Interaction of Biological Molecules with Clay Minerals: A Combined Spectroscopic and Sorption Study of Lysozyme on Saponite

The interaction of hen egg white lysozyme (HEWL) with Na- and Cs-exchanged saponite was investigated using sorption, structural, and spectroscopic methods as a model system to study clay–protein interactions. HEWL sorption to Na- and Cs-saponite was determined using the bicinchoninic acid (BCA) assay, thermogravimetric analysis, and C and N analysis. For Na-saponite, the TGA and elemental analysis-derived sorption maximum was 600 mg/g corresponding to a surface coverage of 0.85 ng/mm² with HEWL occupying 526 m²/g based on a cross-sectional area of 13.5 nm²/molecule. HEWL sorption on Na-saponite was accompanied by the release of 9.5 Na⁺ ions for every molecule of HEWL sorbed consistent with an ion exchange mechanism between the positively charged HEWL (IEP 11) and the negatively charged saponite surface. The <i>d</i>-spacing of the HEWL–Na-saponite complex increased to a value of 4.4 nm consistent with the crystallographic dimensions of HEWL of 3 × 3 × 4.5 nm. In the case of Cs-saponite, there was no evidence of interlayer sorption; however, sorption of HEWL to the “external” surface of Cs-saponite showed a high affinity isotherm. FTIR and Raman analysis of the amide I region of the HEWL–saponite films prepared from water and D₂O showed little perturbation to the secondary structure of the protein. The overall hydrophilic nature of the HEWL–Na-saponite complex was determined by water vapor sorption measurements. The clay retained its hydrophilic character with a water content of 18% at high humidity corresponding to 240 H₂O molecules per molecule of HEWL

Second-Sphere Effects on Methane Hydroxylation in Cu-Zeolites

Two [Cu₂O]²⁺ cores have been identified as the active sites of low temperature methane hydroxylation in the zeolite Cu-MOR. These cores have similar geometric and electronic structures, yet different reactivity with CH₄: one reacts with a much lower activation enthalpy. In the present study, we couple experimental reactivity and spectroscopy studies to DFT calculations to arrive at structural models of the Cu-MOR active sites. We find that the more reactive core is located in a constricted region of the zeolite lattice. This leads to close van der Waals contact between the substrate and the zeolite lattice in the vicinity of the active site. The resulting enthalpy of substrate adsorption drives the subsequent H atom abstraction stepa manifestation of the “nest” effect seen in hydrocarbon cracking on acid zeolites. This defines a mechanism to tune the reactivity of metal active sites in microporous materials

Identification of α‑Fe in High-Silica Zeolites on the Basis of ab Initio Electronic Structure Calculations

α-Fe is the precursor of the reactive Fe^IVO core responsible for methane oxidation in Fe-containing zeolites. To get more insight into the nature and stability of α-Fe in different zeolites, the binding of Fe­(II) at six-membered-ring cation exchange sites (6MR) in ZSM-5, zeolite beta, and ferrierite was investigated using DFT and multireference ab initio methods (CASSCF/CASPT2). CASPT2 ligand field (LF) excitation energies of all sites were compared with the experimental DR-UV–vis spectra reported by Snyder et al. From this comparison it is concluded that the 16000 cm^–1 band of α-Fe, observed in all three zeolites, can uniquely be assigned to a high-spin square-planar (SP) Fe­(II) located at a 6MR with an Al–Si–Si–Al sequence, where the Al atoms are positioned opposite in the ring and as close to each other as possible. The stability of such conformations is also confirmed by the binding energies obtained from DFT. The bands at 10000 cm^–1 in the experimental spectra, assigned to spectator Fe­(II), are attributed to six-coordinated trigonal-prismatic Fe­(II) species, as calculated for the γ-site in ZSM-5. The entatic effect of the zeolite lattice on the stability of the SP sites was investigated by making use of the unconstrained Fe­(II) model complex FeL₂ (with L = [Al­(OH)₄]⁻). The SP conformer is approximately 2 kcal/mol more stable than the tetrahedral form, indicating that the SP coordination environment of α-Fe is not imposed by the zeolite lattice but rather electronically preferred by Fe­(II) in the environment of four O ligands. A significant contribution to the stability of the SP conformer is provided by mixing of the doubly occupied 3d_<i>z</i>² orbital with the higher lying 4s

