CN105968016A - Carbonic ester type photosensitive reagent and preparation method thereof as well as preparation method of 5'-photolabile dN - Google Patents
Carbonic ester type photosensitive reagent and preparation method thereof as well as preparation method of 5'-photolabile dN Download PDFInfo
- Publication number
- CN105968016A CN105968016A CN201610325613.1A CN201610325613A CN105968016A CN 105968016 A CN105968016 A CN 105968016A CN 201610325613 A CN201610325613 A CN 201610325613A CN 105968016 A CN105968016 A CN 105968016A
- Authority
- CN
- China
- Prior art keywords
- preparation
- nitrophenyl
- ortho
- nucleoside
- ethyl acetate
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C201/00—Preparation of esters of nitric or nitrous acid or of compounds containing nitro or nitroso groups bound to a carbon skeleton
- C07C201/06—Preparation of nitro compounds
- C07C201/12—Preparation of nitro compounds by reactions not involving the formation of nitro groups
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
- C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
- C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
- C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
- C07D213/62—Oxygen or sulfur atoms
- C07D213/63—One oxygen atom
- C07D213/64—One oxygen atom attached in position 2 or 6
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D233/00—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, not condensed with other rings
- C07D233/54—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, not condensed with other rings having two double bonds between ring members or between ring members and non-ring members
- C07D233/56—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, not condensed with other rings having two double bonds between ring members or between ring members and non-ring members with only hydrogen atoms or radicals containing only hydrogen and carbon atoms, attached to ring carbon atoms
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H19/00—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
- C07H19/02—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof sharing nitrogen
- C07H19/04—Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
- C07H19/06—Pyrimidine radicals
- C07H19/073—Pyrimidine radicals with 2-deoxyribosyl as the saccharide radical
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H19/00—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
- C07H19/02—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof sharing nitrogen
- C07H19/04—Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
- C07H19/16—Purine radicals
- C07H19/173—Purine radicals with 2-deoxyribosyl as the saccharide radical
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Saccharide Compounds (AREA)
Abstract
The invention relates to a carbonic ester type photosensitive reagent and a preparation method thereof as well as a preparation method of 5'-photolabile dN. The structural formula of the carbonic ester type photosensitive reagent is shown in the specification, wherein X is -OX', imidazolyl or substituted imidazolyl. The preparation method of 5'-photolabile dN comprises the following steps: preparing a crude product of 5'-photolabile dN from 2'-deoxynucleoside and 2-o-nitrobenzene-1-propyl carbonic ester type photosensitive reagent and triethylamine; and purifying the crude product to obtain the 5'-photolabile dN. According to the preparation method of 5'-photolabile dN, the selectivity is improved by 3.4 times on average, the purifying process is simplified, the yield is increased, the production cost is greatly reduced, and the use of virulent diphosgene is avoided.
Description
Technical field
The invention belongs to technical field of organic synthesis, be specifically related to a kind of carbonates photosensitive reagents and preparation method thereof, the preparation method of 5'-protection against light sensitivity nucleoside.
Background technology
Biochip technology is a kind of High biotechnology utilizing making nucleic acid molecular hybridization principle to combine with microelectric technique and being formed.Can be widely applied to medical diagnosis on disease, drug screening, bioscience research, many fields such as the preferred good child-rearing of crops, judicial expertise, Food Hygiene Surveillance, environment measuring.Owing to original position photoetching technique can prepare high density oligonucleotide gene chip; at present; gene chip based on original position photoetching technique is widely adopted, and 5 '-protection against light sensitivity nucleoside and derivant 5 '-photosensitive phosphoramidite monomer thereof are one of critical materialses preparing such gene chip.But, in the method (as follows) of existing preparation 5 '-protection against light sensitivity nucleoside,
nullThere are two common difficult problems: (1)、When introducing 5 '-photosensitive group，Owing to using acyl chloride photosensitive reagents，The selectivity that reaction lack is enough，React the most on a large scale，The more difficult control of selective response condition of acyl chloride photosensitive reagents，While generating 5 '-protection against light sensitivity nucleoside (5 '-Photolabile dN)，There is considerable amount of by-product 3 '-protection against light sensitivity nucleoside (3 '-Photolabile dN) and 3 ',5 '-bis-protection against light sensitivity nucleoside (3 ',5 '-diPhotolabile dN) generate,Different according to base，The principal product 5 '-protection against light sensitivity nucleoside generated and by-product (3 '-protection against light sensitivity nucleoside and 3 ',5 '-bis-protection against light sensitivity nucleoside etc.) ratio be generally 5:1 to 2:1 (list of references: Sigrid Buhler,Irene Lagoja etc.，Helvetica Chimica Acta,2004,v87(3),620-659).Owing to 3 '-protection against light sensitivity nucleoside and 5 '-protection against light sensitivity nucleoside separating property are the most close, make separation 5 '-protection against light sensitivity nucleoside and 3 '-protection against light sensitivity nucleoside extremely difficult.Even if separating, owing to both easily intersect, causing the loss of 5 '-protection against light sensitivity nucleoside product relatively big, productivity is on the low side.Therefore, find and the primary (1 °) alcohol and secondary (2 °) alcohol are had the reagent of high selectivity, improve the extremely important (list of references: Abbas-Alli G.Shaikh of selectivity of reaction, Chem Review, 1996, v96,951-976)；(2) surpalite (diphosgene) (list of references: Woffgang Pfleiderer&Heiner Giegrich, must be used when preparing acyl chloride photosensitive reagents, 1998, United States Patent (USP), US5763599), surpalite toxicity is the biggest, operate very inconvenient, and serious environment pollution, should avoid as far as possible aborning using.
