* 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, 1957, vol. 79, p. 4686,4689
[2] Bioorganic and Medicinal Chemistry Letters, 2011, vol. 21, # 24, p. 7355 - 7358
[3] Journal of Medicinal Chemistry, 2013, vol. 56, # 6, p. 2374 - 2384
2
[ 13139-15-6 ]
[ 36077-41-5 ]
Reference：
[1] Russian Journal of Organic Chemistry, 2015, vol. 51, # 5, p. 615 - 618[2] Zh. Org. Khim., 2015, vol. 51, # 5, p. 637 - 640,4
3
[ 13139-15-6 ]
[ 32991-17-6 ]
Reference：
[1] Helvetica Chimica Acta, 1994, vol. 77, # 4, p. 1124 - 1165
[2] Bulletin of the Chemical Society of Japan, 1983, vol. 56, # 5, p. 1527 - 1531
[3] Bioorganic and Medicinal Chemistry, 2014, vol. 22, # 11, p. 3055 - 3064
4
[ 13139-15-6 ]
[ 27532-96-3 ]
[ 32991-17-6 ]
Reference：
[1] Angewandte Chemie - International Edition, 2018, vol. 57, # 30, p. 9456 - 9460[2] Angew. Chem., 2018, vol. 130, # 30, p. 9600 - 9604,5
5
[ 13139-15-6 ]
[ 74-88-4 ]
[ 53363-89-6 ]
Yield
Reaction Conditions
Operation in experiment
98%
Stage #1: With sodium hydride In tetrahydrofuran; mineral oil at 0℃; for 0.5 h; Inert atmosphere Stage #2: at 0 - 20℃; for 4.5 h; Inert atmosphere
To a solution of Boc-L-Leu (5.0 g, 21.6mmol) in THF (73 mL) wasadded NaH (60percent dispersion in mineral oil, 10.0 g, 0.13 mol) inportions at 0 oC. After stirring at 0 oC for 30 min, MeI (4.1 mL,66.0 mmol) and DMF (3.7 mL) were successively added. Thereaction was stirred for 30 min at 0 oC and additional 4 h atrt, until it was quenchedwith H2O (50 mL). The mixture was extracted by EtOAc (2×100 mL). The aqueoussolution was then acidified with 10percent aqueous KHSO4 solution to pH 5 and extractedwith ethyl acetate. The combined EtOAc extracts were washed with 5percent Na2S2O4solution and brine, and dried over Na2SO4, filtered and evaporated to give 13 (5.2 g,98percent yield) as a colorless oil.
95%
With sodium hydride In tetrahydrofuran at 0 - 20℃; for 26 h; Inert atmosphere
General procedure: (S)-2-(Tert-butoxycarbonyl(methyl)amino)-3-hydroxypropanoic acid(6a) Gummy substance (This compound was prepared byadding neat sodium hydride (10 equiv.) in portion wiseover a period of 2.0 h to a cooled (0 °C) solution of (S)-2-(tert-butoxycarbonylamino)-3-hydroxypropanoic acid (1equiv.) and iodomethane (10 equiv.) in dry THF under astream of nitrogen. The reaction mixture was stirred at room temperature for 24 h under nitrogen atmosphere andthen diluted with ether (20 mL) and quenched with water(30 mL). The layers were separated and the aqueous layerwas extracted with ether (2 x9 15 mL), acidified to pH 3with a 20 percent aqueous solution of citric acid and extractedwith EtOAc (3 x 20 mL). The combined organic phasewas dried over Na2SO4 and evaporated to afford thecorresponding N-methylated product in 90 percent yield asGummy substance.)
94%
Stage #1: With sodium hydride In tetrahydrofuran at 0℃; for 1 h; Inert atmosphere Stage #2: at 0 - 20℃; Inert atmosphere
Boc—Leu—OH (4.7 g, 30 mmol) was dissolved in 150 mL of THF in an inert atmosphere. Sodium hydride (3.2 g, 133 mmol, 4.0 equiv.) was added at 0°C in portions. The reaction mixture was agitated 1 hour at low temperature before adding iodomethane (14.2 g, 100 mmol, 5.0 equiv.). The reaction was left under agitationovernight at ambient temperature and then neutralised with 100 mL of water and washed 3 times with 200 mL of EtOAc. The aqueous phase was brought to pH 3 with iN HC1 and then extracted 3 times with 100 mL of EtOAc. The organic phases were combined, washed once with 300 mL of NaC1—saturated aqueous solution, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure yielding 4.7g (94 percent) of compound 52A in the form of a red oil.
