New sensitive method for HEPES quantification in (68)Ga-radiopharmaceuticals

BACKGROUND: The introduction of a GMP-certified (68)Ga-generator spurred the application of (68)Ga-radiopharmaceuticals. Several radiosynthesis of (68)Ga-radiopharmaceuticals are more efficient and robust when performed with 2-[4-(2-hydroxyethyl)piperazin-1-yl] ethanesulfonic acid (HEPES) buffer, which is considered as an impurity in the quality control (QC) procedure. Thus, prior to clinical use, QC must be conducted to ensure that HEPES does not exceed the maximum dose of 200 μg/V (Injected) as described in European Pharmacopoeia (Ph Eur) for edotreotide. However, when applying the thin-layer chromatography (TLC) method described in the Ph Eur to quantify the HEPES amount present in the (68)Ga-octreotide or in the remaining (68)Ga-radiopharmaceuticals that were tested, no amount was detectable after 4 min of iodine incubation. Here we tested our modified TLC method and validate a new high-performance liquid chromatography (HPLC) method to quantify HEPES in (68)Ga-radiopharmaceuticals and compare it to the TLC-method described in Ph Eur. In addition, samples collected from various institutes were tested to evaluate whether the synthesis of different (68)Ga-radiopharmaceuticals or the use of different synthesis methods could affect the amounts of HEPES. RESULTS: HEPES could not be detected by the TLC method described in the Ph Eur within 4 min incubation in an iodine-saturated chamber. As for our modified TLC method, only after 2 h, spots were only visible > 1 mg/mL. The HPLC method had a limit-of-quantification (LOQ) of 3 μg/mL and a limit-of-detection (LOD) of 1 μg/mL. From the three (68)Ga-radiopharmaceuticals tested, only in the [(68)Ga]Ga-NODAGA-Exendin samples exceeding amounts of HEPES were found and its concentration in the [(68)Ga]Ga-NODAGA-Exendin was significantly higher, when compared to [(68)Ga]Ga-DOTATOC and [(68)Ga]Ga-PSMA-11. CONCLUSION: The TLC method described in Ph Eur and our modified TLC method may not be sufficiently sensitive and thus unsuitable to use for QC release. The new HPLC method was sensitive, quantitative, reproducible and suitable for QC release. With this method, we were able to determine that some (68)Ga-radiopharmaceuticals may exceed the HEPES limit of 200 μg/ V (Injected). This new analytical system would allow correcting for the maximum injected dose in order not to exceed this amount.