CN107619397A - Compound with FRET performance and application thereof - Google Patents
Compound with FRET performance and application thereof Download PDFInfo
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Abstract
The present invention relates to a kind of compound with FRET performance and application thereof.The compound is built-up by appropriate linking arm by trimethine cyanine and Cyanine dyestuff.Compound provided by the invention can be used for detection active oxygen and active nitrogen, not by concentration and probe concentration, the influence of the factor such as background fluorescence and instrument error, and have higher FRET efficiency.
Description
Technical field
The present invention relates to a kind of compound with FRET (FRET) performance and application thereof；Specifically
Say, be related to a kind of compound by trimethine cyanine (Cy3) and Cyanine dyestuff (Cy5) structure with FRET performances and
Its purposes.
Background technology
Active oxygen is the oxygen radical and its derivative directly or indirectly formed by electronics transfer by oxygen.Common
Active oxygen (ROS) and active nitrogen (RNS) include：Hypochlorite (ClO-), peroxynitrite (OONO-), hydrogen peroxide
(H2O2), singlet oxygen (1O2), superoxide anion (O2-), nitric oxide (NO), t-butylperoxy (ROO), hydrogen-oxygen freedom
Base (OH) etc..
In vivo, the main source of active oxygen is the mitochondria of neutrophil leucocyte, phagocyte etc., endoplasmic reticulum and thin
After birth.In organism, ROS and RNS can be produced, shift and consumed by the way that aerobic activity is lasting, be in nuclear body
The main molecules source of redox reaction.They can react with various biomolecules, on the one hand can regulate and control physiology
The signal transduction of process, on the other hand will also result in many pathology damages, for example, the too high levels of active oxygen or its result from
The balance of elimination is destroyed all to cause the oxidative damage of DNA, protein etc., so cause diabetes, artery sclerosis, cancer and
The diseases such as nervus retrogression.
Therefore, the fluorescence probe for developing reliable, sensitive, high selectivity detection ROS and RNS will be helpful to preferably study
ROS and RNS distributions in vivo and operative condition, and potential provide research tool for the early diagnosis of relevant disease.It is standby
Paid close attention to by scientist.
However, existing detection ROS and RNS fluorescence probe relies on single channel detection mostly, the increasing of fluorescence intensity is only relied on
By force, it is larger by the interference of concentration and probe concentration, cell micro-environment etc., therefore such probe still has room for improvement.
The content of the invention
Screening of the present inventor through substantial amounts of parent compound, and the parent compound obtained to screening is fitted
When chemical modification after, obtain a kind of compound.Such compound has FRET (FRET) effect, can compare
Rate type detects ROS and RNS, not by concentration and probe concentration, the influence of the factor such as background fluorescence and instrument error, and has higher FRET
Efficiency.
A purpose of the invention is, there is provided a kind of structure it is novel and the compound with higher FRET efficiency.
Described compound is compound shown in formula Z or its pharmaceutically acceptable salt：
In formula Z, R1~R16It is respectively and independently selected from：Hydrogen (H), halogen (F, Cl, Br or I), amino (- NH2), C1~C6Straight chain
Or branched alkyl, C1~C6Straight or branched alkoxyl, carboxyl (- COOH) or sulfonic group (- SO3H it is a kind of in)；
RaAnd RbIt is respectively and independently selected from：C1~C6Straight or branched alkyl or by carboxyl (- COOH) or/and sulfonic group (-
SO3H) the C of substitution1~C6It is a kind of in straight or branched alkyl；
X is NH, O, S, methylene (CH2) or by C1~C3The methylene of straight or branched alkyl substitution；
M is 1~5 integer；N is 1~5 integer；Y is the heterocyclic radical containing at least two nitrogen (N) of divalence；P for it is cloudy from
Sub- acid group (such as halogen (F, Cl, Br or I) or oxygen-containing acid group).
Another object of the present invention is, discloses a kind of purposes of the above-mentioned compound with FRET performances, i.e.,：Formula Z institutes
Show compound or its pharmaceutically acceptable salt prepare detection active oxygen (ROS) and active nitrogen (RNS) fluorescence probe in
Application；And compound shown in Formulas I or its pharmaceutically fluorescence probe of acceptable salt as detection ROS and RNS
Using.
