Developing reliable LC‑MS methods for polar analytes often presents challenges—insufficient retention, poor peak shape, or unstable ionization. When using a Cogent TYPE‑C™ column, such as the Cogent Diamond Hydride™, Aqueous Normal Phase (ANP) provides a highly effective mode for separating these compounds, particularly in positive‑ion electrospray workflows.

The following enhanced guidance expands on the original MICROSOLV technical note to help analysts establish robust, reproducible ANP gradients.

1. Prepare Solvents for Positive‑Ion LC‑MS

Proper mobile‑phase composition is essential for achieving retention and optimal ionization:

• Solvent A: 50% methanol / 50% DI water + 0.1% formic acid
• Solvent B: 90% acetonitrile / 10% DI water + 0.1% formic acid

These compositions promote strong initial retention of polar analytes while supporting stable protonation efficiency in positive‑ion MS detection.

2. ANP Gradient Setup for Polar Compound Retention

To retain highly polar species, ANP gradients must start with a very high organic content:

• Begin at 100–90% Solvent B This high‑acetonitrile environment enhances surface interactions on TYPE‑C™ silica, enabling polar compounds to retain effectively.
• Gradually reduce the percentage of Solvent B, down to 20% Solvent B This transition is appropriate when Solvent A is 50/50 methanol–water and allows controlled elution of polar compounds without early breakthrough. 

This behavior is unique to ANP and differs significantly from reversed‑phase (RP) or HILIC, making TYPE‑C™ columns particularly powerful for analytes that struggle to retain under traditional methods.

3. Rapid Column Re‑Equilibration—A Major Advantage

One of the operational strengths of Cogent TYPE‑C™ columns is their fast equilibration time:

• Column re‑equilibration: ~5 minutes
• Comparable HILIC columns: 20–30 minutes or more

This substantial time savings improves throughput, reduces idle instrument time, and leads to more consistent retention, especially in high‑volume LC‑MS laboratories. 

4. WHY TYPE‑C™ Columns Excel in ANP for LC‑MS

The silicon‑hydride surface of TYPE‑C™ columns offers several benefits:

• Strong retention of polar molecules in high‑organic environments
• Rapid switching and equilibration between gradient runs
• Cleaner baselines and improved ionization efficiency compared to traditional silica surfaces

This unique behavior helps analysts create robust, reproducible methods with minimal trial‑and‑error.