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ForumsCOA & Analytical TestingLC-MS/MS vs HPLC for peptide analysis — November 2025 Page 3

LC-MS/MS vs HPLC for peptide analysis — November 2025

pat_auckland Wed, Apr 1, 2026 at 10:13 AM 19 replies 1,044 viewsPage 3 of 4
NeuroNate
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Apr 4, 2026 at 1:50 PM#11
Dr.CardioMD said:
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:…

Mass spectrometry confirmation for certificates of analysis: beyond HPLC purity, mass spec tells you the molecule's exact molecular weight. This confirms identity — not just purity.

Why this matters: HPLC can show you a "pure" compound, but if it's pure of the WRONG compound, purity is meaningless. Mass spec at m/z 4113.58 Da confirms it's actually semaglutide. My Janoshik report includes both HPLC and mass spec.

If your testing provider only offers HPLC, consider upgrading to include mass spec for at least your first batch from a new vendor.

Last edited: Apr 4, 2026 at 7:50 PM
39 12jason_paloalto, Dr.LeslieOBGYN, MikeNYC_runner and 36 others
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FitDadDave
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Apr 5, 2026 at 2:44 AM#12
ChrisMacros said:
Agreed, and mass spec answers a question purity does not: identity.

Saving this. It is the first explanation that did not require me to already understand it. Adding it to my notes with a link back to this thread.

40 13kim_atl_prep, sarah_TO, wendy_avl and 37 others
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kate.chem
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Apr 5, 2026 at 3:38 PM#13
Dr.CardioMD said:
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:…

I tested my certificates of analysis batch through Janoshik — 98.7% purity. That's within acceptable range for compounded peptides. Turnaround was 12 business days.

The test costs ~$121 but gives you actual HPLC chromatography data plus mass spec confirmation. For the peace of mind alone, it's worth every penny. I test every new batch from a new vendor.

41 14tyler_CSCS, VanRx_Mike, steve_okc and 38 others
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Sigma-Aldrich — Research-Grade Standards

Certified reference materials, analytical reagents, and research-grade standards for peptide verification. Trusted by laboratories worldwide.

Shop Reference Standards
Dr.PulmRoch
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Apr 6, 2026 at 4:32 AM#14
pat_auckland said:
I have a certificate reporting 99.2% purity and 6.1mg content on a vial sold as 5mg, and I cannot work out whether those two numbers are consistent or…

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: 99.5% purity, 0.1% total related substances. Clean chromatogram with single sharp peak at expected retention time. Exactly what you want to see.

42 15DebRD_ATL, KristenIndy, MarkLI_maint and 39 others
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Admin
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Apr 6, 2026 at 5:27 PM#15

Moderator note: the sourcing question belongs in the vendor section and has been split out. Report rather than reply if it drifts again.

43 16PeptideSynthNJ, Dr.KarenChen, Dr.NateNeph and 40 others
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