Abstract
The objective of this research is to determine whether there are significant differences in the 18F-FDG produced by either the phosphate or the citrate buffer cassettes in the FASTlab synthesizer. Methods: Forty batches of 18F-FDG were produced with each cassette and analyzed retrospectively. The analysis consisted of determining the mean radiochemical yield (RCY)—uncorrected and corrected for decay—radiochemical purity (RCP), pH, and residual solvent content (ethanol and acetonitrile). An independent t test (alpha error [α], 0.05) was performed to determine whether the differences were statistically significant. Results: The mean decay-corrected RCYs for 18F-FDG produced by phosphate and citrate cassettes were 82.9% ± 17.4% and 79.2% ± 5.0%, respectively. The uncorrected RCY was 57.5% ± 16.7% for phosphate- and 58.8% ± 6.0% for citrate-buffered 18F-FDG, leading to a difference of 4.4% and P value of 0.11 for corrected RCY and a difference of 2.2% and P value of 0.32 for uncorrected RCY. Thus, the RCY differences are neither statistically nor clinically significant. The mean RCPs were 99.4% ± 0.2% for the phosphate-buffered 18F-FDG and 99.0% ± 1.1% for the citrate-buffered 18F-FDG. There was a 0.5% difference and a P value of 0.021, meaning that the difference was statistically significant. The average pHs for 18F-FDG produced by phosphate and citrate buffer cassettes were 5.9 ± 0.1 and 5.3 ± 0.2, respectively, resulting in a 9.6% difference and a P value close to zero (2.6 × 10−19)—a statistically significant difference. The difference between ethanol content was also dramatic. Phosphate-buffered 18F-FDG contained 0.08% ± 0.02% ethanol, whereas the citrate-buffered 18F-FDG contained 0.20% ± 0.07%. No difference was found in the acetonitrile content of the 2 cassettes. Conclusion: The differences in yield between cassettes are due to statistical variability. The results confirm our hypothesis that there is no significant difference in RCY. The differences seen in the statistically significant data (those with a P value > 0.05) turn out to be insignificant in a real-world setting because all values fell within the limits set by the United States Pharmacopeia and Food and Drug Administration. Therefore, determining which cassette to use is a matter of the preference of the institution.
For determining metabolism in the brain and detecting various tumors, 18F-FDG is a commonly used radiopharmaceutical in PET imaging. 18F-fluoride is produced in a cyclotron by the following reaction: 18O(p,n)18F. The 18F-fluoride is then substituted onto a deoxyglucose analog by a multistep reaction. The reaction, from introduction of 18F-fluoride to end of synthesis (EOS), can vary depending on the type of PET synthesizer used (1). However, GE Healthcare’s FASTlab synthesizer using an 18F-FDG cassette decreases the reaction time dramatically to around 22–25 min, depending on the cassette used (2,3).
The FASTlab synthesizer allows for single-use cassettes to be inserted into the unit. The cassettes contain all the necessary chemicals for synthesis of bulk 18F-FDG. GE Healthcare manufactures 2 cassette types: one in which 18F-FDG is diluted in 15 mL of phosphate buffer and another in which 18F-FDG is diluted in 22–38 mL, depending on the volume desired, of sodium citrate buffer (2,3). The Mayo Clinic PET Radiochemistry Facility in Rochester, Minnesota, uses 29 mL for the final volume of 18F-FDG produced with the citrate cassette. Each cassette is bar-coded, allowing the radiopharmaceutical being produced to be traced. The synthesis is controlled and monitored with vendor-supplied custom software, which automates the process, reducing exposure to the user while also minimizing or eliminating any variability caused by the operator. It is the objective of this study to determine whether any significant differences exist between 18F-FDG produced by phosphate and by citrate buffer cassettes and to make users of the FASTlab system aware of these differences.
MATERIALS AND METHODS
18F-FDG Production
Phosphate Buffer Cassette.
