Sample Cleanup: Method Development for Solid Phase Extraction and Beyond

Significance of the Topic

The selection and optimization of sample cleanup methods is critical in analytical chemistry to ensure accurate, sensitive, and reproducible measurements. Solid phase extraction (SPE) and related technologies remove matrix interferences such as lipids, proteins, pigments, and salts, enhancing instrument performance and extending column and ion source lifetime. Effective cleanup is especially important in trace analysis of environmental contaminants, pharmaceuticals, PFAS, mycotoxins, and food safety applications.

Objectives and Study Overview

This work aims to guide method development and troubleshooting for SPE and complementary cleanup strategies. Key goals include:

• Reviewing SPE fundamentals, sorbent chemistries, formats, and processing equipment.
• Providing a systematic approach to selecting sorbents based on analyte properties and sample matrix.
• Detailing nonpolar SPE method optimization and common troubleshooting scenarios.
• Highlighting new Agilent cleanup products for lipids, PFAS, mycotoxins, and pigments.

Methodology and Instrumentation

The general SPE workflow consists of:

• Sample pretreatment (pH adjustment, dilution, protein precipitation).
• Conditioning and equilibration of the sorbent (wetting with organic solvent and water/buffer).
• Sample loading under controlled pressure or vacuum.
• Washing to remove weakly bound interferences.
• Elution of target analytes with appropriate solvents.

Sorbent selection factors include analyte polarity (hydrophobic, polar, ionic), molecular size, logP/pKa, and matrix interferences. Common sorbents are C18, C8, polymeric HLB, ion exchangers, mixed-mode phases, and specialty materials like phenylboronic acid or carbon graphitized carbon. SPE cartridges, 96-well plates, pipette-tip formats, and bulk sorbent are used with vacuum or positive-pressure manifolds.

Instrumentation and consumables mentioned:

• Bond Elut SPE cartridges and OMIX pipette-tip sorbents.
• VacElut and PPM positive-pressure manifolds for high-throughput workflows.
• Captiva EMR filters and QuEChERS kits for targeted filtration and dispersive cleanup.

Main Results and Discussion

Nonpolar SPE optimization involves balancing retention and elution to maximize analyte recovery and minimize coextracted matrix. Troubleshooting covers:

• Low retention due to inadequate pH or flow rate—adjust sample pH and slow loading speed.
• Analyte loss during wash—reduce wash solvent strength or split wash steps.
• Incomplete elution—use stronger or multiple elution solvents, aliquot elution volumes, or change sorbent hydrophobicity.
• Dirty extracts—add or modify wash steps, switch to more selective or mixed-mode sorbents.
• Variable recovery—ensure consistent conditioning, adequate sorbent capacity, and sample neutralization.

Example methods:

• Bond Elut PPL for polar organic contaminants in drinking water: 500 mg cartridge, dual-step elution with acidified methanol/acetonitrile.
• Bond Elut Plexa for PAHs, chloropesticides, and triazines in water: single 200 mg method with sequential ethyl acetate and dichloromethane fractions.

New SPE and cleanup products include:

• EMR-Lipid sorbent for lipid removal in small-molecule analysis.
• PFAS WAX cartridges optimized for EPA and ISO methods with minimal background contamination.
• Bond Elut HLB polymeric phases for broad polarity ranges, high capacity, and wide pH stability.
• Carbon S for pigment and planar pesticide removal in food/environmental matrices.
• Captiva EMR PFAS Food I and II formats for mixed-mode cleanup of PFAS in diverse food samples.
• Captiva EMR Mycotoxins cartridges tailored for multiclass mycotoxin analysis.

Benefits and Practical Applications

The described strategies deliver:

• Enhanced analyte recovery and reproducibility across complex matrices.
• Improved instrument uptime through effective matrix removal.
• Streamlined workflows with fewer solvent consumption and reduced sample volumes.
• Compatibility with regulatory methods (EPA 533, 1633; ISO 21675) and high-throughput automation.
• Versatility for environmental monitoring, food safety, pharmaceutical analysis, and emerging contaminant screening.

Future Trends and Applications

Emerging directions include:

• Advanced mixed-mode sorbents combining size exclusion, hydrophobicity, and ion exchange for targeted cleanup.
• Integration of SPE with on-line LC-MS workflows and automation for real-time sample processing.
• Miniaturized formats for microsampling and single-cell analysis.
• Development of sorbents for ultra-trace detection of PFAS, microplastics, and next-generation contaminants.
• Expansion of EMR chemistries for lipidomics, proteomics, and mycotoxin multiplex assays.

Conclusion

Methodical selection and optimization of cleanup techniques, informed by analyte and matrix characteristics, underpin robust analytical protocols. Combining classic SPE fundamentals with novel sorbents and formats enhances cleanup specificity and throughput. Troubleshooting guidelines and example workflows facilitate rapid method development. The latest Agilent technologies address evolving challenges in environmental, food, and clinical analysis, paving the way for higher sensitivity and broader applicability.

References

• Javadi G. Sample Cleanup: Method Development for Solid Phase Extraction and Beyond. Agilent Technologies Application Note DE-001315; September 25, 2024.