Summary of the invention
For the deficiencies in the prior art; the present invention provides a kind of carbonates photosensitive reagents and preparation method thereof, the preparation method of 5'-protection against light sensitivity nucleoside; the present invention synthesizes a kind of new carbonates photosensitive reagents; its with the primary (1 °) alcohol and secondary (2 °) alcohol are had highly reactive selectivity; photosensitive group is selectively introducing 5 '-position of nucleoside so that the productivity of 5'-protection against light sensitivity nucleoside is greatly improved.
The technical solution used in the present invention is:
A kind of carbonates photosensitive reagents, its structural formula is:
Wherein X be-OX ', imidazole radicals, substituted imidazole base, described substituted imidazole base is 2-methylimidazole base, 1,2,4-triazol radicals；In-OX ', X ' is p-nitrophenyl (4-nitrophenyl), pentafluorophenyl group (pentafluorophenyl), 2-pyridine radicals (2-pyridyl), and succinimido (succinimidyl).
A kind of preparation method of carbonates photosensitive reagents, step includes:
A, 2-ortho-nitrophenyl-1-propanol is dissolved in organic solvent, adds carbonic diester class reagent or carbonyl diurethane azole reagent, add triethylamine, stirring reaction 6-10h under room temperature；
B, reacted solution priority water and saturated sodium-chloride water solution being extracted, retain organic solution layer, organic solution is dried with anhydrous sodium sulfate, filters, and filtrate is evaporated, and vacuum drying obtains 2-ortho-nitrophenyl-1-propyl carbonate class photosensitive reagents.
In described step A, organic solvent is selected from dichloromethane, chloroform, the one in ethyl acetate, toluene；
In described step A, 2-ortho-nitrophenyl-1-propanol, carbonic diester class reagent or carbonyl diurethane azole reagent, triethylamine concentration in organic solvent are respectively 0.40-0.65mol/L, 0.50-0.75mol/L, 0.40-0.65mol/L.
Described step A carbonic diester class reagent one in double (p-nitrophenyl) carbonic ester, double (phenyl-pentafluoride) carbonic ester, carbonic acid two-2-pyridine ester, N, N '-succinimidyl carbonates；Carbonyl diurethane azole reagent is selected from N'N-carbonyl dimidazoles, N'N-carbonyl diurethane (2-methylimidazole), the one in N'N-carbonyl diurethane (1,2,4-triazole)；
Described step A ambient temperature is 10-30 DEG C.
A kind of preparation method of 5'-protection against light sensitivity nucleoside, step includes:
A, dry 2 '-deoxynucleoside is dissolved in the mixed solvent of anhydrous pyridine and dichloromethane; it is cooled to-3 DEG C-3 DEG C; it is sequentially added into 2-ortho-nitrophenyl-1-propyl carbonate class photosensitive reagents and triethylamine; it is stirred at room temperature reaction 14-22h; decompression boils off reaction dissolvent; obtain 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-2 '-deoxynucleoside crude product, i.e. 5'-protection against light sensitivity nucleoside crude product；
B, the mixed liquor of 5'-protection against light sensitivity nucleoside crude product ethyl acetate and sodium bicarbonate aqueous solution is extracted; retain ethyl acetate layer; ethyl acetate layer priority water and saturated sodium-chloride water solution washing; then ethyl acetate layer anhydrous sodium sulfate is dried; filter; filtrate is evaporated; product silica gel column chromatography after being evaporated purifies; with methylene chloride/methanol as eluting solvent; collect product component, be concentrated to give 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-2 '-deoxynucleoside i.e. 5'-protection against light sensitivity nucleoside.
In described step a, 2 '-deoxynucleoside is selected from 2 '-deoxyribosylthymine, 2 '-BrdU, N-acetyl group-2 '-deoxycytidine, N-benzoyl-2'-deoxyadenosine, one in N-isobutyryl-2'-deoxyguanosine；
In described step a, 2 '-deoxynucleoside, 2-ortho-nitrophenyl-1-propyl carbonate class photosensitive reagents and triethylamine concentration in the mixed solvent of anhydrous pyridine and dichloromethane are respectively 0.25-0.35mol/L, 0.29-0.39mol/L, 0.26-0.36mol/L.
