These plasma parameters are similar to those attained for typical single-pulse near-infrared laser plasmas but with the advantage of substantially lower pulse energy (by factor of ~ 2.5) in the dual-pulse REMPI case. A combustion study focusing on ignition of propane/air mixtures shows that the dual-pulse REMPI method leads to an extension of the lean flammability limit, and an increase in combustion efficiency near the lean limit, as compared to laser ignition with a single NIR pulse. The measurement results and observed gas dynamics are discussed in the context of their impact on combustion applications.This contribution investigates a novel laser ignition method based on a dual-pulse resonant pre-ionization scheme. The first laser pulse efficiently creates initial gas ionization (seed electrons) through a 2 + 1 resonantly-enhanced multiphoton ionization (REMPI) scheme targeting molecular oxygen (λ ~ 287.6 nm). This pulse is followed by a second non-resonant near-infrared pulse (λ = 1064 nm) for energy addition into the gas via inverse bremsstrahlung absorption. The sequence of two pulses creates a laser induced plasma that exhibits high peak electron number density and temperature (n The technology is of interest to a variety of combustion applications including reciprocating engines, aero-turbines, and rocket engines10. System level features of LI, as contrasted to conventional igniter technology, include: the lack of electrodes (which tend to act as heat sinks) within the combustion chamber, the ability to achieve fine control over ignition timing, flexibility in locating the ignition source (based on focusing optics), and potentially systems with favorable cost, packaging and reliability parameters.The ever-increasing push for energetically efficient and environmentally friendly energy conversion systems, in particular combustion technologies, has driven the interest in laser ignition (LI) as an alternative to conventional ignition systems. LI systems are potentially attractive in this regard, for example, when applied to industrial reciprocating natural gas engines, LI has shown reduction in NO12. In general, short wavelength is better suited for the preionization (owing to more efficient multi-photon ionization) while longer wavelength radiation is better suited for energy addition (owing to more efficient IB). The preionization approach relies on synergistic effects between the two pulses, in particular the increased absorption of the second pulse caused by the preionization. This is distinct from multi-pulse approaches  where the individual pulses each independently cause breakdown14 Experimentally, Yalin et al. showed that preionization due to an ultraviolet (UV) 266 nm beam in air reduces the breakdown requirement of an overlapped near-infrared (NIR) energy addition beam at 1064 nm12. When applied to combustion, Dumitrache et al. showed that the dual-pulse preionization scheme allowed ignition of leaner propane-air mixtures as compared to single-pulse ignition with the same pulse energy15. Subsequent modeling work has further elucidated aspects of the pre-ionization laser plasmas. A computational fluid dynamics (CFD) analysis has shown the role of plasma-driven gas dynamics to enhance the early kernel growth, in particular an ability to tailor the resulting laser plasma  to either suppress or admit the typical 3rd lobe feature which in turn is linked to the ability to ignite lean mixtures16. Kinetic modeling has examined nonequilibrium and electron dynamics in the dual-pulse preionization laser plasmas17.Recent approaches towards LI have examined dual (or multiple) pulse laser plasmas based on pre-ionization, i.e. where a first laser pulse primarily ionizes the gas (to produce seed electrons) and then subsequent pulse(s) primarily heat and add energy to the plasma gas kernel via inverse Bremsstrahlung (IB)18 since the larger volume is statistically more likely to overlap ignitable (well-mixed) regions. Our past work has shown larger kernel volumes  when preionization is employed19. In general, high temperature will benefit ignition owing to promoting chain initiation reactions 20, and higher electron density and temperature can increase the production of radicals and metastables from electron impact reactions21. However, the situation is also complicated by the associated gas dynamics, in particular fluid entrainment and vortex formation/ejection, which has also been shown to play an important role in early kernel growth22. An example of translating to applications is provided by past studies in a rapid compression machine where we showed successful ignition of methane–air mixtures over a range of equivalence ratios with NIR plasmas3. While the present work makes the first examination of the dual-pulse REMPI approach in quiescent, room temperature mixtures, future work should include detailed parametric studies tailored to specific applications.Specifying required plasma parameters for ignition in practical applications is challenging owing to the different possible fuel mixtures and flowfields (e.g. ignition of reciprocating engines versus aero-turbines versus scramjets). The ignition kernel volume, in particular, is expected to be of great importance in combustion devices (e.g. some fuel-injected engines) that exhibit poor mixing of the fuel-and-aire = 1.3–3.7 × 1017 cm−3 using a 3 mJ REMPI pulse compared to ne = 2.5–6.1 × 1016 cm−3 using a non-resonant (1064 nm) 20 mJ pulse with the same beam waists). When the REMPI and NIR pulses are combined, the resonant dual-pulse scheme provides similar gas temperatures and ignition characteristics as compared to the 266/1064 nm scheme but with overall lower energy requirements (EREMPI+NIR = 40 mJ vs. EUV+NIR = 60 mJ). In general, the use of UV wavelengths (harmonic generation) is less energetically efficient on the laser source side. This aspect  should be evaluated at the system level while keeping in mind that in many combustion applications the power of the ignition source is negligible compared with the thermal power of the flame23.In the present contribution, we investigate the use of a resonant preionization step, specifically the use of oxygen resonant enhanced multiphoton ionization (REMPI) at ~ 287 nm, in place of the 266 nm excitation. As will be shown herein, the resonant scheme allows more efficient electron generation (with the preionization) for much lower total pulse energy ) to the intermediate state ) which, when excited by a single additional photon (+ 1) is finally excited to an ionized level of molecular oxygen (O2+ X2Π). At high laser intensities, any molecule reaching the intermediate state will also be subsequently ionized such that, as investigated herein, the overall ionization rate scales as the square of the laser intensity (i.e. step-wise ionization is limited by the 2-photon step)24. In terms of laser plasma formation, this REMPI transition has been investigated by Adams et al. for breakdown and guiding of discharges25 and for thermometry based on microwave scattering from the laser plasma27 . At high laser intensities effects of anomalous laser absorption can also become important29.Figure The remainder of the paper focuses on our investigation of laser plasma formation and ignition using REMPI pre-ionization. We first examine the electron generation (preionization) as a function of laser wavelength in the vicinity of the REMPI transition and confirm resonant 2 + 1 excitation. Next, through the use of Rayleigh and Thomson scattering diagnostics, we consider the temporal profiles of gas temperature and electron density for the dual-pulse preionization plasmas (287 nm followed by 1064 nm pulses). We find that peak temperature and electron density is quite comparable to the corresponding values for single-pulse excitation, but with considerably lower  pulse energy in the dual-pulse REMPI case. In terms of ignition, the dual-pulse REMPI case extends the lean limit for propane-air ignition when compared to a single laser pulse (and again with lower pulse energy). The final investigation examines the gas dynamics and vorticity generation in the laser induced plasma revealing the important role of the 3rd lobe in the flame growth. Conclusions and outlook for future research are also presented.30. Briefly, laser Thomson scattering yields a scattering spectrum due to elastic scattering of photons by electrons which, in the incoherent (non-collective) regime (α << 1), yields a Gaussian spectrum with amplitude proportional to electron density (and probe laser power) and width proportional to the electron temperature. However, for all the scattering measurements reported here a wavelength integrated scattering signal was used. Figure 27 Note that the spectrum here refers to the dependence of the full (wavelength-integrated) Thomson signal as a function of the wavelength of the REMPI laser (not the spectrum of Thomson scattered light). The modeled spectrum requires detailed knowledge of the two-photon line strengths and includes Voigt convolution of laser linewidth with the thermal and natural line-broadening. The spectrum is due to the rotational structure of the vibronic transition from the molecular oxygen ground state to the intermediate state, i.e. C3Πg ←← X3Σ−g , as this is the rate-limiting step of the 2 + 1 step-wise ionization. The reasonable agreement between model and experiment, including the fact that we observe no Thomson signal when detuned from the transition, confirm that ionization is via REMPI, as opposed to via (non-resonant) multi-photon ionization (MPI) and/or EAI. The discrepancies that are present are likely due to a combination of: (1) laser pulse energy  variations over the broad scanning interval and (2) the need to further tune the detailed line strengths and linewidths in the model. The REMPI assumption was further examined by measuring the dependence of the Thomson scatter signal on the energy of the pre-ionization pulse (at fixed wavelength of ~ 287.62 nm). These experiments showed that the Thomson scatter intensity varied with the square of the intensity of the pre-ionization beam (i.e. ne ~ I2) which further confirms plasma excitation via the 2 + 1 REMPI31. This result can be contrasted with similar studies  where, for 266 nm ionization, we found a cubic dependence (i.e. ne ~ I3) indicative of oxygen MPI 19. In summary, these tests confirm the role of the oxygen REMPI pre-ionization in the first step of our dual-pulse scheme. For all