Staphylococcus aureus is the most prevalent cystic fibrosis (CF) pathogen. Several phenotypes are associated with worsened CF clinical outcomes including methicillin-resistance and small-colony-variants. The inoculum effect (IE) is characterized by reduced β-lactam susceptibility when assessed at high inoculum. The IE associates with worse outcomes in bacteremia and other high-density infections, and may therefore be relevant to CF. The prevalence of IE amongst a CF cohort (age ≥18 years), followed from 2013 to 2016, was investigated. Yearly methicillin-sensitive S. aureus (MSSA) isolates were screened at standard (5 × 105 CFU/mL) and high (5 × 107 CFU/mL) inoculum against narrow-spectrum anti-Staphylococcal β-lactams and those with anti-pseudomonal activity common to CF. A ≥ 4-fold increase in minimum inhibitory concentration between standard and high inoculum defined IE. Isolates underwent blaZ sequencing and genotyping and were compared against published genomes. Fifty-six percent (99/177) of individuals had MSSA infection. MSSA was observed at ≥105 CFU/mL in 44.8% of entry sputum samples. The prevalence of the IE was 25.0%-cefazolin; 13.5%-cloxacillin; 0%-meropenem; 1.0%-cefepime; 5.2%-ceftazidime; and 34.4%-piperacillin-tazobactam amongst baseline MSSA isolates assessed. blaZ A associated with cefazolin IE (P = 0.0011), whereas blaZ C associated with piperacillin-tazobactam IE (P < 0.0001). Baseline demographics did not reveal specific risk factors for IE-associated infections, nor were long-term outcomes different. Herein, we observed the IE in CF-derived MSSA disproportionally for cefazolin and piperacillin-tazobactam and this phenotype strongly associated with underlying blaZ genotype. The confirmation of CF being a high density infection, and the identification of high prevalence of MSSA with IE in CF supports the need for prospective pulmonary exacerbation treatment studies to understand the impact of this phenotype. Staphylococcus aureus (SA) is the most prevalent cystic fibrosis (CF) pathogen isolated from >50% of persons with CF (pwCF) (S. aureus isolates with specific phenotypes including methicillin-resistant S. aureus (MRSA) (F (pwCF) , 2. ThisF (pwCF) , 3. Impos (MRSA)  and smalS. aureus (MSSA) is the potential to have reduced susceptibility to β-lactam antibiotics when present at high concentrations/inoculums (inoculum effect” (IE) is predominately attributable to the production and accumulation of a β-lactamase, BlaZ. BlaZ is well-known to hydrolyze penicillin, but can also exert weak hydrolytic activity on a broad range of β-lactams when abundantly present, reducing efficacy  ≥18 years and older. This is the fourth-largest adult CF clinic in Canada. All sputum-derived pathogens from pwCF attending the CACFC are maintained in a prospectively inventoried biobank  is a regional program encompassing all of Southern Alberta, Canada, caring for pwCF ( biobank . We incl5 CFU/mL) and high inoculum  and were used to define IEs (Supplement).The first MSSA per pwCF between 2013 and 2016 was tested for susceptibility with anti-Staphylococcal β-lactams; cefazolin and cloxacillin, and those administered in the management of CF pulmonary exacerbations ; cefepime, ceftazidime, meropenem, and piperacillin-tazobactam. Subsequent yearly isolates were tested only with cefazolin, meropenem, and piperacillin-tazobactam, following observed trends. Minimum inhibitory concentrations (MICs) were reported for each isolate at standard inoculum  versus HI (107 CFU/mL) (An isolate with ≥4 fold difference in MIC at SI (10 CFU/mL) , 15.An isolate susceptible at SI but non-susceptible at HI, using 2009 Clinical and Laboratory Standard Institute (CLSI) breakpoints , 29.blaZ identification via Sanger sequencing. All MSSA isolates were strain-typed by pulsed-field gel electrophoresis, and multilocus sequence type (MLST) was inferred (Supplement). The comprehensive antibiotic resistance database   , and BLAC, and D , 19, 31 blaZ allele types in our cohort were broadly representative, we compared our data with that obtained from other cohorts. We performed analysis of blaZ types from published non-CF cohorts where blaZ alleles were reported, and from published studies including CF-derived MSSA using available de novo assemblies via custom Python scripts (Supplement).To understand if S. aureus, and all co-infections were recorded. Persistent MSSA was defined as an isolate being recovered during a year in which ≥ 50% of sputum cultures were MSSA+. Participant status was defined on 31 December 2016 (or at study exit) as “ongoing follow-up,” “moved/transferred,” “lung transplantation,” or “death.” The primary clinical outcome of interest was the rate of lung function decline  were do decline ), with st-test. For each comparison, p-values were reported with odds ratios (OR) or risk ratios (RR) and 95% confidence intervals (CI), when significant. An alpha of 0.05 was used for significance in all data sets including <20 comparisons. Alpha was Bonferroni-corrected (0.05/number of comparisons) and noted when relevant for data sets with ≥20 comparisons. Assessing the similarity of our data to published cohorts was performed using two-proportion Z-score tests, while MSSA and Pseudomonas aeruginosa density comparisons were performed via the Spearman rank correlation test. Rates of FEV1 and BMI decline were calculated by dividing the sum of all values by each patient’s time in the study. Patients with only one FEV1 or BMI value were excluded from decline analyses. Statistical tests were performed using STATA version 17.0 .Fisher’s exact probability tests were performed for comparisons of categorical variables (1-sided). The normality of distribution was assessed using the Shapiro-Wilk test. Non-normally distributed variables were reported as medians with interquartile ranges (IQR) and compared via the Mann-Whitney rank-sum test. Normally distributed variables were reported as means with standard deviation (SD) and compared using the P = 0.9828) or FEV1 (P = 0.7630) and had only one isolate each. Amongst the 96 participants, 16 left the study prematurely; 14 transferred to another clinic, one received lung transplantation, and one died. The median age at entry was 25.4 years (IQR: 21.3–34.8) and 51.0% (49/96) were male. Eighty-four percent (81/96) had ≥1 F508del mutation. Median predicted FEV1 was 71.0% (IQR: 51.5–86.0) and FVC, 91.5% (IQR: 75.5–103) (Table E4).Between 2013 and 2016, 177 pwCF met study criteria and submitted 2,045 sputum samples. MSSA was cultured in 38.3%  of samples, in 99 (55.9%) pwCF. Participants had a mean 15.1 submitted cultures (SD: 13.2), with a mean of 4.3/year (SD: 0.46). MSSA was recovered from the CACFC biobank in 96/99 of these individuals. The three patients not included did not differ with respect to age (5 CFU/mL (IQR: 104–106CFU/mL) amongst first isolates. Importantly, MSSA exceeded ≥105 CFU/mL in 44.8% (43/96) of initial samples. Those with persistent MSSA infection had higher mean sputum density than those without . Of patients infected with MSSA, 15.6% were co-infected with MRSA, 1.0% with SCV+ MSSA, and 67.7% with P. aeruginosa.In their first year, 62.5% of pwCF (60/96) had persistent MSSA infection (Table E5). Not all individuals had MSSA-positive samples each year—the median yearly isolates/participant was 2 (IQR: 1–3), for a total of 223 isolates. All 96 participants had a first yearly isolate (2013–2016), 70-a second (2014–2016), 40-a third (2015–2016), and 17-a fourth (2016) included. Median MSSA density was 10n = 96). Piperacillin-tazobactam showed the highest prevalence of IEs, followed by cefazolin. As expected, the IE was infrequent with cloxacillin, cefepime and ceftazidime, and absent with meropenem. The pIE was less common than the IE and was predominantly observed with piperacillin-tazobactam. The occurrence of IE or pIE with one antibiotic did not associate with an increased likelihood of IE/pIE with other β-lactam antibiotics (data not shown). Individuals with persistent MSSA were not more likely to have an IE to any antibiotics tested, nor was higher MSSA density associated with any IE (data not shown). As IEs were more common than pIEs, these were the primary focus of further analyses.Susceptibility testing at SI and HI for cefazolin, cefepime, meropenem, ceftazidime, piperacillin-tazobactam, and cloxacillin are outlined in n = 127) were tested for susceptibility to cefazolin and piperacillin-tazobactam, owing to higher IE prevalence with these drugs, while using meropenem as a negative control. IEs were again most common with piperacillin-tazobactam; 42.6% , while 38.6% (86/223) showed pIE. Cefazolin IE prevalence was 22.0% (49/223), and pIE 3.6% (8/223). Consistently, meropenem showed no IEs. Isolates with the cefazolin or piperacillin-tazobactam IE were more likely to also display the corresponding pIE . There was an evident rightward shift in the MIC of all three antibiotics at HI relative to SI (The remaining yearly isolates (ve to SI .n = 96) were subsequently assessed for presence and allele-type of blaZ. Seventy-three (76.0%) were blaZ positive, with blaZ C being most common  and blaZ A associated with isolates having cefazolin IE  (Table E6). Isolates lacking blaZ were less likely to have IEs overall (data not shown).Initial isolates , and a lower proportion of blaZ D (Z = −5.87). These trends did not differ when the USA study alone was considered  . Nine were excluded; fragmented genome (n = 4), multiple blaZ alleles (n = 4), both fragmented and unassignable (n = 1). Overall, CF-published genomes had a slightly lower blaZ prevalence compared to our cohort (Z = −2.63), owing to decreased blaZ D (Z = −3.61). Prevalence of types A, B, and C did not differ  . We also the USA , 35 and  the USA , met crit differ .blaZ A .To establish if IE is intrinsic to individual strains, we examined strain types (MLST) amongst our total 223 MSSA. The most common strain types corresponded to ST-5 (18.4%), ST-15 (15.2%), and ST-30 (14.3%) (Table E7). ST-30 isolates were more likely to have P < 0.001). Likewise, individuals without initial piperacillin-tazobactam IE were more likely to have their second ST-matched strain also negative . The most common ST amongst those isolates that preserved the piperacillin-tazobactam IE was ST-15 (7/18) (Fifty-eight percent (56/96) of pwCF had an ST at baseline that reappeared at least once during follow-up. These STs were most commonly ST-5 (23.2%), ST-15 (16.1%), ST-30 (14.3%), and ST-1 (10.7%). Eighteen of the 56 (32.1%) had isolates that showed piperacillin-tazobactam IE at entry. Such patients were more likely to have their second MSSA of the same ST also be piperacillin-tazobactam IE positive  . Such trNo baseline clinical characteristics were identified associating with the risk of MSSA with cefazolin or piperacillin-tazobactam IE (Table E8). Based on the retrospective nature of this study as well as the fact that IE testing was not performed in real-time (only on representative yearly isolates), we were limited in assessing the most relevant CF outcome— individual PEx events (pulmonary function recovery and reduction in MSSA bioburden). During the four years of study, 122 exacerbations were documented in 38.5% of participants, with a median incidence of 2 PEx/participant . Exacerbations lasted for a median of 13 days (IQR: 12–15d), and median time between individual PEx events was 352 d (IQR: 107–634 d). Twenty-four exacerbations (19.7%) were treated with cefazolin in 12 pwCF and four (3.3%) were treated with piperacillin-tazobactam in four pwCF. Owing to these limitations, we primarily focused on exploring long-term outcomes. Our primary outcome—rate of lung function decline over four years, did not differ based on either cefazolin or piperacillin-tazobactam IE. One patient died (end-stage lung disease), and one was transplanted, independent of IE.