CN100486976C - Fluorescent molecular probe and use for inspecting transient metal and heavy metal ion - Google Patents
Fluorescent molecular probe and use for inspecting transient metal and heavy metal ion Download PDFInfo
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- CN100486976C CN100486976C CNB2006100099258A CN200610009925A CN100486976C CN 100486976 C CN100486976 C CN 100486976C CN B2006100099258 A CNB2006100099258 A CN B2006100099258A CN 200610009925 A CN200610009925 A CN 200610009925A CN 100486976 C CN100486976 C CN 100486976C
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Abstract
A fluorescent molecular probe and its use for inspecting transition metal and heavy metal ion are disclosed. It has better quenching and selective performances and can be used to inspect cobalt ion in cell, biological tissue, sewage water, water solution, emulsion and suspending system.
Description
Technical field
The present invention relates to a kind of fluorescent molecular probe, comprise its design, the invention still further relates to the application of a kind of this fluorescent molecular probe in transition metal and heavy metal ion identification and detection with synthetic.
Background technology
The fluorescent molecular probe technology is applied to the detection of transition metal and heavy metal ion, can realize the in situ detection in the microenvironment.But usually, cause detection sensitivity decline because transition metal and heavy metal ion can make the fluorescent quenching of fluorophor by spin-orbit coupling and electronics or energy transfer.In addition, the solvation tendency of the identification base of metal ion and fluorescent molecular probe in the aqueous solution is better than organic solvent, the bonding properties of fluorescent molecular probe and detected species is descended, and therefore, many probe molecules only just show recognition reaction in organic solvent.But the fluorescent molecular probe that in fact can show recognition performance in the aqueous solution just has wide practical value.
Design, synthesize and in the aqueous solution, to have optionally, and can realize that fluorescence strengthens present or challenging research work of fluorescent molecular probe of identification.The fluorescent molecular probe of having reported to transition metal and heavy metal ion fluorescence enhancing identification in aqueous solution nearly all is to detect to have d
10The probe of the metal ion of electronic structure.Therefore, designing and synthesizing a class can be at various solution, particularly in aqueous solution to non-d
10The fluorescent molecular probe that the transition metal of electronic structure and heavy metal ion selectivity fluorescence strengthen identification is an important topic of current this area.We are once at " J.Am.Chem.Soc " .2004, and 125,2272-2273 has reported 2, and 6-diaminomethyl pyridine is that the fluorescent molecular probe of recognition group can strengthen the identification mercury ion by selectivity fluorescence, and the present invention is the continuation of the research.
Summary of the invention
At first to be that design is synthetic a kind ofly not only have anti-transition metal and heavy metal ion to the cancellation performance of fluorophor and good selectivity to goal of the invention of the present invention, and can also be at various solution, particularly in aqueous solution to non-d
10The transition metal of electronic structure and heavy metal ion selectivity fluorescence strengthen the fluorescent molecular probe of identification, and another object of the present invention provides the utilisation technology of a kind of this fluorescent molecular probe in transition metal and heavy metal ion identification and detection.
Fluorescent molecular probe of the present invention have a following general structure:
In the general formula:
R
1, R
2, R
3Represent hydrogen, alkyl, hydroxyalkyl, carboxyalkyl, polyoxyethylene groups, aryl, aralkyl or unsubstituted.
X
1, X
2Represent nitrogen-atoms or Sauerstoffatom.
X
3Represent nitrogen-atoms, sulphur atom, Sauerstoffatom, phosphorus atom or two ends that Alkylenediamine, two sulfur derivatives in generation are arranged.
F
1And F
2Represent two fluorescence chromophoric groups.
Q
1, Q
2Representative has alkylidene group or two alkylidene group of 1-10 carbon atoms.