subsequent experiments, the wavelength of the REMPI beam was set to ~ 287.62 nm at the approximate peak of the spectrum.The first set of experiments was to confirm the role of the oxygen REMPI in the preionization step based on our experimental scheme using focused nanosecond lasers in air see “” section2 REMPI pulse on its own , single NIR pulse , and the dual-pulse REMPI . The measurements were conducted in a pressure cell filled with zero-grade air at p0 = 1 bar and T0 = 298 K. Even with the relatively low REMPI pulse energy a significant degree of ionization is still achieved. For example, we find electron number density of ne = (1.9 ± 0.8) × 1017 cm−3 at 100 ns after the pulse). In contrast, the non-resonant near-infrared (NIR) pulse does not achieve measurable ionization (breakdown) at such low energies. Indeed, for single-pulse NIR, a pulse energy of 95 mJ was adopted for testing as it was the lowest energy for which consistent (visible) breakdown was achieved. However, once formed, the NIR plasma is found to be almost an order of magnitude denser as compared to the (lower pulse energy) REMPI case, for example ne = (8.3 ± 1.3) × 1017 cm-3 at 110 ns after the pulse. These results are largely explained by the different mechanism involved in non-resonant breakdown at 1064 nm where most of the free-electrons are generated by electron avalanche ionization (as opposed to MPI or REMPI) which also results in a stronger threshold behavior19. For the dual-pulse REMPI case, the resulting plasma is similarly dense as compared to the single-pulse NIR pulse but using approximately 2.5 times less total pulse energy, i.e. 40 mJ (= 3 mJ + 37 mJ) total for dual-pulse REMPI versus 95 mJ for single-pulse NIR. The fast decay in electron density is explained by two main factors: kinetics (rapid recombination and attachment) and gas-dynamics (both experiments and simulations showed that the hot kernel expands very quickly during the first microsecond after plasma formation thus leading to a decrease in density22). In all cases presented in Fig. 1-D Rayleigh and Thomson scattering measurements were performed to determine the plasma temperature and electron number density profiles. Figure 32. Figure 2 REMPI plasma is extremly cold (given the degree of ionization) with a maximum measured temperature of ~ 530 K at 1 μs after the discharge. This is to be expected in a rezonant ionization process where most of the laser energy goes directly into ionization, in this case forming Gas temperature measurements are performed from Rayleigh scattering signals using the same experimental setup as was used for electron density (including the same pulse energies). The Rayleigh signals can only be accurately analyzed (with no Thomson interference) at sufficiently late times after the plasma forming laser pulses0 = 323 K and p0 = 1 bar. Combustion events were monitored based on the temporal evolution of pressure inside the cell. Pressure traces corresponding to ignition of C3H8/Air mixtures via the combined REMPI + NIR pulses are shown in Fig. Laser ignition experiments were conducted in a pressure cell with mixtures of propane-air at initial conditions of Tλ = 287.62 nm, E = 3 mJ and λ = 1064 nm, E = 37 mJ) ignition scheme with past (baseline) measurements of combustion efficiencies for single-pulse NIR  and non-resonant dual-pulse 15 as also shown in Fig. Min = 0.55 for the dual-pulse REMPI and ϕMin = 0.7 for single-pulse NIR, respectively), as well as achieving higher combustion efficiency for the leanest ignited mixtures. Additionally, the REMPI + NIR technique exhibits higher combustion efficiency towards the lean limit as compared to the non-resonant dual-pulse. Ignition using the REMPI pulse on its own (with no accompanying 1064 nm beam) was unsuccessful due to the low temperature generated by the resonant plasma  and a fuel-lean condition of ϕ = 0.7 (bottom) using the dual-pulse REMPI laser technique . At stoichiometric conditions, the flame dynamics exhibit similar behavior as has been observed in the post-discharge of NIR laser plasmas38. Specifically, one observes an asymmetric toroidal flame structure having two side lobes  and a third  lobe that tends to propagate towards the laser source.The flame dynamics induced by the laser plasma kernels are investigated based on OH* chemiluminescence images. The image sequences shown in Fig. rd lobe has been previously observed in single-pulse (NIR) laser breakdown39. However, the regeneration of the third lobe at later times has not, to our knowledge, been reported before. This might suggest that the dual-pulse REMPI breakdown generates stronger (more persistent) vorticity resulting in the observed gas-dynamics in the lean mixtures case, as compared to other LI modes.Interestingly, lean mixtures appear to exhibit a rapid quenching of the third lobe within the first few hundred microseconds  output  from an optical parametric oscillator (OPO) laser system (Continuum Sunlite). The OPO is pumped by a seeded Nd:YAG laser (Continuum Powerlite 8010). For dual-pulse schemes, the energy addition pulse was the fundamental output (λ = 1064 nm) from a second Nd:YAG (New Wave Gemini PIV). The energy of this laser could be changed by a variable attenuator comprised of a half-wave plate and polarizer pair. The two beams are spatially overlapped (with precision ~ 10 f = 300 mm plano-convex lenses (one in each beam path). To vary the offset between the two beams in the axial direction , a translation stage was used to move the focusing lens of the 1064 nm beam. Fine-tuning of the overlap of the two beams (REMPI and NIR) is achieved by minimizing the beam energies required for plasma formation in lab air. For the non-resonant UV + NIR dual-pulse measurements, the UV pulse was obtained from the same Continuum Powerlite 8010 by replacing the third harmonic crystal with a fourth harmonic crystal (266 nm) and overriding the OPO path. For the Rayleigh/Thomson scattering experiments a third probe beam, due to the 532 nm output of another Nd:YAG laser (Spectra-Physics Quanta-Ray), was brought in orthogonally to the two plasma-formation (287 nm and 1064 nm) lasers using a set of dielectric mirrors and a plano-convex lens (f = 250 mm). A series of photodiodes and energy meters were used to monitor laser pulse durations and pulse energies of each beam leg (i.e. 287 (or 266) nm and 1064 nm) both before and after passage through the focal region and combustion chamber. The axial overlap and the beam diameters at the focus were measured using a beam profiler (Ophir SP503). The beam characteristics for each laser beam used are summarized in Table 2 beam quality factor is also provided in Table 40.The experiments were conducted using the basic setup shown in Fig. 42 and here we summarize the essential features. The probe beam is nominally aligned to pass through the center of the laser plasma to allow 1-D (line) measurement of (combined) Rayleigh and Thomson scattering from the laser plasma , and IB, IP, and ID correspond to background counts, plasma luminosity counts, and dark counts, respectively. This approach automatically normalizes for variations in laser spatial intensity along the beam. Finally, the maximum electron number density (or temperature) is extracted from the images by averaging a 4 × 4 pixel matrix around the maximum along the direction of the probe beam.Each scattering measurement is based on a ratio-normalization approach whereby scattering at a given measurement condition  is divided by the scattering from a reference condition . The following relations are used to obtain the plasma electron number density and temperature at each pixel in the images:43 where the ratio approach serves to cancel out constants such as the light collection solid-angle and detector efficiency. The Thomson measurement uses an upper- and lower-bound to yield the corresponding bounds of the electron density estimate. These bounds are shown in Fig. 19. Shot noise (caused by photon counting) is another contribution to the uncertainty. This type of error is more prevalent at the early times when plasma luminosity has to be subtracted from the images  state of the NIR breakdown kernel can be obtained iteratively by fitting the experimental 1-D scattering profiles at early times as shown in Fig. 3. By comparison a dual-pulse kernel takes an almost cylindrical shape with a height of 2 cm and a radius of 0.3 mm, yielding an overall volume of 5.5 mm3. More details about the CFD model and the fitting procedure can be found in our previous work22.In some cases, we also employ a computational fluid dynamics (CFD) model to analyze the temperature evolution of the laser plasmas. The CFD simulations are performed using a Riemann solver developed in-house based on the Navier–Stokes equationsIn particular, CFD is introduced in Fig. 47.Combustion studies were performed in a chamber with central volume of ~ 0.195 L and two side arms of length ~ 20 cm with 2.54 cm diameter circular windows for optical access. Pressure traces were recorded using a piezoelectric transducer (PCB: 113B24) mounted flush on the inner wall of the chamber. At least ten pressure traces were collected for each equivalence ratio, and the results were averaged. The traces shown in Fig. 2 = OH* + CO. The excited OH* emits light at ~ 310 nm as it relaxes to the ground state48. For CL imaging of this transition a 310 nm bandpass filter  was placed in front of the ICCD.Additionally, chemiluminescence images of the OH* radical were acquired with the same ICCD camera used in the scattering experiments. The electronically excited hydroxyl radical is generated during the combustion of hydrocarbon fuels through the chain branching reaction: CH + OThe combustion experiments presented in this study were due to propane-air mixtures at initial pressure of 1 bar. Equivalence ratios ranging from ϕ~0.6–1.0 were tested with mixtures prepared inside the chamber based on partial pressures recorded from a gage (Omega DGP409) mounted downstream of the chamber valve. The equivalence ratio is defined as the ratio of the actual fuel/air ratio to the stoichiometric fuel/air ratio (so that ϕ = 1 corresponds to a stoichiometric mixture and leaner mixtures have ϕ < 1). After filling the chamber, but prior to measurements, we waited at least 10 min to allow thorough mixing of the fuel and air components. The chamber was evacuated and pumped down after each experiment to eliminate any effects of residual gases on subsequent experiments.


