Structure of endo-BCN-NHS carbonate
CAS No.: 1426827-79-3
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CAS No. : | 1426827-79-3 |
Formula : | C15H17NO5 |
M.W : | 291.30 |
SMILES Code : | O=C(ON1C(CCC1=O)=O)OC[C@@H]2[C@@]3(CCC#CCC[C@]23[H])[H] |
MDL No. : | MFCD19705416 |
InChI Key : | SKTDJYHCSCYLQU-FOSCPWQOSA-N |
Pubchem ID : | 75412386 |
GHS Pictogram: |
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Signal Word: | Warning |
Hazard Statements: | H302-H315-H319-H335 |
Precautionary Statements: | P261-P280-P301+P312-P302+P352-P305+P351+P338 |
* All experimental methods are cited from the reference, please refer to the original source for details. We do not guarantee the accuracy of the content in the reference.
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
In dichloromethane; at 20℃; | A solution of 5-((E)-3-aminoprop-1 -en-1 -yl)-5'-0-dimethoxytrityl-2'-deoxyuridine (1 .17 g, 2.0 mmol) (prepared according to the method of Santoro, et al., J. Am. Chem. Soc. 2000, i22(1 1 ), 2433-9)in DCM (10 mL) is treated with a solution of endo- bicyclo[6.1.0]non-4-yn-9-ylmethyl (2,5-dioxopyrrolidin-1-yl) carbonate (Vila) (0.64 g, 2.2 mmol) (obtained from SynAffix, Catalog No. SX-A1028) in dichloromethane (10 mL). After stirring overnight at room temperature, the reaction mixture is washed with saturated aqueous NaHC03 and the organic layer is dried over Na2SC>4. After filtration, the resulting solution is concentrated at reduced pressure. The residue is purified by chromatography on silica gel, eluting with a gradient of 1-5% methanol in dichloromethane. Fractions containing pure 12 are combined and concentrated at reduced pressure. The residue is dissolved in dichloromethane (50 mL) and concentrated again. Drying overnight under vacuum affords 12 as a crisp, colorless foam. MS (AP+): 762 (M+H). |
Yield | Reaction Conditions | Operation in experiment |
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In tetrahydrofuran; at 20℃; | General procedure: (R,S)-3-(3-aminopropoxy)propane-1 ,2-diol (1.64 g, 1 1 mmol) (Prepared according to the method of Misiura and Gait, WO 91 17169 A1 ) and VIIa (2.91 g, 10 mmol) are dissolved in anhydrous tetrahydrofuran (50 ml_) and stirred overnight at roomtemperature. The reaction solution is concentrated at reduced pressure and the residue is partitioned between ethyl acetate (50 ml.) and 5% aqueous Na2C03 (20 ml_). The organic layer is dried over Na2S04, filtered, and concentrated at reduced pressure. The residue is dissolved in anhydrous pyridine (30 ml_) and treated with dimethoxytrityl chloride (3.0 g, 9.0 mmol) and the resulting solution is stirred overnight at room temperature. The reaction is concentrated at reduced pressure and the resulting residue is partitioned between ethyl acetate and saturated aqueous NaHC03. The organic layer is washed with saturated aqueous NaCI, dried over Na2S04, filtered, and concentrated at reduced pressure. Flash chromatography on silica gel, eluting with a gradient of 5% to 45% ethyl acetate in hexanes affords 6a as a sticky foam upon evaporation of solvents at reduced pressure. MS (AP+): 628, (M+H). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With triethylamine; In tetrahydrofuran; water; at 20℃; for 1.5h; | To a solution of H2N-PEG8-COOH (822 mg, 1.86 mmol) in THF:H20 1 : 1 (20 mL) were added BCN-OSu (23) (651 mg, 2.23 mmol) and Et3N (774 μ, 5.59 mmol). The reaction was stirred at r.t. for 1.5 h andd acidified to pH 1 followed by extraction with EtOAc (3 x 35 mL). The combined organic layers were dried over Na2S04, filtered and the solvent removed under reduced pressure. The crude product was then dissolved in dry DCM (20 mL) and subsequently DCC (461 mg, 2.23 mmol) and NHS (257 mg, 2.23 mmol) were added. After stirring at r.t. for 1 h, the reaction was filtered and the filtrate concentrated in vacuo. Flash chromatography (MeCN, MeCN:H20 30: 1) afforded BCN-PEG8-COOSu.1H-NMR (300 MHz, CDC13): δ 5.44 (br s, 1H), 4.14 )d, J= 8.1 Hz, 2H), 3.84 (t, J= 6.3 Hz, 2H), 3.68-3.63 (m, 30H), 3.56 (t, J = 5.2 Hz, 2H), 3.34 (q, J = 5.4 Hz, 2H), 2.90 (t, J = 6.3 Hz, 2H), 2.85 (s, 4H), 2.36-2.14 (m, 6H). 1.72-1.49 (m, 2H), 1.36 (qn, J = 8.7 Hz, 1H), 1.02-0.88 (m, 2H).LRMS (ESI+) calcd for C34H54N2Oi4(M+Na+) 737.35, found 737.3. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
68% | In dichloromethane; for 3.16667h; | To a solution of 2,2'-(ethylenedioxy)bis(ethylamine) (11.78 mL, 80.5 mmol) in DCM (200 mL) was added BCN-OSu 23 (7.82 g, 26.8 mmol) in DCM (100 mL) dropwise over 3 h. After complete addition the mixture was stirred for 10 min followed by washing with saturated aqueous NH4C1 (3χ200 mL). The organic layer was dried over Na2S04, filtrated and concentrated in vacuo. Flash column chromatography (DCM:MeOH 99: 1-93 :7 + 1% Et3N) gave product 24 (5.95 g, 54.7 mmol, 68%). 1H- NMR (300 MHz, CDC13) δ 5.38 (s, 1H), 4.13 (d, J = 8.1 Hz, 2H), 3.59 (s, 4H), 3.56- 3.50 (m, 4H), 3.35 (q, J = 5.1 Hz, 2H), 2.88 (t, J = 5.1 Hz, 2H), 2.32 (br s, 2H), 2.27- 2.15 (m, 6H), 1.62-1.42 (m, 2H), 1.33 (qn, J = 8.7 Hz, 1H), 0.97-0.85 (m, 2H).13C- NMR (CDC13, 75 MHz) δ 156.4, 98.3, 68.7 (2C), 62.2, 45.5, 40.3, 40.2, 28.6, 20.9, 19.6, 17.3. HRMS (ESI+) calcd for C^s^NaC ( +Na+) 347.1947, found 347.1952. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
In water; N,N-dimethyl-formamide; at 55℃; for 4h; | General procedure: The freeze-dried 5-aminopropargyl-2'-deoxyuridine-5'-triphosphate (20 O.D.) was dissolved in 40.0 μL water and 40.0 μL 1.0 M TEAB buffer (pH = 7.5). The active ester (2.0 mg) in DMF (80.0 μL) was added to the triphosphate solution. The mixture was kept at 55oC for 4 h and the solvent was then removed in vacuo. The products were purified by RP-HPLC (eluentA: 0.1 M TEAB buffer (pH = 7.5), B: 60 % acetonitrile in 0.1 M TEAB (pH = 7.5) buffer, 3.5% - 60 % buffer B in 20 min). The pure products were characterized by mass spectrometry. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
