Liquid-Phase Split-Type Combinatorial Synthesis of Tripeptide Derivatives Encoded by Fluorous Fmoc Reagents

Abstract

A liquid-phase mixture synthesis of 18 tripeptides, some of which are analogues of ACE inhibitors, was effectively conducted by using fluorous Fmoc reagents as encoding tags.

Key words

combinatorial chemistry - liquid-phase synthesis - mixture synthesis - fluorous tag - peptides

• References and Notes


For examples, see:
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For examples of peptides synthesis using fluorous tags, see:
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• 6 Analytical Data for f₁₄-Fmoc-Ala-OH: Pale-yellow solid; mp 95.7–96.5 °C. ¹H NMR (270 MHz, CDCl₃): δ = 1.49 (d, J = 7.0 Hz, 3 H), 2.31–2.51 (m, 4 H), 2.96–3.02 (m, 4 H), 4.16–4.21 (m, 1 H), 4.39–4.48 (m, 3 H), 5.28 (d, J = 7.0 Hz, 1 H), 7.25 (d, J = 7.3 Hz, 2 H), 7.42 (s, 2 H), 7.68 (d, J = 8.1 Hz, 2 H). ¹⁹F NMR (466 MHz, CDCl₃): δ = –80.4 (6F), –115.4 (4F), –127.6 (4F); HRMS [FAB+]: m/z calcd for C₂₈H₂₃NO₄F₁₄: 704.1482; found: 704.1456.
• 7 Analytical Data for f₁₈-Fmoc-Phe-OH: White solid; mp 122.2–123.1 °C. ¹H NMR (270 MHz, CDCl₃): δ = 2.33–2.50 (m, 4 H), 2.94–3.00 (m, 4 H), 3.09–3.25 (m, 2 H), 4.13–4.17 (m, 1 H), 4.33–4.47 (m, 2 H), 4.66–4.73 (m, 1 H), 5.21 (d, J = 7.8 Hz, 1 H), 7.12 (d, J = 6.5 Hz, 2 H), 7.23–7.26 (m, 5 H), 7.39 (d, J = 3.8 Hz, 2 H), 7.67 (d, J = 8.6 Hz, 2 H). ¹⁹F NMR (466 MHz, CDCl₃): δ = –80.8 (6F), –114.7 (4F), –124.2 (4F), –125.9 (4F). HRMS [FAB+]: m/z calcd for C₃₆H₂₇NO₄F₁₈: 880.1731; found: 880.1685.
• 8 Analytical Data for f₂₆-Fmoc-Leu-OH: White solid; mp 134.5–135.4 °C. ¹H NMR (270 MHz, CDCl₃): δ = 0.97 (d, J = 5.4 Hz, 6 H), 1.59–1.76 (m, 3 H), 2.32–2.45 (m, 4 H), 2.96–3.02 (m, 4 H), 4.17–4.22 (m, 1 H), 4.39–4.44 (m, 3 H), 5.11 (d, J = 8.3 Hz, 1 H), 7.24 (d, J = 7.3 Hz, 2 H), 7.43 (s, 2 H), 7.67 (d, J = 8.1 Hz, 2 H). ¹⁹F NMR (466 MHz, CDCl₃): δ = –80.6 (6F), –114.4 (4F), –121.7 (4F), –122.7 (4F), –123.3 (4F), –125.9 (4F). HRMS [FAB+]: m/z calcd for C₃₇H₂₉NO₄F₂₆: 1046.1760; found: 1046.1747.
• 9a Konig W, Geiger R. Chem. Ber. 1970; 103: 788
• 9b Konig W, Geiger R. Chem. Ber. 1970; 103: 2024
• 9c Knorrl R, Trzeciak A, Bannwarth W, Gillessen D. Tetrahedron Lett. 1989; 30: 1927
• 10 Mixture Synthesis for Three-Component Mixture of f₁₄-Fmoc-Ala-Ala-OBn, f₁₈-Fmoc-Phe-Ala-OBn, and f₂₆-Fmoc-Leu-Ala-OBn; Typical Procedure: f₁₄-Fmoc-Ala-OH (560.0 mg, 0.79 mmol), f₁₈-Fmoc-Phe-OH (714.4 mg, 0.79 mmol), and f₂₆-Fmoc-Leu-OH (832.0 mg, 0.79 mmol) were mixed and dissolved in DMF (20 mL). To the solution were added HOBt·H₂O (438.8 mg, 2.86 mmol) and HBTU (1086 mg, 2.86 mmol), separately. After stirring for 5 min, Ala-OBn (1.01 g, 2.86 mmol) and DIPEA (974 μL, 5.72 mmol) were added to the above reaction mixture separately. The reaction mixture was stirred for 24 h at room temperature. After the addition of aq 1.0 M HCl and then dilution with ethyl acetate, the organic layer was washed with H₂O, sat. aq NaHCO₃ and brine, dried over Na₂SO₄ and concentrated. The crude residue was purified by silica gel chromatography (CHCl₃–MeOH, 20:1) to give the title compound (2.4 g, 95% based on the average molecular weight of the mixture).
