N-trifluoroacetyl derivatives

The energy barriers and relative conformer stabilities have also been obtained and interpreted for a number of N-trifluoroacetyl derivatives (84BSB927) 119 in all those examined the Z-conformer prevails. 

N-Trifluoroacetyl derivatives were applied in the GC analysis of amino acids in combination with different alkyl esters. TFA anhydride serves as a strong acylating agent, which is very efficient in the derivatization of all protonic groups except carboxyl. An acylation medium, usually a mixture of TFA anhydride and methylene chloride, may be injected into the GC column without any preliminary evaporation. This is a very important fact as TFA derivatives are very sensitive towards moisture and mere evaporation can lead to decomposition, particularly of acylated hydroxy and thiol groups. Possible losses of more volatile derivatives are also eliminated. 

Peptide synthesis. The reagent is used to prepare N-trifluoroacetyl derivatives of amino acids and peptides. The substrate is warmed with 1.2-2 equivalents of the reagent in molten phenol at 120-150° for 2-20 min. Excess reagent and phenol are removed under vacuum or by solution in petroleum ether or carbon tetrachloride, from which the amino acid derivative crystallizes. The main advantage of the N-trifluoroacetyl protective group is that it can be removed under very mild conditions. 1,. Benoiton, H. N. Rydon, andJ. E. Willett, Chem. Ind., 1060(1960)... 

Z2. Zomzely, C., Marco, G., and Emery, E., Gas chromatography of the n-butyl-N-trifluoroacetyl derivatives of amino acids. Anal. Chem. 34, 1414-1417 (1962). 

Nonvolatile analytes must be chemically converted to a volatile derivative before analysis. For example, amino acids are not sufficiently volatile to analyze directly by gas chromatography. Reacting an amino acid with 1-butanol and acetyl chloride produces an esterfied amino acid. Subsequent treatment with trifluoroacetic acid gives the amino acid s volatile N-trifluoroacetyl- -butyl ester derivative. 

A sensitive method for primary amines is shown in reaction 2, leading to the corresponding 7V-benzenesulfonyl-/V-trifluoroacetyl derivatives. These can be determined by GC-ECD using SE-30 columns LOD 1-5 pg, which is about 200 times more sensitive than GC-FID. The method was applied for determination of phenethylamine (33) in urine110. This analysis was performed also by LLE into n-pentane, derivatization to the benzenesulfonamide and GC-FPD using a capillary column recoveries of aliphatic primary amines in urine were 91-107%, RSD 0.2-4.5%111,112. Amines in environmental waters and sediments were determined after LLE with dichloromethane, derivatization with benzenesulfonyl chloride and GC-SIM-MS LOD 0.02-2 pg/L of water and 0.5-50 ng/g of sediment113. 

Finally, a recent example on the biocatalyzed synthesis of new cytotoxic derivatives of the anthracycline doxurobicin (8) deserves to be mentioned. In the presence of Mucorjavanicus lipase or subtilisin Carlsberg ion-paired with Aerosol , the primary C-14 OH was chemoselectively acylated to give the corresponding derivatives, such as the valerate 8a [19]. However, optimization of this enzymatic synthesis did not provide a synthetic approach suitable for the preparative synthesis of gram quantities of Valrubicin (N-trifluoroacetyl doxurobicin-14-valerate, 8b)... 

Acetylation of amino groups is necessary to promote volatility and trifluoroacetyl derivatives (N-TFA) formed from trifluor-oacetic anhydride or acyl derivatives formed from acetic anhydride have both been used with success (Table 9.3). Excess reagent must be removed with care at this point since the derivatives are highly volatile. Gehrke and co-workers have studied a number of stationary phases in detail as summarized in Figure 9.8 (41). They have suggested the use of Apiezon M and a rapid GLC... 

N-trifluoroacetyl amino acids have been reported as stable at -10°C over a 2-week period (53) and the TFA n-butyl esters are stable for about 72 hr when stored at 0 C in a nitrogen atmosphere (32). Reasonable stability of derivatives is necessary for repetitive analyses when many samples are prepared and stored prior to chromatography. Instability of silyl derivatives to hydrolysis has been a major problem which has led to decreased activity in research on these materials. The efficient treatment of amino acids with N,0-bis(trimethylsilyl)acetamide (51) and subsequent GLC of the derivatives is now used only occasionally as a result of instability of the derivatives. 

The naturally occurring amino acids are very polar, and cannot be separated as the free compounds by GC at a temperature below decomposition. If the polar groups in the molecule are chemically modified to produce a more volatile derivative a suitable temperature is then possible. Weinstein (25) reviews all the various derivatives which may be formed from amino acids and the GC conditions necessary to separate them. In actual practice only three derivatives are in popular use. These include the N-heptafluorobutyryl n-propyl ester derivatives, the N-trimethyl-silyl ether derivatives, and the n-trifluoroacetyl n-butyl ester derivatives. 

Azodrin ([3-(dimethoxy-phosphinyl) N-methyl-cis-crotonamide]) Strawberries Solvent extraction GC-MS of trifluoroacetyl derivatives [123]... 

Fig 1 Association of N-trifluoroacetyl (N-TFA) amino acid derivatives and diamide phases (left l,l right l,d )... 

A detailed study of the carbon-13 n.m.r. spectra of aporphines has appeared,37 and an X-ray analysis of N,0-diacetyl-4-hydroxynornantenine (60) has been carried out.38 The gas-chromatographic and/or mass spectral behaviour of a series of aporphines as their trimethylsilyl or trifluoroacetyl derivatives has been... 

