What is HPLC and Why Does It Matter?
High-performance liquid chromatography (HPLC) is the standard analytical method for determining peptide purity. A Certificate of Analysis usually includes a chromatogram that shows how much of the sample corresponds to the target peptide.
Understanding these reports helps researchers assess whether a material is suitable for a given protocol.
Reading a Chromatogram
The Main Peak
The largest peak represents the target peptide. The area under this peak, expressed as a percentage of the total peak area, is the reported purity value. A ≥99% result means the main peak accounts for at least 99% of the total integrated area.
Retention Time
Each compound has a characteristic retention time, meaning how long it takes to pass through the HPLC column. This supports identity confirmation when compared with the expected value.
Impurity Peaks
- Deletion sequences: peptides missing one or more amino acids.
- Truncated sequences: shortened versions of the target peptide.
- Oxidation products: oxidised forms of susceptible amino acids.
- TFA or acetate salts: counter-ions from synthesis and purification.
What Purity Level Do You Need?
- ≥95% purity: often suitable for screening and selected in vitro work.
- ≥98% purity: recommended for most research applications.
- ≥99% purity: preferred for quantitative work and publication-quality studies.
Verifying Your COA
- Check that the purity percentage matches or exceeds the stated specification.
- Compare retention time with the known value for the peptide.
- Look for a clean baseline with minimal noise.
- Confirm that the target peptide dominates as a single main peak.
Conclusion
HPLC reports provide a practical view of peptide purity and analytical quality. Always review the COA and ask suppliers about methods, columns, gradients, and acceptance criteria when the protocol requires tighter control.
For more information, visit our Quality Assurance page.