* 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.
Reference：
[1] Journal of the American Chemical Society, 1952, vol. 74, p. 2002
2
[ 1155-64-2 ]
[ 501-53-1 ]
[ 405-39-0 ]
Yield
Reaction Conditions
Operation in experiment
99%
With sodium carbonate In 1,4-dioxane; water at 20℃; for 1 h;
Preparation of Compound 24e A three-necked flask was loaded consecutively with H2O (40 mL), 1,4-dioxane (70 mL) and H-Lys(Z)-OH (10 g, 35.7 mmol). The mixture was stirred until complete dissolution. The pH was adjusted to about 10.5 by adding of 2 M aqueous Na2CO3. Benzyl chloroformate (6.69 g, 39.2 mmol) was added while maintaining the pH at about 10-11 by adding at the same time 2 M aqueous Na2CO3. After completing addition, the reaction mixture was stirred at 20 °C for 1 hour. Then EtOAc (50 mL) was added and pH of the resulting mixture was adjusted to 2-3 with c-HCl. The organic layer was separated and the aqueous layer was extracted with EtOAc (50 mL). The combined organic layers were washed with brine (50 mL), and dried over Na2SO4. Filtration and concentration under reduced pressure yielded the compound 24e as yellowish oil (14.7g, 99 percent). 1H-NMR (400 MHz, CDC13) δ 7.33-7.27 (m, 10H), 5.07-5.04 (d, 4H), 4.08 (m, 1H), 3.09 (t, 2H), 1.51 (br s, 1H), 1.49 (bs, 1H), 1.47-1.40 (m, 4H).
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3
[ 657-27-2 ]
[ 405-39-0 ]
[ 1155-64-2 ]
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4
[ 24424-99-5 ]
[ 1155-64-2 ]
[ 2389-45-9 ]
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5
[ 1155-64-2 ]
[ 58632-95-4 ]
[ 2389-45-9 ]
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[1] Patent: US4432971, 1984, A,
6
[ 1070-19-5 ]
[ 1155-64-2 ]
[ 2389-45-9 ]
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[4] Recueil des Travaux Chimiques des Pays-Bas, 1968, vol. 87, p. 559 - 571
[5] Indian Journal of Chemistry, 1965, vol. 3, p. 111 - 117
7
[ 18595-34-1 ]
[ 1155-64-2 ]
[ 2389-45-9 ]
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[1] Justus Liebigs Annalen der Chemie, 1968, vol. 716, p. 175 - 185
8
[ 16965-08-5 ]
[ 1155-64-2 ]
[ 2389-45-9 ]
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9
[ 58632-95-4 ]
[ 1155-64-2 ]
[ 2389-45-9 ]
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10
[ 1155-64-2 ]
[ 108-24-7 ]
[ 6367-08-4 ]
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[4] Tetrahedron, 2010, vol. 66, # 10, p. 1859 - 1873
11
[ 67-56-1 ]
[ 1155-64-2 ]
[ 27894-50-4 ]
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[2] Tetrahedron Asymmetry, 1998, vol. 9, # 9, p. 1505 - 1518
[3] Bioorganic and Medicinal Chemistry, 2008, vol. 16, # 10, p. 5473 - 5481
[4] Bioorganic and Medicinal Chemistry, 2011, vol. 19, # 9, p. 2991 - 2996
[5] Letters in Drug Design and Discovery, 2016, vol. 13, # 1, p. 98 - 106
12
[ 1155-64-2 ]
[ 27894-50-4 ]
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13
[ 1155-64-2 ]
[ 28920-43-6 ]
[ 86060-82-4 ]
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[4] Journal of Organic Chemistry, 1989, vol. 54, # 23, p. 5551 - 5558
14
[ 1155-64-2 ]
[ 88744-04-1 ]
[ 86060-82-4 ]
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15
[ 1155-64-2 ]
[ 6366-70-7 ]
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[1] Patent: US1312, 1994, H1,
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[ 1155-64-2 ]
[ 100-51-6 ]
[ 6366-70-7 ]
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17
[ 1155-64-2 ]
[ 13734-28-6 ]
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[1] Journal of Chemical Research, Miniprint, 1991, # 5, p. 914 - 934
With phosgene; In tetrahydrofuran; at 50 - 65℃; for 0.833333h;
EXAMPLE 3: PREPARATION OF N6-CBZ -L-LYSINE N-CARBOXYANHYDRIDE.; [0032] In this Example, a stirred mixture of 3.2 liters anhydrous tetrahydrofuran and 400 g (1.42 moles) N6-CBZ-L-lysine, was purged with EPO <DP n="16"/>nitrogen below liquid surface at 0.5 liters/min and heated to 50C over 30 minutes. The nitrogen purge was increased to 2 liters/min and 282 g (2.85 moles) gaseous phosgene was added at a rate to maintain a reaction mixture temperature of 50-65C over 5 minutes. The phosgene was refluxed back into the reaction vessel using a dry ice/acetone reflux condenser. After the phosgene was added, the reaction mixture was heated at 50-65C for15 minutes until the solids disappeared and a clear solution resulted. The dry ice and acetone were removed from the reflux condenser and the reaction mixture was purged with nitrogen at 4 liters/min and 50-65C for 30 minutes. The tetrahydrofuran and excess phosgene were removed by vacuum stripping at50-65C down to 100 mm Hg whereupon the concentrated residue crystallized. The stripped residue was dissolved in 2.4 liters anhydrous ethyl acetate with warming up to 30C and the cloudy mixture vacuum filtered to remove insoluble solids. 3.2 liters cold (5C), anhydrous hexanes were added gradually with stirring to the filtrate to crystallize the product. The product slurry was stirred for30 minutes and refrigerated at 5C overnight. The product was isolated by vacuum filtration under a nitrogen blanket, washed with a 0.8 liter solution of 4:1 , hexanes:ethyl acetate and with 0.8 liters hexanes, dried in the funnel by pulling vacuum on it for 30 minutes while maintaining a positive nitrogen purge and vacuum dried at room temperature to a constant weight. After drying, 380 g (87.0% yield) N6-CBZ-L-lysine N-carboxyanhydride were produced with a melting point of 98-100C and a chloride content of 0.082%.
With sodium hydroxide In water at 20℃; for 6.5h; Cooling with ice;
II.1 Step 1 : Preparation of Nn-lauroyl-N£-carbobenzoxy-L-Lysine
Step 1 : Preparation of Nn-lauroyl-N£-carbobenzoxy-L-Lysine [0050] 60.0 g of NE-cbz-L-Lysine (cbz is carbobenzoxy) purchased from Atomole Scientific Company, LTD was added to a three-liter beaker along with 1200 imL of RO water and the mixture was stirred. Next, 39 imL of 30% aqueous NaOH was added, resulting in dissolution of the NE-cbz-L-Lysine. The resulting solution was cooled in an ice bath and then 52.5 imL of lauroyl chloride was added. The ice bath was removed 30 minutes later, and stirring continued for an additional six hours, at which time 18 mL of concentrated hydrochloric acid was added. The reaction mixture was then filtered via vacuum filtration, the white solid product washed with 1 M aqueous HCI, and then the solid product was dried in vacuo while heated to approximately 85 °C. 96.5 g of dry white solid product was obtained. The product is further purified by dissolving it in methanol, filtering off any insoluble precipitate, and removing the methanol in vacuo to recover a white solid product (mp 99.5 - 103.0 °C)
With sodium hydroxide In water for 6.5h; Cooling with ice;
II.1 Step 1: Preparation of Nα-lauroyl-Nε-carbobenzoxy-L-Lysine
60.0 g of Nε-cbz-L-Lysine (cbz is carbobenzoxy) purchased from Atomole Scientific Company, LTD was added to a three-liter beaker along with 1200 mL of RO water and the mixture was stirred. Next, 39 mL of 30% aqueous NaOH was added, resulting in dissolution of the Nε-cbz-L-Lysine. The resulting solution was cooled in an ice bath and then 52.5 mL of lauroyl chloride was added. The ice bath was removed 30 minutes later, and stirring continued for an additional six hours, at which time 18 mL of concentrated hydrochloric acid was added. The reaction mixture was then filtered via vacuum filtration, the white solid product washed with 1 M aqueous HCl, and then the solid product was dried in vacuo while heated to approximately 85° C. 96.5 g of dry white solid product was obtained. The product can be further purified by dissolving it in methanol, filtering off any insoluble precipitate, and removing the methanol in vacuo to recover a white solid product (mp 99.5-103.0° C.)
25 tert-Butyl N6-((benzyloxy)carbonyl)-1-lysinate (8).
Nε-((benzyloxy)carbonyl)-1-lysine (12.03 g, 42.92 mmol) was mixed with t-butyl acetate (120 mL) in a 250 mL round bottom flask and concentrated HClO4 (3.90 mL) added to this mixture, producing a clear solution. This solution was stirred for 12 h before extracting with 200 mL H2O, 200 mL 5% HCl, then 200 mL, H2O. The aqueous extracts were combined and extracted with diethyl ether (3×200 mL) after addition of 30% NaOH solution until the aqueous layer was pH 11. The ether extracts were combined and dried over anhydrous MgSO4. The ether was then filtered and concentrated under reduced pressure and dried under a 50 μm vacuum overnight giving a colorless oil. Yield: 9.25 g (63%). 1H NMR (400 MHz, CDCl3): δ 1.30 (s 9H), 1.23-1.50 (m 8H), 2.99 (t 2H), 3.11 (t 1H), 4.91 (s 2H), 5.61 (br 1H), 7.14-7.16 (m 5H), 13C NMR (101 MHz, CDCl3): δ 175.18, 156.34, 142.38, 136.60, 128.31, 127.88, 108.60, 80.74, 77.46, 77.14, 76.82, 66.29, 54.66, 40.66, 34.36, 31.08, 29.53, 27.90, 22.64.
With perchloric acid
With perchloric acid at 23℃; for 14h;
With perchloric acid at 22℃; for 14h; Inert atmosphere;
HClO4 (3.01 mL) was slowly added to a stirred solution of Z-Lys 1 (6.5 g, 23.18 mmol) in t-butyl acetate (80 mL). The mixture was stirred at 22° C. for 14 h before extracting with H2O (150 mL) and 0.5N HCl solution (150 mL). The combined aqueous solutions were treated with 10% K2CO3 to give a solution of pH 9; the basic solution was then extracted with CH2Cl2 (3*100 mL). The combined organic layers were dried over anhydrous MgSO4, filtered, and then concentrated to give 2 as a colorless oil. The oil was dissolved in CH3CN (150 mL) before addition of NaHCO3 (4.29 g, 51.07 mmol) and t-butyl bromoacetate (13.58 g, 69.64 mmol). The mixture was heated at reflux for 15 h, then cooled to 22° C. before concentration of the mixture under reduced pressure and extraction of the residue with ethyl acetate (2*150 mL). The combined organic layers were washed with saturated NaCl solution (2*100 mL), dried over anhydrous MgSO4, and evaporated. The crude residue was purified by silica gel flash chromatography using 4:1 hexane:EtOAc as eluent to give compound 3 (11.62 g, 91%) as a colorless oil. 1H NMR (300 MHz, CDCl3) δ 7.30-7.35 (m, 5H), 5.08 (s, 2H), 3.44 (dd, J=17.4, 11.1 Hz, 4H), 3.30 (t, J=7.35 Hz, 1H), 3.20 (m, 2H), 1.62 (m, 2H), 1.53 (m, 4H), 1.45 (s, 9H), 1.43 (s, 18H); 13C NMR (75 MHz, CDCl3) δ 172.2, 171.2, 157.0, 137.34, 128.9, 128.6, 128.4, 81.64, 81.2, 66.9, 65.6, 54.4, 41.3, 30.6, 29.8, 28.7, 28.6, 23.5; HRMS (ESI): (M+H)+m/z calc'd for C30H48N2O8=564.7107. found 565.0643. The NMR spectra are in agreement with previously published data. (Huang et al., Bioconjugate Chem. 2006, 17, 1592-1600).
