A peptide COA, or certificate of analysis, is the document that links a batch of material to the analytical tests performed on it. For anyone comparing suppliers, being able to read one is essential. This guide walks through the main sections, explains what an HPLC peptide purity figure does and does not tell you, and shows how mass spectrometry confirms that the peptide is the one on the label.
For laboratory research use only. Not for human or veterinary use. See our research use only policy.
At a glance
- Name: Certificate of analysis (COA)
- Synonyms: CoA, analytical certificate, batch certificate
- Purity method: reversed-phase high performance liquid chromatography (RP-HPLC)
- Identity method: mass spectrometry, commonly ESI-MS or MALDI-TOF
- Our purity claim: >99% (HPLC)
- Form: Lyophilised powder
- Storage: 2-8°C, dark, upright - do not freeze. Keep sealed until use.
What a certificate of analysis contains
Layouts vary between laboratories, but most peptide certificates cover the same core information. When reading one, look for:
- Product identity: name, sequence, molecular formula and theoretical molecular weight
- Batch identification and date of analysis
- Appearance: for example white lyophilised powder
- Purity by HPLC, expressed as a percentage, with the chromatogram
- Identity by mass spectrometry, showing the observed mass against the theoretical mass
- The name of the testing laboratory and an authorised signature
Some certificates also report peptide content, water content, counter-ion content or residual solvents. These are separate measurements from purity and are discussed below.
HPLC: what the purity figure means
Reversed-phase HPLC separates the components of a sample according to how strongly they interact with a hydrophobic stationary phase as the proportion of organic solvent in the mobile phase increases. A detector, usually UV absorbance at around 210 to 220 nm where the peptide bond absorbs, records each component as a peak. The area of the main peak divided by the total area of all peaks gives the purity percentage.
Reading the chromatogram
The chromatogram should show one dominant peak at a defined retention time, with any impurities appearing as small peaks either side. Typical impurities in synthetic peptides include deletion sequences missing one amino acid, truncated chains, incompletely deprotected sequences and oxidised or deamidated variants. A clean baseline and a sharp, symmetrical main peak are good signs. A purity figure quoted without a chromatogram is harder to assess.
Limits of an HPLC purity figure
HPLC purity is a relative measure: it describes what proportion of the UV-absorbing material is the main peptide. It does not tell you how much of the powder by weight is peptide, because water and counter-ions such as acetate or trifluoroacetate do not appear as peaks under these conditions. It also cannot by itself prove identity, since an incorrect peptide can still produce a single clean peak. That is why HPLC is paired with mass spectrometry. Our article on how to read a purity figure looks at these points in more detail.
Mass spectrometry: confirming identity
Mass spectrometry measures the mass-to-charge ratio of ionised molecules. For peptides, electrospray ionisation (ESI) often produces several multiply charged ions, which software converts back to a single molecular mass. MALDI-TOF more commonly produces a singly protonated ion, recorded as [M+H]+.
On the certificate, compare the observed mass with the theoretical molecular weight calculated from the sequence. Agreement within the expected tolerance of the instrument supports the conclusion that the material has the correct composition. Be aware of whether the value given is the monoisotopic mass or the average molecular weight, as these differ slightly for larger molecules. For example, our catalogue lists SNAP-8 at 1075.2 g/mol and SS-31 at 639.8 g/mol as average molecular weights.
Other measurements you may see
Peptide content
Net peptide content, often measured by amino acid analysis or elemental nitrogen analysis, reports what percentage of the powder by weight is peptide. Values well below 100% are normal, because lyophilised peptides contain bound water and counter-ions.
Water and counter-ion content
Karl Fischer titration measures water content, and ion chromatography can quantify acetate or trifluoroacetate. These figures explain the gap between purity and peptide content.
A checklist for comparing certificates
- Does the sequence and formula on the COA match the product page?
- Is the HPLC chromatogram included, not just a number?
- Does the observed mass match the theoretical molecular weight?
- Is the testing laboratory named and the document dated and signed?
- Are the units and methods stated clearly?
Our peptides are specified at >99% purity by HPLC. For an overview of how purity, identity and compliance fit together when choosing a supplier, read the research peptides UK buyer's guide. Correct storage of lyophilised peptides then protects that purity after delivery.
View our research peptides in the catalogue.
Frequently asked questions
What is a peptide COA?
A peptide COA, or certificate of analysis, is a document reporting the analytical tests carried out on a batch of peptide, typically HPLC purity and mass spectrometry identity, along with appearance and product details.
What does peptide purity by HPLC mean?
It is the area of the main peptide peak as a percentage of the total peak area in the HPLC chromatogram. It describes the proportion of related impurities, not the peptide content by weight.
Why is mass spectrometry needed as well as HPLC?
HPLC shows how pure a sample is but cannot confirm what the main component is. Mass spectrometry measures molecular mass, which is compared with the theoretical value for the sequence to confirm identity.
Is purity the same as peptide content?
No. Purity describes the proportion of the target peptide relative to peptide-related impurities. Peptide content describes how much of the powder by weight is peptide, after accounting for water and counter-ions.
What purity are your peptides?
Our peptides are specified at >99% purity by HPLC.
References
- 1. Mant CT, Chen Y, Yan Z, Popa TV, Kovacs JM, Mills JB, Tripet BP, Hodges RS. HPLC analysis and purification of peptides. Methods in Molecular Biology. 2007;386:3-55.
- 2. International Council for Harmonisation. ICH Q2(R1): Validation of Analytical Procedures: Text and Methodology. 2005.
- 3. International Council for Harmonisation. ICH Q6B: Specifications: Test Procedures and Acceptance Criteria for Biotechnological/Biological Products. 1999.
- 4. United States Pharmacopeia. General Chapter <621> Chromatography. USP-NF.
For laboratory research use only. Not for human or veterinary use. See our research use only policy.