How to Read a Peptide Certificate of Analysis

A peptide Certificate of Analysis (COA) is a batch-specific document that confirms three things: purity (by HPLC), identity (by mass spectrometry), and endotoxin level (by LAL assay). It is the primary evidence that a peptide is what the label says it is. Without a legitimate COA, every downstream experiment carries an invisible variable.

What does a peptide COA contain?

A legitimate research peptide COA documents six things: batch identifier, compound identity confirmation (mass spectrometry), purity (HPLC), endotoxin level, testing date, and the testing laboratory. Anything missing one of these is incomplete.

Reading the HPLC chromatogram

Reversed-phase HPLC is the standard for peptide purity analysis. The chromatogram shows absorbance (typically at 220 nm) against retention time. The peptide of interest appears as the dominant peak; impurities, truncated sequences, or oxidation products appear as smaller peaks before or after.

Purity is reported as the percentage area under the main peak relative to total integrated area. For research peptides, the practical threshold is 98%. Anything labelled research-grade should clear this. Pillar Research compounds average above 99%.

Look at the chromatogram itself, not just the headline number. A clean main peak with a flat baseline and minimal shoulder peaks is what you want. A messy baseline with multiple impurity peaks even at 98% nominal purity should raise questions.

Mass spectrometry confirmation

Mass spec confirms identity. The COA should report both the theoretical mass (calculated from the sequence) and the observed mass (from MS analysis). The two numbers should match to within experimental error (typically < 1 Da for ESI-MS on peptides under 5 kDa).

A mass mismatch is a red flag. Common causes are oxidation (+16 Da for each added oxygen), incomplete deprotection, or — in the worst case — the wrong compound entirely.

Endotoxin testing

For peptides used in cell culture or any application requiring sterility, endotoxin level matters. The LAL (Limulus Amoebocyte Lysate) assay is the standard, with results reported in endotoxin units per milligram (EU/mg). Sub-1 EU/mg is generally considered low-endotoxin grade.

Red flags

• No batch number on the COA, or a batch number that does not match your vial label
• Manufacturer-only COA with no independent third-party verification
• Missing testing date, or a date older than the manufacturing date
• Purity reported without a chromatogram image
• Mass spectrometry observed mass differing from theoretical by more than expected error
• Identical COAs across multiple batches (suggests batches are not being individually tested)

Verifying a Pillar Research COA

Every Pillar Research compound ships with a batch-specific COA. Match the batch number printed on the vial label to the batch number on the COA, confirm the testing laboratory and date are present, then walk through the HPLC chromatogram, mass spec, and endotoxin sections as above. If anything looks wrong, contact us — we will reship from a different batch.

You can download the batch-specific COA from any of our product pages, and a full overview of our verification methodology lives at /coa.