DCHA salt

The Z-pGlu-OH-DCHA salt (44.4 g, 100 mmol) was stirred with 1M HC1 (300mL) and EtOAc (350 mL) for 1 h. After filtration, the organic phase was washed with H20, dried (Na2S04), concentrated, and the residue recrystallized (EtOAc/petroleum ether) yield 23.2 g (88%) mp 134-135 °C. 

This compound was synthesized as described above for 71, with 2 -(phthalimidomethyl)biphenyl-2-acetic add (68 11.04 g) as the starting material. Concentration of the Et20 layers yielded an oily material yield 6.8 g (67%). The DCHA salt was used for the elemental analysis. 

The tetravalent lysinyl core peptide was synthesized as described previously 130 Briefly, the synthesis of the first level of carrier core to form Boc-Lys(Boc)-Ala-OCH2-PAM-resin was achieved using a fourfold excess of Boc-Lys(Boc)-OH DCHA salt with BOP in CH2C12. The second level of lysine was generated by the same protocol. After Boc deprotection, tetravalent chloroacetyl moieties were introduced to the core peptide by using a tenfold excess of chloroacetic acid via DIC/HOBt activation. The tetravalent (chloroacetyl)lysinyl core peptide was cleaved from the resin by HF/PhOMe (9 1) and finally purified by RP-HPLC. MALDI-MS m/z [M + H]+ calcd, 780.5 found, 780.5 [M+Na —H]+ calcd, 802.5 found, 802.3. 

To a stirred soln of Z-Cl (1 54.4 mmol) in CHCI3 (100 mL) was added HOSu-DCHA salt (obtained as a precipitate in acetone by mixing equimolar quantities of HOSu and DCHA) (54.4 mmol) in portions. After 12 h, the precipitate was filtered off and washed with CHCI3. The filtrate and washings were combined and washed with one-third of the volume each of 10% citric acid, 10% NaHC03, and H2O, and dried. The solvent was removed and the residue was recrystallized (CHCl3/Et20) yield 90% mp 80-81 °C. 

The protection of a-amino groups is usually effected with allyl chloroformate under Schotten-Baumann conditions,P but diallyl dicarbonate (AI0C2O) can also be used.P As with other chloroformates, a severe side reaction is the formation of A -Aloc-protected dipeptides. Since most A -Aloc amino adds are not crystalline compounds, they are preferentially isolated as the corresponding DCHA salts (for A -Aloc amino acids and their analytical characterization, see reff l). 

The DCHA salts were obtained by adding the amine (1 equiv) to a soln of the oily Aloe derivatives in EtOH or EtOAc and precipitation with Et20 or Et20/petroleum ether for yields and analytical characterization, see refP l... 

Method A A soln of amino acid methyl or ethyl ester hydrochloride (lOmmol), TEA (1.01 g, 10 mmol), and Teoc-ONp (2.83 g, 10 mmol) in THF (50 mL) was stirred at rt for 24 h. The mixture was concentrated, and the residue partitioned between EtOAc and H2O. The organic layer was washed with 2% aq KHSO4, 5% aq Na2C03, and H2O, dried, and evaporated. The iV -Teoc amino ester was dissolved in dioxane (50 mL) and treated with 1M NaOH (1 equiv). After completion of the reaction, the bulk of dioxane was removed, H2O was added and the mixture acidified to pH 2-3 with 1M HCl. The product was extracted with EtOAc or Et20 and the combined extracts were washed with H2O, dried, and concentrated. The product was crystallized from a suitable solvent or isolated as its DCHA salt. For example Teoc-Ala-OH DCHA (crystallized from iPr20) yield 66% mp 123-125 °C [a]D —11.7, [aW° -13.9 (c 1, 80% AcOH). 

To a stirred soln of the amino acid (Immol) in HjO (ImL) was added a soln of TEA (152 mg, 1.5 mmol) in dioxane (ImL) followed by solid Teoc-OBt (0.31 g, -1.1 mmol). The mixture was stirred at rt overnight, then diluted with HjO (5mL), acidified with sat. KHSO4, and extracted with Et20 (3 x 15 mL). The combined organic extracts were washed with H2O (4 x 20 mL), dried, and concentrated. The crude products were purified by crystallization or as CHA or DCHA salts. For example Teoc-Ala-OH CHA yield 86% Teoc-Ser-OH CHA (crystallization from CH2Cl2/Et20, 1 1 and hexane) yield 88% mp 120-123 °C [a]o -1-12.8 (c 1.9, MeOH) Teoc-Val-OH- CHA (crystallization from iPr02/hexane) yield 95% mp 127-133 C -2.0 (c 3.7, MeOH). 

The Af -Mbv amino acid derivatives are used as potassium l or DCHA salts,the latter being less hygroscopic and more stable, crystalline compounds soluble in organic solvents. N-Enamine derivatives have also been used for intermediate protection in the synthesis of esters, whereby best results were obtained with the methyl or ethyl acetoacetate derived enamines.f 1... 

A mixture of freshly distilled AC2O (700 mL) and Z-Asp-OH (1.005 kg, 3.75 mol) was stirred at 45 °C for 4h and the solvent removed. The solid residue was triturated with abs MeOH (4L) and heated under reflux for another 4 h. After removal of the solvent, the oil was dissolved in Et20 and extracted with several portions of aq NaHC03, and each fraction was separately acidified and stored in the cold. Fractions in which the Z-Asp-OMe separated in crystalline form were combined and dissolved in Et20. Upon addition of DCHA (1.05 equiv), the DCHA salt was filtered off and recrystallized (EtOH/ petroleum ether) yield 961 g (52%) mp 159-160°C -1-14.9 (c 1, EtOH). 

The 4-sulfobenzyl ester has been proposed as an anionic protecting group by Bindewald et al.P 1 The incorporation of a charged group in totally protected peptides increases the solubility in polar solvents. For the synthesis of 4-sulfobenzyl esters, N-protected anoino acid cesium or DCHA salts are reacted with sodium 4-(bromomethyl)benzenesuIfonate in DMF... 

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