Radioactivity determination tissues

The majority of the 14C-human ADME studies are conducted with a small number of healthy adult subjects (often between 6-8) and if bile collection is needed, a small group of additional subjects are included [228], Traditionally, due to ethical reasons, male subjects are selected for the 14C-ADME studies. Before the start of the 14C-ADME studies, study sponsors have the responsibility to determine stability of the radiolabel, purity of the radiolabel (distinguishing degradants from metabolites is very important), and conduct tissue distribution studies in nonclinical species preferably using quantitative whole-body autoradiography (QWBA) to detect radioactivity in tissues, organs, and excreta to determine the safe radioactivity dose. Nonclinical tissue distribution study data are extrapolated and used to show that radioactivity exposure of a specific tissue/organ will be well below the allowable limits to humans [229,230], Most of the 14C-human ADME studies consider a total radioactivity dose of 100[tCi or less to be safe [231],... 

Autoradiography is particularly well suited for determining the distribution of radioactivity in tissue. In principle, the distribution of radioactivity in a tissue could be assessed by gas flow counting, if the tissue was dissected, its parts weighed and uniformly spread as a dry film on a degassed planchet. However, very often it is difficult to identify exactly the part of the tissue that has been dissected. Furthermore, the fluids which surround the tissue in the body may often be very highly labelled and will contaminate the dissected specimen. The use of autoradiography readily overcomes these difficulties . 

Radioactivity Analysis. Samples of urine, feces, and tissues were combusted to COo and analyzed for radioactivity (5). By using this method the recovery of radioactivity from samples spiked with C was 95 dt 5%. To determine the radioactivity expired as CO2, 5-ml aliquots of the solution used to trap the CO2 were added to 15 ml of a scintillation counting solution containing 4 grams 2,5-diphenyloxazole (PPO) and 0.1 grams l,4-bis-2(5-phenyloxazolyl)-benzene (POPOP) per liter of 1 1 toluene 2-methoxyethanol. Samples were counted for radioactivity in a Nuclear Chicago Mark II liquid scintillation counter. Counting eflSciency was corrected by the internal standard technique. 

The uptake of TRA into cervical tissue was determined by measuring tissue radioactivity following insertion of the collagen sponge cervical cap containing tritium-labeled TRA. The TRA concentrations peaked at 4 hr and then diminished rapidly by 24 hr. Since measurements of blood samples revealed that no systemic absorption had occurred, high local concentrations over an extended period of time may be possible without systemic side effects. 

In the last case, this may be a physical problem resulting from incomplete penetration by the extraction solvent into the matrix. Alternatively, incomplete recovery of the analyte may result from chemical binding between the analyte and a constituent of the matrix. This is particularly important in the determination of drugs in body tissues where binding to proteins is known to occur. Problems of this kind are documented in the literature. If a new procedure is being developed, it is necessary to investigate the extraction step, e.g. by using radioactive tracers. 

After maximum radioactivity incorporation the protein is denatured and generally subjected to HPLC or gel-electrophoresis. Those methods separate the proteins from the specific tissue by size and the radioactivity distribution can be determined among the protein components. The specifically labeled biopolymers are distinguished simply by a competition experiment performed by the addition of excess of non-labeled parent ligand. It eliminates the radioactivity incorporation. 

Distribution. Cyanide is rapidly distributed by the blood throughout the body. In a study using orally administered radioactively labelled potassium cyanide, radioactivity detected in whole blood or plasma decreased rapidly within 6 hours. Of the low levels of radioactivity detected in the red blood cells, about 94% of the radioactivity recovered was found in the hemolysate of which 70% was detected in the heme fraction, 14-25% in globin, and only 5-10% in cell membranes (Farooqui and Ahmed 1982). Yamamoto et al. (1982) determined that the pattern of distribution of cyanide did not vary with the concentration used. Ballantyne (1983b) observed higher cyanide levels in whole blood than in serum in rabbits exposed dermally to hydrogen cyanide, potassium cyanide, and sodium cyanide. See Section 2.3.2.1 for specific studies on cyanide tissue distribution. 

Another important application of cDNAs is to identify specific proteins in a tissue homogenate or tissue section. Since cDNAs undergo complementary base pairing, adding a radioactively labelled cDNA to a homogenate or tissue slice will bind it to the complementary sequence by a process of hybridization. Thus the amount of radioactive cDNA that hybridizes to the tissue or tissue extract is a measure of the amount of mRNA that is complementary to it. When this procedure is undertaken on slices of brain, it is known as in situ hybridization. In this way it is possible to determine the distribution of specific receptors in a tissue by accurately determining the distribution of mRNA that encodes for the receptor protein. This is a particularly valuable technique for the administration of psychotropic drugs. 

Exposure through inhalation As most of the quantity of the cyclic siloxanes evaporate shortly after application of the PCPs or volatilize from electronics, adhesives and sealants, polishes and surface cleaners, etc., the main pathway of exposure is through inhalation. Plotzke et al. [286] studied the inhalation exposure to D4 by exposing rats to C-DA. Using liquid scintillation counting, it was determined that the retention of inhaled D4 in the body of the rats was 5-6%. The radioactivity reached maximum concentrations in the fat 24 h after exposure, but in the plasma and other tissues (except for fat) in only 3 h. The fat tissue acted as a depot because the elimination of the radioactivity from it was slower than from other tissues. 

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