Background <p>¹⁸F-Labeled (<i>E</i>)-1-fluoro-3-((2-(4-(5-(methylamino)pyrazin-2-yl)but-1-en-3-yn-1-yl)benzo[<i>d</i>]thiazol-6-yl)oxy)propan-2-ol ([¹⁸F]SPAL-T-06) and (<i>E</i>)-1-fluoro-3-((2-(4-(6-(methylamino)pyridine-3-yl)but-1-en-3-yn-1-yl)benzo[d]thiazol-6-yl)oxy)propan-2-ol ([¹⁸F]C05-05) are positron emission tomography (PET) imaging agents for α-synuclein pathologies. Recently, we demonstrated the utility of PET imaging with [¹⁸F]SPAL-T-06 for α-synuclein pathologies in patients with multiple system atrophy and with [¹⁸F]C05-05 for α-synuclein pathologies in Parkinson’s disease and related disorders. In this study, to establish [¹⁸F]SPAL-T-06 and [¹⁸F]C05-05 as PET radiopharmaceuticals for clinical investigations, we developed routine radiosynthetic procedures using an ¹⁸F-labeling synthesizer and performed quality control (QC) testing of these two PET radiopharmaceuticals.</p> Results <p>[¹⁸F]SPAL-T-06 and [¹⁸F]C05-05 were synthesized by direct ¹⁸F-fluorination of their respective tetrahydropyranyl-protected tosylated precursors, followed by deprotection, using an ¹⁸F-labeling synthesizer. The non-decay-corrected radiochemical yields of [¹⁸F]SPAL-T-06 and [¹⁸F]C05-05 from ¹⁸F⁻ at the end of synthesis (EOS) were 11 ± 4.6% (<i>n</i> = 19) and 20 ± 6.8% (<i>n</i> = 37), respectively. The radiochemical purities of both PET radiopharmaceuticals exceeded 99%. The molar activities of each PET radiopharmaceutical at EOS exceeded 68&#xa0;GBq/µmol, and the total synthesis times were within 100&#xa0;min after irradiation. In addition, [¹⁸F]SPAL-T-06 and [¹⁸F]C05-05 were efficiently separated from byproducts using a semi-preparative high-performance liquid chromatography system. All QC results for the two PET radiopharmaceuticals complied with in-house QC and quality assurance specifications. The stabilities of the two PET radiopharmaceuticals were evaluated under standard fluorescent light in a general laboratory environment and under UV-cutoff light in a natural light-shielded environment. After 10&#xa0;min of exposure to the standard fluorescent light, the radiochemical purity of each tracer decreased to approximately 86%. In contrast, the radiochemical purity of each tracer exceeded 99% after 60&#xa0;min under UV-cutoff light in a natural light-shielded environment.</p> Conclusions <p>We successfully prepared two PET radiopharmaceuticals, [¹⁸F]SPAL-T-06 and [¹⁸F]C05-05, for clinical investigation. The radiosynthesis and QC findings provide a practical basis for establishing standardized production procedures and facilitating technology transfer to other PET centers. In particularly, all post-deprotection radiosynthesis steps and QC testing for [¹⁸F]SPAL-T-06 and [¹⁸F]C05-05 should be performed under UV-cutoff light in a natural light-shielded environment to prevent photoisomerization.</p>