Mass spectrometry gives information about the molecular weights of individual molecules or their ions. Other techniques merely separate compounds. It can be employed for peptide identity validation, modification detection and impurity investigation.
How does a mass spectrometer work?
The first step in the process is that the mass spectrometer turns the sample into electrically charged molecules. Next, the ions are sorted by their mass-to-charge ratio and detected by the instrument.
The output is a mass spectrum. Usually this is a graph with m/z on the abscissa and signal intensity on the ordinate. All of the large peaks corresponded to detected ions of a particular mass-to-charge ratio.
For peptides, techniques such as electrospray ionisation (ESI) and matrix-assisted laser desorption/ionisation (MALDI) are often used. These techniques can analyse fairly large molecules without fragmenting them too much.
MS tells you about a peptide.
Verification of the molecular mass is one of the most important applications of MS in peptide research. Using the amino acid sequence and any chemical modifications, the mass of a peptide can be predicted. This value can then be compared to the measured mass.
Mass does not always prove the sequence in toto, but a close match can support the identity of the peptide. There are some cases that two different compounds have the same or very similar molecular masses.
Mass spectrometry can also be used for identifying chemical modifications. Modifications such as acetylation, amidation or oxidation can alter the mass that is being measured, enabling investigators to see whether a modification is present.
MS versus MS/MS
Mass spectrometry (MS) is a measurement of the mass-to-charge ratio of ions. Tandem mass spectrometry (MS/MS) takes it a step further by selecting an ion and breaking it into smaller fragments.
The resulting fragment pattern can give information about the order of amino acids in a peptide. This is why MS/MS is often performed in conjunction with the original MS measurement when researchers need more information about peptide identity or sequence.
MS and peptide purity
Mass spectrometry and HPLC are often used in conjunction, but they provide different types of information.
HPLC can separate the components in the sample. It can be used to estimate purity. Mass spectrometry detects the mass of the ions and helps identify the compounds present. Thus a peptide can be of high HPLC purity, but MS is still necessary to confirm that the measured mass is that of the expected peptide.
MS results may be reported with HPLC purity, sequence information and other analytical data when reviewing a peptide Certificate of Analysis (CoA).
Related terms: Mass-to-Charge Ratio · m/z · Mass Spectrum · Molecular Weight · MS/MS · Electrospray Ionisation · MALDI · HPLC · Purity · Sequence · Certificate of Analysis
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Research Use Only. This glossary explains terminology used in published research. It is not medical advice and does not describe any product for human or animal use.