Quantitation of Ethylene Glycol in Postmortem Specimens by GC-MS/MS

Significance of the Topic

Ethylene glycol is commonly used in antifreeze but poses a severe poisoning risk when ingested. Accurate quantitation in postmortem samples is vital for forensic toxicology, clinical diagnosis and legal investigations due to its rapid metabolism and life-threatening effects.

Objectives and Study Overview

The study aimed to develop and validate a GC-MS/MS method using a deuterated internal standard (EG-d4) to simultaneously identify and quantify ethylene glycol in postmortem specimens following SWGTOX guidelines. This approach addresses limitations of earlier GC-FID and single-quadrupole GC-MS methods.

Methodology

Blood and biological specimens (200 µL) were spiked with EG-d4 and proteins were precipitated with cold acetonitrile. After centrifugation, the supernatant was reacted with 2,2-dimethoxypropane:DMF, dried under nitrogen at 80 °C, derivatized with MTBSTFA/1% t-BDMCS, and extracted with ethyl acetate. A 1 µL aliquot was injected for GC-MS/MS analysis under time-scheduled MRM.

Used Instrumentation

• Bruker 436 GC equipped with a 30 m × 0.25 mm i.d. × 0.25 µm DB-5MS column (Agilent)
• Carrier gas: Helium at 1 mL/min; splitless injection at 250 °C; oven program: 80 °C (1 min), ramp 20 °C/min to 315 °C (1.25 min)
• Bruker EVOQ GC-TQ triple-quadrupole mass spectrometer; source 200 °C; CID gas 1.8 mTorr; MS WorkStation software

Main Results and Discussion

The method exhibited linearity from 25 to 2000 mg/L (1/x2 weighting), with an LOD of 10 mg/L and LOQ of 25 mg/L. Inter- and intra-day precision at 50 and 1000 mg/L were ≤1% RSD, with biases of –2.0% and +2.6%. No carryover was detected after a 2000 mg/L calibration standard. Dilution integrity at 1:1 remained within 5% bias and precision. No interference or cross-talk was observed among 80+ compounds, and samples were stable for 12 h at ambient temperature.

Benefits and Practical Applications

This GC-MS/MS approach offers enhanced specificity, sensitivity and throughput for forensic and clinical laboratories. The use of a deuterated standard and MRM transitions ensures robust quantitation in complex biological matrices. Implementation can streamline toxicology workflows and improve result reliability.

Future Trends and Potential Applications

Further developments may include expansion to other polyol toxins, integration with high-resolution mass spectrometry for non-target screening, miniaturized sample preparation and automation. Adaptation to field-deployable instruments could enhance rapid on-site forensic analysis.

Conclusion

A validated GC-MS/MS method using EG-d4 provides accurate, precise and interference-free quantitation of ethylene glycol in postmortem specimens. Adopted by the Miami-Dade Medical Examiner Department, this protocol improves forensic casework and proficiency testing since 2012.

Reference

• Porter W, Rutter P, Yao H. Simultaneous Determination of Ethylene Glycol and Glycolic Acid in Serum by Gas Chromatography-Mass Spectrometry. J Anal Toxicol. 1999;23:591-597.
• Rosano T, Swift T, Kranick C, Sikirica M. Ethylene Glycol and Glycolic Acid in Postmortem Blood from Fatal Poisonings. J Anal Toxicol. 2009;33:508-513.
• Sun Q, Wang K. Demonstration of Fast Scan Time and No Cross-Talk on SCION TQ Triple Quadrupole Mass Spectrometer. Bruker App Note CA 281752; 2012.
• SWGTOX. Standard Practices for Method Validation in Forensic Toxicology. J Anal Toxicol. 2013;37:452-474.