−2 versus 0.2 kg/m2/year, and −7.6 × 10−2 versus 0.2 kg/m2/year, respectively) throughout the study duration. No differences in BMI change were observed between those treated with cefazolin, or piperacillin-tazobactam for their PEx versus not (data not shown). However, patients having at least one piperacillin-tazobactam IE positive strain were more likely to have P. aeruginosa isolated from their sputum during their time in the cohort , though this was not noticed with cefazolin . P. aeruginosa density was not associated with an increased MSSA density amongst first isolates .The rate of BMI decline was greater for patients having at least one cefazolin or piperacillin-tazobactam IE +strain  was confirmation of IE persistence over time within serial isolates from patients infected with the same strain. Amongst pwCF whose strains were preserved, isolates with initial piperacillin-tazobactam IE were nearly 10X more likely to retain this phenotype. Such findings further suggest a strain-dependent nature underlying the IE. Persistence of the cefazolin IE over time was not confirmed , though associations with th ST-15 — furtherblaZ allelic distributions in our cohort to published studies involving general non-CF and CF-derived MSSA from other parts of the world. Relative to non-CF cohorts, only minor differences in blaZ B and D were observed. As neither of these are associated with cefazolin nor piperacillin-tazobactam IE, these differences are thought to be negligible. Overall, we demonstrated only modest differences between ours, and CF cohorts from the USA  and modest number of exacerbations experienced in this cohort, we did not attempt an exploratory analysis including PEx outcomes. To best assess the impact of IE/pIE on CF-related outcomes, a properly powered prospective pulmonary exacerbation treatment study is required. Such a study could assess for differential bacterial killing and lung function recovery in the context of PEx based on whether an individual was treated with an antibiotic to which their MSSA had inoculum-related reduced susceptibilities. The importance of study design is well-illustrated by early work on MRSA. Initial investigations to assess for impacts of MRSA on CF outcomes failed to observe differences in the rate of FEVcontrols , while lcontrols . Subsequseverity , and greseverity .P. aeruginosa S. aureus is now the most prevalent CF pathogen and other phenotypes have significant impacts on CF-outcomes, our study serves as a call-to-action to begin further CF-IE/pIE investigations, given their prevalence.In this first study of the IE/pIE in CF, we have established that MSSA within CF airways exists at high-density, and that many strains display the phenotype of inoculum-related reduced susceptibility—particularly to cefazolin and piperacillin-tazobactam. As


Designing and synthesizing materials with smart hydrophobicity against an external magnetic field for efficient oil/water separation is of great importance due to the increasing problems caused by oil pollution. Here, the nanocomposites were fabricated based on graphene and different iron oxides exhibit smart hydrophobicity against an external magnetic field and they are in powder form eliminating the requirement for a substrate employing a facile and echo friendly method. The results prove that autoclaving of graphene leads to its ferromagnetic property; then it is attached to iron oxides by magnetic attraction and a nanocomposite is produced. The magnetic property of the resulting nanocomposite is higher than the magnetic property of its individual components. In addition, following nanocomposite formation, its hydrophobicity and surface area also change. FESEM images were taken from the nanocomposites to study their surface morphology, and EDS-MAP analysis to observe the elemental distribution uniformity of the nanocomposites. Also, to measure the surface area and pore size, BET analysis has been performed on pure materials and graphene-black iron oxide nanocomposite (graphene@black iron oxide). The results show that the specific surface area of black iron oxide increases after being composited with graphene dispersed at 5000 rpm. Indeed, graphene forms a composite by binding to iron oxide, and therefore, its specific surface area increases compared to iron oxide and graphene alone. These results show an increase in oil sorption and better separation of oil from water by the prepared nanocomposite. Also, to measure the magnetic properties of pure materials, graphene@black iron oxide, and ferromagnetic graphene at 3000 and 5000 rpm, the Vibrating Sample Magnetometer analysis has been performed. The results have proven that the nanocomposite powder prepared by a simple method obtained from cost-effective and available materials is hydrophobic and becomes more hydrophobic by applying an external magnetic field. Due to the ease with which oil can be readily removed from the nanocomposite by eliminating the external magnetic field, this nanocomposite is an excellent choice for the separation of oil from water. These spills can come from a variety of places, including oil rigs, carriers, and undersea pipelines. Studies have suggested a range of ways for cleaning up oil spills, including absorption2, dispersants3, solidifiers4, and controlled burning5.  Due to its high efficiency, ease of use, and versatility in combining with other techniques, absorption is one of the most often employed techniques separation and recovery of split oil from water.Aquatic ecosystems have suffered negative consequences as a result of oily wastewaters, petroleum industry spills, and shipping mishaps6, layer by layer deposition8, plasma etching9, chemical vapor deposition (CVD)10, template-based extrusion11, sol–gel process12, electrospinning13 self-assembly14 and extrusion15.Up to now, all methods that have used the phenomenon of hydrophobicity or lipophilicity to separate water from oil have been based on the fabrication of surfaces that have used surface engineering to create roughness and reduce the energy of these surfaces. Unfortunately, the methods used to create roughness and cover the surface with hydrophobic compounds suffer from being costly and time-consuming. These fabrication methods of superhydrophobic surfaces include lithographic methods16, alumina17, zeolites18, silica19, polypropylene20, and polystyrene21 are a few examples of these absorbents. Some of the limitations of such sorbents that lessen their absorption efficacy are their low sorption capacities, low recovery efficiencies, generation of toxic wastes, and weak buoyant qualities. It is crucial to provide lightweight materials with environmental friendliness, high sorption capacity, and chemical inertness for oil spill response activities. Surface wettability is among the most important factors in the oil–water separation process. One phase may be allowed to permeate while the other is repelled by such regulation of wettability22. Thus, superhydrophobic/superhydrophilic membranes and hydrophilic/underwater superhydrophobic membranes are the two main groups of materials or membranes used for water/oil separation. It is water permeable and oil-resistant. For mild oil/water separations, this kind is most effective. Nonetheless, applications for separations involving heavy oil/water are feasible when using materials and membranes with superhydrophobicity. If light oil/water mixes are separated one way alone, it is claimed that light oil molecules will become caught between the water layer and the water-resistant membranes. Therefore, the need for inverse wettability membranes was emphasized as being essential for effective and continuous oil/water separation22. Such membranes were made by coating various materials, such as metal oxides, sponges, polymers, metal meshes, and many more.Some of the distinguishing characteristics that oil absorbents should have are high oil sorption capacity, quick oil sorption rate, low density, low cost, and reusability. So far, a wide range of advanced and innovative adsorbents with hydrophobic and oleophilic surfaces have been synthesized to separate and recover oil from water. Fiber sheets26. The word “composite” is frequently used to refer to materials whose characteristics, such as the elastic modulus, differ considerably from those of homogeneous materials due to the separation of different phases on a scale bigger than atoms. Composites have properties that cannot be represented by any of the ingredients alone. In other words, these components have distinctly dissimilar chemical or physical characteristics, and when combined, they produce a product with attributes distinct from those of the constituent parts. What distinguishes composites from mixtures and solid solutions is that the components of composites remain separate and distinct in the final structure on a macroscopic or microscopic scale. In addition to a vast array of other items, composites are used in pipelines, buildings, roads, bridges, and airplanes. To create composites that are robust, durable, affordable, and light, researchers are figuring out how to increase composites quality30.Solids called composites comprise two or more different constituents or phases on a scale greater than an atom. These materials are distinguishable from each other in macroscopic dimensions31. The first phase’s strength, electrical conductivity, resistance, magnetic characteristics, etc. are increased in the second phase by dispersing nanometer-scale particles as reinforcements or fillers. Types of nanocomposites include polymer-based nanocomposites, ceramic-based nanocomposites, and metal-based nanocomposites34. Among nanocomposites, the most attention is paid to polymer-based nanocomposites. For example, Nine et al. developed a method for the producing modified polymer-based nanocomposites by using graphene. These modified nanocomposites have strong mechanical strength and are self-cleaning, therefore can be used to improve superhydrophobic surfaces, which often suffer from mechanical instability after abrasion/scratching, for self-cleaning applications35.One of the disadvantages of composites is the concentration of stress at the boundary between their construction phases, which can lead to the destruction of the composite in that area. The structure of the composites has been modified by nanotechnology and the fabrication of nanocomposites. A nanocomposite is a composite in which one or more components are less than 100 nm in size. You can find two phases in nanocomposites. Typically, the nanocomposite’s foundation material or matrix, which can be comprised of a polymer, metal, or ceramic, has a crystalline structure in the first phase38. While a sheet of graphene weighs only 0.0077 g, it can tolerate up to four kilograms39. In other words, although it is thin and light, it is extremely strong. It also has a large surface, excellent thermal conductivity, and electricity and many incredible features. Scientists called graphene as “miracle substance” of the century and predicted that it can cause a huge revolution in various industries. Studies show that graphene has a myriad of unprecedented properties, any of which can be used to make extraordinary composites. The addition of graphene can improve bulk materials’ conductivity and strength and assistance in the development of composites of better ability. Additionally, it may be used to make conductive and heat-resistant composites by mixing it with ceramics, polymers, and metals42.Superhydrophobic composite coatings with high mechanical strength, self-cleaning, and