The temperature of the 2D electrons was inferred from the electronic noise in a gold wire, connected to the 2DEG by a metallic ohmic contact. With effective screening and filtering, we demonstrate a temperature of 0.9 ± 0.1 mK, with scope for significant further improvement. This platform is a key technological step, paving the way to observing new quantum phenomena, and developing new generations of nanoelectronic devices exploiting correlated electron states.Two-dimensional electron gases (2DEGs) with high mobility, engineered in semiconductor heterostructures host a variety of ordered phases arising from strong correlations, which emerge at sufficiently low temperatures. The 2DEG can be further controlled by surface gates to create quasi-one dimensional systems, with potential spintronic applications. Here we address the long-standing challenge of cooling such electrons to below 1 mK, potentially important for identification of topological phases and spin correlated states. The 2DEG device was immersed in liquid  Cooling electrons into the microkelvin temperature range is of interest both for practical purposes and fundamental studies, but current demonstrations are limited to small, specific devices. Here, the authors achieve sub-millikelvin temperatures in a large-area, two-dimensional electron gas. This requires the high crystal perfection of MBE growth and the use of modulation doping 3 has led to extensive studies of two-dimensional electron systems in perpendicular magnetic field B. The lowest Landau level (LL), with filling factors ν < 1  for the lowest spin branch is described by composite fermion (CF) theory4. It exhibits a rich variety of quantum states: fractional quantum Hall states for odd fractions, while ν = 1/2 is a composite fermion Fermi sea5. A key question is the role of residual CF interactions in giving rise to exotic states6. A Wigner solid is found for small ν, which also exhibits re-entrance with a maximum melting temperature of 50 mK above the ν = 1/3 FQHE state7. Interactions are also believed to play a role at other fractional fillings such as ν = 4/11, which has a measured activation energy of 5 mK8.The lowest possible electron carrier temperature coupled to the highest sample quality is key to discovering and elucidating new ground states in such systems. The observation of the integer quantum Hall effect9 required electron temperatures as low as 8 mK10. Nevertheless the topology of the ν = 5/2 ground state  remains an open question. Observations are highly sensitive to disorder12. It has been shown that a quantum phase transition to a quantum Hall nematic phase can be induced by applied pressure in both the 5/2 and 7/2 state13. The importance of such topological ground states is reinforced by the proposal that they offer a route to topological quantum computing using non-Abelian anyons14. A non-Abelian phase is also predicted for the 12/5 state.The nature of quantum states in the second LL is in many cases not resolved, and this 2D electron system is of particular interest for study to lower temperatures than hitherto achieved. We discuss a few examples here. Even denominator states forming in the second Landau level are thought to arise from pairing of composite fermions. Confirmation of exact quantization of the 5/2 state first discovered by Willett et al.16 found that the gaps of the ν = 2 + 6/13 (10 mK) and 2+2/5 (80 mK) states do not follow the CF hierarchy, leading to the proposal of a parton state17. The 2+3/8 state also has a reported gap of only 10 mK. Furthermore, there are indications of novel topological order in the upper spin branch of the second LL, at fillings 3+1/3 (37 mK) and 3+1/5 (104 mK)18.Many of these states have small energy gaps. The 12/5 state has a gap of order 30 mK, which exhibits non-trivial tilt dependence of the magnetic field. Studies of the energy gaps at other fractional fillings20 and the Kondo effect22. Furthermore the conduction electrons in GaAs-based 2DEGs are expected to couple to the nuclear spins, undergoing a collective ferromagnetic transition mediated by RKKY interactions which is expected at electron temperatures in the mK regime23. The hyperfine interaction is also predicted24 to create new topological phases in a semiconductor 1D wire coupled to a superconductor and magnetic moments. A transition from metal- to insulator-like behaviour with decreasing temperature has been reported in a 2D hole gas in GaAs25, potentially related to many-body localization. Recently, Wigner crystal-like zigzag chains have been discovered in 1D wires at low electron densities26. At lower temperatures there is potential for quantum communication through 1D spin chains, coupling high fidelity quantum dot qubits27.New states arising from strong correlations in the low-dimensional electron system are also expected in low magnetic fields. Electrostatic confinement of the 2DEG to form 1D wires and zero-dimensional (0D) quantum dots has led to studies of the many-body physics of Luttinger liquids28, with prospects of commercial solutions in the microkelvin range30. However, cooling electrons in semiconductor devices is challenging. In situ cooling via the electron–phonon coupling is severely limited by its strong temperature dependence, while the electrons are susceptible to heat leaks arising from electromagnetic noise. Electromagnetic shielding and ultra-sensitive measurement schemes with low or no electrical excitation33 are common strategies for minimising the heat leak. Cooling via the electrical leads has been widely implemented36, commonly employing immersion in liquid helium. For 2D electrons, this has so far been limited, for many years, to 4 mK, see ref. 34 and references therein. Primary thermometers within samples confirmed 2D electron temperatures of 6 mK36.Advances in cryogen-free technologies now make low millikelvin temperature platforms widely accessible37. More recently this has been extended by incorporating nuclear refrigeration elements into the device itself. For this on-chip cooling the Coulomb blockade device used is itself a thermometer, providing an accurate measurement of the electron temperature in mesoscopic metallic islands as low as 0.5 mK39.In response to the challenge to cool nano-electronic devices to ultra-low temperatures (ULT), new approaches have been introduced which focus on refrigeration by nuclear demagnetisation of each lead connected to the sample2) 2DEG. Our approach to cooling is motivated by the requirement for flexibility to cool a wide range of devices in different sample environments. We report an ultimate electron temperature in the 2DEG of 0.9 ± 0.1 mK, cooled in a 3He immersion cell, achieved after improvements in electromagnetic shielding. The 2DEG device is immersed in liquid 3He and cooled through metallic ohmic contacts, which are coupled to the 3He via compact heat exchangers made of sintered Ag powder. Cooling of devices by the immersion cell technique, as compared to on-chip cooling, is remarkably versatile due to modularity of the design. The cooling is provided by a physically remote copper nuclear adiabatic demagnetization stage module, so the magnetic field applied to the sample can be independently adjusted. A broad class of devices can potentially be cooled in this way, and in some cases the 3He liquid pressure can be used as an experimental control parameter40.In contrast, and complementary to this approach, the focus of our work is the cooling of a relatively large area  of the 2DEG and ohmic contacts, and reducing the heat leak to the device. We fabricated a high-mobility GaAs-based 2DEG with sub-1 Ω AuNiGe ohmic contacts, see Fig. 41. Due to the fundamental Nyquist relation the NT operates over 5 orders of magnitude in temperature with a single point calibration. It can be configured as a primary thermometer42, and as such underpins the recent redefinition of the Kelvin.A key feature of the experiment was the choice of thermometer. We use a current sensing noise thermometer (NT) as an external module attached to the 2DEG via an ohmic contact. A SQUID current sensor is used to read out the voltage fluctuations across a gold wire3He in which it is immersed. We have constructed a thermal model which demonstrates that at the lowest temperatures the NT accurately reflects the electron temperature in the 2DEG. The ability to alter the resistance of the 2DEG in situ by optical illumination, due to persistent photoconductivity, was used to experimentally validate the model.The noise thermometer dissipates no power in the device, but we must take account of both the inevitable residual heat leak into the thermometer, and the potential parallel cooling channel by direct coupling to the 3He bath with a photon-tight environment. Filtered connections to room-temperature electronics are provided to probe electrical resistances in the device and to operate heaters in thermal transport experiments. Two NTs are connected to the 2DEG and the demagnetisation refrigerator. Illumination by a red light-emitting diode (LED) facing the 2DEG allows us to increase the 2D carrier density and mobility43. Preliminary studies were conducted in a plastic cell shown in Fig. 34. See Figure 44 to be a superconducting transition in the ohmic contacts.Figure Te to the 2DEG NT temperature TNT. We consider two sources of heat: The use of an external thermometer in our setup requires an investigation of the heat flow in the system in order to relate the electron temperature in the 2DEG 3He bath, as inferred from measurements of TNT as a function of 3He bath temperature Tbath. Figure GΣ = Ge + GHe, Ge via electrons in the 2DEG and ohmic contacts, and the Kapitza boundary conductance GHe between the NT assembly and the 3He bath45. The illumination of the 2DEG increases Ge, allowing us to separate it from GHe. To reduce GHe the heat exchangers in the immersion cell were plated with approx. 30 μmol/m2 4He coverage before loading 3He46.Figure T → 0 limit52. The WF law predicts that RNT to IV = 27 Ω (6 Ω) is the electrical resistance from the NT ohmic contact to I± and V± connected in parallel, measured before (after) illumination, where 47. Figure 53 in the ohmic contacts44. To the leading order the change in GΣ due to illumination is ΔGΣ = ΔGe ∝ T; to explain this observation we propose a model of partially-superconducting ohmic contacts, see Fig. Tc, Ge = L0T/Re is described by a resistance Re ≥ RNT to IV. The data are well described byRe = 59 Ω (27 Ω) before (after) illumination and illumination-independent GHe = K2T2 + K3T3. Similarly the response to X4, but with different values of α before and after illumination. The X4T4 term describes an additional cooling mechanism in the 2DEG or ohmic contacts, insignificant below TNT = 3 mK, where X4T4 ≪ GΣ(T). In this low temperature regime, Eq.  law, ubiquitous in electronic transport in the ment see , allowinX4T4 term in Eq.  for Tph = 0.3 mK and We demonstrate that at 1 mK the dominant cooling of the 2D electrons is by the ohmic contacts, which have a thermal conductance of X4T4 term in Eq. , GNT(T) and GIV(T), see Fig. G ∝ T behaviour, if the effective contact resistance Te for the preliminary experiments shown in Fig. X4T4 term, a more detailed thermal model, beyond the scope of this work, should take into account the distribution of The m in Eq.  points tsee Fig.  is not cRe ≫ RNT to IV. Therefore the thermal conductance through the I± and V± contacts in parallel, GIV = Ge/α, /α = 2.1 times higher than that through the NT ohmic, GNT = Ge/(1 − α), despite being 15 times larger in circumference. This suggests that the large I± ohmics were rendered thermally inactive due to low thermal conductance along them, as the gold wires were bonded 0.5 mm away from their front edges, Fig. α)/α = 3.2 before illumination is consistent with the I± ohmics being more active when the 2DEG resistance is higher.We note that after illumination the thermal resistances are dominated by the ohmic contacts, since see Fig. , is onlyA natural question to ask is whether even lower electron temperatures can be achieved. First, the heat leak may be reduced by optimising the device geometry  embedded in Ag epoxy was inserted between 2DEG NT and its SQUID sensor. This design was chosen for having zero DC resistance and for inducing sufficiently low amounts of noise in the SQUID so as to not mask the NT signal. This filter was mounted directly on top of the immersion cell and sealed to the cell wall with In to maintain a photon-tight enclosure. Another multi-line Ag epoxy filter at 1.5 K was inserted between the SQUID sensor and its room temperature electronics. All remaining filters were mounted at the mixing chamber plate of the dilution refrigerator.60 and were calibrated against a primary magnetic field fluctuation thermometer at 10–100 mK61.Both NTs were read out with integrated 2-stage SQUID current sensorswere performed using a mains-powered Stanford Research Systems SR-124 analogue lock-in amplifier using its internal oscillator. Below 20 mK measurement currents between 1 and 10 nA were used to ensure sub-fW Joule heating.TNT  = Ta,b at different levels of Tbath as GN(T). Below 3 mK the 2DEG NT responded slowly, Fig. Tbath could not be kept constant sufficiently long due to the single-shot operation of the demagnetisation cooling, so Ta was inferred from measurements before and after Tb, see Fig. dTNT/dt = 0, see Fig. TNT > 1.2 mK.The thermal conductance TNT was too high to ignore the X4T4 term in GN. Here the thermal model given by Eqs. (4He plating GN(T) was measured separately in each experiment, see In the early experiments shown in Fig.  by Eqs.  and 4)  TNT was Supplementary InformationPeer Review File