In described step a, anhydrous pyridine is 4-6:1 with the volume ratio of dichloromethane；
In described step a, ambient temperature is 10-30 DEG C；
In described step b, ethyl acetate and sodium bicarbonate aqueous solution volume ratio are 1-2:1；Sodium bicarbonate aqueous solution mass percentage concentration is 3-5%；
In described step b, in eluting solvent, dichloromethane and methanol volume ratio are 20-50:1.
The reaction principle of the preparation method of 5'-protection against light sensitivity nucleoside is as follows:
In formula, BRFor thymus pyrimidine (Thymine is called for short T), N4-acetylcytosine (N4-Acetyl Cytosine, is called for short Ac-C), N6-benzoyl adenine (N6-Benzoyl-Adenine, is called for short Bz-A), and N2-isobutyryl guanine (N2-ibu-Guanine, is called for short ibu-G) etc..Carbon atom owing to being connected with ortho-nitrophenyl in 5 '-(2-ortho-nitrophenyl-1-propyl carbonate) has chirality, and gained 5'-protection against light sensitivity nucleoside product should be the mixture that ratio is about two diastereomers of 1:1, to product1During H HMR and HPLC analyzes, the existence of two diastereomers can be clearly observed.
nullThe present invention uses has highly reactive selective carbonates photosensitive reagents to the primary (1 °) alcohol and secondary (2 °) alcohol，Primary (1 °) alcohol 5 '-OH reaction in the deoxynucleoside optionally protected with deoxynucleoside or base，Selectively produce 5'-protection against light sensitivity nucleoside，Required product (5 '-protection against light sensitivity nucleoside) and by-product (3 '-protection against light sensitivity nucleoside and 3 ' will be generated,5 '-bis-protection against light sensitivity nucleoside) ratio bring up to 14:1 to 10:1 (average out to 12:1) from 5:1 to 2:1 (average out to 3.5:1),Selectivity averagely improves 3.4 times，Simplify purification process，Improve productivity，Greatly reduce production cost，Avoid the surpalite using severe toxicity simultaneously,It is widely used in the 5'-protection against light sensitivity nucleoside of biochip technology for preparation and derivant provides the new process of a kind of highly effective，There is the highest using value.
Detailed description of the invention
Embodiment 1:
5’-NPPOC-dT(5) preparation
The preparation reactions steps of 1a. photosensitive reagents:
Room temperature 25 DEG C, in reaction vessel, by 2-ortho-nitrophenyl-1-propanol1[2-(2-nitrophenyl) propan-1-ol, 54.4g, 0.30mol] is dissolved in dichloromethane (544ml), adds double (p-nitrophenyl) carbonic ester2[Bis (4-nitrophenyl) carbonate, 0.33mol], adds triethylamine (0.303mol)；Reaction system is stirred at room temperature, and after 6 hours, HPLC shows that reaction is completely, stopped reaction, (2x 300ml, represents after extracting with 300ml water reaction solution successively use water, extract once with 300ml water again, coextraction 2 times) and saturated sodium-chloride water solution (300ml) extraction, retaining organic (dichloromethane) layer, dichloromethane layer anhydrous sodium sulfate is dried, filter, filtrate is evaporated, vacuum drying, obtains p-nitrophenyl carbonic acid (2-ortho-nitrophenyl)-1-propyl ester3, 87.3g, HPLC purity is 96%, and productivity is 84%.
1b. selects coupling reaction step:
By dry 2 '-deoxyribosylthymine4(Thymidine, 0.20mol) is dissolved in anhydrous pyridine (485ml) and dichloromethane (97ml), cooling, is sequentially added into Nitrobenzol carbonic acid (2-ortho-nitrophenyl)-1-propyl ester 0 DEG C of priority3(0.224mol) with triethylamine (0.204mol), finish, reaction system is stirred room temperature 25 DEG C, after 14 hours, HPLC shows reaction completely, stopped reaction, decompression boils off reaction dissolvent, obtains 5 '-[p-nitrophenyl carbonic acid (2-ortho-nitrophenyl)-1-propyl carbonate]-2 '-deoxyribosylthymine5(5 '-NPPOC-dT) reacting coarse product.Now, HPLC shows that product (5 '-protection against light sensitivity nucleoside) is 14:1 with the ratio of by-product (3 '-protection against light sensitivity nucleoside and 3 ', 5 '-bis-protection against light sensitivity nucleoside).
1c, purification step:
Above-mentioned 5 '-NPPOC-dT reacting coarse product ethyl acetate (500ml) and 3% sodium bicarbonate aqueous solution (300ml) are extracted, retain ethyl acetate layer, ethyl acetate layer successively use water (300ml) and saturated sodium-chloride water solution (300ml) washing, then ethyl acetate layer anhydrous sodium sulfate is dried, filter, filtrate is evaporated, and obtains 5 '-NPPOC-dT crude products.5 '-NPPOC-dT crude product purified by silica gel column chromatographies purify, and with methylene chloride/methanol as eluting solvent, collect product component, are concentrated to give 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-2 '-deoxyribosylthymine5(5 '-NPPOC-dT), 73.7g, HPLC purity is 98.3%, and productivity is 82%.5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-2 '-deoxyribosylthymine5The chemical constitution of (5 '-NPPOC-dT) is passed through1H NMR and LC-MS (M+) confirm.