There is growing interest in assessing psychological well-being in patients after anterior cruciate ligament (ACL) reconstruction. It is unknown whether an assessment of psychological outcome in addition to tests of muscle function can facilitate decisions on return to sport (RTS). Therefore, the aim of this study was to evaluate passing rates in different physical RTS test batteries, with and without the inclusion of psychological outcome measures 1 year after ACL reconstruction.. Passing rates on different muscle function (MF) test batteries (with results presented as Limb Symmetry Index (LSI)), consisting of knee extension and flexion strength tests, 3 hop tests, and 2 psychological patient-reported outcomes (PROs); Quality of Life subscale from the Knee injury and Osteoarthritis Outcome Score (KOOS QoL) and ACL Return to Sport after Injury (ACL-RSI), were evaluated 1 year after ACL reconstruction. Muscle function test batteries comprised: 2 MF tests ; 2 MF tests and 2 PRO , 5 MF tests , and 5 MF tests and 2 PRO .In this cross-sectional cohort study a total of 320 patients (51% men) aged 18–65 years were included 1 year after ACL reconstructionPassing rates in the different test batteries were 47% for 2 MF tests, 19% for 2 MF tests and 2 PROs, 29% for 5 MF tests and 13% for 5 MF tests and 2 PROs. The use of psychological PROs together with tests of muscle function gave the lowest passing rate (13%). There was a very strong correlation between passing 2 hop tests and 2 PROs and passing 5 MF tests (rφ = 0.41) as well as passing 5 MF tests and 2 PROs (rφ = 0.79).The use of hop tests together with psychological PROs provides a clinician-friendly RTS test battery for assessment 1 year after ACL reconstruction as the passing rate was 19% when using 2 hop-tests combined with 2 PROs, compared with 29% when using 5 tests of MF requiring advanced testing equipment. After an anterior cruciate ligament (ACL) reconstruction, up to 30% of patients suffer a second knee injury within 5 years from surgery . ConsequIn a recent systematic review of clinical practice guidelines (CPG) for rehabilitation after ACL reconstruction, the evaluation of psychological measures was recommended in 2 out of 5 CPGs . DespiteResults from only hop performance can be insufficient as RTS criteria. Furthermore, as demands increase by adding more MF tests, the passing rate  is reduced –18. It iThe aim of this study was therefore to evaluate passing rates in different physical RTS test batteries, with and without the inclusion of psychological outcome measures 1 year after ACL reconstruction.This cross-sectional study was based on data extracted from a rehabilitation outcome registry, Project ACL, on 8 February 2019. Project ACL was established in 2014 and aims to improve the care of patients with an ACL injury through the use of regular assessments as well as to provide patients and clinicians with treatment feedback. Data are collected prospectively at predefined follow-ups with ACL injury or ACL reconstruction as baseline . The folIn the present study, data from the 1-year follow-up were extracted for analysis. Patients included in the registry were eligible if: aged 18–65 years, had undergone a unilateral ACL reconstruction and attended Project ACL’s 1-year follow-up. Patients were excluded if any of the following criteria was met; registered with a second ACL injury, had not performed 1 or more of the 5 tests in the battery of MF tests, or had not responded to the Knee injury and Osteoarthritis Outcome Score, subscale Quality of Life (KOOS QoL) or the ACL Return to Sport after Injury scale (ACL-RSI).The tests of MF comprised of 2 strength and 3 hop tests. Patients are required to go through a detailed familiarization procedure with their responsible physical therapist before they are tested in Project ACL. Before testing, patients performed a standardized warm up of 10 min on a stationary bike and sub maximum trials on each test . The Biodex dynamometer is reliable for testing muscle strength . Peak toHop performance is measured with 3 single-leg hop tests: vertical hop , hop for distance and a 30-s side-hop test. Each hop test was performed with the patients holding their hands behind their back. For the vertical hop, the time from take-off to landing was converted into hop height in centimeters. In the hop for distance test, the distance between top of the toes at take-off to heel at landing was measured in centimeters. For the 30 s side hop test, one trial per leg was allowed, where the patient was instructed to hop as many times as possible over 2 lines 40 cm apart. The number of hops was recorded. The hop tests have good validity and reliability for measuring hop performance in patients with an ACL injury or reconstruction .The results of the tests are presented as the Limb Symmetry Index (LSI), which is the result for the injured leg, divided by the result for the uninjured leg, multiplied by 100 and expressed as a percentage.The KOOS is valid and reliable for patients with an ACL injury . The KOOThe ACL-RSI has been developed to measure an athlete’s psychological readiness to return to sport. The ACL-RSI is reliable, valid, and widely used to predict return to sport , 26. EacThe Tegner Activity Scale (Tegner) is meant to reflect how strenuous a physical activity is for the knee . The scaThe PROs were chosen as the ACL-RSI is specifically developed for patients with ACL injuries, and has been reported with the highest methodological quality to assess patients with ACL reconstruction . The QoLIn this study, 4 different test batteries were evaluated. The names of the test batteries subsequently used in this paper are presented in Table For the 2 MF tests, the vertical hop and the hop for distance were chosen as Abrahams et al.  reportedPassing the tests of MF was defined as achieving an LSI value of ≥90% . When 2 For the psychological PROs, Muller et al.  suggeste. Alpha was set at < 0.05. To test correlations, the Phi coefficient was used for binary variables. Reference values used for the Phi coefficient were: > 0.05 = weak; > 0.10 = moderate; > 0.15 = strong; > 0.25 = very strong [Statistical analysis was performed with the Statistical Package for Social Sciences (SPSS) . Mean values, standard deviations, counts and percentages were calculated and presented for demographic data. To compare passing rates between the different test batteries, the sign test was usedy strong .A total of 320 patients 51% men) met the final inclusion criteria for the study   of the patients passing 2 MF tests (LSI ≥90%), compared with 19% (n = 61) passing 2 MF tests and 2 PROs (p ≤ 0.001). There were 29% (n = 92) passing 5 MF tests, compared with 13% (n = 41) passing 5 MF tests and 2 PROs (p ≤ 0.001)  on 2 MF tests and 2 PROs were significantly (p ≤ 0.001) lower than passing rates (29%) on 5 MF tests.There were 47% n = 15 of the pn = 152) of the patients met the cut-off for ACL-RSI, while 62% (n = 198) met the cut-off for KOOS QoL (p ≤ 0.001). When adding ACL-RSI or KOOS QoL to 2 MF tests, the passing rates decreased from 47% (n = 152) to 20% (n = 65) (ACL-RSI) and 31% (n = 98) (KOOS QoL), respectively (p ≤ 0.001). When adding ACL-RSI or KOOS QoL to 5 MF tests, the passing rates decreased from 29% (n = 92) to 14% (n = 44) (ACL-RSI) and 20% (n = 63) (KOOS QoL), respectively (p ≤ 0.001)  as well as passing 5 MF tests and 2 PROs  . Interestingly, there was a very strong correlation [2 hop tests and 2 psychological PROs as criteria for RTS. The passing rates are comparable or even lower than the passing rates of a comprehensive battery of 5 MF tests (strength and hop). There was no significant correlation between passing MF test batteries and reaching cut-offs for either KOOS QoL or ACL-RSI, which indicates that the use of only MF tests or only psychological outcomes is likely insufficient as RTS criteria.The main finding of this study was that using an RTS test battery comprising relation  between Our results suggest that 1 in every 2 patients passed the RTS criteria and achieved symmetrical knee function, when the decision was based on reaching leg symmetry in 2 unilateral hop tests. With test batteries that comprise more tests, the passing rates decreased, in agreement with the literature –18. MoreCurrent recommendations for RTS evaluation are strongly supported by results from the present study, suggesting that batteries of tests should comprise strength and hop tests, as well as PROs , 37. In The results of low psychological readiness to RTS and unacceptable low knee-related QoL suggest that some patients have recovered MF without recovering psychological impairments. Psychological factors are important during rehabilitation , 39, wheIn this study, a smaller proportion of patients met the criteria for an acceptable ACL-RSI compared with KOOS QoL. The ACL-RSI was developed to assess psychological readiness to RTS . HoweverGiven the high rate of new knee-related injuries in patients after ACL reconstruction , 44 and A limitation of this study is that we did not determine the different test batteries effectiveness to reduce the risk of a second ACL injury. Even though there is evidence , 45 suggThe use of the LSI is a limitation since the patients’ healthy limb can have reduced strength after ACL reconstruction , meaningPatients who suffered a second ACL rupture were excluded from the present study in order to create a group of patients that was as homogeneous as possible. Future studies will show how the different batteries of tests assessed in this study affect the risk of a new ACL injury.In this study, the primary outcomes were results from muscle function tests and PROs. Cases where patients were unable to RTS due to unresolved impairments to the somatosensory system could therefore not be studied.The primary strength of this study is the relatively large number of patients included. Another strength is the choice of PROs, since the KOOS and the ACL-RSI was used, and these have high methodological quality .The use of hop tests together with psychological PROs provides a clinician-friendly RTS test battery for assessment 1 year after ACL reconstruction as the passing rate was 19% when using 2 hop-tests combined with 2 PROs, compared with 29% when using 5 tests of MF requiring advanced testing equipment.


Additive manufacturing promises a major transformation of the production of high economic value metallic materials, enabling innovative, geometrically complex designs with minimal material waste. The overarching challenge is to design alloys that are compatible with the unique additive processing conditions while maintaining material properties sufficient for the challenging environments encountered in energy, space, and nuclear applications. Here we describe a class of high strength, defect-resistant 3D printable superalloys containing approximately equal parts of Co and Ni along with Al, Cr, Ta and W that possess strengths in excess of 1.1 GPa in as-printed and post-processed forms and tensile ductilities of greater than 13% at room temperature. These alloys are amenable to crack-free 3D printing via electron beam melting (EBM) with preheat as well as selective laser melting (SLM) with limited preheat. Alloy design principles are described along with the structure and properties of EBM and SLM CoNi-base materials. Additive manufacturing promises a major transformation of the production of high economic value metallic materials. Here, the authors describe a new class of 3D printable superalloys that are amenable to crack-free 3D printing via electron beam melting as well as selective laser melting. However, only a limited number of existing alloys are amenable to the complex thermal conditions present during metal-based AM, where layer-by-layer growth of the component is achieved through local melting of metal powder by either a laser or electron beam energy source7. AM of metals is fundamentally a repeated welding process, in which a directed energy source is used to locally melt and join material. Candidate materials for AM therefore tend to be weldable alloys, which are less susceptible to cracking mechanisms that originate in the liquid phase, such as liquation cracking or hot tearing, or due to stresses that