Dried Blood Spot Analysis for Rat and Dog Studies: Validation, Hematocrit, Toxicokinetics and Incurred Sample Reanalysis

Wickremsinhe, Enaksha

doi:10.4155/bio.15.12

Cited by 12 publications

(6 citation statements)

References 45 publications

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“…The used blood volume for spotting was 30 µL. According to Wickremsinhe (Wickremsinhe, 2015) there was no difference between 10, 20 or 30 µL spot volume. Thus, 30 µL was used for more sensitive determination of caffeine.…”

Section: Quantification Of Caffeinementioning

confidence: 99%

“…A 30-µL aliquot of whole venous blood was directly spotted on Whatman paper cards for DBS as already done by Wickremsinhe (Wickremsinhe, 2015). Blank blood samples for calibration standards were collected before drug administration and spotted directly.…”

Section: Rat Plasma Samplingmentioning

confidence: 99%

“…Thus, 30 µL was used for more sensitive determination of caffeine. The whole spot was punched avoiding variations in spread area caused by different hematocrit values (Wickremsinhe, 2015).…”

Section: Quantification Of Caffeinementioning

confidence: 99%

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Cytochrome P450 inhibition potential of new psychoactive substances of the tryptamine class

et al. 2016

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R., Maurer, Hans H., Cytochrome P450 inhibition potential of new psychoactive substances of the tryptamine class.Toxicology Letters http://dx.doi.org/10.1016/j.toxlet. 2015.11.013 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.  IC50 values were comparable to that of clinically relevant inhibitors. Members of the DALT family were strong inhibitors of CYP1A2 and CYP2D6 activity. 5-MeO-DALT showed in vivo inhibition against CYP1A2 activity. ABSTRACTNew psychoactive substances (NPS) are not tested for their cytochrome P450 (CYP) inhibition potential before consumption. Therefore, this potential was explored for tryptamine-derived NPS (TDNPS) including alpha-methyl tryptamines (AMTs), dimethyl tryptamines (DMTs), diallyl tryptamines (DALTs), and diisopropyl tryptamines (DiPTs) using test substrates preferred by the Food and Drug Administration in a cocktail assay. All tested TDNPS with the exception of DMT inhibited CYP2D6 activity with IC50 values below 100 µM. DALTs inhibited CYP2D6 activity similar to paroxetine and quinidine and CYP1A2 activity comparable to fluvoxamine. 5-Methoxy-N,N-diallyltryptamine reduced in vivo the caffeine metabolism in rats consistent with in vitro results. Five of the AMTs also inhibited CYP1A2 activity comparable to amiodarone. AMT and 6-F-AMT inhibited CYP2A6 activity in the range of the test inhibitor tranylcypromine. CYP2B6 activity was inhibited by 19 tryptamines, but weakly compared to efavirenz. CYP2C8 activity was inhibited by five of the tested TDNPS and three showed values comparable to trimethoprim and gemfibrozil. Six tryptamines inhibited CYP2C9 and seven CYP2C19 activities comparable to fluconazole and chloramphenicol, respectively. Nineteen compounds showed inhibition of CYP2E1 and 18 of CYP3A activity, respectively. These results showed that the CYP inhibition by TDNPS might be clinically relevant, but clinical studies are needed to explore this further.

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Section: Quantification Of Caffeinementioning

confidence: 99%

Section: Rat Plasma Samplingmentioning

confidence: 99%

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Cytochrome P450 inhibition potential of new psychoactive substances of the tryptamine class

et al. 2016

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“…However, the effect/impact of hematocrit on assay bias is minimized in nonclinical studies using purpose-bred animals, for which inter-animal variability is expected to be low relative to clinical populations (2,(22)(23)(24), but may be important when conducting studies in diseased animals where blood counts could be altered. Finally, in rare instances the stability of the analyte(s) in blood may interfere with the assay results, in which case the stability of suspect structures would need to be evaluated up front.…”

Section: Introductionmentioning

confidence: 99%

Evaluation and Optimization of Blood Micro-Sampling Methods: Serial Sampling in a Cross-Over Design from an Individual Mouse

et al. 2016

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-Purpose: Current practices applied to mouse pharmacokinetic (PK) studies often use large numbers of animals with sporadic or composite sampling that inadequately describe PK profiles. The purpose of this work was to evaluate and optimize blood microsampling techniques coupled with dried blood spot (DBS) and LC-MS/MS analysis to generate reliable PK data in mice. In addition, the feasibility of cross-over designs was assessed and recommendations are presented. Methods: The work describes a comprehensive evaluation of five blood microsampling techniques (tail clip, tail vein with needle hub, submandibular, retro-orbital, and saphenous bleeding) in CD-1 mice. The feasibility of blood sampling was evaluated based on animal observations, ease of bleeding, and ability to collect serial samples. Methotrexate, gemfibrozil and glipizide were used as test compounds and were dosed either orally or intravenously, followed by DBS collection and LC-MS/MS analysis to compare PK with various bleeding methods. Results: Submandibular and retro-orbital methods that required non-serial blood collections did not allow for inter-animal variability assessments and resulted in poorly described absorption and distribution kinetics. The submandibular and tail vein with needle-hub methods were the least favorable from a technical feasibility perspective. Serial bleeding was possible with cannulated animals or saphenous bleeding in non-cannulated animals. Conclusions: Of the methods that allowed serial sampling, the saphenous method when executed as described in this report, was most practical, reproducible and provided for assessment of inter-animal variability. It enabled the collection of complete exposure profiles from a single mouse and the conduct of an intravenous/oral cross-over study design. This methodology can be used routinely, it promotes the 3Rs principles by achieving reductions in the number of animals used, decreased restraints and animal stress, and improved the quality of data obtained in mouse PK studies.

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“…The expected hematocrit level in Sprague Dawley rats is within the range of 40 to 50 %167 . A hematocrit level of 45 % was selected for the preparation of control DBS samples to represent the average value expected in the Sprague Dawley rats used in this study168 . Blank rat whole blood was centrifuged at 7000 × g for 4 min and the plasma generated transferred to a clean eppendorf.The red blood cell (RBC) suspension and plasma was mixed in proportions (45:55, v/v) to give whole blood with an adjusted hematocrit of 45 %.…”

mentioning

confidence: 99%

Evaluation of paper spray mass spectrometry for direct analysis of small molecule drugs and hemoglobin constant spring peptide markers

Takyi-Williams¹

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Dried Blood Spot Analysis for Rat and Dog Studies: Validation, Hematocrit, Toxicokinetics and Incurred Sample Reanalysis

Abstract: Successfully validated and adopted DBS for preclinical GLP studies.

Cited by 12 publications

References 45 publications

Cytochrome P450 inhibition potential of new psychoactive substances of the tryptamine class

Cytochrome P450 inhibition potential of new psychoactive substances of the tryptamine class

Evaluation and Optimization of Blood Micro-Sampling Methods: Serial Sampling in a Cross-Over Design from an Individual Mouse

Evaluation of paper spray mass spectrometry for direct analysis of small molecule drugs and hemoglobin constant spring peptide markers

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