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

Aug 27, 2016Journal of chromatography. A

Using a fluorescent method with water-based extraction to measure butyrobetaine, l-carnitine, and acetyl-l-carnitine in human blood plasma

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Abstract

The detection limit for l-carnitine and acetyl-l-carnitine was 4.5 fmol.

A novel method utilizing aqueous solvent-based dispersive liquid-liquid microextraction enhances the detection of hydrophilic compounds.
Linear response for l-carnitine and acetyl-l-carnitine spans from 10 to 2000 nM with a determination coefficient greater than 0.998.
Good reproducibility is indicated by a relative standard deviation of better than 7.50% for slope and 9.06% for intercept in linear regression.
The method successfully characterizes butyrobetaine, l-carnitine, and acetyl-l-carnitine in human plasma with determination coefficients ranging from 0.996 to 0.999.
The approach requires a small sample volume and reduces the use of toxic solvents while maintaining high sensitivity.

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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.

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

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