SOLAμ for pre-analysis sample concentration

Significance of the Topic

Achieving low limits of detection in bioanalytical workflows often requires effective sample concentration and cleanup strategies. Conventional solid phase extraction (SPE) techniques demand time-consuming evaporation and reconstitution steps that can lead to analyte loss and extended turnaround times. The SOLAµ micro-elution SPE platform addresses these challenges by enabling high pre-analysis concentration in a simplified workflow, thereby enhancing sensitivity and preserving analyte integrity.

Objectives and Study Overview

The primary aim of this application note was to evaluate the performance of Thermo Scientific™ SOLAµ™ SPE for pre-analysis concentration of niflumic acid in human plasma. Specific goals included:

• Demonstrate up to 20-fold sample concentration without additional evaporation steps.
• Assess accuracy, precision, recovery, and matrix effects across a broad calibration range (40–40 000 pg/mL).
• Validate a rapid LC-MS/MS method using Accucore™ HPLC columns and TSQ Vantage™ mass spectrometry.

Methodology and Instrumentation

The workflow combined SOLAµ WAX 2 mg micro-elution SPE in a 96-well format with direct injection of a low-volume eluent:

• Sample pretreatment: 500 µL human plasma diluted 1:1 with 4% phosphoric acid.
• SPE protocol: Condition with methanol and phosphoric acid, load at 0.5 mL/min, wash with ammonium acetate and 70% methanol, elute in two 12.5 µL aliquots of 50/50 methanol/acetonitrile with 2% ammonia.
• Chromatography: Dionex™ UltiMate™ 3000 RSLC with Accucore™ RP-MS 50 × 2.1 mm, 2.6 µm column; 3-minute gradient (70:30 to 10:90 A/B) at 0.75 mL/min, 30 °C.
• Mass spectrometry: TSQ Vantage™ triple quadrupole with HESI in positive mode; optimized voltages and gas settings; MRM transitions for niflumic acid (m/z 283→265) and d5 internal standard (m/z 288.8→271.1).
• Data processing: LC QUAN™ v2.6 quantitative software.

Main Results and Discussion

By concentrating 500 µL of plasma into 25 µL eluent, a 20-fold enhancement was achieved without evaporation. Key findings included:

• Calibration linearity: 40–40 000 pg/mL with r2 = 0.998.
• Limit of quantitation: Clear chromatographic peaks at 40 pg/mL above acceptable signal-to-noise thresholds.
• Accuracy and precision: Calibration standards showed ≤7.4% deviation, QC samples at 400, 20 000, and 30 000 pg/mL exhibited accuracy within ±5% and RSD ≤1.31% (n=18).
• Recovery: >89.9% at low QC and >94.0% at high QC.
• Matrix effects: <9% at both QC levels, indicating effective sample cleanup.

Benefits and Practical Applications

The SOLAµ SPE approach offers several operational advantages:

• Workflow efficiency: Eliminates lengthy evaporation/reconstitution steps.
• Enhanced sensitivity: Facilitates up to 20-fold concentration for trace analyte quantitation.
• Robust analytical performance: High reproducibility at low elution volumes minimizes sample failures.
• Broad applicability: Suitable for volume-limited samples in clinical, pharmaceutical, and environmental analyses.

Future Trends and Potential Applications

Micro-elution SPE technologies like SOLAµ are expected to evolve alongside high-throughput and automation trends. Potential developments include:

• Integration with robotic liquid handlers for fully automated sample prep.
• Expansion to diverse SPE chemistries targeting polar metabolites, peptides, and intact biomolecules.
• Application in emerging fields such as metabolomics and exposomics requiring ultra-sensitive quantitation.
• Hybrid workflows coupling online SPE with UHPLC-MS/MS for seamless, real-time concentration and analysis.

Conclusion

This study confirms that SOLAµ micro-elution SPE reliably delivers up to 20-fold sample concentration while maintaining excellent accuracy, precision, recovery, and minimal matrix effects. The streamlined protocol significantly reduces processing time and preserves analyte integrity, offering a robust solution for high-sensitivity bioanalytical applications.

References

Thermo Fisher Scientific. Application Note 20941: SOLAµ for Pre-Analysis Sample Concentration. 2014.