5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-2 '-deoxyribosylthymine5(5’-NPPOC-dT):1H NMR(CDCl3null,400MHz)δ(ppm)8.65(s,NH,Diastereomer),8.63(s,NH,Diastereomer),7.76(m,1-Ar-H,H-NPPOC),7.57(t,1-Ar-H,H-NPPOC),7.40(m,2Ar-H,H-NPPOC),7.31(s,H-6,Diastereomer),7.28(s,H-6,Diastereomer),6.33(t,H-1’,Diastereomer),6.29(t,H-1’,Diastereomer),4.31-3.87(m,6H,H-3’,H-4’,2H-5’,-the CH2 of NPPOC),3.76(m,1H,-the CH of NPPOC),2.61-2.58(dd,1H,3’-OH),2.39-2.18(m,2H,H-2’),1.85,1.74(2s,3H,5-CH3),1.38,1.35(2d,3H,-the CH3 of NPPOC)；LC-MS:m/z for(M+H)+, value of calculation C20H24N3O9,450.41.；Measured value 450.4.
Embodiment 2:
5’-NPPOC-N-Ac-dC(9) preparation
The preparation reactions steps of 2a. photosensitive reagents:
At room temperature 20 DEG C, in reaction vessel, by 2-ortho-nitrophenyl-1-propanol1[2-(2-nitrophenyl) propan-1-ol, 0.30mol] is dissolved in chloroform (544ml), adds double (phenyl-pentafluoride) carbonic ester6[Bis (pentafluorophenyl) carbonate0.33mol], adds triethylamine (0.303mol).Reaction system is stirred at room temperature, and after 9 hours, HPLC shows reaction completely, stopped reaction, and reaction solution successively use water (2x 300ml) and saturated sodium-chloride water solution (300ml) extraction retain organic (chloroform) layer；Chloroform layer is dried with anhydrous sodium sulfate, filters, is evaporated, and vacuum drying obtains phenyl-pentafluoride carbonic acid (2-ortho-nitrophenyl)-1-propyl ester7, 95.1g, HPLC purity is 97%, and productivity is 81%.
2b. selects coupling reaction step:
N-acetyl group-2 by dry '-deoxycytidine8(N-Ac-dC, 0.20mol) is dissolved in anhydrous pyridine (540ml) and dichloromethane (108ml), cooling, is sequentially added into phenyl-pentafluoride carbonic acid (2-ortho-nitrophenyl)-1-propyl ester 0 DEG C of priority7(0.224mol) with triethylamine (0.204mol); finish; reaction system is stirred room temperature 20 DEG C; after 18 hours; HPLC shows that reaction is completely; stopped reaction, decompression boils off reaction dissolvent, obtains 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-acetyl group-2 '-deoxycytidine9(5 '-NPPOC-N-Ac-dC) reacting coarse product.Now, HPLC shows that product (5 '-protection against light sensitivity nucleoside) is 12:1 with the ratio of by-product (3 '-protection against light sensitivity nucleoside and 3 ', 5 '-bis-protection against light sensitivity nucleoside).
2c. purification step:
Extract by above-mentioned 5 '-NPPOC-N-Ac-dC reacting coarse product ethyl acetate (500ml) with 3% sodium bicarbonate aqueous solution (300ml), retain ethyl acetate layer, ethyl acetate layer successively use water (300ml) and saturated sodium-chloride water solution (300ml) washing, then ethyl acetate layer anhydrous sodium sulfate is dried, filter, filtrate is evaporated, and obtains 5 '-NPPOC-N-Ac-dC crude products.5 '-NPPOC-N-Ac-dC crude product purified by silica gel column chromatographies purify, and with methylene chloride/methanol for eluting solution, collect product component, are concentrated to give 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-acetyl group-2 '-deoxycytidine9(5 '-NPPOC-N-Ac-dC), 75.3g, HPLC purity is 98.1%, and productivity is 79%.5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-acetyl group-2 '-deoxycytidine9The chemical constitution of (5 '-NPPOC-N-Ac-dC) is passed through1H NMR and LC-MS (M+) confirm.
5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-acetyl group-2 '-deoxycytidine9(5’-NPPOC-N-Ac-dC):1H NMR(CDCl3null,400MHz)δ(ppm)8.92(br,Acetyl group-NH,Diastereomer),8.89(br,Acetyl group-NH,Diastereomer),7.92(d,H-6,Diastereomer),7.89(d,H-6,Diastereomer),7.74(m, 1-Ar-H,H-NPPOC),7.55(t,1-Ar-H,H-NPPOC),7.38(m,2Ar-H,H-NPPOC),7.29(d,H-5,Diastereomer),7.26(d,H-5 diastereomer),6.24(t,H-1’,Diastereomer),6.19(t,H-1’,Diastereomer),4.32-3.89(m,6H,H-3’,H-4’,2H-5’,-the CH2 of NPPOC),3.75(m,1H,-the CH of NPPOC),2.62-2.59(dd,1H,3’-OH),2.38-2.17(m,2H,H-2’),2.14,2.11(2s,3H,CH3C=O),1.40,1.37(2d,3H,-the CH3 of NPPOC)；LC-MS:m/z for (M+H)+, value of calculation C21H25N4O9,477.44.；Measured value 477.5.
