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Synlett 2006(14): 2235-2238  
DOI: 10.1055/s-2006-949632
LETTER
© Georg Thieme Verlag Stuttgart · New York

Mio- and Dio-Fmoc - Two Modified Fmoc Protecting Groups Promoting Solubility

Pablo Wessig*, Sylvia Czapla, Kristian Möllnitz, Jutta Schwarz
Institut für Chemie, Humboldt-Universität zu Berlin, Brook-Taylor-Str. 2, 12489 Berlin, Germany
Fax: +49(30)20937450; e-Mail: pablo.wessig@chemie.hu-berlin.de;
Further Information

Publication History

Received 25 June 2006
Publication Date:
24 August 2006 (online)

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Abstract

Two novel Fmoc-derived protecting groups, Mio-Fmoc and Dio-Fmoc, were developed, which cause dramatically enhanced solubility of the corresponding derivatives in most organic solvents. Furthermore, the suitability of these groups in solid-phase peptide synthesis was highlighted.

Key words

protecting groups - solid-phase synthesis - solubility - peptides

5

Dtb-Fmoc-Cl is available from Sigma Aldrich.

9

We found that the sequence consisting of carboxylation of lithiated fluorenes14 followed by the reduction of the carboxylic acid with borane15 gives clearly better yields than the commonly employed treatment of the lithiated fluorenes with formaldehyde or paraformaldehyde.