Chryseobacterium polytrichastri uncovered its diterpene synthase activity, yielding polytrichastrene A and polytrichastrol A with new skeletons, besides known wanju‐2,5‐diene and thunbergol. The enzyme mechanism was investigated by isotopic labeling experiments and DFT calculations to explain an unusual ethyl group formation. Rationally designed exchanges of active site residues showed major functional switches, resulting for I66F in the production of five more new compounds, including polytrichastrene B and polytrichastrol B, while A87T, A192V and the double exchange A87T, A192V gave a product shift towards wanju‐2,5‐diene.A reinvestigation of the linalool synthase from  Chryseobacterium polytrichastri was found to have diterpene synthase activity and produced diterpenes with new skeletons containing a rare ethyl group. Its formation was mechanistically studied through isotopic labeling and DFT calculations. Exchange of active site residues resulted in a functional switch leading to additional novel diterpenes. This work lays the ground for future rational engineering of terpene synthases.Linalool synthase from  S. clavuligerus are rare.S. mobaraensis is the only known bacterial type I sesterterpene synthase (StTPS).Terpenoid biosynthesis starts with the cyclization of an acyclic oligoprenyl diphosphate into a structurally complex terpene hydrocarbon, which can be followed by several functionalizations to introduce bioactivity. The cyclization step is catalyzed by a single enzyme, a terpene synthase (TPS), and proceeds by substrate ionization through abstraction of diphosphate (type I TPSs) or protonation (type II) to initiate a cationic cascade reaction.S. pristinaespiralis (SdS) in complex with the substrate surrogate 2,3‐dihydrofarnesyl diphosphate (DHFPP).While the overall structures of type I TPSs are highly conserved and exhibit a similar α‐helical fold as avian farnesyl diphosphate synthase,VvdW in Å3) can be calculated according to Abraham and co‐workers (Table S1).VvdW (ΣVvdw) is given in Table S2. The spanned regions and mean ± standard deviation for each group of TPSs are summarized in Figure Vvdw ranging from 907 Å3 for MTPSs to 733 Å3 for StTPSs, with an average decrease of 58 Å3 between two consecutive TPS groups. This value is similar to the increase of the substrate size of 84 Å3 for each isoprene unit (Table S1), suggesting that the active site space becomes larger according to the requirements of the substrate. We have recently described a TPS from Chryseobacterium polytrichastri as linalool synthase (CpLS), based on the observed activities towards different substrates in a mixed Tris/phosphate buffer.Vvdw=550 Å3, which is the smallest value among all TPSs (red entry in Table S2) and did not seem to fit for a MTPS, but would rather suggest a function as DTPS or StTPS.Several non‐polar active site residues enclose DHFPP in the SdS structure that define the substrate conformation and the available space within the active site Figure . Their v1) and polytrichastrol A (2), both featuring novel skeletons with an ethyl group (Scheme 3), known from the C. wanjuense wanju‐2,5‐diene synthase (CwWS),4)Chryseobacterium polytrichastriPolytrichastrene Synthase (CpPS). The absolute configurations of 1 and 2 were determined by stereoselective deuteration using dimethylallyl diphosphate (DMAPP) and (R)‐ or (S)‐isopentenyl diphosphate (IPP)Streptomyces cyaneofuscatusE)‐ and (Z)‐IPP,1 and 2.A reinvestigation of this enzyme indeed resulted under optimized conditions with a solely Tris‐based incubation buffer in the efficient conversion of GGPP into several unknown diterpenes, while GFPP did not yield any products and GPP and FPP gave only acyclic terpenes . The main hydrocarbon and a minor alcohol were isolated and their structures elucidated by NMR spectroscopy , resulting in polytrichastrene A ‐GLPP by syn‐allylic transposition of diphosphate with a specific stereochemical fate for the 1‐pro‐R and the 1‐pro‐S hydrogen of GGPP, ending up in the 1E and 1Z positions of GLPP, respectively. After vinyl group rotation around C2‐C3 the 1,14‐cyclization continues with attack of C14 at C1 from the Si face  to install the correct configuration at C14 in A, which also turns H1S to the bottom. This hydrogen is now closer to the cation at C15 than H1R and shifts into the iPr group in B, as confirmed experimentally with (R)‐ and (S)‐(1‐2H)GGPP . For the enantiomers of 1–4 the migration of H1R into the iPr group would be expected, and thus the migration of H1S versus H1R can give evidence for the absolute configurations of terpenes, as demonstrated previously for other sesqui‐ and diterpenes.[4 from CpPS  discloses that this is the same enantiomer as in Pseudotsuga menziesii .The cyclization mechanism by CpPS requires GGPP isomerization to GLPP to explain the 2B is a first branching point and can be attacked by water to yield 4, while the steps from B to F1 towards 3 (shown in blue) were supported by previous labeling experiments.C1 to D1 and from E1 to F1 occur that were both supported for 1 with GGPP and GGPP, enzymatically prepared from GPP13C,2‐2H)FPP13C‐NMR spectrum . The steps from F1 towards 1 may include a dyotropic rearrangementH (orange arrow) to explain the Et group formation. Stereoselective deuterations introduced at C12 using DMAPP plus (E)‐ or (Z)‐IPP and GGPPS revealed the specific shift of the 12‐pro‐S hydrogen, while the usage of IPPJ. Deprotonation to 1 with cyclopropanation proceeds with loss of the 5‐pro‐R hydrogen, as demonstrated by stereoselective deuterium labeling introduced with (R)‐ and (S)‐IPP . Another WMR from J results in K, the precursor of 2 that is formed by quenching with water. The overall mechanism towards 1–4 is also in line with 13C‐labeling experiments, in which each of the twenty carbons of GGPP were labeled individually .Cation 1, but no 2 and slightly increased 3 and 4. In addition, several new compounds were obtained. A preparative scale incubation allowed the isolation and structure elucidation by NMR spectroscopy  of wanju‐2,6‐diene (5), wanju‐2‐en‐6α‐ol (6), polytrichastrol B (7), bonn‐2‐en‐11α‐ol (8) and polytrichastrene B (9). Their absolute configurations were determined by analogous stereoselective deuterium labeling experiments as described above for 1–4, giving uniform results for all nine compounds .For a deeper understanding of the role of active site residues in TPS catalysis , several CpPS variants targeting positions homologous to SdS active site residues were constructed (Tables S7 and S8). The purified enzyme variants were adjusted to same concentration , followed by conversion of GGPP and GC/MS analysis of the products . The position equivalent to I66 in CpPS . A 1,4‐hydride shift from F2 to M, 2,14‐cyclization and deprotonation can lead to 9. Notably, F2 can also be linked to F1 through WMR (purple), so that the cyclization cascade along B, the red branch to F2, the purple link to F1 and the downstream reactions to L contain all intermediates in one direct line to explain all products by simple deprotonation or attack of water (only 9 requires a short side branch via M and N). The labeling experiments performed here cannot distinguish between the blue and the red pathway, because both pathways contain analogous elementary steps, so that all substrate atoms end up in the same positions.Alcohol A to B proceeds smoothly, ending in a B conformer that allows both cyclization reactions to C1 and C2 through similar transition state (TS) barriers (Scheme C1 to F1 (blue) are all associated with low TS barriers or even barrierless, while the step from C2 to D2 has a fairly high barrier . The conversions from D2 to F2 are more feasible, and also the purple link between F1 and F2 can be passed in both directions. This analysis suggests that the blue pathway may be preferred for the formation of 1–7, and also 9 can be reached via this pathway and the purple link to F2, as the 1,4‐hydride transfer to M can be realized computationally. Ring closure of M to N was only possible with assistance of a base (MeNH2) in contact with Me20 to suppress spontaneous ring opening of N to M, ultimately leading to 9. An alternative two‐step process with 1,3‐ and 1,2‐hydride transfer is geometrically not possible, similar to 1,3‐hydride transfers in guaiane biosynthesis that are rare and only allowed for the few stereoisomers with suitable geometry.8 may be the only case for which the red path is entered from B, because the step from F2 to E2 has an even higher TS than the C2 to D2 conversion. The high TS barriers to reach E2 from any side may explain the observation of 8 only as a minor compound in the I66F variant.To distinguish between these pathways, DFT calculations were performed . The 1,3‐hydride shift from F1 to H dyotropic rearrangement discussed above seems not possible , but assistance by a catalytic base, for which MeNH2 was selected as a model , allows a stepwise process with low barriers through deprotonation of H12S and fragmentation to G followed by back transfer of the same proton to C11 and ring closure to H.Based on the calculations the concerted 3, while A87F and A91F were inactive, presumably because of steric interference with GGPP. The M308W exchange did not yield a soluble protein, suggesting a major impact on the enzyme structure.The SdS positions I75, L78, L79 and F297 are also often occupied by aromatic residues in DTPSs. In the corresponding positions of CpPS small  or non‐polar (Met) residues are found that were exchanged to obtain the A87F, G90F, A91F, and M308W variants. G90F showed a decreased activity (23±2 %) with main product 3 of CpPS is the main product of CwWS, and these enzymes differ in their active site residues in only two positions: A87 and A192 in CpPS are taken by T86 and V191 in CwWS (Table S2). The importance of active site residues to control the product profile of a TPS is impressively demonstrated by the A87T, A192V and—with two simultaneous exchanges—the A87T,A192V enzyme variants, all of which gave 3 as main product. The A192V variant showed an increased activity (176±8 %), but also the single A87T and the double A87T,A192V exchange did not disturb substrate acceptance .Side product In summary, we have characterized a bacterial DTPS that converts GGPP into diterpenes with novel skeletons. An unusual ethyl group formation was studied in detail by isotopic labeling experiments and DFT calculations, revealing a two‐step rather than a concerted dyotropic rearrangement likely assisted by an active site base that needs to be identified by future structural work ideally in conjunction with QM/MM simulations. We have also identified 13 active site residues that are promising candidates for functional switches within TPSs, which allows for a rational design of engineered biocatalysts in future work.The authors declare no conflict of interest.As a service to our authors and readers, this journal provides supporting information supplied by the authors. Such materials are peer reviewed and may be re‐organized for online delivery, but are not copy‐edited or typeset. Technical support issues arising from supporting information (other than missing files) should be addressed to the authors.Supporting InformationClick here for additional data file.