Embodiment 3:
5’-NPPOC-N-Bz-dA(13) preparation
The preparation reactions steps of 3a. photosensitive reagents:
At room temperature 25 DEG C, in reaction vessel, by 2-ortho-nitrophenyl-1-propanol1[2-(2-nitrophenyl) propan-1-ol, 0.30mol] is dissolved in ethyl acetate (540ml), adds N'N-carbonyl dimidazoles10(1,1 '-Carbonyldiimidazole, 0.33mol), adds triethylamine (0.303mol).Reaction system is stirred at room temperature, and after 7 hours, HPLC shows reaction completely, stopped reaction, and reaction solution successively use water (2x 300ml) and saturated sodium-chloride water solution (300ml) extraction retain organic (ethyl acetate) layer；Ethyl acetate layer is dried with anhydrous sodium sulfate, filters, is evaporated, and vacuum drying obtains TMSIM N imidazole carbonic acid (2-ortho-nitrophenyl)-1-propyl ester11, 69.4g, HPLC purity is 96%, and productivity is 84%.
3b. selects coupling reaction step:
By dry N-benzoyl-2'-deoxyadenosine12(N-Bz-dA, 71.1g, 0.20mol) is dissolved in anhydrous pyridine (710ml) and dichloromethane (142ml), cooling, is sequentially added into TMSIM N imidazole carbonic acid (2-ortho-nitrophenyl)-1-propyl ester 0 DEG C of priority11(0.224mol) with triethylamine (0.204mol), finish, stir reaction system, after 16 hours, HPLC room temperature 25 DEG CTableBright reaction is complete, stopped reaction, and decompression boils off reaction dissolvent, obtains 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-benzoyl-2'-deoxyadenosines13(5 '-NPPOC-N-Bz-dA) reacting coarse product.Now, HPLC shows that product (5 '-protection against light sensitivity nucleoside) is 11:1 with the ratio of by-product (3 '-protection against light sensitivity nucleoside and 3 ', 5 '-bis-protection against light sensitivity nucleoside).
3c. purification step:
Extract by above-mentioned 5 '-NPPOC-N-Bz-dA reacting coarse product ethyl acetate (600ml) with 3% sodium bicarbonate aqueous solution (400ml), retain ethyl acetate layer, ethyl acetate layer successively use water (400ml) and saturated sodium-chloride water solution (400ml) washing, then ethyl acetate layer anhydrous sodium sulfate is dried, filter, filtrate is evaporated, and obtains 5 '-NPPOC-N-Bz-dA crude products.5 '-NPPOC-N-Bz-dA crude product purified by silica gel column chromatographies purify, and with methylene chloride/methanol as eluting solvent, collect product component, are concentrated to give 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-benzoyl-2'-deoxyadenosines13(5 '-NPPOC-N-Bz-dA), 90.1g, HPLC purity is 98.2%, and productivity is 80%.5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-benzoyl-2'-deoxyadenosines13The chemical constitution of (5 '-NPPOC-N-Bz-dA) is passed through1H NMR and LC-MS (M+) confirm.
5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-benzoyl-2'-deoxyadenosines13(5’-NPPOC-N-Bz-dA): 1H NMR(CDCl3null,400MHz)δ(ppm)9.29(br,Benzoyl-N H,Diastereomer),9.25(br,Benzoyl-N H,Diastereomer),8.61(s,H-2,Diastereomer),8.58(s,H-2,Diastereomer),8.19(s,H-8,Diastereomer),8.16(s,H-8,Diastereomer),7.86(m,2H,The Ar-H of benzoyl),7.75(m,1-Ar-H,H-NPPOC),7.56(t,1-Ar-H,H-NPPOC),7.45-7.44(m,3H,The Ar-H of benzoyl),7.37(m,2Ar-H,H-NPPOC),6.35(t,H-1’,Diastereomer),6.31(t,H-1’,Diastereomer),4.31-3.88(m,6H,H-3’,H-4’,2H-5’,-the CH2 of NPPOC),3.74(m,1H,-the CH of NPPOC),2.83-2.80(dd,1H,3’-OH),2.39-2.17(m,2H,H-2’),1.41,1.38(2d,3H,-the CH3 of NPPOC)；LC-MS:m/z for (M+H)+, value of calculation C27H27N6O8,563.53.；Measured value 563.5.
