Radioactive compounds, analysis

Another technique which has much potential in analysis, but which to date has had only limited use, is the formation of radioactive derivatives of non-radioactive compounds for quantitation by radiocounting. A radiolabeled reagent is used to form the derivative. This approach has been of use in combination with chromatography. The advantage of this technique is that it avoids problems of sample background which are often associated with spectrophotometric methods. The 14C-methylation of carboxylic acids and the 14 C-acetylation of hydroxyl groups have been studied [39,40]. These methods are quantitative and the sensitivity is dependent on the activity of the radioactive group added to the molecules. The radioactive derivatization of lipids has been reviewed [41]. 

Radiochemical detection is another approach to analysis. The selectivity of such a system makes it very useful for radioactive compounds or suitable derivatives. 

Dilution analysis can also be used to determine how much of a radioactive compound is present in a mixture, if the specific activity of the compound is known. For example, if an organism is grown on labeled or uniformly... 

The European directorate for the quality of medicines (EDQM), in charge of the secretariat of the European Pharmacopeia Commission, is responsible initially for finding and compiling information, and it asks the mannfactnrers and snppliers for samples for analysis and in some cases, for nse as reference snbstances. In the case of radioactive compounds, the samples are sent directly to the experts, as the laboratory of the EDQM is not allowed to handle radioactivity. 

FIGURE 10.5 Analysis of two radioactive compounds using stop flow HPLC-RFD (a) nonstop mode (b) stop by fraction mode and (c) stop by level mode. Radioactivity in stop by fraction and stop by level analyses were counted for 1 min each fraction. 

Inverse isotope dilution analysis. This variation is a method to determine not an inactive compound hut a radioactive one. To he determined is the mass X of a certain radioactive compound (or ion), whose specific activity is known to be So, in a sample. To the sample solution is added a solution containing W of the same but inactive compound (spike). If the specific activity of the compound after this addition and thorough blending is taken to be S, then... 

The weak point of the above inverse isotope dilution analysis is that the specific activity So of the compound to be determined in the original sample solution must be known. This can be avoided by double isotope dilution analysis. In this type of analysis, two portions of exactly the same amount are taken from the sample solution. The mass of the radioactive compound in each portion is taken to be X. To the portions are added different masses Wi and W2 of the inactive compound. Taking the specific activities of the compound in the portions after this to be Si and S2,... 

It may be pointed out here that non-radioactive compounds can be detected autoradiographically after neutron activation or in the form of labelled derivatives. These procedures are discussed in more detail in section V, Analysis by Means of Radioisotopes . 

This procedure is used especially in biochemistry for the quantitative determination of small amounts of compounds which cannot be performed accurately by any other method. The substance to be determined must be available in radioactive form and its activity must be known. A weighed amount of the radioactive compound is mixed with the sample and some of the compound to be determined is then isolated in a pure state. The amount of compound originally present M-j) in the mixture for analysis can be calculated from the formula ... 

The radioactive compounds were revealed by autoradiography after exposure to Kodak Regulix films, and analysed by scanning (ScanJet II C-Hewlett Packard - Logiciel Desk scan II, Scan analysis). 

ImmunO lSS iy. Chemiluminescence compounds (eg, acridinium esters and sulfonamides, isoluminol), luciferases (eg, firefly, marine bacterial, Benilla and Varela luciferase), photoproteins (eg, aequorin, Benilld), and components of bioluminescence reactions have been tested as replacements for radioactive labels in both competitive and sandwich-type immunoassays. Acridinium ester labels are used extensively in routine clinical immunoassay analysis designed to detect a wide range of hormones, cancer markers, specific antibodies, specific proteins, and therapeutic dmgs. An acridinium ester label produces a flash of light when it reacts with an alkaline solution of hydrogen peroxide. The detection limit for the label is 0.5 amol. 

Radiochemical tracers, compounds labeled with radioisotopes (qv), have become one of the most powerful tools for detection and analysis in research, and to a limited extent in clinical diagnosis (see Medical IMAGING TECHNOLOGY). A molecule or chemical is labeled using a radioisotope either by substituting a radioactive atom for a corresponding stable atom in the compound, such as substituting for H, for or for P, and for for... 

One of the first decisions to be made when designing an experiment is the method of detection to be used with a particular solute. If radiolabeled material is available, a simple method of analysis is to count the radiolabel appearing in the receiver compartment as a function of time. While convenient, this can be a dangerous practice. Depending upon the type of radioisotope, its position in the molecule, and its specific activity, radiolabeled compounds can be subject to a variety of chemical and solution-catalyzed degradation pathways. If the stock solution contains a significant amount of radioactive impurities or generates them as a result of solution instability, then the possibility for preferential transport of... 

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