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Purity is not potency
HPLC, mass spectrometry and Karl Fischer each answer a different question. Three suppliers quoting "99%" may be quoting three different things.
Three numbers get used interchangeably in this market and they are not interchangeable. Knowing which one a supplier means is most of the work of comparing two of them.
| Assay | Question it answers | What it cannot tell you |
|---|---|---|
| RP-HPLC, 214 nm | What fraction of the peptide present is the one you wanted | How much peptide is present at all |
| Mass spectrometry | Is this the molecule you ordered | How much of it there is |
| Karl Fischer | How much water is in the solid | Anything about the peptide |
| Net peptide content | What fraction of the solid is peptide | Whether it is the right peptide |
None of the four is a sterility or endotoxin test; those are separate assays.
The gap that matters
Purity is a ratio between peptides. Net content is a ratio between peptide and everything else in the vial. A sample can be 99 per cent pure and 74 per cent peptide simultaneously, and a supplier quoting the first number without the second is not lying — they are answering a question you did not ask.
Reading a chromatogram
At 214 nm the detector is reading the peptide bond, so it sees peptides and is close to blind to salt and water. One dominant peak with a flat baseline is what you want. Shoulders on the main peak usually mean deletion sequences that co-elute; a cluster of small peaks late in the run often means oxidation.
The quality section of the home page shows a trace, and the purity figure on each certificate in our testing library is the peak area purity for that lot rather than a house average.
Common questions
What is the difference between peptide purity and net peptide content?
Purity is a ratio between peptides, and net peptide content is a ratio between peptide and everything else in the vial. A sample can be 99 per cent pure and 74 per cent peptide at the same time, so a supplier quoting the first number without the second has answered a different question.
What does mass spectrometry tell you about a peptide?
It tells you whether this is the molecule you ordered. It cannot tell you how much of it there is. That makes it a separate assay from RP-HPLC, which reports what fraction of the peptide present is the intended one, and from Karl Fischer, which reports how much water is in the solid.
Do HPLC purity and mass spectrometry cover sterility or endotoxin?
No. None of the four measurements compared in this article, RP-HPLC purity, mass spectrometry, Karl Fischer water and net peptide content, is a sterility or endotoxin test. Those are separate assays that answer their own questions.
Why does an HPLC detector at 214 nm not see salt and water?
It is because at 214 nm the detector is reading the peptide bond. It sees peptides and is close to blind to salt and water, which is why a purity figure describes the peptides present and cannot say how much peptide is present at all.
For laboratory research use only. Not a drug, not a supplement, and nothing here is a claim about what any of this material does in a person or an animal.