Reference：
[1] Tetrahedron Letters, 2014, vol. 55, # 44, p. 6109 - 6112
[2] Helvetica Chimica Acta, 1994, vol. 77, # 4, p. 1124 - 1165
[3] Medicinal Chemistry Research, 2016, vol. 25, # 6, p. 1148 - 1162
[4] Patent: WO2014/174060, 2014, A1, . Location in patent: Page/Page column 112
[5] International Journal of Molecular Sciences, 2017, vol. 18, # 3,
[6] Bioscience, Biotechnology, and Biochemistry, 1994, vol. 58, # 6, p. 1193 - 1194
[7] Tetrahedron, 2006, vol. 62, # 2-3, p. 264 - 284
[8] Canadian Journal of Chemistry, 1977, vol. 55, p. 906 - 910
[9] Tetrahedron, 1991, vol. 47, # 29, p. 5453 - 5462
[10] Tetrahedron Letters, 1998, vol. 39, # 40, p. 7373 - 7376
[11] Current Medicinal Chemistry, 2013, vol. 20, # 9, p. 1183 - 1194
[12] Journal of Organic Chemistry, 2014, vol. 79, # 17, p. 8491 - 8497
6
[ 13139-15-6 ]
[ 77-78-1 ]
[ 53363-89-6 ]
Reference：
[1] Organic letters, 2003, vol. 5, # 2, p. 125 - 128
[2] Indian Journal of Chemistry, Section B: Organic Chemistry Including Medicinal Chemistry, 1995, vol. 34, # 1, p. 45 - 47
(S)-2-[(S)-3-(4-Amino-phenyl)-2-((S)-2-amino-propionylamino)-propionylamino]-4-methyl-pentanoic acid [(S)-1-((S)-1-carbamoyl-4-guanidino-butylcarbamoyl)-3-methyl-butyl]-amide[ No CAS ]
(S)-2-[(S)-2-((S)-2-Amino-propionylamino)-4-phenyl-butyrylamino]-4-methyl-pentanoic acid [(S)-1-((S)-1-carbamoyl-4-guanidino-butylcarbamoyl)-3-methyl-butyl]-amide[ No CAS ]
(S)-2-[(S)-2-((S)-2-Amino-3-hydroxy-propionylamino)-3-(4-amino-phenyl)-propionylamino]-4-methyl-pentanoic acid [(S)-1-((S)-1-carbamoyl-4-guanidino-butylcarbamoyl)-3-methyl-butyl]-amide[ No CAS ]
(S)-2-[(S)-2-((S)-2-Amino-3-hydroxy-propionylamino)-4-phenyl-butyrylamino]-4-methyl-pentanoic acid [(S)-1-((S)-1-carbamoyl-4-guanidino-butylcarbamoyl)-3-methyl-butyl]-amide[ No CAS ]
(S)-2-((S)-3-(4-Amino-phenyl)-2-{(S)-3-(4-fluoro-phenyl)-2-[(E)-(3-phenyl-acryloyl)amino]-propionylamino}-propionylamino)-4-methyl-pentanoic acid ((S)-1-carbamoyl-4-guanidino-butyl)-amide[ No CAS ]
(S)-2-((S)-2-{(S)-3-(4-Fluoro-phenyl)-2-[(E)-(3-phenyl-acryloyl)amino]-propionylamino}-4-phenyl-butyrylamino)-4-methyl-pentanoic acid ((S)-1-carbamoyl-4-guanidino-butyl)-amide[ No CAS ]
A round-bottom flask was charged with Boc-Ala-OH (1.0 g, 5.3 mmol), HOBt (0.72 g, 5.3 mmol, 1.0 eq) and HBTU (3.0 g, 7.9 mmol, 1.5 eq) under Ar(g). To this was added anhydrous DMF (5 mL) and DIEA (2.7 mL, 15.8 mmol, 3 eq.). The reaction mixture was allowed to stir at room temperature for 15 min upon which the solution turned to a pale yellow. A solution of 3-butenylamineHCl (0.85 g, 7.9 mmol, 1.5 eq) in DMF (2 mL) was added and the reaction mixture heated to 50 C. and allowed to stir for 1 h. The solution was cooled to room temperature and H2O (20 mL) was added, followed by CH2C12 (50 mL). The organic layer was removed and the aqueous layer was extracted with CH2C12 (5*50 mL). The combined organic layers were washed with brine (5*50 mL), and dried over Na2SO4. The solvent was removed in