In the fluorescent molecular probe of above-mentioned general formula, alkyl can also be straight chain or the isomery alkyl with 1-20 carbon atoms, as short-chain alkyls such as methyl, ethyl, n-propyl, sec.-propyls; Can be chain alkyls such as dodecyl, octadecyl also, chain alkyl helps to improve the fat-soluble of probe.Hydroxyalkyl is meant the hydroxyalkyl with 1-12 carbon atoms, and as methylol, hydroxyethyl, hydroxypropyl, dihydroxypropyl, 4-hydroxybutyl etc., the short chain hydroxyalkyl helps improving the solvability of probe molecule in polar solvent.Carboxyalkyl is meant the carboxyalkyl with 1-12 carbon atoms, and as carboxymethyl, propyloic, carboxylic propyl group etc., carboxyalkyl is introduced probe molecule, and to be less than 7 short chain derivative with carbonatoms best.Polyoxyethylene groups is meant the polymerization degree in 1-40 polyoxyethylene groups, as two polyoxyethylene groups, three polyoxyethylene groups, and oxozone ethene etc., the introducing of polyoxyethylene group helps improving the water-soluble of probe molecule.Aryl is meant benzene, naphthalene, anthracene, pyrene, pyridine, naphthalimide, benzo furazan etc.Aralkyl is meant phenmethyl, furfuryl, menaphthyl, anthracene methyl, pyrene methyl, thenyl, quinoline methyl, phenanthroline methyl, picolyl, styroyl, furans ethyl, naphthalene ethyl, anthracene ethyl, pyrene ethyl, thiophene ethyl, quinoline ethyl, phenanthroline ethyl, pyridine ethyl, N-ethyl naphthalene imide, N-Yi Ji perylene tetracid imide, 4-ethylamino naphthalimide etc., wherein containing heteroatomic aralkyl helps to improve identification selection, and the aralkyl with fluorescent emission function helps to improve the identification susceptibility.
In the fluorescent molecular probe of above-mentioned general formula, fluorophor is meant 4-amino-1,8-naphthalimide, naphthalene, anthracene, pyrene, benzo furazan, 3,4,9,10-perylene tetracid imide, tonka bean camphor, dansyl amide, fluorescein, rhodamine, fluorine boron pyrroles, benzoglyoxaline, carbazole, benzoxazoles etc. have the organic group of fluorescent emission function.
In the fluorescent molecular probe of above-mentioned general formula, the linking group Q between fluorophor and recognition group
1, Q
2Representative has alkylidene group or two alkylidene group of 1-10 carbon atoms, as methylene radical, 1,1-ethylidene, ethylene, 1,1-propylidene, 1,3-propylidene, 1,2-propylidene, 1,1-butylidene, 1,2-butylidene, 1,3-butylidene, tetramethylene etc. are until the alkylidene group of 10 carbon atoms; Also can constitute this linking group by two alkylidene groups in the above-mentioned alkylidene group, as two ethylene, methylene radical and ethylene, methylene radical and 1,3-ethylidene, ethylene and 1,3-ethylidene etc.
In the fluorescent molecular probe of above-mentioned general formula, the linking group X between two pyridines
3Except that representing heteroatomss such as nitrogen-atoms, sulphur atom, Sauerstoffatom, phosphorus atom, comprise that also there are Alkylenediamine, two sulfur derivatives of replacement at two ends, as quadrol, diethylenetriamine, triethylene tetramine, 1, the inferior propylene diamine, 1 of 3-, the inferior propylene diamine, 1 of 2-, 2-Aden diamines, 1,3-Aden diamines, 1,4-Aden diamines, piperazine, 1,3-encircles pentamethylene diamine, 1,3-cyclohexanediamine, 1,2-dithio ethane, 1,3-dithio propane, 1,2-dithio propane, 1,2-dithio butane, 1,3-dithio butane, 1,4-dithio butane etc.
In the fluorescent molecular probe of above-mentioned general formula, two (2, the 6-lutidine) derivative is the recognition group with a plurality of ligating atoms, helps discerning the metal ion of certain radius and electronic structure; The two ends of molecule are connected with fluorophor, and these two fluorophors are controlled by part nitrogen-atoms in the recognition group by photoinduction transfer transport (PET).When the nitrogen-atoms in the identification base combined with metal ion, the PET process was blocked, and the fluorescence of two or more fluorophors strengthens simultaneously, helped making identification sensitivity to improve.
With 4-amino-1,8-naphthalimide fluorophor is an example, and such probe can obtain by following typical synthetic route.
R in the molecular formula, R
7And R
8Can be the alkyl and the aryl of hydrogen atom, alkyl, replacement.
4-amino naphthalenes imide is one to have by force and pushes away-fluorescent chemicals of La electron system, has advantages such as fluorescence quantum yield height, stokes (Stokes) displacement is big, the fluorescent emission wavelength is moderate, and the design that is often used in fluorescent molecular probe is with synthetic.By connecting base and identification division, the acceptor portion that promptly is identified thing links to each other at the imido electron donating group of 4-amino naphthalenes position, and such structure helps electron donor to chromophoric transfer, strengthens the PET process.
In addition, the present invention also designs, to have synthesized with anthracene, 4-amino-7-nitro benzo furazan etc. be that two pyridines of fluorophor are discerned basic fluorescent molecular probe, generally all connect hydroxyl isopolarity group on these molecules, the wetting ability that helps whole fluorescent probe molecule, be convenient to the application of fluorescent probe in the aqueous solution, and further deriving and the immobilized reaction active groups that provides for molecular probe.
