Woodward’s Reagent K

The amino add analysis of all peptide chains on the resins indicated a ratio of Pro Val 6.6 6.0 (calcd. 6 6). The peptides were then cleaved from the resin with 30% HBr in acetic acid and chromatogra phed on sephadex LH-20 in 0.001 M HCl. 335 mg dodecapeptide was isolated. Hydrolysis followed by quantitative amino acid analysis gave a ratio of Pro Val - 6.0 5.6 (calcd. 6 6). Cycll2ation in DMF with Woodward s reagent K (see scheme below) yielded after purification 138 mg of needles of the desired cyc-lododecapeptide with one equiv of acetic add. The compound yielded a yellow adduct with potassium picrate, and here an analytically more acceptable ratio Pro Val of 1.03 1.00 (calcd. 1 1) was found. The mass spectrum contained a molecular ion peak. No other spectral measurements (lack of ORD, NMR) have been reported. For a thirty-six step synthesis in which each step may cause side-reaaions the characterization of the final product should, of course, be more elaborate. 

Woodward s Reagent K N-Ethyl-5-phenylisoxazolium-3 -sulfonate, sodium salt MW 176... 

Figure 3.11 Woodward s reagent K undergoes a rearrangement in alkaline solution to form a reactive ketoket-enimine. This active species can react with a carboxylate group to create another active group, an enol ester derivative. In the presence of amine nucleophiles, amide bond formation takes place.

Boyer, T.D. (1986) Covalent labeling of the nonsubstrate ligand-binding site of glutathione S-transferase with bilirubin-Woodward s reagent K./. Biol. Chem. 261, 5363. 

Dunn, B.M., and Affinsen, C.B. (1974) Kinetics of Woodward s reagent K hydrolysis and reaction with staphylococcal nuclease./. Biol. Chem. 249, 3717. 

Johnson, A.R., and Dekker, E.E. (1996) Woodward s reagent K inactivation of Escherichia coli L-threo-nine dehydrogenase Increased absorbance at 340-350 nm is due to modification of cysteine and histidine residues, not aspartate or glutamate carboxyl groups. Protein Sci. 5, 382-390. 

Isoxazolium salts, such as the well-known Woodward s Reagent K , have found significant use as reagents in the preparation of activated enol esters suitable for peptide coupling reactions. Since this aspect of isoxazole chemistry has been extensively surveyed, this topic will not be treated in further detail here (B-74MI11601). 

Woodward s reagent K is N-ethyl-3-phenylisoxazolium-3 -sulfonate, a zero-length cross-linking agent able to cause the condensation of carboxylates and amines to form amide bonds (Woodward et al., 1961 Woodward and Olofson, 1961). The reaction... 

Some examples of the isoxazolium ions which have been tried as peptide coupling reagents are compounds 56-59. The sulfonium inner salt 56 (Woodward s Reagent K) is widely used because the by-product (60) in Scheme 10 is water soluble it is stated to be superior to diimides as far as racemization is concerned.168,170,171 (V-Arylisoxazolium salts such as 57... 

Woodward s reagent K or NEPIS 84 is a zwitterionic isoxazolinium that reacts with A-protected amino acids in presence of triethylamine to form the activated enol ester 85 (108) (see Fig. 16). This intermediate can be reacted without additional purification with an amine to yield the desired amide and the sulfonate by-product that can be easily removed by aqueous extraction. 

N -ethyl-phenylisoxazolium-3 -sulphonate, also known as Woodward s reagent K N -methylmorpholine pentafluorophenyl para-nitrophenyl... 

Condensation of 5,5-dimethylhexan-l,3-dione (dimedone) with amino acid esters leads to Af -5,5-dimethyl-3-oxocyclohex-l-enyl derivatives 128 as particularly stable enamines that are converted into the free acids by saponification, for coupling by carbodiimides, carbonyldiimidazole, or Woodward s reagent K, or are transformed into the hydrazides for coupling via the azide method (Scheme... 

If cross-linking is not expected to occur, then Woodward s reagent K may be included in an enzyme solution, and a single-step immobilization reaction may be performed. 

Woodward s Reagent K 830 Active ester Water-soluble by-products ease work-up. 

Peptide synthesis. Izumiya and Muraoka1 examined the extent of isomerization with commonly used coupling reagents (isobutyl chloroformate, DCC, N-ethyl-5-phenylisoxazolium-3 -sulfonate, and EEDQ) and of these found that EEDQ caused the least racemization (0.2%) in the particular test chosen. The next most satisfactory was N-ethyl-5-phenylisoxazolium-3 -sulfonate (Woodward s Reagent K, 1.7-1.8%). 

Woodward s Reagent K. 2-Ethyl 5-m sutfophen-ylisoxazoltiim hydroxide, fit nor salt N-ethyl-5-pbenylisoxa-zolium-3 -sulfonate. CnH-.NO.S mol wt 253,29. C 52.16%, H 4.38%, N 5.53%, O 25.27%, S 12.66%. Prepd by reacting 5-phenylisoxazole with chlorosulfonic acid, followed by alkylation with triethyloxonium fluoroborate, and acid hydrolysis Woodward et ai. J. Am. Chem. Soc. 83, 1010 (I960... 

Polymers with intramolecular cross-links have organized spatial structures, and are good models for the tertiary structure of proteins (26,27,1 ). In the studies discussed here poly Glu52Lys33Tyr15 (No. 3) was internally cross-linked with Woodward s reagent K or ionic carbodiimide and fractionated on Sephadex G-100. The number of cross-links in each fraction was determined by amino acid analysis. In addition, the parent polymer was fractionated in the same way to provide standards for comparison with the cross-linked polypeptides. 

Figure 4. Immobilization of N-acetulated diaminoalkane agarose gels using N-ethyl-5-phenylisoxazolium-3f -sulfonate (Woodward s Reagent K) activated heparin carboxylic groups.

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