A purity percentage tells you how much of a sample is the target molecule — but not whether that target molecule is really in the vial. Mass spectrometry answers that question. This guide explains how it works and why the two measurements need each other.
Mass spectrometry weighs molecules: it measures the mass-to-charge ratio of ionised particles, and the molecular weight follows. If that matches the sequence, identity is confirmed. It does not produce a purity percentage — that remains HPLC's job.
What is mass spectrometry?
Mass spectrometry (MS) is a technique that ionises molecules and then sorts them by their mass-to-charge ratio (m/z). The result is a spectrum: a series of signals along the m/z axis, where the position of a signal reveals the mass of the corresponding ion.
For peptides a soft ionisation technique is typically used, usually electrospray ionisation (ESI). Soft means the molecule enters the gas phase and picks up a charge without falling apart. Exactly what you want when the molecular weight of the intact peptide is the question.
How does a molecular weight follow from that?
The molecular weight is calculated back from the measured m/z values and the number of charges, then compared with the weight implied by the amino acid sequence. Those two have to agree within a small margin. Deviate further and something other than what the label says is in the vial — or not only that.
Under ESI peptides often pick up more than one charge, so the same molecule appears at several positions in the spectrum: at its mass divided by one charge, by two, by three. That is not contamination but a property of the technique, and it is precisely that series which makes the calculation reliable.
The molecular weight in the specifications on a product page is the theoretical value from the sequence. The measured value for a specific batch is on the certificate.
HPLC and MS: what measures what?
The two techniques answer different questions, and a batch is only characterised once both have been done.
Which is why a serious certificate of analysis contains both. More on the separation side is in what is HPLC; our thresholds are on quality standards.
What mass spectrometry does not tell you
A mass spectrum cannot distinguish substances that share a molecular weight. Two molecules with the same formula but a different structure — including a swap in the amino acid order — weigh exactly the same. Seeing through that requires fragmentation, where the molecule is deliberately broken apart and the fragments reveal the sequence.
And just like a purity measurement, a spectrum says nothing about sterility, endotoxins or suitability for any application. Why we have those measurements done by a third party is on Janoshik Analytical; check a specific batch via verify a lot number.
This information is general and educational in nature. Peptide Lab supplies research peptides for laboratory and research purposes only. Not for human or animal consumption, not a medicine and not medical advice.
Frequently asked questions about mass spectrometry
What does mass spectrometry do for a peptide?
Mass spectrometry measures the mass-to-charge ratio of ionised molecules, and the molecular weight follows from that. If the measured weight matches the weight implied by the amino acid sequence, that is the confirmation that the intended molecule is in the vial.
What is the difference between HPLC and mass spectrometry?
HPLC separates the components of a sample and determines what percentage is the target peptide — how much. Mass spectrometry confirms molecular identity and molecular weight — what. They measure different things and do not replace each other.
Why are there several peaks in the mass spectrum of a single peptide?
Peptides often pick up multiple charges during ionisation. One molecule then appears at several positions in the spectrum, each time at its mass divided by the number of charges. A single molecular weight is calculated back from that series.
Does mass spectrometry prove a peptide is pure?
No. A spectrum confirms identity, not a purity percentage. Two substances with the same molecular weight cannot be told apart by an ordinary measurement. HPLC is what produces the purity figure.
