Trityl radical ox063

Manufactured by Oxford Instruments

Sourced in United Kingdom, United States

Trityl radical OX063 is a stable free radical commonly used as a polarizing agent in dynamic nuclear polarization (DNP) experiments. It serves as a source of unpaired electrons to enhance the nuclear spin polarization of samples, enabling improved signal-to-noise ratios in nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR) spectroscopy.

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Lab products found in correlation

Hypersense dnp system, by Oxford Instruments (5 mentions) 1 13c pyruvic acid, by Merck Group (4 mentions) 1 13c pyruvic acid, by Cambridge Isotopes (3 mentions) Hypersense, by Oxford Instruments (3 mentions) Dotarem gd chelate, by Guerbet (2 mentions) Hypersense polarizer, by Oxford Instruments (2 mentions) Gadolinium dotarem, by Macrocyclics (2 mentions) Dotarem, by Guerbet (2 mentions) Spinlab hyperpolarizer, by GE Healthcare (1 mentions) 3t mri scanner, by GE Healthcare (1 mentions) Hypersense dnp instrument, by Oxford Instruments (1 mentions) Ethylenediaminetetraacetic acid edta, by Merck Group (1 mentions) Spinlab, by GE Healthcare (1 mentions) Hypersense dnp, by Oxford Instruments (1 mentions) Gadoterate meglumine, by Guerbet (1 mentions) 6 13c arginine, by Cambridge Isotopes (1 mentions) Trizma base, by Merck Group (1 mentions) Hypersense dissolution dnp system, by Oxford Instruments (1 mentions) Gd dota, by Guerbet (1 mentions)

21 protocols using trityl radical ox063

Dynamic Nuclear Polarization of Fructose

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Dynamic nuclear polarization was performed with a 5.0-T SpinLab Hyperpolarizer (GE Healthcare). A 2.0 M solution of 3 dissolved in water containing 15 mM OX063 trityl radical (Oxford Instruments) was polarized in a 5.0-T SpinLab Hyperpolarizer for 3 hours. The hyperpolarized substrate was ejected from the polarizer via dissolution with a large excess of superheated D₂O to a final concentration of 25 mM with a resultant pH of 7.2. Polarization quantification was carried immediately after dissolution. One milliliter of the hyperpolarized dissolution was added to an NMR tube and transferred to a 1-T Spinsolve ¹³C NMR spectrometer (Magritek, NZ). NMR spectra were acquired every 3 s with a 5° excitation over 3 min. Apparent relaxation time (T₁) was fitted to a mono-exponential curve and corrected for flip angle. Thermal polarization was determined from the average spectrum of 1024 scans acquired with 90° flip angle every 10 s. Final polarization values after correcting for flip angles and the apparent T₁ were calculated to be 5 to 10% for all HP fructose dissolutions. Substrate concentration was measured by ¹³C NMR at 14.1 T in the presence of 1 mM Gd-DOTA and 15 mM [¹³C]-acetate standard. The integral of the C₂ carbon of fructose was compared to that of the acetate standard to calculate concentrations.

Tee S.S., Kim N., Cullen Q., Eskandari R., Mamakhanyan A., Srouji R.M., Chirayil R., Jeong S., Shakiba M., Kastenhuber E.R., Chen S., Sigel C., Lowe S.W., Jarnagin W.R., Thompson C.B., Schietinger A, & Keshari K.R. (2022). Ketohexokinase-mediated fructose metabolism is lost in hepatocellular carcinoma and can be leveraged for metabolic imaging. Science Advances, 8(14), eabm7985.

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Cryogenic OX063 Trityl Radical Preparation

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A 20 mM solution of OX063 trityl radical (Oxford Instruments, UK) in neat 1-¹³C pyruvic acid (Sigma Aldrich, US) was prepared. A volume of 80 μL of this solution was pipetted into a bullet that is then immersed in liquid nitrogen to freeze the solution.

Kouřil K., Kouřilová H., Bartram S., Levitt M.H, & Meier B. (2019). Scalable dissolution-dynamic nuclear polarization with rapid transfer of a polarized solid. Nature Communications, 10, 1733.

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Hyperpolarized Pyruvate Metabolite Imaging

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28.5 mg [1-¹³C]pyruvic acid (Cambridge Isotope Laboratories) were polarized to ~30% at 1.42 K and 94 GHz with a HyperSense DNP system (Oxford Instruments). Before insertion into the DNP system, the neat pyruvic acid was mixed with 1.7% by weight OX063 trityl radical (Oxford Instruments) and Dotarem^™ Gd chelate (Guerbet) was added to achieve a total concentration of 15 mmol per mole pyruvate. 4 mL Tris-buffered saline was heated to 190°C at 10 bar, and was used to rapidly dissolve the frozen sample. This solution was diluted using sufficient oxygenated Krebs-Henseleit buffer, as above but without BSA, to yield the desired hyperpolarized pyruvate concentration. The solution was mixed thoroughly and injected using syringe pump in lieu of the steady-state perfusate until the hyperpolarized solution was gone (32 mM) or until 20 ml had been injected (all other hyperpolarized pyruvate concentrations). Note that the dilution buffer was prepared with additional lactate such that when mixed with the hyperpolarized solution the resulting injectate maintained the same lactate concentration as the steady-state perfusion buffer.