Fluorescent molecular probe involved in the present invention, also can be by the fluorescent chemicals and 2 of reaction active groups such as other band primary amine groups, secondary amine, 6-dichloromethyl pyridine, 2,6-two bromo methyl cycloheptapyridines or 2, the derivative condensation of 6-dimethyl methyl acid esters picoline etc. and these pyridines thereof obtains.
Fluorescent molecular probe of the present invention not only has anti-transition metal and heavy metal ion to the cancellation performance of fluorophor and good selectivity, and can also be at various solution, particularly in aqueous solution to non-d
10The transition metal of electronic structure and heavy metal ion selectivity fluorescence strengthen identification, can discern transition metal ion in the mixing solutions as methyl alcohol, ethanol, acetonitrile, DMSO etc. and water and organic solvent at water, organic solvent.As embodiment 2 synthetic fluorescent molecular probe P-2 in the mixed solvent of second alcohol and water (1:1) (in Tutofusin tris-hydrochloric acid buffer solution of 0.01mol/L, pH=7.3), in multiple metal ion (as Hg
2+, Cd
2+, Pb
2+, Ag
+, Zn
2+, K
+, Na
+, Mg
2+, Ca
2+, Cr
3+, Fe
3+, Cu
2+, Co
2+, Ni
2+), to Co
2+The selectivity fluorescence that shows strengthens identification.So its cobalt ion that can be applicable in the microenvironments such as biological tissue and cell detects, the cobalt ion that also can be used in water, sewage and the aqueous solution, emulsion, the suspension system etc. detects.By The above results design of the present invention as can be seen, synthetic Co
2+Fluorescent molecular probe is very successful.As embodiment 12 synthetic fluorescent molecular probe P-12 acetonitrile, dimethyl sulfoxide (DMSO) and water (4:1:15, pH=8.6 buffer concentration are in Tutofusin tris-hydrochloric acid buffer solution of 0.01mol/L), in multiple metal ion (as Hg
2+, Cd
2+, Pb
2+, Ag
+, Zn
2+, K
+, Na
+, Mg
2+, Ca
2+, Cr
3+, Fe
3+, Cu
2+, Co
2+, Ni
2+), to Hg
2+And Cd
2+Show selectivity fluorescence and strengthen identification.
Design of the present invention, synthetic one class two 2,6-diamines pyridine derivatives fluorescent molecular probe have structure simple relatively, be easy to synthesize, in the aqueous solution, the selective fluorescence of transition metal ion strengthened recognition reaction, and can be further used for the making of immobilized identification susceptible device susceptor, can be it is immobilized in polymkeric substance or solid surface as fluorescent molecular probe of the present invention by the reactive group of molecule, realize device, be used for the key component of the fluorescent optical sensor device of above-mentioned purpose.The identification and the detection of transition metal ion in various solution and the dispersion system both can be applied to; Also can be used for the identification and the detection of transition metal ion in the biological sample.
Description of drawings
Fig. 1 is Co
2+The relation of concentration and fluorescence intensity, wherein ordinate is a fluorescence intensity, abscissa is fluorescent emission wavelength (λ
Em/ nm, excitation wavelength is 410nm).
Fig. 2 is the selectivity that fluorescent molecular probe P-2 discerns various common metal ion, and wherein ordinate is the fluorescence intensity of maximum emission wavelength place (540nm), and abscissa is different metal ion.
Embodiment
Embodiment 1
With N-butyl-4-(2-(2-hydroxyethylamino) ethylamino)-1,8-naphthalimide 180mg (0.385mmol), 2,6-dichloromethyl pyridine 70mg (0.40mmol) is dissolved in the 50ml acetonitrile, and about 8h refluxes.Steam solvent.Separate to such an extent that intermediate (P-1-1) is the deep yellow thick liquid through silica gel column chromatography, productive rate: 69% (130mg).
Above-mentioned intermediate P-1-1130mg (0.262mmol), monoethanolamine 8mg (0.13mmol) are added in the 50ml acetonitrile solvent N
2Protection, the about 7h of reflux is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, deep yellow thick liquid (P-1), productive rate: 55% (70mg).
1H-NMR(400MHz，CDCl
3)δ：8.46(d，J＝7.2Hz，2H)，8.27(d，J＝8.4Hz，2H)，8.15(d，J＝8.2Hz，2H)，7.44(m，4H)，6.95(m，4H)，6.53(br，2H)，4.16(t，J＝7.6Hz，4H)，3.6-3.8(m，14H)，3.20(br，4H)，2.98(br，6H)，2.75(br，4H)，1.70(m，4H)，1.44(m，4H)，0.97(t，J＝6.9Hz，6H).
