Novel methods for the quantification of toxic, residual phase transfer catalyst in fluorine-18 labeled radiotracers.

INTRODUCTION: Fluorine-18 labeled radiopharmaceuticals undergo quality control testing for residual phase-transfer-catalyst content. The almost universally used quality-control test is a silica plate spot-test comparison of the radiopharmaceutical beside a 50-ppm standard. Once developed by staining, the radiopharmaceutical spot must be of equal or less intensity to pass the test. There is currently a need for a quantitative, inexpensive, and less subjective quality control method that allows the automatic incorporation of the acquired measurement directly into electronic batch reports. RESULTS: In the developed method, a resazurin test solution is mixed with an aliquot of the radiopharmaceutical analyte along with dichloromethane (DCM). The mixture is vortexed. The potassium resazurin-phase transfer catalyst complex solubilizes into the DCM imparting a blue color. The organic layer is then removed for analysis. Three measurement methods were utilized: visual colorimetry against pre-prepared standards, spectrophotometric measurement of transmittance, and electrical conductance. A simple prototype spectrophotometer and an electrical test cell were constructed to acquire data. Sodium Resazurin dye was found to be a suitable test chromophore for residual phase transfer catalyst analysis of aqueous solutions. Quantitative spectrophotometric measurements are possible in the 0-100-ppm range (18-crown-6) and 0-150-ppm range (Kryptofix® or tetrabutylammonium). Electrical resistance measurements of the phase transfer-catalyst resazurin complex in DCM are also a viable method, allowing quantitative phase transfer catalyst measurements in the 0-100-ppm range. CONCLUSION: The methodologies developed are more quantitative alternatives to the current spot-test method. The spectrophotometric method was determined to be the most accurate method.

Synthesis of [(18)F]FMISO in a flow-through microfluidic reactor: Development and clinical application.

INTRODUCTION: The PET radiotracer [(18)F]FMISO has been used in the clinic to image hypoxia in tumors. The aim of the present study was to optimize the radiochemical parameters for the preparation of [(18)F]FMISO using a microfluidic reaction system. The main parameters evaluated were (1) precursor concentration, (2) reaction temperature, and (3) flow rate through the microfluidic reactor. Optimized conditions were then applied to the batch production of [(18)F]FMISO for clinical research use. METHODS: For the determination of optimal reaction conditions within a flow-through microreactor synthesizer, 5-400 µL the precursor and dried [(18)F]fluoride solutions in acetonitrile were simultaneously pushed through the temperature-controlled reactor (60-180 °C) with defined flow rates (20-120 µL/min). Radiochemical incorporation yields to form the intermediate species were determined using radio-TLC. Hydrolysis to remove the protecting group was performed following standard vial chemistry to afford [(18)F]FMISO. RESULTS: Optimum reaction parameters for the microfluidic set-up were determined as follows: 4 mg/mL of precursor, 170 °C, and 100 µL/min pump rate per reactant (200 µL/min reaction overall flow rate) to prepare the radiolabeled intermediate. The optimum hydrolysis condition was determined to be 2N HCl for 5 min at 100 °C. Large-scale batch production using the optimized conditions gave the final, ready for human injection [(18)F]FMISO product in 28.4 ± 3.0% radiochemical yield, specific activity of 119 ± 26 GBq/µmol, and >99% radiochemical and chemical purity at the end of synthesis (n = 4). CONCLUSION: By using the NanoTek microfluidic synthesis system, [(18)F]FMISO was successfully prepared with good specific activity and high radiochemical purity for human use. The product generated from large-scale batch production using flow chemistry is currently being used in clinical research.

Synthesis of radioiodinated aryl iodides via boronate precursors.

Arylboronate esters are converted to iodine-123 labeled aryl iodides using no-carrier-added iodine-123 labeled sodium iodide in the presence of chloramine-T. High yields of radiochemically pure products are obtained.

A facile synthesis of radioiodinated (Z)-vinyl iodides via vinylboronates.

A direct radioiodination of (Z)-vinylboronic acid esters to the corresponding vinyl iodides using Na(123)I and chloramine-T is described. The boronates were prepared from vinyl iodides via palladium coupling reactions.

A rapid microfluidic synthesis of [18F]fluoroarenes from nitroarenes.

Microfluidic radiofluorodenitrations have been successfully performed using a commercially available microfluidic synthesis system. Reactions of nitroarenes with para-substituted electron withdrawing groups provide incorporation yields ranging from 43 to 97%. Ortho- and meta-substituted nitroarenes provided incorporation yields up to 35%. The reactions were conducted using dry, no-carrier-added [(18)F]-fluoride and K(2)CO(3)/K(222) dissolved in N,N-dimethylformamide or dimethyl sulfoxide with total synthesis times of less than five min. The methodology developed in these studies can be applied to the synthesis of a variety of fluorine-18 labeled radiotracers and radiolabeled prosthetic groups from nitroarene precursors.

Synergistic effects of leucine and resveratrol on insulin sensitivity and fat metabolism in adipocytes and mice.

BACKGROUND: Sirtuins are important regulators of glucose and fat metabolism, and sirtuin activation has been proposed as a therapeutic target for insulin resistance and diabetes. We have shown leucine to increase mitochondrial biogenesis and fat oxidation via Sirt1 dependent pathways. Resveratrol is a widely recognized activator of Sirt; however, the biologically-effective high concentrations used in cell and animal studies are generally impractical or difficult to achieve in humans. Accordingly, we sought to determine whether leucine would exhibit synergy with low levels of resveratrol on sirtuin-dependent outcomes in adipocytes and in diet-induced obese (DIO) mice. METHODS: 3T3-L1 mouse adipocytes were treated with Leucine (0.5 mM), ß-hydroxy-ß-methyl butyrate (HMB) (5 µM) or Resveratrol (200 nM) alone or in combination. In addition, diet-induced obese mice were treated for 6-weeks with low (2 g/kg diet) or high (10 g/kg diet) dose HMB, Leucine (24 g/kg diet; 200% of normal level) or low (12.5 mg/kg diet) or high (225 mg/kg diet) dose resveratrol, alone or as combination with leucine-resveratrol or HMB-resveratrol. RESULTS: Fatty acid oxidation, AMPK, Sirt1 and Sirt3 activity in 3T3-L1 adipocytes and in muscle cells, were significantly increased by the combinations compared to the individual treatments. Similarly, 6-week feeding of low-dose resveratrol combined with either leucine or its metabolite HMB to DIO mice increased adipose Sirt1 activity, muscle glucose and palmitate uptake (measured via PET/CT), insulin sensitivity (HOMAIR), improved inflammatory stress biomarkers (CRP, IL-6, MCP-1, adiponectin) and reduced adiposity comparable to the effects of high dose resveratrol, while low-dose resveratrol exerted no independent effect. CONCLUSION: These data demonstrate that either leucine or its metabolite HMB may be combined with a low concentration of resveratrol to exert synergistic effects on Sirt1-dependent outcomes; this may result in more practical dosing of resveratrol in the management of obesity, insulin-resistance and diabetes.