CN115490742A - Method for synthesizing On-DNA aromatic tertiary amine - Google Patents

Method for synthesizing On-DNA aromatic tertiary amine Download PDF

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CN115490742A
CN115490742A CN202110672177.6A CN202110672177A CN115490742A CN 115490742 A CN115490742 A CN 115490742A CN 202110672177 A CN202110672177 A CN 202110672177A CN 115490742 A CN115490742 A CN 115490742A
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李进
杜甜
伍荣峰
高森
刘观赛
万金桥
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Hitgen Inc
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Abstract

The invention relates to a method for synthesizing On-DNA aromatic tertiary amine, which takes On-DNA alpha, beta-unsaturated carbonyl compounds and aryl tertiary amine compounds as raw materials, obtains the On-DNA aromatic tertiary amine through photocatalytic reaction in alkaline environment, can be carried out in a mixed aqueous phase of an organic solvent/aqueous phase, has simple operation and environmental protection, and is suitable for synthesizing a DNA coding compound library by using a porous plate.

Description

Method for synthesizing On-DNA aromatic tertiary amine
Technical Field
The invention belongs to the technical field of coding compound libraries, and particularly relates to a method for realizing carbon-carbon bond construction by visible light catalysis of alpha-hydrocarbon activation of aromatic tertiary amine and reaction of On-DNA olefin in construction of a DNA coding compound library.
Background
In drug development, especially new drug development, high-throughput screening for biological targets is one of the main means for rapidly obtaining lead compounds. However, traditional high throughput screening based on single molecules requires long time, large equipment investment, limited number of library compounds (millions), and the building of compound libraries requires decades of accumulation, limiting the efficiency and possibility of discovery of lead compounds. The recent emergence of DNA-encoded compound library technologies (WO 2005058479, WO2018166532, CN 103882532) combined with combinatorial chemistry and molecular biology technologies, adding a DNA tag to each compound on the molecular level, and being able to synthesize up to one hundred million levels of compound libraries in a very short time, is a trend toward the next generation of compound library screening technologies, and has begun to be widely applied in the pharmaceutical industry, producing many positive effects (Accounts of Chemical Research,2014,47,1247-1255).
The DNA coding compound library can rapidly generate a giant compound library through combinatorial chemistry, and can screen out a lead compound with high flux, so that the screening of the lead compound becomes unprecedented rapidness and high efficiency. One of the challenges in constructing libraries of DNA-encoding compounds is the need to synthesize chemically diverse small molecules on DNA in high yields. Because DNA can be kept stable under certain conditions (solvent, pH, temperature and ion concentration), the On-DNA reaction applied to the construction of the DNA coding compound library also needs higher yield. Therefore, the reagent type, reaction type and reaction condition of the chemical reaction (On-DNA reaction for short) carried out On DNA directly influence the richness and selectivity of the DNA coding compound library. Therefore, the development of chemical reactions compatible with DNA is also a long-term research and research direction of the current DNA coding compound library technology, and the application and commercial value of the DNA coding compound library are directly influenced.
The aromatic tertiary amine is an important medicine compound skeleton structure, however, no method for synthesizing the On-DNA aromatic tertiary amine by the On-DNA alkenyl compound is reported at present. In addition, the small molecule building blocks commonly used in DEL technology all have obvious functional groups available for attachment, for example: carboxyl, aldehyde (ketone), primary (secondary) amine, boric acid (ester), halogen and other functional groups. The tertiary amine structure small molecule module cannot be directly used in the past DNA compatible reaction, and can be merged into the DNA coding compound library only by the guidance of other functional groups (such as carboxylic acid, aldehyde, primary (secondary) amine and the like), thereby limiting the chemical space and topological structure of the tertiary amine structure in the DNA coding compound library. Therefore, a new synthesis method of On-DNA aromatic tertiary amine suitable for large-scale multi-plate operation is developed, and a tertiary amine structure is introduced through direct functionalization so as to increase the diversity of a DNA coding compound library and further improve the application value of the DNA coding compound library technology.
Disclosure of Invention
In order to solve the problems, a synthesis method of a DNA coding compound library is developed, wherein the synthesis method has the advantages of stable storage of raw materials, mild reaction conditions, good substrate universality and small damage to DNA and is suitable for batch operation by using a porous plate, and the On-DNA alpha, beta-unsaturated carbonyl compound library can be quickly converted into the On-DNA aromatic tertiary amine through one-step reaction.