In addition, the method that the present invention also provides compound shown in a kind of formula Z, methods described comprise the following steps：
(1) compound (one of raw material) prepares intermediate as shown in formula A：Shown in formula B the step of compound；
(2) compound (the two of raw material) prepares intermediate as shown in formula C：Compound shown in compound shown in formula D and formula E
Step；
(3) as shown in formula F compound (the three of raw material) withReaction, shown in obtained formula G the step of compound；
(4) compound is reacted with compound shown in formula D as shown in formula G, shown in obtained formula H the step of compound；
(5) compound shown in formula B is reacted with compound shown in formula H, shown in obtained Formulas I the step of compound；
(6) as shown in formula J compound (its prepare referring to step (2)) withReaction, chemical combination shown in formula K is made
The step of thing；
(7) compound (it is prepared referring to step (1)) is reacted with compound shown in formula K as shown in formula L, is made shown in formula M
The step of compound；With.
(8) compound shown in compound shown in formula M and Formulas I is reacted, the step of object (compound shown in formula Z) is made.
Brief description of the drawings
Fig. 1 are that compound Z-1 (probe PNcy3cy5) (contains 0.2%DMF) in pH=7.4 phosphate buffer solution
Detect OONO-Ultraviolet spectra response diagram.
Fig. 2 are that probe PNcy3cy5 (2 μM) (containing 0.2%DMF) in pH=7.4 phosphate buffer solution is detected
OONO-The fluorescence emission spectrum response diagram (excitation wavelength 530nm) of (0-5eq).
Fig. 3 are that compound I-1 (probe cy3BOC) (containing 0.2%DMF) in pH=7.4 phosphate buffer solution is examined
Survey ClO-Ultraviolet spectra response diagram.
Embodiment
In a preferable technical scheme of the invention, R1~R16It is H.
In presently preferred technical scheme, RaAnd RbIt is respectively and independently selected from：C1~C3Straight or branched alkyl
Middle one kind；
Further preferred technical scheme is：RaAnd RbIt is respectively and independently selected from：It is a kind of in methyl or ethyl.
In a further preferred technical solution of the present invention, X is C (CH3)2。
In a further preferred technical solution of the present invention, Y is 5~6 circle heterocycles bases containing at least two N of divalence；
Further preferred technical scheme is：Y is 5~6 circle heterocycles bases containing at least two N of divalence；
Technical scheme still more preferably is：Y is the 6 circle heterocycles bases containing at least two N of divalence；
The Y that the present invention recommends be divalence piperazine (Curve mark is substitution position).
In a further preferred technical solution of the present invention, active oxygen (ROS) described previously includes：From hypochlorous acid or it is common
Yoke alkali (ClO-/ HClO), peroxynitrite or its conjugate base (HOONO/OONO-), hydrogen peroxide (H2O2), singlet oxygen (1O2),
Superoxide anion (O2-), nitric oxide (NO), t-butylperoxy (ROO) and hydroxyl free radical (OH).
In a further preferred technical solution of the present invention, active nitrogen (RNS) described previously includes：Peroxynitrite or its
Conjugate base (HOONO/OONO-) and nitric oxide (NO).
Below by implementing to be further elaborated the present invention, its purpose, which is only that, is best understood from present disclosure.Cause
This, the cited case does not limit the scope of the invention.
Embodiment 1
(1) synthesis (compound D-1) of 2,3,3- trimethyls -3H- indoles
30mL absolute ethyl alcohol, phenylhydrazine (9.82mL, 0.1mol) (compound C- are sequentially added in 100mL single port bottle
1), 3- methyl -2- butanone (12mL, 0.11mol), 3 drop glacial acetic acid is added, are heated to reflux, TLC tracks to raw material conversion, cold
But ethanol is evaporated under reduced pressure afterwards, obtains red oil；Continue to add 30mL glacial acetic acid in red oil, argon gas is protected
Shield, is heated to reflux, and TLC tracks to raw material conversion, and water pump, which is evaporated under reduced pressure, after cooling removes most of glacial acetic acid, is then slowly added into
The NaHCO of saturation3The aqueous solution, regulation pH to neutrality.Extracted (3 × 20mL) with absolute ether, merge organic layer, anhydrous Na2SO4
Dry, obtain dark red oily crude product, column chromatography for separation (PE/EA=40:3, v/v；Rf=0.15 pale yellowish oil) is obtained
Liquid 12.7g (compound D-1), yield 80%.