Embodiment 4:
5’-NPPOC-dT(5) preparation
The preparation reactions steps of 4a. photosensitive reagents:
At room temperature 26 DEG C, in reaction vessel, by 2-ortho-nitrophenyl-1-propanol1[2-(2-nitrophenyl) propan-1-ol, 54.4g, 0.30mol] is dissolved in dichloromethane (540ml), adds carbonic acid two-2-pyridine ester14(Di-2-pyridyl carbonate, 0.33mol), adds triethylamine (0.303mol).Reaction system is stirred at room temperature, and after 10 hours, HPLC shows reaction completely, stopped reaction, and reaction solution successively use water (2x 300ml) and saturated sodium-chloride water solution (300ml) extraction retain organic (dichloromethane) layer；Dichloromethane layer is dried with anhydrous sodium sulfate, filters, is evaporated, and vacuum drying obtains (2-pyridine) carbonic acid-2-ortho-nitrophenyl-1-propyl ester15, 74.4g, HPLC purity is 97%, and productivity is 82%.
4b. selects coupling reaction step:
By dry N-isobutyryl-2'-deoxyguanosine16(N-ibu-dG, 0.20mol) is dissolved in anhydrous pyridine (670ml) and dichloromethane (134ml), cooling, is sequentially added into (2-pyridine) carbonic acid-2-ortho-nitrophenyl-1-propyl ester 0 DEG C of priority15(0.224mol) with triethylamine (0.204mol); finish; reaction system is stirred room temperature 26 DEG C; after 22 hours; HPLC shows that reaction is completely; stopped reaction, decompression boils off reaction dissolvent, obtains 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-isobutyryl-2'-deoxyguanosines17(5 '-NPPOC-N-ibu-dG) reacting coarse product.Now, HPLC shows that product (5 '-protection against light sensitivity nucleoside) is 10:1 with the ratio of by-product (3 '-protection against light sensitivity nucleoside and 3 ', 5 '-bis-protection against light sensitivity nucleoside).
4c. purification step:
Extract by above-mentioned 5 '-NPPOC-N-ibu-dG reacting coarse product ethyl acetate (600ml) with 3% sodium bicarbonate aqueous solution (400ml), retain ethyl acetate layer, ethyl acetate layer successively use water (400ml) and saturated sodium-chloride water solution (400ml) washing, then ethyl acetate layer anhydrous sodium sulfate is dried, filter, filtrate is evaporated, and obtains 5 '-NPPOC-N-ibu-dG crude products.5 '-NPPOC-N-ibu-dG crude product purified by silica gel column chromatographies purify, and with methylene chloride/methanol as eluting solvent, collect product component, are concentrated to give 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-isobutyryl-2'-deoxyguanosines17(5 '-NPPOC-N-ibu-dG), 82.8g, HPLC purity is 98.0%, and productivity is 76%.5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-isobutyryl-2'-deoxyguanosines17The chemical constitution of (5 '-NPPOC-N-ibu-dG) is passed through1H NMR and LC-MS (M+) confirm.
5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-N-isobutyryl-2'-deoxyguanosines17(5’-NPPOC-N-ibu-dG): 1H NMR(CDCl3null,400MHz)δ(ppm)10.25(s,Guanine-NH,Diastereomer),10.21(s,Guanine-NH,Diastereomer),9.28(br,Isobutyryl-NH,Diastereomer),9.24(br,Isobutyryl-NH,Diastereomer),7.97(s,H-8,Diastereomer),7.94(s,H-8,Diastereomer),7.74(m,1-Ar-H,H-NPPOC),7.57(t,1-Ar-H,H-NPPOC),7.38(m,2Ar-H,H-NPPOC),6.19(t,H-1’,Diastereomer),6.16(t,H-1’,Diastereomer),4.33-3.90(m,6H,H-3’,H-4’,2H-5’,-the CH2 of NPPOC),3.73(m,1H,-the CH of NPPOC),2.77-2.74(m,1H,-the CH of isobutyryl),2.61-2.58(dd,1H,3’-OH),2.39-2.17(m,2H,H-2’),1.42,1.39(2d,3H,-the CH3 of NPPOC),1.21,1.18(dd,6H,-the CH3 of isobutyryl)；LC-MS:m/z for (M+H)+, value of calculation C24H29N6O9, 545.51, measured value 545.5.
Claims (10)
1. a preparation method for 5'-protection against light sensitivity nucleoside, step includes:
A, dry 2 '-deoxynucleoside is dissolved in the mixed solvent of anhydrous pyridine and dichloromethane, is cooled to-3 DEG C-3 DEG C,
It is sequentially added into 2-ortho-nitrophenyl-1-propyl carbonate class photosensitive reagents and triethylamine, is stirred at room temperature reaction 14-22h, decompression
Boil off reaction dissolvent, obtain 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-2 '-deoxynucleoside crude product, i.e. 5'-protection against light sensitivity
Nucleoside crude product；
B, 5'-protection against light sensitivity nucleoside crude product ethyl acetate and sodium bicarbonate aqueous solution are extracted, retain ethyl acetate layer, second
Ethyl acetate layer priority water and saturated sodium-chloride water solution washing, then ethyl acetate layer anhydrous sodium sulfate is dried, and filters, filter
Liquid is evaporated, and the product silica gel column chromatography after being evaporated purifies, and with methylene chloride/methanol as eluting solvent, collects product component,
It is concentrated to give 5 '-(2-ortho-nitrophenyl-1-propyl carbonate)-2 '-deoxynucleoside i.e. 5'-protection against light sensitivity nucleoside.
