← Research Library · Published August 30, 2026
Almost every meaningful peptide Certificate of Analysis rests on two measurements: a purity estimate from high-performance liquid chromatography (HPLC), and an identity confirmation from mass spectrometry (MS). They answer different questions, and a COA that reports only one is only half an analysis.
Reversed-phase HPLC — the purity question
In reversed-phase HPLC, the dissolved sample is pushed through a column packed with a hydrophobic stationary phase while the solvent composition changes over time. Components elute at different points depending on how strongly they interact with the column. A detector records each as a peak. The target peptide is the main peak; synthesis by-products, truncated sequences, and other impurities show up as separate, usually smaller, peaks.
Purity is calculated as the area of the main peak as a fraction of the total peak area, expressed as a percentage. Because it is an area measurement of what the detector sees, the method parameters matter: the column, the gradient, the detection wavelength, and the run time all affect what is resolved and what is reported. A credible COA lists them.
Mass spectrometry — the identity question
A mass spectrometer ionises molecules and sorts the ions by mass-to-charge ratio. For peptides, electrospray ionisation (ESI) and matrix-assisted laser desorption/ionisation (MALDI) are the common front ends. The instrument reports the measured mass, which is then compared to the theoretical mass calculated from the intended amino-acid sequence. When the observed value falls within a small tolerance of the expected value, identity is confirmed.
Mass spec does not tell you how pure the sample is — a small amount of the right molecule and a large amount of it produce the same mass reading. That is why it is paired with HPLC.
Why the two are complementary
- HPLC without MS: you know most of the sample is one compound, but not that it is the right one.
- MS without HPLC: you know the target molecule is present, but not whether it is 60% or 99% of the material.
- Both together: a defensible statement that the sample is predominantly the intended peptide.
What neither method covers
HPLC and MS characterise the peptide. They are not designed to quantify bacterial endotoxin, residual solvents, or microbial contamination — those require their own assays. For most in-vitro laboratory work the identity-and-purity pair is the baseline; whether additional testing is needed depends on the study.
Applying this when you buy
When a COA is available for a Summit Labs product it is linked on the product page and, where applicable, reports the methods used. Use the COA reading guide to work through a document field by field, and the supplier vetting checklist to decide how much weight to give it.
FAQ
Is in-house testing as good as third-party testing?
Independent third-party testing removes the conflict of interest in a supplier grading its own material. When evaluating a vendor, ask which laboratory ran the analysis and whether the method and raw data are available on request.
Do HPLC and mass spec detect every possible contaminant?
No. They characterise purity and identity of the peptide itself. Separate assays — for example endotoxin or sterility testing — address contaminants those two methods are not designed to catch.
For Research Use Only. All compounds sold by Summit Labs are intended solely for in vitro research and laboratory use. Not for human or veterinary consumption. Must be handled by qualified researchers in compliance with all applicable regulations.