1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide Hydrochloride1

[25952-53-8]  · C8H18ClN3  · 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide Hydrochloride  · (MW 191.74) (base)

[1892-57-5] (.MeI)


(peptide coupling reagent;1b,2 amide formation;3 ester formation;4 protein modification;5 mild oxidations of primary alcohols6)

Alternate Name: 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide; water-soluble carbodiimide; EDC; EDCI.

Physical Data: free base is an oil, bp 47-48 °C/0.27 mmHg; HCl salt is a white powder, mp 111-113 °C; MeI salt mp 97-99 °C.

Solubility: sol H2O, CH2Cl2, DMF, THF.

Form Supplied in: commercially available as HCl salt and as methiodide salt that are white solids. Reagents are >98% pure; main impurity is the urea that can form upon exposure to moisture.

Analysis of Reagent Purity: IR: 2150 cm-1 (N=C=N stretch); urea has C=O stretch near 1600-1700 cm-1.

Purification: recrystallization from CH2Cl2/ether.

Handling, Storage, and Precautions: EDC is moisture-sensitive; store under N2 in a cool dry place. It is incompatible with strong oxidizers and strong acids. EDC is a skin irritant and a contact allergen; therefore avoid exposure to skin and eyes.

Peptide Coupling Reagent.

This carbodiimide (EDC) reacts very similarly to 1,3-Dicyclohexylcarbodiimide and other carbodiimides. The advantage EDC has over DCC is that the urea produced is water soluble and, therefore, is easily extracted. Dicyclohexylurea is only sparingly soluble in many solvents and is removed by filtration which may not be as effective as extraction. A typical example of EDC for peptide coupling is shown in (eq 1).2

The problems associated with carbodiimide couplings are mainly from the O-acylisourea intermediate (1) having poor selectivity for specific nucleophiles. This intermediate can rearrange to an N-acylurea (2), resulting in contamination of the product and low yields. Additionally it can rearrange to 5(4H)-oxazolones (3) that tautomerize readily, resulting in racemization. Using low dielectric solvents like CH2Cl2 or additives to trap (1) minimizes these side reactions by favoring intermolecular nucleophilic attack on (1).1b

When low dielectric constant solvents are used, the carboxylic acid tends to dimerize, thus promoting symmetrical anhydride formation. This intermediate is stable enough to give good yields of the desired product. High dielectric solvents such as DMF retard acylation of amino acids, and N-acylurea can be a major byproduct.1b Unfortunately, many starting materials require polar solvents for dissolution.

Addition of trapping agents such as N-Hydroxysuccinimide7 and 1-Hydroxybenzotriazole (HOBt)8 reduce the extent of many side reactions, especially N-acylurea formation. Also, racemization is suppressed when these additives are present. The latter reagent eliminates the intramolecular dehydration of o-amides of asparagine and glutamine that occurs with carbodiimides (eq 2).8,9

With EDC the addition of Copper(II) Chloride suppresses racemization to <0.1% compared to 0.4% with HOBt under ideal conditions.10 Combinations of HOBt and CuCl2 are also useful.10b The suggested stoichiometry for the highest optical purity and yield is 2 equiv of HOBt and 0.25-0.5 equiv of CuCl2 in DMF.

A practical example of EDC's utility is the solution synthesis of human epidermal growth factor, a 53-residue protein.11 This included several couplings of fragments ranging from three to six residues in length. All couplings were performed with EDC/HOBt in DMF or NMP. At the completion of the synthesis, no racemized material could be detected by HPLC.

Amide Formation.

Formylated amino acids and peptides are prepared in high yields by forming the acid anhydride (eq 3).3 These products are pure without requiring chromatography or recrystallization.

Another method for formylation of amines is with p-nitrophenyl formate, which usually gives products in high yield. However, removing the last traces of the p-nitrophenol is difficult.12

Treating carbon dioxide and amines with EDC gives symmetrical ureas (eq 4).13 DCC with CO2 at ambient pressure works equally well.

EDC facilitates the synthesis of xanthine analogs by condensing a diamine with water-soluble acids (eq 5).14 The use of water as the solvent precludes the use of DCC in this case.

Ester Formation.

Esters of N-protected amino acids are prepared in high yield with EDC and 4-Dimethylaminopyridine (eq 6).4 DMAP causes extensive racemization if not used in a catalytic amount.15 However, when esterifying the a-carboxyl of b- and g-benzyl esters of aspartyl and glutamyl derivatives, extensive racemization was observed even with DMAP present in catalytic amounts. It is postulated that the side-chain esters contribute in some fashion to the lability of the a-H.

Carbodiimides including EDC are also useful in preparing active esters such as the p-nitrophenyl, pentafluorophenyl, and N-hydroxysuccinimide esters.1b Numerous additional methods have been reported.16

Protein Modification.