2. preparation method as claimed in claim 1, it is characterised in that: in described step a 2 '-deoxynucleoside selected from 2 '-
Deoxyribosylthymine, 2 '-BrdU, N-acetyl group-2 '-deoxycytidine, N-benzoyl-2'-deoxyadenosine, N-isobutyryl
One in base-2'-deoxyguanosine.
3. preparation method as claimed in claim 1, it is characterised in that: 2 '-deoxynucleoside, 2-neighbour's nitre in described step a
Base benzene-1-propyl carbonate class photosensitive reagents and triethylamine concentration in the mixed solvent of anhydrous pyridine and dichloromethane are respectively
0.25-0.35mol/L、0.29-0.39mol/L、0.26-0.36mol/L。
4. preparation method as claimed in claim 1, it is characterised in that: anhydrous pyridine and the body of dichloromethane in described step a
Long-pending ratio is 4-6:1.
5. preparation method as claimed in claim 1, it is characterised in that: in described step b, ethyl acetate and sodium bicarbonate are water-soluble
Liquid volume ratio is 1-2:1；Sodium bicarbonate aqueous solution mass percentage concentration is 3-5%；Dichloromethane and methanol volume in eluting solvent
Ratio is 20-50:1.
6. preparation method as claimed in claim 1, it is characterised in that: described 2-ortho-nitrophenyl-1-propyl carbonate class is photosensitive
The structural formula of reagent is:
Wherein X be-OX ', imidazole radicals, substituted imidazole base；In-OX ', X ' is p-nitrophenyl, pentafluorophenyl group, 2-pyrrole
Piperidinyl and succinimido.
7. preparation method as claimed in claim 6, it is characterised in that:
The preparation process of described 2-ortho-nitrophenyl-1-propyl carbonate class photosensitive reagents includes:
A, 2-ortho-nitrophenyl-1-propanol is dissolved in organic solvent, adds carbonic diester class reagent or carbonyl diurethane azole reagent, then
Add triethylamine, stirring reaction 6-10h under room temperature；
B, reacted solution priority water and saturated sodium-chloride water solution are extracted, retain organic solution layer, organic solution nothing
Aqueous sodium persulfate is dried, filters, and filtrate is evaporated, and vacuum drying obtains 2-ortho-nitrophenyl-1-propyl carbonate class photosensitive reagents.
8. preparation method as claimed in claim 7, it is characterised in that: in described step A, organic solvent is selected from dichloromethane, and three
One in chloromethanes, ethyl acetate or toluene.
9. preparation method as claimed in claim 7, it is characterised in that: 2-ortho-nitrophenyl-1-propanol, carbon in described step A
Acid diesters class reagent or carbonyl diurethane azole reagent, triethylamine concentration in organic solvent be respectively 0.40-0.65mol/L,
0.50-0.75mol/L、0.40-0.65mol/L。
10. preparation method as claimed in claim 7, it is characterised in that: described step A carbonic diester class reagent is selected from double (right
Nitrobenzol) carbonic ester, double (phenyl-pentafluoride) carbonic ester, carbonic acid two-2-pyridine ester, in N, N '-succinimidyl carbonate one
Kind；Carbonyl diurethane azole reagent is selected from N'N-carbonyl dimidazoles, N'N-carbonyl diurethane (2-methylimidazole), N'N-carbonyl diurethane (1,2,4-
Triazole) in one.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610325613.1A CN105968016A (en) | 2015-02-12 | 2015-02-12 | Carbonic ester type photosensitive reagent and preparation method thereof as well as preparation method of 5'-photolabile dN |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510075776.4A CN104628574A (en) | 2015-02-12 | 2015-02-12 | Carbonic ester photosensitive reagent as well as preparation method thereof and method for preparing 5'-photosensitive protected nucleoside |
| CN201610325613.1A CN105968016A (en) | 2015-02-12 | 2015-02-12 | Carbonic ester type photosensitive reagent and preparation method thereof as well as preparation method of 5'-photolabile dN |
Related Parent Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510075776.4A Division CN104628574A (en) | 2015-02-12 | 2015-02-12 | Carbonic ester photosensitive reagent as well as preparation method thereof and method for preparing 5'-photosensitive protected nucleoside |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN105968016A true CN105968016A (en) | 2016-09-28 |
Family
ID=53207871
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610325613.1A Pending CN105968016A (en) | 2015-02-12 | 2015-02-12 | Carbonic ester type photosensitive reagent and preparation method thereof as well as preparation method of 5'-photolabile dN |
| CN201510075776.4A Pending CN104628574A (en) | 2015-02-12 | 2015-02-12 | Carbonic ester photosensitive reagent as well as preparation method thereof and method for preparing 5'-photosensitive protected nucleoside |
