Evaluating Quality Control of Radiotracer 18F-FDG: Experience in the United Hospital, Bangladesh

Radionuclide sample is not 100% pure and contains some contaminants which arise from the production process or the decay of the primary radioisotope. Radionuclide impurities increase the radiation dose received by the patient and degrade the quality of any imaging procedure performed. The presence of ¹⁸F radionuclide in sample is determined by the radionuclide purity analysis.

In British pharmacopeia, the presence of ¹⁸F radionuclide can be identified by half-life measurement and the gamma spectrum detection. Though first one is more reliable method, the second one reveals the percentage purity [7]. In quality control laboratory, CRC®-25PET Dose Calibrator (Capintec, USA) [8] is used for half-life measurement and Osprey^™ multi-channel analyzer (MCA) with Thallium-doped sodium iodide NaI (Tl) scintillation detector (Canberra, USA) [9] is used to detect gamma spectrum.

A tiny amount, <1 µl of ¹⁸F-FDG solution with dead time <15% is put in the test tube inside the Scintillation detector. Then the interaction of the emitted positron of ¹⁸F with the atoms in the crystals of the Scintillation detector emits 511 keV gamma photons as shown in figure 1. These Gamma photons are directed to the photomultiplier which converts gamma photons into electrons by photoelectric effect. After that, multiplications of electrons occur in the series of dynode. Then preamplifier produces voltage pulse from these electrons. The Multichannel Analyzer (MCA) reshapes this voltage pulse and converts this electrical signal into a digital signal [10]. The signal sent by MCA is stored, displayed & analyzed by operation computer.

Dose Calibrator is used to check the radionuclide identity of ¹⁸F-FDG. A glass vial containing (<10 µl) ¹⁸F-FDG is placed in the Argon gas filled chamber of Dose Calibrator which consists of anode and cathode those are connected to an external power source. Radioactive decay of ¹⁸F-FDG produces high energy photon. Ion pairs are produced due to the interaction between high energy photons and Argon gas atoms. These ion pairs migrate towards the anode and cathode and produce current. This current is proportional to the radioactivity of ¹⁸F. When isotope selector button is pressed, the result of the division of current and activity conversion factor of ¹⁸F is found which represents the radio activity of ¹⁸F [11]. After that, we take an average value of the observed half-life of Dose calibrator which is as same as the half-life of ¹⁸F.
 

In radiopharmacy, biodistribution of the radiopharmaceutical is determined by radiochemical purity. Radiochemical impurity obscures the diagnostic image obtained and renders the investigation meaningless due to its different patterns of biodistribution.

In British pharmacopeia, both thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) are accurate and reliable method to determine the radiochemical purity and radiochemical identity of ¹⁸F-FDG. Though TLC is easier, it takes longer time [7]. To check the radiochemical purity of ¹⁸F-FDG by TLC method, Peak Simple Chromatography Data System (SRI Instrument, USA) [12] is used in United Hospital quality control lab.

In TLC, according to the monographs, most usually <1 μl of ¹⁸F-FDG is applied with the aid of micropipette on the TLC strip. When the solution dries, the plate is put into 95% Acetonitrile solution. The solvent migrates up the plate (usually 10 cm from the spot) as shown in figure 2. The strip is removed when the solvent reaches the front.

The dried TLC strip is placed into the chromatographic ruler (platform) between two folia strips and insert the ruler into the lead shield of the detector.

A slit-collimated NaI(TI) scintillation radiation detector is used to quantify the regional distribution of radioactivity on the strip. The radioactivity distribution between the start and the solvent front points is recorded and plotted as radioactive peaks by this detector. The potential radiochemical impurities such as free ¹⁸F-FDG and incompletely hydrolyzed intermediate are easily separated from the authentic ¹⁸F-FDG using silica gel thin-layer chromatography.
 

In ¹⁸F-FDG, the mannose triflate precursor is the main chemical contaminant and organic solvent but it is not analyzed in this laboratory because of its non-toxic characteristics. Acetonitrile & Ethanol are also possible chemical contaminants and to ensure the chemical purity of ¹⁸F-FDG the amount of them has been checked. 430-Gas Chromatograph ( Bruker, USA) [13] is used to check the chemical purity of ¹⁸F-FDG.