other special qualities may be made using graphene, a two-dimensional matrix of carbon atoms in a honeycomb network44. In addition, magnetized graphene can also be used to prepare magnetic nanocomposites. In our previous work, we were able to magnetize graphene at room temperature with simple methods. Furthermore, we looked into how magnetic field, pressure, and temperature changes affected the ferromagnetic characteristics of graphene46. We can also use this graphene, which is magnetized by a simple and cheap method, to prepare smart magnetic nanocomposite.The magnetic and electrical characteristics of graphene can be enhanced by doping the surface with metal nanoparticles and other organic and inorganic species. The entire nanocomposite can be magnetized if these particles have magnetic characteristics49.Graphene-based composites can contain the element iron in its primary form, or core–shell nanoparticles, and or iron oxides. The presence of oxygen-containing groups on the graphene surface, the employment of reducing agents, and other factors affect the fraction of iron nanoforms in various stages of oxidation. Because epoxy, carboxyl, and hydroxyl functional groups are present, magnetic nanoparticles incorporating iron are deposited on graphene oxide sheetsDue to some problems in the absorption of oil and because the methods that have been presented so far all need a substrate for absorption, in this work, it has been tried to use smart superhydrophobic nanocomposite of graphene and iron oxide with high absorption capability and appropriate recyclability.50.The membrane’s hydrophobic characteristics, however, might result in the fouling phenomenon, which reduces the membrane’s lifespan. Hence, it is essential to modify the membrane, particularly to increase its hydrophilicity, in order to maximize its functionality. Due to its exceptional thinness and distinct layered structure, graphene and its derivatives have attracted the interest of numerous researchers working on membranes for water treatment during the past several years. These distinctive characteristics make it ideal for oil/water separation by increasing the penetration fluxes and enhancing the hydrophilic and physicochemical qualities51. This means that surface modification is necessary to increase the sponges’ hydrophobicity and decrease their water-wettability in order to create the requisite superoleophilic and superhydrophobic oil sorbent. Sponge surfaces are frequently covered with a variety of substances, such as polyaniline, polypyrrole, carbon nanotubes, graphene, nanofibers, and nanocrystals, to boost their superoleophilicity and superhydrophobicity52. The typical characteristics of carbon materials are high capacity, high porosity, mechanical integrity, and large pore volume53. Much emphasis has been given to the creation of carbon materials with elevated hydrophobicity, oleophilicity, functional qualities, and distinctive thermal properties54. The graphene-based sponge (GS), which is both hydrophobic and oleophilic, has a high capacity for absorbing oils and other organic liquids and is very recyclable. These excellent outcomes make the GS an excellent choice for use in oil and water separation55. Aerogel and hydrogel materials, as well as foams based on graphene, have all received significant interest in the subject of water/oil separation58. An efficient way to remove oils from water by employing graphene-coated cotton fibres has also been proposed, which can be used to separate all types of oils and organic solvents from water. The development of useful materials that can successfully separate oil from water seems important59.Some of the biggest difficulties in using graphene derivatives or their composite membranes for oil/water separation are linked to how well the nanoparticles decompose on the membrane surface. More study is still required to determine how the ratio of graphene derivative-based nanoparticles affects the membrane matrix. The greatest performance of membranes requires optimization of the concentration and distribution of the components based on graphene. Moreover, manufacturing the graphene composite membranes on a large scale presents the largest obstacle to a successful use. If these challenges are solved, successful use for water/oil separation and other water treatment applications may be possible. Studies suggest that because of their significant surface area, porous sponges that are three-dimensional (3D) have considerable promise as high-capacity sorbents. Nevertheless, the majority of sponges sold commercially are not extremely hydrophobic, which restricts their use for recovering and cleaning up oil spillsDespite reports of significant progress and promising applications, the oil/water separation technique still has many hurdles to overcome. More problems are related to the efficiency and productivity of the industry. One of the biggest problems is how graphene and its derivatives are attached to the substrate because they can easily detach from it during use. This is explained by weak van der Waals forces, which are responsible for keeping graphene and its derivatives on the substrate surface. Therefore, it is very important to develop preparation techniques and perhaps use powders that no longer require a substrate.The last point is related to the industrialization of the water and oil separation processes by graphene and its derivatives. So far, no research has been conducted on industrial scale studies. Instead, all studies have focused solely on small-scale laboratory systems. Scalability tests are required to assess the effectiveness and acceptability of the system at greater flow rates and under more accurate circumstances. However, recent developments in sponges with graphene attached are still weak and poor chemical and mechanical stability. Consequently, in this work, we tried to use magnetized graphene without the need for a substrate.The most importance novelty of our research is using ferromagnetic graphene in the composite which can help in the preparation of stable nanocomposites. In fact, ferromagnetic graphene can be attracted Iron particles by magnetic attraction. Our group recently searched the preparation of ferromagnetic graphene and reported several methods. This research is the first application of ferromagnetic graphene that is investigated in our group. On another hand, as a great result, these nanocomposites are smart and their interaction with water can change in the presence of magnets.Seeking an alternative route to improve the separation quality, we used powder instead of the surface, to separate water from oil. For this purpose, the powders of smart superhydrophobic nanocomposites were prepared which respond to a magnetic field. Nanocomposite powders can be easily separated after absorbing oil (petroleum contaminants) by using a magnet. These nanocomposites become more hydrophobic in the presence of magnets; consequently, their ability is increased to absorb oil. As the hydrophobicity of these nanocomposites increases in the presence of a magnetic field, it will be easier to completely separate them from water.The ethanol and graphite (99.9%) were bought from Merck Company. The industrial Iron powder (IP) and black iron oxide (BI), red iron oxide (RI) and yellow Iron oxide (YI) were obtained from Sanat Hamoon Company. Sulfuric acid (> 99%), sodium dichromate (98%) and Sodium hydroxide (99%) were purchased from Merck for use in alkaline and chemical engraving of aluminum alloys. Finally, an OES water filtration system  provided deionized water.61. Before each measurement, the platinum ring was carefully cleaned and flame-dried. Vibrating sample magnetometer  measurements were made to determine the magnetization of the produced nanocomposite. Raman spectroscopy  was used to acquire Raman spectra. FESEM (TESCAN) was used to analyse the morphologies. Analyses of the FTIR (8400S) spectrophotometer were carried out by the Shimadzu instrument. Using N2 adsorption at 77 K and a surface area analyzer , the Brunauer–Emmett–Teller (BET) technique was used to determine the specific surface area. The Data Physics OCA15 plus contact angle metre was used to measure the contact angle (CA) on the surface of the samples. Furthermore, the Digimizer programme was used to measure the contact angle.Larger graphene flakes were collected by centrifuge and they were removed (Hettich EBA20). Graphite impurities were quantified by ICP spectrometry (ICPS-7000 Shimadzu). A Sigma 700 tensiometer was used to measure the surface tension of an ethanol solution at 298.15 K using the ring technique at atmospheric pressure63. The liquid-phase exfoliation process in an ethanol/water solution was employed to disperse graphene63. The key factor for attaining a high yield of graphene dispersion was identified as adjusting the solution’s surface tension in the range of 40–50 mN/m62. Various liquids and surfactants are employed to modify surface tension67. In our previous work, graphene was dispersed using aqueous solutions of two different surfactants as well as their mixtures66. In this investigation, a 20:80 ethanol/water ratio was utilised to achieve a surface tension of 45 mN/m. In a mixture of 20:80 ethanol and water, the graphite (5 g/l) was ultrasonically sonicated for 30 min at high power (400 W). The suspensions had been centrifuged at 3000 and 5000 rpm for 10 min to get rid of the bigger flakes. The supernatant from the centrifugation procedure has been decanted and kept in a container in order to employ dispersed graphene in the creation of the nanocomposite. The nanocomposites were prepared in two separate steps with the centrifuged graphene at 3000 and 5000 rpm. According to Table By using a process called liquid-phase exfoliation, graphene suspensions were created 70. To this end, 50 ml of 2% solution of sodium hydroxide is poured into a beaker and the aluminum sheet, which has already been washed and dried with acetone in an ultrasonic bath, is immersed in a sodium hydroxide solution, and place the beaker for 2 min in an oil bath that has already been heated to 60 °C. Then, we wash and dry the alkaline aluminum sheet with deionized water. This method causes roughness on the surface of aluminum sheets and oxidation of the aluminum surface.A 30 mm × 30 mm × 0.3 mm aluminium sheet was first degreased in acetone, followed by three minutes of ultrasonic, a rinse in ethanol, and drying. To create a mechanical lock between the coating and the substrate and increase the surface adhesion, the alkali and chemical etching of Al sheet was performed by using sodium hydroxide solution and sulfochromic solution, respectivelyThe sulfochromic solution was prepared according to ASTM D2674 standard. 