A mixture of C6-C12) 50 mg, 0.033 mmol, as prepared in US patent No. 8,673848), sodium bicarbonate (18 mg, 0.215 mmol) and water (40 uL) in DMF (0.5 mL) was stirred at RTfor 10 mins, then (IR,8S)-bicyclo[6.1.0]non-4-yn- 9-ylmethyl succinimidyl carbonate (Berry &associates, 18 mg, 0.065 mmo[) was added. The reaction mixture was stirred at RT for 90 mins. A mxture of + I and +2 additions was observed by LCMS, so mixture was purified by mass triggered HPLC (Peptide Method 5 25- 50% ACN 5mm gradient: Conditions: Sunfire 3OxSOmm 5um column ACN/H20 w/ 0.1%TFA75m1/minl.5m1 injection): it 3.2mm (+1), it 4.65 mi 4.9mm (+1 and +2 mixture). LCMS confirms desired ±1 product in 61% yield and ±1, ±2 mixture in 18% yield. LCMS: (Basic Eluent A: Water + 5mM Ammonium Hydroxide Eluent B: ACN Acidic Column: Sunfire C18 3.Spm 3.Ox3Omm - 40C Basic Column: XBridge C18 3.Spm 3.Ox3Omm - 40C) Retention time: 0.98 mins; MS [M-i-2]2: observed: 856.0, calculated: 865.0245. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With N-ethyl-N,N-diisopropylamine; In tetrahydrofuran; N,N-dimethyl-formamide; at 20℃; for 0.5h; | Example 53: (S)-2-((2R,3R)-3-((S)-1 -((3R,4S,5S)-4-((S)-2-((1 R,3S,4S)-2-(6- ((((1 R,8S,9s)-Bicyclo[6.1 .0]non-4-yn-9-ylmethoxy)carbonyl)amino)hexanoyl)-2- azabicyclo[2.2.1 ]heptane-3-carboxamido)-N,3-dimethylbutanamido)-3-methoxy-5- methylheptanoyl)pyrrolidin-2-yl)-3-methoxy-2-methylpropanamido)-3-phenylpropanoic acid (75)To (S)-2-((2R,3R)-3-((S)-1 -((3R,4S,5S)-4-((S)-2-((1 R,3S,4S)-2-(6-aminohexanoyl)-2- azabicyclo[2.2.1 ]heptane-3-carboxamido)-N,3-dimethylbutanammethylheptanoyl)pyrrolidin-2-yl)-3-methoxy-2-methylpropanamido)-3-phenylpropan acid (Step 2, Example 44) (5 mg, 0.005 mmol) in DMF-THF (1 :1 , 2 ml) was added (1 R,8S,9s)-bicyclo[6.1 .0]non-4-yn-9-ylmethyl (2,5-dioxopyrrolidin-1 -yl) carbonate (1 .5 mg, 0.005 mmol) and DIEA (0.0025 ml, 0.014 mmol). The reaction mixture was stirred at rt for 30 min and then purified by preparative HPLC (40-65% acetonitrile-H20 containing 0.05% TFA) to obtain compound (75). MS m/z 1031 .6 (M+H). Retention time 1 .337 min. | |
With N-ethyl-N,N-diisopropylamine; In tetrahydrofuran; N,N-dimethyl-formamide; at 20℃; for 0.5h; | To (S)-2-((2R,3R)-3-((S)- 1 -((3R,4S,5 S)-4-((S)-2-((1R,3 S,4S)-2-(6-aminohexanoyl)-2- azabicyclo [2.2.1 jheptane-3 -carboxamido)-N,3 -dimethylbutanamido)-3 -methoxy-5 - methylheptanoyl)pyrrolidin-2-yl)-3 -methoxy-2-methylpropanamido)-3 -phenylpropanoic acid (Step 2, Example 3-44) (5 mg, 0.005 mmol) in DMF-THF (1:1, 2 ml) was added (1R,85,9s)-bicyclo[6.1.Ojnon-4-yn-9-ylmethyl (2,5-dioxopyrrolidin-1-yl) carbonate (1.5 mg, 0.005 mmol) and DIEA (0.0025 ml, 0.0 14 mmol). The reaction mixture was stirred at rt for 30 mm and then purified by preparative HPLC (40-65% acetonitrile-H20 containing 0.05% TFA) to obtain compound (75). MS m/z 1031.6 (M+H). Retention time 1.337mm. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
