One‐step methodology for the synthesis of FA picolinyl esters from intact lipids

Abstract

Picolinyl derivatives are used for structural determination of FA by GC-MS. Although they provide reliable diagnostic fragments, the usual multistep methodologies applied for their preparation require TAG hydrolysis or acid chloride formation prior to picolinyl synthesis. These reaction conditions may result in the presence of artifact molecules in the samples and thus compromise analytical quality and accuracy. To address these problems, a rapid, simple and quantitative methodology for the synthesis of FA picolinyl esters from intact lipids was developed. It involves their transesterification under basecatalyzed conditions using 3-potassiooxamethylpyridine in methylene chloride. The catalyst was prepared by proton exchange between potassium tert-butoxide and anhydrous 3-hydroxymethylpyridine. Mild reaction conditions allowed complete derivatization of TAG and phospholipids in 2 min at room temperature, and of FAME in 15 min at 45°C. The proposed procedure, which can be used on a routine basis, was applied to Ipomoae imperialis seed lipids and used to confirm occurrence of γ-linoleic acid at a level of 0.9%.Paper no. J10108 in JAOCS 79, 253-256 (March 2002). KEY WORDS:Alkoxide, fatty acid picolinyl ester, gas-liquid chromatography, Ipomoae seed oil, γ-linolenic acid, mass spectrometry, mass spectroscopy, potassium tert-butoxide, transesterification.Derivatization is a necessary step prior to structural determination of FA by GC-MS. However, mass spectra of FAME, the usual derivatives for GLC-FID analysis, are devoid of sufficient information for identification of structural isomers. This is mainly due to the high sensitivity of the carboxyl group to fragmentation and to double bond migration (1,2). However, stabilization of the carboxyl group by the formation of a derivative containing a nitrogen atom results in mass spectra that allow structural determination for most FA (1). A first such derivative used in GC-MS analysis was pyrrolidide (3), but structural determination of FA by this technique is now usually performed using picolinyl and 4,4-dimethyloxazoline derivatives. Indeed, these FA derivatives provide diagnostic fragments that allow accurate structure determination (1). However, the usual multistep methodologies for the synthesis of picolinyl esters require hydrolysis of the lipid sample prior to final derivatization, either by condensation of the formed acid chloride with 3-hydroxymethylpyridine (1) or through an imidazole intermediate (4). The latter methodology was developed for the analysis of sensitive FA containing polyethylenic bond systems or an epoxy group. Both procedures give accurate qualitative and quantitative results but require long reaction times, which do not always lead to complete derivatization, and moderate to high temperatures that may result in artifacts. The aim of this study was to develop a short, simple, and quantitative procedure for the synthesis of FA picolinyl esters in mild conditions and from O-acetylated acids. The proposed methodology was applied to confirm occurrence ...

Vertebrate fatty acyl desaturase with Δ4 activity

“…FAME were prepared, extracted, and purified by thin-layer chromatography (19), and picolinyl esters were prepared from FAME (48). Briefly, FAME samples dissolved in 1 mL dry dichloromethane were added to a mixture of 0.1 mL potassium tert-butoxide in tetrahydrofuran (1.0 M) and 0.2 mL 3-pyridylcarbinol.…”

Molecular Analysis of a Bifunctional Fatty Acid Conjugase/Desaturase from Tung. Implications for the Evolution of Plant Fatty Acid Diversity

“…The column temperature was programmed as described for GC/FID. GC/MS analysis of picolinyl ester derivatives was performed as described (Destaillats and Angers, 2002). For HPLC/PDA analysis, FAME were separated on an HPLC system (Waters Corp., Milford, MA) equipped with a multisolvent delivery system (model 600E), autosampler (model 712), tunable absorbance detector (UV; model 486), and PDA (model 996).…”

Picolinyl ester fragmentation mechanism studies with application to the identification of acylcarnitine acyl groups following transesterification

“…The method of Destaillats and Angers [11] for lipids was adapted for transesterification of acylcarnitines. Briefly, a solution (200 L) containing 1 mg/mL of an acylcarnitine in methanol was evaporated to dryness under nitrogen at room-temperature.…”

“…However, the presence of organic acids in urine makes questionable the interpretation of the source of acyl groups as being exclusively from acylcarnitines. Although acyl picolinyl esters usually are produced from organic acids, transesterification has been reported to produce acyl picolinyl esters from lipids by direct reaction with 3-pyridylcarbinol, thus eliminating the need to hydrolyze the lipids first [11].In this paper, we propose mechanisms for the formation of the fragment ions of acyl picolinyl esters based on experiments using deuterium-labeled organic acids. We applied these results to the characterization of acyl groups of several acylcarnitine species following their transesterification to acyl picolinyl esters.…”

“…However, the presence of organic acids in urine makes questionable the interpretation of the source of acyl groups as being exclusively from acylcarnitines. Although acyl picolinyl esters usually are produced from organic acids, transesterification has been reported to produce acyl picolinyl esters from lipids by direct reaction with 3-pyridylcarbinol, thus eliminating the need to hydrolyze the lipids first [11].…”