The technical scheme of the invention is as follows:
a method for synthesizing On-DNA aromatic tertiary amine uses On-DNA alpha, beta-unsaturated carbonyl compound and aryl tertiary amine compound as raw materials, and On-DNA aromatic tertiary amine is obtained by photocatalysis reaction in alkaline environment; wherein the structural formula of the On-DNA alpha, beta-unsaturated carbonyl compound is shown in the specification
Figure BDA0003118525830000021
Figure BDA0003118525830000022
The structural formula of the aryl tertiary amine compound is
Figure BDA0003118525830000023
Wherein the DNA in the structural formula comprises a single-stranded or double-stranded nucleotide chain obtained by polymerizing artificially modified and/or unmodified nucleotide monomers, and the nucleotide chain is connected with a nitrogen atom through one or more chemical bonds or groups;
the size of the DNA is 10 to 200 bases.
Wherein, the DNA in the structural formula is connected with the nitrogen atom through one chemical bond or a plurality of chemical bonds. One chemical bond means that the DNA in the structural formula is directly connected with a nitrogen atom; when multiple chemical bonds are present, they refer to the DNA and nitrogen atoms in the structural formulaAre connected with each other by a plurality of chemical bonds, e.g., a methylene group (-CH) between the DNA and the nitrogen atom 2 -) are linked, i.e. linked by two chemical bonds; or the DNA and the nitrogen atom are connected with the amino of the DNA through a carbonyl (-CO-) and also through two chemical bonds; or DNA and nitrogen atom through a methylene carbonyl group (-CH) 2 CO-) is attached to the amino group of the DNA, again by three consecutive chemical bonds.
R 1 、R 2 、R 6 Selected from hydrogen or a group having a molecular weight of 1000 or less directly bonded to a carbon atom; or R 1 、R 2 And the attached carbon atom to form a ring; or R 1 、R 6 And the attached carbon atom to form a ring;
R 3 、R 4 selected from groups having a molecular weight of 1000 or less which can be directly bonded to a nitrogen atom;
or R 3 、R 4 And the nitrogen atom to which they are attached form a heterocyclic ring;
R 5 selected from hydrogen or a group having a molecular weight of 1000 or less directly bonded to a nitrogen atom; or R 5 、R 2 And the attached atoms form a ring;
ar is selected from aryl or heteroaryl.
Preferably, ar is selected from the following groups:
Figure BDA0003118525830000031
x is O, S, NH or any one of alkyl substituted imino, and the number of the substituent groups for substituting Ar is one or more; the substituent of the substituted Ar is one or more of hydrogen, halogen, carboxyl, nitryl, alkoxy and aldehyde group which are mutually independent.
Preferably, R is 1 、R 2 、R 3 、R 4 、R 5 、R 6 Are respectively selected from alkyl, substituted alkyl, alkoxy, 5-10 membered aryl, substituted 5-10 membered aryl, 5-10 membered aromatic heterocyclic group and substituted 5-10 membered aromatic heterocyclic group; wherein the alkyl is C 1 ～C 20 An alkyl group;
the number of substituents of the substituted alkyl group is one or more; the substituent of the substituted alkyl is one or more independently selected from halogen, carboxyl, nitro, alkoxy, halogenated phenyl, phenyl and alkyl phenyl;
the number of the substituent for substituting the 5-to 10-membered aryl is one or more, and the substituents for substituting the 5-to 10-membered aryl are independently selected from halogen, cyano, nitro, carboxyl, alkoxy and C 1 ～C 20 One or more of alkyl and trifluoromethyl;
the number of the substituent(s) for substituting the 5-to 10-membered aromatic heterocyclic group is one or more, and the substituent(s) for substituting the 5-to 10-membered aromatic heterocyclic group are independently selected from the group consisting of halogen, cyano, nitro, carboxyl, alkoxy, C 1 ～C 20 One or more of alkyl and trifluoromethyl.
Preferably, the method comprises the following steps: said R 1 、R 2 And the attached carbon atoms form a 3-10 membered unsaturated carbocyclic ring, a 3-10 membered unsaturated heterocyclic ring.