Embodiment 2
With N-hydroxyethoxy ethyl-4-piperazinyl-1,8-naphthalimide 401mg (1.09mmol) and 2,6-dichloromethyl pyridine 229mg (1.30mmol) is added in the 50ml acetonitrile solvent, N
2Protection, the about 6h of reflux is cooled to room temperature.Reaction solution is spin-dried for, separates, obtain faint yellow solid (P-2-1) with silica gel column chromatography.Fusing point: 119.3-120.1 ℃, productive rate: 53% (290mg).
P-2-1203mg (0.398mmol), monoethanolamine 11mg (0.18mmol) are added in the 30ml acetonitrile solvent N
2Protection, the about 6h of reflux is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, yellow solid (P-2).Fusing point: 89.0～90.1 ℃, productive rate: 45% (80mg).
1H-NMR(400MHz，CDCl
3)δ：8.57(d，J＝7.2Hz，2H)，8.50(d，J＝8.4Hz，2H)，8.40(d，J＝8.4Hz，2H)，7.66(m，4H)，7.38(d，J＝7.2Hz，2H)，7.30(d，J＝10.8Hz，2H)，7.20(d，J＝8.4Hz，2H)，4.43(t，J＝5.6Hz，4H)，4.00(s，4H)，3.85(t，J＝5.6Hz，4H)，3.72(t，J＝3.2Hz，4H)，3.68(t，J＝3.2Hz，4H)，2.89(br，10H).
Embodiment 3
With N-butyl-4-piperazinyl-1,8-naphthalimide 380mg (1.126mmol) and 2,6-dichloromethyl pyridine 200mg (1.136mmol) is added in the 50ml acetonitrile solvent, N
2Protection, the about 7h of reflux is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, obtaining intermediate is yellow-green colour solid (P-3-1).Fusing point: 147.0～147.8 ℃, productive rate: 58% (290mg).
Get above-mentioned intermediate P-3-1172mg (0.360mmol), monoethanolamine 10mg (0.164mmol) is added in the 30ml acetonitrile solvent, N
2Protection, the about 6h of reflux is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, golden yellow solid (P-3).Fusing point: 85.8～86.9 ℃, productive rate: 60% (92mg).
1H-NMR(400MHz，CDCl
3)δ：8.57(d，J＝7.2Hz，2H)，8.50(d，J＝8.0Hz，2H)，8.40(d，J＝8.4Hz，2H)，7.68(t，J＝8.0，2H)，7.63(t，J＝8.0，2H)，7.39(d，J＝7.6Hz，2H)，7.30(s，2H)，7.21(d，J＝8.0Hz，2H)，4.16(t，J＝7.4Hz，4H)，4.02(br，4H)，3.85(s，4H)，3.73(t，J＝4.6Hz，2H)，3.32(br，8H)，2.94(br，2H)，2.87(br，8H)，1.70(m，4H)，1.44(m，4H)，0.97(t，J＝7.4Hz，6H).
Embodiment 4
P-3-1112mg (0.234mmol), amino ethoxy ethanol 11mg (0.104mmol) are added in the 30ml acetonitrile solvent N
2Protection adds the about 6h of hot-fluid, is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, golden yellow solid (P-4).Fusing point: 80.5～81.8 ℃, productive rate: 67% (69mg).
1H-NMR(400MHz，CDCl
3)δ(
＊10
-6)：8.57(d，J＝7.6Hz，2H)，8.50(d，J＝8.0Hz，2H)，8.39(d，J＝7.6Hz，2H)，7.70(t，J＝8.0，2H)，7.67(t，J＝8.0，2H)，7.53(d，J＝7.6Hz，2H)，7.38(d，J＝7.2Hz，2H)，7.20(d，J＝8.0Hz，2H)，4.16(t，J＝7.6Hz，4H)，3.99(br，4H)，3.82(s，4H)，3.72(t，J＝3.6Hz，2H)，3.68(t，J＝4.8Hz，2H)，3.54(t，J＝4.0Hz，2H)，3.31(br，8H)，2.89(br，10H)，1.70(m，4H)，1.44(m，4H)，0.97(t，J＝7.4Hz，6H).
Embodiment 5
Get NBD-P 200mg (0.80mmol) and 143mg (0.81mmol) 2,6-dichloromethyl pyridine is dissolved in the 60mL acetonitrile, at N
2Back flow reaction 8h under the gas.Decompression steams solvent, and silica gel column chromatography separates, and gets red powder (P-5-1), fusing point: 190 ℃ of decomposition, productive rate 56% (174mg).
