Carbodiimides carboxylic acid amides

In 1955 Sheehan and Hess andKhorana showed independently that suitably blocked amino acids are condensed with amino acids in the presence of carbodiimides to produce carboxylic acid amides 662 under mild conditions. 

Mechanism of amide formation by reaction of a carboxylic acid and an amine with dicyclohexyl-carbodiimide (DCC). 

N-substituted carbodiimides can react with carboxylic acids to form highly reactive, o-acylisourea derivatives that are extremely short-lived (Reaction 11). This active species then can react with a nucleophile such as a primary amine to form an amide bond (Reaction 12)... 

Cascade Blue cadaverine and Cascade Blue ethylenediamine both contain a carboxamide-linked diamine spacer off the 8-methoxy group of the pyrene trisulfonic acid backbone. The cadaverine version contains a 5-carbon spacer, while the ethylenediamine compound has only a 2-carbon arm. Both can be coupled to carboxylic acid-containing molecules using a carbodiimide reaction (Chapter 3, Section 1). Since Cascade Blue derivatives are water-soluble, the carbodiimide EDC can be used to couple these fluorophores to proteins and other carboxylate-containing molecules in aqueous solutions at a pH range of 4.5-7.5. The reaction forms amide bond linkages (Figure 9.39). 

Despite continnons progress in amide bond formation, the acylation of hydroxylamine nnder carbodiimide promotion is often contaminated by Af,0-diacylation even with sub-stoichiometric amounts of acids. Appendino and colleagues have developed a practical solution to the problem by combining the in situ activation of carboxylic acids 102 with the cyclic phosphonic anhydride PPAA (103), and the generation of hydroxylamine from its corresponding hydrochloride to form 104 (Scheme 54). 

Couplings with N-protected isoxazolidine-3-carboxylic acid can be performed without particular restrictions of the method, and the use of a mixed anhydride for acylation of amino acids or amide formation has been reported. 179 A similarly free choice of coupling methods is available for the acylation of isoxazolidine-3-carboxylic acid derivatives so far the use of carbodiimide and mixed anhydrides have been reported. 168179 Isoxazolidine-3-carboxylic acid derivatives are listed in Table 7. 

The mechanism of carbodiimide-mediated amide formation is outlined in Figure 13.4. The first intermediate formed during the reaction of a carbodiimide with a carboxylic acid (or an ammonium carboxylate [56]) is an O-acylisourea [1]. Under acidic conditions [57,58], this intermediate reacts with acids to yield anhydrides, and with alcohols, phenols, HOBt, or other compounds containing hydroxyl groups, to yield the corresponding esters. The carbodiimide is thereby transformed into a urea. 

For the solid-phase synthesis of amides, it makes a significant difference whether the amine or the acid is linked to the support. Resin-bound amines are readily acy-lated by adding first a carboxylic acid and then a carbodiimide (Table 13.3). The acid/ carbodiimide ratio is not critical, because both the O-acylisourea (ratio 1 1) and the symmetric anhydride (ratio 2 1) will lead to N-acylation. It should, however, be borne in mind that the half-lives of O-acylisoureas are shorter than those of anhydrides, and for difficult couplings it might be advantageous to acylate with a symmetric anhydride (two equivalents of acid and one of carbodiimide). 

The esterification of support-bound carboxylic acids has not been investigated as thoroughly as the esterification of support-bound alcohols. Resin-bound activated acid derivatives that are well suited to the preparation of esters include O-acylisoureas (formed from acids and carbodiimides), acyl halides [23,226-228], and mixed anhydrides (Table 13.15). A-Acylurea formation does not compete with esterifications as efficiently as it does with the formation of amides from support-bound acids. Esters can also be prepared from carboxylic acids on insoluble supports by acid-catalyzed esterification [152,229]. Alternatively, support-bound carboxylic acids can be esteri-fied by O-alkylation, either with primary or secondary aliphatic alcohols under Mitsu-nobu conditions or with reactive alkyl halides or sulfonates (Table 13.15). 

A carboxylate-reactive cross-linking compound typically contains a primary amine functional group that can be coupled to a carboxylic acid group in a protein or other molecule through the use of a suitable activating agent, such as a carbodiimide. The carbodiimide forms an active ester intermediate that then reacts with the amine to create an amide bond (Chapter 3, Section 1). Recent reported use of diazoalkyl deriva-... 

---