Preferably, the method comprises the following steps: said R 3 、R 4 And the attached nitrogen atom form a 3-10 membered saturated heterocyclic ring; the number of the substituent groups of the 3-10 membered saturated heterocyclic ring is one or more, and the substituent groups for substituting the 3-10 membered saturated heterocyclic ring are one or more independently selected from hydrogen, halogen, carboxyl, nitro and alkoxy.
Preferably, the method comprises the following steps: r is 5 、R 2 And the attached atoms form a 3-10 membered unsaturated heterocyclic ring.
Preferably, the method comprises the following steps: said R 1 、R 6 And the carbon atoms connected form a 3-10 membered carbocyclic ring and a 3-10 membered heterocyclic ring.
More specifically: the On-DNA alpha, beta-unsaturated carbonyl compound is selected from:
Figure BDA0003118525830000032
Figure BDA0003118525830000033
Figure BDA0003118525830000041
preferably, the method comprises the following steps: the tertiary aryl amine compound is selected from:
Figure BDA0003118525830000042
Figure BDA0003118525830000043
preferably, the method for synthesizing the On-DNA aromatic tertiary amine comprises the following steps: adding 10-1000 times molar equivalent of aryl tertiary amine compound and 10-1000 times molar equivalent of alkali into an On-DNA alpha, beta-unsaturated carbonyl compound solution with molar equivalent of 1 and molar concentration of 0.5-5mM, finally adding 1-10 times molar equivalent of catalyst, and carrying out light reaction at 10-100 ℃ for 0.5-16 hours until the reaction is finished.
Further, the base is selected from sodium borate, lithium hydroxide, sodium hydroxide, potassium hydroxide, cesium hydroxide, sodium carbonate, potassium carbonate, cesium carbonate, sodium phosphate, potassium phosphate, sodium hydrogen phosphate, potassium hydrogen phosphate, dipotassium hydrogen phosphate, N-methylmorpholine, triethylamine, diisopropylethylamine, DBU (1,8-diazabicycloundec-7-ene), 4-dimethylaminopyridine, 2,6-dimethylpyridine, N-methylimidazole or hydrazine hydrate; preferably, the base is dipotassium hydrogen phosphate.
Further, the reaction is carried out in a solvent, wherein the solvent is a water-containing mixed solvent of any one or more of water, methanol, ethanol, acetonitrile, dimethyl sulfoxide, N-dimethylformamide, N-dimethylacetamide, an inorganic salt buffer solution, an organic acid buffer solution and an organic base buffer solution; preferably, the reaction solvent contains water and dimethyl sulfoxide.
Further, the reaction solvent has a pH of 5 to 11; preferably, the pH is 9.
Further, the catalyst for the reaction is selected from Ir [ dF (CF) 3 )ppy] 2 (dtbbpy)PF 6 、([Ir(dtbbpy)(ppy) 2 ][PF 6 ])、Ir[p-F(Me)ppy] 2 (dtbbpy)PF 6 、Ir(ppy) 2 (bpy)PF 6 Or 4-CzIPN; preferably, the reaction catalyst is Ir [ dF (CF) 3 )ppy] 2 (dtbbpy)PF 6 。
Further, the illumination power of the reaction is 0 volt, 7.5 volts or 13.8 volts; preferably, the reflected light power is 13.8 volts.
Further, the reaction time of the reaction is 0.5 to 16 hours; preferably, the reaction time is 2 hours.
Furthermore, in the method, the equivalent of the On-DNA alpha, beta-unsaturated carbonyl compound is 1, the molar equivalent of the aryl tertiary amine compound is 10-1000, the molar equivalent of the alkali is 10-1000, and the molar equivalent of the catalyst is 1-10; preferably, the molar equivalent of the aryl tertiary amine compound is 50, 100, 200, 300, 400, 500, 600, 800, 1000, the molar equivalent of the base is 50, 100, 120, 200, 300, 400, 500, 600, 800, 1000, the molar equivalent of the catalyst is 1, 5, 10; most preferably, the molar equivalent of the aryl tertiary amine compound is 100, the molar equivalent of the base is 120, and the molar equivalent of the catalyst is 1.
Further, the above method is used for batch multi-well plate operations.
Further, the above method is used for the synthesis of libraries of DNA encoding compounds for multiwell plates.