The cyclotron was used to irradiate 18O-enriched water with 16.5-MeV protons, producing 18F-fluoride. The 18F-fluoride was then transferred into the FASTlab synthesis unit. There, the water containing 18F-fluoride was passed through an anion exchange column, to which it became bound. A solution composed of aminopolyether (Kryptofix, K222; GE Healthcare) and potassium carbonate in acetonitrile and water was then passed through the exchange column and eluted the 18F-fluoride, which was collected in a reaction vessel. Approximately 1.7 mL of acetonitrile was added to the reaction chamber, and the solvents were evaporated by heating to 125°C. The reaction vessel contents were dried using nitrogen and the application of a vacuum for approximately 9 min. Mannose triflate, dissolved in acetonitrile, was then added to the reaction chamber while being heated for approximately 2 min, allowing the nucleophilic substitution of the triflate group for 18F-fluoride, forming 18F-fluoro-tetra-acetyl-glucose (18F-FTAG). The contents of the reaction chamber were then mixed with water and passed through a tC18 cartridge (Sep-Pak; Waters), which retained the 18F-FTAG. The unbound materials were washed through the cartridge into a waste container. Sodium hydroxide was then passed through the cartridge to elute and hydrolyze the 18F-FTAG. The elution was mixed with water and the pH adjusted using phosphate buffer. The buffered solution was then passed through tC18 and alumina cartridges (Sep-Pak; Waters) to remove any remaining impurities. Finally, the synthesized 18F-FDG was passed through a 0.22-μm filter and collected in a sterile vial. The synthesis time was around 22 min (3). The quantity of 18F-FDG was determined by measuring the bulk quantity with a dose calibrator (CRC15-PET; Capintec). This measurement was deemed to be EOS. The radiochemical yield (RCY) was determined by the fraction of activity present at EOS divided by the activity entering the synthesis unit after the end of bombardment (EOB). In essence, (activity at EOS/activity at EOB) × 100 = uncorrected RCY percentage. RCY can be corrected for the decay during synthesis with the application of the decay equation.
Citrate Buffer Cassette.
For the citrate buffer cassette, the same procedure was followed as for the phosphate buffer cassette; however, 18F-FDG was diluted in sodium citrate and pushed through reversed-phase and alumina cartridges (Sep-Pak; Waters). The synthesis time was generally 25 min (2).
Quality Control of 18F-FDG
In compliance with the guidelines of the Food and Drug Administration (FDA) for PET drug current good manufacturing practice and the United States Pharmacopeia (USP) monograph, quality control (QC) procedures were performed on newly synthesized 18F-FDG to ensure that it not only meets safety and purity requirements but also has the correct identity, strength, and quality, (4,5).
Radiochemical Purity (RCP).
RCP was determined using radio–thin-layer chromatography (TLC) in compliance with the USP monograph for 18F-FDG (4). QC samples were analyzed using aluminum-backed silica gel plates (Whatman) in acetonitrile and water 90:10. The plates were then analyzed with a radio-TLC chromatograph (AR-2000; Bioscan). The acetonitrile causes 18F-FDG to travel along the TLC plate, whereas free 18F-fluoride remains at the origin. The radiochromatogram typically contains 2 peaks: one at origin (18F-fluoride) and one at an Rf of approximately 0.6 (18F-FDG). By comparing the areas under the radiopeaks, the RCP of 18F-FDG can be calculated. The current USP monograph allows up to 10% 18F-fluoride (4,5).
pH.
The pH was measured using indicator strips with a range of 4.0 to 7.0 ± 0.2. The acceptable range for pH, as stated in the USP monograph, is between 4.5 and 7.5 (4).
Residual Solvent Content.
Residual solvent content was determined using gas chromatography (model 8610C; SRI Instruments) in compliance with the USP monograph (4). A 0.5-μL sample was pushed through the gas chromatography column (inside diameter, 30 m × 0.53; MXT Crossbond Carbowax [Restek]) at 10 mL/min using helium gas. The temperature was initially 40°C for 2 min and then increased 20°C per minute up to 130°C. The gas chromatograph allowed for determination of the amount of ethanol and acetonitrile in the final 18F-FDG product. The maximum allowable content of ethanol and acetonitrile as stated by guidelines of the USP and FDA are 0.5% and 0.04%, respectively (4,5).
Data Analysis
Data for 40 batches of 18F-FDG for both the phosphate and citrate buffer cassettes were analyzed retrospectively. The analysis consisted of determining the mean RCY (uncorrected and corrected for decay), RCP, appearance, RSC, and pH. An independent t test (alpha error [α], 0.05) was performed to determine whether any differences were statistically significant.
RESULTS
The results for RCY, RCP, pH, and RSC are shown in Table 1.
