Determination of Drugs in Human Brain via Bidirectional Solid-Phase Extraction

Significance of the Topic

In forensic toxicology, determining drug presence in post-mortem brain tissue is critical for understanding cause of death and drug distribution. Brain matter presents high fat and protein content, challenging conventional extraction and requiring robust, contamination-free methods. Automated bidirectional solid-phase extraction (BD-SPE) offers a novel approach to enhance reliability and throughput in such demanding matrices.

Objectives and Overview of the Study

This study aims to develop and validate an automated BD-SPE workflow using the FREESTYLE system to extract acidic, neutral, and basic drugs from human brain tissue. The approach focuses on minimizing cross-contamination by reversing sample flow through the SPE sorbent and on preserving sample integrity for reanalysis.

Used Methodology and Instrumentation

Sample Preparation:

• Homogenize 2 g of brain tissue with phosphate buffer (pH 7.4).
• Spike samples with internal standards and centrifuge.

BD-SPE Workflow:

• Condition, wash, and dry 3 mL SPE cartridges.
• Aspirate homogenate through the sorbent from bottom to top, then elute back through the sorbent.
• Repeat bidirectional loading for quantitative transfer.

Elution and Analysis:

• Evaporate eluates, reconstitute in ethyl acetate, and derivatize with MSTFA.
• Analyze samples by GC-MS using an Agilent 6890/5973 system and a Zebron ZB-5MSi column.

Main Results and Discussion

The BD-SPE method achieved mean recoveries of 28–75% for a broad analyte panel (n=6). Repeatability RSD ranged from 1.4% to 7.6% and reproducibility RSD from 3.6% to 18.3% (n=8), demonstrating robustness in the complex brain matrix. The bidirectional loading effectively excluded cross-contamination and allowed sample reuse.

Benefits and Practical Applications

Key advantages of the automated BD-SPE workflow include:

• Elimination of sample cross-contamination by preventing direct contact with automation components.
• High throughput and consistency for routine forensic screening.
• Sample preservation for confirmatory or additional analyses.

Future Trends and Potential Applications

Future developments may explore extending BD-SPE to other complex biological matrices (e.g., liver, fat tissue) and coupling with advanced analytical platforms such as LC-MS/MS. Miniaturization of SPE cartridges and integration with machine learning for data interpretation offer additional potential for forensic and clinical investigations.

Conclusion

The automated bidirectional SPE method on the FREESTYLE platform provides a fit-for-purpose solution for reliable, high-throughput extraction of drugs from challenging brain tissue samples. Its robust performance, avoidance of contamination, and sample preservation make it well-suited for forensic laboratories.

Reference

Reiter B., Stimpfl T. Determination of Drugs in Human Brain via Bidirectional Solid-Phase Extraction. Application Note AN0022, LCTech GmbH, November 2019.