Fluorescent derivatization combined with aqueous solvent-based dispersive liquid-liquid microextraction for determination of butyrobetaine, l-carnitine and acetyl-l-carnitine in human plasma

J Chromatogr A. 2016 Sep 16:1464:32-41. doi: 10.1016/j.chroma.2016.08.030. Epub 2016 Aug 22.

Abstract

A novel aqueous solvent-based dispersive liquid-liquid microextraction (AS-DLLME) method was combined with narrow-bore liquid chromatography and fluorescence detection for the determination of hydrophilic compounds. A remover (non-polar solvent) and extractant (aqueous solution) were introduced into the derivatization system (acetonitrile) to obtain a water-in-oil emulsion state that increased the mass transfer of analytes. As a proof of concept, three quaternary ammonium substances, including butyrobetaine, l-carnitine and acetyl-l-carnitine, were also used as analytes and determined in pharmaceuticals, personal care products, food and human plasma. The analytes were derivatized with 4-bromomethylbiphenyl for fluorescence detection and improved retention in the column. The linear response was 10-2000nM for l-carnitine and acetyl-l-carnitine with a good determination coefficient (r(2)>0.998) in the standard solution. The detection limit for l-carnitine and acetyl-l-carnitine was 4.5 fmol. The method was also successfully applied to a 1μL sample of human plasma. In the linearity calculations for determining butyrobetaine, l-carnitine and acetyl-l-carnitine in human plasma, the determination coefficients ranged from 0.996 to 0.999. Linear regression exhibited good reproducibility and a relative standard deviation better than 7.50% for the slope and 9.06% for the intercept. To characterize highly hydrophilic compounds in various samples, the proposed method provides good sensitivity for a small sample volume with a low consumption of toxic solvents.

Keywords: Aqueous solvent-based dispersive liquid-liquid microextraction; Fluorescence derivatization; Narrow-bore liquid chromatography; l-carnitine.

Publication types

  • Evaluation Study

MeSH terms

  • Betaine / analogs & derivatives*
  • Betaine / blood
  • Betaine / chemistry
  • Betaine / isolation & purification
  • Carnitine / blood*
  • Carnitine / chemistry
  • Carnitine / isolation & purification*
  • Chromatography, Liquid / methods
  • Fluorescence
  • Humans
  • Hydrophobic and Hydrophilic Interactions
  • Limit of Detection
  • Liquid Phase Microextraction / instrumentation
  • Liquid Phase Microextraction / methods*
  • Reproducibility of Results

Substances

  • Betaine
  • gamma-butyrobetaine
  • Carnitine