Influence of Co-Dosed Lipids from Biota Extracts on the Availability of Chemicals in In Vitro Cell-Based Bioassays

Abstract

Extraction of chemicals from biota leads to co-extraction of lipids. When dosing such extracts into in vitro bioassays, co-dosed lipids act as an additional phase that can reduce the bioavailability of the chemicals and the apparent sensitivity of the assay. Equilibrium partitioning between medium, cells, and co-dosed lipids was described with an existing equilibrium partitioning model for cell-based bioassays extended by an additional lipid phase. We experimentally investigated the influence of co-dosed lipids on the effects elicited by four test chemicals of different hydrophobicity in two bioassays, indicative of the aryl hydrocarbon receptor and oxidative stress response (AREc32). The partitioning model explained the effect of the test chemicals in the presence of spiked triolein within a factor of 0.33−5.83 between the measured and predicted effect concentration (EC). We applied the model to marine mammal blubber extracted with silicone. Extracts dosed in the AREc32 bioassay showed a linear increase of apparent EC with increasing lipid fraction. The partitioning model was used to interpret the role of the co-extracted lipid. A quantitative lipid correction of bioassay results in the presence of co-dosed lipids was possible for known compounds and defined mixtures, while we could only estimate a range for mixtures of unknown chemicals.

Kinetics of Equilibrium Passive Sampling of Organic Chemicals with Polymers in Diverse Mammalian Tissues

“…There is one caveat, though, when the extract with the coextracted lipid is dosed into bioassays; there is also partitioning between the codosed lipid and the bioassay medium, and the overall sensitivity of the bioassay is decreased in comparison to dosing the same amount of the extract but without the codosed lipid . As Figure B demonstrates, there is little difference in analytically measured PCB126 concentrations and their bioanalytical equivalents PCB126-EQ, but this can change in samples with lower levels of contamination, where a non-negligible amount of lipids is dosed to the bioassay …”

“…Even if that ratio could be decreased to 1:1, sampling would remain nondepletive. This is not a problem, provided that the detection limits are low enough and the small fraction of coextracted lipids does not interfere with the bioassays …”

“…The extraction method is ultimately aimed at extracting complex mixtures from tissues for analysis with in vitro bioassays. Therefore, uptake kinetic experiments from the liver tissue in PDMS were conducted with PCB126 as a bioactive hydrophobic model chemical, which acts as an agonist in the AhR CALUX (chemically activated luciferase expression) bioassay. − The homogenized liver tissue was spiked with PCB126 at concentrations of 240–570 ng PCB126 g liver –1 , and passive sampling was carried out statically as well as dynamically with the described stirrer. The concentrations of PCB126 were quantified in the extracts with GC-MSD (Text S6), and the extracts were submitted to the AhR CALUX in vitro bioassay for the activation of the aryl hydrocarbon receptor (AhR) assay (Text S7).…”

“…48 As Figure 6B demonstrates, there is little difference in analytically measured PCB126 concentrations and their bioanalytical equivalents PCB126-EQ, but this can change in samples with lower levels of contamination, where a non-negligible amount of lipids is dosed to the bioassay. 48 Recommendations for Unbiased Extractions of Mixtures of Organic Chemicals. Passive equilibrium sampling has been applied so far only to tissue sampling of POPs, but we were able to demonstrate that the range of applicability can be extended to cover eight orders of magnitude of hydrophobicity expressed by log K ow with very little difference in extraction efficiency expressed by log K PDMS/lipid or log K PDMS/tissue .…”

Bioanalytical Tools in Water Quality Assessment

“…It must be noted, however, that even small fractions of coextracted lipids decrease the sensitivity of the in vitro bioassays (Reiter et al, 2020). This is caused by decreased bioavailability of the contaminants, as they are retained in the co-dosed lipids in a similar manner as the serum-mediated passive dosing (SMPD) described in Chapter 9.…”