Epr finger dewar

Resonance Raman (rR) samples were prepared by addition of Cu(II) and Cu(I) to apoprotein. The rR samples were excited at 840 nm using continuous wave excitation from a Lighthouse Photonics Sprout-G pumped SolsTiS-PX Ti: Saph laser with a power of ~100 mW at the sample. The sample was immersed in a liquid nitrogen cooled (77 K) EPR finger dewar (Wilmad). Data were recorded while rotating the sample manually to minimize photodecomposition. The spectra were recorded using a Spex 1877 CP triple monochromator with a 1200 grooves/mm holographic grating and detected by an Andor iDus CCD cooled to −80 °C. Spectra were energy calibrated with citric acid. The protein spectra were normalized to the ice peak at ~230 cm⁻¹. The rR data was processed using SpectraGryph version 1.2 (Dr. Friedrich Menges Software-Entwicklung, Oberstdorf, Germany) and Origin 2018b software.

All reagents and solvents purchased and used were of commercially available quality except as noted. MeTHF and THF were distilled over Na/benzophenone under Ar. The ligands ^XTMPA (X = −H, −NH₂, −NH₂,CH₃, −CH₃, and −(CH₃)₂) and complexes [(^XTMPA)Cu^I]BAr^F (BAr^F = B(C₆F₅)4⁻), (F₈)⁵⁶Fe^II, (F₈)⁵⁷Fe^II, and d₈-F₈Fe^II were synthesized as previously described.^{107 (link),108 ,123 (link),124 (link),138 (link)–142 (link)} Preparation and handling of air-sensitive complexes were performed in a Vac atmosphere OMNI-LAB drybox or under argon atmosphere using standard Schlenk techniques. Solvent deoxygenation was achieved by bubbling Ar through the desired solvent for ≥45 min via an addition funnel connected to a receiving Schlenk flask. UV–vis measurements were carried out by using a Carey-50 Bio spectrophotometer with a 10 mm path quartz cell. The spectrometer was equipped with Cary WinUV Scanning Kinetics software and a Unisoku thermostat cell holder for low-temperature experiments. All NMR spectra were recorded in 9 in., 5 mm o.d. NMR tubes on Bruker 300 NMR instrument equipped with a tunable deuterium probe to enhance deuterium detection. The ²H chemical shifts are calibrated to natural abundance deuterium solvent peaks. Resonance Raman samples were excited at a variety of wavelengths (413 nm excitation unless otherwise noted with data), using either a Coherent 190C-K Kr⁺ ion laser, a Coherent 25/7 Sabre Ar⁺ ion laser, or an Ar⁺ pumped Coherent Ti:Saph laser while the sample was immersed in a liquid nitrogen cooled (77 K) EPR finger Dewar (Wilmad). For data collection at temperatures between −110 °C and −90 °C, the finger dewar was filled with various solvent/N₂ cooling baths, using a thermocouple thermometer to monitor the temperature at sample point in situ. Power was ~2 mW at the sample for the high-energy lines and >200 mW at the low-energy lines. Data were recorded while rotating the sample to minimize photodecomposition. The spectra were recorded using a Spex 1877 CP triple monochromator with either a 600, 1200, or 2400 grooves/mm holographic spectrograph grating and detected by an Andor Newton CCD cooled to −80 °C (for high-energy excitation) or an Andor IDus CCD cooled to −80 °C (for the low-energy excitation). Spectra were calibrated on the energy axis to toluene. Excitation profiles were intensity calibrated to the solvent (MeTHF) by peak fitting in the program Origin. EPR spectra were collected with an ER 073 magnet equipped with a Bruker ER041 X-Band microwave bridge and a Bruker EMX 081 power supply: microwave frequency = 9.42 GHz, microwave power = 0.201 mW, attenuation = 30 dB, modulation amplitude = 10 G, modulation frequency = 100 kHz, temperature = 10 K.

