A Comprehensive Comparison of Solid Phase Extraction (SPE) vs. Solid Liquid Extraction (SLE) vs. Liquid Liquid Extraction (LLE) Sample Prep Techniques in Bioanalysis and Forensic Toxicology Analyses

Importance of the Topic

Solid-phase extraction (SPE), supported liquid extraction (SLE), and liquid-liquid extraction (LLE) are cornerstone sample-preparation techniques in bioanalysis and forensic toxicology. Efficient cleanup of complex biological matrices such as plasma and urine is critical to achieve accurate quantification, minimize matrix effects, and streamline high-throughput workflows. A direct comparison of these approaches highlights their relative strengths, limitations, and suitability for diverse classes of analytes including pharmaceuticals, drugs of abuse, steroids, and synthetic cannabinoids.

Objectives and Overview of the Study

This study evaluated a streamlined three-step SPE protocol using Waters® Oasis PRiME HLB μElution plates against traditional SLE and LLE workflows. Two matrix types—plasma spiked with 22 analytes spanning acidic, basic, and neutral chemistries, and enzymatically hydrolyzed urine containing 23 drugs of abuse—were processed. Key performance metrics included extraction recovery, matrix effects, processing time, and method simplicity. The goal was to determine whether a single generic SPE workflow could deliver superior or equivalent performance compared to more laborious SLE and LLE methods.

Methodology and Instrumentation

All extracts were analyzed by UPLC-MS/MS under uniform chromatographic and detection conditions.

• UPLC System: ACQUITY UPLC I-Class with ACQUITY UPLC CORTECS C18 column (2.1 × 100 mm, 1.6 μm) at 40 °C.
• Mobile Phase: Gradient elution of 0.1% formic acid in water (A) and acetonitrile (B) at 0.6 mL/min, 6.5-min total run.
• MS Detection: Xevo TQ-S triple quadrupole, ESI+ mode, MRM transitions optimized per analyte.
• Sample Prep: Oasis PRiME HLB μElution plates for SPE; inert support SLE plates; MTBE-based LLE in centrifuge tubes. SPE used direct load-wash-elute without conditioning or evaporation.

Main Results and Discussion

In plasma, Oasis PRiME HLB SPE yielded average recoveries of 98 ± 8% across all 22 analytes and mean absolute matrix effects of 6%. SLE delivered 89 ± 7% recoveries but exhibited matrix effects up to 26%, with poor extraction (<30%) of acidic analytes. LLE recoveries averaged 70 ± 10% and showed variable matrix suppression. In urine, SPE achieved >75% recovery for 21/23 compounds (average 86 ± 6.6%) and mean matrix effects of 12%. SLE and LLE protocols required pH adjustments to recover polar amines, which compromised acidic metabolite recovery or increased ion suppression. Processing 96 plasma samples took 15 min by SPE versus 40 min (SLE) and 60 min (LLE).

Benefits and Practical Applications

The Oasis PRiME HLB SPE approach offers:

• High and consistent recoveries for broad analyte classes with minimal method development.
• Cleans extracts by removing proteins, salts, and phospholipids, reducing matrix effects.
• Elimination of conditioning, evaporation, and reconstitution steps in μElution format.
• Substantial time savings and simplified operation supporting high-throughput forensic and clinical workflows.

Future Trends and Applications

Emerging sample-prep innovations will focus on further automating SPE platforms, integrating on-line extraction with UPLC-MS/MS, and developing universal sorbents tailored for ultra-polar or labile compounds. Miniaturized formats, green solvents, and multimodal sorbents promise to enhance selectivity and throughput. Applications are likely to expand into metabolomics, therapeutic drug monitoring, and point-of-care testing, leveraging streamlined SPE protocols.

Conclusion

A direct comparison confirms that generic Oasis PRiME HLB SPE outperforms or matches SLE and LLE in recovery, cleanliness, and speed for both plasma and urine matrices. Its ability to extract diverse analytes with a single three-step protocol underscores its value in bioanalytical and forensic toxicology laboratories.

References

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