vacuo and the crude residue was purified by flash chromatography (SiO2, 0% to 50% EtOAc in hexanes) to provide 1.16 g (91%) of 3 as a white solid: 1H NMR (300 MHz, CDCl3) delta 6.51 (bs, 1H), 5.72 (ddt, J=17.1, 10.2, 6.8 Hz, 1H), 5.22 (d, J=7.7 Hz, 1H), 5.12-4.96 (m, 2H), 4.12 (q, J=7.6 Hz, 1H), 3.38-3.18 (m, 2H), 2.22 (qt, J=6.9, 1.3 Hz, 2H), 1.40 (s, 9H), 1.31 (d, J=7.0 Hz, 3H); 13C NMR (126 MHz, CDCl3) delta 172.71, 155.48, 135.04, 117.09, 79.83, 50.02, 38.41, 33.67, 28.30 (3C), 18.64. HRMS (ESI) m/z calcd for C12H22N2O3 [M+H]+: 243.1630. found 243.1626.
1-(4-fluorophenyl)-2-methyl-2-propyl 2-[(N-Boc)amino]-4-methylpentanoate[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
0.14 g (29%)
With sodium hydrogencarbonate; In dicyclohexylcarbodiimide (DCC); benzene;
Example 20 1-(4-Fluorophenyl)-2-methyl-2-propyl (S)-2-amino-4-methylpentanoate. N-tert-Butoxycarbonyl-L-leucine (N-Boc-L-leucine) (0.3 g, 1.3 mmol) 1-(4-fluoro-2-methyl-2-propanol (0.39 g, 2.6 mmol) and <strong>[823-39-2]dimethyl aminopyridine</strong> (0.015 g, 0.13 mmol), were dissolved in 3 ml of benzene. The solution was cooled in an ice bath and dicyclohexylcarbodiimide (DCC) (0.28 g, 1.35 mmol) was added under stirring. The solution was stirred during 48 h at room temperature. Dicyclohexylurea was removed by filtration and washed with benzene (2x5 ml). The filtrate was washed sucessively with HCl (0.5 M, 10 mlx3), NaHCO3 (5 percent, 10 mlx3) and water (10 mlx2). The organic phase was dried and the solvent was evaporated. The resulting ester (0.24 g) was purified by flash chromatography with hexane: ethylacetate 9:1 as eluent to yield 0.14 g (29 percent) of 1-(4-fluorophenyl)-2-methyl-2-propyl 2-[(N-Boc)amino]-4-methylpentanoate. 1H NMR (200 MHz CDCl3) delta ppm 7.15(dd, 2, 3,5-phenyl, J=8.9 Hz, 5.4 Hz), 6.97(t, 2, 2,6-phenyl, J=8.8 Hz), 4.85(d, 1, NH, J=8 Hz), 4.18(m, 1, CCH), 3.09, 2.93(AB, 2, benzylic, J=15 Hz), 1.62(m, 3, CH2CH), 1.46(s, 3, CCH3) 1.45(s, 9, (CH3)3), 1.42(s, 3, CCH3), 0.93(d, 2, CH CH 3, J=5.4 Hz), 0.89(d, 2, CH CH 3, J=5.6 Hz).
With (benzotriazo-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate; N-ethyl-N,N-diisopropylamine; In N,N-dimethyl-formamide; for 48h;
Structure 2 This structure was created by using a step-by-step synthesizing process. First Boc-Leu-Pro-OMe was synthesized using the following method. Proline methyl ester hydrochloride (322 mg, 2 mmol) and Boc-Leu-OH (2 mmol, 498 mg) dissolved in DMF (20 mL) and BOP (2 mmol, 884 mg) added with stifling. DIEA (14 mmol, 2.43 mL) was added dropwise and mixture stirred for 48 h. Reaction mixture quenched by addition to water (200 mL) and extracted with ethyl acetate (4×100 mL). Combined organic phases washed with water (3×150 mL), 2N hydrochloric acid (100 mL), saturated sodium bicarbonate solution (60 mL) and brine (60 mL), then dried over sodium sulfate. Evaporation of solvent gave a colorless oil (596 mg, 87%) which was used without further purification.
