Research resource
A plain-English guide to High-Performance Liquid Chromatography (HPLC) for peptide purity verification, what the percentage actually measures, how to read a Janoshik Analytical lab report, and why a batch-specific Certificate of Analysis (COA) matters for research-grade compounds.
HPLC is a separation technique. A small sample of the peptide is dissolved in a mobile-phase solvent and pumped at high pressure through a column packed with a stationary phase (typically silica with a chemically bonded C18 hydrocarbon coating, for reversed-phase HPLC). Different molecules travel through the column at different speeds depending on their interaction with the stationary phase. A detector at the column outlet records each molecule as it elutes, producing a chromatogram - a 2D plot of detector response (UV absorbance at 214 nm or 220 nm) against time.
The pure target peptide produces a single sharp peak. Any impurities - truncated synthesis byproducts, deamidated variants, oxidised residues, or unrelated contaminants - produce additional peaks at different retention times. The purity percentage is calculated as the area under the target peak divided by the total area under all peaks in the chromatogram, multiplied by 100. A 99% purity result means the target peak accounts for 99% of all detector signal in the chromatogram.
The difference between 95% and 99% purity is not just a five-percent shift - it is a five-fold difference in the impurity load. A 95% pure peptide has 5% impurities; a 99%+ pure peptide has less than 1%. For in-vitro receptor assays, especially competition-binding studies and dose-response curves, that impurity differential can produce measurably different results. Synthesis byproducts often include truncated peptide variants that compete for the same receptor - an impurity present at 5% can shift apparent EC50 values significantly.
For published research-quality data, the standard is 99%+ HPLC purity verified by an independent lab. Every batch of every compound in our catalogue meets this standard.
A supplier can self-test, but the results carry no independent verification. The supplier has commercial incentive to pass their own batches. This is the same conflict of interest that applies to any quality-control system - which is why pharmaceutical and clinical laboratories use independent third-party testing for release.
Janoshik Analytical (based in the Czech Republic) has emerged as the standard third-party HPLC testing laboratory for the European research-peptide industry. Janoshik is independent of the suppliers it tests, runs reversed-phase HPLC + mass spectrometry on submitted samples, and issues batch-specific lab reports with unique verification codes. Buyers can match the verification code on their COA against Janoshik's published records to confirm authenticity.
We use Janoshik for batch HPLC testing on the compounds in our catalogue. Where a report has been issued for the current batch, it ships with the order. View sample reports →
A typical Janoshik COA contains the following elements:
For three real examples see the Purity page, where we publish a sample of recent Janoshik reports for Tirzepatide, BPC-157 and other compounds.
Purity is measured at the point of testing. After that, several factors can reduce purity in storage:
For these reasons, lyophilised peptides stored at 2–8 °C have 24+ month shelf lives, while reconstituted solutions typically expire after 14–30 days at the same temperature. See the Research Handling Guide for compound-specific storage recommendations.
HPLC measures separation and quantification of peaks - it does not directly measure biological activity. Two batches with identical 99% HPLC purity could have subtly different bioactivity if the active conformation differs. For most research peptides this is not an issue (the standard chemistries are well-validated), but it is worth knowing the limit. For receptor-binding assays where bioactivity matters most, additional functional QC tests can complement HPLC - though for in-vitro reference reagents 99%+ HPLC is the standard sufficient quality bar.