Elexsys e500

A Bruker CW-ESR spectrometer (Elexsys E500, Bruker, Karlsruhe, Germany) driven by a PC running the XEpr program and equipped with a Super-X microwave bridge (ER 049X) operating at 9.3–9.5 GHz and a SHQE probe head were used throughout the work. All frozen solution ESR spectra of Cu²⁺ complexes were recorded at 150 K by means of an ER4131VT variable temperature unit. A small amount of methanol (up to 10%) was added to the Cu²⁺-Ctr1 samples in order to increase spectral resolution. Parallel spin Hamiltonian parameters were taken directly from the experimental spectra and were always calculated from the 2nd and 3rd line to get rid of errors coming from second order effects [55 (link)]. The resolution of the ESR spectra [56 (link)] was improved by using isotopically pure ⁶³Cu(NO₃)₂ (0.05 M). In order to achieve a better determination of the magnetic parameters, some of the experimental spectra were simulated by the program Monoclin [57 (link),58 (link)], which is able to discriminate one or more species [57 (link)]. Instrumental settings of frozen solution ESR spectra were as follows: number of scans, 4; microwave frequency, 9.425–9.429 GHz; modulation frequency, 100 kHz; modulation amplitude, 0.7 mT; time constant, 163 ms; sweep time, 2.8 min; microwave power, 20 mW; receiver gain, 60 dB.

Two continuous wave EPR spectrometers were employed: A Bruker Elexsys E500 with Bruker ER4102ST rectangular cavity and a Varian E-9 magnet with E-101 microwave bridge (X-band, ∼9.5 GHz) with super-high Q ER4122SHQE cylindrical cavity. Oxford ESR910/ESR900 Liquid He cryostat operating below 2 and 4 K were used with the two spectrometers, respectively.

Cu(II)-loaded PlAA10 was produced as previously described.^{15 (link)} Protein samples (concentration around 200 μM)
for continuous-wave EPR were prepared in either 50 mM MES buffer (pH
6.5) or 50 mM Tris·H₂SO₄ buffer (pH 8.5).
EPR spectra were recorded on a Bruker Elexsys E500 spectrometer (Bruker,
Karlsruhe, Germany) operating at X-band at 120 K (BVT 3000 digital
temperature controller) with the following acquisition parameters:
modulation frequency, 100 kHz; modulation amplitude, 5 G; conversion
time, 90 ms; sweep time, 92.1 s; and microwave power, 20 mW. EPR spectra
were simulated using the EasySpin toolbox developed for MATLAB.^{27 (link)} The optimum Hamiltonian parameters have been
obtained using the second order perturbation, and then, an exact diagonalization
has been used for the final simulations. Pseudo-modulation treatment
of the spectra was performed to graphically extract the exact number
of nitrogen centers contributing to the superhyperfine pattern.^{28 (link),29 (link)}A_N constants were considered isotropic
and used in the final simulation of the first-derivative spectra.
The Hamiltonian used for the simulations is the following equation with i =
the number of nitrogen
centers.

The morphologies and elemental mapping of series catalysts were observed by scanning electron microscopy (SEM, Zeiss, sigma500, Jena, Germany) with an Oxford Ultim Max Large Area SDD EDS detector. The size of catalysts was measured by a laser particle size analyzer (Malvern, MS-2000, Malvern, UK). The crystallographic structures of catalysts were characterized by a high-resolution transmission electron microscope (HRTEM, JEM-2100, Tokyo, Japan) and X-ray diffraction (XRD, Rigaku Corporation, XRD-6000, Tokyo, Japan). The specific surface area and pore size distribution of samples were analyzed by Brunauer–Emmett–Teller (BET) and Barrett–Joyner–Halenda (BJH) methods, respectively. A Fourier-transform infrared spectrophotometer (FT-IR, American Nicolet Corp. Model 170-SX, Green Bay, WI, USA) was used to investigate the chemical structures of the catalyst. X-ray photoelectron spectroscopy (XPS, Thermo Fisher Scientific, ESCALAB250Xi, Waltham, MA, USA) was utilized to study the surface chemical composition of catalysts. To study the catalytic mechanism of CDM, an electron paramagnetic resonance spectrometer (Bruker ELEXSYS E500, Karlsruhe, Germany) was used to obtain the EPR signals. The leaching of Cu and Mn in wastewater was detected by an inductively coupled plasma spectrometer (Agilent 5800 ICP-OES, Santa Clara, CA, USA).