In the era of semantic web, life science ontologies play an important role in tasks such as annotating biological objects, linking relevant data pieces, and verifying data consistency. Understanding ontology structures and overlapping ontologies is essential for tasks such as ontology reuse and development. We present an exploratory study where we examine structure and look for patterns in BioPortal, a comprehensive publicly available repository of live science ontologies.We report an analysis of biomedical ontology mapping data over time. We apply graph theory methods such as Modularity Analysis and Betweenness Centrality to analyse data gathered at five different time points. We identify communities, i.e., sets of overlapping ontologies, and define similar and closest communities. We demonstrate evolution of identified communities over time and identify core ontologies of the closest communities. We use BioPortal project and category data to measure community coherence. We also validate identified communities with their mutual mentions in scientific literature.With comparing mapping data gathered at five different time points, we identified similar and closest communities of overlapping ontologies, and demonstrated evolution of communities over time. Results showed that anatomy and health ontologies tend to form more isolated communities compared to other categories. We also showed that communities contain all or the majority of ontologies being used in narrower projects. In addition, we identified major changes in mapping data after migration to BioPortal Version 4. Ontologies are used for tasks such as the standardization of terminology, the verification of data consistency, and the integration of heterogeneous databases. Ontologies have been actively applied to areas including, but not limited to Biology and Medicine , Crisis ontology mappings identify existing concepts with similar meaning. These ontology mappings are useful in tasks like finding new annotations, supporting other data integration methods, combining related ontologies, or ontology reuse. When ontologists build new ontologies they often search for existing ontologies to avoid redundancy of concepts as recommended, for example, by the OBO Foundry principles (Life science inciples . Identifinciples .evolution of ontologies (communities) that tend to overlap more often than others. Please note that in this paper we use the word community for a set of ontologies that tend to overlap. In contrast, the OBO  project defines a community as a set of ontologies that work together and reduce mutual overlap. We also recognize hub ontologies, i.e., ontologies that connect many other ontologies/communities. Since BioPortal data often changes , we analyse the mapping data at different time points. We propose an alignment of similar communities, define stable communities and perform a time transition analysis. Our work aims to answer questions like “In my area of interest, what ontologies already exist and how are they related to each other?” or “In my area of interest, which sub-areas are stable and which are not in terms of ontology development?”. Answering these questions can assist in tasks like ontology reuse and development. The results of the data gathered and analysed at five different time points are presented.In this paper we analyse and evaluate NCBO BioPortal  ontologytologies . To helpRelated work can be categorized in the following (often overlapping) two groups: (1) analysis of ontology mappings and (2) evolution of ontologies. Below we introduce the most relevant papers from these groups.Similar to our work, Changes in ontologies have been previously studied and tools such as GOMMA  have beestable and similar communities), which is the focus of this paper. We also perform analysis of project and category data and discuss alignment with identified communities. In addition, we try to validate generated clusters with information found in MEDLINE abstracts : •NCBO mappings are periodically calculated with a computer algorithm. The algorithm finds mappings for terms with close lexical match or mappings for terms with the same URI from different ontologies. The majority of the mappings is from this group. •Unified Medical Language System (UMLS) mappings link terms with the same UMLS concept unique identifier (CUI) or terms from the UMLS MRMAP.RRF data. •Mappings between ontology terms related by an OBO  xref property.test in their full name, and (3) restricted or private ontologies. From data gathered in December 2013 and July 2014 we removed summary ontologies . The filtered data was then processed with Gephi  ontologies with the retired or alpha status, (2) ontologies that contain the keyword th Gephi , an openfromOntology: the name of the source ontology,(1) toOntology: the name of the target ontology(2) numberOfMappings: number of directed mappings between source and target ontology.