3.3.3. Determination of In Vitro Stability and log D7.4 of [18F]**1**

In vitro stability was investigated by incubation of about 5 MBq of [18F]**1** in phosphate-buffered saline (PBS, pH 7.4), *n*-octanol, saline and ethanol (500 μL) at 40 ◦C up to 60 min. Samples were taken at 60 min and the radio-TLC and radio-HPLC analyses were performed.

The lipophilicity of [18F]**1** was experimentally determined by partitioning a small amount of the radiotracer between *n*-octanol and PBS (pH 7.4) at room temperature using the conventional shake-flask method. The radiotracer (15 μL, ~800 kBq) was added to the tubes containing a mixture of *n*-octanol and PBS (3.0 mL, n = 4). After shaking for 20 min using the mechanical shaker (HS250 basic, IKA Labortechnik GmbH & Co. KG, Staufen, Germany), the samples were centrifuged for 5 min at 5000 rpm, followed by separation of the phases. Aliquots were taken from the organic and the aqueous phases (1 mL of each) and activity was measured in a γ-counter (PerkinElmer Wallac Wizard 1480 Gamma Counter, manufactured by WALLAC, Turku, Finland). For the second extraction, another 1 mL aliquot of the organic layer was mixed with 2.0 mL *n*-octanol and 3.0 mL of PBS and was subjected to the same procedure. The distribution coefficient (D) was calculated as [activity (cpm/mL) in *n*-octanol]/[activity (cpm/mL) in PBS, pH 7.4] stated as the decade logarithm (log D7.4).

#### *3.4. Animal Studies*

#### 3.4.1. General Information

All experimental work including animals has been conducted in accordance with the national legislation on the use of animals for research (Tierschutzgesetz (TierSchG), Tierschutz-Versuchstierverordnung (TierSchVersV)) and has been approved by the responsible research ethics committee (TVV 18/18, DD24.1-5131/446/19, Landesdirektion Sachsen, 20 June, 2018). Female CD-1 mice, 10–12 weeks, were obtained from the Medizinisch-ExperimentellesZentrum at Universität Leipzig. For the time of the experiments, the animals were kept in a dedicated climatic chamber with free access to water and food under a 12:12 h dark:light cycle at a constant temperature of 24 ◦C.
