A certificate of analysis (COA) is the single most useful document a peptide supplier can give you. It ties a specific production lot to measured data. Knowing how to read one lets you judge whether the material in your vial is what the label says, and how pure it is.
The header: product, lot and date
The top of a COA identifies the compound, its sequence, the theoretical molecular weight, the lot number and the analysis date. Check two things first: that the lot number matches your vial label, and that the test date is recent enough to reflect the material you received. A COA without a lot number, or one reused across many lots, documents nothing about your specific vial.
HPLC purity
Reverse-phase HPLC separates the target peptide from closely related impurities. A UV detector, typically at 214 or 220 nm where the peptide bond absorbs, records a chromatogram: a trace with peaks for each separated component.
Purity is reported as the area of the main peak divided by the total area of all peaks. A result of 99.2% means 99.2% of UV-absorbing material at that wavelength eluted as the target.
| What to look for | Why it matters |
|---|---|
| The chromatogram itself, not just a number | A single sharp main peak with small, well-separated minor peaks is what a clean product looks like. |
| Method details (column, gradient, wavelength) | Purity depends on method. A shallow gradient resolves impurities that a steep one can hide. |
| Integration of minor peaks | Every peak should be accounted for in the calculation. |
Mass spectrometry identity
HPLC tells you how pure something is, not what it is. Mass spectrometry (typically ESI-MS or MALDI-TOF) confirms identity by measuring the molecular mass. Larger peptides often appear as multiply charged ions (for example [M+3H]3+), and the deconvoluted mass should match the theoretical molecular weight within the instrument's tolerance.
This is the section that catches substitutions. For example, material sold as TB-500 may be full-length thymosin β4 (about 4,963 Da) or only the LKKTETQ fragment (under 900 Da). The mass spectrum tells you which you have.
Net peptide content
Lyophilized peptides contain the peptide itself plus counter-ions (acetate or TFA) and residual water. Net peptide content (NPC), measured by amino acid analysis or elemental nitrogen analysis, reports the fraction that is actually peptide. If a 10 mg vial has an NPC of 80%, it contains about 8 mg of peptide.
NPC matters for quantitative analytical work, such as preparing reference standards. Not every supplier reports it.
Other tests you may see
- Appearance: color and form, such as white lyophilized powder (or blue, for GHK-Cu).
- Endotoxin: bacterial lipopolysaccharide content, measured by the LAL test.
- Water content: by Karl Fischer titration.
- Counter-ion content: acetate or TFA, by ion chromatography.
Red flags
- No chromatogram or mass spectrum, only a summary table
- No lot number, or a lot number that does not match the vial
- The same COA used for years of production
- Mass spectrum peaks that do not match the theoretical molecular weight
- No identification of the testing laboratory
Our release process and the tests behind every lot are described on our quality standards page.
Worked example: checking a mass spectrum
Electrospray ionization adds protons to the molecule, so peaks appear at a mass-to-charge ratio (m/z) rather than at the molecular weight directly. For a peptide of mass M carrying n extra protons, each about 1.008 Da:
m/z = (M + n × 1.008) ÷ n
For BPC-157 (average MW 1419.5), expect [M+H]+ near m/z 1420.5 and [M+2H]2+ near m/z 710.8. For a larger peptide such as MOTS-c (MW 2174.6), the most intense peaks usually carry more charges: [M+2H]2+ near m/z 1088.3 and [M+3H]3+ near m/z 725.9. If a COA shows a deconvoluted mass, it should land on the theoretical value within the stated instrument tolerance.
Common impurities and their signatures
| Impurity | Mass shift | What it means |
|---|---|---|
| Deletion sequence | Minus one residue mass | A coupling step failed during synthesis |
| Methionine oxidation | +16 Da per oxidized Met | Exposure to air or oxidants, often during storage |
| Deamidation | +1 Da (0.98) | Asn or Gln converted to Asp or Glu, faster at neutral to basic pH |
| tert-Butyl adduct | +56 Da | Incomplete removal of a side-chain protecting group |
| Pbf on arginine | +252 Da | Incomplete removal of the arginine protecting group |
| TFA adduct | +114 Da | Trifluoroacetate associating in the ion source; usually an artifact rather than a covalent impurity |
Small minor peaks in the chromatogram are normal. What matters is that they are small, accounted for in the purity calculation, and do not correspond to a substituted or truncated product being sold as the target.
Questions to ask a supplier
- Can I see the full chromatogram and spectrum for the lot I will receive?
- Which laboratory ran the analysis, and is it independent?
- What HPLC method and wavelength were used?
- Is net peptide content or counter-ion content reported?
- How long after the test date was the lot shipped, and how was it stored?
Published April 2, 2026 by the RWP Laboratories research team. This article covers chemistry, analytical testing and storage for laboratory use only.
For research use only. All products are sold strictly for laboratory research and analytical use and are not for human consumption. They are not drugs, foods, cosmetics or supplements. We do not sell syringes or needles.