For both low and high relative Ca concentrations,the Ca ions are found to be octahedrally coordinated by the NH3 solvent, and these hexammine units are spatially correlatedout to lengthscales of ∼7.4–10.3 Å depending onthe concentration, leading to pronounced ordering in the bulk liquid.We further demonstrate that this liquid order can be progressivelydisrupted by the substitution of Ca for Na, whereby a distortion ofthe average ion primary solvation occurs and the intermediate rangeion–ion correlations are disrupted.Thelocal and intermediate range ordering in Ca–NH Unusually, this immiscibility region isextremely pronounced in the Ca–NH3 phase diagram,with the electrolytic state only surviving to a maximum Ca concentrationof 1.68 mole percent metal (MPM) at the consolute point or a criticaltemperature of 290 K with the onset of phase separation at <0.1MPM at 200 K.4 For comparison, the Na–NH3 consolute point is 4.12 MPM and 231 K with phase separationbeginning at 1 MPM at 220 K, and in Li–NH3, theconsolute point is 4.35 MPM at 210 K with phase separation beginningat 2 MPM at 220 K.5 Such early onset andpronounced liquid–liquid phase separation clearly indicatethe stability of, or energetic drive toward, the metallic liquid inCa–NH3 solutions.The dissolution of electropositive groupI and group II metalsin liquid NH3 solvent can all be varied. Some metal–ammoniasystems, including Li–NH3 and Ca–NH3, have been shown to possess strong intermediate range ordering between∼6 and 10.5 Å in their metallic phases, whereas othersystems such as Na–NH3 and K–NH3 do not.9 Interestingly, only those systems which exhibit such strong orderingin their liquid state are known to crystallize as solid phases attheir concentration limit. All other metal–ammonia or metal–aminesystems precipitate out the metal upon cooling and do not form solidmetal–amine phases. Taking only the examples of the Ca–NH3 and Na–NH3 systems, it is notable that,despite their similar ionic-radii,10 Ca–NH3 possesses distinct intermediate range ordering in the liquidand crystallizes as the expanded metal Ca(NH3)6,11 whereas no such liquid order nor any crystallinephases have been witnessed in Na–NH3.8Previous studies have soughtto understand the link between thedifferent electronic states of metal–ammonia systems and theirliquid structure, whereby the charge of the metal cation, the extentof screening by the itinerant electron, and the secondary solvationeffects from the NH12 to provide a more detailed study of the Ca–NH3 system in the metallic phase than has been performed previously.Both the local and intermediate solvation environments of the Ca ionsare determined alongside Ca–Ca correlations in the liquid phase.We also demonstrate that the intermediate range order of the Ca–NH3 solution can be successively disrupted by the introductionof Na.Herein, we use neutron scattering withcombined H/D isotopic substitutionsand empirical-potential structure refinements–5 mbar. Appropriate ratios of isotopicallyunique ammonia (NH3/ND3) were premixed to theexact volume required and cryogenically pumped onto the metal samples.The samples were isolated, warmed, and monitored for homogeneity.No notable sample decomposition occurred as monitored by lack of H2 evolution.Stoichiometric amounts of mechanicallycleaned metal were loadedunder argon into a sealed flat-plate null-coherent scattering Ti/Zrcontainer of size 15 mm × 30 mm × 1 mm. The container andsample were attached to a stainless steel gas rig and evacuated to<103 andmixed metal NaCa–NH3 solutions were performed usingthe SANDALS and NIMROD diffractometers, respectively, at the ISISSpallation Neutron and Muon Source, UK.14H/D substitution experiments on the Ca–NH16 The resultant totalstructure factor can be expressed asci and bi are theatomic fraction and bound coherent neutron scattering length of species i, respectively. Sαβ(Q) is the Faber–Ziman partial structurefactor, and Q is the magnitude of the scatteringvector.Thesample data were corrected for sample containment, multiplescattering, and incoherent scattering according to standard proceduresusing the Gudrun package.G(r)corresponding to F(Q) is obtainedby replacing Sαβ(Q) with the partial pair distribution functions gαβ(r) by the signed Fouriertransform relationship to giveThe real space function G(r)can be written as a sum of three partial pair distribution functionsbX and atomicconcentration cX are defined asIn the current context, cH = 1 – cX. Then, the gHH(r) term in x is the fraction of protonated ammonia in themixture sample. ThenThe label H refers to substituted atoms in the sample, and X refersto unsubstituted atomic species. It follows from gHH(r) – 1 in eq The remaining partial pair distribution functions can be calculatedin an analogous way to 12 This techniquerefines a structural model based upon an equilibrated Monte Carlosimulation using seed pair-potentials. These potentials are iterativelymodified through the introduction of an empirical potential basedupon the difference between simulated and measured, isotopically uniqueneutron scattering structure factors. Simulation boxes consisted of6000 molecules within the cubic cell of length 64.588 and 66.674 Åfor the 4 and 10 MPM systems, respectively, constructed using thereported densities for the Ca–NH3 system. The seedLennard-Jones plus Coulomb potentials used in the refinement are reportedin 3 with an adjustableCoulomb term to ensure net neutrality in the simulations of the itinerantelectron systems.17 The reduced chargeof Ca ions was taken as those reported from ab initio simulations for single ions in NH3 solutions18 and was found to be necessary for subsequentstructural refinement to the experimental neutron data.Structural models for the liquid systems at4 and 10 MPM were refinedto the respective neutron datasets using the empirical potential structurerefinement (EPSR) software.F(Q), for metallic Ca–NH3 solutionsat 4 and 20 MPM are presented in Q-values is characteristicof those metallic solutions which possess a strong intermediate rangeorder of the solvated Ca2+ cationic centers with increasingmetal content. Their occurrence at 0.61 and 0.85 Å–1 indicate Ca–Ca correlations at ∼7.4 and ∼10.3Å in the 10 and 5 MPM solutions, respectively. Our measurementsare thus in agreement with previous neutron measurements of Ca–NH3.9 It can also be seen that the principal diffractionpeak in the 2.5–3.0 Å–1 region displaysconsiderable broadening with increasing metal concentration, and thefeature between 4.0 and 10.0 Å–1 is distinctlystructured in the more concentrated Ca–NH3 solution.The experimental neutron total structure factors, G(r), for the 4 and 10 MPM Ca–NH3 data, respectively,as well as the extracted partial pair distribution functions, GHH(r), GXH(r), and GXX(r) as defined in GXH(r) and GXX(r) partials are far more structured in the 10 MPM Ca–NH3 solution than they are at 4 MPM in the 2.5–6.0 Åregion, indicating a distinctly ordered solvation environment of theCa ions. In particular, we can assign the feature at 2.52 Å in GXX(r) to the Ca–N distance,being far more pronounced in the 10 MPM solution. Likewise, the narrowingin the most intense feature to 3.2 Å and the appearance of apeak at 5.0 Å can be assigned to cis- and trans-N–N distances,which would be consistent with an octahedral arrangement of NH3 around Ca. It is interesting to compare these liquid, metallicCa–NH3 solutions to solvated, aqueous calcium solutions,19 whereby the Ca–O distances reported areonly slightly shorter at 2.46 Å despite the aqueous ions being8-coordinate as opposed to 6-coordinate for the amines. Turning tothe GXH(r) partials,we see again that the 10 MPM system is more structured with correlationspresumably being successive Ca–H, N–H distances.The origin of these features is clearer when lookingat the realspace transforms for the neutron data. 3 solutions.9 The refinements for both 4 and 10 MPM solutions are well describedeven down to the low-Q region where the relativeintensities of the prepeak in F(Q) are matched across datasets, albeit slightly over-fit in the 10MPM system. Likewise, the full extent of the structuring of the featurebetween 4.0 and 10.0 Å–1 in the more concentratedCa–NH3 solution is not fully captured by the EPSRmodel. We can see clearly where this discrepancy arises by examiningthe simulated GXX(r).The Ca–N distance is correct in the EPSR refinement, but thefeature is sharper than in the measured neutron data, where the intensityis moved into the peak wings. This is most likely due to the insufficienciesof the seed and empirical potentials being too “hard”for a good description of the Ca ions in a metallic solution. Nevertheless,the high level of agreement between our structural models and theneutron data lends credence to our extracted pair distributions functionspresented in In order to extract more informationfrom the experimental datasets,individual pair distribution functions were determined from EPSR modelsfor both 4 and 10 MPM systems, with refinements reported alongsidethe experimental data in GXH(r) partials in the 10 MPMsolution can now be interpreted further as the increasing contributionfrom the Ca–H correlations at 3.0 Å as the Ca concentrationis increased. Only a minor low-r shoulder is seenin the N–H partials, in agreement with the large disruptionof any hydrogen bonding in these metallic systems, a commonality ofall measured metal–amine systems in their metallic phases.The EPSR models also allow us to determine correlations which areonly a minor contribution to the experimental data, such as the secondarysolvation of Ca(NH3)6 in the 4 MPM solution.Clear secondary and tertiary solvation distances are found at 4.8and 8.2 Å.The pair distribution functions extracted from the EPSR model allowus to confirm our assignment of the Ca–N and N–N distancesfrom the measured neutron data and the origin of the prepeak as beingdue to Ca–Ca correlations. The structuring of the measured 3 system, 3 solutions. As expected from our assignments of the experimentaldata, the inner coordination sphere (corresponding to the sharp firstpeak in gCaN(r) in 3 coordinatedtrans to each other. The second coordination shell (correspondingto the broader gCaN(r) peak in the range of 4.0–7.0 Å) contains those NH3 molecules not directly bound to the Ca ions in solution anddemonstrate a drastic change between those located up to ∼5Å and those beyond. The second peak in gCaN(r) for the 4 MPM solutions occurs at 4.8Å, where there is only a minor contribution in the 10 MPM system.This can be assigned to the hydrogen-bonded, secondary solvation ofthe Ca centers and can be seen in 3)6 species. The 10 MPM systemexhibits its secondary solvation maximum in gCaN(r) at 5.9 Å, at which distance thereis very little angular dependence in the spatial density. This demonstratesthat neighboring Ca(NH3)6 units are only weaklycorrelated in terms of their relative orientations despite the strongradial correlation between Ca ions in the metallic solutions. Whilein the 4 MPM solutions, there is no angular correlation in the distributionof Ca ions, in the 10 MPM solution, we do find a slight dominancefor a facial approach of one Ca(NH3)6 unit tothe next, consistent