A very small amount approximately 1µl of ¹⁸F-FDG sample is injected into a hot injector port (which is covered by a rubber septum) of the GC. The temperature of the injector is higher than the components boiling points. Inside the hot injector chamber, the solvent components of ¹⁸F-FDG evaporate into the gas phase. The gaseous components of the ¹⁸F-FDG are pushed by carrier gas helium on to the GC capillary column. Sample passing rate through the column is directly proportional to the temperature of the column. The separation of the organic compounds take place between the carriers gas (the mobile phase) and the high boiling liquid (the stationary phase) within the GC capillary column. Each compound passes through a detector just before exits the GC instrument. The mixture components reach the detector at varying times due to differences in the partitioning between mobile and stationary phases. The signal is sent by this detector to the recorder through amplifier .The result with all peaks is sent into the system computer which stores, displays, and analyzes the data by using the Galaxie^™ Chromatography Data Software System(Bruker, USA) [13].
 

For testing the pH value of ¹⁸F-FDG, British pharmacopeia does not specify any method [7]. pH value of ¹⁸F-FDG can be measured by using pH meter or pH paper with pH reference standards. Narrow band pH paper which is verified by standard pH buffers displays a colour change for each 0.5 pH unit and gives an approximate value of pH. However, pH meter gives specific value [14-19]. In United Hospital quality control laboratory (UHL), properly calibrated SevenMulti pH meter (Mettler- Toledo, Switzerland) and Merck pH paper both are used to check the pH. The physiological range for pH of ¹⁸F-FDG is 4.5-8.5 as per European Pharmacopoeia acceptance criteria and it is consistent from batch to batch.
 

OsprayTM MCA with NaI (TI) scintillation detector [19-22] was used to check the radionuclide purity by gamma spectrum detection. The horizontal position of the peak as shown in figure 3 was determined by the gamma ray energy and the area of the peak was determined by the intensity of the gamma ray and the efficiency of the detector. 511 keV peak of gamma spectrum produced by ¹⁸F was detected in this detector. This single peak represented that ¹⁸F-FDG solution contained >99% ¹⁸F. No other significant amount of radionuclide impurity was present here.

CRC®-25PET Dose Calibrator was used to check the radionuclide identity of ¹⁸F-FDG by half-life measurement because gamma spectrum detection was not enough to confirm the presence of ¹⁸F. As per instruction from the manual supplied by the company the parameters tabulated below were measured [6] (Table 1).

From the observed output of Dose Calibrator as shown in table 2, it was found that the radioactivity decreased gradually when time increased. At the initial the radioactivity of ¹⁸ F was 16.34 mCi and after 10 min it was 15.44 mCi. The average value of the observed half-life (110.26min) was nearly same as the half-life (109.7 min) of ¹⁸F which was shown in table 3.
 

In quality control laboratory, TLC method was used to check the radio chemical purity of ¹⁸F-FDG. The retention factors of the free ¹⁸F-, ¹⁸F-FDG, and unhydrolysed ¹⁸F-FDG were about 2.0 to 2.3, 5.2 to 5.7 and 8.0 to 8.2 respectively. Retention factor (Rf) is the distance traveled by the individual components from the starting point to the solvent front. TLC result is varied according to different brands of TLC plates and operation conditions.

In figure 4, no peak was found in between 2 to 2.3 and 8 to 8.2, so the free ¹⁸F- and unhydrolysed ¹⁸F-FDG concentration were not detectable here. It was approximately found < 0.9% whereas peak in between 5.2 to 5.7 implied the presence of ¹⁸F-FDG. From the above illustration, it was found that more than 99% ¹⁸F-FDG was present in this sample.
 

The chemical purity of ¹⁸F-FDG was checked by Gas Chromatography method in quality control laboratory. The amount of analyte, e.g., Ethanol and Acetonitrile (ACN), present in a sample was determined by the area under the peak of chromatogram.

% component of the analyte had been worked out by the following formula

Area = (height) x (width at ½ height)

%Component 1 = [(area under peak 1)/ (total area)] x 100%

From chromatogram (Figure 5), the area of residual Acetonitrile and Ethanol were found (Table 4).

The calibration curve (Figure 6) of residual solvents, Acetonitrile and Ethanol, showed linear relation of area and concentration. According to the graph, when the area of residual Acetonitrile was 3500 µV.Min, the concentration was 400ppm.

From the observed results of chromatographic spectrum (Table 4), the area of Acetonitrile was found 1¹⁸ µV.Min which correspond very low residual concentration (<70 ppm). Same was seen in case of Ethanol (EtOH), the area of EtOH was found 235.8 µV.Min from the chromatographic spectrum (Table 4), which also correspond very low residual concentration (<50 ppm).

As per European Pharmacopoeia acceptance criteria, residual solvents concentrations in ¹⁸F-FDG were given in table 5. From the above discussion, it was found that the observed residual concentration of Acetonitrile and Ethanol are within the acceptance limit.
 

Digital pH meter and narrow-band pH paper were used to measure the pH of ¹⁸F-FDG. The range of pH was found within 4.5 to 6.5.