“≥99% purity” is printed on nearly every research peptide label, but the number has a specific technical meaning. Understanding it helps you judge documentation critically and compare materials fairly.
How HPLC purity is measured
Reversed-phase HPLC separates the components of a sample as they pass through a column under a solvent gradient. A UV detector, usually set at 214–220 nm, records a peak for each component as it elutes. Purity is reported as area percent: the area of the main (target) peak divided by the total area of all detected peaks.
So “99% by HPLC” means 99% of the UV-detected, column-eluting material is the target peptide. It is a relative measure of related impurities, not an absolute measure of what is in the vial.
What HPLC purity does not include
- Water: lyophilized peptides retain bound water.
- Counter-ions: peptides are salts, commonly acetate or trifluoroacetate (TFA), and the counter-ion adds mass.
- Non-UV-absorbing substances and anything that doesn’t elute under the method’s conditions.
Purity vs. net peptide content
Because of water and counter-ions, a peptide with 99% HPLC purity might have a net peptide content of roughly 70–90% by weight. Net peptide content is a separate measurement, typically by amino acid analysis or nitrogen determination. When a vial is described as containing a given mass of peptide, it’s worth knowing whether that figure refers to the gross powder weight or to net peptide.
Common peptide impurities
Most research peptides are made by solid-phase peptide synthesis (SPPS), building the chain one amino acid at a time. Typical impurities include:
- Deletion sequences: chains missing one residue because a coupling step was incomplete.
- Truncated sequences: chains that stopped growing early.
- Incompletely deprotected peptides: chains still carrying a protecting group from synthesis.
- Oxidized or deamidated forms of the target peptide.
- Diastereomers from racemization of an amino acid during synthesis.
Why identity testing matters too
A clean chromatogram shows the sample is mostly one substance. It doesn’t prove which substance. Mass spectrometry confirms identity by matching the observed molecular mass to the theoretical mass of the sequence. A credible COA reports both. Read more in How to Read a Peptide Certificate of Analysis.
Purity grades at a glance
- Crude: unpurified synthesis product, used for screening.
- >90–95%: common for many in-vitro applications.
- >98–99%: used where related impurities could confound results, such as quantitative or analytical work.
Lot-specific purity and identity results for our materials are published in the COA Library.
The Pure Vial Co. supplies research materials for laboratory, analytical, and scientific research use only. Nothing in this guide is medical advice or an indication that any material is suitable for human or veterinary use. See our Research Disclaimer.
Leave a Reply