with the bcc arrangement ofthe solid calcium hexammine structure, Ca(NH3)6, formed at the pseudo-eutectic point whereby cooling any metallicCa–NH3 solution to 185 K excess NH3 crystallizesout leaving only the expanded metal.21 This expanded-metalhexammine crystallizes in the Im3m space group with a lattice constant, ao = 9.12 Å, and an octahedral orientation of NH3 moleculesaround each Ca atom.20 Bond distances in the crystalhave been determined as r(Ca–N) = 2.56 Åand r(Ca–D) = 3.12 Å, correlations whichwe have now shown to persist into the liquid state, even upon furtherdilution by additional NH3 as in the 4 MPM solution.In order to build up a three-dimensional pictureof the Ca–NH3 and metal. Our findings forthe Ca–NH3 system further strengthen this viewpointand certainly provide a solution-phase limit in the future searchfor new, crystalline expanded metals.We have previously suggested that for a metal–amine systemto crystallize into an expanded metal upon cooling, there must bea substantial order already present in the liquid both in terms ofprimary solvation and intermediate range order of the metal centers.This leads to the stability of the pseudo-eutectic and prevents theformation of crystalline NH3 system can be disrupted by examiningthe effect of progressive substitution of Ca by Na. The ionic radiiof Ca2+ and Na+ are similar, and both have beendetermined to be octahedrally coordinated in aqueous salt solutions,but in the case of Na+, there is some uncertainty in this,with the experimentally determined coordination numbers ranging from4 to 8.23 We should also note that when consideringNaCa–NH3 solutions in their metallic phase, despitemaintaining the concentration of metal ions (MPM), the density ofconduction electrons in the solution (sometimes referred to as mole-percentelectrons) can vary up to a factor of 2 due to the difference in valencebetween Na and Ca. This conduction electron density is believed tolie predominantly between solvated ion units in metal–aminesolutions, and we can thus determine the effect on the liquid structureof varying this electron density through control of the Na/Ca content.We can investigate themanner in which the structural order inthe liquid Ca–NH3 system at various Na/Ca ratios. We canclearly see that the pronounced prepeak in Ca–NH3 rapidly decreases in intensity as the Na content increases, indicatingthat ion–ion correlations are decreasing, and both the widthof the principal peak and structuring of the feature from 4.0 to 10.0Å–1 decrease. This would indicate a disruptionof the octahedral arrangement of the solvated metal ions, which canbe confirmed by examining the equivalent data in real space. Interestingly,while the M–NH3 correlations are still found at∼2.5 Å, we see a slight shift to shorter bond lengthsas the Na content increases, despite the lower charge of these ions,and there is a progressive disruption of the structure in the 3.0–5.0Å range. This shorter bond is consistent with the Na–Ndistances previously reported for pure Na–NH3 solutionsin their metallic phase.8 It is likelythat the longer Ca–N distances in Ca–NH3 isa reflection of the highly structured Ca(NH3)6 units being more “molecular” in nature than the Na–NH3 analogue. It is clear that the incorporation of Na ions intothe Ca–NH3 solution disrupts the local solvationstructure of the metal ions, with the distinctly octahedral coordinationof Ca being lost upon substitution by Na. This in turn leads to aloss of intermediate range order in the mixed-ion solutions.3 system is structurally quite simple. There is a strong local orderingwhereby the octahedral coordination of Ca by NH3 comprisethe solvated ion cores for both the most dilute and more concentrateditinerant electron liquids. The approach of additional NH3 is toward the octahedral faces, being driven by the formation ofhydrogen bonding interactions with the solvated ion cores. Beyondthis, further NH3 molecules still exist in distinct solvationshells but are spatially isotropic. It is striking to note that evenat low metal concentrations of 4 MPM, a significant intermediate rangeorder exists in these solutions, with the solvated ion cores beingcorrelated out to >10 Å. This becomes more pronounced andwitha shift to a shortened distance of 7.4 Å for the 10 MPM solution.As we have previously noted, this structuring in the liquid stateseems to be a feature of metal–amine systems, which undergocrystallization at their concentration limit, and is significantlyreduced or absent in those which do not.24Despite its molecular nature and the large concentration rangeover which a metallic phase occurs, the liquid Ca–NH3 enables the local and intermediate structureof the bulk liquid to be controlled as a function of the conductionelectron density. Sequential disruption of the primary solvation environmentupon incorporation of Na is evident from the neutron scattering data,which in turn is reflected in the intermediate range, ion–ioncorrelations being reduced. Whether this has an effect on the electronicproperties of the liquids, or indeed one which can be carried acrossinto any crystalline material, is yet to be determined. It is nowclear that, at the very least, the liquid structure for metal–NH3 and other metal–amine systems is amenable to chemicalcontrol.Furthermore, we have showed that the miscibility of Na–and Ca–NH


Human parvovirus B19 (B19V) and human parvovirus 4 (PARV4) are known to infect humans and transmit through contaminated blood and blood products. Globally, three genotypes of B19V, as well as PARV4, have been identified, respectively. The existence of different B19V genotypes in Chinese plasma donors has been investigated, however, the data regarding PARV4 were not available. The main objective of this study is to identify the genotypes of PARV4 circulating in Chinese plasma donors. By using a duplex quantitative polymerase chain reaction assay adapted for all genotypes of B19V and PARV4, 78 source plasma pools for fractionation were screened and quantified. Results showed that positive rates of B19V and PARV4 DNA in plasma pool samples were 25.64% and 14.10%, respectively. PARV4 sequences in two positive samples were next genotyped, and these two sequences belonged to PARV4 genotypes 1 and 2, respectively. In conclusion, the data present demonstrate the existence of PARV4 genotypes 1 and 2 in Chinese plasma donors for the first time and also show the relatively lower prevalence and level of PARV4 DNA in Chinese plasma donors in comparison with that of B19V DNA. At least two PARV4 genotypes, 1 and 2, were currently present in China.The prevalence and level of PARV4 DNA in Chinese plasma donors were relatively lower in comparison with that of B19V DNA.The rate of B19V and PARV4 coinfection in Chinese source plasma pools was low. Viral safety of the products is the top priority of the blood products industry.Parvoviridae, can infect humans and can be transmitted through blood components and blood products. B19V is much concerned in the fields of blood products because (1) it can present at extremely high titers in plasma of acutely infected but asymptomatic donors, thereby resulting in the highly contaminated source plasma pools for fractionation, (2) it is highly resistant to virus inactivation or removal methods used in the manufacture of blood products, and (3) sometimes it can cause severe diseases in at‐risk recipients.Parvoviruses are characterized by small‐sized nonenveloped single‐stranded DNA viruses.cts, and  sometimeSimilar to B19V, PARV4 is also a frequent contaminant of source plasma pools for the production of blood products and the final products.In our previous article, the prevalence of PARV4 in Chinese plasma pools has been reported, but the genotypes of PARV4 were not identified.22.1A total of 78 source plasma pool samples (each comprising 2000–3000 donations) from one Chinese blood product manufacturer were analyzed in this study. All of such plasma samples were sourced from plasma donors in Central China and collected between April 2017 and June 2018. The collections of plasma samples were approved by the National Health Commission of the People's Republic of China, and all of the plasma donors provided informed consent. Every single plasma donation has gotten tested for infectious agents before and after the pooling and was confirmed to be qualified, according to the requirements of Pharmacopeia of the People's Republic of China.2.2Viral DNA was isolated from a volume of 200 μl of each plasma sample with the High Pure Viral Nucleic Acid Kit (Roche Diagnostics) according to the manufacturer's instructions. The concentration and purity of extracted DNA were measured using the GeneQuant 1300 spectrophotometer .2.3The DNA samples were initially tested for presence and quantities of B19V and PARV4 DNA using a duplex quantitative polymerase chain reaction (qPCR) assay, which has been proved to be able to simultaneously detect and quantify all the known genotypes of B19V and PARV4, with an equal limit of quantification of 5 copies/ml.2.4A 161‐bp region related to the NS1 gene of PARV4 (positions 1564–1724 in AY622943) was amplified by nested PCR with the primers described previously.2.5The PARV4 genotypes were determined by phylogenetic tree analysis based on the 161‐nt NS1 region (positions 1564–1724 in AY622943) using MEGA version 6.0. Twenty‐four PARV4 nonredundant sequences spanning this region were downloaded from GenBank (May 2020) and worked as the reference sequences. As reported previously, genetic distances of the sequences were calculated using the Kimura two‐parameter method, and phylogenetic trees were constructed by the neighbor‐joining method with 1000 bootstrap replicates.2.6The nucleotide sequences of the partial NS1 gene of the PARV4 have been lodged with the GenBank sequence database under the accession numbers MT873797 and MT873798.33.1Of 78 source plasma pool samples tested, 20 (25.64%) were positive for B19V DNA, 11 (14.10%) were positive for PARV4 DNA, only 1 (1.28%) was identified positive for both viral DNA, and 48 (61.54%) had no detectable levels of B19V and PARV4 nucleic acid.3.22 to 2.30 × 109 copies/ml plasma. Levels of B19V DNA was as high as 2.30 × 109 copies/ml plasma, although 70% of the positive samples were at low levels (102–104 copies/ml plasma). For PARV4, the level of virus DNA was lower than that of B19V: viral loads ranged from 2.42 × 102 to 2.28 × 105 copies/ml plasma, and most samples contained 102–103 copies/ml plasma. In the one coexistence sample, levels of B19V and PARV4 were low, equal to 1.31 × 103 copies/ml plasma and 2.42 × 102 copies/ml plasma, respectively.The levels of B19V and PARV4 DNA in source plasma pools were shown in Figure 3.32 and 9.75 × 103 copies/ml plasma, respectively. While no targeted PCR products were amplified from five samples containing greater than 5.28 × 102 copies/ml plasma of PARV4 DNA. These unexpected results might attribute to the incomplete sequence of PARV4 genome in these samples, in view of the different target regions between PARV4 qPCR and nested PCR.The genotypes of PARV4 sequences in source plasma pool samples were identified. Of the 11 samples found to be positive