Stage #1: L-Lysine hydrochloride; benzyl chloroformate With sodium hydroxide; sodium hydrogencarbonate; copper(II) sulfate In water at 0℃; for 3h;
Stage #2: With hydrogenchloride; ethylenediaminetetraacetic acid In water
Stage #1: L-Lysine hydrochloride; benzyl chloroformate With copper(ll) sulfate pentahydrate; sodium hydrogencarbonate; sodium hydroxide In water at 0 - 20℃; Inert atmosphere;
Stage #2: With disodium ethylenediamine tetraacetic acid In water for 10h; Inert atmosphere; Reflux;
With sulfuric acid In 1,4-dioxane at -78 - 20℃; Inert atmosphere;
73%
With sulfuric acid In 1,4-dioxane for 18h;
55%
With sulfuric acid In 1,4-dioxane for 4h; Ambient temperature;
With sulfuric acid In 1,4-dioxane
With sulfuric acid In 1,4-dioxane for 6h; dry ice-acetone cold;
16.7 gm (64%)
With sodium hydroxide; sulfuric acid In 1,4-dioxane
IV EXAMPLE IV
EXAMPLE IV Into two separate 500 ml hydrogenation vessels each containing 100 ml of dioxane and 9.0 ml (160 mmol) of concentrated sulfuric acid was placed 11.0 gm (39.2 mmol) and 10.6 gm (37.8 mmol) of Nε-benzyloxycarbonyl-L-lysine. The two solutions were cooled to -78° C. and 140 ml of condensed (-78° C.) isobutylene was added to each vessel. The mixtures were then mechanically shaken (Parr shaker) at room temperature for 4 hours (26 psi). The reaction mixtures were combined and poured into 1000 ml (1.0 mol) of an ice cold solution of 1.0N aqueous sodium hydroxide and subsequently extracted three times with ether. The organic portions were combined (ca. 3000 ml), washed once with brine, dried over magnesium sulfate, filtered and concentrated in vacuo to give 16.7 gm (64%) of t-butyl Nε-benzyloxycarbonyl-L-lysinate (IV-A) as an oil.
16.7 gm (64%)
With sodium hydroxide; sulfuric acid In 1,4-dioxane
IV EXAMPLE IV
EXAMPLE IV Into two separate 500 ml hydrogenation vessels each containing 100 ml of dioxane and 9.0 ml (160 mmol) of concentrated sulfuric acid was placed 11.0 gm (39.2 mmol) and 10.6 gm (37.8 mmol) of Nε-benzyloxycarbonyl-L-lysine. The two solutions were cooled to -78° C. and 140 ml of condensed (-78° C.) isobutylene was added to each vessel. The mixtures were then mechanically shaken (Parr shaker) at room temperature for 4 hours (26 psi). The reaction mixtures were combined and poured into 1000 ml (1.0 mol) of an ice cold solution of 1.0N aqueous sodium hydroxide and subsequently extracted three times with ether. The organic portions were combined (ca. 3000 ml), washed once with brine, dried over magnesium sulfate, filtered and concentrated in vacuo to give 16.7 gm (64%) of t-butyl Nε-benzyloxycarbonyl-L-lysinate (IV-A) as an oil.
The N-carboxyanhydride of epsilon-carbobenzyloxylysine was synthesized from epsilon-carbobenzyloxylysine by phosgenation in dry tetrahydrofuran (THF). A 1,000 mL round bottom, four-necked flask with a thermocouple well was fitted with a calibrated addition funnel that was topped with a small dry ice cold finger. A second, larger cold finger was also fitted to the flask. A magnetic stirring bar was placed in the flask, which was charged with 500 mL of dry THF and 50 g (71 mmol) of epsilon-carbobenzyloxylysine (Aldrich, Product No. C8008). The contents of the flask were heated to 50 C. prior to phosgene addition. Liquid phosgene (25.4 mL, 142 mmol) was added in two equal portions to the reaction and the reaction mixture was stirred until a slightly cloudy solution was obtained. The solvent was then vacuum distilled at 40 C. or less to dryness and the product was resuspended in 150 mL of dry THF. The solution was then transferred to a dry box and filtered through activated charcoal (Aldrich; product no. 16,1551) and Celite Diatomaceous Earth (World Minerals Inc., Santa Barbara, Calif.) in separate steps. Then, hexanes (EM Science, Gibbstown, N.J.) were added to initiate crystallization. The solution was placed in a refrigerator at -20 C. for 24 h. The resulting white crystals (2 crops) were collected by vacuum filtration. The resulting yields were 47.32 g and 4.32 g respectively. The correct structure was confirmed by proton NMR in deuterated DMF.
With sodium hydrogencarbonate In water; <i>tert</i>-butyl alcohol Heating;
96%
With sodium hydroxide In water; <i>tert</i>-butyl alcohol at 20 - 40℃; for 18h;
92%
With sodium hydroxide In 1,4-dioxane; water at 20℃; for 16h; Inert atmosphere;
89%
With sodium hydroxide In <i>tert</i>-butyl alcohol Ambient temperature;
With sodium hydroxide; triethylamine In methanol for 2h;
4
As illustrated in Scheme VI, L-lysine reacts with N benzyloxycarbonyloxy-5-norborene-2,3-dicarboximide to provide N6- (benzyloxycarbonyl)-L-lysine (1). The N2-AMINO group is protected with TERT- butoxycarbonyl using di-tert-butyl dicarbonate to give the fully protected L-lysine (2), which condenses with L-proline TERT-BUTYL ester in the presence of N, N1- DICYCLOHEXYLCARBODIIMIDE to generate N-[N2-(TERT-BUTOXYCARBONYL)-N6- (BENZYLOXYCARBONYL) -L-LYSYL]-L-PROLINE TERT-BUTYL ESTER (3). THE TERT- butoxycarbonyl and tert-butyl ester protecting groups in compound (3) can be Scheme VI 0 NHz O Ph^O''NH J X f O 0 J H2N COOH 1 1 A o 1 lao pu nu JL o Ph O NH 1 0lolo N (OCC) 0 L A 1/'-' . 0 po H 0I'N COOH O H O O--f- 3 g TFA, 1 h, r. t. 0 0'in Ph^0I'NH Ph"0"NH J ho . O-N I vHCOOH 5 COOH N N NÇ OOH HzN H O O COOH COOH. p-N /6 1. 30-32% HBr/AcOH 2. 2% pyridine-H20 NH2 COOL COOL _ O COOH . O-N N 7 removed by treating compound (3) with trifluoroacetic acid providing N- [N6- (BENZYLOXYCARBONYL)-L-LYSYL]-L-PROLINE (4). Reductive coupling of (4) with 2- OXO-4- (2-OXY-1, 1,3, 3-tetramethyl-2, 3-DIHYDRO-1 H-ISOINDOL-5-YL) BUTYRIC acid (5) from Scheme 2 using sodium cyanoborohydride yields N-{N2-[L (S)-CARBOXY-3-(2- oxy-1,1, 3,3-tetramethyl-2, 3-DIHYDRO-1 H-ISOINDOL-5-YL) PROPYL]-N6-BENZYLOXYCARBO NYL-L-LYSYL}-L-PROLINE (6). Removal of the benzyloxycarbonyl group is achieved by treating compound (6) with 30-32% HBr in glacial acetic acid and then 2% pyridine/H20 solution generating the final product N-{N2-[L (S)-CARBOXY-3-(2- oxy-1,1, 3,3-tetramethyl-2, 3-DIHYDRO-1 H-isoindol-5-yl) propyl]-L-lysyl}-L-proline (7). The nitroxide (5), 2-oxo-4- (2-oxy-1, 1, 3,3-tetramethyl-2, 3-DIHYDRO-1H- isoindol-5-yl) butyric acid, may be synthesized by the method in Scheme 2:
With sodium hydroxide In tetrahydrofuran; water at 0℃; for 12h;
With sodium hydroxide In 1,4-dioxane; water at 20℃; for 3h; Cooling with ice;
12.1 Step 1 Synthesis of (2S)-6-(benzyloxycarbonylamino)-2-(tert-butoxycarbonylamino)hexanoic acid
To (2S)-2-amino-6-(benzyloxycarbonylamino)hexanoic acid (10.0 g, 35.7 mmol) was added 1,4-dioxane/water (80 mL/80 mL), 1M aqueous sodium hydroxide solution (35.6 mL) was added under ice-cooling, a 1,4-dioxane solution (50 mL) of di-tert-butyl dicarbonate (8.5 g, 39.2 mmol) was added with stirring and the mixture was stirred at room temperature for 3 hr. The reaction mixture was concentrated under reduced pressure, diluted with water, and acidified with 2N potassium hydrogensulfate aqueous solution. The aqueous layer was extracted with ethyl acetate, and the organic layer was dried over anhydrous sodium sulfate and concentrated to give the title compound (13.5 g, yield quantitative) without purification. 1H-NMR (300 MHz, DMSO-d6) δ12.40 (s, 1H), 7.35-7.29 (m, 5H), 7.24-7.23 (m, 1H), 7.04 (d, J=7.8 Hz, 1H), 4.99 (s, 2H), 3.83-3.76 (m, 1H), 2.98-2.92 (m, 2H), 1.62-1.50 (m, 2H), 1.42-1.37 (m, 13H).