The method can realize the On-DNA aromatic tertiary amine compound obtained by the On-DNA alkenyl compound in the DNA coding compound library, can be widely applied to various On-DNA alkenyl substrates, and can introduce various substituted aryl tertiary amine compounds as synthesis modules in a large scale. The method has high yield and single product, can be carried out in the mixed aqueous phase of an organic solvent/aqueous phase, has simple operation and environmental protection, and is suitable for synthesizing the DNA coding compound library by using a porous plate.
Definitions of terms used in connection with the present invention: the initial definitions provided herein for a group or term apply to that group or term throughout the specification unless otherwise indicated; for terms not specifically defined herein, the meanings that would be given to them by a person skilled in the art are to be given in light of the disclosure and the context.
"substituted" means that a hydrogen atom in a molecule is replaced by a different atom or molecule.
The minimum and maximum values of the carbon atom content in the hydrocarbon group are indicated by a prefix, e.g. prefix (Ca-C) b ) Alkyl means any alkyl group containing from "a" to "b" carbon atoms. Thus, for example, C 1 ～C 12 Alkyl refers to a straight or branched chain alkyl group containing 1 to 12 carbon atoms.
Alkyl means a straight or branched hydrocarbon radical in an alkane molecule, e.g. methyl-CH 3 ethyl-CH 2 CH 3 methylene-CH 2 -; the alkyl group may also be part of another group, such as C 1 ～C 6 Alkoxy radical, C 1 ～C 6 An alkylamino group.
Carbocyclic ring: refers to saturated or partially saturated rings having multiple carbon atoms and no ring heteroatoms, and having a single ring or multiple rings (including fused, bridged, and spiro ring systems).
Cycloalkyl groups: refers to saturated or partially saturated cyclic groups having multiple carbon atoms and no ring heteroatoms, and having a single ring or multiple rings (including fused, bridged, and spiro ring systems).
Heterocyclic ring: is a saturated or unsaturated monocyclic or polycyclic ring carrying at least one atom of 3 to 10 atoms selected from O, S, N.
Heterocycloalkyl group: is a saturated or unsaturated monocyclic or polycyclic hydrocarbon group carrying at least one atom of 3 to 10 selected from O, S, N.
The halogen is fluorine, chlorine, bromine or iodine.
Alkoxy groups: means that the alkyl radical is bound to an oxygen atom to form a substituent, e.g. methoxy is-OCH 3 。
Halogenated phenyl group: refers to a group formed by substituting H on a phenyl group with a halogen.
Alkyl phenyl: refers to a group formed by substituting H on a phenyl group with an alkyl group.
Aryl is an aromatic monocyclic or multicyclic radical composed of C atoms and containing no heteroatoms.
The term "arylheterocyclyl" refers to a single cyclic group or a plurality of cyclic groups having aromaticity composed of 5 to 10 atoms such as C, O, S, N.
It will be apparent that various other modifications, substitutions and alterations can be made in the present invention without departing from the basic technical concept of the invention as described above, according to the common technical knowledge and common practice in the field.
The present invention will be described in further detail with reference to the following examples. This should not be understood as limiting the scope of the above-described subject matter of the present invention to the following examples. All the technologies realized based on the above contents of the present invention belong to the scope of the present invention.
Drawings
FIGS. 1 to 2: 42 On-DNA aromatic tertiary amine compounds corresponding conversion rate distribution maps are obtained in the embodiment 1 of the invention.
Detailed Description
The present invention will be described in further detail with reference to the following examples, but it should not be construed that the scope of the above subject matter is limited to the following examples. All the technologies realized based on the above contents of the present invention belong to the scope of the present invention.
The raw materials and equipment used in the invention are known products and are obtained by purchasing commercial products.
DNA-NH in the present invention 2 Is formed by single-stranded or double-stranded DNA and a linker group and has-NH 2 DNA Structure of linker, e.g. DNA-NH of "compound1" in WO2005058479 2 And (5) structure. Also for example the following DNA structure:
Figure BDA0003118525830000061
wherein A is adenine, T is thymine, C is cytosine, and G is guanine.
DIPEA: n, N-diisopropylethylamine; DIC: n, N' -diisopropylcarbodiimide; HATU:2- (7-azabenzotriazole) -N, N' -tetramethyluronium hexafluorophosphate; DMA: dimethylacetamide; DMSO (dimethylsulfoxide): dimethyl sulfoxide; DDTC: sodium diethyldithiocarbamate.