0.33 g of sodium dichromate powder was poured into a beaker, first, 8.17 ml of deionized water and then 1.83 ml sulfuric acid 98% was added to sodium dichromate. Treated aluminum sheets were immersed in the sulfochromic solution and the beaker was placed in an oil bath that had already reached a temperature of 60 °C for 30 min. Then, the acidified aluminum sheets were taken out and rinsed with deionized water. The oxidized aluminum surface of the alkaline stage was corroded by the acid at this stage, creating micrometer pores in it, which increases the roughness and mechanical locking between the substrate and the composite coating.After end of autoclave process, the prepared nanocomposites were dropped on the surface of the chemically etched aluminum sheet and dry at room temperature. Indeed, we prepared a layer of the composites of graphene@iron powder (IPG), graphene@black iron oxide (BIG), graphene@red iron oxide (RIG), graphene@yellow iron oxide (YIG) with the centrifuged graphene at 5000 rpm. In each step, a 10-µl drop was placed on the surface of the aluminum sheet, after drying; the next drop was placed on the previous dried drop until the surface of the sample has been covered with composite. After drying, we measured the contact angle of the composites deposited on the aluminum substrate with deionized water.In order to better observe the surface changes, FE-SEM images have been prepared from aluminum alloy before etching, after alkaline etching and at the last stage after chemical etching. In the image of the aluminum alloy after alkaline etching Figure , it is oThe test was performed qualitatively by pouring deionized water into the beaker, and then 20 µl of oil on the surface of the water was placed. One of the nanocomposites was ground into powder, and the oil absorption by the powder was observed after it was placed on the water surface. The magnet was then brought close to the outer wall of the beaker (from the outside); this was done separately for each of the prepared nanocomposites, and it was looked at how the oil was absorbed by the nanocomposites and how the magnetic field affected the absorption of oil.−1 have been observed. The produced powder was combined with KBr in a weight-to-weight ratio of 1:4 to create the pellet, which was then compressed. The spectra obtained for the nanocomposites show that graphene isn’t functionalized with iron or its oxides and no new bonds are formed in the graphene structure. In fact, graphene is magnetized in the autoclave46, and as a result, it will be connected to ferromagnetic iron compounds through magnetic attraction and form a composite.On a NICOLET 380 FT-IR spectrometer, Fourier transform-infrared (FT-IR) spectra between 4000 and 400 cm−1 in BI is connected to the bending vibration of the Fe–O bond71. Two peaks at 472.28 and 570.61 cm−1 in the FT-IR spectra of RI are associated with the transverse stretching vibrations of Fe–O73. Two strong bands at 528 cm−1 and 433 cm−1 in the FT-IR spectra of hematite nanoparticles are attributed to Fe–O deformation in the Fe2O3 rhombohedral lattice of hematite. The graphene sheet was incorporated into hematite nanoparticles (Fe2O3) during the formation of the composite of RIG, as shown by the shift of these bands from 528 to 570.61 cm−1 and 433 to 472.28 cm−1 in the FT-IR spectrum. The reason for this is the high absorption property of hematite compared to other iron oxides74. Additionally, YI provides two peaks in the ranges of 458.156 cm−1 and 607.168 cm−1 that are connected to the Fe–O bond. A little quantity of water has been absorbed on the samples, as evidenced by the peak in the range of 3300–3500 cm−1 in the spectrum of iron and its different oxides, which is connected to the stretching vibration of water. Due to intermolecular hydrogen bonding between FeOOH (YI) and the absorbed water, the surface hydroxyl band in YI is more intense and has moved to 3150 cm−175. The structure of FeOOH for YI is clearly shown by the band at 3150 cm−176. Eventually, the results obtained from FT-IR analysis confirm the formation of graphene composite with various iron oxides.FT-IR spectrum of pristine materials  can be seen in Fig. −1, which denotes the existence of defects in the graphene sheet, as well as the peak at 1582 cm−1, which is associated with the G band and denotes the vibration of the sp2 carbon sheet. It is also connected to the two-phonon resonance mechanism that the 2D band seen at 2627 cm−1 exists. Any factor that can affect the crystal lattice of graphene and its electronic structure can lead to the change and displacement of all three peaks G, D, and 2D. In other words, due to the exposure of the sample to these factors, the peaks are shifted, broadened or split. Additionally, in most of the spectra 46. In this work, autoclave has been used to prepare nanocomposites, since pressure has a major contribution in changing the electronic and vibrational structure of graphene. As a result of this structural disorder, it leads to a disorder in the Raman spectrum and even to the creation of new vibrational modes. In Fig. 64, there are defects mainly in the edges of the flakes and the basal plan is without defects. The presence of peaks at a wavenumber less than 1000 cm−1, which is related to iron oxides, confirms the formation of graphene nanocomposites with iron oxides. The presence of water molecules in the samples may be the cause of the wide peak that was formed in all spectra in the range of 3500–4100 cm−1.Typically, two Raman and infrared (IR) spectrometers are used to examine the bond vibrations of the manufactured materials. The Raman device picks up non-polar vibrations, while the IR device picks up polar vibrations. Defects in the structure of graphene both before and after the creation of the nanocomposite may be examined using Raman spectroscopy. The most noticeable Raman peak Fig.  is assocSee Fig. , structuIn this work, EDS analysis was prepared from pristine IP, BI, RI and YI Figure . EDS-MAPIron (Fe) and carbon (C) are the sole elements present in the corresponding IPG EDS spectrum Figure , which pThe increase in the size of RI and BI particles in the composite prepared with 3000 rpm graphene, which has more layers, leads to an increase in the distance between the particles. The majority of these particles are spherical, as shown in Fig. 46. In addition, iron compounds naturally have magnetic properties. Therefore, the combination of ferromagnetic graphene and magnetic irons can produce a stable composite in which all particles are bound together by magnetic attraction. In this study, the magnetic properties of all components are investigated by the vibrating sample magnetometer (VSM) at ambient temperature, in the field range of − 15 kOe < H <  + 15 kOe.In our previous study, we showed that the temperature and pressure inside the autoclave lead to the magnetization of grapheneAs shown in Fig. 46. In this study, the centrifuge at 5000 rpm is used for preparing graphene which its magnetization is more than 3000 rpm sample , and reactions at the interface between these iron oxides  with the solution affect their crystallisation and dissolution, stability, and rheology, as well as their interaction with other species. Surface Fe atoms are Lewis acids and react with Lewis bases because iron oxides have empty atomic orbitals. They consequently coordinate with water molecules or hydroxyl ions in aquatic settings where Fe and these ions share ion–electron pairs. Normally, the water molecules split apart during adsorption, covering the surface with hydroxyl groups that are coupled to the underlying Fe atoms. Iron oxides undergo a quick process called hydroxylation that takes minutes or hours to complete. It is followed by further water molecule adsorption to the surface OH groups through hydrogen bonding. Adsorbed water layers have different characteristics from bulk water; an “ice-like” structure has been proposed. Figure 72. Iron oxide surfaces can have three different types of OH groups with charges of − 0.5, 0, and + 0.5 if a charge of + 0.5 is taken into account for each Fe–O bond (under the assumption of six-fold coordination). Both the crystal structure and the rate at which certain crystal faces form determine the density of these groups. Consequently, it differs slightly from oxide to oxide and is also impacted by crystal shape. The geminal group, which consists of one iron atom and two OH groups, is the fourth kind of group72. For this reason, the behaviors of iron compounds are different in terms of hydrophilicity and hydrophobicity. IP and BI are hydrophobic, which shows the hydroxyl group on their surface is coordinated with two iron atoms or is geminal type. YI and FI are also hydrophilic, which indicates coordination with one or three Fe atoms77. Additionally, all substances exhibit magnetically smart behaviours, meaning that when magnetic fields are present, their contact angles vary. In the presence and absence of magnets, respectively, nearly BI and YI in powder form had the largest and lowest contact angles, as illustrated in Fig. The contact angle of all iron compounds is determined and the results show that IP and BI are hydrophobic and RI and YI are hydrophilic. Following thorough research, it was discovered that iron oxides often consist of tiny crystals with sizes ranging from a few tens of nanometers to a few microns. Because of this, these crystals have a large surface-to-volume ratio, and a large portion of their atoms are on their surface. These crystals have a high surface free energy  measurements demonstrate that an increase in the number of graphene layers has a significant impact on the morphology of different iron oxide particles, including their size and the distance between them when they form a composite with graphene. Multilayer graphene also produces particles with larger sizes and distances84.Investigating the change in the physicochemical properties of graphene by changing the number of its layers is one of the most important recent research works. Studies have shown that the shape of the metal oxide composited with graphene is influenced by the number of graphene layersFigure To determine the specific surface area and pore size, which are two effective parameters in oil absorption, BET analysis was performed for BIG nanocomposite, in addition to the pure materials . Thus, the efficiency of this nanocomposite for oil absorption was investigated. The findings in Table 0 is the powder weight before oil adsorption and m1 is the powder weight after oil absorption. The absorption capacity was obtained 2.8 g g−1.After pouring the BIG nanocomposite powder on the beaker surface containing oil and water, the oil on the water surface was absorbed by the nanocomposite powder. Then, by bringing the magnet closer, the rate of oil absorption by the nanocomposite increased so that the oil was completely absorbed by the nanocomposite Fig. . Video 1Graphene nanocomposites/iron powder and graphene nanocomposites/black, red and yellow iron oxides were prepared. The contact angle data showed that these nanocomposites are highly-hydrophobic and some of them are smart with respect to the magnetic field. Among these nanocomposites, the nanocomposite prepared with 3000 rpm graphene and RI showed the highest response to the magnetic field. So that by applying the magnetic field, the nanocomposite becomes more hydrophobic and absorbs more oil, and the oil is separated from it after removing the field. The prepared nanocomposites can absorb oil both when they are placed on the substrate and when they are used in the form of powder (without the need for a substrate). For oil absorption by the nanocomposites prepared in this work, unlike other methods that have been presented so far, no substrate is needed, and as a result, both time and money are saved. Nanocomposites are made from environmentally friendly materials and do not pollute the environment. Easily and with the help of magnetic field (magnet), nanocomposite powder can be separated from the water after absorbing oil. These nanocomposites are smart and most of them become more hydrophobic in the presence of a magnet and thus have a higher ability to absorb the oil. The magnetic field applied by a magnet to these nanocomposites, in addition to increasing the hydrophobicity and as a result of more absorption of oil by the nanocomposite, causes their complete separation from water. This study is ongoing to enhance absorption capacity while employing a straightforward approach for large-scale implementation in industries. Additionally, a combination of various iron oxides is being employed to achieve improved outcomes.Supplementary Information.