at 20 - 25℃; Alkaline conditions;
2 N2-[(1,1-dimethylethoxy)carbonyl]-N6-[(phenylmethoxy)carbonyl]-L-lysine
A 100 L jacketed reactor was equipped with an overhead stirrer, thermocouple probe and a reflux condenser. To the reactor was added about 3.8 kilograms of N6-[(phenylmethoxy)carbonyl]- L-lysine, about 40 liters of deionized water, and about 2.05 kilograms (1.1 equiv.) of potassium carbonate. In a separate container was added about 3.1 kilograms (1.05 equiv.) of di-tert- butyldicarbonate and about 16 liters of 1,4-dioxane. The di-tert-butyldicarbonate solution was then gradually added to the solution of N6-[(phenylmethoxy)carbonyl]-L-lysine over 1 hour while the temperature of the reaction mixture was maintained at a temperature between about 20°C and 25°C, and the resulting reaction mixture was stirred until less than 1% of the N6- [(phenylmethoxy)carbonyl]-L-lysine material was determined to be present. The resulting mixture was concentrated under reduced pressure using a rotary evaporator, having a bath temperature of about 45 oC, to a volume of approximately 20 liters, following which about 7 liters of deionized water were added. The pH of the resulting mixture was then adjusted to a pH of approximately 3 to 4 by the gradual addition of 1 N hydrochloric acid (approximately 22.4 liters). To the resulting mixture was added about 20 liters of ethyl acetate, the mixture was agitated, the phases were allowed to separate, the aqueous phase was separated and placed into clean containers and the remaining organic phase was placed into clean containers. The aqueous phase was returned to the reactor, an additional portion (about 20 liters) of ethyl acetate were added, the mixture was agitated, the phases were allowed to separate, and the aqueous and organic layers were separated. This process was repeated a second time, after which the aqueous phases were discarded. The combined organic phases were returned to the reactor, about 13 liters of 1 N hydrochloric acid were added, the resulting mixture was agitated, the phases were allowed to separate, and the aqueous phase was removed and discarded. To the remaining organic phase was added about 13 liters of deionized water, the resulting mixture was agitated, the layers were allowed to separate, and the aqueous layer was separated and discarded. The organic phase was added about 10 liters of saturated, aqueous sodium chloride, the mixture was agitated, the phases were allowed to separate, and the bottom aqueous phase was removed and discarded. Another portion of about 10 liters of sodium chloride were added, the mixture was agitated, the phases were allowed to separate, and the bottom aqueous phase was removed and discarded. To the remaining organic phase were added about 2 kilograms of anhydrous magnesium sulfate, the mixture was stirred with a glass rod and then allowed to stand for not less than an hour. The resulting mixture was filtered through a medium porosity filter and the filter cake was washed with several volumes of ethyl acetate. The resulting solution was concentrated on a rotary evaporator, having a bath temperature of about 45 oC (maximum bath temperature 60 oC), under reduced pressure until no further weight loss was observed to afford N2- [(1,1-dimethylethoxy)carbonyl]-N6-[(phenylmethoxy)carbonyl]-L-lysine as a solid. The resulting solid was dried, with occasional stirring of the solid, in a vacuum oven having an oven temperature of about 50 oC until loss on drying was less than about 0.2% per hour.
With methyl-phenyl-thioether In trifluoroacetic acid at 25℃; for 3h; Yield given;
With trifluoroacetic acid In dichloromethane at 20℃;
With methyl-phenyl-thioether; trifluoroacetic acid at 25℃; conversion as a function of time, also in the case of other reagents: TFA/DMSO, TFA/anisole;
With methyl-phenyl-thioether; trifluoroacetic acid at 28℃; for 24h; var. reag.: dimethylsulfide, ethanedithiol, 2-mercaptoethanol; var. time;
98 % Chromat.
With cell extract of Sphingomonas paucimobilis SC 16113 In water at 42℃; for 20h; Enzymatic reaction;
With hydrogen bromide; potassium bromide; sodium nitrite In water at -13 - 0℃; for 6.5h;
2
(S)-6-[(Benzyloxy)carbonyl]amino}-2-bromohexanoic acid, Compound IV.(S)-2-Amino-6-[(benzyloxy)carbonyl]amino}hexanoic acid, 20.0 g (0.14 mole), and potassium bromide, 28.9 g (0.24 mole), were dissolved in 80 mL of -6N aqueous HBr, previously chilled to 00C. A stream of nitrogen was bubbled through the solution and the mixture was chilled to -10 0C by means of a cold bath. Sodium nitrite, 5.9 g (0.86 mole) was added in portions over 30 min with stirring, while maintaining the temperature of the reaction from -13°C to -10°C. After 6 hours stirring at -10°C, the mixture was extracted with ethyl ether, 4x50 mL and the resulting solution dried with magnesium sulfate. The mixture was filtered to remove the drying agent and evaporated to give an orange oil. The oil was purified by flash chromatography, chloroform-hexanes to give a clear and colorless oil. The yield was 20.0 g, i.e., 81 %, based on starting 2-amiϖo-6-[(benzyl-oxy)carbonyl]- aminojhexanoic acid. NMR is consistent with structure. (Nicolaides et al.)
79%
With hydrogen bromide; sodium nitrite In water at 0 - 20℃; for 2.5h;
26
Λ^-carbobenzyloxy-L-lysine 1 (commercial product) (15g; 0.052 mol) was dissolved in 6M HBr (45 mL) at O0C. NaNO2 (3.97 g; 0.057 mol) was added in little portions over 30 min. The reaction solution was stirred at room temperature for 2 h, then extracted with ethyl acetate (3 x 100 mL). The organic phase was dried (Na2SO4), filtered and evaporated under vacuum. The crude was purified by column chromatography to give the compound 2 (14.06 g) as an orange oil. Yield 79%.
23.2 g
With hydrogen bromide; sodium nitrite for 0.0833333h;
With sodium hydroxide In water 1.) 0 deg C, 30 min; 2.) up to RT, 30 min;
In sodium hydroxide
2.a (a)
(a) N2 -Benzoyl-N6 -[(phenylmethoxy)carbonyl]-L-lysine To an ice-cold solution of N6 -[(phenylmethoxy)carbonyl]-L-lysine (10.09 g., 36 mmole) in aqueous sodium hydroxide (1N, 26 ml.) is added benzoyl chloride (5 ml., 43.2 mmole) and aqueous sodium hydroxide (4N, 10.8 ml.) simultaneously in 5 portions over a period of 30 minutes. The ice bath is removed and stirring is continued for an additional 1.5 hours at room temperature. The reaction mixture is then extracted with ethyl acetate (discarded), the aqueous mother liquor is acidified with dilute hydrochloric acid and extracted with ethyl acetate. The ethyl acetate extract is concentrated and the residue crystallized from ethyl acetate-hexane to give 12.9 g. of N2 -benzoyl-N6 -[(phenylmethoxy)carbonyl]-L-lysine; m.p. 110°-112° (109°).
4 g
Stage #1: N6-[(benzyloxy)carbonyl]-L-lysine With sodium hydroxide In water at 0℃; for 1h; Inert atmosphere;
Stage #2: benzoyl chloride In water for 9h;
32 32. Preparation of N-(7-amino-l,l,l-trifluoro-2-oxoheptan-3-yl)benzamide
[0344] To ε-Cbz lysine (2.4 g, 8.56 mmol, 1 equivalent) in H20 (6 mL) was added NaOH (684.93 nig, 17.12 mrnol, 2 equivalent) in one portion at 0 °C under N2. The mixture was stirred at 0 °C for 1 hour. The mixture was then added compound benzoyl chloride (1.20 g, 8.56 mmol, 994.64 μ^, 1 equivalent) dropwise, and the mixture was stirred for 9 hours. The reaction mixture was filtered and the solid was washed with EtOAc (10 ml) to give 36.1 (4 g) as a white solid; LCMS [M + H]: 385; RT = 0.77 min.
With hydrogenchloride; sodium hydrogencarbonate In toluene at 25℃; for 24h;
80%
Stage #1: L-lysine With L-serin; L-leucine; fipronilβ-cyclodextrin In aq. buffer at 20℃; for 0.0833333h;
Stage #2: benzyl chloroformate In aq. buffer at 20℃; chemoselective reaction;
General procedure: β-Cyclodextrin (0.1 mmol) was dissolved in 0.1 M carbonate buffer (pH = 8)(10 mL) at room temperature, the L-lysine (1 mmol) and other amino acids (1 mmol)were added, and the reaction mixture was stirred for 5 min. Cbz-Cl (1 mmol) wasadded and stirring was continued at room temperature until the reaction was complete.The reaction mixture was extracted with ethyl acetate (2 × 5 ml). The solvent wasremoved under vacuum and the products were obtained.
78.6%
Stage #1: L-lysine With sodium carbonate In water for 1h;
Stage #2: benzyl chloroformate In water at 25℃; for 3h;
2
SiO2CD (40mg) was added to sodium carbonate aqueous solution (4mL), the pH value was about 9.5.After stirring until uniformly dispersed, L-lysine (0.1 mmol) was added. After stirring for 1 h, A1(benzyl chloroformate, Cbz-Cl, 0.09 mmol) wasadded dropwise.After reacting for 3 hours, stop the reaction, transfer the reaction solution to a separatory funnel, let stand for 1 hour, take the upper product, add water to wash and separate for 3 times, filter, and then wash with ethyl acetate (3mL) to obtain the final product B1, namely Nε -cbz-L-lysine, the yield is 78.6%, and the selectivity is >99%.
54%
Stage #1: L-lysine; benzyl chloroformate With 1,2,3-Benzotriazole; sodium hydroxide In tetrahydrofuran; water at 0℃;
Stage #2: With hydrogenchloride In water
7
A solution of benzyl chloroformate (1.84 g, 10.8 mmol) in THF (5 mL) was slowly added, stirring at 00C and maintaining pH 12 with aqueous sodium hydroxide, into a solution of L-lysine (1.7 g, 11.6 mmol) and benzotriazole(1.42 g, 11.9 mmol) in water (7 mL) and THF (11 mL) at pH12, obtaining a suspension. After adding a further 25 mL of THF and 25 mL of water to obtain a clear solution, the reaction mixture was stirred overnight, concentrated under vacuum, extracted with ethyl acetate, treated with hydrochloric acid to pH 6 and filtered. The product (1.64 g, 54%) is the expected H-Lys(Z)-OH (1H-NMR, 13C-NMR, ESI- MS) .
With copper(II) carbonate; sodium hydroxide
Stage #1: L-lysine With hydrogenchloride; copper carbonate hydroxide In water at 80 - 90℃; for 2h;
Stage #2: benzyl chloroformate With sodium hydrogencarbonate at 25℃; for 12h;
Stage #3: at 25℃; for 24h;
Stage #1: L-lysine With hydrogenchloride; copper (II) carbonate hydroxide In water at 80 - 90℃; for 2h;
Stage #2: benzyl chloroformate With sodium hydrogencarbonate at 25℃; for 12h;
Stage #3: With ethylenediaminetetraacetic acid at 25℃; for 24h;
The synthesis was adapted from literature and modified [37]. In a pre-dried three-necked flask equipped with a reflux condenser, dropping funnel and a septum, 14.00 g (50 mmol) Z-protected lysine were suspended in 100 ml of THF and the suspension was heated to 70 C. 6.6 ml (55 mmol) diphosgene were added over 30 min. The solution was heated until all solid disappeared (30-60 min). Dry nitrogen was then bubbled through the solution for 2-3 h to remove excess HCl and phosgene. The solution was concentrated in vacuo and dry hexane was added to precipitate the NCA. The suspension was stored in the fridge for 1 h. The solid was collected by filtration in an inert atmosphere and washed with hexane. It was then dissolved in the smallest possible volume of THF and again hexane was added to precipitate the product. The suspension was left in the fridge overnight. The solid was then collected by filtration in an inert atmosphere and dried in a stream of dry nitrogen. The recrystallization was repeated and AgNO3 (0.1 M in water) was added to the filtrate, to confirm the absence of chloride ions (AgCl would immediately precipitate as a colorless solid). The Lys(Z)NCA was dried and the colorless solid (13.23 g, 43.2 mmol, 86%) was transferred to a Schlenk tube and stored at -80 C. mp 99.1 C (lit: 101 C) [38]. 1H NMR (400 MHz, DMSO-d6) delta [ppm]=9.09 (s, 1H, CONH-Calpha), 7.40-7.25 (m, 6H, Ar H, NH(Z)), 5.01 (s, 2H, NHCH2Ph), 4.43 (t, 1H, CalphaH), 2.99 (q, 2H, CH2NH), 1.74-1.63 (m, 2H, CH-CH2), 1.44-1.31 (m, 4H, CH2-CH2); 13C NMR (75MHz, DMSO-d6) delta [ppm]=171.61 (CalphaCOOC=NH), 156.06 (COOBn), 151.94 (CalphaNHCOO), 137.21 (Ar), 128.29 (Ar), 127.67 (Ar), 65.09 (PhCH2), 56.97 (Calpha), 39.88 (Cepsilon), 30.58 (Cbeta), 28.72 (Cdelta), 21.55 (Cgamma).
