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Absorption: In Vivo Tests (Radiolabeled)

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Drug Discovery and Evaluation: Safety and Pharmacokinetic Assays

Abstract

The use of radiolabeled molecules allows a drug and its labeled metabolites to be followed throughout the body and excreta over time. The radioactivity concentration can be tracked in blood and plasma as well as in tissues. Whether the drug with its specific radioactivity administered to the body is completely captured can be proven by calculating the so-called mass balance.

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Notes

  1. 1.

    The estimation of the drug absorption (not to be mixed up with bioavailability) using non-radiolabeled drugs and not using the mass balance approach would be much less reliable, since the entity of metabolites cannot be captured in the matrices necessary to be followed, normally.

  2. 2.

    Half-life of 5,730 year for 14C, making an half-life correction unnecessary. As a weak β – radiator (up to 156 keV) the risk of handling is an acceptable compromise in the laboratory (protection area!).

  3. 3.

    Imagine the case of a projected nonabsorbable drug, for instance, for topical application, and the situation of having detected 3% absorption with a radioanalytical purity of the labeled drug of 97%. Or imagine the case of a minor part of radioactivity with a long terminal half-life suggesting a metabolite with an accumulation potency. The radioactivity represents the sum of the original compound and/or radioactive-labeled metabolites and not to forget possible synthetic side-products which can be present in traces (depending on the purity and content of the synthetic material). Discussing traces of radioactivity, for instance, traces crossing the placenta, keep in mind that these traces may be due to synthetic side-products. Thus, whenever possible try to use radiolabeled compound as clean as possible.

  4. 4.

    For instance, in case of urine samples when the quench is in the range of the calibration curve or bile samples.

  5. 5.

    For instance, with Solvable® from Perkin Elmer for dissolution and H2O2 for discoloration.

  6. 6.

    For instance, with a Tri-Carb® 307 combuster which can be equipped with a robot unit for automatic sample handling from Perkin Elmer. Since carry-over effects are not negligible in case a low radioactivity sample follows a high radioactivity sample, a reasonable arrangement of samples in a sequence is essential.

  7. 7.

    It might be necessary to use a normalized apparent collection interval for graphical reasons in case of dissimilar collection intervals.

  8. 8.

    Instead of the collection interval, the mean time of the collection period is often used in the graphic presentations.

  9. 9.

    Via determination of the rate constant which is calculated by linear regression of ln concentration on time using a least three data points which appeared to be randomly distributed about a single straight line. Half-lives were calculated as ln2/rate constant.

  10. 10.

    The following equation describes the percent of total radioactivity in plasma relative to total radioactivity in blood

    $$ \begin{array}{ll} {\text{Plasma/Blood}}\,{(}\% {)} & = \frac{{({V_B}) \times (P \times [1 - HCT])}}{{({V_B} \times B)}} \times 100 \\ & = \frac{{P \times (1 - HCT)}}{B} \times 100 \end{array} $$

    where

    • VB = volume of blood

    • P = drug concentration in plasma

    • B = drug concentration in blood

    • HCT = hematocrit

  11. 11.

    An example where this assumption is not fulfilled is described by Okuyama et al. (1997); consequently the authors only mention the ratio of AUCs after oral and intravenous administration and do not correlate this ratio to absorption.

  12. 12.

    Mass balance results in rats, mice, dogs, monkeys, and humans after administration of the same drug, including bile excretion results from rat, dog, monkey, and human (!), are described by Donglu Zhang.

  13. 13.

    For an example of a mass balance study in mice (besides rat and dog) refer to Miraglia L.

  14. 14.

    It might be necessary to use a normalized apparent collection interval for graphical reasons in case of dissimilar collection intervals.

  15. 15.

    Instead of the collection interval, the mean time of the collection period is often used in the graphic presentations.

  16. 16.

    Via determination of the rate constant which is calculated by linear regression of ln concentration on time using a least three data points which appeared to be randomly distributed about a single straight line. Half-lives were calculated as ln2/rate constant.

  17. 17.

    The following equation describes the percent of total radioactivity in plasma relative to total radioactivity in blood:

    $$ \begin{array}{ll}{\text{Plasma/Blood}}\,{(}\% {)} & = \frac{{({V_B}) \times (P \times [1 - HCT])}}{{({V_B} \times B)}} \times 100 \\ & = \frac{{P \times (1 - HCT)}}{B} \times 100 \end{array} $$

    where:

    • VB = Volume of blood

    • P = Drug concentration in plasma

    • B = Drug concentration in blood

    • HCT = Hematocrit

  18. 18.

    For instance, blood of exsanguination collected after heart puncture; jugularis puncture, jugularis or carotis catheter, retroorbital blood, blood from the vena femoralis, sublingual blood after short narcosis or blood from the tail vein.

  19. 19.

    Caution: The administration should not be done at the site of sample collection to avoid contamination. For instance, an i.v. administration into the vena femoralis after a short transient anesthesia can be recommended.

  20. 20.

    Instead of an oral an intra-duodenal administration should be chosen, when using anesthetized animals.

  21. 21.

    The procedure as described in the main part still has supporters. See Hoehle et al. (2009).

  22. 22.

    Can be easily constructed connecting a water-jet pump with a microwash bottle; an Eppendorf vessel is placed under the inlet tube; at the other end the inlet tube is fitted with a polyethylene tube and a microfunnel.

    The “microwash bottle” can be assembled from a 15-mL scintillation vessel with two openings in the lid and polyethylene tubing.

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  1. DI & A DMPK, Sanofi Pharma Deutschland GmbH, Industriepark Hoechst, Building G 876, 65926, Frankfurt, Germany

    Dr. Volker Krone

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    Jochen Maas

  2. Dieburg, Germany

    Franz J. Hock

  3. Idstein, Germany

    Dieter Mayer

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Krone, V. (2013). Absorption: In Vivo Tests (Radiolabeled). In: Vogel, H.G., Maas, J., Hock, F.J., Mayer, D. (eds) Drug Discovery and Evaluation: Safety and Pharmacokinetic Assays. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-25240-2_33

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