HPLC and mass spectrometry.

Two analyses appear on almost every peptide report. They measure different things and complement each other.

HPLC measures how pure

High performance liquid chromatography pushes the dissolved compound under high pressure through a column with a packing that slows components down differently. Small or less bound molecules come out earlier than large or more strongly bound molecules.

At the end a detector measures what passes. That produces a graph with peaks. The ratio between the area of the main peak and the total is the purity percentage.

The limitation: the detector only sees what it can see. Salts, solvent residues and moisture do not always show up and so do not count in the percentage.

Mass spectrometry measures what it is

Mass spectrometry ionises the molecules and measures their mass divided by their charge. The result is compared with the theoretical mass that follows from the amino acid sequence.

If the measured mass matches, the identity is confirmed. If it deviates, there is something else in the vial than what is on the label, however high the HPLC percentage.

Why you need both

A compound can be 99 percent pure and still be the wrong compound. HPLC alone does not rule that out. Conversely the identity can be right while the batch is full of related chains. Only together do the two measurements give a complete picture.

And heavy metals

That is a third, separate measurement, usually with ICP-MS. It looks for traces of metals that can come from equipment or raw materials. We have it done as standard.