(3) Modularity Analysis  to the data. Modularity Analysis (MA) is a measure of structure in graphs. Gephi implements Louvain method  metric  metric  to identWe used information from MEDLINE abstracts , to analWe downloaded the 2016 version of MEDLINE in XML format and developed an algorithm to find pairs of ontology names in all abstracts published before August 2014 (our latest version of the graph is July 2014). Ontology names and abstracts were transformed to lower case characters before the comparison. Simple exact string matching was used to look for ontology names mentions. For example, in the following text “… we introduce GoPubMed, a web server which allows users to explore PubMed search results with the Gene Ontology…”, Gene Ontology would be identified.t contain at least some ontologies from the previous time point t − 1. The challenge is to align similar communities to compare graphs at multiple time points. With aligned communities we can identify communities that changed their size, new communities, or disappearing communities.Running the community detection algorithm at five time points provides us with different number of communities for each time point. Our previous research  showed tThere are several ways to find similar communities in evolving graphs  and no mcommunity core ontologies. In addition, we should consider ontologies that are not shared between two communities. Based on these issues, we first define several terms that are explained in the following paragraphs.Therefore, we wish to use a more stable method for identifying similar communities. We expect that ontologies with the highest BC scores play an important role in BioPortal as they will likely stay in the repository in the future. We call these ontologies C1 contains communities identified at time point t1 and C2 contains communities identified at time point t2 (t2 > t1). First, we define the importance score of ontology o as: BCo,t1 and BCo,t2 represent BC scores for ontology o at time points t1 and t2 respectively.Let us imagine that we identified two groups of communities where group similarity score SCcx,cy between two communities cx ∈ C2 and cy ∈ C1. The similarity score is based on a weighted version of the Dice coefficient  communities and their main hub ontologies, present results of validation with MEDLINE abstracts, analyse transition (evolution) between different time points, analyse the coherence of communities, and present results of measuring effects of ontology sizes on community detection. •MAV represents Modularity Analysis values, •#All is the number of all ontologies in the graph, •#Map is the number of ontologies with at least one mapping (source or target), •%Map is percentage of ontologies with at least one mapping (source or target), •#NoMap is the number of ontologies with no mappings, •%NoMap is the percentage of ontologies with no mappings, •x < 8) values represent the number of ontologies in each identified community. The number of ontologies in each community orders communities.#Com is the number of identified communities. The #Cx , we can notice that pairs where each ontology belongs to a different community tend to be found in lower number of abstracts . On the other hand, ontology pairs that can be found together in large numbers of abstracts  tend to belong to the same community.We found 3,020 ontology pairs  that were mentioned together in at least one abstract. These results imply that identified communities contain ontologies that appear more often together in the literature. However, since most ontologies pairs were not found in the abstracts, different methods should be explored . This is an area for future investigations.NoMap row represents ontologies that had no mappings in the previous version of the graph but are members of one of the communities in the newer version. The New row represents ontologies that did not exist in the previous version of the graph.When observing heat maps on For example, let us consider the heat map A . Row names represent hub ontologies for the old version (Oct12), while column names represent ontologies for the new version (Feb 13) of the graph. The third row shows that all ontologies from the UBERON community stayed in the same community, i.e., the community did not split. On the other hand, observing row 2 shows that although the majority of Oct12 NCIT ontologies stayed in the closest community in Feb13 , some ontologies also migrated into the NIF (column 1), UBERON (column 3), RADLEX (column 4) and SNOMEDCT (column 5) communities. The third column illustrates merging of parts of three different communities  into the new UBERON community. The heat map A also shows that the new identified community  mainly consists of ontologies from the old EP (row 1) community and some ontologies that had no mappings in Oct12 (row 6).With the heat maps we can find pairs of closest communities, which are identified with the most intensive shades of green in each column. For example, on the heat map A, the NIF column contains three coloured squares. However, the square in the EP row is the most intensive shade of green, which identifies the closest community to NIF. In An interesting community is the NCBITaxon community, which appears the Feb13 and