• 11 Wuts PG. M, Greene TW. Green’s Protective Groups in Organic Synthesis . Wuts PG. M, Greene TW. Wiley-Interscience; New Jersey: 2007. 3rd ed., 415
• 12 Analytical and preparative f-HPLC were conducted by using FluoroFlash® columns purchased from Fluorous Technologies Inc. The corporation is no longer trading; however, fluorous column (Wakopak® Fluofix-II 120E) with almost the same separation ability is available from Wako Pure Chemical Industries, Ltd.
• 13 The f-HPLC separation could also be applied to a tripeptide that was derived from hydrophilic amino acids such as glycine. The f₁₄-Fmoc-Gly-Gly-Gly-OMe, f₁₈-Fmoc-Gly-Gly-Gly-OMe and f₂₆-Fmoc-Gly-Gly-Gly-OMe appeared at 2.4, 3.0, and 6.4 min, respectively, in the analytical f-HPLC trace under the same conditions.
• 14 Analytical Data for f₁₄-Fmoc-Ala-Ala-Ala-OBn: White solid; mp 153.8–154.6 °C. ¹H NMR (270 MHz, CDCl₃): δ = 1.36–1.42 (m, 9 H), 2.31–2.44 (m, 4 H), 2.96–3.02 (m, 4 H), 4.16–4.19 (m, 1 H), 4.22–4.28 (m, 1 H), 4.39–4.50 (m, 3 H), 4.56–4.61 (m, 1 H), 5.13 (d, J = 11.9 Hz, 1 H), 5.19 (d, J = 12.7 Hz, 1 H), 5.44 (d, J = 7.3 Hz, 1 H), 6.55–6.62 (m, 2 H), 7.24 (d, J = 9.9 Hz, 2 H), 7.31–7.36 (m, 5 H), 7.42 (s, 2 H), 7.68 (d, J = 8.1 Hz, 2 H). ¹⁹F NMR (466 MHz, CDCl₃): δ = –80.3 (6F), –115.4 (4F), –127.6 (4F). HRMS [FAB+]: m/z calcd for C₄₁H₄₀N₃O₆F₁₄: 936.2694; found: 936.2704.
• 15 Analytical Data for f₁₈-Fmoc-Phe-Ala-Ala-OBn: White solid; mp 171.5–172.4 °C. ¹H NMR (270 MHz, CDCl₃): δ = 1.30 (d, J = 6.8 Hz, 3 H), 1.40 (d, J = 7.3 Hz, 3 H), 2.34–2.44 (m, 4 H), 2.95–3.02 (m, 4 H), 3.07–3.11 (m, 2 H), 4.10–4.15 (m, 1 H), 4.30–4.46 (m, 4 H), 4.52–4.58 (m, 1 H), 5.13 (d, J = 11.9 Hz, 1 H), 5.19 (d, J = 12.7 Hz, 1 H), 5.35 (d, J = 6.8 Hz, 1 H), 6.38 (d, J = 5.9 Hz, 1 H), 6.45 (d, J = 7.0 Hz, 1 H), 7.13–7.16 (m, 2 H), 7.23–7.40 (m, 12 H), 7.68 (d, J = 7.8 Hz, 2 H). ¹⁹F NMR (466 MHz, CDCl₃): δ = –80.6 (6F), –114.5 (4F), –124.1 (4F), –125.8 (4F); HRMS [FAB+]: m/z calcd for C₅₂H₄₅N₃O₅F₁₈: 1112.2943; found: 1112.2969.
• 16 Analytical Data for f₂₆-Fmoc-Leu-Ala-Ala-OBn: White solid; mp 179.9–181.0 °C. ¹H NMR (270 MHz, CDCl₃): δ = 0.94 (d, J = 5.4 Hz, 6 H), 1.36–1.42 (m, 6 H), 1.50–1.70 (m, 3 H), 2.35–2.48 (m, 4 H), 2.96–3.02 (m, 4 H), 4.15–4.20 (m, 2 H), 4.36–4.59 (m, 3 H), 4.45–4.59 (m, 1 H), 5.16–5.22 (m, 3 H), 6.48 (d, J = 6.5 Hz, 2 H), 7.32–7.43 (m, 9 H), 7.68 (d, J = 7.2 Hz, 2 H). ¹⁹F NMR (466 MHz, CDCl₃): δ = –80.6 (6F), –114.4 (4F), –121.6 (4F), –122.6 (4F), –123.2 (4F), –125.9 (4F). HRMS [FAB+]: m/z calcd for C₅₂H₄₆N₃O₆F₂₆: 1278.2971; found: 1278.3009.
• 17 Analytical data for these tripeptides are provided in the Supporting Information
• 18a Perich JW, Johns RB. Tetrahedron Lett. 1988; 29: 2369
• 18b Wuts PG. M, Greene TW. Green’s Protective Groups in Organic Synthesis . Wuts PG. M, Greene TW. Wiley-Interscience; New Jersey: 2007. 3rd ed., 506