HPLC vs UPLC for peptide purity — method comparison study

Dr.EndoEP said:

The provenance questions are more useful than the numbers.

That reframing is the part I needed. Sending this to two other people who asked me the same thing last week.

From the other side of the consultation, briefly.

My HPLC chromatogram analysis for certificates of analysis — I sent my batch to Janoshik and here's what the results showed:

Main peak retention time: 16.7 minutes (consistent with semaglutide reference standard)
Peak purity: 98.5%
Related substances: 0.6% (single impurity at 10.4 minutes — likely a synthesis intermediate)
Mass spec: m/z 4113.5 (matches expected molecular weight)

This is a clean result. The mass spec confirmation is particularly important — it proves the molecule is actually semaglutide and not a substitution.

chris_chi24 said:

Reconstituted peptide stability data for certificates of analysis: I had Janoshik test my reconstituted vial at day 1, day 14, and day 28 stored in…

HPLC results interpretation for certificates of analysis: when you get your Janoshik report back, here's what the numbers mean:

• Purity %: Should be >97%. Below 95% is concerning.
• Retention time: Should match reference standard for the peptide
• Related substances: Degradation products should be <2% total
• Single impurity: No individual impurity should exceed 1%

My last batch: 99.4% purity, 0.0% total related substances. Clean chromatogram with single sharp peak at expected retention time. Exactly what you want to see.

kate.chem said:

Inter-laboratory variation comes from real methodological differences, not sloppiness: which reference standard was used and how it was itself…

My vendor review relevant to certificates of analysis: ordered 3 times from my current compounder. Consistency has been excellent.

Shipping: Next-day with cold pack
Packaging: Professional, discreet, proper labeling with lot/batch numbers
Purity: 99.0% average across orders (Janoshik verified)
Customer service: Responsive within 2 hours
Price: $144/month for semaglutide 1.7mg equivalent

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