90% | In aq. phosphate buffer; dimethyl sulfoxide; at 20℃; for 5h;pH Ca. 7.6; | To (7.24 mg, 95 nmole) of IDS (1) in phosphate buffer 20 mM at pH 7.6, 380 nmole (4 equiv) of the BCN-N-hydroxysuccinimide ester (2) (from stock solution prepared as follows: 5.82 mg dissolved in 1000 μl of anhydrous DMSO) was added at RT for 5 h with occasional manual shaking. The modified IDS 3a, 70-56-1A was purified from the excess reagent by gel filtration with HiPrep 26/10 desalting column at 5 mL/minute with phosphate buffer 20 mM pH 7.6. The collected fractions were concentrated by Amicon ultra centrifugal filter (limit 10 kDa, 3000 rpm) to 3.8 mL (6.5 mg, yield 90%). The modified IDS 70-56-1A (3a) was recovered and was used for the next conjugation step with azido-An2 (N-terminus) (4). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
95% | In aq. phosphate buffer; dimethyl sulfoxide; at 20℃; for 5h;pH 7.6; | To (152 mg, 1992 nmole) of IDS in Phosphate buffer 20 mM at pH 7.6, 11952 nmole (6 equiv) of the BCN-N-hydroxysuccinimide ester (20 mmolar in anhydrous DMSO) was added at RT for 5 h with occasional manual shaking. The modified IDS 70-56-2B was purified from the excess reagent by gel filtration with desalting column Sephadex G25 at 15 mL/minute with phosphate buffer 20 mM, pH 7. The collected fractions (65 mL) were concentrated by Centricon Plus-70 (10 kDa, 3100 rpm) centrifugal filters to 36 mL, (145 mg, yield 95%). The modified IDS was recovered for the next conjugation step with azido An2 (N-terminus). To modified IDS (130 mg, 1704 nmole), 12 equiv of azidoAn2 (62 mg) were added. The solution was manually shaken, wrapped with aluminum foil and left overnight at RT. Conjugate was purified on a Q Sepharose 20 mL column using 20 mM TRIS buffer at pH 7 for binding buffer and 20 mM TRIS with 500 mM NaCl at pH 7.0 for eluent buffer. Conjugate (90 mL) was isolated, concentrated to 30 mL using Centricon Plus-70 (10 kDa, 3100 rpm) and exchanged with IDS buffer (1×: 137 mM NaCl, 17 mM NaH2PO4, 3 mM Na2HPO4, at pH 6) by washing (4×35 mL IDS buffer). The material was then concentrated to 33 mL with Centricon Plus-70 (10 kDa, 3100 rpm) columns and sterile filtered to obtain 70-56-2B (114 mg, 88%). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
100% | In aq. phosphate buffer; dimethyl sulfoxide; at 20℃; for 5h; | To (14.5 mg, 190 nmole) of IDS (1) in phosphate buffer 20 mM at pH 7.6, 1520 nmole (8 equiv) of the BCN-N-hydroxysuccinimide ester (2) (from stock solution prepared as follows: 5.82 mg dissolved in 1000 μl of anhydrous DMSO) was added and stored at RT for 5 h with occasional manual shaking. The modified IDS (10) was purified from the excess reagent by gel filtration with HiPrep 26/10 desalting column at 5 mL/minute with phosphate buffer 20 mM pH 7. The collected fractions were concentrated by Amicon ultra centrifugal filter (limit 10 kDa, 3000 rpm) to 4 mL (14.5 mg, yield 100%). The modified IDS was recovered and was used for the next conjugation step with An2-azido (C-terminus). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With N-ethyl-N,N-diisopropylamine; In N,N-dimethyl-formamide; at 20℃; for 1h; | Example 82: Synthesis (S)-2-((bis(dimethylamino)methylene)amino)-1-(((3R,4S,5S)-1- ((S)-2-((1R,2R)-3-(((S)-N-1-(3-((1R,8S,9s)-bicyclo[6.1.0]non-4-yn-9- ylmethyloxycarbonyl)aminopropylsulfonamido)-1-oxo-3-phenylpropan-2-yl)amino)-1- methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxoheptan-4- yl)(methyl)amino)-3-methyl-1-oxobutane (CL-33)A solution of (S)-2-((bis(dimethylamino)methylene)amino)-1-(((3R,4S,5S)-1-((S)-2- ((1R,2R)-3-(((S)-N-1-(3-aminopropylsulfonamido)-1-oxo-3-phenylpropan-2-yl)amino)-1- methoxy-2-methyl-3-oxopropyl)pyrrolidin-1-yl)-3-methoxy-5-methyl-1-oxoheptan-4- yl)(methyl)amino)-3-methyl-1-oxobutane (CL-32) (15 mg, 14 Pmol) and DIEA (12 Pl) in DMF (1 mL) was added to (1R,8S,9s)-bicyclo[6.1.0]non-4-yn-9-ylmethyl (2,5- dioxopyrrolidin-1-yl) carbonate (4.1 mg, 14 Pmol) and DIEA (12 Pl) in DMF (1 ml) . The reaction was covered by aluminum foil and stirred for 1 h at rt. Purification by preparative HPLC (20-70% acetonitrile-H2O containing 0.05% TFA) afforded compound (CL-33). MS m/z 101 3.5(M+1 ). Retention time 1.203 min. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With N-ethyl-N,N-diisopropylamine; In N,N-dimethyl-formamide; at 20℃; for 1h; | Example 83: Synthesis of (1 R,8S,9s)-bicyclo[6.1 .0]non-4-yn-9-ylmethyl (3-((S)-2- ((2R,3R)-3-((S)-1 -((3R,4S,5S)-4-((S)-2-((bis(dimethylamino)methylene)amino)-N,3- dimethylbutanamido)-3-methoxy-5-methylheptanoyl)pyrrolidin-2-yl)-3-methoxy-2- methylpropanamido)-3-phenylpropanamido)propyl)carbamate (CL-34)Step 4The product obtained in step 3 (13.3 mg, 15 Pmol) was dissolved in DMF (1 ml) and DIEA (13 Pl) was added. (1 R,8S,9s)-Bicyclo[6.1.0]non-4-yn-9-ylmethyl (2,5-dioxopyrrolidin-1 -yl) carbonate, , (4.4 mg, 15 Pmol) was dissolved in DMF (1 ml), and DIEA (13 Pl) was added. The two solutions were combined. The reaction mixture was stirred for 1 h at rt, and then purified by preparative HPLC (20-70% acetonitrile-H2O containing 0.05% TFA) to obtain compound (CL-34). MS m/z 949.6(M+1 ). Retention time 1.190 min. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
Step 1 :A mixture of pE-R-P-R-L-C*-H-K-G-P-Nle-C*-F-OH(Disulfide C6-C12) 50 mg, 0.033 mmol, as prepared in US patent No. 8,673848), sodium bicarbonate (18 mg, 0.215 mmol) and water (40 uL) in DMF (0.5 mL) was stirred at RT for 10 mins, then (1 R,8S)-bicyclo[6.1.0]non-4-yn-9- ylmethyl succinimidyl carbonate (Berry &associates, 18 mg, 0.065 mmol) was added. The reaction mixture was stirred at RT for 90 mins. A mxture of + 1 and +2 additions was observed by LCMS, so mixture was purified by mass triggered HPLC (Peptide Method 5 25-50% ACN 5 min gradient: Conditions: Sunfire 30x50mm 5um column ACN/H20 w/ 0.1 %TFA 75ml/min1.5ml injection): rt 3.2min (+1 ), rt 4.65 min, 4.9min (+1 and +2 mixture). LCMS confirms desired +1 product in 61 % yield and +1 , +2 mixture in 18% yield. LCMS: (Basic Eluent A: Water + 5mM Ammonium Hydroxide Eluent B: ACN Acidic Column: Sunfire C18 3.5μm 3.0x30mm - 40C Basic Column: XBridge C18 3.5μm 3.0x30mm - 40C) Retention time: 0.98 mins; MS [M+2]2+ observed: 856.0, calculated: 865.0245. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
