WATERS COLUMNS AND ANALYTICAL STANDARDS AND REAGENTS SELECTION GUIDE

Significance of the Topic

Modern liquid chromatography demands a versatile selection of stationary phases to address a wide range of analytes and application requirements. Waters’ comprehensive portfolio of UPLC, UHPLC, HPLC, and preparative LC columns combined with QC reference materials allows laboratories to achieve consistent, high-resolution separations under diverse conditions. This guide supports analytical chemists in selecting the optimal column chemistry to improve method robustness, throughput, and reproducibility.

Objectives and Overview of the Article

The primary goal is to present an organized classification of Waters’ column families and corresponding analytical standards and reagents. The article highlights key performance characteristics, operating pH and temperature ranges, available particle sizes and packing materials, and recommended QC reference materials for each column type.

Methodology and Instrumentation

Waters conducted systematic evaluation of column performance across multiple chemistries:

• Stationary Phases: charged-surface silica, fused-core (CORTECS), ethylene bridged hybrid (BEH), high-strength silica (HSS), and polymeric ion-exchange resins.
• Ligands: C18, C8, phenyl, fluoro-phenyl, T3, amide, HILIC, mixed-mode, SEC, protein- and oligonucleotide-specific phases.
• Instruments: UPLC (1.6–1.8 µm), UHPLC (2.5–2.7 µm), HPLC (2.7–10 µm), preparative chromatography setups.
• Quality Control: neutrals QC, reversed-phase QC, HILIC QC, peptide and protein standards, glycan test standards.

Main Results and Discussion

The survey identified distinct performance and selectivity profiles:

• C18 and Shield RP18 phases offer general-purpose reversed-phase separations with wide pH (1–12) and temperature (up to 80 °C) stability. Charged-surface hybrid technology improves peak shape for basic compounds.
• C8 and BEH Phenyl columns deliver shorter retention and enhanced π-π interactions beneficial for aromatic analytes. BEH phases maintain performance at extreme pH.
• Aqueous-phase compatible T3 and HSS C18 phases ensure balanced retention of polar and non-polar analytes under high-water-content mobile phases.
• HILIC and BEH Amide stationary phases excel in retaining highly polar and water-soluble compounds, carbohydrates, and glycans, with extended stability at elevated pH.
• Specialized phases for biomolecules: peptide BEH C18 columns demonstrate robust performance with tryptic digests; SEC columns cover proteins from small peptides to antibodies; IEX columns (SP, CM, DEAE) enable high-resolution separation of charged biomolecules; RP mAb Polyphenyl columns address mAb and ADC analysis.

Benefits and Practical Applications of the Method

• Scalability from analytical to preparative scale using the same phase chemistry.
• Reproducible method development supported by QC reference materials and performance standards.
• Enhanced peak shapes and loading capacities for challenging compounds (basic, polar, high-molecular-weight).
• Extended operational life of columns under aggressive pH and temperature conditions.
• Compatibility with multiple detectors (UV, MS, ELSD, fluorescence) for comprehensive analyte detection.

Future Trends and Opportunities for Application

• Development of hybrid and mixed-mode stationary phases to further expand selectivity options.
• Integration of AI-driven method scouting and gradient optimization with next-generation stationary phases.
• Advancements in micro- and nanoLC for single-cell and limited-sample analyses.
• Green chromatography initiatives focusing on reduced solvent consumption and sustainable materials.
• Enhanced column technologies for emerging biomolecule classes (glycoproteomics, oligonucleotides, biopharmaceuticals).

Conclusion

Waters’ extensive range of column chemistries and QC reference materials provides analytical laboratories with the tools needed for robust, high-resolution separations across small molecules, peptides, proteins, and glycans. By matching stationary phase characteristics to specific application requirements, chemists can achieve reliable performance, efficient method development, and scalable workflows from UPLC to preparative LC.

Reference

Waters Corporation. Columns and Analytical Standards and Reagents Selection Guide. February 2020.