for PARV4 sequences by qPCR  clustering together with genotype‐1 reference sequences belonged to PARV4 genotype 1, and the other one sequence (pool‐14) clustering together with genotype‐2 reference sequences belonged to PARV4 genotype 2 (Figure 4Since discovered in 2005, PARV4 has received much attention.This is the first report to identify the genotypes of PARV4 circulating in Chinese plasma donors. Source plasma pools are ideal materials for blood‐borne virus genotype studies. In our previous studies, the source plasma pools were used as the samples for B19V genotyping, and first reported the cocirculation of B19V 1a, B19V 1b, and B19V 3b, as well as the putative B19V 1/3 recombinant and new strains in Chinese plasma donations.These results also showed that the positive rates and levels of B19V DNA in Chinese plasma pool samples were relatively higher than that of PARV4 DNA, indicating that the prevalence of B19V DNA in the Chinese population might be higher than that of PARV4, consistent with results detected in the general population from a previous report.In our study, out of the 30 parvovirus DNA‐positive source plasma pool samples, only one was identified positive for both B19V and PARV4 DNA. The low rate of coinfection indicated that the PARV4 was not associated with the infection of B19V. Thus, the implementation of B19V NAT assays was not able to reduce the possible risk of PARV4 transmission by blood products. The transfusion‐mediated transmission of PARV4 remains a concern. Given that associated disease or specific symptoms have not yet been confirmed, there is no need to exclude PARV4 from source plasma pools, at least for the time being.It should be noted that although the use of source plasma pools as materials in this study increases the potential for the detection of PARV4, the data regarding PARV4 containing plasma pools were still relatively limited. In further research, screen and sequence more source plasma pool samples by extracting larger volumes of plasma or concentrating virions by immune adsorption could address this limitation.5In conclusion, the data present demonstrate the existence of PARV4 genotype 1 and 2 in Chinese plasma donors for the first time and also show the relatively lower prevalence and level of PARV4 DNA in Chinese plasma donors in comparison with that of B19V DNA.The authors declare that there are no conflict of interests.Junting Jia searched data and drafted the manuscript. Junting Jia and Yadi Zhong performed the experiments and analyzed the data. Yuyuan Ma, Jingang Zhang, and Deqing Wang designed the study and provided critical revision of the article. Huan Zhang, Dian Yuan, and Limin Ma provided help during the process of experiments. All authors reviewed and approved the final manuscript.


The calcium sites with different coordination environments lead to different reaction pathways and free energy barriers. The low free energy barriers result in that the detachment of the calcium ion is a ligand exchange and auto-catalytic process. Moreover, the water adsorption, proton exchange and diffusion of water into the surface layer accelerate the leaching of the calcium ion from the surface step by step. The discovery in this work thus would be a landmark for revealing the mechanism of tricalcium silicate hydration.Dissolution of minerals in water is ubiquitous in nature and industry, especially for the calcium silicate species. However, the behavior of such a complex chemical reaction is still unclear at atomic level. Here, we show that the ab initio molecular dynamics and metadynamics simulations enable quantitative analyses of reaction pathways, thermodynamics and kinetics of the calcium ion dissolution from the tricalcium silicate (Ca Dissolution of minerals in water is ubiquitous in nature, its mechanism at the atomic level still under debate. Here, the authors investigate the dissolution mechanism of tricalcium silicate at early stage by ab initio molecular dynamics and metadynamics simulations. Calcium silicate is an essential constituent in many natural minerals and has been used in a variety of fields from building materials9 to pharmaceutical products10. Because of its bioactivity, biocompatibility and hydraulic nature, it is also a candidate for drug delivery12, filling and regeneration materials in dentistry14 and bone tissue15. Above all, its application in cement is of great interest due to huge amount of usage in world widely. Tricalcium silicate (Ca3SiO5) is the main and most reactive calcium silicate species in ordinary Portland cement (OPC)6. It is well known that the cement hydration is stimulated by the dissolution of calcium ions from the Ca3SiO5 surfaces accompanied by the precipitation of lamellar calcium-silicate-hydrate (C–S–H), which is responsible for the cohesivity, durability and mechanical properties of concrete16.Exploring the kinetics of dissolution and dynamic properties at the water/solid interface on the atomic scale is of great significance to understand the natural process and instruct the industrial production at macroscopic scale. This has been at the heart of numerous research fields, such as geochemistry3SiO5 hydration exhibits clear stages  and is governed by multiple coupled parameters diverging in different time scales (from fs to years) and space scales , which is extremely complex to depict precisely. The experimental studies found that during the dissolution process the surface topography undergoes a complicated transformation with the formation of etch pits, point defects and screw dislocation17. Besides, the hydrated silicate species above the surfaces reconstruct with the remaining Ca ions after the detachment of Ca ions19. In general, the dissolution rate is well accepted to be affected by the grain particles size, overall reactive surface area, temperature, components of solution and dislocations on the solid surface20 on the macroscopic scale. Alongside these, the global dissolution rate is also controlled by the slowest step, which depends on the individual stage during reaction. However, the case would be more intricate for Ca3SiO5 due to the coupling effect with the precipitation of hydrate product. It has been observed the dissolution rate of Ca3SiO5 is extremely fast initially and then decreases dramatically from the highest to the lowest21. The reasons for this phenomenon are still on debate. Firstly, the hydroxylation prior to dissolution may stabilize the surface and therefore lower the solubility of Ca3SiO5, as is the case for other minerals20. Furthermore, the dissolution theory17 implies the driving force for the initial swift dissolution rate is the high degrees of undersaturation as it is energetically favorable for etch pits to form. When the composition of the solution is very close to the solubility equilibrium of Ca3SiO5, the etch pits no longer form and even step retreat, thus limiting the dissolution rate rather severely22, like the natural weathering and other mineral hydration23. Moreover, there may also be an electrical double layer24 and a metastable hydrate phase barrier21 formed on the Ca3SiO5 surface.The Ca3SiO5 interface using atomistic simulations can provide some new insights on the Ca3SiO5 dissolution. The adsorption of water on the Ca3SiO5 surface with molecular and dissociative mode25 is the first step of Ca3SiO5 hydration, which happens even before contacting the bulk water due to the strongly hydrophilic nature of Ca3SiO526. After the surface hydroxylation and the proton hopping into the surface27, the Ca ion will dissolve into the solution destroying the initial surface topology and promoting the further water penetration27, which is a key step for advancing the Ca3SiO5 hydration. For this process, the density functional theory (DFT)-based geometry optimization calculations28 indicated the adsorption of water on the Ca ion impairs the bond strength between the calcium and oxygen ions on the surface. Recently, reactive MD simulations have been widely used to study the Ca3SiO5 dissolution and successfully obtain several new perception on dissolution process. Manzano et al.27 found the Ca ion desorbs quickly and tends to accumulate as inner- and outer-sphere complexes at the Ca3SiO5 (111) surface. Qi et al.29 showed a more easier Ca dissolution from the Ca3SiO5 (010) surface than the Ca2SiO4 (100) surface due to the higher surface hydroxylation degree. Sun et al.30 did not observe dissolution of Ca ions from the (010) surface even after 10 ns at 300 K, but after raising the temperature to ~1000 K, the dissolution rate increases five times than that of room temperature. Claverie et al.31 first investigated the Ca3SiO5 hydration using ab initio molecular dynamics (AIMD) simulations and found that the hydroxides formed on superficial oxide ions are highly stable. However, they did not observe an obvious vertical displacement of Ca ions relative to their initial positions. In fact, it is very hard to probe a complete calcium dissolution process at the atomic level using the AIMD simulations32 with small timescale (i.e. within 100 ps). The breakage of Ca–Os  bonds and formation of Ca–Ow (Ow indicates the oxygen ion in water) bonds is indeed a rare event for not only AIMD, but also the reactive MD, which calls for the cooperation with the enhancing sampling method, such as metadynamics. Uddin et al.33 used reactive forcefield (ReaxFF) combined with metadynamics to calculate the free energy changes of dissolution of Ca ions from various Ca3SiO5 surfaces along the reaction coordinate of the distance between the center of mass and the selected calcium ion. However, this collective variable cannot illustrate the nature of dissolution clearly, which is well accepted as a ligand exchange reaction34. Moreover, the chemical reactions at water/Ca3SiO5 interface are typically accompanied by electron transfer. Hence, it is indispensable to give an ab initio description of such a fundamental reaction.Understanding the interfacial reactions at the water/Ca3SiO5 at early stage with ab initio method. We calculated the reaction pathways, free energy changes and free energy barriers of Ca3SiO5 dissolution using the ab initio metadynamics simulations. We show that the calcium dissolutions at different sites have different reaction pathways and the less coordinated Ca is easier to escape from the surface. Furthermore, we found that water molecules can reduce the dissolution free energy barriers not only by attractive effect through adsorbing on Ca, but also by repulsive effect through proton penetrating into the surface and water diffusion into the original Ca site. Our findings pave the way to the atomistic understanding of surface reaction for the Ca ion dissolution from Ca3SiO5.Here, we uncover the dissolution mechanism of Ca3SiO5 hydration, especially the dissolution of Ca ions, is a process of breaking the old Ca–Os bonds and forming new Ca–Ow bonds. Therefore, we probe into the coordination environment of the Ca ion to calculate the full dissolution pathways. The Ca3SiO5 dissolution rates at different surface sites  are typically different due to the different chemical environments around the Ca ion. The coordination environments