Validation of a high performance liquid chromatography-tandem mass spectrometry method for β-hydroxy fatty acids as environmental markers of lipopolysaccharide

• β-Hydroxy dodecanoic and tetradecanoic acids are LPS markers in GN strains.
• A new method is validated to quantify β-hydroxy-fatty acids in bacterial suspensions.
• Results of HPLC-MS/MS and LAL-test on 4 bacterial strains are linearly correlated.
• Results are not affected by β-glucans, nor by repeated sample freezing and thawing.
• It is useful for the assessment of the occupational exposure to airborne endotoxins.

The most commonly analytical method used for the assessment of the occupational exposure to airborne endotoxins is the LAL test, but other methods are reported in the literature.The objective of this work was to develop and validate a method using HPLC coupled to triple quadrupole mass spectrometer for the quantitative determination of β-hydroxy-dodecanoic and β-hydroxy-tetradecanoic acids as chemical markers of lipopolysaccharide in Gram-negative strains. Matrix effect was significant but compensated by the use of internal standard; accuracy was always more than 85% and variability always less than 10%. LOD and LLOQ were respectively 2.5 μg/l and 10 μg/l for β-hydroxy-dodecanoic acid, 1 μg/l and 5 μg/l for β-hydroxy-tetradecanoic acid.The analysis of different bacterial strains showed that samples contain β-hydroxy-dodecanoic and β-hydroxy-tetradecanoic acids in variable ratio, and therefore the sum of their concentrations was considered the chemical marker. The method showed a significant correlation of results with those of LAL-test on four bacterial strains.