[(P^ImH)Fe^III−(O₂^•−)] (2). In the glovebox, 2 mM solutions of an (P^ImH)Fe^II in THF were prepared and transferred to rR tubes and capped with tight-fitting septa. The sample tubes were placed in a cold bath (dry ice/acetone) and oxygenated using ¹⁶O₂ or ¹⁸O₂ gases. The oxygenated samples were set in a cold bath for 10 min, after which the sample tubes were frozen in liquid N₂ and sealed by flame. rR samples were excited at 413 nm, using either a Coherent I90C–K Kr⁺ ion laser as the sample was immersed in a liquid-nitrogen-cooled (77 K) EPR finger Dewar (Wilmad). Power was ~2 mW at the sample, which was continuously rotated to minimize photodecomposition. The spectra were recorded using a Spex 1877 CP triple monochromator, and detected by an Andor Newton CCD cooled to −80 °C. rR [Fe^III−(O₂^•−)] (2): ν_O−O, 1171 cm⁻¹ (Δ¹⁸O₂, −61 cm⁻¹); ν_Fe−O, 575 cm⁻¹ (Δ¹⁸O₂, −24 cm⁻¹).
[(P^ImH)Fe^IICu^I]⁺(3), [(P^ImH)Fe^III−(O₂²⁻)−Cu^II]⁺(3a), and [(DCHIm)(P^ImH)Fe^III−(O₂²⁻)−Cu^II]⁺(3b). In the glovebox, 5 mM solutions of an equimolar mixture of (P^ImH)Fe^II and [Cu^I(CH₃CN)₄](B(C₆F₅)₄) in THF were prepared and transferred to rR tubes and capped with tight-fitting septa. The sample tubes were placed in a cold bath (dry ice/acetone) and oxygenated using ¹⁶O₂ and ¹⁸O₂. The labeled gases were cooled in dry ice for 5 min and injected through the solution by using a Hamilton gastight syringe. The oxygenated samples were set in a cold bath for 10 min, after which the sample tubes were frozen in liquid N₂ and sealed by flame. To the cold THF solution (at −80 °C, acetone–dry-ice bath) of 3a was added 1.5 equiv of DCHIm (3.5 mg of DCHIm in 0.5 mL of THF) for complex 3b. rR [(P^ImH)Fe^III−(O₂²⁻)−Cu^II]⁺ (3a): ν_O−O, 799 cm⁻¹ (Δ¹⁸O₂, −48 cm⁻¹); ν_Fe−O, 524 cm⁻¹ (Δ¹⁸O₂, −23 cm⁻¹). [(DCHIm)(P^ImH)Fe^III−(O₂²⁻)−Cu^II]+ (3b): ν_O−O, 817 cm⁻¹ (Δ¹⁸O₂, −55 cm⁻¹); ν_Fe−O, 610 cm⁻¹ (Δ¹⁸O₂, −26 cm⁻¹).

All reagents and solvents were of commercially available quality except as noted. Acetone was distilled from Drierite under argon atmosphere. [(MeAN)Cu^I](BAr^F) (BAr^F: B(C₆F₅)₄⁻) was synthesized as previously described.^{24 (link)} Sodium phenolates were obtained using a synthetic method similar to previous results.^{23 (link)} All UV–vis measurements were carried out using a Hewlett-Packard 8453 diode array spectrophotometer with a 10 mm quartz cell. The spectrometer was equipped with HP Chemstation software and a Unisoku cryostat for low temperature experiments. ¹H NMR spectra were recorded on a Bruker 400 instrument. Resonance Raman (rR) samples were excited using a Coherent I90C–K Kr⁺ ion laser at 413.1 or 568.2 nm while the sample was immersed in a liquid-nitrogen-cooled (77 K) EPR finger dewar (Wilmad). Power was ~20 mW at the sample for the 413.1 nm line and ~130 mW at 568.2 nm. Data were recorded while rotating the sample to minimize photodecomposition. The spectra were recorded using a Spex 1877 CP triple monochromator with 600, 1200, or 2400 grooves/mm holographic spectrograph grating and detected by an Andor Newton CCD cooled to −80 °C (413.1 nm) or an Andor IDus CCD cooled to −80 °C (568.2 nm). Spectra were calibrated on the energy axis to toluene at room temperature.