1H NMR (400MHz, CDCl3)：δ 7.55 (d, J=8.0Hz, 1H), 7.26~7.29 (m, 2H), 7.20 (dd, J=
8.0Hz, 1H), 2.28 (s, 3H), 1.29 (s, 6H).
13C NMR (100MHz, CDCl3)：δ 15.18,23.01,53.54,119.79,121.26,125.16,127.57,
145.52 153.32,188.11.
GC-MS：Calculated value C11H13N [M]=159.1048, experiment value [M]=159.1.
(2) synthesis of 1- ethyls -2,3,3- tri-methyl indoles (compound E-1)
Sequentially add compound D-1 (4.773g, 30mmol) in 250mL single port bottle, iodoethane (24.11mL,
300mmol), acetonitrile 60mL, argon gas protection, is heated to reflux, and TLC tracks to raw material conversion, and iodoethane is spin-dried for after cooling, filters,
Filter cake is washed with acetone, collects filter cake, and ethyl alcohol recrystallization obtains yellow flat crystal 4.246g (compound E-1), yield 45%.
1H NMR (400MHz, CDCl3)：δ 7.77 (d, J=4.0Hz, 1H), 7.60~7.61 (m, 3H), 4.76 (q, J=
8.0Hz, 2H), 3.16 (s, 3H), 1.62~1.66 (m, 9H).
13C NMR (100MHz, CDCl3)：δ 13.70,17.00,23.10,45.37,54.66,115.39,123.49,
129.54,130.16,140.49,141.66,195.38.
LC-MS：Calculated value C13H18N+I-[M-I]=188.1434, experiment value：188.1.
(3) N, N '-amitraz diphenylurea synthesis
Sequentially add aniline (18.3mL, 200mmol) in 100mL single port bottles, triethyl orthoformate (16.6mL,
100mmol), heating reflux reaction, TLC track to raw material conversion, a large amount of solids are separated out after cooling, filter, the anhydrous second of filter cake
Alcohol washs, and collects filter cake, and absolute ethyl alcohol is recrystallized to give white needles 14.7g (title compound), yield 75%.
1H NMR (400MHz, CDCl3)：δ 10.31 (s, 1H), 8.33 (s, 1H), 7.38 (t, J=8.0Hz, 4H), 7.17
(t, J=8.0Hz, 2H), 7.13 (d, J=8.0Hz, 4H).
13C NMR (100MHz, CDCl3)：δ 119.31,123.41,129.46,145.43,150.34.
LC-MS：Calculated value：C13H12N2[M+H]=197.1000, experiment value：[M]=197.1.
(4) synthesis of 1- ethyls -2- (β-anilino-) vinyl -3,3- dimethyl indoles (compound B-1)
Compound E-1 (4.05g, 12.8mmol), N, N are sequentially added in 250mL single-necked flasks '-amitraz diphenylurea
(2.76g, 14.1mmol), glacial acetic acid 60mL, argon filling gas shielded, it is heated to reflux, TLC tracks to raw material conversion, and decompression rotation is steamed
Hair removes most of glacial acetic acid, uses CH2Cl2Extract (3 × 70mL), merge organic layer, anhydrous Na2SO4Dry, column chromatography for separation
(PE/EA=2:1, DCM/EA=3:1, v/v) light yellow bits shape solid 1.38g (compound B-1), yield 26%, are obtained.
1H NMR (400MHz, DMSO)：δ 11.92 (s, 1H), 8.70 (d, J=12.0Hz, 1H), 7.71 (d, J=
8.0Hz, 1H), 7.46~7.59 (m, 6H), 7.29~7.37 (m, 2H), 6.27 (d, J=16.0Hz, 1H), 4.19 (d, J=
8.0Hz, 2H), 1.72 (s, 6H), 1.36 (t, J=8.0Hz, 3H).
13C NMR (100MHz, DMSO)：δ 12.34,28.10,50.06,91.33,112.11,118.68,123.1,
126.5,129.06,130.3,138.79,141.5,141.6,152.18,177.73.
HRMS calculated values C20H23N2 +I-The experiment value of [M-I]=291.1856 291.1869.