3.6 g
With pyrographite; In tetrahydrofuran; at 60℃;
N6-carbobenzyloxy-L-lysine (5.0 g, 17.8 mmol) and activated charcoal (0.3 g) were suspended in tetrahydrofuran(70 mL). TCF (3.5 mL, 25 mmol) was added to the suspension under vigorous stirring. The temperature was increased to 60 C, and the reaction was continued until the N6-carbobenzyloxy-L-lysine dissolved completely. The solution was filtered through Celite using a glass filter,and the filtrate was concentrated under vacuum to givea brown oil. The crude product was crystallized from ethyl acetate and hexanes. Yield = 3.6 g. 1H NMR (ppm):7.39 (m, -C6H5), 6.40 (s, -C(O)NHCH-), 5.14 (s, -OCH2(C6H5)), 4.88 (s, -CH2NHC(O)-), 4.31 (s, -C(O)NHCH-),3.24 (t, -CH2NHC(O)-), 1.86 (d, -CHCH2CH2CH2CH2NH-), 1.59 (d, -CHCH2CH2CH2CH2NH-), 1.45(s, -CHCH2CH2CH2CH2NH-)
With sodium nitrite In water at 0℃; for 0.333333h;
54%
With sulfuric acid; sodium nitrite In acetonitrile at 0 - 20℃;
With sulfuric acid; acetonitrile; sodium nitrite
Multi-step reaction with 2 steps
1: L-amino acid oxidase from Providencia alcalifaciens, 0.06percent H2O2, phosphate buffer pH 7.4 / H2O / 51 h / 30 °C
2: L-HIC dehydrogenase, formate dehydrogenase, sodium formate, NAD / H2O / 64 h / 28 °C
With water; acetic acid; sodium nitrite at 0℃; for 0.666667h;
With acetic acid; sodium nitrite at 0℃; for 2h;
With sulfuric acid; water; sodium nitrite In acetonitrile at 0℃; for 5h;
Synthesis of Poly[α-(4-aminobutyl)-L-glycolicacid](Nε-cbz-PAGA)
Briefly, a solution of NaNO2 in water was added during 3 h to ice-cooled and stirred solution of N-cbzlysinein 2N H2SO4/acetonitrile (1:1). The mixture was stirred for an additional 2 h. After concentrating in vacuo,the resulting clear solution extracted with ether and was then recrystallized from ether/petroleum ether to give a white crystal (s)-6-[[(Phenylmethoxy)carbony]amino]-2-hydroxyhexanoic acid (PCAHA). The white crystal PCAHA was added into an oven-dried reaction tube and nitrogen-purged three times. High vacuum was applied at150 C. After 5 days of reaction, the crude polymer was dissolved in chloroform and purified by excess methanol to give a white powder N-cbz-PAGA.
N2-[4-(N-benzyloxy-N-methylcarbamoyl)-n-butanoyl]-N6-benzyloxycarbonyl-L-lysine[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
70%
Stage #1: glutaric acid 5-N-benzoyloxy-N-methyl monoamide With 4-methyl-morpholine; isobutyl chloroformate In tetrahydrofuran at -10℃; for 0.75h;
Stage #2: N6-[(benzyloxy)carbonyl]-L-lysine With triethylamine In tetrahydrofuran at -10 - 20℃; for 2h;
glutaric acid 5-N-benzoyloxy-N-cyclohexyl monoamide[ No CAS ]
[ 452303-41-2 ]
Yield
Reaction Conditions
Operation in experiment
70%
Stage #1: glutaric acid 5-N-benzoyloxy-N-cyclohexyl monoamide With 4-methyl-morpholine; isobutyl chloroformate In tetrahydrofuran at -10℃; for 0.75h;
Stage #2: N6-[(benzyloxy)carbonyl]-L-lysine With triethylamine In tetrahydrofuran at -10 - 20℃; for 2h;
With sodium hydroxide In water at 0 - 20℃; for 14h;
86.5%
With sodium hydroxide at 0 - 70℃;
80%
With sodium hydroxide In water at 20 - 50℃; for 21h;
80%
With water; sodium hydroxide at 0 - 60℃;
80%
With sodium hydroxide at 0 - 60℃; for 26h;
79.5%
Stage #1: bromoacetic acid With sodium hydroxide at 0℃;
Stage #2: N6-[(benzyloxy)carbonyl]-L-lysine With sodium hydroxide at 0 - 35℃;
Stage #3: With hydrogenchloride In water
6.1; 6.2; 6.3; 6.4
(1) Prepare a sodium bromoacetate solution.The bromoacetic acid and the sodium hydroxide solution with a mass concentration of 2M were mixed and stirred, andcooled to 0 ° C in an ice bath to obtain a sodium bromoacetate solution. The molar mass ratio of the lysine compound to the bromoacetic acid was 1:1.5,bromoacetic acid. The molar ratio to the sodium hydroxide solution is 1:1;(2) Dissolving the lysine compound in a 2M sodium hydroxide solution, and slowly adding it to thesodium bromate solution of the step (1) in anice bath, the lysine compound and the sodium hydroxide solution. The molar ratio was 1:2.5, and the reaction was carried out at 0 ° C for2 h, and then stirred at room temperature overnight;(3) The reaction liquid obtained in the step (2) is reacted in a water bath at 35 ° C for 2 h, then stirred to add a hydrochloric acid solution, cooled andfiltered to obtain a white solid, and the hydrochloric acid solution is equal to the molar amount of the sodium lysate solution in which the lysine is dissolved;(3) obtained in step (4) as a white solid was dissolved in a concentration of 1M sodium hydroxide solution, and then added with stirring and the likeamount of the concentration of 1M hydrochloric acid solution and cooled to precipitate a white solid, after drying in vacuo 24h, to give Compound A .The yield was79.5%.
With sodium hydroxide at 20 - 80℃; for 14h;
With sodium hydroxide In water at 0 - 50℃;
With sodium hydroxide
3
Na,Na-bis(carboxymethyl)-Ne-benzoxycarbonyl-L-lysine (Z-NTA) was synthesized from bromoacetic acid and Ne-benzoxycarbonyl-L-lysine. The protecting group Z (benzoxycarbonyl) was then removed by hydrogenation resulting in Na,Na-bis(carboxymethyl)-L-lysine (NTA) from which MANTA was synthesized as follows. NTA (0.262 g, 1 mmole) was dissolved in sodium hydroxide solution (aqueous, 3 mmole, 5 ml) together with tiny amount of inhibitor, and was cooled to 0° C. To this solution was added methacryloyl chloride (0.048 g, 1.2 mmole), and with slight delay, sodium hydroxide solution (aqueous, 1.2 mmole, 2 ml). Then the reaction was allowed to progress with stirring at room temperature for 2 hrs followed by extraction with equal volume of diethyl ether (anhydrous) to remove unreacted acid chloride. The water layer was acidified to pH23 with hydrochloric acid (aqueous, 6M) followed by repeated extraction with ethyl acetate (10 ml per time for 15 times) while being saturated by sodium chloride solid. Organic layers were combined, and dried over magnesium sulfate (anhydrous) overnight. After the drying agent was filtered off, the organic solvent was evaporated to obtain white solid, which was recrystallized from tetrahydrofuran. About 0.23 g of MANTA was obtained (yield 70%) with a melting point of 105107° C. TLC: MeOH/H20 (4/1), Rf=0.72. MW: 330.38 Da. C14H22N2O7. C %: 50.90 (50.86); H %: 6.71 (6.63); N %: 8.48 (8.41). 1H-NMR (DMSO-D6, 200 MHz): d=1.32-1.58 (m, 6H, CH2-(CH2)3); d=1.89 (s, 3H, CH3); d=3.04 (m, 2H, CO-NH-CH2); d=3.32 (t, 1H, CH2-CH(COOH)-N); d=3.46 (s, 4H, N-(CH2-COO)2); d=5.28, 5.60 (t, s, 2H, CH2); d=7.65 (t, 1H, CO-NH); d=12.40 (s, 2.7H, (COOH)3)
9.57 g
With sodium hydroxide In water at 0 - 50℃;
With sodium hydroxide In water at 18 - 50℃; for 18h;
12.4 g
With sodium hydroxide at 20 - 70℃; Cooling with ice;
1.1.1 (1) Synthesis of (1S)-N-(5-Carbobenzyloxyamino-1-carboxypentyl)iminodiacetic Acid (Nε-Cbz-NTA):
Weigh 8.4g Nε-Cbz-Lys (30mmol), add 45mL 2M sodium hydroxide solution to dissolve; then weigh 8.34g bromoacetic acid (60mmol) dissolved in 30mL 2M sodium hydroxide solution, under ice bath conditions, A sodium hydroxide solution of Nε-Cbz-Lys was added dropwise; the reaction was carried out overnight at room temperature, and then the reaction was heated to 70°C for two hours. After the reaction was completed, the reaction was cooled in an ice bath, and 90 mL of 1N hydrochloric acid solution was added, and a large amount of white precipitate was deposited in the solution; the precipitate was collected by suction filtration, washed twice with water, drained, and dried in vacuum to obtain 12.4 g of product Nε-Cbz -NTA.
