Flow-through radioactivity detectors

LC-ARC is a novel online RAD method that is 10-20 times more sensitive than the conventional flow-through radioactivity detectors [219] and manufactured by Aim Research Company. It is a powerful technique to detect compounds with low... 

Following its emergence from the other end of the column, the eluent flows through the detector. Detectors operate on various principles. For example, some monitor the ultraviolet, visible, or fluorometric properties of molecules others monitor radioactivity and still others monitor differences in oxidation-reduction potential and refractive index. These detectors are listed in Table 2.1 together with some examples of the specific reactions with which they have been used. 

Radiolabeled products were separated from substrates by chromatography on a Merck Qg column (5 /an). The mobile phase contained 0.1 M sodium acetate, 0.1 M citric acid, 0.1 mAf sodium octylsulfate, 0.15 mAf EDTA, and 0.2 mAf dibutylamine in 10% methanol (v/v). The pH was 4 for the monoamine oxidase assay and 3.7 for phenol sulfotransferase. A flow-through radioisotope detector was used to quantitate the amount of radioactivity in the eluted peaks. 

The minimum amount of radioactivity that can be detected by a flow-through radiochemical detector is a subject of continuing debate. It is generally accepted that a fairly sharp peak that contains counts that are at least twice background can be detected. One formula for calculating the minimum detectable activity (MDA) is given by... 

The method is based on the international standard ISO 4053/IV. A small amount of the radioactive tracer is injected instantaneously into the flare gas flow through e.g. a valve, representing the only physical interference with the process. Radiation detectors are mounted outside the pipe and the variation of tracer concentration with time is recorded as the tracer moves with the gas stream and passes by the detectors. A control, supply and data registration unit including PC is used for on site data treatment... 

LC-ARC-MS is a novel radiochemical detection system that is designed to detect samples with very low levels of radioactivity (Lee et al., 2000 Lee, 2003). Similar to the conventional flow through radiochemical detection method, LC-ARC is an in-line detection technique that allows real-time display of metabolite peaks. LC-ARC can either be set up as a stand-alone system or be coupled with a mass spectrometer to become LC-ARC-MS. Thus, the combination of ARC and MS enhances the sensitivity of peak detection and also provides mass spectral information for structural elucidation of metabohte(s). Other interfaces, coupled with the LC-ARC system, are also available for example, LC-ARC/RD-MS/FC is a system of LC-ARC which couples with a radiochemical detector (RD), a mass spectrometer, and fraction collector (FC) (Lu et al., 2002). 

Regardless of cell size, the faster you push a radioactive peak through the cell, the smaller that peak will appear to the radioactivity detector (Figure 5). Thus low level samples cannot be run at high flow rates. With the usual 4.6mm x 25cm HPLC columns, flow rates of 0.5-1.0ml/min. are used routinely with no difficulty. 

Kynurenine was separated from L-3-hydroxykynurenine by chromatography on a Waters /uBondapak Qg column (8 mm X 100 mm, 10-/i,m), using a flow rate of 3 mL/min. The mobile phase was 0.02 M sodium acetate (pH 5.5) containing 2% methanol. Radioactivity was quantitated with a flow-through detector, using scintillation fluid at 3 mL/min. 

Radiolabeled UDP-glucose and sucrose-P were separated on a Selectispher-10 boronate column (5 mm x 250 mm). The mobile phase was 0.12 M sodium phosphate (pH 7.6) delivered at a rate of 1 mL/min. The column eluent was monitored for absorbance at 262 nm, and for radioactivity by a radioactive flow-through detector. 

A radiometric detector, also called a radio-flow detector, is used to measure radioactivity of radioactive analytes in the HPLC eluent passing through a flow cell. Most are based on liquid scintillation technology to detect phosphors caused by the radioactive nuclides. A liquid scintillator can be added post-column with a pump or a permanent solid-state scintillator can be used around the flow cell. This detector is specific only to radioactive compounds and can be extremely sensitive. This detector is used for experiments using tritium or C-14 radiolabeled compounds in toxicological, metabolism, or degradation studies. 

In on-line flow-through detectors, the counting time is the mean residence time of radioactive atoms in the detector and can be increased only by either decreasing the flow-rate or increasing the detector volume. Both will influence the precision and accuracy of the chromatographic separation by influencing the resolution attained. 

FIGURE 9.6 Comparative in vitro profile in liver microsomes and hepatocytes of rat, monkey, and human incubated with [ " CJdasatinib. The profiles are radiochrotogram of HPLC eluant flowing through an online IN/US P-Ram radioactivity detector. 

Detection can be carried out either with an online detector coupled to the eluent flow, or by collection and subsequent analysis of discrete fractions. For collected fractions, a range of analytical methods can be used, both quantitative (e.g., radioactive isotope labeling and metal analysis) and more qualitative (e.g., microscopic techniques). Online detectors suitable for coupling to the FFF channels include both non-destructive flow through cell systems and destructive analysis systems. It is often desirable to use online detection if possible since the total analysis time is much less than for discrete fraction analysis. Regardless of detector type, the dead volumes and flows in the system between the FFF channel and detector or fraction collector must be accurately determined and corrected for. 

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