(5) synthesis of 4- (the bromo- acetyl of 2-)-piperazine -1- acetic acid-tetrabutyl ester (compound G-1)
Boc- piperazines (compound F-1,0.558g, 3mmol) are weighed in 25mL beaker and are dissolved in the anhydrous CH of 10mL2Cl2
In, claim anhydrous Na2CO3(0.848g, 8mmol) pours into the two in 100mL single port bottles in 7mL deionized waters, is stirred under ice bath
20min is mixed, bromoacetyl bromide (0.568mL, 6.6mmol) is dissolved in the anhydrous CH of 10mL2Cl2In, it is quick to instill in single port bottle, it is added dropwise
After add DMAP (36.7mg, 0.3mmol), continue to stir, TLC track to raw material conversion, be transferred to room temperature reaction 1h.
Use CH2Cl2Extract (3 × 30mL), merge organic layer, anhydrous Na2SO4Dry, column chromatography for separation (CH2Cl2/ MeOH=60:1, v/
V) Off-white solid 598mg (compound G-1), yield 65% are obtained.
1H NMR (400MHz, CDCl3)：δ 3.85 (s, 2H), 3.56 (t, J=8.0Hz, 2H), 3.47 (d, J=4.0Hz,
2H), 1.44 (s, 9H).
13C NMR (100MHz, CDCl3)：δ 25.62,28.35,41.93,46.55,80.45,154.45,165.42.
GC-MS：Calculated value C11H19BrN2O3[M]=306.0579, experiment value [M]=306.0.
(6) compound H-1 synthesis：
Compound D-1 (477mg, 3mmol), compound G-1 (1.38g, 4.5mmol), second are added in 50mL single port bottles
Nitrile 15mL, argon filling gas shielded, is heated to reflux, and TLC tracks to raw material conversion, and decompression rotary evaporation removes acetonitrile, and residue is used up
Measure few CH2Cl2Dissolving, a large amount of ether are added, separate out solid, solid is carefully sucked into conical flask, solid must be made
Surface still with the presence of ether, is immediately dried up ether with argon gas, and conical flask is preserved with ParafilmTM isolation air.Product is
Pink solid 700mg (compound H-1), yield 50%.
1H NMR (400MHz, CDCl3)：δ 7.53 (s, 4H), 6.37 (s, 2H), 3.45~3.89 (m, 8H), 2.98 (s,
3H), 1.65 (s, 6H), 1.47 (s, 9H).
LCMS calculated values C22H32N3O3 +Br-The experiment value of [M-Br]=386.2438 386.2.
(7) compound I-1 synthesis：
Sequentially add compound B-1 (1.57g, 3.76mmol) in 50mL single-necked flasks, compound H-1 (2.17g,
4.67mmol), aceticanhydride 8mL, pyridine 8mL, it is heated to reflux under argon gas protection, TLC tracks to raw material conversion, and rotary evaporation removes vinegar
Acid anhydride and pyridine, column chromatography for separation (EA, Rf=0.2), obtain red bits shape solid 615mg (compound I-1), yield 28%.
1H NMR (400MHz, CD3OD)：δ 8.51 (t, J=4.0Hz, 1H), 7.54 (t, J=4.0Hz, 2H), 7.26~
7.44 (m, 6H), 6.72 (d, J=16.0Hz, 1H), 6.60 (d, J=12.0Hz, 1H), 5.28 (s, 2H), 4.24 (d, J=
8.0Hz, 2H), 3.76 (d, J=8.0Hz, 4H), 3.60 (s, 2H), 3.51 (s, 2H), 1.79 (s, 6H), 1.73 (s, 6H),
1.49 (s, 9H), 1.40 (t, J=8.0Hz, 3H).
13C NMR (100MHz, CD3OD)：δ 11.68,26.84,27.20,27.32,39.31,41.86,44.66,
45.69,49.20,49.36,80.30,102.85,103.00,110.68,111.09,122.00,122.26,124.99,
125.59,128.39,128.67,140.27,141.09,141.44,142.65,151.08,154.83,163.88,174.62,
175.76。
HRMS calculated values C36H47N4O3 +[M+]=583.3643, experiment value [M+]=583.3655.
(8) synthesis of compound (compound I-1a) shown in Formulas I -1a：
Compound I-1 (500mg, 0.7mmol), anhydrous CH are sequentially added in two mouthfuls of flasks of 50mL2Cl210mL, applying argon gas
Protection, by the CH of trifluoroacetic acid2Cl2Solution (1:5, v/v) slowly it is added dropwise as in flask, TLC tracks to raw material conversion, decompression rotation
Turn evaporation, column chromatography for separation (CH2Cl2/ MeOH=20:1v/v) obtain red bits shape product 300mg (compound I-1a), yield
70%.