CBZ-protected Lys (2.5 g, 8.91 mM) was treated with Triphosgene (1.4 g, 4.45 mM) in dry THF at 0 C. The solution was heated to 45 C. for 12 hrs. Any visible precipitates were removed by filtration and the solvent was removed in vacuo. Recrystallization was carried out with Hexane/THF (1:1; v/v) to yield 8. 1H NMR (300 MHz, CDCl3), delta 7.37-7.29 (s, 5H), 5.13 (s, 4H), 4.39-4.3 (d, 2H), 3.6 (s, 2H), 3.23 (s, 2H), 1.9-1.5 (m, 8H)
Ca. 80%
In tetrahydrofuran; at 65℃; for 2h;Inert atmosphere;
epsilon- (0228) (Z)-L-Lysine and D,L-Valine NCAs were synthesized as per the previous examples, with the inclusion of an additional purification step to remove hydrochloride impurities from the reaction. Dried H-Lys(Z)-OH (2 g, 7.14 mmol) or D,L-Valine (2 g, 17.0 mmol)) were suspended in anhydrous THF (50 ml_) in a three-necked round bottomed flask under argon. Triphosgene (lys: 0.85 g, 2.86 mmol, 1 .2 equiv. phosgene; val: 2.0 g, 6.74 mmol, 1 .2 equiv. phosgene) was then added and the mixture was refluxed at 65 C for 2 h with continuous stirring. After cooling to room temperature, the reaction mixture was sparged with argon for 45 mins into a sat. NaOH solution, then solvent removed completely in vacuo to a white solid. The solid was then suspected in EtOAc (anhydrous), chilled and placed into a separator funnel where the crude NCA solution was gently washed with chilled saturated brine solution (50 ml_), and 0.5 % w/v NaHCOs solution (50 ml_). The organic phase was then dried with MgS04, filtered and concentrated to an oil under low heat, and re-crystalized (x2) from EtOAc (anhydrous) and n-pentane (anhydrous). The resulting crystals were then filtered and washed with n-pentane (dry), then re- precipitated and washed (x 2) with dry n-pentane to afford white powder solids (Yields: -80 %) 1 H NMR (CDCIs): (Z)-L-Lysine NCA 1H NMR (400 MHz, CDCI3): deltaEta 1 .40-1.60 (m, 4H, NH-CH2-CH2-CH2-CH2-), 1 .81 -1.94 (m, 2H, NH-CH2-CH2-CH2-CH2-), 3.18 (m, 2H, NH-CH2-CH2-CH2-CH2-), 4.25 (t, 1 H, C/-/N), 4.97 (s, 1 H, side chain NH), 5.09 (s, 2H, CH2-ArH), 7.04 (s, 1 H, ring NH), 7.3-7.4 (m, 5H, ArH). D,L-Valine NCA 1 H NMR (400 MHz, CDCI3): deltaEta 1 .02 (d, 3H, J= 7.0 Hz, CH3), 1.08 (d, 3H, J= 7.0 Hz, CH3), 2.25 (m, 1 H, CH(CH3)2), 4.22 (d, 1 H, J = 4.4 Hz, CH-NH), 6.95 (s, 1 H, CO-NH).
78%
In tetrahydrofuran; at 50℃; for 2h;Inert atmosphere;
Lys(Z)-NCA was synthesized by referring to the literature procedure[30]. Briefly, in a round-bottom flask fitted with a stir bar, L-lysine beta-benzyl ester (1 g, 3.57 mmol) was suspended in anhydrous THF bubbledwith argon flux. After adding 0.5 equiv. of triphosgene, the suspensionwas stirred at 55 C for 2 h. Then, the filtrate of reaction mixture wasconcentrated and crystallized in hexane to achieve the purified Lys(Z)-NCA (yield: 78%). 1H NMR(400 M, DMSO-d6, ppm): 9.09 (s, 1H), 7.32 (m, 5H), 5.00 (s, 2H), 4.41 (m, 1H), 2.99 (m, 2H), 1.69 (m, 2H),1.39-1.29 (m, 4H)
In tetrahydrofuran; at 50℃;
Weigh 11.2g Nepsilon-benzyloxycarbonyl-L-lysine solid and dissolve in 180 ml anhydrous tetrahydrofuran; Weigh 5.2g bis(trichloromethyl) carbonate solid and dissolve in 40 ml anhydrous tetrahydrofuran. Poured into constant pressure funnel. Under oil bath reaction, the reaction temperature is maintained at around 50 C. The already dissolved bis(trichloromethyl) carbonate solution was drop by drop dropped into Nepsilon-benzyloxycarbonyl-L-lysine. 1-2h dropping complete. After completion of dropping, react for 0.5h. The reaction product and steaming and evaporation and concentration to 20 - 30 ml. For 200 ml anhydrous n-butane precipitation products, -4 C recrystallization 6h filtered, to remove the filtrate. The filtering the resulting solid in the drying in the vacuum drying box 12h, stored at -20 deg.C ;
In tetrahydrofuran; at 50℃;
Weigh 11.2gN -benzyloxycarboxy-L-lysine solid was dissolved in 180 mL of anhydrous tetrahydrofuran;5.2 g of bis (trichloromethyl) carbonate was dissolved in 40 mL of anhydrous tetrahydrofuran and poured into an isocratic funnel. In the oil bath reaction,The reaction temperature was maintained at about 50 C.The dissolved bis (trichloromethyl) carbonate solution was dropped dropwise dropwise into N-epsilon -benzyloxycarbonyl-L-lysine,1 ~ 2h drops finished, completely dripping and then reaction 0.5h.The reaction product was evaporated to evaporation to 20 to 30 mL.The product was precipitated with 200 mL of anhydrous n-butane and recrystallized at -4 C for 6 h to remove the filtrate.The resulting solidified by suction filtration was dried in a vacuum oven for 12 h at -20 C;
In 1,4-dioxane; at 50℃; for 2h;
The carboxy benzyloxycarbonyl lysine anhydride (N-carboxy- (Nepsilon-benzyloxycarbonyl)-L-lysine anhydride) was prepared by dissolving 9 mM benzyloxycarbonyllysine and 30 mM triphosgene in 30 mL of anhydrous dioxane and reacting at 50 C for 2 hours and then It was synthesized by precipitation with an excess of hexane
With sodium nitrite In water; acetic acid at 0 - 20℃; for 18.5h;
14
Add within 30 min, 400 mL of 2M NaNO2 solution (5.0 g; 70.0 mmol) to a suspension of L-CBzLysine (10.0 g; 35.7 mmol, Bachem) in 200 mL of a H2O/AcOH 1/1 mixture cooled on an ice bath. The reaction medium is stirred at room temperature for 18 hours and is then homogenous. Add 200 mL of water and extract with ethyl acetate (3×150 mL). Collect the organic phases and wash them with brine (3×100 mL). Dry on sodium sulfate. Evaporate the solvents and then take up the oily residue in toluene in order to remove a maximum of acetic acid. Evaporate to dryness in order to obtain 11.5 g of yellow oil (the O-acetylated derivative in majority). Perform diacetylation in 100 mL of a MeOH-H2O 1/1 mixture in the presence of 6.0 g of potassium carbonate.(pH 9-10). Reaction is completed after 3 hrs 30 mins. Add 50 mL of water and evaporate the methanol. Extract the aqueous phase with 100 mL of ethyl acetate. Acidify the aqueous phase (pH 2) and extract with ethyl acetate (2×100 mL). Wash with brine and dry on sodium sulfate. Evaporate in order to obtain 8.70 g of a sticky whitish powder. Triturate in 100 mL of pentane. Remove the solvent in order to obtain a white powder (7.22 g; 72%). 1H NMR (CDCl3-300 MHz): 1.51 (m, 4H), 1.72-1.76 (m, 2H), 3.18 (m, 2H), 4.25 (m, 1H), 5.11 (s, 2H), 7.34 (m, 5H). 13C NMR (CDCl3-75 MHz): 21.9; 29.3; 33.5; 40.9; 66.9; 70.1; 128.0-128.5; 136.4; 158.0; 177.7. Melting point: 74-76° C. (bed 79-81° C.).
1
One embodiment of the present invention includes the incorporation of a catalyst into the above-noted reaction to increase the speed of the reaction and to generate higher yield percentages of the present invention. The three following reagents were evaluated for their usefulness as catalysts for this dehydrative cyclocondensation reaction: sulfuric acid (H2SO4), phosphoric acid (H3PO4), and phosphorus pentoxide (P2O5) using m-cresol as a solvent. The term “substantially complete” as used herein with regard to the synthesis of the present invention refers to at least about 80% of the starting material being consumed within the prescribed time period. Reactions were carried out with each of the potential catalysts. The consumption of Cbz-L-lysine and the appearance of bis-3,6-[(N-benzyloxycarbonyl)-4-aminobutyl]-2,5-diketopiperazine (DKP1) were monitored by high performance liquid chromatography (HPLC) at time points throughout this reaction. At selected time points DKP1 was isolated and its yield calculated. The presence of sulfuric acid as a catalyst resulted in a yield of about 35%, which is within the range of the uncatalyzed reaction. However, this 35% yield was achieved in only 4 hours (see FIG. 3) as compared to the conventional uncatalyzed reaction times of from about 18 hours to about 33 hours to achieve a similar reaction yield percentage. The phosphoric acid catalyzed reaction was substantially complete in that it had consumed at least about 80% of the Cbz-L-lysine by about three hours. The DKP1 was stable when evaluated within 4 hours of the reaction and the yield was found to be about 55% (see FIG. 4).
48.9%
In 3-methyl-phenol at 165℃; for 0.5 - 4h;
1; 2; 3; 7
A 1-L round bottom flask equipped with a mechanical stirrer, short-path distillation apparatus, a thermocouple temperature read-out/controller, and a nitrogen inlet was charged with Cbz-L-lysine (100 g), m-cresol (200 g) and phosphorus pentoxide (7.5 g). The reaction mixture was stirred, heated to a target temperature of 165° C., and held at temperature for 1.5 hours. See FIG. 7. After completion of the heating period, the reaction solution was allowed to cool to room temperature (22-25° C.) and quenched with a solution of deionized water (100 mL) and MeOH (400 mL). The resulting suspension was stirred for 30-60 minutes and then filtered through a medium porosity sintered glass funnel. The filter cake was washed sequentially with 250 mL MeOH, 2×250 mL deionized water, and 2×250 mL MeOH. The wet cake was dried in a vacuum oven at 50° C. The recovered material was analyzed by HPLC and recrystallized from glacial acetic acid (3 mL/gram of crude product) according to the following procedure. The crude dry DKP1 in acetic acid was heated to reflux in an Erlenmeyer flask equipped with a thermometer and a reflux condenser with nitrogen head and outlet connected to a caustic scrubber. After 5 to 10 minutes (to ensure that complete dissolution occurred), the heat was removed and the mixture cooled to a temperature of less than 100° C. Deionized water (1 mL/gram of crude product) was added to the flask and the resulting suspension was cooled to room temperature with stirring over a period of 6-18 hours. The precipitated solid was filtered through a medium porosity sintered glass funnel and washed with deionized water (3×100 mL) and acetone (3×100 mL). The recrystallized wet cake was dried at 50° C. in a vacuum oven. Example 3 Optimization of Phosphorus Pentoxide Catalyzed Synthesis of DKP1 The dehydrative coupling and cyclization of Cbz-L-lysine to form DKP1 was evaluated as a function of reaction heating time and P2O5 charge. Reactions were carried out using 5%, 7.5% or 10% P2O5 (wttwt based on Cbz-L-lysine) with various heating times (Table 2). The data indicated that yield and purity were maximized using 7.5% P2O5. Subsequent experiments were designed to identify optimal conditions for the 7.5% phosphorus pentoxide catalyzed conversion of Cbz-L-lysine to DKP1. Additionally, these reactions were monitored for the disappearance of Cbz-L-lysine. FIG. 5 shows data obtained from a study in which the Cbz-L-lysine/cresol/P2O5 (7.5%) reaction mixture was monitored for DKP1 formation for 3 hours. During this time the reaction temperature was maintained at 165° C.+/-5° C. Aliquots (3 mL) were removed at specified times throughout the reaction, quenched with deionized water/methanol, and the product isolated by filtration. HPLC analysis of these samples suggested that the yield of DKP1 reached a maximum after heating for 1 to 3 hours (Table 2). After 4 hours at 165° C.+/-5° C., a 7-8% decrease in DKP1 yield was observed. Having identified 1-3 hours as the optimal reaction time, several reactions (100 g scale) were conducted to measure isolated DKP1 yields within this time period. FIG. 8 shows that the maximum DKP1 isolated yield (60.9%) was obtained after 1.5 hours. HPLC analysis showed this material to be >99.5% DKP1. This reaction was repeated with comparable results, namely DKP1 (100.0 area %) was isolated in a 60.3% yield. The reaction was also performed using heat ramps of 4 hours and 6 hours to simulate the large volume heating times. These reactions gave yields of 59.6% and 63.1%, respectively (Table 2).; The optimized conditions (7.5% P2O5, 165° C., 1.5 hours) were then demonstrated twice at the 5-L scale. Again a 4-hour and a 6-hour ramp were employed to stimulate the heating profile employed in production volumes. Both reactions reproduced the results obtained at the 1-L scale. The reactions have overall yields of 58.9% and 58.4% with purity comparable to the current process (99.8%).
