Simultaneous determination of perfluoroalkyl substances and bile acids in human serum using ultra-high-performance liquid chromatography–tandem mass spectrometry

Abstract

There is evidence of a positive association between per-and polyfluoroalkyl substances (PFASs) and cholesterol levels in human plasma, which may be due to common reabsorption of PFASs and bile acids (BAs) in the gut. Here we report development and validation of a method that allows simultaneous, quantitative determination of PFASs and BAs in plasma, using 150 μL or 20 μL of sample. The method involves protein precipitation using 96-well plates. The instrumental analysis was performed with ultraperformance liquid chromatography-tandem mass spectrometry (UHPLC-MS), using reverse-phase chromatography, with the ion source operated in negative electrospray mode. The mass spectrometry analysis was carried out using multiple reaction monitoring mode. The method proved to be sensitive, robust, and with sufficient linear range to allow reliable determination of both PFASs and BAs. The method detection limits were between 0.01 and 0.06 ng mL −1 for PFASs and between 0.002 and 0.152 ng mL −1 for BAs, with the exception of glycochenodeoxycholic acid (0.56 ng mL −1). The PFAS measured showed excellent agreement with certified plasma PFAS concentrations in NIST SRM 1957 reference serum. The method was tested on serum samples from 20 healthy individuals. In this proof-of-concept study, we identified significant associations between plasma PFAS and BA levels, which suggests that PFAS may alter the synthesis and/or uptake of BAs.

Associations between perfluoroalkyl substances and serum lipids in a Swedish adult population with contaminated drinking water

“…The comparison of serum lipid levels between the exposed and control areas was based only on water districts, not serum PFAS levels. Therefore, it is not affected by potential confounding related to intestinal reabsorption of PFAS and bile acids [15,16]. However, the participants from the two areas may differ in other aspects than water quality.…”

“…The issue of potential confounding by dietary factors, or reverse causality (by binding of PFAS to lipoproteins), was discussed extensively by EFSA [7]. Moreover, confounding by variability in intestinal reabsorption of PFAS and bile acids, which in turn is associated with serum cholesterol, remains an important issue when exposure to PFAS is assessed by serum PFAS levels [15,16]. PFAS are excreted in bile, but reabsorbed in the intestine to a high extent, and relatively high reabsorption should be associated with higher steady state PFAS levels in serum.…”

Bile Acids Quantification by Liquid Chromatography–Tandem Mass Spectrometry: Method Validation, Reference Range, and Interference Study

“…The relative low sensitivity is a possible limitation of this approach. Recently published methods providing concentration of samples by nitrogen-drying achieved detection limit below 1 ng/mL for most BA, which is >10-fold lower than the LOQ that can be achieved with our assay [30]. However, due to the purpose of obtaining a suitable method for future clinical studies, sensitivity has been outweighed by fast and high throughput analyses.…”

“…Conversely, a more recent investigation, including 20 healthy individuals [11], found that the median values of most BA were higher than those observed in our study. Nonetheless, this difference could be attributable to the different age of the population, which has not been clearly reported in the previous investigation of Salihović et al [30]. Along with the validation procedure and establishment of reference ranges, we also performed interference studies aimed at assessing the matrix effect from in vitro hemolysis, lipemia, and hyper-bilirubinaemia.…”

Perfluoroalkyl Substances are increased in patients with Late-Onset Ulcerative colitis and induce Intestinal Barrier defects ex vivo in Murine Intestinal tissue

“…coupled to a triple quadrupole mass spectrometer as previously described. 22 Briefly, all For statistical analyses of PFAS and BA concentrations, the data was zero imputed, log2…”