{(S)-2-Amino-1-[(S)-1-((S)-1-{(1S,2R)-1-benzyl-2-hydroxy-2-[3-(5-thioxo-4,5-dihydro-[1,2,4]oxadiazol-3-yl)-phenylcarbamoyl]-ethylcarbamoyl}-3-methyl-butylcarbamoyl)-2-methyl-propylcarbamoyl]-ethyl}-carbamic acid tert-butyl ester[ No CAS ]
{(S)-2-Amino-1-[(S)-1-((S)-1-{(1S,2R)-1-benzyl-2-hydroxy-2-[3-(5-oxo-4,5-dihydro-[1,2,4]oxadiazol-3-yl)-phenylcarbamoyl]-ethylcarbamoyl}-3-methyl-butylcarbamoyl)-2-methyl-propylcarbamoyl]-ethyl}-carbamic acid tert-butyl ester[ No CAS ]
General procedure: Boc-D-homophenylalanine (0.43 g, 1.5 mmol), TBTU (0.49 g, 1.54 mmol), DIEA (0.36 mL, 2.8 mmol) and dimethylformamide (DMF, 7 mL) were mixed in a round bottom flask at 0 C. After 10 minutes, a solution of 4-amino-7-chloroquinoline (3) in DMF (7 mL) was added and the reaction was allowed to proceed for one day at room temperature. Following, 14 mL of water were added to the reaction mixture and the product was extracted with dichloromethane (3×14 mL); the organic layers were pooled and sequentially washed with 1% aq. HCl (3×14 mL) and 5% aq. Na2CO3 (3×14 mL). Finally, the organic layer was dried over anhydrous Na2SO4, filtered, and evaporated to dryness under reduced pressure. The crude product was purified by column liquid chromatography on silica, using dichloromethane:acetone (DCM/Me2CO) 6:1 v/v as eluent. The resulting product was obtained as a white solid and corresponded to the target compound (4) (218 mg, 35%); Compound 4 (0.13 g, 0.3 mmol) was reacted with neat trifluoroacetic acid (TFA, 1 mL) for 2 at room temperature, under constant stirring. Then, the pH was adjusted to 12 with 30% aq. Na2CO3 and the solution extracted with DCM. The organic layer was dried with anhydrous Na2SO4 and evaporated to dryness under reduced pressure. Compound 5 was thus obtained as a yellow oil (85.3 mg, 85%); Boc-D-leucine (0.13 g, 0.38 mmol), PyBOP (0.22 g, 0.42 mmol), DIEA (0.162 mL, 1.25 mmol) and DCM (2 mL) were mixed in a round bottom flask at 0 C and put under stirring for 20 minutes. Then, a solution of 5 in DCM (2 mL) was added and the reaction allowed to proceed for one day at room temperature. Following, the reaction mixture was diluted with 14 mL of DCM and sequentially washed with 1% aq. HCl (3×18 mL) and 5% aq. Na2CO3 (3×18 mL). Finally, the organic layer was dried with anhydrous Na2SO4, filtered, and evaporated to dryness. The crude product was purified by column liquid chromatography on silica, using DCM/Me2CO 6:1 v/v as eluent. The target compound, 6, was isolated as a yellowish oil (122 mg, 58%); Compound 6 (0.122 g, 0.22 mmol) was reacted with neat trifluoroacetic acid (TFA, 1 mL) for 2 at room temperature, under constant stirring. Then, the pH was adjusted to 12 with 30% aq. Na2CO3 and the solution extracted with DCM. The organic layer was dried with anhydrous Na2SO4 and evaporated to dryness under reduced pressure. Compound 7 was thus obtained as a yellowish oil (72 mg, 87%); The relevant cinnamic acid (1.1 eq), PyBOP (1.1 eq), DIEA (2 eq.) and DCM (2 mL) were mixed in a round bottom flask and put under stirring for 20 min. Then, a solution of 7 in DCM (2 mL) was added and the reaction allowed to proceed for three days. Precipitation was observed and the precipitate was collected by suction filtration, washed with ice-cold DCM, dried and identified as the pure target compound, except in the case of 8l (p-nitrocinnamic acid derivative), which was further submitted to liquid chromatography on silica, using DCM/Me2CO 6:1 (v/v) as eluent. The synthesis details, analytical and spectroscopic data of intermediate compounds 3 to 7 and compound 10a are given in detail in SI.