develop in the solid state, resulting in, for example, strain-age cracking and ductility-dip cracking8.Metal-based additive manufacturing (AM), or three-dimensional (3D) printing, has enabled the fabrication of near net shape metal components with optimized geometries that are not achievable through conventional manufacturing techniques. The promise of increased design flexibility has led to significant interest in applying 3D printing methods to commercial alloys currently used in biomedical, automotive, and aerospace applications9. These alloys consist of a high volume fraction (>0.6) of sub-micron size cuboidal precipitates of the 3, L12) that are coherent with a solid solution-strengthened matrix, or γ phase . However, many of the highest-performing Ni-base superalloys are observed to be non-weldable due to the rapid precipitation of the 10. This behavior is captured in Prager–Shira weldability diagrams that show how increasing Al or Ti content, which both increase the volume fraction of the 11.Due to their excellent mechanical properties at elevated temperatures, Ni-base superalloys are the material of choice for structural components such as single crystal (SX) turbine blades and vanes used in the hot sections of aircraft engines and land-based natural gas turbinesγ phase solidifies, the liquid becomes locally enriched by the rejection of 12. This solute segregation lowers the local liquidus temperature, creating solute-enriched liquid films between solid dendrites that contract at differential rates in the melt pool during cooling, resulting in tensile stresses and cracking15. This susceptibility to liquid-mediated cracking can be influenced by control of the liquid composition and liquid fraction at a given temperature via changes in the alloy composition. Just below the melting point, stresses can be accommodated by plastic deformation in the solid state, which is very sensitive to the temperature at which the strengthening precipitates appear. Therefore, many of the desirable high As the 20 represents the major barrier to the use of these alloys for AM components in critical applications. For alloys that operate at relatively low temperature, such as high strength aluminum alloys, control of grain nucleation in the melt pool via functionalization of powder surfaces can mitigate the cracking problem21. However, this results in a small grain size, which is unfavorable for high-temperature operation. Thus, innovative alloy designs are needed for AM, especially for more severe environments22.The cracking susceptibility of high-performance engineering alloys including high 23. By increasing the solid solution strengthening elements in the Ni-base superalloy Hastelloy X within the existing commercial alloy composition range, a reduction in microcracking in AM was observed24. To control material anisotropy, Haines et al. performed a sensitivity analysis with a focus on adjusting alloy composition to control the columnar to equiaxed transition in Ni-base alloys25. Similarly, control of the columnar to equiaxed transition through AM process control has been employed by Kontis et al. to successfully fabricate a non-weldable Ni-base superalloy through atomic-scale grain boundary engineering26. Additionally, AM allows for the mixing of alloy powders before printing, resulting in the fabrication of metal–metal composites with unique microstructures that would be difficult to fabricate by other means27. Since the γ-Several different strategies have recently been pursued for the development of alloys for AM28. These Co-base alloys are morphologically identical to their Ni-base counterparts, except the 3. The 32.Recent interest in Co-base superalloys commenced with the study of Sato et al. that revealed the possibility of precipitation strengthening in the Co–Al–W ternary systemHere we present a CoNi-base superalloy that can be processed through both selective laser melting (SLM) and electron beam melting (EBM) manufacturing pathways, resulting in crack-free components in spite of the presence of a high volume fraction of the desirable 33. In these alloys the Ni3Al phase is thermodynamically stable up to a few degrees below the solidus temperature, thus it was hypothesized that a gap of  >50 °C between the solidus and solvus temperatures would increase cracking resistance while still maintaining a high volume fraction of precipitates at elevated temperatures.The primary goals for alloy design were good high-temperature strength, achieved via stabilization of a high volume fraction of the 34. This has enabled a suite of tools to be developed and integrated for exploration of the large multicomponent composition-space necessary to discover alloys for AM. These tools were used to expand compositions from the original Co–Al–W ternary to the multicomponent composition of the printed alloy investigated here. Below are brief descriptions of the tools and their role in the alloy design process.The emergence of this class of cobalt-base alloys coincides with a recent thrust to develop computational and high-throughput alloy design tools2 γ, creating either superlattice intrinsic stacking faults (SISFs) or antiphase boundaries (APBs). To assess the role of solute on these planar fault energies, density functional theory (DFT) calculations using the Vienna ab initio simulation package (VASP) along with special quasi-random structures (SQSs), the axial next-nearest neighbor Ising (ANNNI) model36, and the diffuse multi-layer fault (DMLF) model with a proximate structure for the (111) APB38 were conducted. Calculations indicated that Ti, Ta, Nb, and Ni were favorable alloying additions. Creep experiments on several SX variants of these materials confirmed that alloys with these elements possessed creep strengths that exceeded first generation SX Ni-base superalloy levels40.The strength of the two-phase 42.To validate fault energy assessments and understand the complex precipitate shearing mechanisms observed across individual and groups of precipitates following mechanical testing, ab-initio calculations of the generalized-stacking-fault (GSF) potential energy were used for phase field dislocation calculations44. CASM includes cluster expansions for configurational disorder in multi-component solids and lattice-dynamical effective Hamiltonians for vibrational degrees of freedom involved in structural phase transitions. CASM was used to assess the intrinsic thermodynamic stability of the L12 phase at elevated temperatures, to calculate the ternary Co–Al–W phase diagram and to assess the finite temperature fault energies and the driving forces for segregation at planar faults47.Interfacing with first-principles electronic structure codes, a clusters approach to statistical mechanics (CASM) software package automates the construction and parameterization of effective Hamiltonians and subsequently builds highly optimized kinetic Monte Carlo codes to predict finite-temperature thermodynamic and kinetic properties3 phase was developed and subsequently incorporated into the CompuTherm PanCobalt database48. Thermodynamic calculations were used to adjust the Ni content to increase the L12 solvus temperature to the desired range, to predict phases present across composition space in the combinatorial studies described below and to predict solidification paths and their likely influence on printability.An early thermodynamic database in the compositional space adjacent to the Co50. Because first principles modeling of non-stoichiometric oxides is extremely challenging, a combinatorial library approach was developed. Ion plasma deposition (IPD) using five cathodes of different alloy compositions was used to create three libraries covering a spectrum of Co–Ni–Al–W–Cr–Ta alloy compositions with 234 samples. The combinatorial approach52 was coupled with rapid oxide screening based on photo-stimulated luminescence spectroscopy (PSLS)53 and the Calphad database48 to outline regions of composition space giving rise to α-Al2O3 scales and two-phase Ternary Co–Al–W alloys have limited oxidation resistance due to the formation of non-protective CoO and mixed spinels40. After analysis of microstructure and phases present, a final composition, SB-CoNi-10, possessing a desirable two-phase microstructure was selected for (a) SX growth to measure basic properties and (b) 3D printing to assess its behavior in both laser and electron beam based PBF methods.Starting from the composition of an alumina-forming IPD alloy of composition Co–32.4Ni–11.7Al–4.4W–3.3Cr–1.5Ta (at.%), three additional alloys with small variations in composition were arc melted into 40 g buttons40. It possesses a high volume fraction of −3), and is able to form environmentally protective α-Al2O3 after high-temperature exposure in air54. The γ and an equilibrium freezing range of 52 °C.The SB-CoNi-10 alloy has a nominal composition of Co–36.4Ni–13.2Al–6Cr–3.5Ta–1W (at.%) along with minor additions of 0.08B–0.08C–0.018Hf–0.002Y which are included for grain boundary strengthening, carbide formation, and oxide scale adhesionA 136 kg lot of SB-CoNi-10 powder was fabricated by vacuum induction melting and argon gas atomization. A powder size range of 15–53 μm was used for SLM printing, while a larger size distribution of 53–177 μm from the same heat was used for the EBM process, as shown in Fig. The 3D printing trials with both the EBM and SLM approaches resulted in crack-free blocks and blade-shaped samples in the as-printed state Fig. c–f. Due The as-printed grain structure characterized by electron backscatter diffraction (EBSD) in the scanning electron microscope (SEM) is shown for both the SLM and EBM processes with inverse pole figure (IPF) maps in Fig. XY-plane, i.e. perpendicular to the build direction, from cross-sections taken 1 mm below the final build layer  along the yer Fig. . By collQuantitative chemical maps of the EBM and SLM material are provided in Fig. 56. Investigations at varying heights in the EBM build  resulted in a microstructure that was free from porosity and also resulted in recrystallization of the microstructure into a coarse-grained equiaxed microstructure  reduces the thermal stresses that develop during solidification, with low orientation gradients present within individual grains  does not promote the formation and coarsening of the 57. Samples tested with the tensile axis along the build direction are indicated as Z-orientation, whereas samples that were tested perpendicular to the build direction are indicated as XY-orientation. Typically, additively manufactured alloys that contain a high density of cracks after printing fail shortly after the yield point. The SB-CoNi-10 alloy demonstrates significantly improved ductility and resistance to cracking compared to similar Ni-base superalloys produced by PBF, as shown by the engineering stress–strain curves in Fig. 59. EBSD maps along the build direction of the post-mortem EBM specimens reveals this grain rotation by showing how the 〈001〉 crystallographic direction of the columnar grains are now more strongly aligned with the loading direction after the test ; thus the computational suite of design tools was essential for efficient exploration and selection of a composition likely to be favorable for AM. By fabricating the alloy SB-CoNi-10 with both SLM and EBM methods, it is suggested that this region of compositional space may yield alloys that are easier to process through AM compared to current Ni-base superalloys. Importantly, it has been demonstrated that the SB-CoNi-10 alloy can be printed in a simple bar form , the structure is refined further still. Finer scale chemical maps acquired by transmission electron microscopy are required to conclusively measure the degree, if any, to which segregation is suppressed in the SLM processed samples.The EPMA grid technique described above is often used on SX castings of superalloys fabricated using the Bridgman method to study the microsegregation of various elements during directional solidification, which strongly affects the liquid density during solidification and can promote casting defects such as stray grains and freckle chains64. The microstructure after EBM resembles the columnar grain structure formed in directionally