13

Friedel-Crafts Acylation of Fluorenes; General Procedure A To a stirred mixture of fluorene (3 or 5a, 60.2 mmol) and AlCl3 (8.8 g, 66.0 mmol, 1.1 equiv) in anhyd CH2Cl2 (200 mL) 2-ethylhexanoyl chloride (11 mL) was added at 0 °C. After stirring overnight, ice and concd HCl were poured into the mixture until all the solids had been dissolved. The phases were separated, the aqueous phase was extracted twice with dichloromethane, the combined organic phases were dried, and concentrated. The residue was purified by flash chromatography (silica). 2-(2-Ethylhexanoyl)fluorene ( 4a)
Flash chromatography (PE-CH2Cl2, 1:1) gave 15.1 g of 4a (51.7 mmol, 86%) as a yellow solid; R f 0.5 (PE-CH2Cl2, 1:1). 1H NMR (CDCl3, 300 MHz): δ = 0.81-0.93 (m, 6 H), 1.19-1.35 (m, 4 H), 1.46-1.64 (m, 2 H), 1.73-1.88 (m, 2 H), 3.36-3.45 (m, 1 H), 3.92 (s, 2 H), 7.25 (dd, 3 J = 6.8 Hz, 4 J = 1.0 Hz, 1 H), 7.36 (dt, 3 J = 7.4 Hz, 4 J = 1.5 Hz, 2 H), 7.80 (d, 3 J = 7.6 Hz, 2 H), 7.99 (dd, 3 J = 8.1 Hz, 4 J = 1.6 Hz, 1 H), 8.13 (d, 4 J = 0.8 Hz, 1 H). 13C NMR (CDCl3, 75 MHz): δ = 12.0, 13.9, 22.9, 25.6, 29.8, 31.9, 36.9, 47.7, 119.7, 125.2, 127.0, 127.5, 127.9, 136.3, 140.5, 143.3, 144.5, 146.1, 204.5. 2-(2-Ethylhexanoyl)-7-(2-ethylhexyl)fluorene ( 6)
Flash chromatography (PE-EtOAc, 10:1) gave 6.81 g of 6 (16.83 mmol, 95%) as a yellow oil; R f 0.5 (PE-EtOAc, 20:1). 1H NMR (CDCl3, 300 MHz): δ = 0.77-0.89 (m, 12 H), 1.17-1.31 (m, 12 H), 1.39-1.60 (m, 3 H), 1.68-1.83 (m, 2 H), 2.49-2.61 (m, 2 H), 3.31-3.40 (m, 1 H), 3.86 (s, 2 H), 7.13 (dd, 3 J = 7.9 Hz, 4 J = 1.2 Hz, 1 H), 7.30 (s, 1 H), 7.67 (d, 3 J = 7.8 Hz, 1 H), 7.93 (dd, 3 J = 8.1 Hz, 4 J = 1.5 Hz, 1 H), 8.07 (s, 1 H). 13C NMR (CDCl3, 75 MHz): δ = 10.8, 12.0, 13.9, 14.1, 22.9, 23.0, 25.4, 25.6, 28.8, 29.8, 31.9, 32.3, 36.8, 40.4, 41.3, 47.6, 119.3, 120.4, 124.8, 126.0, 127.5, 128.1, 135.9, 138.1, 142.3, 143.3, 144.6, 146.4, 204.5. Reduction of 2-Ethylhexanoyl Groups; General Procedure B
AlCl3 (12 g, 90.0 mmol, 1.7 equiv) was placed in a 500-mL round-bottom flask and anhyd Et2O (300 mL) was added cautiously dropwise with external cooling with an ice bath and stirring. A solution of 4a (51.7 mmol) or 6 in anhyd Et2O (100 mL) was added dropwise. After stirring for 30 min LiAlH4 (3.42 g, 90.1 mmol, 1.7 equiv) was added portion-wise. The mixture was refluxed for 90 min and then allowed to cool to r.t. The excess of LiAlH4 was destroyed with EtOAc and then dil. HCl was added until gas evolution ceased. The phases were separated, the organic phase was dried with MgSO4, and evaporated. The residue was purified by flash chromatography (silica). 2-(2-Ethylhexyl)fluorene ( 5a)
Flash chromatography (PE-CH2Cl2, 100:3) gave 8.49 g of 5a (30.5 mmol, 59%) as a yellow solid; R f 0.4 (PE-EtOAc, 100:3). 1H NMR (CDCl3, 300 MHz): δ = 0.99-1.09 (m, 6 H), 1.36-1.51 (m, 8 H), 1.69-1.80 (m, 1 H), 2.68-2.80 (m, 2 H), 3.98 (s, 2 H), 7.29 (d, 3 J = 7.8 Hz, 1 H), 7.39 (dt, 3 J = 7.4 Hz, 4 J = 1.2 Hz, 1 H), 7.45 (s, 1 H), 7.48 (t, 3 J = 6.9 Hz, 1 H), 7.64 (d, 3 J = 7.4 Hz, 1 H), 7.81 (d, 3 J = 7.8 Hz, 1 H), 7.87 (d, 3 J = 7.4 Hz, 1 H). 13C NMR (CDCl3, 75 MHz): δ = 10.8, 14.2, 23.1, 25.4, 28.9, 32.4, 36.8, 40.3, 41.3, 119.4, 119.5, 124.9, 125.8, 126.2, 126.6, 127.8, 139.2, 140.7, 141.8, 143.1, 143.3. 2,7-Bis(2-ethylhexyl)fluorene ( 5b)
Flash chromatography (PE) gave 4.47 g of 5b (11.44 mmol, 68%) as a colorless oil; R f 0.6 (PE). 1H NMR (CDCl3, 300 MHz): δ = 0.85-0.93 (m, 12 H), 1.29-1.38 (m, 16 H), 1.62-1.64 (m, 2 H), 2.55-2.67 (m, 4 H), 3.85 (s, 2 H), 7.15 (d, 3 J = 7.7 Hz, 2 H), 7.32 (s, 2 H), 7.65 (d, 3 J = 7.7 Hz, 2 H). 13C NMR (CDCl3, 75 MHz): δ = 10.8, 14.2, 23.1, 25.4, 28.9, 32.3, 36.7, 40.3, 41.4, 119.0, 125.8, 127.7, 139.4, 140.1, 143.2. Fluorene-9-carboxylic Acids; General Procedure C1
