Nitrogenases are responsible for biological nitrogen fixation, a crucial step in the biogeochemical nitrogen cycle. These enzymes utilize a two-component protein system and a series of iron–sulfur clusters to perform this reaction, culminating at the FeMco active site (M = Mo, V, Fe), which is capable of binding and reducing N2 to 2NH3. In this review, we summarize how different spectroscopic approaches have shed light on various aspects of these enzymes, including their structure, mechanism, alternative reactivity, and maturation. Synthetic model chemistry and theory have also played significant roles in developing our present understanding of these systems and are discussed in the context of their contributions to interpreting the nature of nitrogenases. Despite years of significant progress, there is still much to be learned from these enzymes through spectroscopic means, and we highlight where further spectroscopic investigations are needed.

[FeFe] hydrogenases are extremely active H₂ converting enzymes. Their mechanism remains highly controversial, in particular, the nature of the one-electron and two-electron reduced intermediates called H_redH⁺ and H_sredH⁺. In one model, the H_redH⁺ and H_sredH⁺ states contain a semi-bridging CO, while in the other model, the bridging CO is replaced by a bridging hydride. Using low temperature infra-red (IR) spectroscopy and nuclear resonance vibrational spectroscopy (NRVS), together with density functional theory (DFT) calculations, we show that the bridging CO is retained in the H_sredH⁺ and H_redH⁺ states in the [FeFe] hydrogenases from Chlamydomonas reihnhardtii (CrHydA1) and Desulfovibrio desulfuricans (DdHydAB), respectively. Furthermore, there is no evidence for a bridging hydride in either state. These results agree with a model of the catalytic cycle in which the H_redH⁺ and H_sredH⁺ states are integral, catalytically competent components. We conclude that proton-coupled electron transfer between the two subclusters is crucial to catalysis and allows these enzymes to operate in a highly efficient and reversible manner.

Herein, we study the mechanism of iron-catalyzed direct synthesis of unprotected aminoethers from olefins by a hydroxyl amine derived reagent using a wide range of analytical and spectroscopic techniques (Mössbauer, Electron Paramagnetic Resonance, Ultra-Violet Visible Spectroscopy, X-ray Absorption, Nuclear Resonance Vibrational Spectroscopy, and resonance Raman) along with high-level quantum chemical calculations. The hydroxyl amine derived triflic acid salt acts as the “oxidant” as well as “amino” group donor. It activates the high-spin Fe(II) (S_t = 2) catalyst [Fe(acac)₂(H₂O)₂] (1) to generate a high-spin (S_t = 5/2) intermediate (Int I), which decays to a second intermediate (Int II) with S_t = 2. The analysis of spectroscopic and computational data leads to the formulation of Int I as [Fe(III)(acac)₂-N-acyloxy] (an alkyl-peroxo-Fe(III) analogue). Furthermore, Int II is formed by N–O bond homolysis. However, it does not generate a high-valent Fe(IV)(NH) species (a Fe(IV)(O) analogue), but instead a high-spin Fe(III) center which is strongly antiferromagnetically coupled (J = −524 cm^–1) to an iminyl radical, [Fe(III)(acac)₂-NH·], giving S_t = 2. Though Fe(NH) complexes as isoelectronic surrogates to Fe(O) functionalities are known, detection of a high-spin Fe(III)-N-acyloxy intermediate (Int I), which undergoes N–O bond cleavage to generate the active iron–nitrogen intermediate (Int II), is unprecedented. Relative to Fe(IV)(O) centers, Int II features a weak elongated Fe–N bond which, together with the unpaired electron density along the Fe–N bond vector, helps to rationalize its propensity for N-transfer reactions onto styrenyl olefins, resulting in the overall formation of aminoethers. This study thus demonstrates the potential of utilizing the iron-coordinated nitrogen-centered radicals as powerful reactive intermediates in catalysis.