The number printed largest on a vial is the one that matters least for a calculation. “10 mg” on a label is gross mass of the lyophilized cake. The peptide inside it is less than that, sometimes much less, and a stock solution made from the gross figure will be wrong by the same margin.
What is in a lyophilized cake
Freeze-drying leaves the peptide together with everything that came with it out of purification:
- Counter-ions. Peptides are purified as salts, most often trifluoroacetate (TFA) or acetate. Each basic residue carries one, and for a short peptide the counter-ion can be 10–30% of the mass.
- Water. Lyophilized peptides are hygroscopic. Residual water is typically a few percent and rises every time the vial is opened.
- Everything HPLC counted as “other”. A 99% purity figure leaves 1% of UV-absorbing impurities, which also weigh something.
Net peptide content is the assayed fraction of the cake that is peptide. Typical values run from about 70% to 90% depending on sequence and salt form. It is measured by amino-acid analysis, nitrogen determination or a calibrated UV method; the certificate should say which.
The arithmetic
Stock concentration is peptide mass divided by solvent volume. With net content the mass term is corrected:
peptide mass = gross mass × net content e.g. 10 mg × 0.82 = 8.2 mg concentration = peptide mass ÷ solvent volume e.g. 8.2 mg ÷ 2 mL = 4.1 mg/mL (not 5 mg/mL)
An 18% error at the stock carries through every dilution unchanged. The concentration calculator takes net content as its first field so the correction is made once, at the top.
When the certificate has no net content line
Not every lot is assayed for it; the test costs more than HPLC and MS. Where the line is absent, the honest options are to run the assay yourself, to use a literature value for the same salt form as an estimate and say so in your notes, or to treat results from that lot as relative rather than absolute. What not to do is read “10 mg” as 10 mg of peptide.
Salt form matters for more than mass
TFA salts can affect cell-based assays at the concentrations a stock solution reaches. If the certificate states the salt form, note it; if your assay is sensitive, an acetate salt or a TFA-removal step may be appropriate. This is a property of the material, not advice about its use.
Aureum publishes net peptide content on a lot’s COA page whenever the lot has been assayed for it. Research materials are supplied for in-vitro laboratory use only, not for human or veterinary use.