1H NMR (400MHz, CD3OD)：δ 8.57 (t, J=12.0Hz, 1H), 7.56 (d, J=4.0Hz, 1H), 7.53 (d,
J=8.0Hz, 1H), 7.46 (t, J=8.0Hz, 1H), 7.32~7.41 (m, 3H), 7.28 (t, J=8.0Hz, 2H), 5.27 (s,
2H), 4.25 (q, J=8.0Hz, 2H), 4.03 (s, 2H), 3.89 (s, 2H), 3.55 (s, 2H), 1.82 (s, 6H), 1.76 (s,
6H), 1.42 (t, J=8.0Hz, 3H).
13C NMR (100MHz, CD3OD)：δ 11.50,26.74,38.73,39.21,41.72,42.79,43.01,
45.27,49.19,49.43,102.63,102.90,110.60,111.11,121.95,122.22,125.01,125.69,
128.36,128.68,140.22,141.12,141.40,142.57,151.20,163.98,174.89,175.82.
HRMS calculated values C31H39N4O+[M+]=483.3118, experiment value 483.3138
(9) synthesis of 1- (ε-carboxylic amyl group) -2,3,3- tri-methyl indoles (compound K -1)
6- bromocaproic acids (4.85g, 25mmol) are weighed, are dissolved in 10mL o-dichlorohenzenes, addition compound D-1 (1.59g,
110 DEG C 10mmol) are heated to, argon gas protection is lower to react, and TLC tracks to raw material conversion, after cooling, carefully pours out adjacent dichloro
Benzene, residue are dried in infrared baking oven, obtain dark brown product 1.3g (compound K -1), yield 37%.
1H NMR (400MHz, DMSO)：δ 12.04 (s, 1H), 8.01 (d, J=4.0Hz, 1H), 7.86 (d, J=4.0Hz,
2H), 4.48 (t, J=8.0Hz, 2H), 2.88 (s, 3H), 2.22 (t, J=8.0Hz, 2H), 1.84 (t, J=8.0Hz, 2H),
1.54 (s, 8H), 1.44 (s, 2H).
13C NMR (100MHz, DMSO)：δ 14.73,22.49,24.50,25.86,27.43,33.86,47.99,54.64,
116.02,124.02,129.39,129.83,141.51,142.33,174.79,197.00.
HRMS calculated values C17H24NO2 +Br-[M-Br]=274.1802, experiment value 274.1806.
(10) synthesis of 1- ethyls -2- (δ-anilino-) vinyl -3,3- dimethyl indoles (compound L -1)：
Compound E-1 (6.3g, 20mmol), β-anilino- malonaldehyde dianil salt are sequentially added in 250mL single port bottles
Hydrochlorate (5.69g, 22mmol), glacial acetic acid 50mL, argon filling gas shielded, is heated to reflux, and TLC tracks to raw material conversion, decompression rotation
Evaporation removes most of glacial acetic acid, uses CH2Cl2Extract (3 × 100mL), merge organic layer, anhydrous Na2SO4Dry, column chromatography point
From (EA, Rf=0.2), blackish green bits shape solid 1.678g (compound L -1), yield 20% are obtained.
1H NMR (400MHz, DMSO)：δ 12.24 (s, 1H), 8.87 (d, J=12.0Hz, 1H), 8.36 (t, J=
12.0Hz, 1H), 7.62 (d, J=4.0Hz, 1H), 7.41~7.46 (m, 6H), 7.21~7.27 (m, 2H), 6.38~6.47
(m, 2H), 4.16 (q, J=8.0Hz, 2H), 1.67 (s, 6H), 1.27 (t, J=8.0Hz, 3H).
13C NMR (100MHz, DMSO)：δ 12.85,27.58,49.47,101.50,111.46,117.88,122.96,
125.29,125.73,128.91,130.18,140.25,141.57,141.88,156.16,157.89,174.11.
HRMS calculated values C22H25N2 +I-The experiment value of [M-I]=317.2012 317.2017.
(11) synthesis of the carbon cyanines (compound M-1) of 1- (ε-carboxylic amyl group) -1 '-ethyl -3,3,3 ' 3 '-tetramethyl indoles two：
Sequentially add compound L -1 (1.005g, 2.2mmol) in 100mL single port bottles, compound K -1 (1g,
2.8mmol), aceticanhydride 10mL, pyridine 10mL, argon filling gas shielded, it is heated to reflux, TLC tracks to raw material conversion, depressurizes rotary evaporation
Remove aceticanhydride and pyridine, column chromatography for separation (CH2Cl2/ MeOH=30:1, v/v) blueness bits shape solid (compound M-1) is obtained
466mg, yield 42%.