Kadlecek S., Shaghaghi H., Siddiqi S., Profka H., Pourfathi M, & Rizi R. (2014). The effect of exogenous substrate concentrations on true and apparent metabolism of hyperpolarized pyruvate in the isolated perfused lung. NMR in biomedicine, 27(12), 1557-1570.

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Hyperpolarized 13C MRI of Diabetic Mice

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The db/db mice and their control littermates db/m mice were fasted for 6 h prior to image acquisition. A tail vein catheter was placed for intravascular access. A 2.2 mol/L solution of [1-¹³C]DHA in dimethyacetamide containing 15 mmol/L OX063 trityl radical (Oxford Instruments) was HP on a HyperSense dynamic nuclear polarization instrument (Oxford Instruments). The frozen sample was dissolved in distilled water containing 0.3 mmol/L EDTA. Imaging was performed using a 3-T MRI scanner (GE Healthcare, Waukesha, WI) equipped with a multinuclear spectroscopy hardware package. The radiofrequency coil used in these experiments was a dual-tuned ¹H-¹³C coil with a quadrature ¹³C and ¹H channels. Prior to ¹³C studies, three-plane T₂-weighted images were acquired for anatomic localization (echo time 100 ms; repetition time 4 s; six averages) using a standard fast spin echo sequence. ¹³C MR spectroscopic imaging studies were then acquired 25 s postinjection of 15 mmol/L HP ¹³C DHA, at 5-mm isotropic resolution, as previously published (21 (link),24 (link)).

Keshari K.R., Wilson D.M., Sai V., Bok R., Jen K.Y., Larson P., Van Criekinge M., Kurhanewicz J, & Wang Z.J. (2014). Noninvasive In Vivo Imaging of Diabetes-Induced Renal Oxidative Stress and Response to Therapy Using Hyperpolarized 13C Dehydroascorbate Magnetic Resonance. Diabetes, 64(2), 344-352.

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Hyperpolarization of 13C1-Pyruvic Acid

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Hyperpolarization of neat ¹³C₁-pyruvic acid (Sigma Isotec) and 15 mM OX-063 Trityl radical (Oxford Instruments) was conducted at 95 GHz with 50 mW microwave powers in a Hypersense polarizer (Oxford Instruments) operating at 1.4 K. In the multi-bolus experiments the solution was stored at ∼1 T in the fringe field; signal lifetimes measured from the decay of the multi-bolus envelope revealed for this in vitro sample a T₁ = 62.89±6.71 s. In vivo hyperpolarized experiments were conducted by injecting 60 mM ¹³C₁-pyruvate neutralized to pH≈7.6, either in single or multi-bolus volumes as indicated in the text.

Leftin A., Roussel T, & Frydman L. (2014). Hyperpolarized Functional Magnetic Resonance of Murine Skeletal Muscle Enabled by Multiple Tracer-Paradigm Synchronizations. PLoS ONE, 9(4), e96399.

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Hyperpolarized [1-13C]Pyruvate Perfusion

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28.5 mg [1-¹³C]pyruvic acid (Cambridge Isotope Laboratories) mixed with 15 mM OX063 trityl radical (Oxford Instruments) and 1.5 mM Dotarem Gd chelate (Guerbet) was polarized to ~30% at 1.42 K and 94 GHz with a HyperSense DNP system (Oxford Instruments). 4 mL of buffer, containing 50mM Tris, 80mM NaOH, and 100 mg/L EDTA, was heated to 190°C at 10 bar, and was used to rapidly dissolve the frozen sample. This sample was further diluted in 6.0 mL oxygenated Krebs-Henseleit buffer (without BSA, which was found to cause unacceptable signal loss during sample transport), yielding a neutral, isotonic solution of 32 mM [1-¹³C] pyruvate. This solution was injected into the perfusate line at 10 mL/min in lieu of the steady-state perfusion buffer. After the 60 seconds required to inject the hyperpolarized solution, lung perfusion was restored.

Shaghaghi H., Kadlecek S., Deshpande C., Siddiqui S., Martinez D., Pourfathi M., Hamedani H., Ishii M., Profka H, & Rizi A.R. (2014). Metabolic spectroscopy of inflammation in a bleomycin-induced lung injury model using hyperpolarized 1-13C pyruvate. NMR in biomedicine, 27(8), 939-947.

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Synthesis and Hyperpolarization of [1-^13C] DHA

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[1-¹³C] DHA (Isotec, Miamisburg, OH) was synthesized using a published method, by air oxidation of [1-¹³C] ascorbic acid in the presence of catalytic amounts of copper (II) acetate38 (link). A 2.2 M solution of [1-¹³C] DHA in dimethyacetamide (DMA) containing 15 mM OX063 trityl radical (Oxford Instruments) was hyperpolarized on a HyperSense DNP instrument (Oxford Instruments) as previously described19 (link). The frozen sample was dissolved in distilled water containing 0.3 mM EDTA.