Example 1A method for synthesizing On-DNA aromatic tertiary amine
Step 1, synthesis of On-DNA alkenyl compound
Figure BDA0003118525830000071
(1) Dissolving 1 in 250mM boric acid buffer solution with pH =9.4 to prepare 1mM concentration solution, mixing 50 times equivalent amount of HATU (concentration of 0.4M dissolved in DMA), 50 times equivalent amount of carboxylic acid reagent (concentration of 0.4M dissolved in DMA), and 50 times equivalent amount of DIPEA (concentration of 0.4M dissolved in DMA) at 0 deg.C in sequence, mixing the mixture thoroughly by vortex oscillation, storing at 0 deg.C for 5 minutes, adding the mixture into 1 solution, mixing well, and reacting at room temperature for 0.5-1 hour.
And (3) after the reaction is finished, carrying out ethanol precipitation: adding a 5M sodium chloride solution with the total volume of 10% into the solution after the reaction, then continuously adding absolute ethyl alcohol with the total volume of 3 times of the total volume, after uniformly oscillating, placing the reaction in dry ice for freezing for 0.5 hour, then centrifuging for half an hour at the rotating speed of 12000rpm, pouring out the supernatant, dissolving the rest precipitate with deionized water to obtain the solutions of the compounds 2, 4, 7 and 9, and after quantifying by an enzyme labeling instrument OD, sending LCMS to confirm that the reaction conversion rate is 60% -90%.
(2) 1 was dissolved in 250mM boric acid buffer solution having a pH of =9.4 to prepare a 1mM solution, and then 50 equivalents of N-hydroxysuccinimide (NHS) (0.4M in DMA), 40 equivalents of 2-bromoacrylic acid (0.2M in DMA), and 20 equivalents of N, N' -Diisopropylcarbodiimide (DIC) (0.4M in DMA) were mixed in this order at 0 ℃, and the mixture was mixed well by vortexing and then stored at 0 ℃ for 5 minutes. Dividing the mixture into two parts, adding one part into the solution 1, mixing uniformly, reacting for 5 minutes at 0 ℃, adding the rest mixture into the solution 1, mixing uniformly, reacting for 5 minutes at 0 ℃, and finally placing the reaction solution at room temperature for reacting for 5 minutes.
And (3) after the reaction is finished, carrying out ethanol precipitation: adding a 5M sodium chloride solution accounting for 10% of the total volume of the solution after reaction, then continuously adding absolute ethyl alcohol accounting for 3 times of the total volume, uniformly oscillating, placing the reaction in dry ice for freezing for 0.5 hour, then centrifuging for half an hour at the rotating speed of 12000rpm, pouring out the supernatant, dissolving the rest precipitate with deionized water to obtain a solution of a compound 3, quantifying by an enzyme labeling instrument OD, and sending to LCMS to confirm that the reaction conversion rate is 56%.
(3) Compound 3 was dissolved in water to prepare a 1mM concentration solution, followed by sequentially adding 100 equivalents of an arylboronic acid compound (concentration 0.2M in DMSO), 100 equivalents of cesium hydroxide (concentration 0.5M in water), and 2.5 equivalents of chlorine (sodium-2-dicyclohexylphosphine-2 ',6' -dimethoxy-1,1 '-biphenyl-3' -sulfonate) [2- (2 '-amino-1,1' -biphenyl) ] palladium (II) (sSPhos Pd G2) (concentration 0.01M in DMSO), mixing well, and reacting at 90 ℃ for 2 hours. After the reaction, 100 times of equivalent weight of DDTC (0.4M in water) was added to the reaction system, mixed well, and reacted at 90 ℃ for 10 minutes.
After the reaction is finished, centrifuging to take the supernatant, and then performing ethanol precipitation: adding 5M sodium chloride solution with the total volume of 10% into the solution after reaction, then continuously adding absolute ethyl alcohol with the total volume of 3 times of the total volume, after uniformly oscillating, placing the reaction in dry ice for freezing for 0.5 hour, then centrifuging for half an hour at the rotating speed of 12000rpm, pouring out supernatant, dissolving the rest precipitate with deionized water to obtain the solution of the On-DNA alpha, beta-unsaturated carbonyl compound 5, and sending LCMS to confirm that the conversion rate of the reaction is 70-90% after the quantification by an enzyme labeling instrument OD.
