Sample Prep for Chromatography

Importance of the Topic

Sample preparation is a critical step in chromatographic analysis, directly impacting sensitivity, specificity and laboratory throughput. Effective cleanup and concentration of analytes reduce matrix effects, extend column lifetime, and support reliable quantification in fields such as bioanalysis, environmental monitoring, food safety and pharmaceutical research.

Objectives and Overview of the Study

This presentation from the 2010 Innovation Seminar Series at Supelco (Sigma-Aldrich) aims to review advanced sorbents, devices and techniques for improving chromatographic sample preparation. Key goals include comparing general SPE theory, exploring three main cleanup strategies (bind-elute, interference removal, fractionation), and highlighting three device classes—hybrid SPE particles, molecularly imprinted polymers, and solid-phase microextraction fibers—with practical examples.

Methodology and Instrumentation

The methods described combine traditional SPE cartridges and plates, 96-well formats, and microextraction fibers. Main instrumentation includes:

• HPLC and UHPLC systems equipped with C18 or RP-Amide columns
• GC/MS with split/splitless inlets and quadrupole detectors
• LC-MS/MS with electrospray ionization (ESI) sources
• 96-well vacuum manifolds and robotic samplers for high-throughput formats

Method steps typically involve sample loading, washing, elution or desorption, solvent evaporation (if required), and direct analysis by LC/GC or MS.

Key Results and Discussion

Bind-Elute SPE enables concentration of analytes by retaining targets on sorbent, washing off matrix, and eluting under controlled conditions.
Interference Removal SPE, exemplified by HybridSPE™, selectively binds phospholipids via zirconia-coated silica to eliminate ion-suppression in LC-MS bioanalysis. HybridSPE-PPT achieved consistent backpressure, extended column lifetime and eliminated gradient cleaning, increasing injections per day from ~70 to ~700.
Molecularly imprinted polymers (SupelMIPs) offer highly selective binding sites tailored to target molecules. In honey analysis of chloramphenicol, MIPs reduced matrix interference versus liquid–liquid extraction, enabling ppb–ppt detection.
Solid-phase microextraction (SPME) fibers achieve solvent-free enrichment via equilibrium partitioning into a polymer coating. DVB/PDMS and Carboxen/PDMS fibers demonstrated ppt-level detection of odorous compounds in water and headspace profiling of volatile flavor components in food.

Benefits and Practical Applications

• HybridSPE-PPT combines protein precipitation simplicity with SPE selectivity, improving LC-MS sensitivity in bioanalysis and reducing maintenance.
• SupelMIP cartridges deliver rapid, high-specificity cleanup for antibiotics, PAHs, NSAIDs and other analytes in complex matrices, without extensive method development.
• SPME fibers support field-deployable, solvent-minimized sampling for environmental monitoring, forensic screening, flavor analysis and in vivo pharmacokinetics, lowering solvent use and animal costs.

Future Trends and Applications

Emerging directions include biocompatible SPME probes for serial in vivo sampling, integration of automated high-throughput SPE platforms, development of new MIP chemistries for broader analyte classes, and hybrid devices combining molecular recognition with microextraction. Advances in low-dead-volume connectors and on-line coupling to HRMS will further streamline workflows.

Conclusion

Innovations in sample preparation—hybrid SPE, molecularly imprinted sorbents and microextraction fibers—address key challenges of matrix effects, analyte concentration and throughput. These techniques enhance chromatographic performance, support trace-level analysis across diverse fields and pave the way for fully integrated, automated workflows in analytical laboratories.

Instrumentation Used

• HybridSPE™-PPT 96-well plates and cartridges
• SupelMIP™ SPE plates and cartridges
• SPME fiber assemblies and autosamplers
• HPLC/UHPLC with C18, RP-Amide columns
• GC/MS with splitless injection and quadrupole detectors
• LC-MS/MS with ESI interfaces