High-throughput analysis using non-depletive SPME: challenges and applications to the determination of free and total concentrations in small sample volumes

Abstract

In vitro high-throughput non-depletive quantitation of chemicals in biofluids is of growing interest in many areas. Some of the challenges facing researchers include the limited volume of biofluids, rapid and high-throughput sampling requirements, and the lack of reliable methods. Coupled to the above, growing interest in the monitoring of kinetics and dynamics of miniaturized biosystems has spurred the demand for development of novel and revolutionary methodologies for analysis of biofluids. The applicability of solid-phase microextraction (SPME) is investigated as a potential technology to fulfill the aforementioned requirements. As analytes with sufficient diversity in their physicochemical features, nicotine, N,N-Diethyl-meta-toluamide, and diclofenac were selected as test compounds for the study. The objective was to develop methodologies that would allow repeated non-depletive sampling from 96-well plates, using 100 µL of sample. Initially, thin film-SPME was investigated. Results revealed substantial depletion and consequent disruption in the system. Therefore, new ultra-thin coated fibers were developed. The applicability of this device to the described sampling scenario was tested by determining the protein binding of the analytes. Results showed good agreement with rapid equilibrium dialysis. The presented method allows high-throughput analysis using small volumes, enabling fast reliable free and total concentration determinations without disruption of system equilibrium.

Solid-phase microextraction: a fit-for-purpose technique in biomedical analysis

“…While HS-SPME is well-established and has become a standard approach in volatilome analysis, direct-immersion SPME (DI-SPME) is still only used irregularly, despite findings showing its ability to fill the gaps associated with “traditional” approaches. One of SPME’s main advantages is that it allows the same biological system to be sampled multiple times thanks to its low invasiveness and negligible depletion (under certain conditions) [ 7 ]. In the recent work, time-course analysis of the cellular secretome using two- and three-dimensional cell models (2D — cells grow on flat surfaces as monolayers, and 3D — e.g., hanging drops, spheroids, or more complex systems mimicking in vivo conditions), as well as an in vivo model of mouse melanoma, was performed using biocompatible probes with 2 mm long hydrophilic-lipophilic balanced (HLB) coatings [ 8 ].…”

Bridging the Data Gap From in vitro Toxicity Testing to Chemical Safety Assessment Through Computational Modeling

“…Therefore the chemical concentration, particularly the free concentrations, in the culture media and cells during the course of the in vitro experiment should be determined whenever possible. As high-throughput non-depletive methods of quantifying chemicals in miniaturized assays improve, determining chemical concentration kinetics is becoming increasingly practical ( 44 ). In some cases, an in vitro kinetic model describing the chemical concentration changes in the culture medium and cells over time can be constructed (Figure 1 ).…”

Aroma Clouds of Foods: A Step Forward to Unveil Food Aroma Complexity Using GC × GC

“…This technique was developed by Pawliszyn, in the beginning of the 1990 decade ( Pawliszyn, 2009 ), and may be used to extract directly analytes from liquid, solid, and gaseous samples. SPME is a non-exhaustive extraction technique since it only extracts a small analyte fraction, which may be used to characterize the global composition of analytes in the free form ( Vas and Vékey, 2004 ; Pawliszyn, 2009 ; Souza-Silva et al, 2015 ; Boyacl et al, 2018 ). These characteristics allow electing the SPME as a technique especially well positioned to extract the odorant molecules present in the food aroma clouds.…”