Get above-mentioned intermediate P-5-1 174mg (0.45mmol), monoethanolamine 13mg (0.21mmol) is added in the 30mL acetonitrile solvent, N
2Protection, the about 6h of reflux is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, red solid (P-5).Fusing point: 146.9～148.3 ℃., productive rate: 60% (96mg).
1H?NMR(400MHz，DMSO-d
6)δ：8.48(d，J＝9.2Hz，2H)，7.76(t，J＝7.6Hz，2H)，7.41(d，J＝9.2Hz，2H)，7.35(d，J＝9.2Hz，2H)，6.65(d，J＝9.2Hz，2H)，4.16(br，8H)，4.01(s，4H)，3.74(t，J＝4.6Hz，2H)，3.70(s，4H)，2.93(br，2H)，2.70(br，8H).
Embodiment 6
With 0.324g (1.15mmol) Nap-A and 0.22g (1.25mmol) 2; 6-dichloromethyl pyridine joins in the 70mL acetonitrile; stirring and refluxing under the nitrogen protection; analyze through TLC; when reaction mixture no longer during considerable change; reaction solution is spin-dried for, separates obtaining intermediate (P-6-1) with silica gel column chromatography.Directly mix with 18mg (0.30mmol) thanomin in the 50mL acetonitrile the P-6-1 intermediate is broad then, back flow reaction is analyzed through TLC, when reaction mixture no longer during considerable change, reaction solution is spin-dried for, separate with silica gel column chromatography, obtain target product (P-6), pale yellow powder.Fusing point: 138～140 ℃, productive rate: 32% (80m).
1H-NMR(400MHz，CDCl
3)δ：8.30(d，J＝9.2Hz，4H)，8.24(d，J＝10Hz，4H)，7.38(m，8H)，7.06(t，J＝7.6Hz，2H)，6.81(d，J＝7.6Hz，2H)，6.53(d，J＝7.6Hz，2H)，5.43(s，4H)，4.60(s，4H)，3.98(s，4H)，3.73(d，J＝5.0Hz，2H)，3.56(d，J＝5.0Hz，4H)，2.94(d，J＝5.0Hz，2H)，2.82(d，J＝5.0Hz，4H)
Embodiment 7
With N-butyl-4-(2-hydroxyethylamino)-1,8-naphthalimide 120mg (0.38mmol), 2, the sodium hydride of 6-dichloromethyl pyridine 70mg (0.40mmol) and 15mg (0.38mmol) 60% is dissolved in the 50ml acetonitrile, and about 8h refluxes.Steam solvent.Separate to such an extent that intermediate (P-7-1) is a yellow solid through silica gel column chromatography, productive rate: 57% (98mg).
Above-mentioned intermediate P-7-1 98mg (0.217mmol), monoethanolamine 6mg (0.10mmol) are added in the 50ml acetonitrile solvent N
2Protection, the about 7h of reflux is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, yellow solid (P-7), fusing point: 87.0～88.5 ℃, productive rate: 69% (61mg).
1H-NMR(400MHz，CDCl
3)δ：8.57(d，J＝7.2Hz，2H)，8.45(d，J＝7.6Hz，2H)，8.17(d，J＝8.4Hz，2H)，7.6(m，4H)，7.42(d，J＝7.6Hz，2H)，7.39(d，J＝7.6Hz，2H)，6.86(d，J＝8.4Hz，2H)，4.16(t，J＝7.6Hz，4H)，3.8-4.1(br，12H)3.73(t，J＝4.6Hz，2H)，3.62(br，4H)，2.94(br，2H)，1.70(m，4H)，1.44(m，4H)，0.97(t，J＝6.9Hz，6H).
Embodiment 8
With synthetic intermediate P-3-1 320mg (0.671mmol), Na in the foregoing description 3
2S 26mg (0.33mmol) is added in the 50ml acetonitrile solvent, N
2Protection, the about 8h of reflux is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, yellow solid (P-8), fusing point: 137.1～138.5 ℃, productive rate: 35% (106mg).
1H-NMR(400MHz，CDCl
3)δ：8.56(d，J＝7.2Hz，2H)，8.50(d，J＝8.0Hz，2H)，8.39(d，J＝8.4Hz，2H)，7.7(m，4H)，7.39(d，J＝7.6Hz，2H)，7.20(m，4H)，4.16(t，J＝7.4Hz，4H)，4.0(br，4H)，3.32(br，8H)，2.87(br，8H)，1.70(m，4H)，1.44(m，4H)，0.97(t，J＝7.4Hz，6H).