1H NMR (400MHz, CD3OD)：δ 8.25 (t, J=16.0Hz, 2H), 7.49 (dd, J1=4.0Hz, J2=
4.0Hz, 2H), 7.38~7.43 (m, 2H), 7.24~7.30 (m, 4H), 6.64 (t, J=12.0Hz, 1H), 6.30 (dd, J1=
J2=4.0Hz, 2H), 4.15 (q, J=8.0Hz, 2H), 4.10 (t, J=8.0Hz, 2H), 2.25 (t, J=4.0Hz, 2H),
1.83 (t, J=8.0Hz, 2H), 1.72 (s, 12H), 1.69 (q, J=8.0Hz, 2H), 1.49 (t, J=8.0Hz, 2H), 1.38
(t, J=4.0Hz, 3H).
13C NMR (100MHz, CD3OD)：δ 11.28,25.06,26.55,26.65,26.86,35.28,38.66,
43.52,49.16,102.72,163.01,110.43,110.87,122.03,122.08,124.81,124.86,125.37,
128.36,128.39,141.29,141.42,141.69,142.16,154.05,154.15,172.87,731.21,178.75.
HRMS calculated values C33H40N2O2The experiment value of [M+H]=497.3168 497.3179.
(12) synthesis of target compound (compound Z-1)
Sequentially add compound M-1 (253mg, 0.51mmol) in 100mL there-necked flask, HBTU (580mg,
1.53mmol), dry DMF 10mL, argon filling gas shielded, 30min is stirred under ice bath；Weigh Compound I-1a (467mg,
0.77mmol), it is dissolved in 10mL dry DMFs, and is added dropwise under ice bath in there-necked flask；Weigh again DIPEA (0.18mL,
1mmol), it is dissolved in 5mL dry DMFs, and is added dropwise under ice bath in there-necked flask.After stirring 6h under ice bath, ice bath, room temperature are removed
It is stirred overnight at 25 DEG C, TLC tracks to raw material conversion, and decompression rotary evaporation removes most of DMF, uses CH2Cl2Extraction (3 ×
40mL), organic layer, anhydrous Na are merged2SO4Dry, column chromatography for separation (CH2Cl2/ MeOH=100:1v/v) obtain violet solid
(compound Z-1) 200mg, yield 35%, purity (HPLC) are 99%.
1H NMR (400MHz, CDCl3)：δ 8.42 (t, J=12.0Hz, 1H), 7.94 (t, J=12.0Hz, 2H), 7.34~
7.39 (m, 8H), 7.05~7.27 (m, 8H), 6.30 (s, 2H), 4.11 (q, J=8.0Hz, 2H), 4.03 (t, J=8.0Hz,
2H), 3.63~3.80 (m, 8H), 2.45 (q, J=8.0Hz, 2H), 1.85 (m, 2H), 1.77 (s, 6H), 1.72 (s, 5H),
1.70 (s, 11H), 1.54 (t, J=8.0Hz, 2H), 1.40 (q, J=8.0Hz, 6H).
13C NMR (100MHz, CDCl3)：δ 175.62,174.01,173.03,172.60,171.63,163.04,
153.39,153.32,151.15,142.46,141.97,141.58,141.47,141.40,141.27,140.83,139.89,
129.04,128.96,128.72,128.65,125.87,125.68,125.27,125.19,122.36,122.29,121.89,
110.98,110.87,110.71,110.38,103.86,103.79,103.47,103.35,103.10,49.43,49.40,
49.28,49.24,45.31,44.86,44.55,44.08,42.15,40.93,39.52,39.15,32.79,32.53,
29.69,28.27,27.91,27.86,27.75,27.71,26.51,24.86,24.71,12.43,12.25.
HRMS calculated values C64H78N6O22 2+[M2+]=962.6175, experiment value [m/z]=481.2429.
Detection of the probe to active oxygen
Embodiment 2
By probe PNcy3cy5 (compound Z-1) after vacuum drying and weighing, it is placed in 10mL volumetric flasks, with N, N-
Dimethylformamide constant volume, is made into 10-3M solution is as probe mother liquor.It is configured to determine UV absorption light after 2 μM of solution respectively
Spectrum and fluorescence emission spectrum.Concrete outcome is shown in Fig. 1 and Fig. 2.
