Activation by carbodiimides

Reduction of uronic acids Uronic acids (UA) were converted to the corresponding neutral sugars (NS) by carbodiimide activation of the carboxyl groups followed by a reduction with NaBD4 according to the method of Kim and Carpita [5]. In order to achieve a complete reduction of the uronic acids the procedure was repeated once. 

PGIP, purified fi om P.vulgaris hypocotyls [11], was immobilized to the sensor ch via amine coupling. A continuous flow of HBS buffer (5 pl/min) was mantained over the sensor surface. The carboxylated dextran matrix of the sensor surface was first activated by a 6-min injection of a mixture of N-hydroxy-succinimide and N-ethyl-N - (3-diethylaminopropyl) carbodiimide, followed by a 7-min injection of PGIP (lOng/pl in 10 mM acetate, pH 5.0). Hie immobilization procedure was con leted by a 7-min injection of 1 M ethanolamine hydrochloride to block the remaining ester groups. 

Two equivalents of the tertiary amine base are required, and a significant improvement in the diastereoselectivity was observed with TMEDA over DIPEA. Purification and further enrichment of the desired RRR isomer to >98% ee was achieved by crystallization. Oxidative removal of the chiral auxiliary followed by carbodiimide mediated amide formation provides (3-keto carboxamide 14 in good yield. Activation of the benzylic hydroxyl via PPha/DEAD, acylation, or phosphorylation, effects 2-azetidinone ring-closure with inversion of stereochemistry at the C4 position. Unfortunately, final purification could not be effected by crystallization and the side products and or residual reagents could only be removed by careful chromatography on silica. 

Unfortunately, A-(9-fluorenylmethoxycarbonyl)aziridine-2-carboxylic acid cannot be used in peptide synthesis, since N-deprotection of the respective peptides with secondary amines leads to oxazoline or dehydroamino acid side products. Similarly, N-(tert-butoxy-carbonyl)aziridine-2-carboxylic acid is inappropriate due to the instability of the aziridine moiety to TFA treatment. Attempts to convert A-tritylaziridine-2-carboxylic acid into homogenous and stable active esters as useful intermediates in peptide synthesis leads to positive results only in the case of the pentafluorophenyl ester. 47 Consequently, this active ester seems to be the method of choice for acylating peptides. The related Abhydroxysuc-cinimide and A-3-hydroxy-4-oxo-3,4-dihydro-l,2,3-benzotriazine ester could not be isolated in pure form and have therefore been used as crude products. 47 Access to 2-carbonylazir-idine peptides is also possible by carbodiimide-mediated coupling. Additionally, alkylamides of A-tritylaziridine-2-carboxylic acid are prepared by the azide method,1 5 yet this method fails in peptide coupling steps. 85 ... 

The modification of carboxyl groups has been carried out (1) by esterification with dry methanol and HC1, (2) by esterification with aliphatic diazo compounds, (3) by the formation of adducts with carbodi-imides, or (4) by the formation of amides through activation with carbodiimides. Both complete and, apparently specific, partial modification of the 11 free carboxyl groups have been obtained. In general, the first method suffers from the denaturing medium, the second from incomplete reaction, and the third from the uncertain nature of the products. The fourth procedure is perhaps subject to the least question. There are a total of 11 free carboxyl groups in native RNase-A l (Val), 5/ (Asp), 5y(Glu). A summary of the derivatives is given in Table V. 

The combination of carboxyl activation by a carbodiimide and catalysis by DMAP provides a useful method for in situ activation of carboxylic acids for reaction with alcohols.10 The reaction proceeds at room temperature. Carbodiimides are widely applied in the synthesis of polypeptides from amino acids. The proposed mechanism for this esterification reaction involves activation of the acid via isourea 28 followed by reaction with another acid molecule to form anhydride... 

Equimolar quantities of benzhydrol and the phosphorane (3a) were also reacted in dimethyl sulphoxide solution. Apart from bis(benzhydryl) ether (18%) and catechol monobenzhydryl ether (39%), a small amount of benzophenone (17%) was obtained. It was shown that, in the absence of phosphorane, benzhydrol is not oxidised to benzophenone by dimethyl sulphoxide. Reaction similar to that outlined in Scheme 1 is a likely possibility. Again, the alcohol is activated by reaction with the phosphorane toward nucleophilic attack, in this case by dimethyl sulphoxide. Significantly, oxidation of alcohols by dimethyl sulphoxide is usually carried out using the Pfitzner-Moffatt reagent (dicyclohexyl carbodiimide and anhydrous phosphoric acid in dimethyl sulphoxide) (13) whereas the reaction using the phosphorane (3a) is carried out under neutral conditions. Unfortunately, however, attempts to improve the yield of benzophenone have hitherto failed. 

The surface matrix of a carboxymethylated sensor chip CMS (Pharmacia Biosensor, Uppsala, Sweden) was activated by injection of 35 pL of 0.05 M N-hydroxy-succinimide (NHS)/0.2 M N-ethyl-N (dimethylaminopropyl)carbodiimide (EDC). The BIAcore IFC forms four parallel flow cells on the sensor chip. Three of them were derivatized with 40 mM cystamin-dihydrochloride/ethanolamine in the ratios... 

Ketenes readily react with carbodiimides to yield 1 1 adducts (equation 41). The formation of 2 1 adducts (18) is observed when the C=N double bond of the 1 1 adduct is activated by strain... 

However, the isolation of optically pure oxazol-5(4//)-ones from Boc- or Z-protected amino acids by carbodiimide, mixed anhydride, or acyl chloride activation contradicts the above assumptions. It was suggested that these oxazol-5(4//)-ones are 2-alkoxyoxazol-5(4//)-ones and are chirally stable (Scheme... 

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