Marfey s reagent

Marcus equation Marezine Marfey s reagent Margaric acid [506-12-7]... 

Fig. 7-6). Two unichiral amides which have been known capable of this reaction are 1-phenylethylamine [15] and l-(l-naphthyl)ethylamine [16]. Marfey s reagent [N-a-(2,4-dinitro-5-fluorophenyl)-L-alaninamide] was introduced as a reagent to deriva-tize amino acids with cyclopentane, tetrahydroisoquinoline or tetraline structures [17]. Simple chiral alcohols such as 2-octanol can also be used to derivatize acids such as 2-chloro-3-phenylmethoxypropionic acid [18]. 

A simple and rapid method of separating optical isomers of amino acids on a reversed-phase plate, without using impregnated plates or a chiral mobile phase, was described by Nagata et al. [27]. Amino acids were derivatized with /-fluoro-2,4-dinitrophenyl-5-L-alanine amide (FDAA or Marfey s reagent). Each FDAA amino acid can be separated from the others by two-dimensional elution. Separation of L- and D-serine was achieved with 30% of acetonitrile solvent. The enantiomers of threonine, proline, and alanine were separated with 35% of acetonitrile solvent and those of methionine, valine, phenylalanine, and leucine with 40% of acetonitrile solvent. The spots were scraped off the plate after the... 

JG Adamson, T Hoang, A Crivici, GA Lajoie. Use of Marfey s reagent to quantitate racemization upon anchoring of amino acids to solid supports for peptide synthesis. Anal Biochem 202, 210, 1992. 

The enantiomeric purity of protected amino acids used in peptide synthesis can be determined by pre-column partial deprotection followed by derivatization with Marfey s reagent (116). The Marfey diastereoisomers can be easily resolved and determined by RP-HPLC using an ODS-Hypersil column288. Fifteen amino acids collected from mammalian tissues were derivatized with Marfey s reagent and subjected to two-dimensional TLC. Each individual spot (enantiomeric mixture of a diasteroisomer) was then resolved by RP-HPLC. Except for tyrosine (46) and histidine (117), subnanomole quantities of enantiomers could be analyzed289,290. 

Direct and indirect chromatographic methods were developed and compared in systematic examinations for the enantioseparation of P-amino acids direct separation of underivatized analytes involved the use of commercially available Crownpak CR(-I-), teicoplanin, and ristocetin A CSPs [148], while indirect separation was based on precolumn derivatization with 2,3,4,6-tetra-G-acetyl-f)-D-glucopyranosyl isothiocyanate (GITC) or A - a-(2,4-dinitro-5-fluorophenyl)-L-alaninamide (EDAA, Marfey s reagent), with subsequent separation on a nonenantioselective column. 

CE has been applied extensively for the separation of chiral compounds in chemical and pharmaceutical analysis.First chiral separations were reported by Gozel et al. who separated the enantiomers of some dansylated amino acids by using diastereomeric complex formation with Cu " -aspartame. Later, Tran et al. demonstrated that such a separation was also possible by derivatization of amino acids with L-Marfey s reagent. Nishi et al. were able to separate some chiral pharmaceutical compounds by using bile salts as chiral selectors and as micellar surfactants. However, it was not until Fanali first showed the utilization of cyclodextrins as chiral selectors that a boom in the number of applications was noted. Cyclodextrins are added to the buffer electrolyte and a chiral recognition may... 

Fluoro-2,4-dinitrophenyl-5-L-alanine amide (Marfey s reagent) and 2..3.4.6-tetra-0-aeetyl-P-D-gluer>pyranosyl isothkKyanatc... 

Marfey s Reagent Based HPLC Method General Procednre ... 

Determination of D- and L-His content in peptides was carried out using Marfey s reagent, l-fluoro-2,4-dinitrophenyl-5-I-alanine amide (FDAA) (9). Peptides were first hydrolyzed for 18 hr in 6 N HCl under reduced pressure. To ensure that racemization during hydrolysis would be minimized, this time was determined by evaluation of a time course of hydrolysis on the reference peptides. After incubation with 10 mM FDAA in 0.1 M sodium bicarbonate for 1 hr at 40°, the hydrolysates were acidified with 0.2 N HCl before injection on a Hypersil... 

There was generally agreement between the results from determination of racemization by AAA following reaction with Marfey s reagent, by HPLC, with somewhat less agreement by CPA/MALDI-MS. However, in some cases the three analytical methods yielded divergent results. Therefore, the possible sources of discrepancy among the methods were also evaluated as part of this... 

In several instances, AAA following reaction with Marfey s reagent revealed higher racemization than shown by either HPLC or CPA/MALDI-MS this most frequently occurred when there was a failed synthesis, such as samples 6410 and 7818 (see below). Information can be obtained from AAA following reaction with Marfey s reagent even if the synthesis fails. In contrast, HPLC and CPA/MALDI-MS are more dependent on the presence of the correct sequence. In order to effectively utilize the HPLC and CPA/MALDI-MS methods with less pure samples, the corresponding signature ions and retention times of the modified products must be aseertained. 

The purity of protected amino acids is especially important for the synthesis of longer peptides. Standard techniques such as melting point determination, nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry, and optical rotation are effective means of characterization. The optical purity can also be evaluated by high-performance liquid chromatography (HPLC) after derivatization with Marfey s reagent [216,217]. The advanced Marfey method refers to analysis by mass spectrometry after derivatization with Marfey s reagent [218-221]. Purification of side-chain protected amino acids by recrystallization is usually sufficient. 

Figure 6.38 The separation of (a) d- and L-amino acids, and (b) d- and L-dopa enantiomers after reaction with Marfey s reagent. Peaks (a) 1, t-Gen 2, o-Gm 3, Marfey-OH 4, t-Met 5, o-Met 6, L-Phe. (b) L, L-dopa D, o-dopa M, Morfey-OH. (Reproduced by permission of Pierce...

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