solidified (DS) castings used in the aerospace industry for superalloy turbine blades. The elimination of transverse grain boundaries in DS castings significantly improves the high temperature creep response compared to equiaxed castings, which extends the usable life of alloys employed where a combination of high temperature and stress are present. Since these columnar grain structures are industrially relevant, this motivated the printing of the limited amount of EBM powder available for this study in the Z-orientation for microstructural investigations and mechanical testing.The alloys described above are readily modified through standard post-processing such as HIP and heat treatment to remove microstructural inhomogeneities present after printing, such as chemical segregation and porosity, and to promote a more equiaxed grain structure after recrystallization. In the case of EBM, where long columnar grains aligned with 〈001〉 along the build direction are observed in the as-printed state, modified heat treatments may be desirable to preserve the as-printed grain structure while removing as-printed segregation and pores, as has been demonsrated in EBM processed CMSX-4Z samples with the tensile axis oriented along the build direction  and XY samples with the tensile axis oriented normal to the build direction (SLM), demonstrating good properties both parallel and transverse to the axis of the columnar microstructure. In spite of their high UTS after significant strain-hardening during plastic flow, tensile ductilities in excess of 32% are observed for EBM SB-CoNi-10, which is higher compared to the standard precipitation strengthened Ni-base alloy CM 247Ⓡ (CM 247Ⓡ is a registered trademark of Cannon-Muskegon Corporation) fabricated by EBM33. For the SLM SB-CoNi-10 specimens, tensile ductilities in excess of 13% are observed, which is again greater than the standard precipitation strengthened Ni-base alloy IN738LC manufactured by SLM57. An anisotropy in the tensile properties is apparent in the SLM samples that depends on build orientation. The as-printed XY-orientated samples exhibit higher strength and lower ductility than their Z-oriented counterparts, which has been observed in other additively manufactured superalloys with columnar microstructures65. This behavior is often reversed when a high density of cracks is present in the as-printed microstructure, resulting in reduced yield strengths as the print orientation changes and preexisting cracks become oriented perpendicular to the loading direction66. The SX Ni-base superalloy CMSX-4 has recently been printed in cylindrical bar form by EBM, producing a SX core with equiaxed grains at the surface67. Removing samples from the as-printed SX core yielded a room temperature yield strength of 829 MPa and elongation of 6.1%, with no apparent cracking. However, it is unclear whether the techniques used to create the single crystalline core could be used to print this alloy into more complex blade shapes. While the SB-CoNi-10 alloy has a lower yield strength compared to CMSX-4, the CoNi-base superalloy compositional space is under-explored and it is likely that future alloy compositions could be designed with higher yield strengths.The mechanical response at room temperature demonstrates the excellent printability of the SB-CoNi-10 alloy. The ultimate tensile strength (UTS) and elongation at failure was high with both In conclusion, a recently developed CoNi-base superalloy, SB-CoNi-10, has been successfully printed using both EBM and SLM. Compositional mapping of the as-printed microstructures reveals that favorable solute partitioning combined with favorable −1/1137 mm s−1, using a bidirectional raster scan strategy. The EBM specimens underwent HIP at 1245 °C for 4 h under 103.4 MPa of isostatic pressure in Ar, followed by a SHT at 1245 °C for 2 h and aging at 1000 °C for 50 h under vacuum. The EBM fabricated samples were cooled by furnace shut-off quenching after each heat treatment step. The SLM specimens underwent HIP at 1177 °C for 4 h under 103.4 MPa of isostatic pressure in Ar, followed by a SHT at 1245 °C for 1 h and aging at 1000 °C for 50 h. In the case of the SLM samples, these samples were cooled by an oil quench after the SHT, followed by a furnace quench after the aging heat treatment.Metal powder of SB-CoNi-10 for the AM trials was produced in a 136 kg batch by vacuum induction melting and argon gas atomization. Powder was separated into size ranges of −53/+15 μm for SLM and −177/+53 μm for EBM. The EBM fabricated samples were produced on an Arcam Q10+ system running 4.2.89 EBM control software under a controlled He vacuum with an applied preheat temperature of 1000 °C. Other EBM printing details include: max current = 18 mA, speed function = 63 (unitless), hatch spacing = 125 μm, layer thickness = 75 μm, using a standard Arcam raster scan strategy. The SLM fabricated samples were produced on a SLM Solutions SLM125 with an applied preheat of 200 °C in an inert gas atmosphere. Other SLM printing details include: layer thickness = 30 μm, laser power = 134 W/176 W, hatch spacing = 80 μm, laser speed = 744 mm s68 and Hough indexing within OIM Analysis™ v8 software. Fracture surfaces were imaged using the aforementioned SEMs at similar accelerating voltages.Samples were prepared for microscopy using standard metallographic techniques consisting of grinding with SiC papers down to 1200 grit followed by polishing down to a 50 nm colloidal alumina suspension using a Vibromet polisher for 4 h. Scanning electron microscopy with secondary electron (SE) imaging and backscattered electron (BSE) imaging was performed on a ThermoFisher Apreo C at accelerating voltages between 5 and 20 kV using a Schottky field emission gun. EBSD maps were acquired using an EDAX Velocity EBSD camera in a FEI Versa3D microscope at an accelerating voltage of 30 kV. All IPF maps are defined such that the 〈001〉 is aligned with the build direction. The collected diffraction patterns were indexed by spherical indexing using the EMSphInx v0.2 software package6 thermionic emission gun. To quantify the amount of chemical segregation present after printing in both the EBM and SLM samples, a 20 × 20 grid scan of evenly spaced points with long dwell times (1 min of collection time per point) was collected in a 100 × 100 micron area on cross-sections taken 1 mm below the final build layer perpendicular to the build direction. Chemical maps with a reduced dwell time were used for visualization of this chemical segregation. Probe Image and Probe for EPMA software packages developed by Probe Software, Inc. were used for quantification of the collected spectra. All EPMA data underwent standards-based quantification with a combination of pure alloy standards and a reference alloy sample with composition Co–6.7W–8.9Al–3.3Cr–1.5Ta (at.%) that was previously characterized to high accuracy by inductively coupled plasma compositional analysis.EPMA was performed using a Cameca model SX100 with five wavelength dispersive spectrometers and a CeBfs) from 0 to 1, as shown in Fig. fs = 0, since Co partitions to the dendrite core during solidification, as shown in Fig. k, for each major alloying addition, the sorted concentration of the solid vs. apparent fraction solidified curves were fit using the Scheil equation55Cs is the concentration of the solid, Co is the starting composition of the melt, fs is the fraction solidified, and the distribution coefficient is defined as k = Cs/Cl. Curve fits to the Scheil equation were made using a least-squares regression using data with apparent fraction solidified values between fs = 0.1 and fs = 0.9 for both the EBM and SLM datasets. These curve fits provided values for Co and k using the values of Cs and fs as inputs determined experimentally by the EPMA grid scan.The collected EPMA grid scan data were sorted from highest Co concentration to lowest and assigned an apparent fraction solidified (Z-orientation) at a strain rate of 1 × 10−4 s−1 using an Instron 5582 universal testing machine. The SLM manufactured specimens were mechanically tested in the Z- and XY-orientation according to ASTM E8/E8M using machined cylindrical threaded dogbones with a gauge diameter of 6.35 mm. The strain rate was 3 × 10−5 s−1 before yield and 1 × 10−4 s−1 after yield. All mechanical tests were performed at room temperature and taken to rupture. EBSD maps of post-mortem specimens were collected after cross-sectioning the samples in half along the build direction.Sheet dogbones for mechanical testing were removed from the EBM-fabricated bars using electric discharge machining. The dogbones were 44 mm long with gauge dimensions of 2 mm × 3 mm × 14 mm. To remove the EDM-affected zone on the surface of the specimens, the gauge sections were ground down to 1200 grit using SiC grinding papers. Tensile tests on the EBM manufactured specimens were performed along the build direction (i.e. Peer Review File


Postexercise total mood disturbance score was significantly decreased for both conditions  and AME baseline 22.4 ± 4.4 vs. postexercise 13.1 ± 5.2, P < 0.05). These senses of changes in feelings were associated with significant physiological increases in V.O2 and MCAv during both PME and AME (P < 0.05). These results demonstrate that physiological and mood responses to passive and active motion cycling exercise are not different. Future studies should determine whether passive motion cycling exercise is a useful preventive medicine strategy for overcoming various disease-related exercise limitations and counteracting the adverse effects of sedentary lifestyles.Previous studies suggest that passive motion exercise (PME) may be useful for overcoming exercise limitations associated with a sedentary lifestyle, orthopedic disorders, and various other debilitating conditions. Negative mood response is one of the factors that limit a person's ability to exercise. Therefore, this study tests the hypothesis that the mood response associated with PME is not different than the mood response associated with active motion exercise (AME). Eight women and seven men participated in the study and were administrated the Profile of Mood States (POMS) during modes of PME and AME in a randomized order. Outcome of the POMS consisted of the total mood disturbance score [(feelings of tension + depression + fatigue + anger + confusion) − vigor]. ANOVA was used to determine significance of differences in total mood disturbance, oxygen uptake (V.O The positive effects of acute physical activity on mood are well established. The available evidence supports the hypothesis that acute exercise can ameliorate a depressed mood, reduce anxiety and enhance cognitive functions, increase perceptions of energy, and decrease those of fatigue –5. A perSubjects were college students who volunteered following recruitment using an institutionally approved flyer posted throughout the university. Before participation, interested persons completed a medical history form, International Physical Activity Questionnaire (IPAQ) , and sigThis study was designed as a crossover experiment wherein one group was subjected to conditions of performing a 30 min bout of passive motion exercise (PME) and a 30 min bout of active motion exercise (AME). The participants were randomly assigned, by a coin toss, to the order of performing the two exercise protocols, PME and AME. The randomly assigned bouts of PME and AME were performed at the same time of day separated by one week. The research assistant was not blinded to the test conditions.2) was measured using an indirect calorimetric method. Respiratory gas concentrations of expired O2 and CO2 were measured through a mouthpiece connected to an O2 and CO2 analyzer . Minute ventilation and expired gas volumes were measured using a flow turbine . The gas analyzers and gas volume turbine were calibrated before each test. End-tidal CO2 (ETCO2) was measured through a sample line using a CAPNOMAC ll capnometer . Middle cerebral artery blood flow velocity (MCAv) was assessed by transcranial Doppler 6 ultrasonography , using a 2 MHz probe placed over the temporal window. The right MCAv signal was identified and tested according to standard criteria guided by signal depth, velocity, and