Family Applications After (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510075776.4A Pending CN104628574A (en) | 2015-02-12 | 2015-02-12 | Carbonic ester photosensitive reagent as well as preparation method thereof and method for preparing 5'-photosensitive protected nucleoside |
Country Status (1)
| Country | Link |
|---|---|
| CN (2) | CN105968016A (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116836089A (en) * | 2023-06-14 | 2023-10-03 | 哈工大郑州研究院 | Compound for cytosolic delivery and preparation method thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6756492B1 (en) * | 1999-04-08 | 2004-06-29 | Deutsches Krebsforschungszentrum Stiftund Des Offentlichen Rechts | Nucleoside derivatives with photo-unstable protective groups |
| CN102702010A (en) * | 2012-05-23 | 2012-10-03 | 北京大学 | Photosensitive functionalized solid-supported phase, preparation method and application thereof |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP5754685B2 (en) * | 2010-12-02 | 2015-07-29 | 国立研究開発法人農業・食品産業技術総合研究機構 | How to synthesize a build-up combinatorial library on a substrate |
-
2015
- 2015-02-12 CN CN201610325613.1A patent/CN105968016A/en active Pending
- 2015-02-12 CN CN201510075776.4A patent/CN104628574A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6756492B1 (en) * | 1999-04-08 | 2004-06-29 | Deutsches Krebsforschungszentrum Stiftund Des Offentlichen Rechts | Nucleoside derivatives with photo-unstable protective groups |
| CN102702010A (en) * | 2012-05-23 | 2012-10-03 | 北京大学 | Photosensitive functionalized solid-supported phase, preparation method and application thereof |
Non-Patent Citations (1)
| Title |
|---|
| ROCK J. MANCINID等: "Synthesis of a Photo-caged Aminooxy Alkane Thiol", 《ORGANIC & BIOMOLECULAR CHEMISTRY》 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104628574A (en) | 2015-05-20 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104334570B (en) | The method for preparing the nucleotide analog of substitution | |
| SK17432000A3 (en) | Novel nucleosides having bicyclic sugar moiety | |
| IKEHARA et al. | Studies of nucleosides and nucleotides. LXXXII. Cyclonucleosides.(39). Synthesis and Properties of 2'-halogeno-2'-deoxyadenosines | |
| CN101497639B (en) | Preparation of decitabine | |
| AU2008327061A1 (en) | Methods for preparing capecitabine and beta-anomer-rich trialkyl carbonate compound used therein | |
| DE69401136T2 (en) | Modified oligodeoxyribonucleotides, their preparation and their therapeutic use | |
| Fraser et al. | Synthesis and conformational properties of 2′‐deoxy‐2′‐methylthio‐pyrimidine and‐purine nucleosides: Potential antisense applications | |
| EP1995253B1 (en) | Method for detaching protecting group on nucleic acid | |
| JP3042073B2 (en) | Nucleoside derivative and method for producing the same | |
| CN105968016A (en) | Carbonic ester type photosensitive reagent and preparation method thereof as well as preparation method of 5'-photolabile dN | |
| DE68915067T2 (en) | Process for the preparation of 2'-deoxy-beta-adenosine. | |
| CN109053839A (en) | The novel processing step of 3 '-O-CH2N3-2 '-O-Me- cytidine of nucleosides modifier | |
| CN115181147A (en) | Preparation method of C4 '-trifluoromethylthio modified nucleoside and C4' -trifluoromethylthio modified nucleic acid | |
| Heidenhain et al. | Improved methods for the preparation of 2′-deoxyribonucleoside and ribonucleoside 3′-phosphoramidites with allylic protectors | |
| CN1982301B (en) | A manufacturing process of 2',2'-difluoronucleoside and intermediate | |
| US6013790A (en) | Heavily fluorinated sugar analogs | |
| CN111875646B (en) | Preparation method of 2' -oxygen-propynyluridine | |
| CN114315935B (en) | Phosphoramidite nucleoside derivative and synthesis method, application and kit thereof | |
| CN117700369A (en) | Preparation method of 2- (2-chloropyrimidine-5-yl) acetic acid/ester | |
| KR100741310B1 (en) | Naphthalene 2-carboxylate derivative useful for synthesizing gemcitabine and a method for preparing the same | |
| JP2009256335A (en) | Preparation method of ribonucleic acid having alkyl protective group at position 2' | |
| JP2008508261A (en) | 1-α-halo-2,2-difluoro-2-deoxy-D-ribofuranose derivative and method for producing the same | |
| KR100957756B1 (en) | Process for preparing of 2'-deoxy-2'2'-difluorocytidine | |
| JP3025559B2 (en) | Adenosine derivatives | |
| EP2298782A1 (en) | Method for producing pyrazole glycoside derivatives |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20160928 |