With (benzotriazo-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate; N-ethyl-N,N-diisopropylamine; In dichloromethane; at 20℃;
General procedure: To a solution of pyrrole amino-ester 11 (1equiv) in DCM, DIEA (2equiv), the appropriate Boc-protected amino acid (1.2equiv) and BOP (1.2equiv) are added successively. The mixture was stirred for 3-4h at room temperature. The reaction was monitored by HPLC. Once the starting material has completely disappeared, the DCM was evaporated to give a yellow solid, which was dissolved in AcOEt:water 1:1. The organic layer was washed with KHSO4 1M (2×10mL), saturated NaHCO3 (2×20mL) and brine, dried over anhydrous Na2SO4 and evaporated under vacuum to give a yellow solid. The crude product was purified by silica gel flash chromatography using methanol and dichloromethane as eluents.
2-((1-(4,4-dimethyl-2,6-dioxocyclohexylidene)ethyl)amino)hexadecanoic acid[ No CAS ]
Ac-KLIPNASLIENCTKAELK(Ac-KQAEDKVKASREAKKQVEKALEQLEDKVK)-SS-(2-amino-D,L-hexadecanoyl)-2-amino-D,L-hexadecanamide[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
12.5%
General procedure: LCP 1 was synthesised by coupling Dde-C16 onto pMBHA resin(0.45 mmol/g; 0.2 mmol scale) by using HBTU (4.0 equiv) andDIPEA (6.2 equiv) in DMF (2 1 h). Removal of the Dde-protectinggroup was performed by treatments with 2% hydrazine in DMF(2 30 min). The remaining LCP sequence was synthesised bymicrowave-assisted Boc-SPPS. Amino acid activation was achievedby dissolving Boc-amino acids (4.2 equiv) in 0.5 M HBTU(4.0 equiv) in DMF, followed by the addition of DIPEA (6.2 equiv).The amino acid coupling cycle consisted of Boc-deprotection withneat TFA at room temperature (2 1 min), 2 min DMF flow wash,followed by 10 min coupling with the pre-activated amino acid.The temperature was set at 70 C (35 W, 10 min) for all aminoacids, except for Cys and His, which were coupled at 50 C (35 W,10 min). N-terminal acetylation and final cleavage from resin wasperformed as reported previously.29 The lyophilised products werepurified by RP-HPLC. The collected fractions were analysed byESI-MS and RP-HPLC. Where appropriate, fractions were combinedand lyophilised to yield pure product.For the synthesis of LCP 2, Dde-C20 was coupled onto pMBHAresin (0.45 mmol/g; 0.2 mmol scale). The remaining proceduresfollowed that of the synthesis of LCP 1, as mentioned above.LCP 1 yield: 12.5%. Purity: 90%. Molecular weight: 6086.4. ESIMS:[M+4H]4+ m/z 1523.4 (calcd: 1522.6), [M+5H]5+ m/z 1218.4(calcd: 1218.3), [M+6H]6+ m/z 1015.7 (calcd: 1015.3), [M+7H]7+m/z 870.6 (calcd: 870.4), [M+9H]9+ m/z 677.5 (calcd: 677.3), [M+10H]10+ m/z 608.8 (calcd: 609.5). Rt = 25.0 min (0-100% solventB, 40 min, C4-column).
