Why is molar activity important? Is high molar activity always the goal?
There are different situations that will determine if high Am (or As) is a critical factor or not for the study. Three categories can be identified:
High Am is critical when studying:
· bioactive or toxic molecules, as the quantity of injected non-radioactive compound can produce undesired pharmacodynamic or toxicological effects.
· low density receptors. The injection of an excess of the non-radioactive compound can have an effect on the receptor occupancy and, therefore, alter the apparent receptor characteristics. The receptor occupancy with the non-radioactive compound should be kept below 5% in order to avoid pharmacological or pharmacodynamic effects.
· kinetic modelling parameters, as low Am can lead to an unfavourable signal-to-noise ratio within the image. Am can have substantial impact on the quantitative analysis and qualitative interpretation of nuclear medicine images.
High Am is not critical when studying:
· endogenous compounds normally found in high concentrations in the body (e.g., [15O]water, [11C]acetate, [11C]glucose)
· processes in which the biological target or function is not easily saturated (e.g. glucose metabolism, hypoxia, enzyme activity). Nevertheless, the Am can dramatically impact the image outcome, e.g. when the patient eats candies directly before measuring the glucose metabolism.
Intermediate to low Am is necessary when studying:
· antibodies, due to liver depletion which reduces the biological half-time of small amounts of antibodies in the blood stream.
· some peptides or proteins, to optimize the biodistribution properties in terms of reducing radiation burden in specific organs (mostly emunctories) by low organ-to-tumour ratios.
Table 1. Examples of radiotracers falling into the three categories of Am.
Low Am | Intermediate Am | High Am |
[18O]H2O | [11C]PIB | [11C]PHNO |
[11C]Methionine | [11C]DASB | |
[18F]FDOPA | [11C]mHED | |
[68Ga]PSMA | | |
What is the maximum theoretical molar activity?
Theoretical Am values for PET isotopes are very high (e.g. 341.1 TBq/µmol for carbon-11, 63.3 TBq/µmol for fluorine-18) however these values are very far from those obtained in laboratories (e.g. 10-5000 GBq/µmol for carbon-11, 10-1000 GBq/µmol for fluorine-18).
Table 2. Half-life and maximum theoretical Am of some PET radionuclides.
Radionuclide | Half-life (minutes) | Maximum theoretical Am (TBq/µmol) | Examples of radiotracers produced with high Am (TBq/µmol) |
11C | 20.4 | 341.1 | 4.9 ± 2.4 ([11C]Raclopride) at end of synthesis (EOS)1 |
18F | 109.8 | 63.3 | 4.4 ([18F]DCFPyL) at EOS2 |
15O | 2.0 | 3394.0 | |
13N | 10.0 | 699.3 | |
68Ga | 68.0 | 102.3 | |
Low Am might be due to a dilution process with the non-radioactive isotope.
Am is decreased by:
(i) the generation of the radioactive precursor in the cyclotron. Impurities potentially come from the target gas, the valves, pressure regulators, seals, target body, target windows or even residues from cleaning solvents.
(ii) the transfer of the irradiated gas/liquid to the hot cells. Long-time processes decrease Am. Keeping the lines pressurized with high purity gases and avoiding fluorine-rich materials during the preparation of 18F radiotracer or carbon-rich materials for 11C-radiotracer should help to maintain Am.
(iii) the synthesis process. The use of high purity reagents and precursors is the most important factor. Reagents should be prepared and stored under adequate inert conditions to prevent contamination. The synthesis unit should be also cleaned and dried thoroughly and kept isolated from atmosphere/contaminants;
Am can be increased either by:
(i) increasing beam time and/or current (up to a certain level), loading/unloading the target before irradiation, discarding the first 1-3 irradiations of the day and keeping the target under pressure between runs.
(ii) decreasing the amount of non-radioactive sources that might potentially contaminate the reaction. Contaminants from the atmosphere, or undesired chemicals released from tubes, etc. or the presence of impurities in reagents could contribute to decrease of Am.
How to calculate the molar activity.
The methods for measuring the Am will depend on the physical state of the radioactive material. Gaseous and liquid materials can often be analysed with the use of radiogas chromatography, high pressure liquid chromatography (HPLC), ion chromatography or mass spectrometry.
Here we give an example of a PROCEDURE to calculate the Am for a 11C-radiotracer using HPLC.
What you need: HPLC, analytical column, pipettes, falcon tubes, 12C-reference compound, PET dose calibrator, a solution of a 11C-radiotracer, laptop, solvents.
1) Develop a HPLC method to identify the reference compound using an analytical HPLC column and define the:
· UV wavelength (measured at the maximal UV absorption);
· mobile phase;
· column temperature;
· flow rate;
· injection volume (e.g. 10 µL).
2) From a stock solution of the 12C-reference compound (e.g. 1 µM), prepare seven standard solutions in a non-volatile solvent (e.g. 200 nM, 100 nM, 50 nM, 25 nM, 12.25 nM, 6.13 nM, 3.06 nM). From the concentration of the standard solutions calculate the amount (µmol) present in the HPLC injected volume (e.g. 10 µL). Inject the standard solutions into the HPLC resulting in chromatograms with a single peak.
3) From the UV detector chromatogram, determine the area under the curve (AUC) of the peak corresponding to the compound. (Figure 1A). Enter these values and amount of reference compound (µmol) into a computer spreadsheet. Create a calibration curve by plotting the AUCs as a function of injected amount of reference compound (in µmol) (Figure 1B). Analyse the data by applying a linear regression analysis (linear least squares fit). This yields the equation y = mx + y0, where y is the area, m represents the slope, and y0 is a constant that describes the background. The amount of compound (x expressed in µmol) present in a radioactive sample may be calculated from this equation.