(4) Compound 4 was dissolved in water to prepare a 1mM solution, followed by sequentially adding 10 equivalents of pinacol ester of vinylboronic acid (concentration 0.2M in DMSO), 100 equivalents of cesium hydroxide (concentration 0.5M in water), 2.5 equivalents of chlorine (sodium-2-dicyclohexylphosphine-2 ',6' -dimethoxy-1,1 '-biphenyl-3' -sulfonate) [2- (2 '-amino-1,1' -biphenyl) ] palladium (II) (sSPhos Pd G2) (concentration 0.01M in DMSO), mixing well, and reacting at 100 ℃ for 30 minutes. After the reaction, 100 times of the equivalent of sodium diethyldithiocarbamate (DDTC) (concentration of 0.4M, dissolved in water) was added to the reaction system, mixed well, and reacted at 100 ℃ for 10 minutes.
After the reaction is finished, centrifuging to take the supernatant, and then performing ethanol precipitation: adding 5M sodium chloride solution with the total volume of 10% into the solution after reaction, then continuously adding absolute ethyl alcohol with the total volume of 3 times of the total volume, after uniformly oscillating, placing the reaction in dry ice for freezing for 0.5 hour, then centrifuging for half an hour at the rotating speed of 12000rpm, pouring out supernatant, dissolving the rest precipitate with deionized water to obtain the solution of the On-DNA alpha, beta-unsaturated carbonyl compound 6, and sending LCMS to confirm that the conversion rate of the reaction is 70-90% after the quantification by an enzyme labeling instrument OD.
(5) Compound 7 was dissolved in 250mM boric acid buffer solution having pH =9.4 to prepare a 1mM concentration solution (20. Mu.L, 20 nmol), benzaldehyde (4000nmol, 200 equivalents, 200mM DMSO), and potassium hydroxide (10000nmol, 500 equivalents, 1000mM double distilled water) were sequentially added to the solution, and they were mixed well and reacted at 30 ℃ for 1 hour.
And (3) after the reaction is finished, carrying out ethanol precipitation: adding a 5M sodium chloride solution with the total volume of 10% into the solution after the reaction, then continuously adding absolute ethyl alcohol with the total volume of 3 times of the total volume, after uniformly oscillating, placing the reaction in dry ice for freezing for 0.5 hour, then centrifuging for half an hour at the rotating speed of 12000rpm, pouring out the supernatant, dissolving the rest precipitate with deionized water to obtain a solution of the On-DNA alpha, beta-unsaturated carbonyl compound 8, and sending LCMS to confirm that the conversion rate of the reaction is 90% after the quantification by an enzyme labeling instrument OD.
(6) Compound 9 was dissolved in 250mM, pH =9.4 boric acid buffer to prepare a 1mM concentration solution (20. Mu.L, 20 nmol), cyclohexanone (2000 nmol,100 equivalents, 200mM DMSO), potassium hydroxide (12000nmol, 600 equivalents, 1000mM double distilled water) were sequentially added to the solution, and the mixture was mixed well and reacted at 30 ℃ for 1 hour.
And (3) after the reaction is finished, carrying out ethanol precipitation: adding a 5M sodium chloride solution with the total volume of 10% into the solution after the reaction, then continuously adding absolute ethyl alcohol with the total volume of 3 times of the total volume, after uniformly oscillating, placing the reaction in dry ice for freezing for 0.5 hour, then centrifuging for half an hour at the rotating speed of 12000rpm, pouring out the supernatant, dissolving the rest precipitate with deionized water to obtain a solution of the On-DNA alpha, beta-unsaturated carbonyl compound 10, and sending LCMS to confirm that the conversion rate of the reaction is 72% after the quantification by an enzyme labeling instrument OD.
