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jag009 ★★★ NJ, 2024-08-12 15:17 (59 d 06:03 ago) Posting: # 24146 Views: 1,528 |
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Hi all, A dumb question since I am not an bioanalytical guy… Is there a way to find out whether the total drug concentration is measured from a bioanalytical report (no method development report, just bioanalytical and validation reports) if there is no sentence/discussion about that aspect? I read through a report that I was given and I didn't find info on this. The study is a typical BA/BE study comparing a few formulations vs a reference product under fast and fed. Thx J |
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Helmut ★★★   Vienna, Austria, 2024-08-12 15:45 (59 d 05:35 ago) @ jag009 Posting: # 24147 Views: 1,379 |
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Hi John, ❝ A dumb question … ❝ Is there a way to find out whether the total drug concentration is measured from a bioanalytical report (no method development report, just bioanalytical and validation reports) if there is no sentence/discussion about that aspect? I read through a report that I was given and I didn't find info on this. Are you talking about a drug with protein binding? That’s easy. Protein binding is caused by Van der Waals force, which is extremely weak. In a nutshell, in any extraction (liquid-liquid, solid-phase), or protein-precipitation the protein-drug bonds will be practically immediately cleaved and therefore, from the extract you get always the total concentration. If you are interested in the free concentration, you would have to use ultra-filtration or -centrifugation of the same sample, measure this concentration and the total. The protein binding: $$\small{PB(\%)=100\frac{C_\text{total}-C_\text{free}}{C_\text{total}}}$$ Important for new drugs but completely uninteresting in BE. Liposome encapsulated drugs and their relatives are different beasts. — Dif-tor heh smusma 🖖🏼 Довге життя Україна! ![[image]](https://static.bebac.at/pics/Blue_and_yellow_ribbon_UA.png) Helmut Schütz ![[image]](https://static.bebac.at/img/CC by.png) The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes |
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jag009 ★★★ NJ, 2024-08-12 16:35 (59 d 04:46 ago) @ Helmut Posting: # 24148 Views: 1,350 |
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Hi Helmut, ❝ ❝ A dumb question … ❝ You know that there are no dump questions. Just a way to catch your attention  ❝ Are you talking about a drug with protein binding? That’s easy. Protein binding is caused by Van der Waals force, which is extremely weak. In a nutshell, in any extraction (liquid-liquid, solid-phase), or protein-precipitation the protein-drug bonds will be practically immediately cleaved and therefore, from the extract you get always the total concentration. Yes, according to the text it was protein-precipitation extraction from plasma during sample pre-treatment phase. It's a drug w high protein binding. So it's total drug? Thx J |
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Helmut ★★★ Vienna, Austria, 2024-08-12 17:01 (59 d 04:20 ago) @ jag009 Posting: # 24149 Views: 1,346 |
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Hi John, ❝ ❝ Are you talking about a drug with protein binding? In […] protein-precipitation the protein-drug bonds will be practically immediately cleaved and therefore, from the extract you get always the total concentration. ❝ Yes, according to the text it was protein-precipitation extraction from plasma during sample pre-treatment phase. It's a drug w high protein binding. So it's total drug? — Dif-tor heh smusma 🖖🏼 Довге життя Україна! Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes |
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ElMaestro ★★★ Denmark, 2024-08-12 17:13 (59 d 04:08 ago) @ Helmut Posting: # 24150 Views: 1,338 |
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Hi, ❝ I know it’s Monday, but… Accumulation in rbcs? — Pass or fail! ElMaestro |
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Helmut ★★★ Vienna, Austria, 2024-08-12 17:30 (59 d 03:51 ago) @ ElMaestro Posting: # 24151 Views: 1,332 |
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Hi ElMaestro, ❝ Accumulation in rbcs? May I quote John? ❝ ❝ … it was protein-precipitation extraction from plasma Erys are ☣️ and disposed. — Dif-tor heh smusma 🖖🏼 Довге життя Україна! Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes |
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Ohlbe ★★★ France, 2024-08-12 23:37 (58 d 21:44 ago) @ jag009 Posting: # 24152 Views: 1,293 |
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Hi John, Define free and total... The draft FDA dabigatran product-specific guidance uses "free" to designate non-conjugated dabigatran and "total" for the sum of non-conjugated + conjugated dabigatran, measured after hydrolysis of the conjugates. I can live with "total", but "free" is a misnomer IMHO. In this particular case you will get 2 bioanalytical reports, one for "free" and one for "total", which should be clearly identified. — Regards Ohlbe |
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jag009 ★★★ NJ, 2024-08-13 08:27 (58 d 12:53 ago) @ Ohlbe Posting: # 24153 Views: 1,278 |
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Hi Ohlbe, Free = unbound drug concentration Total drug = unbound + bound (to plasma protein). FYI, FDA has a BEBA guidance for this drug but it doesn't ask to measure free drug or total drug. The reason I asked this q is because I emailed the lab and they told me they measured unbound drug but I have a feeling that they answered my question incorrectly. Thx J |
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Helmut ★★★ Vienna, Austria, 2024-08-13 09:31 (58 d 11:50 ago) @ Ohlbe Posting: # 24155 Views: 1,258 |
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Dear Ohlbe, ❝ The draft FDA dabigatran product-specific guidance uses "free" to designate non-conjugated dabigatran and "total" for the sum of non-conjugated + conjugated dabigatran, measured after hydrolysis of the conjugates. I can live with "total", but "free" is a misnomer IMHO. Reminds me on this ‘goody’. — Dif-tor heh smusma 🖖🏼 Довге життя Україна! Helmut Schütz The quality of responses received is directly proportional to the quality of the question asked. 🚮 Science Quotes |
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jag009 ★★★ NJ, 2024-08-26 18:19 (45 d 03:02 ago) @ Helmut Posting: # 24161 Views: 762 |
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Hi all, Update to this "story". It seems like the lab followed the definition of "free" from the draft FDA dabigatran guidance that Ohlbe cited, which was not correct (at least for me LoL). All good now. Thx J |
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Achievwin ★★ US, 2024-08-27 05:02 (44 d 16:18 ago) @ jag009 Posting: # 24164 Views: 752 |
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Total drug is usually referred for drugs that are conjugated and unconjugated. In case of highly protein bound drugs (e.g. Warfarin) they report free drug (taken by centrifugation or dialysis) and total (determined after protein precipitation) Way back in 80s there was an article in Journal of Chromatography, someone published a paper different precipitating agents (methanol, acetonitrile, TCA etc.) and recommended proportions of precipitating agents required for more than 95% protein precipitation and liberating the bound drug. Plasma + Acetonitrile (1:1) Plasma + MeOH (1:2) Plasma + TCA (1:4 I guess) recommended ACN is the best protein precipitating agent by denaturalization and therefore if you use 1:1 ratio you are certain aliquot contains 100% total drug, being so picky I used two step extraction 1:1 first and on the sediment repeat the precipitation again not to loose any drug in the sediment. Is 2 times sample prep for 1+1 precipitation @x (1:1: plasma: ACN). This was critical while working on Warfarin PK work. Long wobble for a Dumb question (Another product that require this kind of analysis is Iron product PK) |
Ing. Helmut Schütz