characteristics  was used for analyzing the mood response to both PME and AME. Mood and total mood disturbance scores were inversely related; decreased total mood disturbance score was indicative of improvement in mood.2, and ETCO2 and then connected to a motorized leg-cycle ergometer (MOTOmed ®). After 10 min rest, baseline measures of MCAv, ETCO2, V.O2, and POMS were recorded. Immediately following baseline measurements, each subject performed 30 min of completely motor-supported leg cycling (PME) at a pedal rate of 60 rpm or 30 min of voluntary leg cycling (AME) at a pedal rate of 60 revolutions per min (rpm) with the motor disengaged. During AME, the desired pedal rate of 60 rpm was maintained by requiring the subjects to pedal to the audio cues of a metronome. The 30 min bout of cycling was supervised by a research assistant.Participants were instructed to enter the laboratory after a six-hour fast and no physical exercise for 24 h. Further instructions provided to the female participants included testing limited to the luteal phase of the menstrual cycle to limit possibly hormonal confounding effects. Following 10 min of seated rest (baseline), baseline values were recorded for 1 min; then, participants performed one of the two-randomized conditions: passive cycling (PME) or active cycling (AME). Prior to the baseline period, participants were instrumented for measurements of MCAv, V.Oα level of P < 0.05 was the cutoff for significance. Data are presented as means ± standard errors. The average baseline POMS total mood disturbance scores of the present study group, before performance of the PME and AME protocols, were compared to those of a normative study group of college students reported by Nyenhuis and associates [P-value and confidence interval (CI) of the mood disturbance scores difference was computed as the probability of obtaining the difference between two observed means based on the pooled standard deviations of the two independent samples. The P-value was computed to be the area of the t distribution with n1 + n2 − 2 degrees of freedom that falls outside ± t. P < 0.05 indicated that the two means were significantly different [2 and MCAv and the decrement in total mood disturbance scores were different for the two exercise conditions, PME vs. AME . To determine whether the coin toss randomization with crossover experimental design was effective at limiting bias with respect to the mood disturbance scoring, we grouped the participants into subgroups PME and AME according to which of the two exercise conditions was performed first (with the other exercise condition performed one week later). The t-test for independent samples and unequal variances was used to determine whether there was a significant difference in the baseline mood disturbance scores of subgroups PME and AME based on which exercise condition was performed first.Data analyses were performed using the IBM SPSS Statistics® 19.0 (IBM Corporation) software package. We performed a repeated measures ANOVA with Newman-Keuls post hoc testing to compare the baseline values and the differences between baseline and PME and between baseline and AME measurements. Two-tailed ifferent . Baselin2, and total mood disturbance score made before the bouts of PME and AME were not significantly different  and following AME  (P > 0.1).The total mood disturbance score decreased significantly following PME (postexercise) compared to baseline  . The tot2 increased significantly from baseline to the last minute of PME  and from baseline to the last minute of AME  (2 tended to be higher compared to PME (4.2 ± 0.1 vs 3.6 ± 0.3 mL kg−1·min−1), but the difference was not significant (P > 0.1).Body V.O < 0.05) . The AMEP < 0.05, P < 0.05, P > 0.1).There was a 17.4% increase in MCAv from baseline to the last minute of exercise . That significant changes and condition-related differences in ETCO2 values were not found indicates that changes in the MCAv cerebral blood flow measurements were physiological, resulting from the exercise interventions, not from breathing pattern, ventilation, or PCO2.There were no significant differences in ETCOn = 132) compared to those of the present study group, 26 ± 19 (n = 15). The mean intergroup difference was −13.0 ± 9.72 (SEM) points less for the present study group compared to the normative study group, 95% CI from −33.20 to 5.20 .The POMS total mood disturbance scores of the present study group of 28-to-30-year-old college students were not significantly different than those of a normative study group of 18–24-year-old college students reported . The meaP=0.8).The subgroup that performed the PME first and the AME one week later consisted of 3 females and 2 males; their mean ± SEM baseline mood disturbance score was found to be 24.4 ± 11.2. The subgroup that performed the AME first and the PME one week later consisted of 5 females and 5 males and their mean ± SEM baseline mood disturbance score was 26.1 ± 4.3. There was no significant difference between the baseline mood disturbance scores of the aforementioned subgroups  than the POMS mood disturbance scores of 40 ± 37 (SD) for a normative group of 132 mixed-gender, 13% African-American, nonathlete 18–24-year-old college-age subjects [To our knowledge, this is the first investigation to show mood improvements based on decrements in POMS mood disturbance scores following both passive and active cycling exercise. It is noteworthy that the POMS total mood disturbance scores for our mixed-gender, 100% African-American nonathlete, sedentary study group of fifteen 28–30-year-old college students is 26 ± 19 (SD). This range is not significantly different (In contrast to previous work that compared light-intensity active cycling to passive cycling, Lindheimer et al.  found neThe results of this study should be interpreted cautiously because it did not control for placebo effects on mood responses. It is plausible that the mood disturbance scores reported herein are biased by the subjects' knowledge that a goal of the study was their self-reporting of emotional variables. As mentioned previously, one approach to limiting the placebo effect is to intentionally misrepresent the purpose of the study. Indeed, this was cleverly done in at least one study showing no significant changes in either cognitive or mood responses after passive, as well as after active light-intensity cycling . These fThe present study describes, for the first time, a mood-enhancing effect of passive motion exercise which appears to be no different than the mood-enhancing effects of active motion exercise. These mood-enhancing effects are accompanied by equal increments in oxygen consumption and cerebral blood flow velocity which, by virtue of physiological measurements, support the hypothesis that passive cycling motion exercise elicits a mood response that is not different than the mood response to active motion exercise. These results imply that passive motion exercise may be a useful preventive medicine strategy for overcoming various disease-related exercise limitations and counteracting the adverse effects of sedentary lifestyles.


Parthenium argentatum), a shrub native to the arid region of the U.S. southwest and Mexico belonging to the Asteraceae family, is a source of high quality, hypoallergenic natural rubber with applications in pharmaceutical, tire, and food industries. Production of rubber results in a substantial amount of resin-containing residues which contain a wide variety of secondary metabolites . In order to enhance the economic viability of guayule as an industrial crop, value-added use of the residues is needed and has the potential to reduce gross rubber production costs. The main objective of this research is the characterization of guayule resin using rapid and accurate analytical techniques to identify compounds of potential commercial value. Guayule resin is inherently complex and includes many high-molecular-weight and non-volatile compounds that are not easy to observe using traditional chromatographic techniques. The combination of two mass spectroscopy techniques: gas chromatography mass spectroscopy (GC–MS) and high-resolution Fourier transform ion cyclotron resonance mass spectroscopy (FT-ICR MS), were used to characterize the composition of the extracted resin from guayule (Parthenium argentatum). FT-ICR MS was used to characterize hundreds of compounds with over a wide range of molecular weights and degrees of aromaticity at higher levels of mass accuracy than other forms of mass spectrometry. GC–MS was used to identify volatile compounds like mono- and sesquiterpene compounds.Guayule ( Specifications TableValue of the Data•This dataset represents a comprehensive characterization of guayule resin using several complementary analysis methods.•Researchers working on natural resin characterization, processing, and utilization can benefit from this data.•Quantitative and qualitative composition data for guayule resin can be used to select future separation techniques for analysis and to identify potential applications of separated resin fractions based on their expected compositions.•Data from GC–MS is commonly available for resin samples and provides a common basis for comparison between this data and previously collected data for other resin samples.•The FT-ICR MS data provides information about a wider range of molecular weights than GC–MS data due to the wider range of molecules that can be detected using the ionization method.1Data here includes: a table of terpene molecules identified within the guayule resin sample by GC–MS with the parameters used for identification ; a table2Guayule resin from pilot-scale bulk (solvent) rubber extraction was acquired from the Bridgestone Americas Biorubber Processing Research Center , and characterized as received. The guayule plants were harvested at 24–36 months old, field dried for 1–7 days  to 10–15% moisture, and milled to pass a ¼ in. (6.4 mm) screen. A miscella of rubber and resin was extracted from the whole ground guayule using a mixture of acetone and hexane. Rubber was precipitated from the miscella with addition of excess acetone. Resin was concentrated by evaporation of the solvent from the miscella; after cooling to room temperature, the resin was nearly solid. The resin was collected into barrels, and stored at ambient temperature and humidity  for up to two weeks prior to shipping for analysis. Upon receiving, resin samples were stored in plastic containers at room temperature.3Guayule resin was analyzed by GC–MS to characterize terpene composition. A 10% (w/v) solution was prepared by dissolving resin sample in carbon disulfide. The analyses were performed using a GC–MS system  equipped with a DB-5 MS capillary column . The injection volume of 1 µL was made in a splitless mode. Helium was used as the carrier gas at a flow-rate of 1 mL/min. Oven conditions were initial temperature 50 °C, 1 min isothermal, 3 °C /min ramp to 320 °C, 10 min isothermal. MS conditions included an EI ion source temperature of 250 °C, an ionization energy of 70 eV, and a mass scan range of 50–550 amu. For the identification of terpene compounds, GC–MS libraries including ADAMS and NIST MS Search 2.0 were used, along with a comparison of the linear retention indices with those reported in Adams library 450%  ≥ 400,000) and mass accuracy (ppm) for detection and identification of thousands of compounds within a single mass spectrum. This technique typically used to analyze complex natural organic mixtures such as petroleum, biofuels, dissolved organic matter, lipids, and proteins 2) and is passed under a krypton VUV lamp producing 10 eV photons (120 nm). Toluene was used to increase ionization efficiency through dopant-assisted photoionization.Fourier transform ion cyclotron resonance mass spectroscopy (FT-ICR MS) is currently the only analytical technique providing the required resolving power  for accumulation in the second quadrupole. Finally, ions were collisionally cooled with helium gas (∼4 − 5 × 10−6 Torr at gauge) before optimized passage 2) [Data collection was facilitated by a modular ICR data acquisition system (PREDATOR) 2) , 9. DataThe authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.