Large language models exhibiting human-level performance in specialized tasks are emerging; examples include Generative Pretrained Transformer 3.5, which underlies the processing of ChatGPT. Rigorous trials are required to understand the capabilities of emerging technology, so that innovation can be directed to benefit patients and practitioners.Here, we evaluated the strengths and weaknesses of ChatGPT in primary care using the Membership of the Royal College of General Practitioners Applied Knowledge Test (AKT) as a medium.AKT questions were sourced from a web-based question bank and 2 AKT practice papers. In total, 674 unique AKT questions were inputted to ChatGPT, with the model’s answers recorded and compared to correct answers provided by the Royal College of General Practitioners. Each question was inputted twice in separate ChatGPT sessions, with answers on repeated trials compared to gauge consistency. Subject difficulty was gauged by referring to examiners’ reports from 2018 to 2022. Novel explanations from ChatGPT—defined as information provided that was not inputted within the question or multiple answer choices—were recorded. Performance was analyzed with respect to subject, difficulty, question source, and novel model outputs to explore ChatGPT’s strengths and weaknesses.P=.04 and .06). ChatGPT’s performance varied with subject category (P=.02 and .02), but variation did not correlate with difficulty . The proclivity of ChatGPT to provide novel explanations did not affect accuracy (P>.99 and .23).Average overall performance of ChatGPT was 60.17%, which is below the mean passing mark in the last 2 years (70.42%). Accuracy differed between sources (Large language models are approaching human expert–level performance, although further development is required to match the performance of qualified primary care physicians in the AKT. Validated high-performance models may serve as assistants or autonomous clinical tools to ameliorate the general practice workforce crisis. Deep learning is a form of artificial intelligence (AI), which facilitates the development of exquisitely organized processing within an artificial neural network architecture, composed of multiple layers of interlinked perceptron nodes . During Generative Pretrained Transformer 3.5 (GPT-3.5) is a large language model (LLM), trained on a data set of over 400 billion words from articles, books, and other forms of media on the internet . ChatGPTThe Applied Knowledge Test (AKT) of the Membership of the Royal College of General Practitioners (RCGP) must be passed for GPs to complete their training in the United Kingdom. A total of 200 questions—mostly multiple choice but with occasional requirement to input numbers or select from a longer list of potential answers—must be answered in 190 minutes by candidates at a computer workstation. Questions test mostly clinical knowledge (80%), as well as evidence-based practice (10%) and primary care organizational and management skills (10%). All questions are designed to test higher-order reasoning rather than simple factual recall.Before trials of clinical applications of NLP chatbots can be designed, the proposed purpose of applications such as ChatGPT must be established, requiring thorough investigation of their strengths and weaknesses. To evaluate the utility of ChatGPT in primary care settings, we used the AKT as an existing standard met by all UK GPs. The distinct sections of the AKT enabled the investigation of the opportunities afforded by ChatGPT (and LLMs more broadly), as well as the limitations of currently available technology. Through this work, we aimed to provide suggestions as to how clinical and computational research should proceed with the design and implementation of NLP chatbots, supported by empirical data.AKT questions were sourced from the RCGP’s GP SelfTest platform , as wellEvery eligible question was inputted into ChatGPT  on 2 separate occasions between January 30 and February 9, 2023, in separate sessions to avoid the second trial from being influenced by previous dialogue. ChatGPT’s answer was recorded, and its whole reply to each question was recorded for further analysis. If ChatGPT failed to provide a definitive answer, the question was retrialed up to 3 times, after which ChatGPT’s answer was recorded as “null” if no answer was provided. Correct answers  was defined as the answers provided by GP SelfTest and the practice papers—these were recorded for every eligible question. ChatGPT’s responses were screened for “novel explanations”—defined as any information provided that was not included in the question or multiple choice answers—by a single researcher.The scores required to pass the AKT in every examination undertaken in the last 2 years were collected from RCGP examiners’ reports for the AKT between 2018 and 2022 . AdditioP values, with statistical significance concluded where P<.05. Statistical analysis was conducted in R , and figures were produced using Affinity Designer .ChatGPT’s answers in both trials were compared to the correct answers to gauge performance and were compared to recent pass marks to assess ChatGPT’s prospects of passing the AKT. ChatGPT’s answers were compared between the 2 trials to measure the consistency of its responses. Performance was analyzed with respect to difficulty, explanation novelty, source, and subject to explore the strengths and weaknesses of ChatGPT. Nonparametric statistical analysis was undertaken due to the nonrandom nature of question design and small number of questions in some subjects. Effect sizes were reported with 95% CI and Ethics approval was not required for this study as human participants were not involved.In total, 720 questions were identified, which increased to 733 questions after multipart questions were separated into distinct entries. In total, 674 unique questions were ultimately inputted into ChatGPT after duplicate and incompatible questions were excluded . IncompaP=.04) but not the first  trial. This indicates that question difficulty (for ChatGPT) differed between sources, although differences in performance were not large  and second  trials. Subject variation did not correlate with the difficulty indicated by the frequency of recommendations of “room for improvement” by the RCGP  or second  iterations (ChatGPT provided novel explanations in response to 58 (8.61%) questions in the first run and 66 (9.79%) questions in the second run. A novel explanation was provided in response to just 18 (2.67%) questions in both runs, illustrating significant stochasticity in the relationship between prompt and output. The proclivity of ChatGPT to provide a novel explanation had no bearing on accuracy in the first , and semiautonomous completion of administrative tasks such as clinic notes, discharge summaries, and referral letters ,22. FurtChatbots leveraging advanced NLP models are an exciting innovation with the potential to ameliorate staffing pressures that disproportionately affect deprived areas . HoweverEffective applications must be rigorously trialed in the same context as the one they are intended to be deployed in the future ,29. As e


Stacking of graphene with hexagonal boron nitride (h-BN) can dramatically modify its bands from their usual linear form, opening a series of narrow minigaps that are separated by wider minibands. While the resulting spectrum offers strong potential for use in functional (opto)electronic devices, a proper understanding of the dynamics of hot carriers in these bands is a prerequisite for such applications. In this work, we therefore apply a strategy of rapid electrical pulsing to drive carriers in graphene/h-BN heterostructures deep into the dissipative limit of strong electron-phonon coupling. By using electrical gating to move the chemical potential through the “Moiré bands”, we demonstrate a cyclical evolution between metallic and semiconducting states. This behavior is captured in a self-consistent model of non-equilibrium transport that considers the competition of electrically driven inter-band tunneling and hot-carrier scattering by strongly non-equilibrium phonons. Overall, our results demonstrate how a treatment of the dynamics of both hot carriers and hot phonons is essential to understanding the properties of functional graphene superlattices. Significant attention has been devoted to understanding the low-electric-field properties of carriers in moiré graphene, but high-electric-field transport has not been as well explored. Here, the authors find non-monotonic transport behavior at moiré minigaps due to competition between inter-band tunneling and coupling to out-of-equilibrium phonons. By combining different semiconductors in periodic superlattices, artificial band structures can moreover be implemented. This technology has been widely used to realize active optoelectronic devices, with Bloch oscillators4 and quantum-cascade lasers5 providing just a couple of important examples. The recent emergence of atomically thin two-dimensional (2D) materials, combined with the capacity to stack6 these materials in multilayered “van-der Waals heterostructures”, has now opened up the possibility of extending traditional concepts of band-engineering to a whole new class of materials and structures. Atomically thin materials offer many natural advantages over their bulk counterparts, including phononic, excitonic, and plasmonic energies that are enhanced by quantum carrier confinement. By combining the best qualities from different 2D materials, the potential exists to enhance the performance of semiconductor technology well beyond its present capabilities.The dynamics of hot carriers in the energy bands of semiconductors have long been exploited in electronic and optoelectronic technology. Notable examples include light-emitting diodes and lasers7 remarkable progress in realizing new physics in multilayered 2D materials. Graphene and hexagonal boron nitride (h-BN) constitute the workhorse for much of this research, in large part because their honeycomb crystal structures are only slightly mismatched from one another (the lattice constant of h-BN is larger than that of graphene by <2%). Their multilayers can therefore exhibit periodic Moiré structures, whose details depend upon the rotational (or twist) angle between the graphene and h-BN sheets15 . These details have been confirmed by scanning-tunneling microscopy9, and from signatures of Hofstadter’s butterfly in quantum-Hall states22. Elsewhere, the implications of Moiré-miniband formation have also been confirmed in transport experiments performed in magnetic-focusing23 and cyclotron-resonance24 geometries. Most dramatic of all has been the discovery that, for a “magic” rotational angle of ~1.1°, superconductivity can even be induced in the 2D carbon layer25. Collectively, these works reveal an unprecedented capacity to engineer electronic states in 2D heterostructures.Recently, there has been26. Yet, while significant attention has been devoted to understanding the low-field (near-equilibrium) properties of carriers in graphene’s Moiré minibands, the corresponding problem of high-field transport has not been as well explored. Only a few studies30 have addressed how far-from-equilibrium carrier distributions may be generated via optical excitation in graphene/h-BN heterostructures. The number of reports concerning hot-carrier action under strong electrical driving is even smaller32, and these works do not consider how this transport is modified in the presence of graphene’s Moiré minibands.While the band engineering of 2D materials offers enormous promise for the design of tailored electronic systems, realizing this potential requires a proper understanding of hot-carrier dynamics in the artificial bands of their heterostructures. To be clear, this problem of high-field (>kV/cm) transport typically bears little relation to its low-field counterpart. Instead, it is necessary to consider a strong coupling of the electronic and phononic systems, which arises as both are driven far from equilibrium33, are taken into account  with the average source–drain electric field (note that 10 kV/cm = 1 V/μm), as determined by transient pulsing for a heterostructure of this kind .  