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c/hplc·posted 2 years ago by u/zeynep_zielinski

[Lab] QSC cagri — Janoshik came back 98.6% against a claimed 98.5%

Lab Cold Box ×4 Well Actually ×2 Long Haul ×3

Numbers in the title, numbers in the post. QSC cagri — Janoshik came back 98.6% against a claimed 98.5%.

Pulling the figures out of the title: 98.6% and 98.5%. All of it is written down as it happened rather than reconstructed.

System suitability — repeat injections, tailing factor, plate count, RSD on area — is what tells you the instrument was fit for the measurement that day. Without it, the purity figure is unanchored.

UV response is not uniform across species. At 214nm you are looking at the peptide bond, which is why it is the working wavelength for this class; at 280nm you only see tryptophan, tyrosine and phenylalanine.

Happy to answer the boring questions. Those are usually the ones worth asking.

56,569 up / 42,617 down57% upvoted36 commentsid vpo0fx9 Mar 2024
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36 comments

6 in this archive, depth 4

best — the order this archive was captured in

u/micro_bump_mick2.5k points·2 years ago

Reading a trace posted here, in the order I actually look at things.

Axes first — if the wavelength and the time axis are not labelled, I stop. Then the baseline: where has it been drawn, and does the drawing absorb anything. Then peak shape: fronting, tailing, a shoulder that never resolves. Then the blank, if one was run, for carryover.

Only after all of that do I look at the percentage, and by then I usually know how much weight it deserves. The number is the last thing on the page and the first thing everybody argues about, which is exactly backwards.

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u/ahmed_rasmussen1.1k points·2 years ago·edited

Push back: a longer run is not automatically better resolution. You are trading peak width for time and the ratio is what matters.

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u/rasmus_petrescu1.7k points·2 years ago

Area percent is the integrated area of your peak over the total integrated area at one wavelength on one gradient. Change any of those and the number changes without the sample changing.

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u/baseline_drifteranalytical-5 points·2 years ago

Disagree. Inter-lab spread of a point or two on this assay is ordinary and calling it a discrepancy misleads people.

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u/salma_vasquez1 point·2 years ago

axis labels or the trace is decoration

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u/baseline_drifteranalytical1 point·2 years ago

214nm sees the peptide bond, 280nm sees the aromatics

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About c/hplc

The analytical deep end: RP-HPLC methods, gradient design, integration decisions, LC-MS confirmation, amino-acid analysis, water content, and why two labs can report 98.1% and 96.4% on the same vial and both be right. Post the chromatogram, not the summary line.

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