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

am I the only one who found method development harder than the injections

Lab Clean Column ×7 Cold Box ×3 Long Haul ×3

Been lurking in c/progresspics for about 3 months and finally have something worth posting.

Short version: week 82, 7.5mg, and gradient is the bit I still cannot get a straight answer on. I have read the wiki, I have read the last 9 threads, and the answers split roughly down the middle.

What I actually want to know is whether the people saying it does not matter have measured it, or whether it is just repeated confidently. Happy to be told I am overthinking this — I probably am — but I would rather overthink it now than at week 76.

15,631 up / 1,754 down90% upvoted41 commentsid 1rmnrp28 Sep 2023

41 comments

7 in this archive, depth 3

best — the order this archive was captured in

u/b12_baseline1.4k points·2 years ago

Strongly agree. Largest single impurity tells a different story than total purity.

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u/hplc_hobbyistMOD1.1k points·2 years ago

Retitled to remove editorialising. Put the evidence in the body.

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u/hamza_weiss812 points·2 years ago

Yeah, the chromatogram profile matters more than the headline number.

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u/week_four_wall531 points·2 years ago·edited

What was the column and what gradient?

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u/clara_danquah273 points·2 years ago·edited

instrument quality is less important than analyst consistency

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u/dizzy_on_standing741 points·2 years ago

Dead space in the needle hub holds a small but non-trivial volume. On low-volume draws that can be several units.

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u/elodie_grimaldiOP389 points·2 years ago

Ran the same vial on two columns, different purity, both right, different separation. Method matters more than equipment.

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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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