By sweeping the voltage (Vg) applied to the top gate  heterostructure with no clear commensurability between its different layers. In this situation, there is no obvious Moiré-miniband structure and the only role of the encapsulating h-BN is to provide smooth and stable surfaces in contact with the graphene236, yet consistent with prior work on graphene/h-BN32, the saturation is never fully complete. Consequently, the current density reaches ultimate values larger than 10 mA/μm (near the highest average field of 80 kV/cm), significantly exceeding the levels reported in previous work. This observation is consistent with the capacity of fast pulsed measurements to attain high current densities, well beyond those realizable in DC measurements, by suppressing self-heating38 of the system under study.When the gate voltage is used to drive the Fermi level away from the Dirac point the current–voltage curves of Fig. R, determined by dividing the steady stage voltage drop across the device by the corresponding value of the transient current, see Vg. At the lowest average field of 5 kV/cm, the Dirac point (defined to occur when Vg ≡ VD) can be clearly identified (near Vg = −3 V) and the relative overall resistance variation induced by the gate voltage corresponds to ~150%. With an increase in the electric field, the size of this variation shrinks and the Dirac peak becomes strongly broadened around its center point. . At Vg ≤ –3.75 V, the current reaches another local minimum, where it once again exhibits a quasi-linear dependence on electric field  is associated with the usual Dirac-point crossing, while the second is located on the hole side (near Vg = 3.7 V) of this peak. Although the hole-side feature is clearly visible at this relatively high temperature (300 K), there is no sign of any corresponding peak on the electron side. This is a common observation in many prior studies of linear transport in Moiré bands18, and is understood to result from the presence of a smaller Moiré gap for electrons than for holes10  Moiré structure arises from the coupling between the graphene and just one of the h-BN layers.To reveal the influence of the Moiré spectrum of GhBN2 more prominently, in Fig. 7 to be associated with the opening of  gaps in the conduction and valence bands, and at the Dirac point itself  for GhBN2 . Referring to the behavior exhibited around the primary Dirac peak (near Vg = 0.5 V), the local resistance minima that flank this feature should correspond to a situation in which the Fermi level lies away from any minigap, being located instead within either the conduction or the valence miniband. Consequently, in Fig. The multi-peaked structure seen at low fields in Fig. see Fig. . Consequh in Fig. Next, we turn to the corresponding behavior exhibited in the vicinity of the hole-related peak, where the Fermi level should lie in the minigap Δe) is much smaller here, typically exhibiting only a nascent character . Consequently, when we plot . Electron–phonon coupling is primarily considered to involve the optical phonons (with energy ħωph = 210 meV) that are known to dominate the high-field transport of graphene. The electrons and phonons are treated on an equal footing, accounting self-consistently for their self-energies within the dynamical mean-field approximation. To describe dissipation in the DC limit, we allow the electrons and phonons to decay into fermionic and phononic baths (at respective rates Γ and τp−1), which are kept at the initial (equilibrium) substrate temperature  and accounts naturally for both the emission and absorption of phonons by hot carriers. The parameter τp may be viewed as representing the lifetime of non-equilibrium phonons, generated by emission from the hot carriers. In a situation in which all other parameters are held fixed, increase of τp will lead to both a rise in the effective temperature (Tph) of these phonons and to increased carrier scattering. We refer the reader to the “Methods” section and to the Motivated by the capacity of transient measurements to probe the intrinsic properties of small devices, while strongly suppressing extrinsic effects due to self-heatinggep2/ħωph) of the electron–phonon coupling. In the cold-phonon model of Fig. 35 and so leads to the decrease of resistance with electric field shown in panel (a). The situation is very different in Fig. gep2/ħωph > 3.7 eV, the resistance now increases with the electric field. Basically, what is happening here is that the increased electron–phonon coupling is allowing hot-phonon effects to dominate over Landau–Zener tunneling, resulting in a very different variation of resistance. In Fig. gep2/ħωph = 3.91 eV and calculate the variation of resistance with an electric field for various initial values of μ in the conduction band. These results capture the effect of varying the gate voltage in the experiment and demonstrate that a proper treatment of hot-phonon effects, and their competition with Landau–Zener tunneling, is needed to accurately describe transport in graphene under non-equilibrium. At higher electric fields (>30 kV/cm) the calculations show a good correspondence with those of Fig. μ = 0), the resistance varies only slowly with an electric field, reflecting the weak influence of electron-phonon effects in the presence of the small Fermi surface. As we move away from the Dirac point, however, the resulting increase in the Fermi surface allows non-equilibrium hot phonons to enhance electron scattering, leading to an increased variation of the resistance. This behavior not only resembles that found for un-gapped graphene but is also like that obtained away from the Moiré resistance peaks of Fig. To provide a comparison with the experimental results of Fig. Moving to the behavior expected when the Fermi level lies within a Moiré gap, in Fig. μ = 0), similar results are also obtained away from this condition. The results of Fig. μ = 18 meV and correspond to a Fermi level that lies well within the conduction band. When 2Δ < 36 meV, this situation mimics the behavior away from either the primary or secondary Dirac peaks  and lattice (Tph) temperatures. These temperatures are deduced in standard fashion, from the form of the electron and phonon distribution functions under non-equilibrium (as described in Eqs. (S18)–(S20) of the 35, in which we artificially impose an instantaneous phonon decay (τp = 0) to hold Tph at its initial (equilibrium) value (Tph = To = 3.5 K). Here we see that the electron temperature increases almost linearly with an electric field, approaching 900 K at the highest fields. Similarly high temperatures have been obtained previously in Monte-Carlo simulations of high-field transport in graphene40. The near-linear variation of Te with an electric field is reasonable for the cold-phonon scenario, for which the supplied electrical power  is dissipated in the carrier system alone. Noting that the energy of a degenerate Fermi gas (at low temperatures) varies as Te2, we might then expect a simple linear dependence of Te with the electric field. This model is clearly unphysical, however, as we have seen already  data points of Fig. Te and Tph appear insensitive to variation of the Fermi level or to whether the graphene is gapped or un-gapped. We attribute this to the fact that transport is dominated by scattering from optical phonons at high fields, independent of the details of low-field transport.As is common in discussions of non-equilibrium carrier phenomena in semiconductors41, novel signatures of the so-called “Dirac fluid” have been studied in graphene superlattices, realized with precise control of the interlayer twist angle. The emergence of controlled mini-gaps in the bandstructure of these systems enables a situation in which the drift velocity of driven carriers can be comparable to the Fermi velocity, leading to a transition to a state in which the filled bands begin to play an essential role in transport. The critical phenomena observed under those conditions were achieved at current densities some two orders of magnitude lower than those reached here, as a result of which the influence of phonon-related effects was not considered. This should therefore be contrasted with our study, performed deep in the dissipative regime, in which we have seen how it is essential to account for the non-equilibrium nature of both the carrier and phonon populations.In very recent workIn this study, we have demonstrated the presence of re-entrant metallic and semiconducting states in a Moiré-gapped h-BN/graphene/h-BN heterostructure. The semiconducting states are observed when the chemical potential is swept through the commensurability induced gaps in the graphene spectrum, while metallic behavior is obtained when this level is located within the minibands bound by these gaps. To account for the experimentally observed behavior, we have developed a model of non-equilibrium transport that accounts self-consistently for the coupling of the carrier and lattice systems. The picture that emerges from this model is one in which the experimentally observed resistance variations can be attributed to a competition between electrically driven inter-band (Landau–Zener) tunneling and nonequilibrium electron–phonon coupling. The latter dominates the metallic resistance variations, found for ungapped graphene or when the Fermi level lies within a Moiré miniband. With the Fermi level in a minigap, however, Landau–Zener tunneling dominates at low fields before hot-phonon effects take over at higher fields. It is this competition that results in the nonmonotonic resistance variations characteristic of the semiconducting states. Overall, our results provide important insight into hot-carrier dynamics in graphene/h-BN heterostructures and the impact of hot phonons on their electrical performance.42. In this approach, graphene flakes and h-BN crystals were exfoliated onto SiO2/Si (300 nm/0.5 mm) chips from bulk crystals. The material used for device fabrication was carefully chosen by means of optical microscopy, Raman spectroscopy, and atomic force microscopy , reactive-ion etching (in a gas mixture of CHF3 and O2), and electron-beam evaporation (Cr/Pd/Au: 3-/15-/80-nm). In this two-step process, the edge contacts were first fabricated using a high-resolution EBL exposure. Next, the large coplanar waveguides were fabricated in a second EBL step, in which a system of on-chip alignment markers was used to accurately overlap the waveguides with the two edge contacts. The extended metallized contacts formed in this way are denoted by white dotted lines in the lower inset to Fig. The van der Waals heterostructures studied here were assembled by the PC (6% poly bisphenol A carbonate dissolved in chloroform)/poly dimethylpolysiloxane- (PDMS) based dry-transfer method44. The measurements were made in a custom-designed setup with full (50-Ω) impedance matching. The device chip was mounted on an FR-4 laminate board, allowing the signal lines of its coplanar waveguide to be connected to semirigid coaxial cables. Single-shot or repetitive pulses were applied with an AVTECH generator, providing a maximum pulse amplitude of 10 V and a repetition frequency that was set at 10 kHz to avoid the influence of self-heating38. The pulse duration could be varied from a few ns to a microsecond and was chosen here to be 60 ns; for this optimal duration, the steady-state current reached with the pulse applied could be accurately identified yet did not differ noticeably from that obtained using shorter pulses . To estimate the twist angle between these two layers, we use the (gate-voltage) separation between the two secondary peaks and the primary Dirac peak , λ is the twist angle between the layers, and θ is defined from the carrier concentration at the secondary peaks according to In GhBN2, the presence of just one secondary peak due to each of the valence and conduction miniband indicatesħωph = 210 meV, is modeled on an infinite honeycomb tight-binding lattice subject to the electrostatic potential V(r) = −eEx . The electron-phonon self-energies are then approximated within the dynamical mean-field theory45. The electron and phonon baths are added by means of dissipative self-energies, facilitating the steady-state DC limit46 [which we impose on the Green’s function via G = G when the position r is displaced by lattice vector a]. Dissipation is controlled by means of the electron dissipation rate Γand the hot-phonon decay rate τp−1 into Ohmic baths. To model transport in graphene with a gap induced by the Moiré effect, we add the energy displacement +Δ and –Δ, respectively, to the A- and B-sublattices of the honeycomb crystal. For further details on the theoretical procedures, we refer the reader to Section We model transport in monolayer graphene by using the non-equilibrium Green’s function method, placing electrons and phonons on an equal footing by self-consistently computing their self-energies. The graphene layer, with on-site optical phonons of energy Further information on research design is available in the Supplementary InformationReporting Summary