at 20℃; | Step 1 : Preparation of NPFF click chemistry handleTo a solution of NPFF (Alfa Aesar, J66509, 5 mg, 3.82 μmοΙ) in DMSO (1 mL) was added triethylamine (5.32 μΙ, 0.038 mmol) and then (1 R,8S,9r)-bicyclo[6.1.0]non-4-yn-9-ylmethyl (2,5- dioxopyrrolidin-1-yl) carbonate (1.335 mg, 4.58 μmοΙ). The reaction mixture was stirred at room temperature. Upon completion the reaction mixture was taken on as crude to the next reaction step. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
87% | With triethylamine; In dichloromethane; at 20℃; for 18h;Inert atmosphere; Darkness; | To a solution of n-decylamine (10 μL, 0.05 mmol) in CH2Cl2 (5 mL) were added Et3N (16 μL, 0.11 mmol) and <strong>[1426827-79-3]BCN-NHS</strong> (15 mg, 0.05 mmol), and the mixture was stirred at room temperature under an inert atmosphere of N2 in the dark for 18 h. The reaction was then quenched with H2O (10 mL). The product was extracted with CH2Cl2 (10 mL*3) and the combined organic extract was dried over MgSO4, filtered, and the solvent was removed under reduced pressure. The residue was purified by column chromatography on silica gel using n-hexane as the eluent. The solvent was removed under reduced pressure and the product was subsequently isolated as yellowish oil. Yield: 16 mg (87%). 1H NMR (400 MHz, CDCl3, 298 K, TMS): δ 4.67 (s, 1H), 4.16 (d, J=8.0 Hz, 2H), 3.18 (d, J=6.4 Hz, 2H), 2.34-2.21 (m, 6H), 1.68-1.59 (m, 2H), 1.52-1.45 (m, 1H), 1.44-1.37 (m, 16H), 0.99-0.94 (m, 2H), 0.89 (t, J=6.8 Hz, 3H). 13C NMR (100 MHz, CDCl3, 298 K, TMS): δ 156.7, 98.8, 62.6, 41.1, 31.9, 30.0, 29.5, 29.3, 29.1, 26.8, 22.7, 21.4, 20.1, 17.8, 14.1. Positive-ion ESI-MS ion cluster at m/z 334 [M+H]+. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
40% | To a solution of amino-dPEG4-acid (1.23 g, 4.23 mmol) in anhydrous DMF (30 mL)were subsequently added 51 (1.02 g, 3.85 mmol) and triethylamine (1.60 mL, 11.53mmol). The reaction mixture was stirred for 3h at rt, after which EDCI.HC1 (0.884 g,4.61 mmol) and NHS (88 mg, 0.77 mmol) were added. The resulting solution was stirredovernight at rt and poured into 100 mL NaHCO3 (sat.) and 150 mL EtOAc. The layerswere separated and the organic phase was washed with sat. NaHCO3 (90 mL) and H20 (75 mL). The organic phase was dried (Na2SO4), filtered and concentrated in vacuo. Gradient flash chromatography (MeCN -* MeCN:H20 30:1) afforded product 60a as colorless oil (800 mg, 1.48 mmol, 40%). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
60% | To a solution of amino-dPEG8-acid (217 mg, 0.492 mmol) in anhydrous DMF (3 mL) were subsequently added 51 (143 mg, 0.492 mmol) and Et3N (204 tL, 1.47 mmol). The reaction mixture stirred for 3h at rt, after which EDCI.HC1 (0.88 g, 4.61 mmol) and NHS (88 mg, 0.77 mmol) were added. The resulting solution was stirred overnight at rt andpoured into 50 mL NaHCO3 (sat.) and 50 mL EtOAc. The layers were separated and the organic phase was washed with sat. NaHCO3 (50 mL) and H20 (30 mL). The organic phase was dried (Na2504), filtered and concentrated in vacuo. Gradient flash chromatography (MeCN - MeCN:H20 30:1) afforded product 60b as colorless oil (212 mg, 0.30 mmol, 60%).‘H NMR (300 IVIFIz, CDC13): (ppm) 4.13 (d, J 8.1 Hz, 2H), 3.84 (t, J 6.3 Hz, 2H),3.68-3.59 (m, 28 H), 3.54 (t, J 5.1 Hz, 2H), 3.36 (q, J= 5.4 Hz, 2H), 2.89 (t, J= 6.3Hz, 2H), 2.82 (s, 4H), 2.35-2.15 (m, 6H), 1.68-1.48 (m, 2H), 1.44-1.23 (m, 1H), 1.00-0.86 (m, 2H). LRMS (ESI+) m/z calcd for C34H54N20,4 (M+Naj = 737.8; found 737.3. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