Embodiment 3
By compound I-1 (probe cy3BOC) after vacuum drying and weighing, it is placed in 10mL volumetric flasks, with N, N- bis-
NMF constant volume, is made into 10-3M solution is as probe mother liquor.Ultra-violet absorption spectrum is determined respectively after being configured to 2 μM of solution
And fluorescence emission spectrum.Concrete outcome is shown in Fig. 3.
The FRET performances of probe
Embodiment 4
Connection arm lengths in probe PNcy3cy5 of the present invention between Cy3 and Cy5 are about 14 angstroms, reported in the literature
It is about 21 angstroms of (Fu, N. to connect arm lengths；Xiong,Y.；Squier,T.C.J.Am.Chem.Soc.2012,134,18530；
Conley,N.R.；Biteen,J.S.；Moerner,W.E.The Journal of Physical Chemistry B 2008,
112,11878)。
For probe PNcy3cy5 of the present invention, existing detection ROS and RNS fluorescence probe, especially
Detect peroxynitrite (OONO-) probe be largely based on single pass, only rely on the rise of fluorescence intensity.And this hair
The bright probe PNcy3cy5 can detect peroxynitrite (OONO by binary channels mode-), realize the detection of Ratio-type.
Claims (10)
1. a kind of compound with higher transferring efficiency of fluorescence resonance energy, it is characterised in that the compound is shown in formula Z
Compound or its pharmaceutically acceptable salt：
In formula Z, R1~R16It is respectively and independently selected from：Hydrogen, halogen, amino, C1~C6Straight or branched alkyl, C1~C6Straight or branched
It is a kind of in alkoxy, carboxyl or sulfonic group；
RaAnd RbIt is respectively and independently selected from：C1~C6Straight or branched alkyl or the C substituted by carboxyl or/and sulfonic group1~C6Straight chain
It is or a kind of in branched alkyl；
X is NH, O, S, methylene or by C1~C3The methylene of straight or branched alkyl substitution；
M is 1~5 integer；N is 1~5 integer；Y is the heterocyclic radical containing at least two nitrogen of divalence；P is anion acid group.
2. compound as claimed in claim 1, it is characterised in that wherein RaAnd RbIt is respectively and independently selected from：C1~C3Straight chain or branch
It is a kind of in alkyl group.
3. compound as claimed in claim 2, it is characterised in that wherein RaAnd RbIt is respectively and independently selected from：One in methyl or ethyl
Kind.
4. compound as claimed in claim 1, it is characterised in that wherein X is C (CH3)2。
5. compound as claimed in claim 1, it is characterised in that wherein Y is 5~6 circle heterocycles containing at least two N of divalence
Base.
6. compound as claimed in claim 5, it is characterised in that wherein Y is the 6 circle heterocycles bases containing at least two N of divalence.
7. compound as claimed in claim 6, it is characterised in that Y is
8. the compound as described in any one in claim 1~7, it is characterised in that wherein, R1~R16It is H.
9. the compound as described in any one in claim 1~8 is preparing the fluorescence probe of detection active oxygen and active nitrogen
In application.
10. application as claimed in claim 9, it is characterised in that wherein described active oxygen includes：From hypochlorous acid or its conjugation
Alkali, peroxynitrite or its conjugate base, hydrogen peroxide, singlet oxygen, superoxide anion, nitric oxide, t-butylperoxy and hydrogen
Oxygen radical；The active nitrogen includes：Peroxynitrite or its conjugate base and nitric oxide.
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| CN108743974A (en) * | 2018-07-07 | 2018-11-06 | 苏州影睿光学科技有限公司 | A kind of live body nano-biosensing system and the preparation method and application thereof |
| CN109651835A (en) * | 2018-12-26 | 2019-04-19 | 云南师范大学 | A kind of IR780 keto-acid Molecule of Cyanine Dyes and preparation method thereof and crystal structure |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108743974A (en) * | 2018-07-07 | 2018-11-06 | 苏州影睿光学科技有限公司 | A kind of live body nano-biosensing system and the preparation method and application thereof |
| CN109651835A (en) * | 2018-12-26 | 2019-04-19 | 云南师范大学 | A kind of IR780 keto-acid Molecule of Cyanine Dyes and preparation method thereof and crystal structure |
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