35%
In 3-methyl-phenol at 165℃; for 4h;
1
One embodiment of the present invention includes the incorporation of a catalyst into the above-noted reaction to increase the speed of the reaction and to generate higher yield percentages of the present invention. The three following reagents were evaluated for their usefulness as catalysts for this dehydrative cyclocondensation reaction: sulfuric acid (H2SO4), phosphoric acid (H3PO4), and phosphorus pentoxide (P2O5) using m-cresol as a solvent. The term “substantially complete” as used herein with regard to the synthesis of the present invention refers to at least about 80% of the starting material being consumed within the prescribed time period. Reactions were carried out with each of the potential catalysts. The consumption of Cbz-L-lysine and the appearance of bis-3,6-[(N-benzyloxycarbonyl)-4-aminobutyl]-2,5-diketopiperazine (DKP1) were monitored by high performance liquid chromatography (HPLC) at time points throughout this reaction. At selected time points DKP1 was isolated and its yield calculated. The presence of sulfuric acid as a catalyst resulted in a yield of about 35%, which is within the range of the uncatalyzed reaction. However, this 35% yield was achieved in only 4 hours (see FIG. 3) as compared to the conventional uncatalyzed reaction times of from about 18 hours to about 33 hours to achieve a similar reaction yield percentage.
25%
In 3-methyl-phenol at 165℃; for 24 - 33h;
1; 2
One embodiment of the present invention includes the incorporation of a catalyst into the above-noted reaction to increase the speed of the reaction and to generate higher yield percentages of the present invention. The three following reagents were evaluated for their usefulness as catalysts for this dehydrative cyclocondensation reaction: sulfuric acid (H2SO4), phosphoric acid (H3PO4), and phosphorus pentoxide (P2O5) using m-cresol as a solvent. The term “substantially complete” as used herein with regard to the synthesis of the present invention refers to at least about 80% of the starting material being consumed within the prescribed time period. Reactions were carried out with each of the potential catalysts. The consumption of Cbz-L-lysine and the appearance of bis-3,6-[(N-benzyloxycarbonyl)-4-aminobutyl]-2,5-diketopiperazine (DKP1) were monitored by high performance liquid chromatography (HPLC) at time points throughout this reaction. At selected time points DKP1 was isolated and its yield calculated. The presence of sulfuric acid as a catalyst resulted in a yield of about 35%, which is within the range of the uncatalyzed reaction. However, this 35% yield was achieved in only 4 hours (see FIG. 3) as compared to the conventional uncatalyzed reaction times of from about 18 hours to about 33 hours to achieve a similar reaction yield percentage.
With sodium hydrogencarbonate In sodium hydroxide; ethanol; water
5 Synthesis of Nα-(4-Nitrophenyl)-Nε-(benzyloxycarbonyl)-L-lysine
EXAMPLE 5 Synthesis of Nα-(4-Nitrophenyl)-Nε-(benzyloxycarbonyl)-L-lysine 5 g (17.83 mmol) of Nε-benzyloxycarbonyl-L-lysine are dissolved in 130 ml of water in the presence of 0.71 g (1 eq) of sodium hydroxide and 2.25 g (1.5 eq) of sodium bicarbonate in a 250 ml three-necked flask equipped with a reflux condenser and a thermometer. A solution of 2.85 ml (1.5 eq) of 4-fluoronitrobenzene in 70 ml of ethanol is run into the reaction mixture, which is subsequently brought to reflux for 48 hours. The mixture is neutralized with 1N hydrochloric acid, evaporated and then extracted with ethyl acetate. The aqueous phase is acidified to pH~3 and again extracted with ethyl acetate. The organic phases are combined, washed with water and dried over sodium sulphate. After evaporation, an orangy-yellow oil is obtained, which oil is purified by flash chromatography (eluent: CH2Cl2 95/CH3OH 5) to give 1.6 g of a yellow crystalline product, i.e. a final yield of 22%.
With hydrogenchloride; sodium hydroxide; N2; toluene-4-sulfonic acid; benzyl alcohol; In water; toluene;
EXAMPLE 1 Preparation of (N-epsilon-carbobenzyloxy)lysine benzyl ester hydrochloride 2 A suspension of 48.69 g (174 mmol) of (N-epsilon-carbobenzyloxy)lysine (Bachem, Inc.) in a solution of p-toluenesulfonic acid (33.04 g, 174 mmol) in toluene (500 mL) and benzyl alcohol (330 mL) was heated to reflux under a Dean-Stark separator. As reaction progressed the solution became homogeneous. Reflux was continued for 22 hr, at which time 6.6 mL of water (106% of theoretical) had collected. The bulk of the toluene was distilled out of the reaction mixture at atmospheric pressure, then the solution was concentrated by distillation under reduced pressure until 750 mL (90%) of the mixture of solvents had been collected. The residue was diluted with EtOAc (800 mL), chilled in an ice bath, and treated with water (200 mL), 2N NaOH (87 mL), and brine (200 mL). A small amount of flocculent precipitate was removed by filtration, and the pH of the aqueous phase was adjusted to 10 with 2N NaOH. The organic phase was washed with brine, dried over MgSO4, filtered to remove dryinq agent, and cooled to ice-bath temperature. Anhydrous HCl was bubbled through the solution for 5 minutes, resulting in the formation of a copious white precipitate. Stirring was continued for an additional 5 minutes, then N2 was bubbled through the mixture for 20 minutes to dispel HCl. The product was collected by filtration and dried under vacuum over P2 O5 to afford a white solid. 1 H NMR (CF3COOD) delta 1.38 (m, 2H), 1.58 (m, 2H), 2.15 (m, 2H), 3.30 (m, 2H), 4.41 (m, 2H), 5.31 (s, 2H), 5.40 (AB, J=5.6 Hz, 2H), 7.42 (m, 10 H).
With triethanolamine In chloroform; ethyl acetate; N,N-dimethyl-formamide
14.a a.
a. N-alpha-(2-Methylsulfonylethyloxycarbonyl)-N-epsilon-benzyloxycarbonyl-L-lysine To a suspension of N-epsilon-benzyloxycarbonyl-L-lysine (20.3 g, 72.25 mmol) in DMF (300 ml) was added 2-methylsulfonylethyl p-nitrophenyl carbonate (20.85 g, 72.25 mmol). The reaction mixture was stirred overnight at room temperature, TEA (10.0 ml, 72.25 mmol) was added and the mixture was stirred for an additional 24 hr. and filtered. The filtrate was concentrated under vacuum. The residue was dissolved in EtOAc (300 ml) and washed with aqueous 1N HCl and brine. The organic phase was dried over Na2 SO4, filtered and concentrated under vacuum to give a viscous yellow oil. The crude product was purified by flash chromatography on silica gel with CHCl3, MeOH:CHCl3 (2:98) and finally MeOH:CHCl3 (5:95) to give the product (20.67 g); TLC, Rf =0.29, silica gel, MeOH:CHCl3:HOAc (9.5:90:0.5).
9.a (a)
(a) N2 -[(1,1-Dimethylethoxy)carbonyl]-N-6 -[(phenylmethoxy)carbonyl]-L-lysine A mixture of N6 -[(phenylmethoxy)carbonyl]-L-lysine (5.6 g., 20 mmole) and triethylamine (4.2 ml., 30 mmole) in dioxane (50 ml.)--water (20 ml.) is treated with 2-(t-butoxycarbonyloxyimino)-2-phenylacetonitrile (5.4 g., 21.9 mmole) in dioxane (30 ml.) and stirred at room temperature under argon. The heterogeneous mixture becomes a clear yellow solution after 2 hours. After 4 hours, the mixture is diluted with water and extracted with ethyl acetate (100 ml.) and then with ethyl acetate (50 ml.)-ether (40 ml.). The aqueous layer is acidified with 5% citric acid to pH4 and extracted twice with ethyl acetate (2*80 ml.). The combined extracts are washed successively with water (2*50 ml.), brine, dried (Na2 SO4), and the solvent stripped to give 7.5 g. of N2 -[(1,1-dimethylethoxy)carbonyl]-N6 -[(phenylmethoxy)carbonyl]-L-lysine as a colorless oil. Tlc (dichloromethane/methanol, 5:1) shows a single spot at Rf 0.32.
With N-ethylmorpholine;; methylamine In tetrahydrofuran; 1,4-dioxane; sodium hydroxide; ethyl acetate
44.i (A)
(i) 21.0 g of N6 -[(benzyloxy)carbonyl]-L-lysine were dissolved in 75 ml of 2M sodium hydroxide solution and 75 ml of dioxane. Thereafter, 18.0 g of di-tert.butyl dicarbonate were added and the mixture was stirred at room temperature for 16 hours. The solution was evaporated in order to remove dioxane, water was added, the solution was extracted with diethyl ether and acidified with 6M hydrochloroic acid. The product was taken up in ethyl acetate, washed with sodium chloride solution, dried over magnesium sulfate and evaporated to give an oil. A solution of this oil in tetrahydrofuran was cooled to -15° C. and treated with 8.51 ml of N-ethylmorpholine, 8.61 ml of isobutylchloroformate and, after 5 minutes, with 10.0 ml of a 40% aqueous solution of methylamine. After stirring at 0° C. for 2 hours, the solvent was removed by evaporation and the residue was dissolved in ethyl acetate. The organic solution was washed with water, 5% citric acid solution, water, 5% sodium hydrogen carbonate solution and sodium chloride solution, dried over anhydrous magnesium sulfate and evaporated. Recrystallization from ethyl acetate yielded 20.5 g of N6 -[(benzyloxy)carbonyl]-N2 -(tert.-butoxycarbonyl)-L-lysine methylamide in the form of a white solid of melting point 100°-102° C.
