In liquid chromatography, the
instrument isn’t the only factor that determines separation quality — the
mobile phase solution plays a crucial role too.
Whether you’re working with HPLC,
UPLC, or LC–MS, the choice of solvents, additives, and gradients can make or
break your analysis.
HPLC (High-Performance Liquid Chromatography)
Typically uses water mixed with
organic solvents such as acetonitrile or methanol.
Buffers like phosphate or acetate are common for pH control.
The focus is on robust,
reproducible separations — ideal for routine assays and quality control.
UPLC (Ultra-Performance Liquid Chromatography)
Uses the same types of solutions
but often requires finer filtration and higher purity solvents due to sub-2 µm
columns and higher system pressures.
Gradient precision and solvent
compatibility are critical to maintain column life and reproducibility.
Slight changes in solvent composition can lead to significant differences in
peak shape and resolution.
LC–MS (Liquid Chromatography–Mass Spectrometry)
Mobile phases must be volatile to
be MS-compatible.
Non-volatile buffers (like
phosphate) are avoided; instead, we use formic acid, ammonium acetate, or
ammonium formate.
The goal is to maintain separation
while ensuring clean ionization and minimal source contamination.
Key Takeaway:
Choosing the right solution depends
on both your separation goal and your detection system.
HPLC: buffered aqueous-organic systems
for robust separations.
UPLC: highly pure, well-filtered
solvents for precision and efficiency.
LC–MS: volatile, MS-friendly mobile
phases for sensitive detection.
Each system has its own “chemistry”
— and understanding your mobile phase is just as important as knowing your
instrument.