Dec13 versions. We already learned that some taxonomy ontologies formed their own community in February 2013 . HoweverConsidering categories such as Anatomy, Health, Ethology, and Gene Product. In addition, information about projects that use BioPortal ontologies is available. We use these two types of information to discuss the coherence of identified communities in our methods. BioPortal groups ontologies into 41 Charts on Another distinct community is G2, where a large proportion of ontologies belongs to the Health category. Almost 70% of Oct12 G2 ontologies are categorised as health ontologies and present the majority in G2 future graphs as well. Health ontologies are distributed through other identified communities in the future and present large portions of G4 and G6. Communities G1 and G3 are more heterogeneous with a mixture of ontologies from all categories in all graph versions.We also investigated the BioPortal project data to analyse its alignment with identified communities. Each project has a list of ontologies that it uses and we investigated how these lists correspond to the identified communities for the Jul14 version. Project with the highest number of ontologies used 35 ontologies, while the majority of projects used a single ontology. We ignored the latter projects in our analysis since it was obvious that they will be aligned with a single community. An important factor that influences the number of mappings between two ontologies is the size  of both ontologies. It is more likely that larger ontologies have higher number of mappings when compared to smaller ontologies. Unfortunately, we did not collect ontology sizes for each time spot in our analysis and historical data is not available through BioPortal’s API. We downloaded old versions of ontologies at the time of writing this paper and tried to manually parse the ontologies with the OWL API to calculate their sizes. The OWL API is a Java API and reference implementation for creating, manipulating and serialising ontologies . HoweverIn this paper we focused on investigating a comprehensive repository of biomedical ontologies  using graph theory concepts. We performed the exploratory study of BioPortal’s mapping data over different time points. As far as we know, this is the first attempt of this kind. With investigating mapping data gathered at five different time points using graph theory methods, we identified similar and closest communities of overlapping ontologies, and demonstrated evolution of communities over time. We also tried to validate communities through mentions of their ontology members in MEDLINE abstracts.The five communities identified in the first version of the graph changed their size. We showed how communities appear, disappear, split or merge over time. Based on similarity scores we determined closest communities between pairs of different graph versions. We then analysed the stability of these closest communities. We discussed how identified communities align with BioPortal’s category and project information. We also identified core ontologies of the closest communities.When studying our conclusions, we should take into consideration some limitations of the work. First, the BioPortal repository can be publicly modified and no evaluation of the uploaded ontologies or mapping data is done. In addition, although we tried to identify them, there are probably some “test” ontologies left in our data. Therefore, we should expect some data noise. Our analysis also showed large differences in data between the Aug13 and Dec13 when BioPortal 4 was announced. Second, our method for identifying communities might favour larger ontologies since we do not consider ontology sizes when calculating edge weights. Although our analysis of data gathered in November 2015 implies that normalising edge numbers results in small changes in final graph, this remains an area for future investigation. Next, due to limitations of BioPortal’s web service API, we were not able to distinguish between different types of ontology mappings in older versions. For example, the MESH and RH-MESH ontologies have same concepts and only differ in syntactic translation, which has not been picked up by our methods. Finally, our observations highly depend on the Louvain method for community detection. We accept this method as a “ground truth” quality metrics of our clusters. The Louvain method was the only available method in Gephi and it is considered as the fastest and most accurate method in terms of modularity score .In the future, we plan to address the above issues, especially distinguishing between different types of mappings and considering ontology sizes. We also plan to consider other graph centrality measures and methods for community detections. Finally, we plan to perform a deeper analysis of changes in the underlying ontologies to investigate how these affect the broader graph clustering patterns.10.7717/peerj.2990/supp-1Supplemental Information 1Mappings between pairs of ontologies for different versions of graphs.Click here for additional data file.


