Radioactivity determination

The reactions were carried out in each case with a 0-1 per cent protein solution in phosphate buffer (pH 6 8), to which the radioactive phosphorofluoridate was added as a concentrated solution in dry ethanol. At the end of the reaction time, the product was dialysed for 20 hr. against running water, and precipitated at 0° by addition of two volumes of acetone. The precipitate was spun off and washed at —5° with ethanol and ether, and dried in air or over sulphuric acid. Samples of 25-50 mg. of dry powder were used for radioactivity determinations, and compared with a standard prepared by hydrolysing a weighed amount (ca. 1 mg.) of the phosphorofluoridate in n sodium hydroxide, neutralizing and drying. 

Four measurements per set per experiment. Radioactivity determined using C-toluene internal standard. 

The specific radioactivities of uracil were 14C, 0.6 mC/mAf T, 7.75 inC/mAf ratio T/14C = 12.9. The concentration of uracil was 10 3M. Photoproducts were isolated by paper chromatography with n-butanol/water, 86.14. Spots were eluted with water and radioactivity determined in a Tri Carb 314 EX (Packard Instruments). 

Reactions of atomic carbon, produced by nuclear reactions, with a number of hydrocarbons have been studied by Wolfgang and his collaborators (69). To minimize radiation induced secondary reactions which occur when use is made of C14, a technique has been developed using short-lived C11 produced by a neutron exchange reaction between a platinum foil and a C12 ion beam from a heavy ion accelerator. Part of the scattered Cu atoms has been allowed to penetrate through the thin brass foil wall of a brass vessel and come in contact with the compound wrhose reaction is studied. Products have been analyzed by gas chromatography using a technique of simultaneous mass and radioactivity determination. 

Cultures were grown for 3 days on medium containing normal calf serum. On the 4th day, the cells were fed medium containing normal or hypothyroid calf serum or without T3 (2 x 10 M) supplementation. On the 10th day in culture the cells were labeled with H2 S0 (400 pCi) (50 pCi/umol) for 16 hrs. The labeled lipids were extracted from the cells and the radioactivity determined. Values are mean s.d. of 4 experiments (22). 

For sample processing, pooled plasma samples (1 and 6 h, 2 ml) were mixed with acetonitrile (6 ml), vortexed vigorously, and centrifuged. The precipitates were again extracted with acetonitrile (2x) and the supernatants from previous extraction were combined. The pooled extracts were evaporated at room temperature under a stream of nitrogen and the residues were reconstituted in HPLC mobile phase. A portion of the sample was used for radioactivity determination, and the remaining sample used for HPLC profiling. 

Three aliquots from each (processed) sample were used for radioactivity determination. The aliquots from the urine samples were measured directly in the liquid scintillation counting procedure after addition of the scintillator Roth-rotiszint eco plus (Roth, Karlsruhe, Germany). The other matrix aliquots taken up on Combusto Cones (Canberra-Packard), weighed, dried at room temperature, were combusted in a Tricarb combuster (Canberra-Packard GmbH, Model 307) and the 14CC>2 formed was absorbed by Carbo-Sorb (Canberra-Packard). The subsequent radioactivity measurements were carried out by the liquid scintillation counting procedure in a ((-spectrometer (Canberra-Packard 2500 TR) after addition of the scintillator Permafluor E+ (Canberra-Packard). 

Samples were processed, the radioactivity determined and data evaluated as described above. 

In the assay described by Beeman and Rossomando (1989), a /xBondapak CI8 column (3.9 mm x 300 mm) was used to separate L-[2,3-3H]ornithine from [1,2-3H]putrescine. The mobile phase contained 0.05 M sodium phosphate (pH adjusted to 3.9 with phosphoric acid) containing 0.01 Af SDS and 36% acetonitrile. Fractions were collected and the radioactivity determined by a liquid scintillation counter The flow rate was 1.0 mL/min, and 0.5 mL fractions were collected. 

In practice the gas is allowed to react with base and the carbonate formed is precipitated as barium carbonate. This precipitate may be collected by filtration and its radioactivity determined by means of a gas flow counter. 

Radioactivity Determinations of Multiply Labeled Samples Using the External Standard Channels Ratio Technique... 

RECOVERY. A dual label recovery experiment, from plasma and PBS, was performed using 3H-all trans and 14C-cis retinoic acid. The normal extraction procedure was followed up to and including the HPLC purification step. No LC/MS analysis was performed. Aliquots were taken and total radioactivity determined after extraction and derivatization. Fractions (0.5 ml) from the HPLC were collected and counted. Counting was performed using a Beckman Model LC3801 liquid scintillation counter. Radioactivity was corrected for spillover and quench. 

Mixtures of radiolabelled tefluthrin (6 pg), water (30 ml) and soil (2.0 g) were placed in centrifuge tubes, shaken (24 hr), the supernatant removed by centrifugation and radioactivity determined as above. The radioactivity of the filtered solids was determined after filtering and treatment with a cocktail specifically formulated to dissolve membrane filters (Zhou et al., 1995c). All experiments were carried out in triplicate and control tubes were used without soil for mass balance and assessment of glass-sorbed tefluthrin. 

P Bore, et al. Pharmacokinetics of a new anticancer drug, navelbine, in patients— Comparative study of radioimmunologic and radioactive determination methods. Cancer Chemother Pharmacol 23 247, 1989. 

It is also necessary to ensure that the amount of radiolabelled internal standard added to each extract is not so small that the percentage error of the radioactivity determination is the limiting factor in the analysis. On the other hand, it is important that the cold carrier associated with the radiolabelled internal standard should amount to, as a rule-of-thumb, no more than 20% of the total lAA in the extract. Otherwise the distinction between endogenous and exogenous lAA will become somewhat blurred. 

---