Chromolith performance rp 18e

The crude product mixtures resulting from direct fluorination reactions were analyzed by liquid chromatography coupled to mass spectrometry (QTrap, Applied Biosystems MIDS SCIEX, Toronto, Canada). The HPLC system used included a Merck Chromolith Performance RP-18e (10 μm, 4.6 × 100 mm) column and an isocratic eluent system consisting of 0.1 % aq. formic acid and CH₃CN (70:30), followed by a 50 % CH₃CN wash. The flow rate was constant at 4 ml/min.

Quantification of the three drugs (ceftazidime, vancomycin and voriconazole) was performed by reversed-phase HPLC (Agilent 1100 Series) using a C18 column (Chromolith^® Performance RP-18e (Merck KGaA, Darmstadt, Germany), 100–3 mm with a linear gradient of 0.1% TFA in water (eluent A) to 0.1% TFA in acetonitrile (eluent B) within 5 min), as previously published for vancomycin and identically applied for ceftazidime and voriconazole [17 (link)]. For all 3 drugs, calibration curves, including the concentration ranges of all tested samples with correlation coefficients of >0.995, were established. All drug solutions were retested after 24 h to guarantee stability of the drugs in the respective solutions until quantification by HPLC. Only the hydrophilic phase was analyzed by HPLC.

Unless otherwise stated all chemicals were purchased from Sigma-Aldrich Chemie (Schnelldorf, Germany) or Merck (Darmstadt, Germany) at analytical grade and were used without further purification. The Ni catalyst (Shimalilte Ni reduced, 80/100 mesh) was purchased from Shimadzu (Kyoto, Japan). The precursor 6-O-desmethyl-elotinib (OSI-420; GMP grade) was purchased from Syncom B.V. (Groningen, Netherlands) and the reference compound erlotinib (N-(3-ethinylphenyl)-6,7-bis(2-methoxyethoxy)-quinazolin-4-amine) was obtained from Apollo Scientific (Bredbury, UK). Semi-preparative high performance liquid chromatography (HPLC) column (Chromolith® SemiPrep RP-18e, 100–10 mm; guard column: 10–10 mm) and analytical HPLC column (Chromolith Performance RP-18e, 100–4.6 mm; guard column: 10–4.6 mm) were purchased from Merck (Darmstadt, Germany). The analytical HPLC column for the optimized quality control (XBridge® Shield RP-18; 2.5 μm; 3.0–50 mm) and C18plus SepPak® cartridges for solid phase extraction (SPE) were purchased from Waters (Waters® Associates Milford, MA, USA). Low-protein binding Millex GS® 0.22 μm sterile filters were purchased from Millipore® (Bedford, MA, USA). Gas chromatography (GC) capillary column (forte GC Capillary Column ID-BP20; 12 m × 0.22 mm × 0.25 μm) was obtained from SGE Analytical Sciences Pty Ltd. (Victoria, Australia).

HPLC-UV analyses were carried out by a Hewlett Packard HP-Ti-1050-Series chromatographic system using a C18 monolithic column (Chromolith Performance RP-18e, 100 × 4.6 mm i.d., Merck KGaA, Darmstadt, Germany). Chromatograms were recorded at 220 nm. Bidistilled water and acetonitrile (AcCN) were used as mobile phase A and B, respectively. A gradient was set as follows: 0–6 min 3% B; 6–15 min from 3 to 7.8% B; 15–21 min from 7.8 to 9% B; 21–27 min from 9 to 13.5% B; 27–37 min from 13.5 to 15.75% B; 37–44 from 15.75 to 18% B; 44–60 min from 18 to 29.1% B; 60–80 min from 29.1 to 47.6% B; 80–82 min from 47.6 to 55% B; flow rate from 0 to 82 min was 1.4 mL/min; 82–83 min 55% B, flow rate increased from 1.4 to 2.0 mL/min; 83–90 min 55% B, flow rate = 2.0 mL/min. The column was re-equilibrated with the starting conditions of analysis for 10 min before the next injection.

F4 was chromatographed on a C18 monolithic column (Chromolith Performance RP-18e, 100 × 4.6 mm i.d., Merck KGaA, Darmstadt, Germany) in order to collect PC3. Chromatographic conditions were as in Section 3.5. Isolation yield = 21.5% on a dry mass basis.

Three Chromolith^® Performance RP-18e (Merck, Darmstadt, Germany) (100 × 4.6 mm) columns were used for chromatographic separations. Chromatograms were recorded in the wavelength range of 220–400 nm. The HPLC conditions were established as follows: Three monolithic columns, a mobile phase consisting of acetonitrile (A), a 15 mM water solution of ammonium acetate adjusted to a pH of 4 with acetic acid (B), and methanol (C). The flow rate was 2 mL/min, and the temperature of the thermostat was 25 °C. The gradient elution program was as follows: From 0 to 20min—15% A, 3% C, and 82% B; from 20.5 to 40.0 min—40% C and 60% B. The identities of the compounds in the plant extracts were confirmed by a comparison of retention times and spectra with corresponding standards.

[⁶⁸Ga]Pentixafor was synthesized using a fully automated Scintomics GRP+ module. The eluate of a ⁶⁸Ge/⁶⁸Ga-generator (GalliaPharm, EZAG, Germany) was concentrated using a Chromafix PS-H⁺ and heated with a solution of 40 µg Pentixafor (unlabeled precursor; GMP quality) in 1.5 mL of HEPES buffer (N-(2-hydroxyethyl)piperazine-N´-(2-ethanesulfonic acid), 1.5 M at 125°C for six minutes. Unbound and colloidal gallium-68 was removed using a SepPak Light C18 cartridge (Waters). The purified product was formulated with PBS buffer. The chemical and radiochemical purity of [⁶⁸Ga]Pentixafor were analyzed by high performance liquid chromatography (HPLC) using an analytic Chromolith® Performance, RP-18e, 100-4.6 mm from Merck (Germany). Free ⁶⁸Ga³⁺-ions and ⁶⁸Ga-colloid were detected using radio-thin-layer chromatography (radio-TLC).
The glucose analogue [¹⁸F]FDG was synthesized using FASTlab FDG cassettes with a phosphate buffer formulation and a GE FASTlab platform (GE Healthcare).