High-resolution transmission electron microscopy (HRTEM) and selected-area electron diffraction (SAED) confirm the formation of the GO layer structure and the hexagonal framework. This is a promising method for fast and effective synthesis of GO from sugarcane bagasse intended for a variety of energy and environmental applications.A new method of graphene oxide (GO) synthesis via single-step reforming of sugarcane bagasse agricultural waste by oxidation under muffled atmosphere conditions is reported. The strong and sharp X-ray diffraction peak at 2θ = 11.6° corresponds to an interlayer distance of 0.788 nm ( Graphenree form , thermalree form , and higree form ,6,7,8.4 and NaNO3 in concentrated H2SO4 acid.Graphene oxide (GO) has also attracted major interdisciplinary attention because of the broad range of envisaged applications across several scientific and engineering fields including physics, chemistry, biology, and medicine ,11,12,132, N2O4, and ClO2, toxic gases which are also explosive. In the past few years, various techniques such as mechanical exfoliation techniques, chemical vapor deposition (CVD) techniques and other chemical techniques have been developed for the synthesis of graphene and graphene oxide materials. However, many of these techniques are highly sophisticated and expensive [All these procedures involve the generation of NOxpensive . Most ofxpensive ,19.Here we report on the synthesis of graphene oxide by directly oxidizing sugarcane bagasse under muffled atmosphere. The GO produced by this method is called SOMA-GO (sugarcane oxidized under muffled atmosphere for graphene oxide). Furthermore, the structural characteristics of the obtained product are confirmed by X-ray diffraction, Fourier transmission infrared spectroscopy, field emission scanning electron microscopy, high-resolution transmission electron microscopy, and Raman spectroscopy.Sugarcane bagasse is a type of agricultural waste (agro waste). After juice was extracted, the remaining fiber was taken. The fiber was crushed and ground well in order to produce powder. This crushing and separating process was repeated several times to obtain fine powder. About 0.5 g of ground sugarcane bagasse powder mixed with 0.1 g of ferrocene was placed in a crucible and put directly into a muffle furnace at 300 °C for 10 min under atmospheric conditions. Next, the as-produced black solid was collected at room temperature. The black solid product was subjected to further analysis .−1 scan rate. Fourier transform infrared spectroscopy (FT-IR), Avatar 370, Thermo Nicolet  was carried out to study the bonding configuration in the synthesized graphene oxide. High resolution transmission electron microscopy (HRTEM) was performed using a JEOL JEM-3000F microscope  operating at 300 kV to study the microstructure of the GO structures. Field emission scanning electron microscopy was carried out using a JEOL JSM 7800F revealed the morphological features of the GO structure.The X-ray diffraction (XRD) pattern was obtained using a Bruker D8 Advance X-ray diffractometer . This device is equipped with a Ni filter, and generates monochromated Cu-Kα radiation (λ = 0.154 nm) operated at 40 kV accelerating voltage and 30 mA current. The samples were scanned in step mode with a 2° minThe structural and chemical analysis of sugarcane bagasse is clearly described by previous investigators ,21,22. I−1 is attributed to stretching vibration modes of C=O in carboxylic acid and carbonyl groups. The peak at 1594 cm−1 is assigned to the skeletal vibrations of un-oxidized graphitic domains. The band at 1060 cm−1 is assigned to C–O (epoxy) groups while the band at 1210 cm−1 is usually attributed to C–OH stretching vibrations [−1 can be attributed to the O–H stretching vibrations of the C–OH groups and water [The FT-IR spectrum of GO shown in brations ,26,27. TThe surface morphology of the graphene oxide (GO) produced by sugarcane bagasse oxidation is identified by FESEM. The SEM image of the GO shown in sp3 hybridized carbon atoms may also be introduced during the oxidation process. HRTEM micrographs of GO are shown in −1, which are attributed to the local defects (found at the edges of graphene sheets) and the 2sp graphitized structure, respectively. The G peak formation is due to the double degenerated zone center E2g mode [sp2 plane domains are possibly due to the extensive oxidation of sugarcane bagasse [D and G bands at 1358 cm−1 and 1585 cm−1 [sp2 carbon hybridization in the observed multi-layer stacks [DI/GI peak intensity ratios are assigned to lower defects/disorders. Raman spectra show an intensity ratio of DI/GI at 0.76 for GO which is in line with previous investigations [The Raman spectrum of graphene oxide synthesized from sugarcane bagasse is shown in E2g mode ,35. The  bagasse . The two585 cm−1  indicater stacks . Thus, Iigations ,37,38.A simple and rapid method is shown here to effectively convert solid sugarcane bagasse waste into value-added graphene oxide. The produced graphene oxide presents a well-graphitized structure. In comparison with the commonly used methods, our method is more environment friendly as it avoids toxic gas emission during the synthesis. This simple and low-cost synthesis process could lead to new opportunities for cost-effective production of graphene-based materials for gas sensors, energy storage, and other functional devices.