Samples were excited at a variety of wavelengths, using either a Coherent I90C-K Kr⁺ion laser, Liconix HeCd laser, a Coherent 25/7 Sabre Ar⁺ ion laser, or a Lighthouse Photonics Sprout-D pumped M-Squared SolsTiS Ti:Saph laser while the sample was immersed in a liquid nitrogen cooled (77 K) EPR finger dewar (Wilmad), or cooled at 110 K through contacted with the liquid nitrogen cooled sample holder. The laser power (<2 mW) and sample spinning conditions were set such that the RR spectra obtain before and after extended data acquisition showed no detectable differences. Resonance Raman sample preparation: In a typical experiment, 0.650 mL of a complex (P^Im)Fe^II (1) (2 mM) solution in THF was placed in a 9 mm economy rubber septum capped NMR tube. After cooling the NMR tube to −80 °C (acetone/N_2(liq) bath), O_2(g) was bubbled through the solution mixture to form the complex [(P^Im)Fe^III(O₂^•−)] (2). Similar to the procedure used for UV–vis experiments, complex [(P^Im)Fe^III(⁻OON═O)] (3) was prepared by removing excess of dioxygen by vacuum/Ar cycles from 2 and careful addition of NO_(g) was carried out using three-way gastight syringe. Complex [(P^Im)Fe^III](NO₃⁻)] (4b) was prepared by thermal decay of complex 3. After generation of all complexes, tubes were frozen in N_2(liq) and flame-sealed. Isotopically labeled (¹⁸O₂/¹⁵N¹⁸O) samples were prepared in analogous manner.

All reagents and solvents used were of commercially available quality and used without further purification except as noted. Inhibitor-free 2-MeTHF was distilled over Na/benzophenone under Ar and deoxygenated with Ar before use. [Cu^I(CH₃CN)₄(BAr^F)] and F₈Fe^II were synthesized as previously described.^{81 (link),82 (link)} All UV–vis measurements were carried out using a Hewlett-Packard 8453 diode Figure 8. DFT-optimized structures of LS-4DCHIm and LS-3DCHIm (porphyrin ArF groups and H-atoms omitted for clarity) depicting the relationship between structure and reactivity toward different acids. array spectrophotometer with a quartz Schlenk cuvette cell to monitor changes in Q-band and with a 0.2 cm path length cuvette to monitor changes in Soret band. The spectrometer was equipped with HP Chemstation software and a Unisoku thermostated cell holder for lowtemperature experiments. Resonance RAMAN samples were excited at a variety of wavelengths, using either a Coherent I90C-K Kr⁺ ion laser, a Coherent 25/7 Sabre Ar⁺ ion laser, or a Lighthouse Photonics Sprout-G pumped SolsTiS-PX Ti:Saph laser, while the sample was immersed in a liquid nitrogen cooled (77 K) EPR finger dewar (Wilmad). Power was ~2 mW at the sample for 413 nm excitation and 150–200 mW for lower energy excitation. Data were recorded while rotating the sample to minimize photodecomposition. The spectra were recorded using a Spex 1877 CP triple monochromator with either a 600, 1200, or 2400 grooves/mm holographic spectrograph grating, and detected by an Andor Newton CCD cooled to −80 °C (for high-energy excitation) or an Andor IDus CCD cooled to −80 °C (for the low-energy excitation). Spectra were calibrated on the energy axis to toluene. Excitation profiles were intensity calibrated to the solvent (MeTHF) by peak fitting in the program Origin.

Samples were prepared by exposure of a 1 mM solution of [(F₈)Fe(II)] and [(AN)Cu(I)]B(C₆F₅)₄ in THF to dioxygen at subambient temperatures (−80°C) in 5 mm rubber septa capped NMR tubes. DCHIm (1eq) was added and the tube was flame sealed. Isotopically labelled (¹⁸O₂) samples were prepared with ¹⁸O₂ in an analogous manner. Samples were excited at a variety of wavelengths, using either a Coherent I90C-K Kr+ ion laser, a Coherent 25/7 Sabre Ar+ ion laser, or an Ar+ pumped Coherent Ti:Saph laser while the sample was immersed in a liquid nitrogen cooled (77 K) EPR finger dewar (Wilmad). The power was ~2 mW at the sample for the high energy lines and >200 mW at the low energy lines. Data were recorded while rotating the sample to minimize photodecomposition. The spectra were recorded using a Spex 1877 CP triple monochromator with either a 600, 1200, or 2400 groves/mm holographic spectrograph grating, and detected by an Andor Newton CCD cooled to −80° C (for high energy excitation) or an Andor IDus CCD cooled to −80°C (for the low energy excitation). Spectra were calibrated on the energy axis to toluene. Excitation profiles were intensity calibrated to the solvent (THF) by peak fitting in the program Origin.