agent.

Images