With chloro-trimethyl-silane; N-ethyl-N,N-diisopropylamine In dichloromethane; water
10 (S)-N6-Benzyloxycarbonyl-N2-mesyllysine
PREPARATION 10 (S)-N6-Benzyloxycarbonyl-N2-mesyllysine (S)-N6-Benzyloxycarbonyllysine (1.5 kg) was slurried in methylene chloride (7.5 l) and chlorotrimethylsilane (1.36 l) added over 10 minutes. The mixture was heated under reflux for 30 minutes to give a solution which was cooled to 3° C. before simultaneously adding diisopropylethylamine (1.87 l) and methanesulphonyl chloride (435 ml) at such a rate as to keep the temperature below 25° C. The reaction was stirred for a further 2.5 hours then poured into 2 M aqueous hydrochloric acid solution. The layers were separated and the methylene chloride phase was washed with 2 M aqueous hydrochloric acid solution followed by water. The solvent was removed under reduced pressure and replaced with n-butyl acetate. The solution was cooled and the resulting crystalline material was collected by filtration, washed with n-butyl acetate and dried under reduced pressure to provide the title compound (1.63 kg), m.p. 83.5-84° C. [α]D25-13.4° (c=1, methanol). Found: C,50.23; H,6.40; N,7.76. C15H22N2O6S requires: C,50.27; H,6.19; N,7.82%. 1H-NMR (300 MHz, d6-DMSO): δ=1.23-1.78(6H,m), 2.85(3H,s), 2.98(2H,q), 3.80(1H,dt), 5.00(2H,s), 7.25(1H,t), 7.30-7.43(5H,m), 7.51(1H,d) ppm.
4 Synthesis of "Nitrilotriacetic Acid" (NTA)
Example 4 Synthesis of "Nitrilotriacetic Acid" (NTA) N,N-Bis-(carboxymethyl)lysine (commonly referred to as "Nitrilotriacetic acid," or "NTA") was synthesised as follows based the procedure reported by Hochuli et al. (Journal of Chromatography, 411:177-184 (1987)). A solution of H-Lys(Z)-OH (42 g; 150 mmol) in 2N NaOH (225 mL) was added drop wise to a solution of bromoacetic acid (42 g; 300 mmol; 2 eq) in 2N NaOH (150 mL) at ~0 to 10° C. White precipitate formed as the solution of H-Lys(Z)-OH added. The reaction continued at room temperature (RT) overnight, after which the temperature was increased to 60° C. and the reaction continued for another 2 h. 1N HCl (450 mL) was added and the mixture was place in a refrigerator for a couple hours. The solid product (Z-protected NTA) was filtered off and recrystallized by re-dissolving the solid in 1N NaOH, then neutralized with the same amount of 1N HCl. The Z-protected NTA was collected by filtration and dried. Z-protected NTA was dissolved in 1N NaOH (130 mL) and 5% Pd/C (~450 mg) was added. The reaction mixture was evacuated and saturated with H2 before being stirred at RT under H2 balloon overnight. The reaction mixture was filtered through a celite bed to remove the Pd/C. The filtrate, containing NTA was collected and water (80 mL) was used to wash the filtering bed. 6N HCl was added to bring the pH down to 7.5-8.0. The collected NTA solution was diluted with water to have the final concentration of ~200 mM.
Stage #1: N6-[(benzyloxy)carbonyl]-L-lysine With sodium hydroxide In water for 1h;
Stage #2: C27H44O7 In water; acetone
Stage #3: With hydrogenchloride In water; acetone at 30℃;
1
Reaction Vessel 2Water (800 ml) was charged to a 2L flask and stirring was commenced. Sodium hydroxide (15 g) was charged to the flask and the reaction was stirred for 10 minutes. Λ/-ε-CBZ-L-Lysine (100 g) was charged to the vessel and the reaction was stirred for a further 60 minutes. The contents of vessel 2 were charged, as rapidly as possible, to vessel 1 followed by stirring for a further 2-4 hours. The pH of the reaction mixture was adjusted to pH 2.0 to 2.5 using 1 M Hydrochloric acid, maintaining the temperature below 3O0C by controlling the addition rate. The contents of vessel 1 were transferred to a suitable separating funnel, ethyl acetate (1.0 L) was charged and the solution was vigorously agitated for 5 minutes. The two layers were separated and the lower aqueous layer was recharged to the separating funnel. Ethyl acetate (1.0 L) was charged and the extraction procedure was repeated.The ethyl acetate layers were combined and recharged to the separating funnel. Water (5.0 L) was charged to the separating funnel and the reaction mixture was vigorously stirred for 5 minutes. The two layers were separated and the upper organic layer was separated and dried with sodium sulphate. The solution was filtered, the filter pad washed with ethyl acetate (1.0 L), and the solvent removed on a rotary evaporator (temp 35 to 5O0C) to afford N-ε-CBZ-cholyl-L-lysine as a white solid.m/z = 371.6 (M+1 ) m.p = 110-1 14 0CTLC (silica gel 60) Rf = 0.15 (20 % Methanol/dichloromethane) IR (KBr) 3340, 2922, 2853, 1703, 1537, 1460, 1376, 1252 cm"1
Stage #1: undec-10-enoyl chloride; N6-[(benzyloxy)carbonyl]-L-lysine With sodium hydroxide In water at 20℃; for 1h;
Stage #2: With hydrogenchloride In water
Stage #1: N6-[(benzyloxy)carbonyl]-L-lysine With calcium hydroxide In water at 20℃;
Stage #2: tert-butyloxycarbonyl-L-alanine p-nitrophenyl ester In N,N-dimethyl-formamide at 25℃; for 24h;
Stage #1: N6-[(benzyloxy)carbonyl]-L-lysine With lithium hydroxide monohydrate In methanol at 20℃; for 0.333333h;
Stage #2: 3-phenyl-propionaldehyde In methanol for 1h;
General synthetic procedure for 5 - 9a - f
General procedure: The amino acid specified below was dissolved in methanol (100 mL). LiOH.H20 was added, and the solution stirred at room temperature for 20 minutes. The corresponding aldehyde was added dropwise, and stirred for 1 hour. Sodium borohydride was then added portionwise. After 30 minutes, the solvent was removed under reduced pressure and the resulting solid was dissolved in water (100 mL). Using 1M HCl, the pH was adjusted to 6.5 and then filtered. The crude white solid was carried on without further purification.
Stage #1: N6-[(benzyloxy)carbonyl]-L-lysine With lithium hydroxide monohydrate In methanol at 20℃; for 0.333333h;
Stage #2: benzaldehyde In methanol for 1h;
General synthetic procedure for 5 - 9a - f
General procedure: The amino acid specified below was dissolved in methanol (100 mL). LiOH.H20 was added, and the solution stirred at room temperature for 20 minutes. The corresponding aldehyde was added dropwise, and stirred for 1 hour. Sodium borohydride was then added portionwise. After 30 minutes, the solvent was removed under reduced pressure and the resulting solid was dissolved in water (100 mL). Using 1M HCl, the pH was adjusted to 6.5 and then filtered. The crude white solid was carried on without further purification.
(2S)-6-[(benzyloxy)carbonyl]amino}-2-hydroxyhexanoic acid[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
With sodium nitrite In water at 0℃; for 0.583333h;
29.I
Example 29: Preparation of (2S)-6-[(benzyloxy)carbonyl]amino}-l-[bis(2- thienylmethyl)amino]-l-oxohexan-2-yl acetate (3-87).I. (2S)-6-[(Benzyloxy)carbonyl]amino}-2-hydroxyhexanoic acid (3-86).[0191] To a suspension of H-Lys(Z)-OH (2.0 g, 7.2 mmol) in aqueous acetic acid (50% v/v, 140 mL) at 0 °C, a solution of sodium nitrite (3.7 g, 54 mmol) in water (10 mL) cooled to 0 °C was added dropwise. The resulting mixture was stirred 35 minutes, and the cold reaction was extracted with diethyl ether 4 times. The combined organic layers were washed twice with saturated brine, dried over magnesium sulfate, filtered, and concentrated under reduced pressure to give the title compound 3-86. This material contained acetic acid and other impurities in addition to 3-86 but was used in the subsequent reaction without purification.
dibenzyl(((2S,5S)-3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))dicarbamate[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
89%
In diethylene glycol dimethyl ether at 160 - 165℃; for 2.5h;
3
[0034] Example 3[0035] Cbz-lysine (10.0 g), diethylene glycol dimethyl ether (diglyme; 50 mL), and a catalyst were charged to a 250 mL round bottom flask. The mixture was heated to 160 - 165 °C for 2.5 hours. The reaction mixture was poured into water and cooled to ambient temperature overnight. The precipitated solid was isolated by filtration, washed with water, and dried in vacuo at 50 °C.
With sodium carbonate; In 1,4-dioxane; water; at 20℃; for 1h;pH Ca. 10 - 11;
Preparation of Compound 24e A three-necked flask was loaded consecutively with H2O (40 mL), 1,4-dioxane (70 mL) and H-Lys(Z)-OH (10 g, 35.7 mmol). The mixture was stirred until complete dissolution. The pH was adjusted to about 10.5 by adding of 2 M aqueous Na2CO3. Benzyl chloroformate (6.69 g, 39.2 mmol) was added while maintaining the pH at about 10-11 by adding at the same time 2 M aqueous Na2CO3. After completing addition, the reaction mixture was stirred at 20 C for 1 hour. Then EtOAc (50 mL) was added and pH of the resulting mixture was adjusted to 2-3 with c-HCl. The organic layer was separated and the aqueous layer was extracted with EtOAc (50 mL). The combined organic layers were washed with brine (50 mL), and dried over Na2SO4. Filtration and concentration under reduced pressure yielded the compound 24e as yellowish oil (14.7g, 99 %). 1H-NMR (400 MHz, CDC13) delta 7.33-7.27 (m, 10H), 5.07-5.04 (d, 4H), 4.08 (m, 1H), 3.09 (t, 2H), 1.51 (br s, 1H), 1.49 (bs, 1H), 1.47-1.40 (m, 4H).
With sodium hydroxide; In tetrahydrofuran; water; at 0 - 20℃;
General procedure: N-Benzyloxycarbonyl alpha-amino acids 1a-f were prepared followingthe method described by Shi et al.19b After usually workup,all products were using direct in the next step without furtherpurification.
1 Example 1: Preparation of N(ε)-benzyloxycarbonyl-L-lysine-N-ring internal anhydride
Mix 1g of the N-benzyloxycarbonyl-L-lysine and 0.6g of bis(trichloromethyl) carbonate at 25°C, add tetrahydrofuran, and heat to 50°C to react for 2h. After the reaction, the reaction mixture Settling in excess petroleum ether, separating, washing, recrystallizing, and drying to obtain N(ε)-benzyloxycarbonyl-L-lysine-N-ring internal acid anhydride.
6''-amino-6''-deoxy-3,2',6',3'',4'''-penta-N-tert-butoxycarbonylarbekacin[ No CAS ]
C61H103N9O22[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
With 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride In tetrahydrofuran; methanol; water at 20℃; for 24h;
6″-N-Alkanoylamino-6″-deoxy-3, 2′, 6′, 3″, 4‴-penta-N-tert-butoxycarbonylarbekacin(5a- z). Amidation by DMT-MM method (5a- l, s - v, x - z).