The HARE chip and a toolbox of helpful instrumentation for time-resolved serial synchrotron crystallography are presented. Serial synchrotron crystallography (SSX) is an emerging technique for static and time-resolved protein structure determination. Using specifically patterned silicon chips for sample delivery, the ‘hit-and-return’ (HARE) protocol allows for efficient time-resolved data collection. The specific pattern of the crystal wells in the HARE chip provides direct access to many discrete time points. HARE chips allow for optical excitation as well as on-chip mixing for reaction initiation, making a large number of protein systems amenable to time-resolved studies. Loading of protein microcrystals onto the HARE chip is streamlined by a novel vacuum loading platform that allows fine-tuning of suction strength while maintaining a humid environment to prevent crystal dehydration. To enable the widespread use of time-resolved serial synchrotron crystallography (TR-SSX), detailed technical descriptions of a set of accessories that facilitate TR-SSX workflows are provided. Taking advantage of the ultrashort time-resolutions accessible at XFELs, many of the initial FEL studies have probed sub-picosecond timescales using liquid-jet delivery systems , approximating the low-damage XFEL structures by using the serial data-collection approach to minimize dose accumulation  experiments for three different proteins . To displace any remaining Tris-HCl from the storage condition the concentrated protein solution was re-diluted and concentrated again for nine consecutive times. Vacuum-induced crystallization in XI crystallization buffer  yielded the microcrystals.Xylose isomerase crystals were obtained from Hampton Research (HR7-102) and dissolved in water. Redissolved crystals were concentrated to 80 mg ml2.2.https://www.embl-hamburg.de/services/mx/P14_EH2/). For sample exchange the chips were mounted on the translation-stage holders as described below and connected to the SmarAct translation stages as described previously . The compartment area is lined with fiducial marks etched into the silicon between the junctions of the outer compartments at a pitch of 4350 µm. The nominal dimensions of the fiducial marks are 86 µm on the top and 20 µm on the bottom and they are situated 850 µm from the centre of the adjacent aperture. Detailed CAD drawings of the HARE chip, which can be used for mask development, can be found in the supporting information .The HARE chip is divided into 6 × 6 compartments (A1–F6) of 3.6 mm × 3.6 mm separated by 900 µm gaps in each direction. For best implementation of the HARE approach, each compartment is subdivided into 24 × 24 crystal wells, resulting in 20 736 crystal wells per chip. This provides a high composite number of crystal wells per row (24), allowing maximum flexibility in specific delay times that can be addressed without any other modification of the experimental parameters, without making the chip too fragile (36 wells per row is too many) (Schulz 3.1.2.a) The production process sequence starts with two layers of monocrystalline silicon in [100] orientation. A 50 µm-thick top layer and 450 µm-thick bottom layer are separated by a thin (300 nm) layer of thermally grown SiO2  [Fig. 2a)]. (b) A thermal oxide of 250 nm is grown on the wafers which acts as the hard mask for the etching in the following steps [Fig. 2b)]. (c) Photoresist is spun on the oxidized wafers on both sides to structure the hard mask [Fig. 2c)]. (d) The photoresist layer is exposed through a glass mask with UV light and the exposed regions on the wafer are then removed in a photoresist development step [Fig. 2d)]. (e) The oxide is wet chemically removed from the wafers in the open regions of the photoresist [Fig. 2e)] and (f) the photoresist layer is stripped [Fig. 2f)]. (g) The wafers with the structured oxide layer are then immersed in a wet chemical bath with TMAH (tetramethyl ammonium hydroxide). The exposed silicon is dissolved while the oxide acts as a hard mask for the etching and the BOX acts as an etch stop for the process [Fig. 2g)]. In this way the etch process is self-limiting and the etching of both sides can be done in one single step. The lateral etch rate is suppressed by the anisotropic etch rates of the TMAH solution. (h) As a last step, the hard mask including the BOX is removed by immersion into hydrofluoric acid [Fig. 2h)] after which the chips can be cut out of the wafer.The HARE chips are manufactured from conventional double side polished silicon-on-insulator wafers and are produced as follows. .Crystal dehydration can also be prevented by humidified environment (Sanchez-Weatherby 3.3.3.3.1.w/v) TERGAZYME (Sigma–Aldrich) or a 1 M HCl solution for 1 h. Subsequently, the chips are rinsed under running hot tap water and then washed with ddH2O. Finally, the chips are washed with isopropanol before being left to dry on the bench for a few hours. To speed up drying, chips can be placed in a custom-built drying station, attached to the pressured air or nitrogen gas feed usually found at synchrotron beamlines, and exposed to pressurized gas for ∼5 min [Fig. 4a)]. The information needed to print the drying station using a 3D printer is available in supporting material 4.The simplest method to clean the chips is extensive incubation in water, which dissolves most residues over time. However, the chips can be more effectively cleaned after each data collection using the following protocol. Directly after use the chips are incubated in a solution of 1%(2O2(30% w/w):H2SO4 in a ratio of 1:2.5 at 100°C], which removes all organic contaminants and leaves the chip surface hydrophilic. We note that piranha solution is extremely caustic and should only be used with appropriate safety precautions and personal protective equipment.Should they be contaminated with stubborn residue, chips can be comprehensively cleaned using hot piranha solution [H3.3.2.supporting material 5.Alteration of the surface charge allows for a more even wetting of the chips, improving crystal-loading homogeneity. As an alternative to chemical treatments, a convenient solution to generate hydrophilic surfaces is glow discharging or plasma cleaning. For a simple surface treatment of the whole chip, individual chips can simply be placed in a glow discharger  one at a time. Chips are then glow discharged for 60 s at 35 mA. To allow a selective wetting of the crystal-well containing compartments on the chip we designed a custom chamber which can hold four chips covered with laser-cut Kapton masks. These masks selectively cover all non-compartment areas of the chips. Closing the chamber presses the Kapton mask onto the chip. The selective wetting of the compartment areas leads to a more homogeneous liquid-layer distribution on the compartments, while the bars between compartments are more hydrophobic and repel the crystal slurry Figs. 4 and 5 ▸.3.3.3.i.e. Agar Scientific Ltd, UK). However, we found that to streamline handling during beam time, increase chip longevity, and safeguard the delicate chips during transport and storage, a custom storage box is extremely helpful. In our design, as in a microscope ‘slide box’, the chips are stored vertically in three arrays of 19 chips each. In comparison with membrane boxes, this minimizes storage space and allows for safe and convenient transport of a large number of chips to the beamline  . This is problematic for TR-SSX as it makes the degree of reaction initiation by either light or on-chip mixing harder to assess.In order to prevent under- or over-loading of the chip the concentration of the protein microcrystals should be assessed. We note that, for the chips described here, crystals should be ≥10 µm to avoid them flowing straight through the chip holes. A convenient and cost-effective way to quickly assess the concentration of protein microcrystal solutions during beam times is to use a simple benchtop light microscope and a Neubauer cell-counting chamber  into which 10 µl of crystal slurry are deposited. At 10×–40× magnification it is possible to not only assess the concentration of the protein microcrystals but also to assess the homogeneity of the crystal size distribution Fig. 8. In our 3.5.2.et al., 2018et al., 2019et al., 2019etc.) the vacuum level dictates how well the samples drain into the crystal wells. If the vacuum level is set too high, the crystals will break or dehydrate and thereby reduce the diffraction hit rate. Therefore, the chip-loading device allows careful optimization of loading parameters for each system, enabling effective loading and preventing dehydration and damage of crystals , vacuum levels between 0.3 and 0.5 bar below atmospheric pressure resulted in efficient loading with no apparent crystal damage, while stronger vacuum levels resulted in visible damage to the crystals and in split diffraction spots  a UV laser pulse has to have a 50 µm diameter, assuming optimal beam alignment and a typical Gaussian profile. In practice, laser beam diameters are determined by the optics used and wavelength, but typically lie between 50 µm and 100 µm (FWHM) , enabling a greater amount of data to be collected per chip  to reproduce the HARE chip and the accessory material. . DOI:


Composite structures have been widely utilized to improve material performance. Here we report a semiconductor-metal hybrid structure (CuO/Ag) for CO oxidation that possesses very promising activity. Our first-principles calculations demonstrate that the significant improvement in this system’s catalytic performance mainly comes from the polarized charge injection that results from the Schottky barrier formed at the CuO/Ag interface due to the work function differential there. Moreover, we propose a synergistic mechanism underlying the recovery process of this catalyst, which could significantly promote the recovery of oxygen vacancy created via the M-vK mechanism. These findings provide a new strategy for designing high performance heterogeneous catalysts. Their high catalytic activity is normally ascribed to the characteristically high charge transfer efficiency between adsorbed gas molecules and the catalyst surface, a quality which is highly dependent on the surface charge density of active sites5. However, precise control of charge location at the atomic level still remains a challenge6. Although tremendous attempts have been made, using methods such as shape-controlling7, chemical doping8, crystal-plane-controlling9, use of an external electric field10, etc., it still remains a considerable challenge to precisely tailor the surface charge state of these solid materials so as to enhance both activity and stability simultaneously12. The fundamental problem is the limited understanding of the nature of surface polarization, a barrier which seriously hinders the further practical application of heterogeneous catalysts.Heterogeneous catalysts of various reactions such as CO oxidation have attracted widespread attention due to their high efficiency, benign environmental impact, and durability13. As shown in Fig. 2O(111) surface was first deposited on a flat Ag(111) substrate, tailored to expose a Cu2O(100) surface. Following this first treatment cycle, the exposed Cu2O(100) surface was then oxidized to form a layer of CuO(100) surface. Compared with previous reported CuO based catalysts18, our design achieves complete conversion of CO at relatively low temperature (150 °C), which performs better than bare CuO/Cu2O14 or CuO15  layers have the same lattice constant with Cu2O surface (a = b = 4.27 Å), which is different from normal CuO according to previous reports14. Such a unique structure may induce an interfacial polarized charge which would then penetrate the CuO(100) surface owing to the very thin atomic thickness along the interfacial line. Our previous results demonstrated that this polarized charge along the interfacial line enables controllable highly efficient CO oxidation with good durability. By maximizing the Ag/CuO interfacial length, the CO conversion rate can be linearly enhanced.To acquire deeper insight into surface polarization, recently we designed a novel semiconductor-metal interfacial structure (CuO/Ag) for CO oxidationHowever, there are still several unsolved problems requiring clarification, such as elucidating the formation process of interfacial polarization, the mechanism of transfer of polarized charge at the interface, and how polarized charge impacts catalyst activity. To gain deeper insight into these inner mechanisms, we now extend our study of this system, so as to provide more confidence and experience in designing new composite functional materials.19. The frozen-core all-electron projector augmented wave (PAW) model20 with Perdew-Burke-Ernzerhof (PBE) functionals21 was employed to describe the interactions between core and electrons. An energy cutoff of 450 eV was used for the