Embodiment 9
With N-butyl-4-(2-ammonia ethylamino)-1,8-naphthalimide 50mg (0.161mmol) and P-2-1 180mg (0.353mmol) are added in the 30mL acetonitrile solvent N
2Protection, heated and stirred, backflow 12h is cooled to room temperature.After reaction solution is spin-dried for, carry out column chromatography and separate, products therefrom solution is spin-dried for, yellow shape solid (P-9).Fusing point: 113.6-114.7 ℃, productive rate: 47.26% (95mg).
1H-NMR(400MHz，CDCl
3)δ(
＊10
-6)：8.61(d，J＝7.2Hz，1H)，8.56(d，J＝6.8Hz，2H)，8.47(m，3H)，8.40(d，J＝8.4Hz，1H)，8.32(d，J＝8.4Hz，2H)，7.68-7.58(m，5H)，7.36(d，J＝7.2Hz，4H)，7.21(s，1H)，7.13(d，J＝8.0Hz，2H)，6.54(d，J＝8.0Hz，1H)，4.43(t，J＝5.2Hz，4H)，4.14(t，J＝7.2Hz，2H)，4.02(s，4H)，3.84(t，J＝5.6Hz，4H)，3.77(s，4H)，3.68-3.66(m，8H)，3.43(s，2H)，3.21(s，8H)，3.09(s，2H)，2.74(s，8H)，1.68(m，2H)，1.42(m，2H)，0.94(t，J＝7.2Hz，3H).
With quadrol 9mg (0.149mmol), P-2-1 350mg (0.687mmol), be added in the 30ml acetonitrile solvent N
2Protection, heated and stirred stops heating behind about 8h that refluxes, be cooled to room temperature.After reaction solution is spin-dried for, carry out column chromatography and separate, yellow solid (P-10).Fusing point: 93.1-94.3 ℃, productive rate: 26% (75mg).
1H-NMR(400MHz，CDCl
3)δ(
＊10
-6)：8.53(d，J＝6.8Hz，4H)，8.47(d，J＝8.0Hz，4H)，8.33(d，J＝8.4Hz，4H)，7.65-7.59(m，8H)，7.45(d，J＝7.6Hz，4H)，7.33(d，J＝6.4Hz，4H)，7.13(d，J＝8.4Hz，4H)，4.42(t，J＝5.6Hz，8H)，3.85-3.83(m，16H)，3.76(s，8H)，3.68-3.65(m，16H)，3.26(s，16H)，2.86(s，4H)，2.80(s，16H).
Embodiment 11
P-2-1242mg (0.475mmol), piperazine 20mg (0.232mmol) are added in the 30mL acetonitrile solvent N
2Protection, heated and stirred, backflow 6h is cooled to room temperature.After reaction solution is spin-dried for, separate with silica gel column chromatography, yellow solid (P-11).Fusing point: 108.3-109.0 ℃, productive rate: 81% (194mg).
1H-NMR(400MHz，CDCl
3)δ(
＊10
-6)：8.53(d，J＝6.8Hz，2H)，8.47(d，J＝8.0Hz，2H)，8.33(d，J＝8.4Hz，2H)，7.65-7.59(m，4H)，7.45(d，J＝7.6Hz，2H)，7.33(d，J＝6.4Hz，2H)，7.13(d，J＝8.4Hz，2H)，4.42(t，J＝5.6Hz，4H)，3.82-3.88(m，20H)，3.74(br，8H)，2.85(br，16H).
Embodiment 12
Pery-A 50mg (0.069mmol) and P-2-1 86mg (0.17mmol) are added among the 20mL DMF N
2Protection, 80 ℃ are reacted 10h down, are cooled to room temperature, and reaction solution is spin-dried for, and separate with silica gel column chromatography, get red solid (P-12).Fusing point: 132.1～133.8 ℃, productive rate: 26% (30mg).
1H-NMR(400MHz，CDCl
3)δ：9.40(d，J＝3.2Hz，2H)，8.77(s，2H)，8.58(d，J＝8.4Hz，2H)，8.49(d，J＝7.2Hz，2H)，8.42(d，J＝8.0Hz，2H)，8.25(d，J＝8.4Hz，2H)，7.58(t，J＝8.0Hz，2H)，7.32(d，J＝5.2Hz，2H)，7.14(d，J＝7.2Hz，4H)，7.09(d，J＝8.0Hz，2H)，4.39(t，J＝5.6Hz，4H)，4.33(t，J＝5.6Hz，4H)，3.93(s，4H)，3.82(t，J＝5.6Hz，4H)，3.70-3.65(m，16H)，3.19(s，8H)，3.06(t，J＝5.6Hz，4H)，2.93(s，4H)，2.72(s，8H).