General procedure: To asolution of 4 (50- 300mg, 0.05 - 0.29mmol) in MeOH-THF-H2O(15:7.5:1, 17.5 ml), a variety of carboxylic acids (1.5 eq) and DMT-MM(2 eq) were added, and the mixture was stirred at room temperature for 1 day.Concentration gave a residue that was extracted with CHCl3. The organicsolution was washed with saturated aqueous NaHCO3 and water, dried overMgSO4 and concentrated. The resulting residue was purified by silica gelcolumn chromatography to provide 5 (41 - 90%) as a colorless solid.
7 Example 7 CD-BisLys(Z) 7
Example 7 CD-BisLys(Z) 7 (0377) (0378) A 50 mL round bottom flask equipped with a magnetic stirbar and a condenser and a septum was charged with 0.124 g (0.443 mmol) of H-Lysine(Z)-OH and 0.15 g (0.111 mmol) of di-iodo-β-cyclodextrin 2 in 5 mL of degassed 0.1M sodium carbonate solution. The solution mixture was heated at 100° C. for 4 h. The solution was then filtered and the pH of the filtrate is adjusted to 8.5 before dialyzing in MWCO 500 membranes for 24 h. The yield of 7 was 0.124 g (68.9%).
With bis(trichloromethyl) carbonate In tetrahydrofuran at 20 - 50℃; for 2h;
4 Example 4: Preparation of N(ε)-benzyloxycarbonyl-L-lysine-N-ring anhydride
1 g of the N(ε)-benzyloxycarbonyl-L-lysine was mixed with 0.6 g of bis(trichloromethyl) carbonate at 25 ° C, tetrahydrofuran was added, and the mixture was heated to 50 ° C for 2 h. After the reaction is completed, the reaction mixture is allowed to settle in excess petroleum ether, separated, washed, recrystallized, and dried to obtain N(ε)-benzyloxycarbonyl-L-lysine-N-ring anhydride. The nuclear magnetic spectrum is shown in Fig. 2. Fig. 2 is a nuclear magnetic resonance spectrum of N(?)-benzyloxycarbonyl-L-lysine-N-ring anhydride prepared in Example 4.
3-(4-fluorophenyl)prop-2-yn-1-amine hydrochloride[ No CAS ]
benzyl (S)-(5-amino-6-((3-(4-fluorophenyl)prop-2-yn-1-yl)amino)-6-oxohexyl)carbamate hydrochloride[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
71%
Stage #1: N6-[(benzyloxy)carbonyl]-L-lysine; 3-(4-fluorophenyl)prop-2-yn-1-amine hydrochloride With dmap; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride; N-ethyl-N,N-diisopropylamine In dichloromethane at 25℃; Inert atmosphere;
Stage #2: With hydrogenchloride In dichloromethane; ethyl acetate at 0℃; Inert atmosphere;
6-{(S)-6-amino-2-[(S)-2,6-diaminohexanamido]hexanamido}hexanoic acid[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
Stage #1: N6-[(benzyloxy)carbonyl]-L-lysine; methyl (2S)-2-amino-6-[(benzyloxy)carbonyl]amino}hexanoate hydrochloride With 4-methyl-morpholine; benzotriazol-1-ol In dichloromethane at 20℃;
Stage #2: With sodium hydroxide In methanol; ethanol for 0.75h; Further stages;
24.4 Step 4. 6-{(S)-6-amino-2-[(S)-2,6-diaminohexanamido]hexanamido}hexanoic acid
To a solution of H-Lys(Z)-OMe hydrochloride (4.0g, 12.09mmol), Z-Lys(Z)-OH (5.61g, 13,54mmol) and HOBt (1.863g, 13.78mmol) in DCM (50mL) and NMM (1.5mL) at rt was added a suspension of EDC (3.94g, 20.56mmol) and NMM (2mL) in DCM (50mL) over lOmin. After stirring overnight, the reaction mixture was concentrated. To a solution of aforementioned in MeOH/EtOH (v/v = 1/1, lOOmL) was added IN NaOH (15mL). After 45min, the hydrolysis was complete, and the pH of the resulting mixture was adjusted to ~7.0 using HC1. Solids were filtered off, and the filtrate was concentrated. To a solution of the aforementioned (7.0g, 10.34mmol) and 6-(benzyloxy)-6-oxohexan-l- aminium 4-methylbenzenesulfonate (4.88g, 12.41mmol) in DMF (200mL) at rt was add HOBt (1.69g, 12.41mmol), EDC (2.379g, 12.41mmol) and DIPEA (3.66mL, 20.69mmol). After stirring at rt for 4h, to the reaction mixture was added 0 (5-6 times volume of the mixture). After stirring at 0°C for lhr, the precipitate was collected and washed with sufficient 0 through filtration. The solids were dissolved in MeOH, and the resulting solution was partitioned between EtOAC (500mL) and IN HC1 (300mL). The organic layer was washed with saturated NaHC03 (300mL), dried over Na2S04 and concentrated. The residue was purified on 220g silica gel with hexanes/EtOAc 0-100% over 60min. A slurry of the aforementioned (6.0g, 6.82mmol) in IPA (lOOmL) and Pd(OH)2 (957mg, 6.82mmol) was degassed and shaken on a Parr shaker under 344.74kPa of Eh. After overnight, catalyst was filtered off through a pad of CELITE and washed with water. The filtrate was freeze-dried to give the title compound.
With bis(trichloromethyl) carbonate In tetrahydrofuran at 50℃; for 1.66667h; Inert atmosphere;
1.1; 2.1; 3.1; 4.1
(1) Polylactic acid polylysine: 0.25g polylactic acid (Mn=5000) is dissolved in 12.5mL chloroform,Add 0.0288g N-hydroxysuccinimide (NHS),0.048g 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC),React with nitrogen at room temperature for 4h; add 158μL of ethylenediamine, react for 48h;Take the supernatant of the reaction solution and spin-evaporate until it becomes turbid, add 10 mL of anhydrous ether, and centrifuge at 7000 rpm for 10 min.Obtain the precipitate; the precipitate is dialyzed with ultrapure water MWCO3500 for 3 days, freeze-dried,Get amino-terminated polylactic acid (PLA-NH2);1.5g N(ε)-Benzyloxycarbonyl-L-lysine dissolved in 20mL tetrahydrofuran,Nitrogen protection reaction at 50°C for 10min,0.79g triphosgene was dissolved in 10mL tetrahydrofuran and then added dropwise to the N(ε)-benzyloxycarbonyl-L-lysine solution,Continue to react for 1.5h; add to 80mL n-hexane after cooling and let stand for 24h,The precipitate was dried in vacuum at room temperature for 24 hours to obtain benzyloxycarbonyl lysine anhydride (Lys(Z)-NCA);Dissolve 0.03g of amino-terminated polylactic acid (PLA-NH2) into 5mL of N,N-dimethylformamide (DMF),Add 0.1g benzyloxycarbonyl lysine anhydride (Lys(Z)-NCA),40°C nitrogen protection reaction for 72h, dialysis on MWCO3500 for 3 days,Freeze-dried polylactic acid-poly(Nε-benzyloxycarbonyl-L-lysine) (PLA-PZLL);Then take 0.3g PLA-PZLL, add 8mL trifluoroacetic acid, and react in an ice-water bath with nitrogen for 30min;Drop 0.6 mL of a solution of acetic acid (HOAc) with a mass concentration of 33% hydrobromic acid (HBr),Continue the reaction for 1 hour, add 20 mL of anhydrous ether dropwise to the reaction system,Centrifuge at 7000 rpm for 10 min to obtain a precipitate. The precipitate was dialyzed with ultrapure water MWCO3500 for 3 days.Freeze drying to obtain polylactic acid-polylysine polymer (PLA-PLL);
Stage #1: N6-[(benzyloxy)carbonyl]-L-lysine With 1H-imidazole; tris[(1,1,1,3,3,3-hexafluoropropan-2-yl)oxy]silane; cesium fluoride In chloroform at 30℃; for 1h; Inert atmosphere; Sealed tube;
Stage #2: With N-methyl-N-tert-butyldimethylsilyl-1,1,1-trifluoroacetamide In chloroform at 30℃; for 1h; Inert atmosphere; Sealed tube;
Stage #3: Ala-OtBu In chloroform at 30℃; for 6h; Inert atmosphere; Sealed tube;
79; 91 2-ethylbutyl (S)-6-(((benzyloxy)carbonyl)amino)-2-(chloro-l5-azanyl)hexanoate.
4 N hydrochloric acid (5 mL) was added to a solution of N6-((benzyloxy)carbonyl)-L-lysine (1 g, 4 mmol) in 2-ethyl-butanol (10 mL) and the resulting mixture was heated to 70 °C. After 3 h, the reaction mixture was concentrated under reduced pressure at 70 °C. The crude solid residue was taken up into hexanes (150 mL) and was stirred for 4 h. The resulting solid was collected by vacuum filtration to afford the product. 1H NMR (400 MHz, methanol-d4) δ 7.38 - 7.25 (m, 5H), 5.06 (s, 2H), 4.26 - 4.12 (m, 2H), 4.03 (t, J = 6.3 Hz, 1H), 3.13 (t, J = 6.7 Hz, 2H), 2.01 - 1.80 (m, 2H), 1.62 - 1.32 (m, 8H), 0.92 (t, J = 7.5 Hz, 6H).
With hydrogenchloride In water at 70℃; for 3h;
79; 91 2-ethylbutyl (S)-6-(((benzyloxy)carbonyl)amino)-2-(chloro-l5-azanyl)hexanoate.
4 N hydrochloric acid (5 mL) was added to a solution of N6-((benzyloxy)carbonyl)-L-lysine (1 g, 4 mmol) in 2-ethyl-butanol (10 mL) and the resulting mixture was heated to 70 °C. After 3 h, the reaction mixture was concentrated under reduced pressure at 70 °C. The crude solid residue was taken up into hexanes (150 mL) and was stirred for 4 h. The resulting solid was collected by vacuum filtration to afford the product. 1H NMR (400 MHz, methanol-d4) δ 7.38 - 7.25 (m, 5H), 5.06 (s, 2H), 4.26 - 4.12 (m, 2H), 4.03 (t, J = 6.3 Hz, 1H), 3.13 (t, J = 6.7 Hz, 2H), 2.01 - 1.80 (m, 2H), 1.62 - 1.32 (m, 8H), 0.92 (t, J = 7.5 Hz, 6H).
With triethylamine In tetrahydrofuran; lithium hydroxide monohydrate at 20℃; for 18h; Inert atmosphere;
1.4 (4) Synthesis of compound 2-5
Weigh compound 2-4 (940 mg, 2.18 mmol) and raw material N6-Cbz-L-lysine (550 mg, 1.96 mmol) and dissolve them in 40 mL of tetrahydrofuran solution, add triethylamine (220 mg, 2.18 mmol), and stir Water was added dropwise until the raw material was completely dissolved and turned into a clear light yellow transparent solution, protected by argon, stirred at room temperature for 18 hours, followed by TLC of the raw material 2-4 until the reaction was complete, the solvent tetrahydrofuran and water were both spin-dried to obtain a light yellow solid, the column layer was Separation and purification (petroleum ether:ethyl acetate=1:1), 550 mg of light yellow oily liquid product was obtained with a yield of 42%.
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