Documentation
How to Read a Peptide Analysis Certificate: Identity, Purity and the Overlooked Line
The two things the document is there to prove, the one number that is routinely misunderstood, and a full walk-through of a realistic example.
A certificate of analysis for a peptide exists to answer two things. Identity: does the vial contain the molecule printed on the label? Purity: of the peptide present, what proportion is that molecule rather than related impurities? Mass spectrometry settles identity and HPLC settles purity. Everything else on the page — the lot number, appearance, the counter-ion, storage advice — is context around those two answers. Once you can read the document properly it stops being a PDF you file away and becomes the single most informative quality indicator a research buyer has. Every peptide in this catalogue comes with a certificate matched to its lot; below we work through such a document section by section.
Header: what product, which lot, when
Before any measurement there are three items to check. First the product name and catalogue identity, which must agree precisely with your vial, including salt form and any modification — an amide, an acetate, a fragment range such as 176-191. Second the lot number, which must be the one printed on the vial's own label; a certificate covering a different lot is advertising, not evidence. Third the date of analysis, plus any retest or expiry date, telling you how recent the data are. Cross-checking the lot in exactly this way is also the core of validating testing by an independent laboratory.
Proving identity: the MS section
The mass-spectrometry block shows two numbers: the mass calculated from the molecular formula, and the mass the instrument actually measured — usually by ESI-MS. Interpretation is a single comparison: the measured figure should sit within the method's tolerance of the calculated value, and for small peptides in research-scale QC that generally means comfortably under 1 Da apart. Consider BPC-157, formula C62H98N16O22, with a mass around 1419.5 g/mol: a measured 1419.6 confirms the identity, while a result 42 higher points to an acetyl group that was never cleaved, and one 18 lower points to dehydration. Seeing multiply charged ions such as [M+2H]²⁺ in an ESI spectrum is entirely normal; what you compare is the deconvoluted mass. Mass spectrometry and peptide identity covers the technical detail.
Purity: the HPLC section
An HPLC purity value is the area of the principal peak expressed as a fraction of the total peak area, generally measured by reversed-phase HPLC with UV detection somewhere around 214–220 nm, the region where the peptide bond absorbs. A result of ≥99% therefore means that all related peptide impurities together — deletion sequences, truncated chains, oxidation products — make up less than 1% of the chromatogram's area. Two good habits: note the conditions used (wavelength, column, gradient), because purity figures are only comparable within a single method, and examine the trace itself when it is included, since a single sharp peak on a flat baseline conveys much more than a percentage on its own. For what the number does and does not tell you, see the discussion set out in HPLC purity explained.
The most misread figure: purity is not net peptide content
An HPLC purity percentage refers to the peptide portion of the sample. The weighed powder also contains bound water, traces of solvent and the counter-ion left behind by purification, usually TFA or acetate. Net peptide content — typically 70 to 90% by weight, measured by amino-acid analysis or a nitrogen assay when it is reported at all — tells you what share of the weighed material is peptide. It is perfectly consistent for a vial to be ≥99% pure while roughly 80% of its milligrams are peptide. Whenever molarity matters, base the calculation on net content if the certificate provides it; molecular weight, moles and molarity works through the arithmetic.
Warning signs on a certificate
- No lot number at all, or a lot number that differs from the vial's — in that case the document tells you nothing about the material you hold.
- Identity asserted purely from an HPLC retention time, with no spectrum. Excluding a wrong or truncated sequence that happens to co-elute is precisely the job MS does.
- A purity figure with no accompanying method: no column, no gradient, no wavelength. An unverifiable number is decoration.
- Paperwork that looks too good — 100.00% purity, identical chromatogram images across different lots, or no analysis date anywhere.
- Relying only on the supplier's own document when the stakes are high. An in-house certificate is the minimum; external confirmation is the step up, which is why third-party verifiability sits near the top of the vendor checklist.
A worked read-through
Picture a certificate accompanying a 10 mg semaglutide vial (CAS 910463-68-2; C187H291N45O59; MW 4113.58). Header: the product and lot match the vial, analysis performed this year. Appearance: white lyophilised powder — as expected. MS: calculated 4113.6, measured 4113.5, so the identity checks out. HPLC: main peak 99.3% of total area, run at 220 nm on a specified C18 gradient, satisfying the ≥99% specification. Net peptide content: 84%. Putting that together: you have the correct molecule at high purity, and for any molar calculation you should treat the vial as holding 8.4 mg of peptide rather than 10 mg. Leaving out that last step is precisely the difference between a certificate that gets filed and one that gets used. The same logic applies across every format we sell, from freeze-dried vials to the bulk material behind sprays and oral capsules.
Questions
What belongs on a good peptide COA?
At minimum: the product name together with its sequence or molecular formula, a lot number that agrees with your vial, when the analysis was performed, a description of appearance, a mass-spectrometry section giving calculated versus measured mass, and an HPLC purity result with the method stated. Better documents also report net peptide content, TFA or counter-ion levels and water content. Two absences make a certificate worthless: no traceable lot, and no MS identity data.
If a COA says 99% purity, is the powder 99% peptide?
No. The HPLC figure applies only within the peptide portion of the sample: of the peptide molecules there, 99% are the intended sequence. The powder as weighed also contains residual solvent, water and counter-ion. The number that tells you how much of the labelled mass is genuinely peptide is net peptide content, which is often between 70 and 90% by weight. So it is entirely possible for one vial to be 99% pure and, at the same time, 80% peptide by mass.
Why is mass spectrometry needed when HPLC is already there?
Because HPLC tells you how clean the sample is, not what the sample is. A completely wrong or shortened peptide can still produce a single sharp peak somewhere on the chromatogram. Mass spectrometry provides independent proof that the molecular mass corresponds to the intended sequence, and typical offsets — −18, +16, +42 — pinpoint specific problems such as loss of water, oxidation, or an acetyl group left in place.
What is the counter-ion line telling me?
Peptides coming off a reversed-phase HPLC column are paired with a counter-ion — acetate or, more often, trifluoroacetate (TFA) — and it remains in the freeze-dried salt. That has two consequences: it contributes to the weight you measure, and leftover TFA can interfere with certain sensitive cell assays, which is why acetate salt is sometimes requested for cell-based work.
How do I establish that a COA is genuine?
Start by confirming the lot on the paper is the lot on the vial. Then look for checkable specifics: the analytical method named, results that look realistic rather than suspiciously tidy, a date of analysis, and chromatograms or spectra that differ between lots rather than being reused. The strongest proof is external: send material from your own lot to an independent laboratory and compare the results. The third-party testing guide lists who performs this work and what it costs.
Are COAs available for sprays and capsules as well?
Yes — the bulk peptide used in those formats undergoes the same testing, HPLC for purity and MS for identity, and the specification for the finished item states the peptide content per unit. Finished formats raise two extra questions, namely content uniformity and stability inside the formulation, which is why our format-specific pages list the purity of the raw material together with the amount per spray or per capsule.