To a solution of amino-dPEG12-acid (43 mg, 0.069 mmol) in anhydrous DMF (1 mL) were added 51 (22 mg, 0.076 mmol) and triethylamine 24 tL, 0.174 mmol). The reaction mixture stirred for 5 h at rt, after which EDCI.HC1 (27 mg, 0.139 mmol) and NHS (8 mg, 0.076 mmol) were added. The resulting solution was stirred overnight at rt and poured into 10 mL sat. NaHCO3 and 10 mL DCM. The layers were separated and the organicphase was washed with H20 (2 x 10 mL). The combined water layers were extracted with DCM (10 mL). The combined organic layers were dried (Na2SO4), filtered and concentrated in vacuo. Gradient flash column chromatography (MeCN - MeCN:H20 20:1, 10:1) afforded BCN-PEG12-C(O)OSu.‘HNIVIR (400 MHz, CDC13): (ppm) 4.14 (d, J 8.0 Hz, 2H), 3.85 (t, J 6.4 Hz, 2H),3.69-3.60 (m, 44 H), 3.56 (t, J 5.2 Hz, 2H), 3.36 (q, J= 5.2 Hz, 2H), 2.91 (t, J= 6.4Hz, 2H), 2.84 (s, 4H), 2.36-2.17 (m, 6H), 1.65-1.51 (m, 2H), 1.44-1.23 (,J 8.4 Hz, 1H), 1.00-0.88 (m, 2H). LRMS (ESI+) m/z calcd for C42H70N2018 (M+Hj = 892.0; found 891.6. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
To a solution of amino-dPEG24-acid (48 mg, 0.042 mmol) in anhydrous DMF (1 mL) were added 51 (14 mg, 0.048 mmol) and triethylamine (17 tL, 0.125 mmol). The reaction mixture was stirred at rt overnight, after which DCM (10 mL) and citric acid (10% aq. sol., 5 mL) were added. The water phase was extracted with DCM (15 mL) and the combined organic layers were dried (Na2SO4). After filtration the solvent was removed in vacuo. The crude product was redissolved in anhydrous DMF (1 mL) and EDCI.HC1 (17 mg, 0.088 mmol) and NHS (8 mg, 0.070 mmol) were added. The resulting solution was stirred overnight at rt, after which an addition equivalent of EDCI.HCL (16mg) and NHS (7 mg) were added. After 6h the reaction mixture was poured into 10 mL sat. NaHCO3 and 15 mL EtOAc. The layers were separated and the organic phase was washed with sat. NaHCO3 (10 mL) and H20 (10 mL). The organic phase was dried (Na2SO4), filtered and concentrated in vacuo. Gradient flash column chromatography (MeCN -* MeCN:H20 20:1, 10:1, 5:1) afforded BCN-PEG24-C(O)OSu as a colorless oil(32 mg, 0.022 mmol, 54%).‘HNIVIR (400 MHz, CDC13): (ppm) 4.08 (d, J 8.0 Hz, 2H), 3.78 (t, J 6.4 Hz, 2H), 3.63-3.54 (m, 92 H), 3.49 (t, J 5.2 Hz, 2H), 3.30 (q, J= 5.2 Hz, 2H), 2.84 (t, J= 6.4 Hz, 2H), 2.78 (s, 4H), 2.29-2.08 (m, 6H), 1.59-1.43 (m, 2H), 1.35-1.23 (m, 1H), 0.93- 0.80 (m, 2H). LRMS (ESI+) m/z calcd for C66H118N2030 (M+Hj = 1420.66; found1420.0. |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