Embodiment 13
Fluorescent molecular probe P-2 among the embodiment 2 is dissolved in the mixed solvent (1:1) of the second alcohol and water that contains Tri(Hydroxymethyl) Amino Methane Hydrochloride (0.01mol/L) buffered soln (pH=7.3), being mixed with concentration is 1.0 * 10
-5Its fluorescence of the mensuration of mol/L (excitation wavelength is 410nm, and maximum emission wavelength is 540nm).Cobalt salt (the CoCl that adds different amounts
26H
2O) fluorescence intensity of solution increases thereupon the time, please refer to Fig. 1.
Embodiment 14
Under the test condition in embodiment 13, research fluorescent molecular probe P-2 discerns the selectivity of various common metal ion.The concentration of fluorescent molecular probe P-2 is 1.0 * 10
-5Mol/L adds Na respectively
+, K
+, Mg
2+, Ca
2+, Cr
3+, Fe
3+, Co
2+, Ni
2+, Cu
2+, Zn
2+, Ag
+, Cd
2+, Hg
2+And Pb
2+Solution, making its concentration is 5.0 * 10
-5Mol/L.By the discovery of comparing with the contrast experiment who does not add metal ion, remove Co
2+The influence of other ion pair fluorescence intensity in addition in one times, Co
2+Make fluorescence intensity of solution be increased to original ten times, see also Fig. 2.This shows that fluorescent molecular probe of the present invention is to Co
2+The good selectivity recognition performance that has.
Below be the application of fluorescent probe P-4 in RESEARCH ON CELL-BIOLOGY.People Hela cell with cultivation is a research object, with Hela cell (1 * 10
4) being inoculated in 24 porocyte culture plates, every hole contains 1mL RPMl1640 substratum.Place CO
2Incubator is cultivated 24 hours or 70% above cell attachment for 37 ℃.Add CoCl
2The aqueous solution is respectively 0.01,0.1,0.5 and 1.0mg/mL to final concentration.Control wells adds equal-volume physiological saline.Continue to cultivate after 30 minutes, add fluorescent probe P-4, make its concentration reach 1 μ M.Observe with the fluorescence inverted microscope, occur obvious fluorescence in the test holes cell, the fluoroscopic image of cell is not seen in contrast.Culture plate is placed Genious, and (TECAN, Swiss) multi-functional plate reading machine excite down at 410nm, record 540nm fluorescent emission intensity.The result shows, 0.01,0.1,0.5 and the fluorescence intensity of 1.0mg/mL culture hole cell apparently higher than control wells, reach 1-10 doubly, and present the concentration dependence.
Embodiment 16
Fluorescent molecular probe P-10 among the embodiment 10 is dissolved in the mixing solutions of second alcohol and water (1:1), contain Tutofusin tris-hydrochloric acid (0.01mol/L, pH=7.9).The concentration of P-10 is 5 * 10
-6Mol/L, concentration of metal ions are 5 * 10
-5Mol/L, in multiple metal ion (as Hg
2+, Cd
2+, Pb
2+, Ag
+, Zn
2+, K
+, Na
+, Mg
2+, Ca
2+, Cr
3+, Fe
3+, Cu
2+, Co
2+, Ni
2+), to Co
2+Show selectivity fluorescence and strengthen identification.Recording its fluorescence (excitation wavelength is 410nm, and maximum emission wavelength is 545nm) intensity increases about 10 times than blank value respectively, and the influence of other metal ion is in 1 times.
Embodiment 17
Fluorescent molecular probe P-11 among the embodiment 11 is dissolved in the mixing solutions of acetonitrile, dimethyl sulfoxide (DMSO) and water (4:1:15), contain Tutofusin tris-hydrochloric acid (0.01mol/L, pH=8.6).The concentration of P-12 is 5 * 10
-6Mol/L, concentration of metal ions are 5 * 10
-5Mol/L, in multiple metal ion (as Hg
2+, Cd
2+, Pb
2+, Ag
+, Zn
2+, K
+, Na
+, Mg
2+, Ca
2+, Cr
3+, Fe
3+, Cu
2+, Co
2+, Ni
2+, Al
3+), to Hg
2+And Cd
2+Show selectivity fluorescence and strengthen identification.Record its fluorescence (excitation wavelength is 410nm, and maximum emission wavelength is 553nm) intensity and increase by 10 times and 2 times than blank value respectively.