of the Ca ions on surfaces are various due to the low symmetry of the M3 type of Ca3SiO5 and the large number of possible surfaces formed during the high-temperature calcination process. For example, the Ca coordination environments at seven low-indexes surfaces range from three to seven (Supplementary Table 3SiO5 (111) surface and they can be classified into three- and five- coordinated Ca species, which are indicated by Caα and Caβ in this work, respectively  free energy surface (FES) , CN(Ca–Ow)), where X indicates the state number on the FES. When the water encounters with the Ca3SiO5 surface, the stable state changes from  (the state before water contacting the substrate) to A  = 3.57 kJ/mol and ΔA⧧(B–C)) = 11.76 kJ/mol), the system comes to the most stable state C with up to four adsorbed water molecules. This high-coordination (seven-coordinated) state compromises the breakage of the original Ca–Os bond but associated with huge free energy barriers (ΔA⧧(C–D)) = 15.71 kJ/mol and ΔA⧧(D–E)) = 14.28 kJ/mol) and a little increase in free energy changes (ΔA(C–D = 4.69 kJ/mol and ΔA(D–E) = 3.48 kJ/mol)). The breakage of the bond between Ca and Osi (the oxygen ion from the silicate group in Ca3SiO5) is earlier than that between Ca and Oi  due to the sequence of reaction pathways from state C to D to E. While the breakage of Ca–Osi bond is more difficult than that of Ca–Oi bond owing to the higher free energy barriers between states C and D. These two sequential steps of breaking Ca–Os bonds decrease the total coordination number from seven to five, making this detached and free Ca ion have more chances to accommodate one more water ligand and reform an octahedral structure, although at this stage it is severely distorted with a trigonal bipyramid (D3h) structure. The free energy barrier and the free energy change for these steps (ΔA⧧(E–F) = 8.70 kJ/mol and ΔA(E–F) = −3.87 kJ/mol) are relatively high compared to the same fivefold to sixfold coordination transition step (A–B).For the dissolution of CaES) Fig. . In addiES) Fig. , c. The  2) Fig. , indicatβ to investigate whether the dissolution pathway alters with initial coordination environment. Obviously, the FES for the detachment of Caβ is different from that for Caα . Albeit the number of water molecule is less than that for Caα, the total coordination number is same with six. However, the reaction pathway for the next step is more complex. The state A has two potential reaction paths to adsorb more water molecules. The first one adsorbs one more water molecule directly without breaking the Ca–Os bond (A–B). While the other one does it by breaking two Ca–Os bonds at the same time (A–D). From a thermodynamic point of view, it is more energetically favorable to pass through state B first due to the larger free energy change between the state A and the next free energy minimum. Nevertheless, from a kinetic point of view, it is faster to react along the pathway of A–D because of its lower free energy barriers (ΔA⧧(A–D) = 6.49 kJ/mol and ΔA⧧(A–B) =  7.01 kJ/mol). The reaction pathway of A–B–C–D for the dissolution of Caβ is similar to the B–C–D–E for the dissolution of Caα, in which the start of Ca–Os bond cleavage is from the sevenfold coordination state. Noticeably, the seven-coordinated species is an essential intermediate in the dissolution of Ca, which is similar to the decomposition of γ-Al2O335. But the free energy barriers along the B–C–D (ΔA⧧(B–C) = 7.38 kJ/mol, ΔA⧧(C–D) = 4.24 kJ/mol) is much smaller than that along C–D–E for Caα. The state D is the most stable state with the same coordinate of the start point, A, on the FES of Caα and is also a new outset for other two different reaction pathways towards breaking one more Ca–Os bond. The reaction would be more likely to proceed in the direction from D to E due to the lower free energy barrier (ΔA⧧(D–E) = 19.60 kJ/mol) compared to the route from D to G (ΔA⧧(D–E) = 29.02 kJ/mol, ΔA⧧(E–F) = 26.11 kJ/mol)) and it is the rate-controlling step among all the reactions. It should be noted that the further Ca–Os bond cleavage from 2 to 0 is not accessible during this simulation due to the large free energy barriers required. Therefore, to uncover the subsequent dissolution mechanism of Ca with less than two- and even zero-coordinated Os, it is necessary to add a ‘wall’ to constrain the CVs in the region of interest.The coordination environments of Ca ions may change the dissolution pathways as well as the thermodynamic and kinetic properties. Thus, we carried out the comparative WT-MetaD simulations for CaCaα Fig.  and becoβ with CN(Ca–Os) from 2 to 0, we assume that the start point is the most stable state H on the FES   and sevenfold state L to the sixfold state K are spontaneous (ΔA(M–K) = −2.09 kJ/mol and ΔA(L–K) = −10.30 kJ/mol), but also because the free energy barriers (ΔA⧧(M–K) = 10.34 kJ/mol, ΔA⧧(L–K) = 6.96 kJ/mol) are lower than those of the reverse reactions, which is confirmed by an additional 30 ps equilibrium AIMD simulations. This final run also shows that the CN(Ca–Ow) increased slightly, and the dissolved Ca forms a more regular octahedral structure with water and hydroxyl with the time evolution  Fig. . The nexe state H, 4 on thz direction on the Ca3SiO5 surface. In the region I (6–8.7 Å), it is obvious that the H ion penetrates into the second layer of the surface (z = 6 Å), due to the escape of the Ca ion and the proton exchange from water molecules to the inner Oi ion. Additionally, the position of the peak of Ow and H is overlapped at 8 Å, indicating one water molecule settles down the original position of the dissolved Ca ion with its dipole moment parallel to the surface. The region II (8.7–11.1 Å) is the chemisorbed water molecule region showing an apparent and strong peak for both Ow and H at nearly the same position, which means the chemisorbed water molecules have a well-ordered structure and tend to orient their dipole moments parallel to the surface. This result is different from our previous work on the Ca3SiO5 surface without Ca dissolution25, which presents a tendency for upright configuration with Ow below H. This divergence indicates the dissolved Ca ion tunes the direction of the chemisorbed water to a flatter configuration. The region III (11.1–14.3 Å) is a mixture of the physisorbed water molecule for the surface and the first hydration shell of the dissolved Ca ion. In this region, the obvious overlap of the peaks for Ow and H disappears, indicating a destruction of the layered water structure. The region IV (14.3–20.8 Å) is the transition layer with a less extent of the structuration and ae oscillating at 0. The region V (20.8–28 Å) is the bulk water layer with ae almost 0, and region VI (28–33 Å) is the water/vacuum layer. The regions of IV, V, and VI are similar to those on the Ca3SiO5 surface without Ca dissolution not only in width but also in intensity. We also calculated the infrared (IR) spectra for the system and extract the parts for Si–OH and Ca–OH. It clearly shows one band at 916 cm−1   Fig.  shows thons Fig. , the Ca3α is easier than the five-coordinated Caβ not only because of its initial less restraint from the Ca3SiO5 surface, but also the smaller free energy barriers along the reaction pathways. In addition, The free energy barriers between the two stable states on the FES of either Caα or Caβ tends to be larger as the number of Ca–Os bonds decreases, which means the water adsorption on the Ca3SiO5 surface is easier than the detachment of Ca and the kinetic rate decreases gradually as this process proceeds. Nonetheless, even the highest free energy barrier is only 29 kJ/mol, which is easy to be crossed, suggesting the Ca dissolution is an auto-catalytic process. Besides, the free energy changes for the detachment of Caα is negative, while for the detachment of Caβ is positive, which indicates that the dissolution of Caα is spontaneous and Caβ is unspontaneous. However, Uddin et al.33 pointed out that the reactivity of different surfaces for Ca dissolution would be different, thus the value of free energy changes and barriers would vary with Miller indices. It should be noted that there is no human intervention during the simulation of crossing over the free energy barriers, which ensures the reliability of our results.The calcium sites with different coordination environments lead to different reaction pathways, free energy barriers, and free energy changes. The dissolution of three-coordinated Ca3SiO5 hydration. In short, the dissolution of Ca can be explained in terms of a ligand exchange process. It initially stimulated by the water adsorption, raising the total coordination number with oxygen ions to a high level (five to seven). The adsorbed water molecules reduce the free energy barriers for breaking the Ca–Os bond, and thus provide an opportunity for Ca to break the Ca–Os bonds with the optimal six coordination number unchanged. In fact, the breakage of Ca–Os bonds is a multi-step and multi-orientation chemical reaction, and every step needs a relatively high free energy barrier, which is strait to cross for traditional AIMD simulations within 100 ps but easy in practice. In addition, the dissolution of Ca is further promoted by the proton exchange and the diffusion of the water molecule from the chemisorbed layer into the second surface layer. On the one hand, the H penetrates into the second layer of the surface and bonds to the Oi previously bonded to the dissolved Ca, leading to a repulsive force pushing Ca out of the surface. On the other hand, this diffused water molecule resides in the position of the Ca before dissolution and forms the hydrogen bond network with Os, Ow, and H, which undermines the attractive force to this Ca ion.Our ab initio WT-MetaD simulations with explicit water solvation highlight the importance of the water molecules on the detachment of Ca during the Ca3SiO5 surface in water solution at the initial stage of Ca3SiO5 hydration is investigated using the ab initio molecular dynamics and metadynamics simulations. We find that the Ca dissolution from the Ca3SiO5 are multi-step and multi-orientation chemical reactions accompanied by the water adsorption, proton exchange, breakage of Ca–Os bonds, and water diffusion. The reaction pathways for Ca in different coordination environments are different and the less coordinated Ca is easier to leach from the surface. The thermodynamic and kinetic analyses show that the detachment of Caα is spontaneous, while the detachment of Caβ is unspontaneous, and the Ca dissolution is an auto-catalytic process with the highest free energy barriers of only 29 kJ/mol. Besides, we find the water molecules provide not only the attractive force pulling Ca out of the surface, but also the repulsive force in filling the previous Ca site and pushing Ca away. The present achievement thus provides an insight into the cement hydration and also predict the evolution of other complex geochemical and catalytic systems.In summary, an atomistic and mechanistic picture of the Ca dissolution from the Ca3SiO5 (111) surface has been studied to have a greater possibility to form in practice37, we took the pure Ca3SiO5 (111) surface from our