Separation and direct detection of long chain fatty acids and their methylesters by the non-aqueous reversed phase HPLC and Silver Ion Chromotography, combined with CO laser pumped thermal lens spectrometry

D.D. Bicanic, G. Mocnik, M. Franko, H.A.G. Niederlander, P. van de Bovenkamp, J.L. Cozijnsen, E.J.C. van der Klift

Research output: Contribution to journalArticleAcademicpeer-review

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Abstract

The potential of the CO laser pumped dual beam thermal lens spectrometer (TLS) used as the detector of infrared (IR) absorbance in non-aqueous reversed-phase high pressure liquid chromatography (NARP-HPLC) and argentation chromatography (Ag-HPLC-TLS) has been investigated. The linoleic acid C18:2 (9,11) cis, cis, the conjugated linoleic acid C18:2 (9,12) cis, cis, and (9,11) cis, cis and oleic acid C18:1 (9) cis free fatty acids present in the test mixture were separated and detected by NARP-HPLC-TLS directly, i.e., without a need for their prederivatization. The limit of detection (LOD) for the present NARP-HPLC-TLS set-up was determined using either oleic acid C18:1 (9) cis or linoleic acid C18:2 (9,12) cis, cis as the only analyte. The LOD obtained at 1750 cm-1 for C18:2 (9,12) cis, cis is 0.14% (v/v) with the standard deviation being 3.8% of the measured value. This is comparable to the LOD attainable by the bulk property refractive index detector (RID). For C18:1 (9) cis the LOD is 0.04% (v/v) with a standard deviation of 2.5%. The unique feature of the NARP-HPLC-TLS is its excellent selectivity; this was demonstrated experimentally by adding alcohols (octanol and decanol) that lack the C&dbnd;O carbonyl group to the aforementioned mixture of three fatty acids. In another experiment, the Ag-HPLC-TLS in IR was used to directly detect two different mixtures of five fatty acid methyl esters (FAMEs) after separation on the column. The results were compared to those achieved by the conventional UV absorbance detector. Despite more favourable excitation cross sections of FAMEs in the IR, the concentration LOD obtained for Ag-HPLC-TLS is inferior to that of the Ag-HPLC-UV. However, the mass LOD (11 µg) achieved by Ag-HPLC-TLS in IR for C18:3 FAME is two times better than that of the Ag-HPLC-UV absorbance detector. High IR absorption of eluent restricts, at present, the sensitivity of the Ag-HPLC-TLS and NARP-HPLC-TLS. Improved LOD's are anticipated by a judicious choice of the mobile phase and after materialising several modifications of the experimental set-up.
Original languageEnglish
Pages (from-to)129-150
Number of pages21
JournalInstrumentation Science and Technology
Volume34
Issue number1-2
DOIs
Publication statusPublished - 2006

Keywords

  • performance liquid-chromatography
  • beta-carotene
  • lipids
  • triacylglycerols
  • deflection
  • oils

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