Although nerve agent use is prohibited, concerns remain for human exposure to nerve agents during decommissioning, research, and warfare. Exposure can be detected through the analysis of the hydrolysis products in urine as well as blood. An analytical method to detect exposure to five nerve agents, including VX, VR (Russian VX), GB (sarin), GD (soman) and GF (cyclosarin), through the analysis of the hydrolysis products, which are the primary metabolites, in serum has been developed and characterized. This method uses solid phase extraction coupled with high performance liquid chromatography for separation and isotopic dilution tandem mass spectrometry for detection. An uncommon buffer of ammonium fluoride was used to enhance ionization and improve sensitivity when coupled with hydrophilic interaction liquid chromatography resulting in detection limits from 0.3–0.5 ng/mL. The assessment of two quality control samples demonstrated high accuracy (101–105%) and high precision (5–8%) for the detection of these five nerve agent hydrolysis products in serum.

An automated dried blood spot (DBS) elution coupled with solid phase extraction and tandem mass spectrometric analysis for multiple fentanyl analogs was developed and assessed. This method confirms human exposures to fentanyl, sufentanil, carfentanil, alfentanil, lofentanil, α-methyl fentanyl, and 3-methyl fentanyl in blood with minimal sample volume and reduced shipping and storage costs. Seven fentanyl analogs were detected and quantitated from DBS made from venous blood. The calibration curve in matrix was linear in the concentration range of 1.0 ng/mL to 100 ng/mL with a correlation coefficient greater than 0.98 for all compounds. The limit of detection varied from 0.15 ng/mL to 0.66 ng/mL depending on target analyte. Analysis of the entire DBS minimized the effects of hematocrit on quantitation. All quality control materials evaluated resulted in <15% error; analytes with isotopically labeled internal standards had <15% RSD, while analytes without matching standards had 15–24% RSD. This method provides an automated means to detect seven fentanyl analogs, and quantitate four fentanyl analogs with the benefits of DBS at levels anticipated from an overdose of these potent opioids.

Biomedical samples may be used to determine human exposure to nerve agents through the analysis of specific biomarkers. Samples received may include serum, plasma, whole blood, lysed blood and, due to the toxicity of these compounds, postmortem blood. To quantitate metabolites resulting from exposure to sarin (GB), soman (GD), cyclosarin (GF), VX and VR, these blood matrices were evaluated individually for precision, accuracy, sensitivity and specificity. Accuracies for these metabolites ranged from 100 to 113% with coefficients of variation ranging from 2.31 to 13.5% across a reportable range of 1-100 ng/mL meeting FDA recommended guidelines for bioanalytical methods in all five matrices. Limits of detection were calculated to be 0.09-0.043 ng/mL, and no interferences were detected in unexposed matrix samples. The use of serum calibrators was also determined to be a suitable alternative to matrix-matched calibrators. Finally, to provide a comparative value between whole blood and plasma, the ratio of the five nerve agent metabolites measured in whole blood versus plasma was determined. Analysis of individual whole blood samples (n = 40), fortified with nerve agent metabolites across the reportable range, resulted in average nerve agent metabolite blood to plasma ratios ranging from 0.53 to 0.56. This study demonstrates the accurate and precise quantitation of nerve agent metabolites in serum, plasma, whole blood, lysed blood and postmortem blood. It also provides a comparative value between whole blood and plasma samples, which can assist epidemiologists and physicians with interpretation of test results from blood specimens obtained under variable conditions.

A method was developed to detect and quantify organophosphate nerve agent (OPNA) metabolites in dried blood samples. Dried blood spots (DBS) and microsampling devices are alternatives to traditional blood draws, allowing for safe handling, extended stability, reduced shipping costs, and potential self-sampling. DBS and microsamplers were evaluated for precision, accuracy, sensitivity, matrix effects, and extraction recovery following collection of whole blood containing five OPNA metabolites. The metabolites of VX, Sarin (GB), Soman (GD), Cyclosarin (GF), and Russian VX (VR) were quantitated from 5.0 to 500 ng mL with precision of ≤16% and accuracy between 93 and 108% for QC samples with controlled volumes. For unknown spot volumes, OPNA metabolite concentrations were normalized to total blood protein to improve interpretation of nerve agent exposures. This study provides data to support the use of DBS and microsamplers to collect critical exposure samples quickly, safely, and efficiently following large-scale chemical exposure events.