AAV6-treated mice fibrotic models, targeting FOXC2 in vivo lead to a reduced degree of liver fibrosis. In sum, our study uncovers a reciprocal crosstalk between matrix stiffness and FOXC2-AS1/FOXC2 axis leading to modulation of HSCs mechanoactivation and liver fibrosis, and present AAV6 shRNA as an effective strategy that targets FOXC2 leading to the resolution of liver fibrosis.Matrix stiffness is a central modulator of hepatic stellate cells (HSCs) activation and hepatic fibrogenesis. However, the long non-coding RNAs (lncRNAs)-regulated transcriptional factors linking matrix stiffness to alterations in HSCs phenotype are not completely understood. In this study, we investigated the effects of matrix stiffness on HSCs activation and its potential mechanism. Through analysis the RNA-seq data with human primary HSCs cultured on 0.4 kPa and 25.6 kPa hydrogel, we identified that forkhead box protein C2 (FOXC2) and its antisense lncRNA FXOC2-AS1 as the new mechanosensing transcriptional regulators that coordinate HSCs responses to the matrix stiffness, moreover, FOXC2 and FOXC2-AS1 expression were also elevated in human fibrosis and cirrhosis tissues. The matrix stiffness was sufficient to activate HSCs into myofibroblasts, resulting in nuclear accumulation of FOXC2. Disrupting FOXC2 and FOXC2-AS1 level abrogated stiffness-induced activation of HSCs. Further mechanistic studies displayed that stiffness-upregulated lncRNA FOXC2-AS1 had no influence on transcription of FOXC2. FOXC2-AS1 exerted its biological function through maintaining the RNA stability of FOXC2, and protecting FOXC2 mRNA from degradation by RNA exosome complex. Additionally, rescue assays confirmed that reintroduction of FOXC2 in FOXC2-AS1-depleted HSCs reversed the repression of FOXC2-AS1 knockdown on stiffness-induced HSCs activation. In  Liver fibrosis is a wound healing process that forms in response to a variety of nosetiologies, including hepatitis virus infection, cholestatic disorders, alcoholic liver disease (ALD), and non-alcoholic steatohepatitis (NASH)Recent evidence indicates that matrix stiffness is central to the persistent HSCs activation that drives progression of fibrosisForkhead box protein C2 (FOXC2), a transcription regulator of the forkhead/winged-helix family, plays critical roles on cell proliferation, apoptosis, lymphangiogenesis and differentiationin vivo suppressed CCl4 or bile duct ligation (BDL)-induced HSCs activation and liver fibrosis. Thus, our studies demonstrated that FOXC2 represents a novel target for mitigating stiffness-induced HSCs activation and liver fibrosis.In this study, through analysis of RNA-seq data of HSCs cultured on polyacrylamide gels with incremental stiffness, we found that matrix stiffness markedly upregulated the expression of FOXC2, and functional experiments elucidated that stiffness-upregulated FOXC2 lead to transcription of α-SMA, Col1A1, CTGF and FN1. Moreover, mechanistic studies revealed that stiffness-induced long non-coding RNA FOXC2-AS1 binds to FOXC2 mRNA and protects it from degradation by the RNA exosome complex to maintain its stability, which plays a crucial role on HSCs activation and liver fibrosis. AAV6 carrying shRNA targeting FOXC2 The primary human HSCs were purchased from ScienCell #5300), and cultured in DMEM (Dulbecco's Modified Eagle's Medium) supplemented with 10% fetal bovine serum, streptomycin and penicillin. The monitoring mycoplasma infection of HSCs were performed by utilizing a MycoAlert detection kit  and these cells were free of infection during the experiments. Human primary HSCs with passage < 8 were utilized for all experiments. The antibodies  used in this study were reported as previous described300, and 63 human liver fibrosis/cirrhosis specimens were collected from patients undergoing liver transplantation in the First Affiliated Hospital of Xi'an Jiaotong University. Patients who were diagnosed with Wilson's disease, Alpha-1 antitrypsin (AAT) deficiency, or other inherited liver diseases based on standard clinical, laboratory and histological assessments were excluded in this study. 21 normal liver specimens were obtained from patients with liver trauma or hepatic hemangioma. All procedures performed in studies involving human participants were in accordance with the ethical standards of the Research Ethics Committee of The First Affiliated Hospital of Xi'an Jiaotong University and with the 1964 Helsinki declaration and its later amendments. All written informed consent to participate in the study was obtained from patients with liver fibrosis, liver trauma or hepatic hemangioma for samples to be collected from them.The generation of RNA-seq data GSE101343) with primary human HSCs seeded on 0.4 kilopascal (kPa) or 25.6 kPa hydrogels and further bioinformatics analysis was performed as previously reported43 with pPackaging of lentiviruses was performed as we previously describedPolyacrylamide hydrogels with incremental stiffness with 0.4 kPa or 25.6 kPa were generated according to the protocols as previously reportedHSCs cultured on polyacrylamide hydrogels with different stiffness (0.4 kPa or 25.6 kPa) or human liver cryosections were prepared for IF staining, as previously describedAfter finishing the designated intervention, the mRNA was extracted from HSCs, human or mice livers by using the Trizol . Equal amounts of mRNA were converted into cDNA (complementary DNA), and quantitative polymerase chain reaction (qPCR) was conducted as we previously describedHSCs, cultured on 0.4 or 25.6 kPa hydrogels, were harvested for ChIP assay, ChIP assay was conducted using an EZ-Magna-ChIP HiSens kit  as previously reportedRIPA buffer  added with a phosphatase inhibitor cocktail  and protease inhibitors  was utilized to lyse the hepatic stellate cells or liver tissues. The protein concentration of the samples was assessed by using a BCA protein assay kit . Western blot assays and densitometric analysis of the designated protein bands were conducted as we previously describedHSCs were incubated with the transcriptional inhibitor actinomycin D  for 0 h, 2 h, 4 h, and 8 h, respectively. The HSCs were collected at the indicated time points, and RNA was isolated. Then, qRT-PCR was performed to assess the relative mRNA level of FOXC2.TM RNA Binding Protein Immunoprecipitation Kit  according to the manufacturer's protocols. Briefly, RIP lysis buffer added with the RNase inhibitors (#3335399001 MilliporeSigma) and protease inhibitor cocktail  was applied to lyse the cells. Then the specific antibody (anti-EXOSC10 or anti-EXOSC4) and protein A/G magnetic beads were placed in cell lysates for overnight at 4℃. On the second day, the RNA/protein complexes binding on the beads were precipitated by magnetic force, then the RNA was extracted using Trizol and determined by qRT-PCR analysis.RIP was conducted by utilizing the EZ-Magna RIPin vitro transcribed from the recombinant plasmids pCMV3-FOXC2-AS1 or pCMV3-FOXC2, and the Biotin RNA Labeling Mix  was utilized to generate biotin-labeled FOXC2-AS1 and biotin-labeled FOXC2 RNA as previously reportedThe FOXC2-AS1 and FOXC2 were The cytosolic and nuclear fractions of hepatic stellate cells were extracted with PARIS Kit  according to the manufacturer's instructions.Nascent RNA of HSCs was extracted by using the Click-iT Nascent RNA Capture Kit  following with the manufacturer's instructions. Then, qRT-PCR was performed to examine the relative expression of FOXC2 nascent RNA.Subcellular localization of FOXC2-AS1 in HSCs was detected by the FISH Kit  following the product's instructions. In brief, HSCs were cultured on coverslips into 24-well plates. Then, the 4% paraformaldehyde was utilized to fix HSCs for 15 minutes. After that, the fixed HSCs were washed with PBS and subjected to permeabilization (0.5%Triton-X PBS for 10 minutes). Subsequently, HSCs were incubated with prehybridization solution and hybridization solution respectively, then hybridized with the cy3-labeled FOXC2-AS1 oligonucleotide probe  overnight at 4℃. HSCs nuclei was visualized with DAPI for 5 min at room temperature. All pictures were taken and recorded by Zeiss Instruments confocal microscope.12 viral genomes·mL-1; Hanbio Biotechnology, Shanghai, China) through tail vein injection, and the detail protocol of AAV6-shFOXC2 injection was reported as previously describedAAV6-shFOXC2 in murine activated HSCs. The detail protocol for murine HSCs isolation was displayed as previously describedAll animal experiments were conducted following the instructions authorized by the ethical committee of Xi'an Jiaotong University. Six- to eight-week male C57BL/6 mice were obtained from Animal Centre of Xi'an Jiaotong University and they were supplied with free access to water and food. Liver fibrosis was induced by intraperitoneal injection of 20% CCl4  in olive oil at 5 μL·g-1 of body weight, twice per week. Two weeks after intraperitoneal injection of CCl4, these mice were administered with 100 μL of AAV6-shControl (non-targeting control) or AAV6-shFOXC2  via tail vein injection. After 3 weeks, mice were sacrificed and the livers obtained for further analysis of fibrogenesis, and IHC staining of CK-19 in BDL model was used to show the proliferation of bile duct epitheliumBDL was conducted as previously reportedP<0.05 was considered as statistically different.Data were displayed as Mean ± S.E.M. The difference between groups was assessed by Student's t-test or ANOVA followed by a post hoc test applying GraphPad Prism 6 software . in vitro culture of HSCsin vivo, CCl4-incuded mice liver fibrotic tissues were collected to examine FOXC2 expression. Corroborating in vitro findings, mice livers harvested after CCl4 injection showed prominently increased mRNA and protein expression of FOXC2  exert vital roles on HSC activationtro Fig. B. Next, vivo Fig C. These However, the function of stiffness-induced FOXC2-AS1 in HSC is still unknown. We next used FOXC2-AS1 shRNA to examine the role of FOXC2-AS1 in mechanotransduction of HSCs. HSCs transduced with lentiviruses encoding FOXC2-AS1 shRNA or non-targeting shRNA (control shRNA) were then plated on 0.4 kPa or 25.6 kPa hydrogels and collected for IF, qRT-PCR and western blot, which corroborated that depletion of FOXC2-AS1 inhibited stiffness-induced upregulation of α-SMA, Col1A1, FN1 and CTGF in HSCs  and a cap AAV6 vector carrying transcription regulators was able to successfully reprogram activated hepatic stellate cells into hepatocytes and thus contribute to mouse hepatic fibrogenesis recoveryet al. uncovered that AAV6 displayed organ tropism for activated HSCs or myofibroblasts in the liver during fibrosis, and the targeting efficacy of AAV6 was reach to 17.04 % in nestin+ murine activated HSCs of CCl4-induced liver fibrosis model AAV vectors have now been considered as a safe and efficient gene delivery tool in clinical trials since they possess low oncogenicity and weak immunogenicityAAV6 vectors carrying shFOXC2 to target activated HSCs and investigate the efficacy of AAV6 shRNA. Our in vitro data present the evidence of the critical involvement of FOXC2-AS1/FOXC2 signaling in stiffness mediated HSCs activation. Consistently, in vivo analyses of targeting FOXC2 by AAV6 shRNA in CCl4 or BDL-induced liver fibrosis model substantiate our in vitro findings. We provided evidence to exhibit that applying with an AAV6 shRNA to effectively impair FOXC2 expression in activated HSCs could mitigate hepatic fibrogenesis in mouse fibrosis model, providing a promising anti-fibrotic strategy. Moreover, besides AAV vectors, liposomes, EV or other new nanomaterial-based targeted delivery tools have been identified in recent years. These vectors have showed better targeting efficiency and great promise for treatment of hepatic fibrosisAAV6 shRNA as an effective strategy that targets FOXC2 leading to liver fibrosis resolution.Recently, the study reported that an Supplementary figures and tables.Click here for additional data file.


