Integrated Exposomics/Metabolomics for Rapid Exposure and Effect Analyses

Abstract

The totality of environmental exposures and lifestyle factors, commonly referred to as the exposome, is poorly understood. Measuring the myriad of chemicals that humans are exposed to is immensely challenging, and identifying disrupted metabolic pathways is even more complex. Here, we present a novel technological approach for the comprehensive, rapid, and integrated analysis of the endogenous human metabolome and the chemical exposome. By combining reverse-phase and hydrophilic interaction liquid chromatography (HILIC) and fast polarityswitching, molecules with highly diverse chemical structures can be analyzed in 15 min with a single analytical run as both column's effluents are combined before analysis. Standard reference materials and authentic standards were evaluated to critically benchmark performance. Highly sensitive median limits of detection (LODs) with 0.04 μM for >140 quantitatively assessed endogenous metabolites and 0.08 ng/mL for the >100 model xenobiotics and human estrogens in solvent were obtained. In matrix, the median LOD values were higher with 0.7 ng/mL (urine) and 0.5 ng/mL (plasma) for exogenous chemicals. To prove the dualcolumn approach's applicability, real-life urine samples from sub-Saharan Africa (high-exposure scenario) and Europe (low-exposure scenario) were assessed in a targeted and nontargeted manner. Our liquid chromatography high-resolution mass spectrometry (LC-HRMS) approach demonstrates the feasibility of quantitatively and simultaneously assessing the endogenous metabolome and the chemical exposome for the high-throughput measurement of environmental drivers of diseases.

Phospholipid Removal for Enhanced Chemical Exposomics in Human Plasma

“…This was likely related to the higher injection volumes used in the exposomics protocol, which led to poorer chromatographic retention for some of these earliest eluting polar compounds, as discussed above for acesulfame and acephate-acetyl- d 3 (non-targeted examples in Figure S18). In contrast with the late eluting hydrophobic features, this early chromatographic region contains very hydrophilic substances which are most likely better monitored in urine samples and with alternative chromatographic modes (e.g., HILIC) . Acesulfame for instance is typically analyzed in urine where it is exclusively excreted in humans. , In agreement with these observations, the majority of level 2 non-phospholipid features that were present only in the control protocol eluted either before 4 min or after 18 min (20 out of 35 features, Table S7).…”

“…Application of both methods to the same samples may be limited by sample volumes, as well as cost and analysis time considerations. Applications of the current exposomics method will depend on case-specific hypotheses and resources, and increased efficiencies might be gained in future by introducing polarity switching, as others have shown for combined exposomics and metabolomics …”

“…Applications of the current exposomics method will depend on case-specific hypotheses and resources, and increased efficiencies might be gained in future by introducing polarity switching, as others have shown for combined exposomics and metabolomics. 12 Compared with the control method, the current exposomics method requires additional steps, cost, and time, but it is only notable that the commercial cartridges used here are already offered in 96 well format. 47 Thus, the current technique is scalable and can feasibly be adapted to liquid handling robotics with some modifications.…”

“…Current analytical trends in chemical exposomics include multiclass targeted methods based on liquid chromatography (LC) or gas chromatography (GC) with mass spectrometry, as well as suspect-screening and non-targeted LC- and GC-high resolution mass spectrometry (HRMS). − Non-targeted HRMS approaches are common in metabolomics and are promising for chemical exposomics because they allow profiling and quantification of known substances, while also acquiring spectra of unknown molecules that may be unanticipated biomarkers of disease. Most chemical exposomics approaches in human studies have so-far been adapted directly from high-resolution metabolomics, , involving only a protein precipitation step and injecting the equivalent of 1–5 μL of plasma on-column. ,,, While these methods have advantages for throughput and unbiased molecular analysis, they do not address the 1,000-fold concentration gap for environmental contaminants in plasma …”

“…Most chemical exposomics approaches in human studies have so-far been adapted directly from high-resolution metabolomics, 13,14 involving only a protein precipitation step and injecting the equivalent of 1−5 μL of plasma on-column. 8,9,11,12 While these methods have advantages for throughput and unbiased molecular analysis, they do not address the 1,000-fold concentration gap for environmental contaminants in plasma. 3 One way to improve sensitivity of chemical exposomics is to introduce more environmental analytes on-column, either through preconcentration steps or large-volume injection.…”