beta-Hydroxymyristic acid as a chemical marker to detect endotoxins in dialysis water


  • Mishra Rupesh K.
  • Robert-Peillard Fabien
  • Ravier Sylvain
  • Coulomb Bruno
  • Boudenne Jean-Luc

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An analytical chemical method has been developed for determination of beta-hydroxymyristic acid (beta-HMA), a component of lipopolysaccharides (LPSs/endotoxins) in dialysis water. In our investigation, the beta-HMA component was used as a chemical marker for endotoxin presence in dialysis water because it is available in the molecular subunit (lipid A) and responsible for toxicity. It is the most abundant saturated fatty acid in that subunit. The developed method is based on fluorescence derivatization with 4-nitro-7-piperazino-2,1,3-benzoxadiazole (NBD-PZ). A high-performance liquid chromatographic separation of the beta-HMA derivative was achieved using an octadecyl silica column in gradient elution. A wide dynamic range of beta-HMA was tested and a calibration curve was constructed with accuracy of 90% and variability of less than 10%. The limits of detection and quantification obtained were 2 and 51 mu M, respectively. The developed method was applied to detect endotoxins in dialysis water by alkaline hydrolysis of LPS using NaOH (0.25 M) at 60 degrees C for 2 h. After hydrolysis, free acid was detected as its NBD-PZ derivative using high-performance liquid chromatography/mass spectrometry (HPLC/MS). Good recovery rates ranging from 98 to 105% were obtained for beta-HMA in dialysis water. (C) 2014 Elsevier Inc. All rights reserved.

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