Step 2, visible light catalysis aromatic tertiary amine alpha-hydrocarbon activation and On-DNA olefin reaction
Dissolving an On-DNA alpha, beta-unsaturated carbonyl compound into water to prepare a 1mM concentration solution, and sequentially adding 100 times of equivalent of an aryl tertiary amine compound (with the concentration of 0.5M and dissolved in DMSO), 120 times of equivalent of dipotassium hydrogen phosphate (with the concentration of 0.3M and dissolved in water) and 1 time of equivalent of a catalyst Ir [ dF (CF) 3 )ppy] 2 (dtbbpy)PF 6 (concentration: 0.01M in DMSO), and DMSO was further added to the reaction mixture to adjust the ratio of the organic phase: water phase volume ratio =3:2, mixing evenly, removing air for 2 hours, and performing light reaction for 2 hours.
And (3) after the reaction is finished, carrying out ethanol precipitation: and adding a 5M sodium chloride solution accounting for 10 percent of the total volume of the solution after the reaction, continuously adding absolute ethyl alcohol accounting for 3 times of the total volume of the solution, uniformly oscillating, placing the reaction in dry ice for freezing for 0.5 hour, centrifuging for half an hour at the rotating speed of 12000rpm, pouring out supernatant, dissolving the rest precipitate with deionized water to obtain a solution of 42 On-DNA products, quantifying by an enzyme labeling instrument OD (optical density measurement) and sending to LCMS (liquid crystal display system) to confirm the conversion rate of the reaction.
In conclusion, the method can obtain the On-DNA aromatic tertiary amine compound by controlling the conditions of solvent, temperature, pH and the like during reaction and reacting the On-DNA alpha, beta-unsaturated carbonyl compound with the aryl tertiary amine compound in the presence of alkali through visible light catalysis. The method has wide substrate application range, can be carried out in a mixed aqueous phase of an organic solvent and an aqueous phase, is simple to operate, is environment-friendly, and is suitable for synthesizing a DNA coding compound library by using a porous plate.

Claims (11)

1. A method for synthesizing On-DNA aromatic tertiary amine is characterized in that: the method takes an On-DNA alpha, beta-unsaturated carbonyl compound and an aryl tertiary amine compound as raw materials, and obtains the On-DNA aromatic tertiary amine through a photocatalytic reaction in an alkaline environment; wherein the structural formula of the On-DNA alpha, beta-unsaturated carbonyl compound is shown in the specification
Figure FDA0003118525820000011
Figure FDA0003118525820000012
The structural formula of the aryl tertiary amine compound is
Figure FDA0003118525820000013
Wherein the DNA in the structural formula comprises a single-stranded or double-stranded nucleotide chain obtained by polymerizing artificially modified and/or unmodified nucleotide monomers, and the nucleotide chain is connected with a nitrogen atom through one or more chemical bonds or groups;
R 1 、R 2 、R 6 selected from hydrogen or a group having a molecular weight of 1000 or less directly bonded to a carbon atom; or R 1 、R 2 With the carbon atom to which it is attached, or R 1 、R 6 And the attached carbon atom form a ring;
R 3 、R 4 selected from groups having a molecular weight of 1000 or less directly bonded to the nitrogen atom;
or R 3 、R 4 And the attached nitrogen atom form a heterocyclic ring;
R 5 selected from hydrogen or a group having a molecular weight of 1000 or less directly bonded to a nitrogen atom; or R 5 、R 2 And the attached atoms form a ring;
ar is selected from aryl or heteroaryl.
2. The method of claim 1, wherein: ar is selected from the following groups:
Figure FDA0003118525820000014
x is O, S, NH or any one of alkyl substituted imino, and the number of the substituent groups for substituting Ar is one or more; the substituent of the substituted Ar is one or more of hydrogen, halogen, carboxyl, nitro, alkoxy and aldehyde group which are independent of each other.