Wilson D.M., Di Gialleonardo V., Wang Z.J., Carroll V., Von Morze C., Taylor A., Sai V., VanCriekinge M., Bok R., Ohliger M.A, & Keshari K.R. (2017). Hyperpolarized 13C Spectroscopic Evaluation of Oxidative Stress in a Rodent Model of Steatohepatitis. Scientific Reports, 7, 46014.

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Hyperpolarized 13C-Pyruvate Imaging

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[1-¹³C]pyruvic acid (Millipore Sigma, St. Louis, MO, USA) was doped with Ox063 trityl radical (Oxford Instruments, Abingdon, UK) to 15 mM concentration. An amount of 20 µL of this solution was mixed with 0.4 µL of 50 mM Gd³⁺ (Bracco Diagnostics, Monroe Township, NJ, USA). This solution was placed in a DNP HyperSense (Oxford Instruments, Abingdon, UK) to polarize for approximately 1 h under microwave irradiation at 94,100 GHz. An average signal enhancement of 20,000-fold was achieved. Once the sample was prepared, it was rapidly heated and dissolved in 4 mL buffer comprised of 40 mM 2-Amino-2-(hydroxymethyl)-1,3-propanediol (TRIS; Millipore Sigma, St. Louis, MO, USA), 80 mM NaOH, 0.1 g/L ethylenediaminetetraacetic acid (EDTA; Millipore Sigma, St. Louis, MO, USA), and 50 mM NaCl. This solution had a final [1-¹³C]pyruvic acid concentration of 80 mM which was then injected intravenously through the tail vein of the mouse.

Salzillo T.C., Mawoneke V., Weygand J., Shetty A., Gumin J., Zacharias N.M., Gammon S.T., Piwnica-Worms D., Fuller G.N., Logothetis C.J., Lang F.F, & Bhattacharya P.K. (2021). Measuring the Metabolic Evolution of Glioblastoma throughout Tumor Development, Regression, and Recurrence with Hyperpolarized Magnetic Resonance. Cells, 10(10), 2621.

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Hyperpolarized Pyruvate MRI Protocol

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Each sample consisted of 36μL of a mixture of 14M
[1,2-¹³C] Pyruvic acid (Sigma-Aldrich Co., St.
Louis, MO) and 15mM OX063 trityl radical (Oxford Instruments, Abingdon, UK). The
samples were polarized using a dissolution dynamic nuclear polarization (DNP)
system (SpinLab, GE Healthcare, Waukesha, WI) for different lengths of time to
achieve approximately 17%, 22% & 40%
liquid-state polarization at the time of dissolution, as calculated by
comparison to the thermal equilibrium signal from each sample. The polarized
sample was dissolved using 15g solution of 0.1g/L Na₂EDTA, 40mM Tris
buffer, pH neutralized using a NaOH solution, and 4ml of the resulting 80mM
solution of hyperpolarized pyruvate with a pH of 7 was used for dynamic MR
measurements.

Datta K, & Spielman D.M. (2016). Doublet asymmetry for estimating polarization in hyperpolarized 13C-pyruvate studies. NMR in biomedicine, 30(2), 10.1002/nbm.3670.

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Hyperpolarized [1-13C]Pyruvate Production

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Dynamic nuclear polarization (Hypersense, Tubney Woods, Abingdon, Oxfordshire, UK) was performed using 30 μL aliquots of [1-¹³C]pyruvic acid (Cambridge Isotope Laboratories Inc., Tewksbury, MA, USA) doped with 15 mM OX063 trityl radical (Oxford Instruments, Concord, MA, USA). Samples were irradiated with microwaves of 94.075 GHz and 100 mW to solid-state polarizations of over 98%. The sample was then rapidly dissoluted in a 4-mL neutralizing solution of 128 mM NaOH, 140 mM Tris buffer, and 88 mg/L EDTA. A dose of 10 μL/g of hyperpolarized sample was injected into the tail vein over 12–15 s during dynamic image acquisition. Liquid-state polarization of the remaining sample was measured in a bench-top polarimeter (¹³C-MQC polarimeter, Oxford Instruments Molecular Biotools Ltd., Abingdon, Oxfordshire, UK), giving polarizations of 18 ± 3% (mean ± SD) at the time of injection. The final [1-¹³C]pyruvate concentration and pH were ~110 mM and 7.8 ± 0.2 (mean ± SD), respectively.

Macdonald E.B., Begovatz P., Barton G.P., Erickson-Bhatt S., Inman D.R., Cox B.L., Eliceiri K.W., Strigel R.M., Ponik S.M, & Fain S.B. (2021). Hyperpolarized 13C Magnetic Resonance Spectroscopic Imaging of Pyruvate Metabolism in Murine Breast Cancer Models of Different Metastatic Potential. Metabolites, 11(5), 274.

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