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Synlett 2004(14): 2591-2593  
DOI: 10.1055/s-2004-834806
LETTER
© Georg Thieme Verlag Stuttgart · New York

Synthesis of New 2,2,5,5-Tetramethyl-2,5-dihydro-1H-pyrrol-1-yloxyl Radicals and 2-Substituted-2,5,5-trimethylpyrrolidin-1-yloxyl Radicals Based α-Amino Acids

Mária Baloga, Tamás Kálaia, József Jekőb, Heinz-Jürgen Steinhoffc, Martin Engelhardd, Kálmán Hideg*a
a Institute of Organic and Medicinal Chemistry, University of Pécs, P. O. Box 99, 7602 Pécs, Hungary
Fax: +36(72)536219; e-Mail: kalman.hideg@aok.pte.hu;
b ICN Hungary Ltd., P. O. Box 1, 4440 Tiszavasvári, Hungary
c Department of Physics, University of Osnabrück, Barbara Str. 7, 49069 Osnabrück, Germany
d Max Planck Institute of Molecular Physiology, P.O. Box 50 02 47, 44202 Dortmund, Germany
Further Information

Publication History

Received 17 August 2004
Publication Date:
20 October 2004 (online)

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Abstract

Unnatural paramagnetic α-amino acids with 2,2,5,5-tetramethyl-2,5-dihydro-1H-pyrrol-1-yloxyl-3-yl radical or 2,5,5-trimethylpyrrolidin-1-yloxyl-2-yl radical side-chains, including a lysine mimic azido precursor and their derivatives, are described. The new set of paramagnetic amino acids presented in this work with different (polar, nonpolar, aliphatic, aromatic, etc.) side-chains offers a useful tool for the ESR study of the protein structure and function after incorporation, fulfilling diverse structural requirements.

Key words

amino acids - azides - free-radicals - O’Donnell synthesis - protecting groups

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Synthesis of 1d: To a stirred solution of 3,4-bis(bromomethyl)-2,2,5,5-tetramethyl-2,5-dihydro-1H-pyrrol-1-yloxyl radical (326 mg, 1.0 mmol) in acetone (10 mL) NaN3 (65 mg, 1.0 mmol) dissolved in H2O (2 mL) was added and the mixture was stirred for 3 h at 40 °C. The acetone was evaporated off and after adding of H2O (5 mL) the mixture was extracted with CHCl3 (2 × 10 mL). The organic layer was separated, dried (MgSO4), filtered and evaporated. Purification of the residue by flash column chromatography (hexane-Et2O) gave compound 1d 106 mg (37%), mp 70-72 °C, R f = 0.28 (hexane-Et2O, 2:1). IR (nujol): ν = 2095 cm-1. MS (EI): m/z (%) = 287/289 (10/10) [M+], 193 (37), 152 (67), 41 (100). The side product is 3,4-bis(azidomethyl)-2,2,5,5-tetramethyl-2,5-dihydro-1H-pyrrol-1-yloxyl radical, 68 mg (27%), mp 90-92 °C, R f = 0.24 (hexane-Et2O, 2:1).

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Compounds were characterized by MS, ESR, IR and elemental analysis. Spectra were consistent in each case with the assigned structures. ESR spectra of all N-Boc protected amino acid were taken in 10-4 M water solution and all monoradicals gave triplett line aN = 15.5-15.8 G.
Representative Synthesis of Compound 5d: To stirred solution of N-diphenylmethylene glycine (801 mg, 3.0 mmol) and compound 1d (864 mg, 3.0 mmol) in CH2Cl2 (20 mL), 10% aq NaOH (3 mL) was added followed by addition of Bu4NHSO4 (508 mg, 1.5 mmol) and the mixture was stirred at r.t. for 2 h. The organic phase was separated, dried (MgSO4), filtered and evaporated to give compound 2d as a yellow oil 740 mg (52%). The crude product was immediately subjected to acidic hydrolysis. Compound 2d was dissolved in EtOH (20 mL), 5% aq H2SO4 (5 mL) was added, the mixture was allowed to stand at r.t. and the mixture was monitored by TLC. After consumption of compound 2d (ca 30 min) H2O (10 mL) was added, and the pH = 8 was adjusted by addition of solid K2CO3, extracted with CHCl3 (2 × 20 mL). Then, the organic phase was separated, dried (MgSO4), filtered, evaporated and the residue was purified by flash column chromatography (CHCl3-MeOH) to give compound 3d (203 mg, 42%) as a yellow oil. IR (nujol): ν = 3350, 3280, 2095, 1730 cm-1. MS (EI): m/z (%) = 310 (3) [M+], 249 (43), 233 (31), 161 (100). Anal. Cald for C14H24N5O3: C, 54.18; H, 7.79; N, 22.56. Found: C, 54.01; H, 7.71; N, 22.40.
To a solution of compound 3d (310 mg, 1.0 mmol) in dry THF (15 mL) t-butoxycarbonyl anhydride (240 mg, 1.1 mmol) was added and the mixture was stirred at 40 °C for 30 min. After cooling, Et2O (20 mL) was added and the organic phase was washed with brine (10 mL). Then, the organic phase was separated, dried (MgSO4), filtered and evaporated to give crude 4d as a yellow solid 279 mg (68%). This crude 4d was dissolved in EtOH (10 mL), then H2O (3 mL) and 10% aq NaOH (1 mL) were added and the mixture was allowed to stand at r.t. and monitored by TLC. After consumption of compound 4d (ca 1 h) the solution was acidified to pH = 3 by cautious addition of 5% aq H2SO4. The aqueous phase was extracted with CHCl3 (2 × 20 mL), the combined organic phase was dried (MgSO4), filtered and evaporated. The residue was purified by flash column chromatography (CHCl3-MeOH) to give compound 5d as a yellow solid 101 mg (39%), mp 160-162 °C. Anal. Calcd for C17H28N5O5: C, 53.39; H, 7.38; N, 18.31. Found: C, 53.43; H, 7.35; N, 18.50. MS was taken with thermospray technique (TSP): m/z = 383 [M + H]+.