With sodium hydrogencarbonate; In water; acetonitrile; for 4h; | To a solution of BCN-OSu derivative 51 in MeCN (5 mL) were added 7-aminoheptanoic acid 50 (145 mg, 1.0 mmol) in 0.1 M aqueous NaHCO3 (30 mL) and MeCN (25 mL). The mixture was stirred for 4 h and partially concentrated. Aqueous saturated NH4C1 (30 mL) was added and after extraction with DCM (2 x 30 mL), the combined organics were dried (Na2SO4) and concentrated. Product 52 was used in the step without furtherpurification. ‘H NIVIR (400 IVIHz, CDC13) (ppm) 4.68 (bs, 1H), 4.14 (d, J 7.9 Hz, 2H),3.17 (dd, J= 12.8, 6.3 Hz, 2H), 2.35 (t, J= 7.5 Hz, 2H), 2.32-2.09 (m, 6H), 1.70-1.25 (m, 1 1H), 0.94 (t, J 9.7 Hz, 2H). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
81% | With triethylamine; In dichloromethane; at 20℃; for 1.5h; | To a solution of 51 (430 mg, 1.48 mmol) in DCM (20 mL) was added a solution of 5-aminopentan-1-ol 55 (152 mg, 1.47 mmol) in DCM (4 mL) and Et3N (619 1iL, 449 mg,4.44 mmol). The mixture was stirred for 1.5 h at rt after which a saturated aqueoussolution of NaHCO3 was added (40 mL). After separation, the organic layer was dried (Na2SO4) and concentrated. The residue was purified by gradient column chromatography (EtOAc/heptane 1/1 -f 3/1). The product 56 was obtained as a colorless sticky liquid (356 mg, 1.27 mmol, 8 1%). ‘H NIVIR (400 IVIFIz, CDC13) (ppm) 4.68 (s,1H), 4.14 (d, J= 8.0 Hz, 2H), 3.65 (dd, J= 11.7, 6.3 Hz, 2H), 3.19 (dd, J 13.2, 6.7 Hz,2H), 2.35-2.15 (m, 6H), 1.66 - 1.30 (m, 7H), 1.02-0.88 (m, 2H). |
Yield | Reaction Conditions | Operation in experiment |
---|---|---|
98% | With N-ethyl-N,N-diisopropylamine; In acetone; at 50℃; for 20h; | DIPEA (52 mg, 0.41 mmol)was added to a solution of amine 6 (58 mg, 0.21 mmol) andcyclooctyne 7 (42 mg, 0.15 mmol) in acetone (5 mL), and thereaction mixture was stirred at 50 C for 20 h. The solvent wasremoved under reduced pressure, and the crude product waspurified by column chromatography (SiO2, 30/1 CHCl3/MeOH, Rf = 0.34). The title compound 8 (65 mg, 0.14 mmol,98%) was obtained as a red solid. Mp: 89 C. 1H NMR (500MHz, DMSO-d6): δ 0.83-0.86 (m, 2 H), 1.24-1.30 (m, 1 H),1.47-1.53 (m, 2 H), 2.10-2.14 (m, 4 H), 2.19-2.24 (m, 2 H),2.69 (s, 3 H), 2.86 (s, 3 H), 3.15 (q, J = 6.0 Hz, 2 H), 3.28 (q, J= 6.1 Hz, 2 H), 3.81 (s, 3 H), 4.04 (d, J = 8.0 Hz, 2 H), 6.38 (brs, 1 H), 6.86 (br s, 1 H), 6.89 (s, 1 H), 7.06 (s, 1 H), 7.24 (t, J =5.7 Hz, 1 H), 8.54 (t, J = 5.4 Hz, 1 H). 13C{1H} NMR (125MHz, DMSO-d6): δ 14.14 (CH), 17.79 (CH), 19.72 (CH),21.01 (CH2), 22.29 (CH2), 28.75 (CH2), 28.90 (CH2), 29.21(CH2), 30.04 (CH3), 30.90 (CH3), 31.49 (CH2), 51.90 (CH3),61.62 (CH2), 99.14 (2 C), 111.86 (CH), 112.21 (CH), 112.88(C), 125.57 (C), 137.95 (C), 143.39 (C), 156.85 (C), 167.85(C), 168.45 (C). IR (ATR): 1/λ 3389 (br, w), 2925 (m), 2853(w), 1694 (m), 1650 (w), 1532 (s), 1227 (s), 633 (m) cm-1. MS (EI, 70 eV): m/z (%) 456 (100) [M+], 324 (3), 306 (14),221 (19), 193 (6). HRMS (EI): calcd 456.2367 (forC24H32N4O5), found 456.2372 [M+]. UV-vis (CH2Cl2): λmax(log ε) 454 nm (3.66). Fluorescence (CH2Cl2): λem 567 nm, λex454 nm, Φ 0.09. C24H32N4O5 (456.53): calcd C 63.14%, H7.06%, N 12.27%; found C 63.19%, H 7.23%, N 11.94%. |
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