Ac-KLIPNASLIENCTKAELK(Ac-KQAEDKVKASREAKKQVEKALEQLEDKVK)-SS-(2-amino-D,L-eicosanoyl)-2-amino-D,L-eicosanamide[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
18.5%
General procedure: LCP 1 was synthesised by coupling Dde-C16 onto pMBHA resin(0.45 mmol/g; 0.2 mmol scale) by using HBTU (4.0 equiv) andDIPEA (6.2 equiv) in DMF (2 1 h). Removal of the Dde-protectinggroup was performed by treatments with 2% hydrazine in DMF(2 30 min). The remaining LCP sequence was synthesised bymicrowave-assisted Boc-SPPS. Amino acid activation was achievedby dissolving Boc-amino acids (4.2 equiv) in 0.5 M HBTU(4.0 equiv) in DMF, followed by the addition of DIPEA (6.2 equiv).The amino acid coupling cycle consisted of Boc-deprotection withneat TFA at room temperature (2 1 min), 2 min DMF flow wash,followed by 10 min coupling with the pre-activated amino acid.The temperature was set at 70 C (35 W, 10 min) for all aminoacids, except for Cys and His, which were coupled at 50 C (35 W,10 min). N-terminal acetylation and final cleavage from resin wasperformed as reported previously.29 The lyophilised products werepurified by RP-HPLC. The collected fractions were analysed byESI-MS and RP-HPLC. Where appropriate, fractions were combinedand lyophilised to yield pure product.For the synthesis of LCP 2, Dde-C20 was coupled onto pMBHAresin (0.45 mmol/g; 0.2 mmol scale). The remaining proceduresfollowed that of the synthesis of LCP 1, as mentioned above.LCP 1 yield: 12.5%. Purity: 90%. Molecular weight: 6086.4. ESIMS:[M+4H]4+ m/z 1523.4 (calcd: 1522.6), [M+5H]5+ m/z 1218.4(calcd: 1218.3), [M+6H]6+ m/z 1015.7 (calcd: 1015.3), [M+7H]7+m/z 870.6 (calcd: 870.4), [M+9H]9+ m/z 677.5 (calcd: 677.3), [M+10H]10+ m/z 608.8 (calcd: 609.5). Rt = 25.0 min (0-100% solventB, 40 min, C4-column).
The titled peptide was synthesized on a model 430A peptide synthesizer (Applied Rio systems, Foster City, Calif., U.S.A.) which was modified to do accelerated Hoc-chemistry solid phase peptide synthesis (Schnolzer, M. et al., mt. J Peptide Protein Res., (1992), 40:180). 4-Methylbenzhydry- lamine (MHHA) resin (Peninsula, Helmont, Calif., U.S.A.), with a substitution of0.91 mmol/g was used. Hoc amino acids (Midwest Hio-Tech, Fishers, Ind., U.S.A.; Novabiochem., San Diego, Calif., U.S.A.) were used with the following side chain protection: Hoc-Ala-OH, Hoc-Arg(Tos)-OH, Hoc-His (DNP)-OH, Hoc-Val-OH, Hoc-Ecu-OH, Hoc-Gly-OH, HocGln-OH, Hoc-Eys(2C1Z)?--OH, Hoc-Ser(Hzl)-OH, Hoc-PheOH, Hoc-Glu(OcHex)-OH and Hoc-Pro-OH. Fmoc-Glu (OtHu)-OH (Novabiochem, San Diego, Calif., U.S.A.) was used for the residue at the 3rd position in the sequence. The synthesis was carried out on a 0.25 mmol scale. The Hoc groups were removed by two treatments with 100percent TFA each lasting one minute. Hoc amino acids (2.5 mmol) were preactivated with HH11J (2.0 mmol) and DIEA (1.0 mE) in 4 mE of DMF and were coupled without prior neutralization of the peptide-resin TFA salt. Coupling times were 5 minutes. At the end of the assembly of the first 25 residues on theAHI 430A® peptide synthesizer and before the coupling of Fmoc-Glu (OtHu)-OH, the protected peptide-resin was transferred into a reaction vessel on a shaker for manual synthesis. After removing the Hoc protecting group with two, one-minute treatments with 100percent TFA and a washing with DMF, the resin was mixed with Fmoc-Glu(OtHu)-OH (2.5 mmol) which was preactivated with HHTU (2.0 mmol), HOHt (2.0 mmol) and DIEA (1.0 mE) in 4 mE of DMF. The mixture was shaken for 2 hours. This coupling step was repeated. After washing with DMF, the resin was treated with a TFA solution containing 5percent water and 5percent TIS for 2 hours to remove the tHu protecting group in the side chain of the Glu residue. The resin was neutralized with 10percent DIEA in DMF and washed with DMF and DCM. The resin was then treated twice with hexylamine (2.0 mmol), DIC (2.0 mmol), HOHt (2.0 mmol) in 5 ml of DCM for two hours per treatment. The resin was washed with DMF and