How to read a Certificate of Analysis

Start with the batch identifiers

Every legitimate COA is tied to a single production lot, not a product line. The document should list a batch or lot number, a test date, and the name of the testing laboratory — ideally an independent third party rather than the manufacturer's own instrument room. Cross-check that the batch number printed on the COA matches the batch number printed on the vial label or shipping insert in front of you.

If a supplier publishes one generic COA for an entire catalog line rather than per-batch documents, that is a structural gap: manufacturing runs vary lot to lot, and a single archived PDF cannot speak for material produced months apart.

• Batch/lot number — must match the physical vial
• Test date — should be reasonably close to the ship date
• Testing laboratory name — look for an independent, named lab rather than 'internal QC'

Purity: HPLC percentage and how it is derived

The headline purity figure on most peptide COAs comes from high-performance liquid chromatography (HPLC), usually reverse-phase. The lab injects a dissolved sample, separates it by hydrophobicity as it passes through a column, and records a chromatogram — a plot of detector absorbance (commonly at 210–220 nm, where the peptide bond absorbs UV light) against retention time.

The purity percentage is the area of the main peak divided by the total area of all peaks in the trace, expressed as a percentage. A single sharp, dominant peak with minimal shoulder peaks or baseline noise indicates a chromatographically clean sample. Multiple smaller peaks represent truncated sequences, deletion products, or degradation byproducts from synthesis.

A COA that reports only a number with no attached chromatogram image gives you far less to verify than one that includes the trace itself — the shape of the peak, baseline stability, and peak symmetry all carry information a bare percentage does not.

Identity confirmation: mass spectrometry

Purity alone does not confirm what the substance is — it only confirms that whatever is in the vial is chromatographically homogeneous. Identity is established separately, usually by mass spectrometry (MS), most commonly electrospray ionization (ESI-MS) or matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF).

The lab compares the observed molecular mass (or mass-to-charge ratio) against the theoretical monoisotopic or average mass calculated from the peptide's amino acid sequence. A match within instrument tolerance (typically a fraction of a percent) supports that the correct sequence is present. A mismatch, or a mass corresponding to a related-but-different sequence, is a material discrepancy worth flagging with the supplier before use.

Secondary fields: TFA content, water content, counter-ion

Solid-phase peptide synthesis typically uses trifluoroacetic acid (TFA) during cleavage and purification, and residual TFA can remain bound to the peptide as a counter-ion (or as free residual solvent) unless it is specifically removed by ion exchange. Some COAs report a distinct TFA content field, usually by ion chromatography or a related analytical method, expressed as a percentage of total mass.

Water content (by Karl Fischer titration) and net peptide content (accounting for water, counter-ion, and other non-peptide mass) are additional fields that better-documented COAs include. Net peptide content is a more complete accounting of what fraction of the vial's total mass is actually the peptide chain itself, separate from bound moisture and salts.

Putting it together

A complete COA lets you verify four separate facts: which batch this document describes, what the chromatographic purity of that batch is, whether the molecular identity matches the labeled sequence, and what non-peptide mass (water, TFA, other counter-ions) is present. Treat a document missing more than one of these fields as incomplete documentation, and always confirm the batch number on the certificate against the vial in hand before relying on it.