Finally, we have encountered situations where carbon-14 from a labeled pesticide has been incorporated into normal natural products including amino acids. We were interested in seeing if we could separate the normal amino acids from a protein hydrolysate on a single column with sufficient resolution to identify separate amino acids. Figure 11 shows a radiochromatogram of the separation of sixteen standard C-amino acids. Were this a product of a metabolism study, we could then isolate any of these fractions, convert the residual amino acid to the N-trifluoroacetyl 0-butyl derivative for gas chromatography ar.d further confirmation of structure. 

The standard method of preparing anhydro-5-hydroxyoxazolium hydroxides (300) is by dehydration of iV-substituted a-acylamino acids, usually with acetic anhydride (equation 154) dicyclohexylcarbodiimide and trifluoroacetic anhydride have also been used. N-Acylglycines (299 R3=H) give rise to unstable mesoionic oxazoles which tend to dimerize (see Section 4.18.3.1.5(0) if such acids are treated with trifluoroacetic anhydride, stable trifluoroacetyl derivatives (300 R3 = CF3CO) are isolated. Numerous unstable betaines (300 R1, R = alkyl or aryl, R -H) have been generated by the action of triethylamine on the corresponding hydroperchlorates, prepared by the method shown in equation (151). 

Of halogenated acetyl derivatives, trifluoroacetyl derivatives are mainly used for the sensitive analysis of amines, particularly owing to their better chromatographic properties despite their ECD response being lower than that of chloroacetyl derivatives (cf., Table 4.7, p. 69). Trifluoroacetyl derivatives were exploited for the GC separation of a mixture of saturated and unsaturated homologues of amines up to C22 on conventional packed columns [71]. Mori et al. [72] developed a method for the quantitative and qualitative GC analysis of m- and p-xylenediamines in polyamides in the form of their t N -trifluoro-acetyl derivatives. An analogous method was elaborated for the analysis of ethanolamine for the presence of mono-, di- and triethanolamine [73]. A 1-ml volume of TFA anhydride is stoppered in a 2-ml vial, which is evacuated with the aid of a 10-ml injection syringe. 

Hajdu P, Schmidt D, Bomm M, Keller A (1984) Determination of (2-[N-[(S)-l-Ethoxycarbonyl-3-phenylpropyl]-L-alanyl]-(1S,3S,5S)-2-azabicyclo[3.3.0]octane-3-carboxylic acid) (HOE 498) and its hydrolysis product in serum and urine. Arznm Forsch/Drug Res 34 1431-1435 Schmidt D, Keller A (1985) Sensitive determination of the ACE-Inhibitor HOE 498 and its metabolites in human urine by capillary GC. Fresenius Z Anal Chem 320 731 Walle T, Ehrsson H, Bogentoft C, Dolby J (1972) Synthesis and structure of an enaminic bis(trifluoroacetyl) derivatives of desipramine. Acta Pharm Suecica 9 509-512... 

The N-H X (X = N, O, S) intramolecular hydrogen bonding in the series of 2-heteroarylpyrroles and their trifluoroacetyl derivatives was examined by the NMR spectroscopy and density functional... 

One of the first chromatographic separation of enantiomers was by Gil-Av, who used gas chromatography to separate the trifluoroacetyl derivatives of some amino acids on an optically active stationary phase consisting of N-trifluoroacetyl-L-phenylalanine cyclohexyl ester. It was noted that the L isomers eluted last on the L stationary phase but first on the D stationary phase. Gas chromatography has not proved the ideal technique for separating optically active isomers as all solutes... 

Oiamide Chiral Separations. The first chiral stationary phase for gas chromatography was reported by Gil-Av and co-workers in 1966 (113) and was based on N-trifluoroacetji (iV-TFA) L-isoleucine lauryl ester coated on an inert packing material. It was used to resolve the trifluoroacetylated derivatives of amino acids. Related chiral selectors used by other workers included -dodecanoyl-L-valine-/-butylamide and... 

Several factors affect the volatility and stability of a peptide derivative, not least of these being the number and nature of the constituent amino acids. Heterocyclic and aromatic amino acids reduce volatility while those containing sulphur tend to decrease the thermal stability. Small naturally occurring peptides which are not derived from proteins often contain only aliphatic amino acids which lack functional groups in the side chains. Peptides of this type of up to about ten amino acids, after conversion to suitable derivatives, are amenable to analysis by mass spectrometry, e.g. [164]. A variety of derivatives has been reported and include N-trifluoroacetyl peptide esters [136,165], N-acetyl peptide esters [166-168], aromatic N-acyl peptide esters [169-172], and per-methylated N-acyl peptides [173]. The principal modes of the electron impact induced fragmentation of these peptide derivatives are well established and have been summarised in recent reviews [174,175]. Although the spectra of the permethylated derivatives [176] are perhaps the simplest and easiest to interpret and are now frequently used, the N-acyl peptide esters have been widely and successfully employed. 

The stable nonvolatile intermediate phenylthiocar-bamoyl derivatives are formed in basic media and can be analyzed directly by reverse-phase high-performance liquid chromatography (RP-HPLC). Their cyclization into hydantoins requires acid catalysis. This mode of derivatization is a very important supplement to the Edman s method of N-terminated sequencing of polypeptides. Before GC analysis, any hydantoins can be converted into N-trifluoroacetyl or enol-O-trimethylsilyl derivatives, which increases the selectivity of their determination in complex matrices. 

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