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c/hplc·submitted 2 years ago by u/ravi_bergstrom

how much of what we believe about HPLC actually comes from gradient threads

Correctionbranch of 13 comments

how much of what we believe about HPLC actually comes from gradient threads, and I want the answer with the reasoning attached rather than just the conclusion. Gave the same trace to two people I trust and got two integrations about a point apart. That was the most educational afternoon I have had here. Carryover…

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13 comments, started 2 years ago
u/yara_boateng463 points·2 years ago

Not convinced by that integration. Dropping the baseline there absorbs part of the shoulder into the main peak.

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u/ravi_bergstromOP315 points·2 years ago

Bought a second-hand instrument and learned more in six months of fixing it than in three years of reading traces.

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u/yara_bakken389 points·2 years ago

integration decisions move the number more than the sample does

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u/ravi_bergstromOP215 points·2 years ago

Small fix — 214nm, not 210. It matters for the comparison you are making with the other run.

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u/step_count_stan123 points·2 years ago

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/insulin_syringe_ian-9 points·2 years ago

I would not blame the sample yet. Everything you have described is consistent with the column rather than the vial.

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u/hugo_bergstrom1 point·2 years ago

the column has a history and it shows in the peak shape

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u/clara_danquah1 point·2 years ago

area percent is relative to what the detector saw and nothing else

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u/ferran_krastev1 point·2 years ago

reproducibility beats resolution if you only get one of them

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u/ravi_bergstromOP1 point·2 years ago

Correcting myself upthread: I said the gradient was linear and looking again it has a hold in the middle.

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u/samir_falk1 point·2 years ago

system suitability before you believe any number on the run

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u/ferran_krastev0 points·2 years ago

Not convinced by that integration.

yara_boateng is right — the integration choice is a decision and it should be stated alongside the result.

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u/erez_yilmaz1 point·2 years ago

Is that baseline drawn by the software or by hand?

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