Fluorene 5a or 5b (19.4 mmol) was dissolved in anhyd THF (30 mL) and cooled to -78 °C. n-BuLi (1.6 M, hexane; 15 mL, 24 mmol, 1.2 equiv) was added dropwise and then the solution was stirred for 1 h. A stream of carbon dioxide, obtained by evaporation of dry ice was passed through a drying jar, into the solution for 30 min, while warming to r.t. The mixture was acidified with dil. HCl to pH 1 and extracted with Et2O. The solvent was evaporated and the residue was purified by flash chromatography. Fluorene-9-methanols; General Procedure C2
Substituted fluorene-9-carboxylic acid (17.24 mmol) was dissolved in anhyd THF (100 mL) and then BH3·Me2S (3.5 mL, 30.9 mmol, 2.1 equiv) was added dropwise at 0 °C. After warming to r.t. the solution was stirred for 2 h; crushed ice and an aq solution of tartaric acid were added cautiously until gas evolution ceased. The mixture was extracted several times with Et2O, the organic phases were washed twice with a sat. aq solution of NaHCO3, dried, and evaporated. The residue was purified by flash chromatography giving the fluorene-9-methanols. Fluorene-9-methyl Chloroformates; General Procedure C3
Fluorene-9-methanol (4.18 mmol) was stirred with a solution of phosgene (10% in toluene, 7.79 mmol, 1.9 equiv) in a sealed flask overnight. The solvent was removed in vacuo and the residue was purified by flash chromatography. 2-(2-Ethylhexyl)fluorene-9-methyl Chloroformate (Mio-Fmoc-Cl) ( 7a)
Instead of flash chromatography the residue of C1 was purified by dissolution in an aq solution of K2CO3 and extracted several times with Et2O. After acidification with dil. HCl to pH 1 the product was extracted with CH2Cl2. The solution was dried and evaporated giving the pure fluorene-9-carboxylic acid. Yields: C1, 90%; C2, 55%; C3, 70%; R f 0.6 (PE-EtOAc, 100:2). 1H NMR (CDCl3, 300 MHz): δ = 0.85-0.89 (m, 6 H), 1.20-1.34 (m, 8 H), 1.56-1.62 (m, 1 H), 2.52-2.64 (m, 2 H), 4.23 (t, 3 J = 7.6 Hz, 1 H), 4.44-4.57 (m, 2 H), 7.18 (d, 3 J = 7.8 Hz, 1 H), 7.27 (dt, 3 J = 7.4 Hz, 4 J = 1.0 Hz, 1 H), 7.33 (s, 1 H), 7.37 (t, 3 J = 7.4 Hz, 1 H), 7.53 (d, 3 J = 7.5 Hz, 1 H), 7.63 (d, 3 J = 7.8 Hz, 1 H), 7.70 (d, 3 J = 7.5 Hz, 1 H). 13C NMR (CDCl3, 75 MHz): δ = 10.8, 14.1, 23.0, 25.4, 28.8, 32.3, 40.3, 41.3, 46.0, 73.6, 119.8, 119.9, 125.0, 125.8, 126.8, 128.1, 129.2, 138.8, 141.4, 141.5, 142.4, 150.7. 2,7-Bis(2-ethylhexyl)fluorene-9-methyl Chloroformate (Dio-Fmoc-Cl) ( 7b)
Yields: C1, 89%; C2, 60%; C3, 97%. 1H NMR (CDCl3, 300 MHz): δ = 0.85-0.90 (m, 12 H), 1.26-1.35 (m, 16 H), 1.54-1.62 (m, 2 H), 2.53-2.66 (m, 4 H), 4.23 (t, 3 J = 7.6 Hz, 1 H), 4.52 (d, 3 J = 7.7 Hz, 2 H), 7.18 (d, 3 J = 7.8 Hz, 2 H), 7.33 (s, 2 H), 7.61 (d, 3 J = 7.8 Hz, 2 H). 13C NMR (CDCl3, 75 MHz): δ = 10.8, 14.1, 23.0, 25.4, 28.8, 32.3, 40.3, 41.3, 45.9, 73.8, 119.5, 125.8, 129.2, 139.0, 141.0, 142.4, 150.7. Solid-Phase Peptide Synthesis
The assembly of the peptides was performed in a mechanical shaker with a fritted glass reactor. Wang resin16 (1 mmol/g, 200-400 mesh) is used as the solid support which is preloaded with Fmoc-Gly-OH.17 The preloaded resins (50 mg, loading 0.89 mmol/g) were swollen in DMF (0.5 mL) for 5 min, and than the excess solvent was removed by filtration. The Fmoc protecting group was removed by treating twice with 20% piperidine in DMF (5 min and 15 min) and then the resin was washed 5 times with DMF. The coupling step was performed by the Fmoc strategy using TBTU/HOBt18 as activation reagents. In a typical experiment, each Fmoc-dipeptide (Table [2] ) was introduced by a common coupling (60 min) using a two-fold excess of the dipeptide, activation of reagents, and a three-fold excess of DIPEA in DMF (0.5 mL). After washing five times with DMF and twice with CH2Cl2 the resin was dried in high vacuum. The substitution levels of the loaded resins were determined spectrophotometrically by Fmoc cleavage.3