Relationship Between RCY, RCP, RSC, and pH of Each Cassette
RCY
Forty batch productions for each FASTlab cassette type were analyzed retrospectively. The mean RCY was calculated to determine the radiochemical efficiency for each cassette. Of the 40 batches of phosphate-buffered 18F-FDG, the mean activity at EOS was 133 GBq (3.6 Ci), ranging from 59 to 263 GBq (1.6–7.1 Ci). The mean activity of the citrate-buffered 18F-FDG at EOS was 133 GBq (3.6 Ci), with a range of 41–244 GBq (1.1–6.6 Ci). There was a 4.4% difference and a P value of 0.11 for the decay-corrected yield and a 2.2% difference and a P value of 0.32 for the uncorrected yield. Because α was set at 0.05, these differences were determined to be statistically insignificant and caused by normal statistical variance.
RCP
Because the P value fell below the α-value (0.05), it can be said that the difference in RCP is statistically significant. However, the difference was clinically insignificant because both cassette means fell well within the limit of at least 90% set by the USP monograph for 18F-FDG (4).
pH
A 9.6% difference in pH existed between the cassettes, with a P value of nearly 2.6 × 10−19. Thus, the pH of the final solution was dependent on which cassette was used in the synthesis of 18F-FDG, and the difference was statistically significant. Again, however, statistical significance did not lead to clinical significance. The pH values of all 80 batches fell within the 4.5–7.5 range stated in the USP monograph for 18F-FDG (4).
RSC
A 35% difference existed in ethanol content between the cassettes, with a statistically significant P value. There was no recorded difference in acetonitrile content between the cassettes. The difference in ethanol content is insignificant because the value falls within the 0.5% allowance stated in the USP monograph (4).
DISCUSSION
The Mayo Clinic PET Radiochemistry Facility switched from producing 18F-FDG with the phosphate buffer cassette to the citrate cassette in December 2011. The change was because of the revised abbreviated new drug application (ANDA) requirements for 18F-FDG set forth by the FDA. The guidance states that the reference listed drug of 18F-FDG is diluted to a concentration of 4.5 mg/mL of sodium chloride and 7.2 mg/mL of citrate in water for injection (6). Thus, the citrate buffer cassette is used to allow PET radiochemistry facilities to submit ANDAs with the same composition as the reference listed drug in the FDA guidance. Hence, one can have peace of mind in switching from phosphate buffer cassettes to citrate cassettes in the FASTlab synthesis unit.
Because USP and FDA requirements for QC procedures for 18F-FDG did not change with the new ANDA requirements, it was the objective of the researchers to ensure that the new citrate cassette produced high-quality batches of 18F-FDG.
The differences seen between the batches produced with the 2 cassettes, as stated previously, are clinically insignificant. The large difference seen in ethanol content can be attributed to GE Healthcare’s striving to produce a more stable end product, because ethanol is a known radiolysis inhibitor at high amounts of radioactivity (7,8). This inhibition may contribute to the large SD seen in the yield of the phosphate-buffered 18F-FDG, because the phosphate buffer cassette has a lower ethanol content and higher radioactive concentration (Table 2). Thus, the citrate cassette has a higher likelihood of better stability at high radioactive concentrations, which are present in PET radiochemistry facilities.
Radioactive Concentration of Each Cassette
Not all QC parameters required by the USP and FDA were analyzed in this research. These QC parameters were selected for evaluation because they are the most likely to be affected by the type of cassette used in the production of 18F-FDG.
CONCLUSION
The RCY, RCP, pH, and RSC of 80 batches of 18F-FDG, produced in GE Healthcare’s FASTlab synthesis unit using the phosphate buffer cassette or citrate buffer cassette, were analyzed and compared to determine whether any significant differences existed between the 2 cassettes. Some statistically significant differences did exist in RCP, pH, and ethanol content. The increased ethanol content of the citrate cassette supports a more stable end product at high concentrations of radioactivity, because of the ability of ethanol to inhibit radiolysis. However, all values for both cassettes fell within acceptable limits set forth by the USP monograph and FDA. Therefore, the biggest differences found were that the citrate buffer is stated explicitly in the reference listed drug composition of 18F-FDG in the FDA guidance on new drug application and ANDA submissions and has a higher likelihood of stability at high radioactive concentrations. Thus, using the citrate buffer cassette to produce 18F-FDG ensures peace of mind for the PET radiochemistry facility.
DISCLOSURE
No potential conflict of interest relevant to this article was reported.
Acknowledgments
The data from this research were presented at the Society of Nuclear Medicine annual meeting held in Miami Beach, Florida, June 9–13, 2012.
Footnotes
Published online Jan. 14, 2013.
- Received for publication August 12, 2012.
- Accepted for publication December 4, 2012.
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