HPLC vs Mass Spectrometry: How Peptide Purity Is Verified
By the Prime Peptide Labs lab team · 17 September 2026 · 7 min read
Two techniques answer two different questions: HPLC tells you how pure a sample is; mass spectrometry tells you whether it is the right molecule. You need both.
Reverse-phase HPLC: measuring purity
The sample passes through a C18 column under a water/acetonitrile gradient. Components separate by hydrophobicity and are detected by UV absorbance (usually 214–220 nm, where peptide bonds absorb).
Purity = main-peak area ÷ total peak area. A clean chromatogram shows one dominant sharp peak with small, well-resolved minor peaks.
Mass spectrometry: confirming identity
ESI-MS or MALDI-TOF measures the mass-to-charge ratio of ionised molecules. The observed molecular weight should match the theoretical value of the sequence within a small tolerance. A 99% pure sample of the wrong peptide is still useless — MS rules that out.
Reading a Certificate of Analysis
Check: compound name and sequence, batch/lot number, theoretical vs observed molecular weight, HPLC purity %, appearance (white lyophilised powder), test date and lab signature. Match the batch number to your vial.
Why ≥98% matters
At 95% purity, one in twenty molecules is something else — often deletion sequences that may be biologically active. For reproducible receptor or cell-based work, ≥98% is the accepted floor.
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