How to Read a Certificate of Analysis

A certificate of analysis is the only document that connects a number on a label to a measurement someone actually made. Most of the trust in a peptide supplier rests on it — and most researchers read only the top line. Here is what each section is telling you.

The header: identity and traceability

Before any figure, check three fields: the product name, the lot number, and the date of analysis. The lot number on the certificate must match the lot number printed on the vial in your hand. If it does not, the document describes a different batch of material and tells you nothing about yours. A supplier that publishes one “representative” certificate for all batches of a compound is publishing marketing, not analytics.

One vial, one lot, one report. Anything else is a category error.

HPLC purity: what the percentage means

Reverse-phase high-performance liquid chromatography separates the sample into its components by how strongly each binds a hydrophobic stationary phase. A detector at a fixed wavelength — usually 214 nm for peptide bonds — measures what comes off the column over time. The result is a chromatogram: a baseline with peaks.

“99.4% purity” means the area under the main peak is 99.4% of the total detected peak area, under those specific conditions. That last clause matters. It is a relative measurement, not an absolute one, and it says nothing about material the detector could not see. Read the wavelength, the gradient and the column stated on the certificate — a purity figure without its method is a number without a unit.

Mass spectrometry: identity, not purity

HPLC tells you how much of one thing is present relative to everything else. It does not tell you what that thing is. Mass spectrometry answers the second question by measuring the mass-to-charge ratio of the ionised molecule; if the observed molecular ion matches the theoretical mass of the declared sequence, identity is confirmed.

A lot can be 99.8% pure and still be the wrong compound. Both tests are required, and a certificate carrying only one of them is incomplete.

Water content

Karl Fischer titration measures residual moisture in the lyophilised cake. This is the figure that quietly governs shelf life: water drives hydrolysis and, in cysteine-containing sequences, oxidation. It also matters for accuracy — if a vial labelled 10 mg contains 8% water, the peptide mass is meaningfully lower than the label states. Researchers weighing material for concentration-sensitive work should correct for it.

Residual solvents and heavy metals

Solid-phase synthesis and preparative purification use organic solvents; trace amounts survive into the final product. Headspace gas chromatography quantifies them against ICH Q3C limits. ICP-MS screens for metal carryover from reagents and hardware. Both are routine, both are boring, and both are absent from a surprising number of certificates.

Endotoxin

Bacterial endotoxin — lipopolysaccharide from Gram-negative cell walls — survives synthesis and sterilisation, and it is biologically active at extremely low concentrations. For any cell-culture work, endotoxin load is often the single most consequential number on the certificate, because contamination produces effects that are easily mistaken for the effect being studied. Look for an LAL result expressed in EU/mg.

What a certificate is not

A certificate of analysis states what was measured, by which method, under which conditions. It is not a statement of fitness for any purpose, it is not a safety assessment, and it is emphatically not a claim about what the material does. Those are separate questions, and a laboratory answers them itself.