plane-wave expansion of the electronic wave function. The force convergence criterion was set to 0.01 eV/Å for pure Ag and Cu2O/CuO unit structures and 0.05 eV/Å for the composite CuO/Ag model, respectively. The energy convergence criterion was set to 10−5 eV. A k-point grid with a 7 × 7 × 1 Monkhorst–Pack mesh for pure Cu2O/CuO and Ag (100) and (111) facets was chosen for sampling the Brillouin zone, while for the CuO/Ag structure, due to the huge size of the model, only Gamma point was performed. A Bader analysis of charge density of atoms was used to analyze the charge distribution22, and the nudged elastic band (NEB) method23 was employed to search the transition states of elementary steps during the CO oxidation reaction.All the calculations were performed at the spin-polarized density functional (DFT) level of theory as implemented in the Vienna ab initio Simulation package (VASP)2O/CuO(100) 6 × 3 supercell structure (the CuO surface was evolved from the Cu2O(100) surface) in Fig. 2O(100), 12 layers of CuO, and 4 transition layers in between14, with the structure being cleaved along the direction of the scissors as indicated. As Cu2O is a cubic lattice, we assume that the CuO lattice evolved from it shares the same cubic geometry, thus we set the cleavage angle to 35.26° and (100) surfaces are minimally influenced by Cu2O. Therefore our final simplified CuO/Ag model does not contain Cu2O. During relaxation all the atoms were relaxed except for the bottom 2 layers.The Cu24, a Schottky barrier should be formed at the metal-semiconductor junction due to the difference in work function between the two materials25. In our case, because the work functions of Ag(111) and CuO(111) differ, surface polarization is inevitably induced at the CuO/Ag interface, as shown in Fig. interfacial polarization.According to the Schottky-Mott rule−. To further elucidate the electrostatic potential difference at the interface, the static potential surface along the direction perpendicular to the interface is given in Fig. 2 molecule and clean unoxidized Ag surface. The real O2/Ag interactions could be far more complicated due to the formation of the epitaxial Ag2O thin oxide overlayers on the surface of Ag26, which could affect the overall catalytic activities. To estimate its impact, we artificially added an Ag3O cluster to mimic the Ag2O layer in the middle between CuO and Ag as shown in Fig. −, indicating that the oxidation layer could reduce the catalytic activity to some extent. Therefore, we strongly suggest that the surface oxidation layer of Ag should be removed first when preparing the CuO/Ag catalyst experimentally.Such a charge polarization is shown in Fig. 27. As shown in Fig. 2O bulk structures are semiconductors according to previous reports14.After the interfacial polarization forms, a new question arises: how do electrons transfer inside the CuO lattice? As work function is defined as the thermodynamic work required to remove an electron from a material to the vacuum near its surface, a crystal facet with a higher work function should be more likely to accumulate electrons29. The CuO(100) surface is terminated by two-coordinated oxygen (O2c) atoms, which can easily combine with CO to form a CO3 surface intermediate. The CO3 intermediate easily decomposes to produce CO2, leaving an oxygen vacancy (Vo) at the active site. In order to evaluate the role of Ag in the CO oxidation, we built a new model CuO* by intentionally removing the Ag substrate from the CuO/Ag model as shown in Fig. 29, we compared our result (0.24 eV) with the barriers of rate-determining steps in other relevant systems, such as CO oxidation on Cu2O (111) (1.63 eV)30, CuO/Cu2O (110) (0.60 eV)31, CuO/Cu2O (100) (1.15 eV)14, and CuO/Cu2O (111) (0.37 eV)14, illustrating the superiority of our CuO/Ag model.Having computed the polarized charge accumulation on the CuO(100) surface, we now move on to study its impact on catalytic activity. CO oxidation on the CuO surface follows the Mars-Van Krevelen (M-vK) mechanism33. Our results show that the CO adsorption energy in the CuO* system is 3.19 eV while in CuO/Ag it is 2.94 eV, indicating that the polarized charge drags the adsorption energy down closer to an optimum value, thereby enhancing catalytic activity.To further understand the enhancement of catalytic activity, we then calculated the CO adsorption energy on a CuO(100) surface in both the CuO* and CuO/Ag models. Normally, optimum activity is obtained at an intermediate adsorption energy according to the volcano shape ruleIn addition, the effect of polarization charge on the lattice oxygen reactivity of CuO(100) was also carefully studied. In the presence of the Ag substrate, the activation energy of a lattice oxygen decreases from 5.34 eV to 5.11 eV Fig. , and theVO is vital in order to enable the next reaction cycle. Our adsorption energy calculations (Table 2 adsorption energy at the oxygen vacancy site on the CuO(111) surface increases from 1.45 eV to 1.64 eV, which should result in a significant improvement in catalytic recovery. CO cannot compete with O2 during the recovery process due to its much lower adsorption energy. Also, because O2 adsorption on Ag is weak, O2 prefers adsorption at the VO site and thus readily heals the defect there. Such a feature agrees well with our previous experimental work13.After CO oxidation, the refilling of ns Table  indicate2 has reoccupied the oxygen vacancy, there should be one additional oxygen atom bound to the CuO(100) surface. This redundant oxygen atom can be much more active than a lattice oxygen14. We calculated the desorption activation energy by removing a redundant oxygen atom gradually from the CuO(100) surface in both CuO* and CuO/Ag models as shown in Fig. After O+) has stronger adsorption energies for both O2 and CO  as mentioned above. Compared with neutral Ag, Ag with a hole  and CuO/Ag (red lines) are summarized as shown in Fig. VO site, making the catalyst more efficient.Keeping these adsorption energies in mind Table , we propion Fig.  shows a VO defect, making the catalyst more durable. These new insights from the case of CuO/Ag can be helpful for studying how the polarization effect may improve other catalytic systems, providing a new approach toward the rational design of more efficient heterogeneous catalysts.In summary, by utilizing an interfacial polarization effect, we have designed a CuO/Ag heterogeneous catalyst for CO oxidation with very promising catalytic activity. The significant improvement in catalytic performance is ascribed to the accumulation of polarization charge on the CuO surface that is induced by the work function difference between CuO and Ag. Interestingly, charge polarization can not only promote CO oxidation, but also promote the healing of the Supporting Information.


Patients with head and neck squamous cell carcinoma (HNSCC) can develop lung squamous cell carcinoma (LuSCC), which could be the second primary tumor or HNSCC metastasis. Morphologically it is difficult to distinguish metastatic HNSCC from a second primary tumor which presents a significant diagnostic challenge. Differentiation of those two malignancies is important because the recommended treatments for metastatic HNSCC and primary LuSCC differ significantly. We investigated if the quantification of the promotor methylation status in HNSCC and LuSCC differs.N = 36) and LuSCC (N = 17) were included in this study. Methylation status in the ASC/TMS1/PYCARD  and MyD88  promoters was analyzed. Bisulfite converted DNA, isolated from tumor tissue was quantified using pyrosequencing. Results of pyrosequencing analysis were expressed as a percentage for each tested CpG site. Receiver-operating characteristic (ROC) curve analysis was used for the evaluation of the diagnostic properties of selected biomarkers.Primary HNSCC (p ≤ 0.0001). By performing ROC curve analysis we showed that corresponding areas under the curve (AUC) were 85–95%, indicating that selected CpG sites are useful for a distinction between primary LuSCC and primary HNSCC.CpG sites located in the promoters of ASC/TMS1/PYCARD_CpG8 (− 65 upstream) and MyD88_CpG4 (− 278 upstream) are significantly hypermethylated in the HNSCC when compared with LuSCC (Results of the present study indicate that there is a significant difference in the methylation status of tested genes between primary HNSCC and LuSCC. However, to prove this approach as a useful tool for distinguishing second primary LuSCC from HNSCC metastasis, it would be necessary to include a larger number of samples, and most importantly, metastatic samples. Patients diagnosed with head and neck squamous cell carcinoma (HNSCC) can present with lung metastases, or develop second primary lung tumors . The occASC/TMS1/PYCARD (apoptosis-associated speck-like protein containing a caspase recruitment domain) and MyD88 (Myeloid differentiation primary response protein 88), could be considered as a promising biomarker in non-small cell lung cancer (NSCLC) in distinguishing healthy from tumor tissue [ASC/TMS1/PYCARD gene was associated with reduced overall survival.In our previous work, we have shown that the methylation status of two genes, r tissue . We founThe fact that there is no fast and reliable diagnostic protocol for distinguishing between second primary LuSCC from HNSCC metastasis to the lung, motivated us to use primary tumor tissues (LuSCC and HNSCC) and investigate if they can be distinguished on methylation level of selected genes. We were using a pyrosequencing approach, still considered a gold standard in methylation study .All the protocols and molecular methods used in this study have been described in detail in our previously published data .N = 17) and head and neck squamous cell carcinoma (HNSCC) (N = 36) were obtained during surgery . Tissue samples were snap-frozen in liquid nitrogen and stored at − 80 °C. Cancer tissues were analyzed by pathologists on hematoxylin and eosin stained slides, and samples with a minimum of 70% of tumor cells were preceded for further analysis. The clinical and pathological features of the patients are shown in Table This study was conducted on freshly frozen primary tumor samples. Early-stage resected squamous cell carcinoma of the lung (LuSCC) N = 7 and heaMyD88 and ASC/TMS1/PYCARD genes in the primary HNSCC and compared them with methylation status in the primary LuSCC samples obtained in the previously mentioned study. We have analyzed 8 different CpG sites in the promoter region of the ASC/TMS1/PYCARD gene and 10 different CpG sites in the MyD88 gene. Locations of the tested CpG sites in the ASC/TMS1/PYCARD gene are as follows: CpG1 + 52; CpG2 + 49; CpG3 + 33; CpG4–134; CpG5–129; CpG6–76; CpG7–71; CpG8–65. Locations of the tested CpG sites in the MyD88 gene are as follows: CpG1–253; CpG2–256; CpG3–267; CpG4–278; CpG5–210; CpG6–216; CpG7–222; CpG8–146; CpG9–151; CpG10–167. Indicated values represent a distance from the transcriptional start site. Schematic locations of the tested CpG sites and tested genes are presented in the Fig. DNA, isolated from freshly frozen tumor tissue samples, was bisulfite converted, amplified, and sequenced. For this study, we have analyzed the methylation status of the ASC/TMS1/PYCARD_CpG8 and MyD88_CpG4), detected for each sample in HNSCC and LuSCC, were plotted.Differences in methylation status were tested using a paired t-test. To define the possible diagnostic applicability of selected CpG sites, receiver operating characteristic (ROC) curves analysis was performed. To assess the diagnostic properties of selected CpG sites, ROC curves, the areas under curves (AUC) together with 95% confidence intervals (95% CI) were plotted. For this analysis values of methylation status (%) of selected CpG sites (ASC/TMS1/PYCARD and MyD88 promoters is different between primary tumors in HNSCC and LuSCC sample cohorts. We have shown that ASC/TMS1/PYCARD_CpG8 (located − 65 upstream) is significantly hypermethylated in primary HNSCC (11.47%) in comparison to LuSCC (4.65%) (p < 0.0001)  is significantly hypermethylated in HNSCC (19.96%) when compared to LuSCC (10.2%) (p < 0.0001) , and the results were the same as presented for the whole cohort. The third, and probably the most important limitation, is that we did not compare lung metastases of HNSCC with primary LuSCC and primary HNSCC. To further define tested loci as potential diagnostic markers, this is necessary.In conclusion, the methylation status of selected genes indicated that it is different in primary tumor tissues of different origins. Further analyses, on a larger number of the samples including metastasis, are needed to confirm its utility for a distinction between second primary LuSCC and metastatic HNSCC in the lung. Results of this pilot study must be considered as preliminary and additional independent studies are necessary to confirm the diagnostic utility of the tested loci/approach.