For the high-power-density operation at 31,000 W/kg , an energy density as high as 11 Wh/kg (or 3.5 Wh/L) was achieved. The results indicated that N-HGE not only improved the conductivity of the composite supercapacitors but also accelerated ion transport by way of shortened diffusion paths through the numerous holes all over the graphene sheets.We present a facile low-cost method to produce nitrogen-doped holey graphene (N-HGE) and its application to supercapacitors. A composite of N-HGE and activated carbon (AC) was used as the electrode active material in organic-electrolyte supercapacitors, and the performances were evaluated. Melamine was mixed into graphite oxide (GO) as the N source, and an ultra-rapid heating method was used to create numerous holes during the reduction process of GO. X-ray photoelectron spectra confirmed the successful doping with 2.9–4.5 at.% of nitrogen on all samples. Scanning electron micrographs and Raman spectra revealed that a higher heating rate resulted in more holes and defects on the reduced graphene sheets. An extra annealing step at 1000 °C for 1 h was carried out to further eliminate residual oxygen functional groups, which are undesirable in the organic electrolyte system. Compared to the low-heating-rate counterpart (N-GE-15), N-HGE boosted the specific capacity of the supercapacitor by 42 and 22% at current densities of 0.5 and 20 A/g, respectively. The effects of annealing time  at 1000 °C were also studied. Longer annealing time resulted in higher capacitance values at all current densities due to the minimized oxygen content. Volumetric specific capacitances of 49 and 24 F/cm Imaterial ,8. Howevmaterial .2/g) of graphene compared to AC, it has many excellent properties, such as high electrical conductivity, thermal conductivity, chemical stability and mechanical strength, which have made them the focus of many EDLC research activities [Despite the slightly smaller SSA  hierarchical porous structures have been utilized by several researchers to overcome the restacking problem. For example, Kim et al.  demonstrn et al.  developeV is smaller than those of organic electrolytes (~2.5–3.0 V). Note that the energy density is proportional to (ΔV)2. Xu et al. [Modifying the morphology of graphene to further improve its electrochemical performance has also been reported by many researchers. Corrugated graphene , curved u et al.  fabricat3) despite its high gravimetric capacitance (F/g). To increase the volumetric capacitance and to extend the ΔV, Huang el al. [3) at high rate charging/discharging (20 A/g).Another important limitation for commercial applications is the low tap density and mass loading for the graphene electrode material, which result in low volumetric capacitance , and then heated to 1000 °C and held for 1 h to further reduce the remaining oxygen functional groups. For N-HGE, the duration of the 1000 °C annealing was also varied  in order to study the effect of residual oxygen functional groups on the performance of SCs. Concerning the yields of the prepared materials, the major material loss occurs in the two heating processes converting MA-GO into N-HGE. First, in the ultra-rapid heating step, typically a weight loss of about 60% occurs; second, during the 0.5 to 2 h of 1000 °C annealing process, the weight loss ranges from 25% to 50%. So the overall yield for N-HGE is around 20–30%.Natural graphite  was first chemically oxidized into graphite oxide (GO) using a modified Staudenmaier method . To prepA scanning electron microscope  was used to examine the microstructure of the graphene. Transmission Electron Microscopy  was used to observe the holes on exfoliated holey graphene sheets. Brunauer–Emmett–Teller (BET) measurements were carried out with a Micromeritics ASAP 2020 surface area and porosity analyzer to evaluate the SSA and pore distribution. To study the thermal decomposition behavior of MA and the thermal reduction process of GO, thermogravimetric analysis (TGA) was performed using a Mettler Toledo TGA/DSC 2-HT thermogravimetric analyzer. To determine the crystal structure of GO and graphene samples, X-ray diffractometry  was performed. Moreover, the D and G bands of graphene in Raman spectroscopy were characterized with a Renishaw H27783 system. Finally, to investigate the chemical composition of the graphene samples, X-ray photoelectron spectra (XPS) were measured using a Thermo Scientific K-Alpha spectrometer. 50 particle size of 6.5 μm and a specific surface area (SSA) of 2100 m2/g. The slurry was uniformly mixed by an ultrasonicator and a magnetic stirrer, and then coated on a 30-μm-thick copper foil with a controlled layer thickness of 200 μm. The obtained electrode was punched into disks with a diameter of 14 mm. After vacuum drying overnight at 90 °C, two same disks divided by an NKK cellulose separator were assembled into a symmetrical supercapacitor inside an argon-filled glove box, and the electrolytes used were 1 M tetraethylammonium tetrafluoroborate (TEABF4) in propylene carbonate (PC) solvent. Cyclic voltammetry (CV) and galvanostatic charge-discharge (GCD) were performed using a Biologic VSP-128 potentiostat with a cell voltage range of 0–2.5 V. And electrochemical impedance spectroscopy (EIS) measurements were performed within a frequency range from 100 kHz to 10 mHz to characterize the cell’s internal impedance. The schematic illustration of the whole experiments is depicted in A standard CR2032 coin cell structure was chosen for electrochemical studies. The electrode slurry was prepared by mixing the active material, carbon black, and polyvinylidene difluoride  with a mass ratio of 80:10:10 in N-methyl-2-pyrrolidone (NMP). The active materials comprised activated carbon (AC) and N-doped GE or HGE in a weight ratio of 10:1. The AC was acquired from China Steel Chemical Corporation  with a DTo evaluate the SSA and pore distribution, BET measurement results for the nanocomposites with various AC-to-holey-graphene ratios are shown in θ = 12–14°. After the heat treatment, the characteristic peaks totally disappeared and a broad and much weaker peak appeared at 2θ ~ 26.4°, indicating that GO had been reduced and exfoliated into few-layer graphene, and the melamine in the mixture had decomposed entirely. The TGA  measurement results for melamine and MA-GO , whereas the D band at 1350 cm−1 is associated with the disorders and/or defects. The Raman spectra in ID/IG ratios, indicating high defect density in the graphene lattice caused by the inserted nitrogen atoms as well as the drastic removal of oxygen functional groups during the 1000 °C post heat treatment. Many authors have reported that higher reduction temperature causes more severe damage to the graphene lattice, hence a higher ID/IG [ID/IG increases gradually with the heating rate, which can be attributed to the increase in defect sites along the hole edges created at higher heating rate. Raman spectroscopy was employed to investigate the lattice defects. The G band at 1580 cmer ID/IG ,33. It cThe XPS analysis of nitrogen-doped graphene is shown in 3)—because the low tap density of graphene results in a low capacitance per volume despite its high capacitance per gram. The coin-cell supercapacitors with these electrodes and 1 M TEABF4/PC organic electrolyte were then assembled and tested. The samples were then mixed with AC (at a ratio of 1:10) to be used as the active materials in the electrode . The reason we added a large percentage of AC rather than using purely graphene or holey graphene is to obtain a higher volumetric specific capacitance  [s values, indicating that the overall resistances for these electrodes are similar. The diameter of the semicircle is attributed to the charge transfer resistance (Rct), which is related to the reaction occurring at the contact with electrolyte and is a limiting factor for the specific power of the supercapacitor [s and Rct values of these cells. The smallest semicircle in the EIS plot of AC/N-HGE implies the smallest Rct, which will result in the highest specific power. The numerous holes on N-HGE enable the ions in electrolyte to effectively diffuse into the interior of the composite, thereby reducing the impedance of ion transfer and increasing the overall specific capacitance. The extracted values of Rct for AC/N-GE-15 and AC/N-GE-60 were 6.78 Ω and 8.6 Ω, respectively. These results are consistent with the oxygen content analysis in XPS  . All curapacitor . Table 3s in XPS , i.e., AE-60 see b–d and TE-60 see  althoughNext we varied the 1000 °C thermal annealing time for N-HGE to control the content of residual oxygen functional groups, which would affect the capacitor performances in organic electrolyte. ID/IG ratio increases slightly with the annealing time, indicating an increase in defect sites due to the reduction of oxygen functional groups at 1000 °C. The XPS analysis of the three N-HGE samples is shown in ct decreases monotonically from 6.72 Ω to 4.19 Ω. This is because of the removal of excess oxygen-containing functional groups, which results in better wetting of the N-HGE in the organic electrolyte, thereby improving the electrical conductivity and the electrochemical activity of the AC/N-HGE composite. The energy density and power density performances of these SCs can be evaluated with the Ragone plot. As shown in Though it is difficult to make direct comparison with other methods, we provide a Ragone plot  comparinN-doped holey graphene was synthesized by a cost-effective ultra-rapid heating approach using melamine as the N source, and organic-electrolyte supercapacitors using the composite of AC and N-HGE were fabricated. It was found that holey graphene can effectively enhance the performances of the SCs compared to the non-holey counterparts owing to the shortened ion diffusion paths. Furthermore, N doping can improve the wettability and conductivity of HGE, thus increasing the electrochemical activity and specific capacitance of the composite. However, the residual oxygen functional groups on N-HGE has unfavorable effects in the organic electrolyte. Increasing the annealing time at 1000 °C from 0.5 h to 2 h improved the specific capacitance by 36% at 0.5-A/g current density, and more than 46% at 20 A/g. The SC with the AC/N-HGE-2hr electrodes exhibited the best specific energy at all power density levels, and retain a high specific energy of 11 Wh/kg (or 3.5 Wh/L) at the largest power density of 31,000 W/kg . We believe that the AC/N-HGE-2hr composite prepared by our simple and low-cost method has great potential for high-performance energy storage applications.