treated with 25percent piperidine in DMF for 30 minutes to remove the Fmoc protecting group. Afier washing with DMF and DCM, the resin was transferred into the reaction vessel on the AHI 430A peptide synthesizer for the assembly of the rest two residues. At the end of the assembly of the whole peptide chain, the resin was treated with a solution of 20percent mercaptoethanol/10percent DIEA in DMF for 2x30 mm to remove the DNP group on the His side chain. The N-terminal Hoc group was then removed by two treatments of 100percent TFA for 2 minutes. The peptide-resin was washed with DMF and DCM and dried under reduced pressure. The final cleavage was done by stirring the peptide-resin in 10 mE of HF containing 1 mE of anisole and dithiothreitol (50 mg) at 0° C. for 75 minutes. HF was removed by a flow of nitrogen. The residue was washed with ether (6x 10 mE) and extracted with 4N HOAc (6x10 mE). This crude product was purified on a reverse-phase preparative HPEC using a colunm (4x43 cm) of C18 DYNAMAX-100A°® (Varian, Walnut Creek, Calif., U.S.A.). The column was eluted with a linear gradient from 75percentAand25percent B to 55percentAand45percent B at flow rate of 10 mE/mm in an hour where A was 0.1percent TFA in water and B was 0.1percent TFA in acetonitrile. Fractions were collected and checked on an analytical HPEC. Those containing pure product were combined and lyophilized to dryness. 31.8 mg of a white solid was obtained. Purity was 89percent based on analytical HPEC analysis. Electro-spray ionization mass spectrometry (ESI MS) analysis gave the molecular weight at 3368.4 (in agreement with the calculated molecular weight of 3368.9).
With 4-methyl-morpholine; benzotriazol-1-ol; 1-ethyl-(3-(3-dimethylamino)propyl)-carbodiimide hydrochloride; In tetrahydrofuran; at 0 - 20℃; for 12h;Inert atmosphere;
General procedure: A mixture of L-valine p-TsOH salt 10a (100 mg, 0.30 mmol) and N-Boc valine (87.12 mg, 0.30 mmol) was taken in 2.5 mL of dry THF and cooled to 0 C under nitrogen atmosphere. After stirring, freshly distilled NMM (0.16 mL, 1.51 mmol) was added via syringe followed by HOBt (46.61 mg, 0.34mmol) and EDCI (65.18 mg, 0.34 mmol) in a single portion. The mixture was stirred overnight slowly warming to room temperature. After 12 hours, it was concentrated by a rotavapor and the residue was taken in EtOAc (20.0 mL), washed with saturated NaHCO3 solution followed by brine. The organic layer was dried over MgSO4, filtered, and then the solvent was evaporated under reduced pressure. The residue was purified by flash column chromatography using hexane-EtOAc (8:2) to afford the title compound 11a
NH<SUB>2</SUB>-(CH<SUB>2</SUB>)<SUB>10</SUB>-CO-Ile-Leu-Pro-D-Phe-Lys-D-Phe-Pro-D-Phe-D-Phe-Pro-D-Phe-Arg-Arg-NH2[ No CAS ]
Yield
Reaction Conditions
Operation in experiment
The synthesis, unblocking, and purification of peptide (2) were conducted using the same method as that for the peptide in Example 1. To protect the Nomega-function of arginine, the nitro group from the omega-amino group of lysine, the chorocarbobenzoxy group, was used. To synthesize the peptide, 0.2 g of N-tert-butyloxycarbonyl-Nomega-nitroarginine methylbenzhydryl amino polymer and 10 ml of dimethylformamide were added to a reaction vessel. The content of the arginine derivative was 1.0 mmol/g of the polymer. (0025) Upon completion of the polypeptide chain assembly, the final unblocking of the protected peptidyl-polymer with simultaneous cleavage of the peptide from the polymer with liquid hydrogen fluoride were conducted. Purification was conducted with an acetonitrile gradient of 10-90% in 0.1% trifluoroacetic acid. According to the optical density data, the content of the main substance, peptide NH2-(CH2)10-CO-Ile-Leu-Pro-D-Phe-Lys-D-Phe-Pro-D-Phe-D-Phe-Pro-D-Phe-Arg-Arg-NH2 was at least 99%. The amino acid composition of the peptide and its molecular weight corresponded to the theoretical values.
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