Carboxyl groups in proteins react with EDC, resulting in an activated group that can be trapped with a nucleophile such as glycine methyl ester.5 The nucleophile can also be amino groups in the protein, causing cross-linking. Applications for carboxylate modification include determining which groups are buried versus exposed, and mechanistic studies for determining which carboxyl groups are essential for an enzyme's activity.

EDC is used extensively in cross-linking proteins to solid supports for affinity chromatography.17

Oxidation of Primary Alcohols.

One example of EDC substituting for DCC in the Pfitzner-Moffatt oxidation is shown in (eq 7).6 The use of DCC in this example resulted in a difficult purification of the product and lower yields.

Miscellaneous Reactions.

EDC is a useful water scavenger as well. An example of this utility is given in (eq 8).18 The carbodiimide is required in this reaction, but DCC does not work well in this transformation.

Another example of EDC having a marked improvement over DCC is the formation of cyanoguanidines (eq 9).19 This was originally attempted with DCC, which gives poor yields even after extended reaction times. It is thought the positively charged nitrogen in EDC facilitates the C-S bond cleavage and since DCC lacks this atom the reaction goes slowly.

Pyrroles are formed when acetylenes undergo cycloadditions with N-acylamino acids in the presence of EDC.20 An N-alkenylmunchnone azomethine ylide is generated in situ and is trapped by the dipolaraphile. Loss of CO2 yields the pyrrole (eq 10).

Related Reagents.

Benzotriazol-1-yloxytris(dimethylamino)phosphonium Hexafluorophosphate; N,N-Carbonyldiimidazole; 1-Cyclohexyl-3-(2-morpholinoethyl)carbodiimide; 1,3-Dicyclohexylcarbodiimide; Diphenyl Phosphorazidate; Di-p-tolylcarbodiimide; N-Ethyl-5-phenylisoxazolium-3-sulfonate; 1-Hydroxybenzotriazole; Isobutene; Isobutyl Chloroformate; 1,1-Thionylimidazole.

1. (a) Kurzer, F.; Douraghi-Zader, K. CRV 1967, 67, 107. (b) Rich, D. H.; Singh, J. The Peptides: Analysis, Synthesis, Biology; Academic: New York, 1979; Vol. 1, pp 241-261.
2. Sheehan, J. C.; Ledis, S. L. JACS 1973, 95, 875.
3. Chen, F. M. F.; Benoiton, N. L. S 1979, 709.
4. Dhaon, M. K.; Olsen, R. K.; Ramasamy, K. JOC 1982, 47, 1962.
5. Carraway, K. L.; Koshland, Jr., D. E. Methods Enzymol. 1972, 25, 616.
6. Ramage, R.; MacLeod, A. M.; Rose, G. W. T 1991, 47, 5625.
7. Wuensch, E.; Drees, F. CB 1966, 99, 110.
8. Konig, W.; Geiger, R. CB 1970, 103, 788.
9. Gish, D. T.; Katsoyannis, P. G.; Hess, G. P.; Stedman, R. J. JACS 1956, 78, 5954.
10. (a) Miyazawa, T.; Otomatsu, T.; Yamada, T.; Kuwata, S. TL 1984, 25, 771. (b) Miyazawa, T.; Otomatsu, T.; Fukui, Y.; Yamada, T.; Kuwata, S. CC 1988, 419.
11. Hagiwara, D.; Neya, M.; Miyazaki, Y.; Hemmi, K.; Hashimoto, M. CC 1984, 1676.
12. Okawa, K.; Hase, S. BCJ 1963, 36, 754.
13. Ogura, H.; Takeda, K.; Tokue, R.; Kobayashi, T. S 1978, 394.
14. Shamim, M. T.; Ukena, D.; Padgett, W. L.; Daly, J. W. JMC 1989, 32, 1231.
15. Atherton, E.; Benoiton, N. L.; Brown, E.; Sheppard, R. C.; Williams, B. J. CC 1981, 336.
16. Greene, T. W. Protective Groups in Organic Synthesis, Wiley: New York, 1981; pp 154-180.
17. Keeton, T. K.; Krutzsch, H.; Lovenberg, W. Science 1981, 211, 586.
18. Tam, T. F.; Thomas, E.; Kruntz, A. TL 1987, 28, 1127.
19. Atwal, K. S.; Ahmed, S. Z.; O'Reilly, B. C. TL 1989, 30, 7313.
20. Anderson, W. A.; Heider, A. R. SC 1986, 357.

Richard S. Pottorf

Marion Merrell Dow Research Institute, Cincinnati, OH, USA

Peter Szeto

University of Bristol, UK

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