3. The method of claim 1, wherein: said R 1 、R 2 、R 3 、R 4 、R 5 、R 6 Are respectively selected from alkyl, substituted alkyl, alkoxy, 5-10 membered aryl, substituted 5-10 membered aryl, 5-10 membered aromatic heterocyclic group and substituted 5-10 membered aromatic heterocyclic group; wherein the alkyl is C 1 ～C 20 An alkyl group;
the number of substituents of the substituted alkyl group is one or more; the substituent of the substituted alkyl is one or more independently selected from halogen, carboxyl, nitro, alkoxy, halogenated phenyl, phenyl and alkyl phenyl;
the number of the substituent for substituting the 5-to 10-membered aryl is one or more, and the substituents for substituting the 5-to 10-membered aryl are independently selected from halogen, cyano, nitro, carboxyl, alkoxy and C 1 ～C 20 One or more of alkyl and trifluoromethyl;
the number of the substituent(s) for substituting the 5-to 10-membered aromatic heterocyclic group is one or more, and the substituent(s) for substituting the 5-to 10-membered aromatic heterocyclic group are independently selected from the group consisting of halogen, cyano, nitro, carboxyl, alkoxy, C 1 ～C 20 One or more of alkyl and trifluoromethyl;
or said R 1 、R 2 And the attached carbon atoms form a 3-10 membered unsaturated carbocyclic ring, a 3-10 membered unsaturated heterocyclic ring;
or said R 3 、R 4 And the nitrogen atom to which they are attached form a 3-to 10-membered saturated heterocyclic ring; the number of the substituent groups of the 3-10 membered saturated heterocyclic ring is one or more, and the substituent groups for substituting the 3-10 membered saturated heterocyclic ring are one or more independently selected from hydrogen, halogen, carboxyl, nitro and alkoxy;
or said R 5 、R 2 And the attached atoms form a 3-10 membered unsaturated heterocyclic ring;
or said R 1 、R 6 And the carbon atoms connected form a 3-10 membered carbocyclic ring and a 3-10 membered heterocyclic ring.
4. The method of claim 1, wherein: adding 10-1000 times molar equivalent of aryl tertiary amine compound and 10-1000 times molar equivalent of alkali into an On-DNA alpha, beta-unsaturated carbonyl compound solution with molar equivalent of 1 and molar concentration of 0.5-5mM, finally adding 1-10 times molar equivalent of catalyst, and carrying out light reaction at 10-100 ℃ for 0.5-16 hours until the reaction is finished.
5. The method of claim 4, wherein: the base is selected from the group consisting of sodium borate, lithium hydroxide, sodium hydroxide, potassium hydroxide, cesium hydroxide, sodium carbonate, potassium carbonate, cesium carbonate, sodium phosphate, potassium phosphate, sodium hydrogen phosphate, potassium hydrogen phosphate, dipotassium hydrogen phosphate, N-methylmorpholine, triethylamine, diisopropylethylamine, 1,8-diazabicycloundecene-7-ene, 4-dimethylaminopyridine, 2,6-dimethylpyridine or N-methylimidazole or hydrazine hydrate.
6. The method of claim 4, wherein: the reaction is carried out in a solvent, and the solvent is a water-containing mixed solvent of any one or more of water, methanol, ethanol, acetonitrile, dimethyl sulfoxide, N-dimethylformamide, N-dimethylacetamide, an inorganic salt buffer solution, an organic acid buffer solution and an organic base buffer solution.
7. The method of claim 4, wherein: the catalyst is selected from Ir [ dF (CF) 3 )ppy] 2 (dtbbpy)PF 6 、
([Ir(dtbbpy)(ppy) 2 ][PF 6 ])、Ir[p-F(Me)ppy] 2 (dtbbpy)PF 6 、Ir(ppy) 2 (bpy)PF 6 Or 4-CzIPN.
8. The method of claim 4, wherein: the illumination power of the reaction is 0 volt, 7.5 volts or 13.8 volts.
9. The method of claim 4, wherein: in the method, the equivalent of the On-DNA alpha, beta-unsaturated carbonyl compound is 1, the molar equivalent of the aryl tertiary amine compound is 50 equivalents, 100 equivalents, 200 equivalents, 300 equivalents, 400 equivalents, 500 equivalents, 600 equivalents, 800 equivalents and 1000 equivalents, the equivalent of the base is 50 equivalents, 100 equivalents, 120 equivalents, 200 equivalents, 300 equivalents, 400 equivalents, 500 equivalents, 600 equivalents, 800 equivalents and 1000 equivalents, and the molar equivalent of the catalyst is 1 equivalent, 5 equivalents and 10 equivalents.
10. The method according to any one of claims 1 to 9, wherein the method is used for multi-well plate operations in batches.
11. The method of any one of claims 1 to 9, wherein the method is used for the synthesis of a library of DNA-encoding compounds for multi-well plates.
CN202110672177.6A 2021-06-18 2021-06-18 Method for synthesizing On-DNA aromatic tertiary amine Pending CN115490742A (en)

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