previous work25, which was cleaved from the M3 polymorph of Ca3SiO5 (obtained from CCSD38), as the M3 polymorph is the most frequently observed in industrial clinkers together with the M1 form39. The details of the DFT-based geometry optimization of the bulk crystal and surface slab were listed in 3SiO5 surface occurs even before contacting bulk water40, we firstly adsorbed isolated water molecules to saturate the dangling bond on the Ca3SiO5 surface  surface  for the dissolution simulations. The lattice parameters were 14.21 Å × 11.72 Å × 36 Å after setting a vacuum of 15 Å along z direction. After the detachment of the Caβ from its initial position to the state of D, we further constructed a new model (14.21 Å × 11.72 Å × 48 Å) by adding another 10 Å thick layer of water on the previous system  to calculate the structural and dynamic properties of the equilibrium state.Because the Caace Fig. . Then, w41 functional and Grimme D3 correction42, which was implemented in the CP2K/Quickstep code43. The Core electrons were described by Goedecker-Teter-Hutter (GTH) pseudopotentials45 and the valence electrons were described by a mixed Gaussian and plane waves basis (GPW)46. The wave functions were expanded on a double-ζ valence polarized (DZVP) basis set along with an auxiliary plane wave basis set at a cutoff energy of 500 Ry. The Brillouin zone was sampled by the gamma approximation. During AIMD, the nuclei were treated within the Born–Oppenheimer approximation with a timestep of 0.5 fs for equilibrium simulation, while 1 fs for metadynamics simulations with the replacement of hydrogen by deuterium to accelerate the structural evolution without energy drifts47. The temperature was maintained at 300 K using a Nosé-Hoover thermostat49 coupled to the system with a time constant of 1000 fs in the Canonical ensemble (NVT). The convergence criterion for energy was set to 10−12 Hartree and for self-consistent field was 10−6 Hartree. All the system were first optimized to a stable state and then thermalized for at least 2.5 ps before the production run for statistical analysis. The production times for different simulations are shown in Table All the AIMD simulations reported in this work were performed within the framework of DFT with the generalized gradient approximation (GGA) using the Perdew-Burke-Ernzerhof (PBE)50 (WT-MetaD) simulations, we utilized a two-dimensional collective variables (CVs) characterized by the coordination number (CN) to monitor the dissolution process. The CN(Ca–Os) is the coordination number of the Ca ion with all oxygen ions from the surface slab, while CN(Ca–Ow) is the coordination number of the Ca ion with all oxygen ions from water molecules. As defined in the PLUMED code51, the CN have the expression as follows:rij is the distance between atom i and atom j. sij(rij) is a rational type of switching function describing the coordination between atom i and j. d0 is the central value of the function. r0 is the acceptance distance of the switching function, where the function well be n/m at d0 + r0. Here, we define d0 is 2.42 Å, which is the equilibrium bond length between the Ca and O ions52; r0 is 0.4 Å, which is around half of the full width at half maximum of the radial distribution function of Ca–O53 and n and m are 6 and 12, respectively.In the well-tempered metadynamicsα and 24 for simulation of Caβ. In addition, for further investigation of Caβ coordinating more water molecules, we added a quadratic wall with the force constant of 500 kJ/mol at the position of CN(Ca–Os) equals to 1.5 to restrict the simulation of further dissolution of Caβ on the regions of free energy surface with CN(Ca–Os) less than 1.5. The time evolutions of the CV1 and CV2, the convergence tests for the free energy surfaces and the errors between the free energy minima were shown in Supplementary Figs. The Gaussian hills were deposited every 30 timesteps with the initial height of 3.5 kJ/mol and width of 0.15 for both CVs. The biasfactor were 15 for simulations of Caw and H, respectively. The negative value for ae indicates an excess of H, while the positive one indicates an excess of Ow.The atomic excess (ae) is defined as:51. The molecular electric properties were calculated every 4 fs (8 timesteps) using the Voronoi integration approach54. The IR spectrum of particular components of a system were computed through the Fourier transform of the molecular dipole autocorrelation function as follows:ω is the frequency, and For the vibrational spectra, we use the last 30 ps equilibrium AIMD trajectory to calculate the infrared (IR) spectrum with the TRAVIS programSupplementary informationPeer Review FileDescription of Additional Supplementary FilesSupplementary Movie 1


Pt, Pd, Ru, Rh, and Ir NPs can also catalyze the dehydrogenation of glucose, except that O2 is preferably reduced to H2O. By the electron transfer feature of noble metal NPs, we overcame the limitation that H2O2 must be produced in the traditional two-step glucose assay and realize the rapid colorimetric detections of glucose. Inspired by the electron transport pathway in the catalytic process of natural enzymes, noble metal NPs have also been found to mimic various enzymatic electron transfer reactions including cytochrome c, coenzymes as well as nitrobenzene reductions.Au nanoparticles (NPs) have been found to be excellent glucose oxidase mimics, while the catalytic processes have rarely been studied. Here, we reveal that the process of glucose oxidation catalyzed by Au NPs is as the same as that of natural glucose oxidase, namely, a two-step reaction including the dehydrogenation of glucose and the subsequent reduction of O 2O2; and that other noble metal NPs also catalyse glucose dehydrogenation, but oxygen is preferably reduced to water.Gold nanoparticles (Au NPs) with enzyme-like activities are useful glucose oxidase mimics, but the insights into the mechanism of this reaction are limited. Here, the authors show that the process of glucose oxidation by Au NPs is analogous to the one catalysed by glucose oxidase, involving dehydrogenation and oxygen reduction to H At present, there are two key problems that existed in the research field of nanozymes, that is, low catalytic activity and poor specificity. To solve these problems, a strategy based on biomimetic simulation of the structure of natural enzymes, i.e., single-atom nanozyme (FeN5) with super high oxidase catalytic activity, and a copper coordinated MOF-818 with high specificity for catechol oxidation, have been demonstrated successfully5. Although the biomimetic strategy has been proved to be very effective for the design of nanozymes, there are only a few nanomaterials with a similar structure to natural enzymes can be found.Nanozymes—nanoparticles (NPs) that exhibit enzyme-like properties—overcome the drawbacks of natural enzymes, such as easy deactivation, high cost, and poor recyclability, and thus are highly promising alternatives to their natural counterparts with superiority2O2 and has attracted substantial interest in biological detection and therapeutics7. Although GOD mimics are of high research significance, their development is very slow. At present, only a few works have reported nanomaterials other than Au present GOD-like properties9. It is noted that these sporadic reports seldom descript the catalytic processes. Given these limitations, the pursuit of understanding the catalytic mechanisms of Au NPs and other NPs has great significance in rationally designing efficient nanozymes with various and specific functions.In particular, Au NPs are structurally different from the active site of glucose oxidase (GOD) but show GOD-like activity, which is highly effective for the oxidation of glucose to produce H2 serves as an oxidant and is incorporated into the reaction products, and (2) dehydrogenation oxidation, in which protons and electrons are removed from the substrate and transferred to an electron acceptor12. Oxygenation is quite common in various reactions, and active intermediates containing oxygen, i.e., superoxide anion radical (O2•−), hydrogen peroxide (H2O2), singlet oxygen (1O2), and hydroxyl radical (•OH), must be produced to oxidize the substrate. For the dehydrogenation reaction, the oxidant plays the role of an electron acceptor and does not react with the substrate directly13. Glucose oxidation in vivo occurs through the dehydrogenation reaction, which is catalyzed by dehydrogenase ) with nicotinamide adenine dinucleotide or nicotinamide adenine dinucleotide phosphate NAD(P) as the electron acceptor or by oxidase  with flavin and O2 as intermediate and terminal electron acceptors, respectively14. GDH and GOD catalyzed glucose oxidation share a lot of similarities, and a difference is that the reduced product of NAD(P) is stable under ambient conditions, while the reduced flavin catalyzed by GOD is sensitive to air and will further donate electrons to O2 to generate H2O2. Understanding the catalytic mechanism of natural enzymes will be helpful to learn the GOD-like activity of Au NPs and further rationally design GOD mimics.Generally, there are two approaches for substrate oxidation by natural enzymes: (1) oxygenation oxidation, in which O+•) instead of O2 as an electron acceptor to study the GOD-like activity of Au NPs. In the presence of glucose, Au NPs can catalyze the reduction of ABTS+•, demonstrating that Au NPs catalyzed the oxidation of glucose by direct dehydrogenation rather than active oxygen radicals. The dehydrogenation was further confirmed by electron paramagnetic resonance (EPR) spectroscopy, as the 5,5-dimethylpyrroline-N-oxide (DMPO)-H adduct was detected during the course of glucose oxidation. These results indicate that Au NPs catalyzes the transfer of electrons from glucose to O2 and the reduction of O2 to H2O2. Similar results were also found with various noble metal NPs , except that the oxygen was reduced to H2O instead of H2O2. In addition, noble metal NPs can catalyze the reactions of a variety of biomolecules, which can be catalyzed by flavoenzymes in organisms. Therefore, we defined these NPs as coenzyme mimics.In this study, we utilized 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) radical (ABTS4 as the reductant. Transmission electron microscopy (TEM) results showed a high degree of NPs dispersion, with mean particle diameters of 5 nm for all samples  as the stabilizer and NaBHes Figs. –s6.2 as a model system to evaluate the catalytic activity of the Au NPs -based colorimetric system using 3,3′,5,5′-tetramethylbenzidine (TMB) as a chromic indicator. After incubation of Au NPs with glucose for 10 min, the characteristic absorbance spectrum of TMBox was observed when HRP and TMB were introduced into the supernatant  as well as to H2O2 (2e− path)15.We used the oxidation of glucose by ONPs Fig. . The gennt Figs.  and s7. 