Liquid-scintillation counting, tritium-labeled

Isotopic dilution techniques were used to determine residual, tritium-labeled gibberellic acid in potatoes, grapes, and various products derived from barley The seeds, the young plants, or the fruit were treated with labeled gibberellic acid and analyzed at the end of the growth period by extraction of the labeled residue in the presence of carrier gibberellic acid, isolation of a pure crystalline specimen, and subsequent assay by liquid scintillation counting ... 

Tritium as a low energy beta-emitter is most conveniently measured by liquid scintillation counting. In general, the tritium measurement is the most complicated of all the radionuclide labels. The difficulties are related to the character of the sample which is tritium labeled. Tritium labeling is not suitable in all cases ir. which the system yields high chemiluminiscence the samples are coloured or turbid. Recently, however, new types of liquid scintillation counters appeared which make it possible to count tritium even under such complicated conditions (Beckman, Packard, Kontron, Berthold). 

C-Carbon is now utilized more often in accordance with the development of new labeling techniques which involve so-call i natural labeling, i.e. replaces by isotopic exchange in molecules of the proteins to be labeled. The application of is more convenient than tritium labeling. Radiocarbon may be measure by various gas flow counters and semiconductor detectors, but liquid scintillation counting is still preferred. 

In these experiments a tritiated substrate was mixed with 1 -labeled protonated (for H/T KIE experiment) or deuterated (for D/T KIE experiment) substrate. The mixture was allowed to react in the presence of enzyme, under defined conditions of pH and temperature, and quenched at different fractional conversions. The quenched mixtures were analyzed by HPLC and liquid scintillation counting to determine the fractional conversion (f, determined from the counting) and tritium to ratio in the products ([ H/I cjy and pH/l4C]oo for the ratio at the time point and the infinity point, respectively). The L/T KIE (also denoted by (V/K)l or kt/kj) were calculated by equation 1 (Melander Saunders, 1987) ... 

In the first experiment, tritium-labeled M-3-G was added to an ongoing fermentation of Cabernet Sauvignon grapes after the third day of the fermentation. This coincided with the maximum level of M-3-G in the must. Figure 1. After 24 hours, a new analysis of both malvidin-3-glucose by chromatography and the concentration of radioactivity by liquid scintillation counting showed decreases. However, the radioactivity had dropped almost by half, while the M-3-G level had decreased only by about 13%. 

Automatic sample preparation methods for the determination of oarbon-14 and/or tritium, and carbon-14 and/or sulfur-36 in dual labelled samples by liquid scintillation counting are presented. The sample is burnt in a stream of oxygen, and the combustion products carrying radioisotopes are subsequently separated and collected for radioactivity determination. Tritium is measured as water, carbon-14 as "carbamate" and sulfur-35 as sulfuric acid. The procedures run automatically, they are free of memory effect and cross contamination, and provide quantitative recovery. 

With this in view, automatic isotcpe analytical processes have been developed. Of these methods, the sample preparation techniques used for simultaneous determination of carbon-14/ tritium and carbon-14/sulfur-35 in dual labelled organic compounds and biological materials hy liquid scintillation counting are presented. For the determinaticai of carbon-14/sulfur-35 an iirproved versicai of a previously published method (Gacs et al., 1978a) is described. The principles of the procedures are shewn in Fig.l and Fig.2. 

In a similar study, Brewis et al prepared sandwiches of a nylon-epoxy adhesive between aluminum adherends and exposed them to water vapor labeled with tritium for up to 6 weeks at 50°C. The bonds were then cut into small rectangular sections and the fractional water uptake measured by liquid scintillation counting. The values were compared with those calculated for diffusion and were found to agree well for the outer pieces. For the inner pieces the measured uptake was less than the predicted value. The adhesive contained a carrier and the results seem to imply that this hinders the passage of water into the adhesive bond. These results show... 

Another radionuclide that has been commonly used in physiological studies of terrestrial animals is tritium. As stated in Section 8.2.1, water can be labeled as HTO and the compound used to study the water pool in plants. Somewhat similarly, total body water and water turnover in mammals can be studied. Longhurst et aL (1970) utilized HTO to compare water kinetics in winter and summer Columbian black-tailed deer (Odocoileus hemionus columbianus) and domestic sheet (Ovis aries). The animals were injected intravenously with 8 mCi HTO, and blood samples were collected periodically and examined for radioactivity with a liquid scintillation counting system. In a water-turnover study in mule deer (Odocoileus hemionus), Knox et aL (1969) also used HTO and observed that the biological half-life of HTO differed in animals maintained in a 0.9 X 1.5 m metabolism stall from that in the same animals maintained in a 4.6 X 5.8 m room. 

It is now possible to obtain the catecholamines and many related compounds in a radioactively labelled form. The introduction of liquid-scintillation counting techniques, together with the availability of tritium-labelled compounds of very high specific activities (> 5 Ci/mmole), has been a very important factor in catecholamine research. The use of such radioactively labelled compounds has been critical for the elucidation of the pathways involved in catecholamine metabolism, and has provided the basis for many other recent advances in this field of research. The sensitivity with which tritiated catecholamines can be detected and accurately measured is very high, with a limit of the order of 1 pg (S x 10 moles). 

The total amount of labeled drug to prepare, in terms of radioactivity, depends on the anticipated studies, the radioisotope, and methods for detecting the radioactivity. In a typical drug-metabolism study Involving 3 animal species and man, using liquid scintillation counting techniques, approximately one mCi of a carbon-14 or sulfur-35 labeled drug will suffice. Similar studies with tritium would require 2 to 4 mCl since the detection efficiency for tritium by the same techniques is only one-half to one-fourth that for carbon-14 and sulfur-35. 

Woods and O Neal (1965) used HTO in a study of water uptake by small oak trees (Quercus laevis, Q, incana, and Q, stellata) in the sandhills of South Carolina. Pipes were driven into the ground at three depths (5, 35.5, and 66 cm), and labeled water was placed within these pipes. Sample plants were at an average distance of 2.1-2.4 m from these pipes. Plastic bags enclosing the ends of branches were used to collect transpired water that was counted for tritium by the liquid scintillation method. 

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