How to read a certificate of analysis (COA): HPLC, mass spectrometry and purity

A certificate of analysis, or COA, is the laboratory's report on one specific batch of a compound. For research peptides it answers two separate questions: is this the right molecule, and how much of the vial is that molecule rather than something else. The two questions need two different methods, and a useful certificate shows both.

Purity: the HPLC chromatogram

High-performance liquid chromatography (HPLC) pushes a dissolved sample through a column that holds back different molecules for different lengths of time. A detector at the end records a signal as each one leaves, and the result is a chromatogram: time along the bottom, signal height up the side. The target peptide should appear as one tall, sharp peak. Impurities — shorter chains left over from synthesis, oxidised or truncated forms — appear as smaller peaks either side of it.

Purity is the area of the main peak as a share of the area of all peaks. A result of ≥99% means that, of everything the detector saw, at least 99% was the main peak. It is a measure of how clean the batch is, not of how much peptide is in the vial — that is a separate figure (net peptide content) that some certificates also report.

Identity: the mass spectrum

HPLC alone cannot prove which molecule the main peak is; a wrong compound can also produce one clean peak. Mass spectrometry (MS) settles identity by measuring the mass of the molecule. The certificate should state the theoretical molecular weight and the measured one, and they should agree closely. For larger peptides the spectrum often shows several peaks for the same molecule carrying different charges; the software combines them into one measured mass.

What a credible certificate includes

• The name of the testing laboratory, and ideally that it is independent of the seller.
• A batch or lot number that matches the batch being sold, and a test date.
• The chromatogram itself, not just a percentage, with the main peak and any impurities visible.
• The mass result: expected against measured molecular weight.
• The method conditions (column, wavelength, gradient) — a sign the test was actually run as described.

Warning signs

• A purity figure with no chromatogram behind it.
• One certificate reused for every size or every batch.
• No laboratory named, or a laboratory that is the seller itself.
• A perfectly flat baseline with a single peak and nothing else — real chromatograms have some noise and small peaks.

BioPeptide publishes the full certificates from an independent laboratory, with both pages, on its product pages and in the Lab Results section, so the figures quoted on the site can be checked against the documents behind them. Certificates describe research material; nothing here is guidance for use in humans or animals.

1. 1.ICH Q2(R2): Validation of Analytical Procedures. International Council for Harmonisation, 2023.
2. 2.United States Pharmacopeia, General Chapter <621> Chromatography.