Magnetic Exchange Coupling in Zeolite Copper Dimers and Its Contribution to Methane Activation

The highly reactive binuclear [Cu2O]2+ active site in copper zeolites activates the inert C–H bond of methane at low temperatures, offering a potential solution to reduce methane flaring and mitigate atmospheric methane levels. While substantial progress has been made in understanding the activation of methane by this core, one critical aspect, the active site’s spin, has remained undetermined. In this study, we use variable-temperature, variable-field magnetic circular dichroism spectroscopy to define the ground state spin of the [Cu2O]2+ active sites in Cu-CHA and Cu-MFI. This novel approach allows for site-selective determination of the magnetic exchange coupling between the two copper centers of specific [Cu2O]2+ cores in a heterogeneous mixture, circumventing the drawbacks of bulk magnetic techniques. These experimental findings are coupled to density functional theory calculations to elucidate magnetostructural correlations in copper zeolites that are different from those of homogeneous binuclear Cu(II) complexes. The different spin states for the [Cu2O]2+ cores have different reactivities governed by how methane approaches the active site. This introduces a new understanding of zeolite topological control on active site reactivity

Tuning Copper Active Site Composition in Cu-MOR through Co-Cation Modification for Methane Activation

The industrial implementation of a direct methane-to-methanol process would lead to environmental and economic benefits. Copper zeolites successfully execute this reaction at relatively low temperatures, and mordenite zeolites in particular enable high methanol production. When loaded to a Cu/Al ratio of 0.45, mordenite (Si/Al 5–9) has been shown to host three active sites: two [CuOCu]2+ sites labeled MOR1 and MOR2 and a mononuclear [CuOH]+ site. Also at low copper loadings (Cu/Al < 0.20), mordenite has been demonstrated to activate methane, but its active site has never been reported. Here, we investigate Na+ mordenite with varying copper loadings to better understand copper speciation in mordenite. At low copper loadings, we uncover an unidentified active site (“MOR3”) with a strong overlap with the [CuOH]+ site’s spectroscopic signal. By changing the co-cation, we selectively speciate more MOR3 relative to [CuOH]+, allowing its identification as a [CuOCu]2+ site. Active site identification in heterogeneous catalysts is a frequent problem due to signal overlap. By changing cation composition, we introduce an innovative method for simplifying a material to allow better analysis. This has implications for the study of Cu zeolites for methane-to-methanol and NOx catalysis, but also for studying and tuning heterogeneous catalysts in general

Spectroscopic Definition of the Copper Active Sites in Mordenite: Selective Methane Oxidation

Two distinct [Cu–O–Cu]²⁺ sites with methane monooxygenase activity are identified in the zeolite Cu-MOR, emphasizing that this Cu–O–Cu active site geometry, having a ∠Cu–O–Cu ∼140°, is particularly formed and stabilized in zeolite topologies. Whereas in ZSM-5 a similar [Cu–O–Cu]²⁺ active site is located in the intersection of the two 10 membered rings, Cu-MOR provides two distinct local structures, situated in the 8 membered ring windows of the side pockets. Despite their structural similarity, as ascertained by electronic absorption and resonance Raman spectroscopy, the two Cu–O–Cu active sites in Cu-MOR clearly show different kinetic behaviors in selective methane oxidation. This difference in reactivity is too large to be ascribed to subtle differences in the ground states of the Cu–O–Cu sites, indicating the zeolite lattice tunes their reactivity through second-sphere effects. The MOR lattice is therefore functionally analogous to the active site pocket of a metalloenzyme, demonstrating that both the active site and its framework environment contribute to and direct reactivity in transition metal ion-zeolites