Multi-Target Screening and Quantitation of 24 Drugs in Blood, Serum and Urine Using Automated Sample Preparation Coupled Directly to LC-MS/MS

Significance of the Topic

The simultaneous screening and quantitation of multiple drugs in biological matrices is crucial for forensic investigations, clinical diagnostics, and workplace testing. By automating sample preparation and coupling it directly to LC-MS/MS, laboratories can significantly reduce analysis time, minimize human error, and improve reproducibility, ultimately enhancing throughput and data reliability.

Objectives and Study Overview

This study aimed to develop and validate a fully automated workflow for the multi-target screening and quantitation of 24 commonly abused drugs in blood, serum, and urine. The platform integrates a CLAM-2030 sample preparation module directly with a Shimadzu LCMS-8060 system. Key goals included method validation, evaluation of quantitative performance, and demonstration of applicability to real postmortem samples.

Methodology and Instrumentation

Automated sample preparation was conducted by the CLAM-2030 module, which executes a series of programmed steps—vial wetting, internal standard addition, dilution, protein precipitation, mixing, and vacuum filtration—in under five minutes per sample. Prepared extracts were injected (2 µL) into the LCMS-8060. Chromatographic separation employed a Restek Raptor Biphenyl column (2.1 mm × 100 mm, 2.7 µm) with a gradient elution of formic acid/ammonium formate in water and methanol at 0.3 mL/min and 50 °C. Detection used positive and negative ESI in MRM mode with synchronized survey scans for MS/MS confirmation.

Used Instrumentation

• Shimadzu CLAM-2030 Automated Sample Preparation Module
• Shimadzu LCMS-8060 Triple Quadrupole Mass Spectrometer
• Restek Raptor Biphenyl Column (2.1 × 100 mm, 2.7 µm)

Main Results and Discussion

LOD and LOQ values were as low as 10 ng/mL for all analytes. Calibration curves from 10–1,000 ng/mL showed linearity (R2 ≥ 0.990 for 22 compounds). Precision and accuracy at 80, 400, and 800 ng/mL met ANSI/ASB 036 criteria (<20% bias and precision), with minor exceptions for naproxen, heroin, and acetylsalicylic acid. No carryover or matrix interferences were observed. Seven postmortem samples analyzed in triplicate yielded %RSD <20% and <20% deviation from target concentrations. Library-based confirmation using similarity scores ≥80 ensured reliable compound identification without false positives or negatives.

Benefits and Practical Applications of the Method

• High throughput: sample prep reduced from >2 hours manually to ~5 minutes automatically.
• Enhanced reproducibility and safety by eliminating manual handling steps.
• Robust quantitative performance across blood, serum, and urine matrices.
• Streamlined method development leveraging a forensic toxicology database.

Future Trends and Potential Applications

Advancements may include expansion of the target list, integration with high-resolution mass spectrometry for untargeted screening, and adoption in clinical toxicology and anti-doping laboratories. Machine learning–driven library matching could further streamline data analysis and improve identification confidence.

Conclusion

The automated CLAM-2030/LCMS-8060 platform provides an efficient, reliable solution for multi-target drug screening and quantitation in biological samples. Its high throughput, minimal hands-on time, and strong analytical performance make it well suited for forensic and clinical applications.

Reference

1. Analysis of Postmortem Samples Using Automated Sample Preparation Coupled Directly to LC-MS/MS, Shimadzu Scientific Instruments, 2019.