2 to generate H2O2 but also to other electron receptors, such as methylene blue (MB+), ferrocene (Fc), and ABTS+•16. Therefore, we utilized ABTS+• instead of O2 as an electron acceptor to reveal the similarity in catalytic activities between GOD and noble metal NPs due to the high reversibility and obvious color change  . Therefore, the oxidation of TMB requires the production of reactive oxygen species such as •O or •OH in the presence of O2 or H2O223. In contrast, the oxidation rate of glucose increased with increasing pH, implying that the oxidation of glucose does not rely on the oxidation ability of O2.In addition to the ability to mimic GOD, the NPs have oxidase-like and peroxidase-like properties; that is, the NPs use OPD) Fig. 20. The rPs Figs.  and s12.Ps Figs. –15. Acco25. Since DMPO cannot dehydrogenate glucose, the reaction proceeds by abstracting hydrogen from glucose to form an Au–H intermediate and further transferring H to DMPO to generate DMPO-H adducts27. The other five noble metal NPs can also catalyze the dehydrogenation of glucose to form DMPO-H adducts, implying the same catalytic pathway applied.The direct dehydrogenation of glucose catalyzed by noble metal NPs was further proven by electron spin resonance (ESR) spectroscopy with DMPO as the spin trapping agent Fig. . The reaAlthough most previous studies merely focused on the GOD-like properties of Au NPs, the catalytic capabilities of Au NPs are beyond simulating GOD. Essentially, the oxidation reaction of glucose involves the dehydrogenation of the hydroxyl groups on glucose to form aldehyde groups. Therefore, we selected other hydroxyl-containing biomolecules to verify the ability of Au NPs to mimic other oxidases. Under the same reaction conditions, galactose was most easily oxidized among different sugars Fig. . We furt2 but also to electrodes, which conduced its wide usage in enzyme fuel cells28. This electron transfer pathway is also found in Au NPs . Because oxygen can be reduced to hydrogen peroxide and water by the two-electron and four-electron processes, respectively, we further used a rotating ring disk electrode (RRDE) to determine the electron transfer number of oxygen reduction on Au surface31.Natural GODs can transfer electrons not only from glucose to ONPs Fig. . In the NPs Fig. . In the −1, O2 was reduced by 4e− to H2O or by fewer electrons to produce H2O2. H2O2 can be collected by the ring electrode and further oxidized to O2. The amount of H2O2 was determined by the oxidation current at the ring electrode32. The electron transfer number of Au NPs for the ORR was calculated to be 2.38, and accordingly, the selectivity of H2O2 was 81% . Then, a direct hydride transfer occurs from the C1 position in glucose to the N5 position in FAD to produce FADH−. FADH− is very sensitive to air and quickly oxidized by O2  to produce H2O2. The reaction path of Au NPs catalysis is the same as that of natural GOD, except that OH− is used as a Brønsted base to abstract H+ from glucose .The gathered results in our study draw a possible GOD-like mechanism for noble metal nanozyme-catalyzed glucose oxidation. For GOD, glucose is first activated by a close His residue (as a Brønsted base) to remove the C1 hydroxyl proton to form an intermediate Fig. 33–35. Thse Figs.  and s21.2O2, and HRP conducts H2O2 to oxidize redox indicators such as ABTS, which indirectly indicates the concentration of glucose38. Accordingly, researchers used Au NPs instead of GOD and peroxidase mimics instead of HRP to realize the quantitative detection of glucose  as the peroxidase mimic, and ABTS as an indicator to detect glucose. To produce considerable H2O2, the first step of glucose oxidation was carried out under alkaline conditions to achieve higher catalytic activity. Thereafter, the mixture solution was added to another acidic buffer solution. By using the peroxidase-like activity of PB, ABTS was oxidized by H2O2 to form ABTS+• . GOD catalyzes the oxidation of glucose to produce Hose Fig. 39–41. HeS+• Fig. , left. T µM Fig. .Fig. 3Tw+• as both an electron acceptor and color indicator to detect glucose in one step  . Figure 2 over time in the presence of Pt NPs and HCOOK. The concentration of FMN maintained in the first 95 s and then rapidly decreased within ca. 10 s. The concentration of oxygen gradually decreased at the beginning of the reaction and was depleted in approximately 120 s. Since the reduction of O2 is favorable over FMN, formic acid firstly consumed O2, and the concentration of FMN remained unchanged because FMNH2 is very sensitive to O2 and easily reoxidized to FMN. Without O2, FMN is reduced to FMNH2, which is stable in the absence of O2. The reduction of FMN catalyzed by Pt NPs and oxidation of FMNH2 by O2 were quite reversible , coenzyme Q, and cytochrome  c) Fig. . Compare2 Figure . The ele2 Figure  and s234ble Fig. . When sh2 can further transfer electrons to ferric Cyt c to form ferrous Cyt c, which is a step in the electron transfer chain. In the presence of HCOOK, Pt NPs, and FMN, the Soret band peak of ferric Cyt c increased and redshifted from 409 to 414 nm and was accompanied by distinctly increased absorption of the α band (550 nm), indicating that ferric Cyt c converted to the ferrous form H as an electron donor. NAD(P)H is generated from the reduction of NAD(P) by dehydrogenases, such as formate dehydrogenase with formic acid as the electron donor Fig. 43. Due tups Fig. . Among tups Fig. . In the oup Fig. . The redsed Fig. . This isrst Fig. 44. When +• instead of O2 as an electron acceptor and demonstrated that the oxidation of glucose proceeds via dehydrogenation on noble metal NPs. The reaction path of Au NPs catalysis is the same as that of natural GOD, except that OH− is used as a Brønsted base to abstract H+ from glucose. The use of OH− as a Brønsted base corresponds to that glucose oxidation and alcohol oxidation are faster under alkaline conditions. Rotating disk tests confirmed that O2 was preferable to be reduced to H2O2 than H2O with Au NPs, and more likely to be reduced to H2O on the other noble metal NPs. These results explain the phenomenon that only Au NPs out of the six noble metal NPs can catalyze the oxidation of glucose and produce H2O2. In terms of the roles of the noble metal NPs, they catalyze both the dehydrogenation of hydroxyl groups and the ORR whose behavior is similar to that of FAD, thus we can define the noble metal NPs as coenzyme mimics. From the perspective of coenzyme mimics, noble metal NPs have been found to mimic various enzymatic electron transfer reactions for Cyt c and coenzymes as well as nitro compound reduction. It is also suggested that elucidating the catalytic process of natural enzymes and hunting for catalysts with similar catalytic functions are rational strategies to design more kinds of and higher efficient nanozymes.In summary, we utilized ABTS4, 98%) was acquired from Aladdin. Glutathione  was bought from Genview. Glucose (99%) and gold chloride hydrate  were purchased from Beijing Chemical Works. All chemicals were used as received without further purification. Ultrapure water was used throughout the experiments.3,3′5,5′-tetramethylbenzidine  was purchased from Sigma-Aldrich. Sodium borohydride  spectrometer with an Al Kα excitation source. UV–Vis absorption measurements were carried out on an Agilent Cary 60 (Varian) UV–Vis-near-infrared (NIR) spectrometer.4 solution (100 mM) was introduced. The reaction mixture was stirred at room temperature for 24 h. In particular, the Ir NPs were synthesized by reacting at 80 °C for 3 h and then stirring at room temperature for 24 h.In a typical experiment, 0.05 mmol metal salt and 20 mg PVP were added to 45 mL water under vigorous stirring. After 5 min, 5 mL NaBH−1) and 20 μL TMB (20 mM in DMSO:EtOH = 1:9) were added into the above solution. UV–Vis absorption measurements were performed within 2 min.In a typical procedure, 50 μL of as-prepared noble metal NPs and 50 μL of glucose (1 M) were sequentially added into a vial containing 860 μL of 100 mM PBS buffer (pH = 7.4), After 20 min incubation at room temperature, 20 μL HRP  were sequentially added into a vial containing 10 mL of 100 mM PBS (pH = 8). After reaction for 2 h, 5 mL of ethyl acetate was added into the mixture solution to extract organic components. The products were detected by GC-MS.2S2O8 (3.5 mM in water) and stored in dark for 12 h to obtain ABTS+•. In total, 50 μL of as-prepared metal NPs, 50 μL of glucose (1 M) and 20 μL of ABTS+• solution were sequentially added into a vial containing 880 μL of 100 mM PBS (pH = 7.4). The reduction of ABTS+• was measured by the adsorption change at 734 nm.In a typical experiment, 1 mL of ABTS (10 mM in water) was mixed with 1 mL of K2O2 (1 M) were sequentially added into a vial containing 900 μL of 100 mM different buffer solutions (acetate buffer (pH = 5-6); PBS (pH = 7-8); carbonate buffer (pH = 9–10)). The decomposition of H2O2 was measured by the decrease of absorbance at 240 nm.In a typical experiment, 50 μL of as-prepared noble metal NPs and 50 μL of H2 for 20 min before recording the EPR spectra.In a typical experiment, 50 μL of as-prepared metal NPs, 50 μL of glucose (1 M) and 20 μL of DMPO solution were sequentially added into a vial containing 880 μL of 100 mM PBS (pH = 7.4). The mixture was deoxygenated by bubbling N2 (or N2) for at least 30 min. The LSV plots were recorded by applying proper potential ranges at the scan rate of 10 mV/s. The electron transfer number (n) and selectivity of the noble metal NPs toward H2O2 formation can be calculated according to the well-known relation  modified with noble metal NPs  on the disk served as the working electrode. Pt foil served as the counter electrode and Ag/AgCl electrode was used as the reference electrode. Before the electrochemical test, the electrolyte solutions were purged with O2CO3–NaHCO3 solution . The reaction mixture was incubated at room temperature for 1 h. Then 500 μL of the above solution, 20 μL of PB (5 mg mL−1) and 20 μL ABTS (20 mM) were sequentially added into a vial containing 460 μL of CH3COOH–CH3COONa buffer . The reaction mixture was further incubated at room temperature for another 1 h before absorption spectroscopy measurement.In the experiment, 50 μL of as-prepared Au NPs and 100 μL of glucose in different concentrations were added into 850 μL of Na+• solution and 100 μL of glucose in different concentrations were added into 830 μL of PBS . The reaction mixture was incubated at room temperature for 30 min, then tested by absorption spectroscopy.In a typical experiment, 50 μL of as-prepared Au or Pt NPs, 20  μL as prepared ABTSc (50 μM) were sequentially added into a vial containing 880 μL of 20 mM HEPES buffer (pH = 7.4). The FMN reduction was monitored by the adsorption change at 450 nm. The Cyt c reduction was monitored by the adsorption change at 550 nm.In a typical experiment, 50 μL of as-prepared noble metal NPs, 50 μL of HCOOK (5 M) and 20 μL of FMN (1 mM) or Cyt p-NP (1 mM) were sequentially added into a vial containing 880 μL of 20 mM HEPES buffer (pH = 7.4). The p-NP reduction was monitored by the absorption change at 400 nm.In a typical experiment, 50 μL of as-prepared noble metal NPs, 50 μL of HCOOK (5 M), and 20 μL of 2 concentration was measured by using a Clark-type oxygen electrode (Hansatech Instruments).In a typical experiment, 2 mL of as-prepared noble metal NPs, 4 mL of HCOOK (5 M) were sequentially added into a vial containing 34 mL of 20 mM HEPES buffer (pH = 7.4). OSUPPLEMENTARY INFOPeer Review File