Embodiment 18
Fluorescent molecular probe P-12 among the embodiment 12 is dissolved in the mixing solutions of acetonitrile, dimethyl sulfoxide (DMSO) and water (4:1:15), contain Tutofusin tris-hydrochloric acid (0.01mol/L, pH=8.6).The concentration of P-12 is 5 * 10
-6Mol/L, concentration of metal ions are 5 * 10
-5Mol/L, in multiple metal ion (as Hg
2+, Cd
2+, Pb
2+, Ag
+, Zn
2+, K
+, Na
+, Mg
2+, Ca
2+, Cr
3+, Fe
3+, Cu
2+, Co
2+, Ni
2+), to Hg
2+And Cd
2+Show selectivity fluorescence and strengthen identification.Record its fluorescence (excitation wavelength is 410nm, and maximum emission wavelength is 545nm) intensity and increase by 4.5 times and 3 times than blank value respectively.
Claims (7)
1, a kind of fluorescent molecular probe is characterized in that this fluorescent molecular probe has following general formula of molecular structure:
In the general formula:
R
1, R
2, R
3Represent hydrogen, carbonatoms be 1-20 alkyl, carbonatoms be 1-12 hydroxyalkyl, carbonatoms be 1-12 carboxyalkyl, the polymerization degree at 1-40 polyoxyethylene groups, aryl, aralkyl, or unsubstituted,
X
1, X
2Represent nitrogen-atoms or Sauerstoffatom,
X
3The carbonatoms of representing nitrogen-atoms, sulphur atom or two ends to have nitrogen to replace is 1-30 Alkylenediamine,
F
1, F
2Represent fluorophor, be respectively 4-amino-1,8-naphthalimide, naphthalene, anthracene, pyrene, benzo furazan, 3,4,9,10-perylene tetracid imide, tonka bean camphor, dansyl amide, fluorescein, rhodamine, fluorine boron pyrroles, benzoglyoxaline, carbazole, benzoxazoles,
Q
1, Q
2Representing carbonatoms is 1-10 alkylidene group.
2, according to the described a kind of fluorescent molecular probe of claim 1, it is characterized in that: described R
1, R
2Represent hydrogen; Q
1, Q
2Represent ethylene group respectively.
3, according to the described a kind of fluorescent molecular probe of claim 1, it is characterized in that: described fluorophor is a 4-amino-1,8-naphthalimide, benzo furazan or anthracene.
4, according to the described a kind of fluorescent molecular probe of claim 1, it is characterized in that: described R
1, R
2Represent hydroxyethyl; Q
1, Q
2Represent methylene radical; F
1And F
2Represent anthracene or pyrene fluorophor.
5, according to the described a kind of fluorescent molecular probe of claim 1, it is characterized in that: described Alkylenediamine is 3,4,9 of quadrol, piperazine or diethylamine replacement, 10-perylene tetracid imide.
6, according to the described a kind of fluorescent molecular probe of claim 1, it is characterized in that: described aralkyl is a 4-amino-1,8-naphthalimide naphthalene, anthracene, pyrene, benzo furazan, tonka bean camphor, dansyl amide, fluorescein, rhodamine or fluorine boron pyrroles.
7, a kind ofly in solution, transition metal and heavy metal ion are carried out application aspect selectivity identification and the mensuration according to the described fluorescent molecular probe of claim 1.
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| CN101440062B (en) * | 2007-11-23 | 2011-09-28 | 齐齐哈尔大学 | Synthesis of N-acyl-8-amino quinoline derivatives and use thereof as fluorescent molecular probe |
| SG183949A1 (en) * | 2010-03-24 | 2012-10-30 | Merck Patent Gmbh | Polymers of 8,9-dihydrobenzo[def]carbazole and their use as organic semiconductors |
| CN101885797B (en) * | 2010-07-09 | 2011-11-23 | 浙江大学 | Functional polymer and preparation method and application thereof |
| CN102072898B (en) * | 2010-08-20 | 2012-08-29 | 沈阳大学 | Self-assembly type fluorescenct chemical sensor |
| CN102507526B (en) * | 2011-11-30 | 2013-11-27 | 齐齐哈尔大学 | Fluorescence detection method of Hg<2+> by riboflavin |
| EP2778161B1 (en) | 2012-03-05 | 2015-05-06 | Dalian University Of Technology | Two-photon fluorescent probe using naphthalene as matrix and preparation method and use thereof |
| CN102735663B (en) * | 2012-07-19 | 2014-07-09 | 贵州大学 | High sensitivity and high selectivity fluorescence quenching spectrum analysis method for cobalt ions |
| CN103013496B (en) * | 2012-12-14 | 2014-10-29 | 江苏大学 | Sulfhydryl group ion fuorescence probe and synthesis method thereof |
| CN103013495B (en) * | 2012-12-14 | 2014-07-30 | 江苏大学 | Copper ion fluorescence probe and synthetic method thereof |
| CN103087346A (en) * | 2013-01-29 | 2013-05-08 | 温州市质量技术监督检测院 | Preparation method of blotting membrane for Hg<2+> detection |
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