Commercialization of graphene based applications inevitably requires cost effective mass production. From the early days of research on graphene, direct liquid phase exfoliation (LPE) of graphite has been considered as the most promising strategy to produce high-quality mono or few-layer graphene sheets in solvent dispersion forms. Substantial success has been achieved thus far in the LPE of graphene employing numerous solvent systems and suitable surfactants. This invited review article principally showcase the recent research progress as well as shortcomings of surfactant assisted LPE of graphene. In particular, a comprehensive assessment of the quality and yield of the graphene sheets produced by different categories of the surfactants are summarized. Future direction of LPE methods is also proposed for the eventual success of commercial applications. Later in 2011, Nuvoli et al. reported an unprecedented graphene concentration as high as 5.33 mg/mL by sonicating a commercially available IL 1-hexyl-3-methyl imidazolium hexafluorophosphate (HMIM)  are semi-organic molten salts, composed of ions, which exhibit highly viscous liquid behaviour . ILs genVery recently, Texter and co-workers developed two excellent water stabilizers for graphene viz., triblock (TB) copolymer and copolymer nanolatex (NL)  to facilitate the phase transfer of graphene sheets from water phase to organic phase via simple sonication   (PS-b-P2VP)   , poly as well. Notley, in a similar study, compared pluronic non-ionic surfactants, F108 (molecular weight ~ 14.6 kDa) and F127 (molecular weight ~ 12.5 kDa) with some ionic surfactants such as CTAB (hexadecyltrimethylammonium bromide), TTAB (tetradecyltrimethylammonium bromide), DTAB (dodecyltrimethylammonium bromide) and SDS   (polyPA-b-polyPEG-A)  polymerization and graphite powder mixed with different amounts of aqueous copolymer solutions were sonicated for 6 h at 30 °C to provide upto 78 % graphene yield when the copolymer to graphite ratio reached 40. This amphiphilic polymer design with multi-pyrene groups was proposed to bind at the graphite surface simultaneously via π-π stacking, working like “sucking discs” to drag the bound surface graphene sheet off the graphite precursor with the assistance of sonication. The as-prepared graphene/polymer composite films also exhibited increased tensile strength and tunable conductivity. Direct exfoliation of graphite flakes in the presence of pyrene-labeled single stranded DNAs yielded highly aqueous dispersible mono- and bi-layer graphene sheets with 100 nm to 4 μm flake size [As mentioned above, Pyrene derivatives have been widely investigated as small molecule conjugate stabilizers. Concurrently, a few reports on pyrene-based polymers are also exploited as efficient graphene exfoliant. One typical report is from Zheng et al., wherein supercritical (SC) COsability . Specialake size . SubsequHighly conductive and transparent graphene films were fabricated by Jo et al. from direct exfoliation of graphite using a non-ionic semiconducting polymer quinquethiophene-terminated PEG (5TN-PEG) . Relevant crucial processing factors such as solvents, external forces like ultrasonication/shear and purification methods by centrifugation have been discussed in detail in association with their influences on the exfoliation results. Significantly, most of the solvents used in the initial studies had revealed significant drawbacks, such high toxicity, high boiling point etc. that prompted the re-direction of research into environment benign less toxic solvents like water. Unfortunately, the surface energy of pure water is too high for graphene exfoliation such that a variety of surfactants have been introduced thus far. We categorized the large spectrum of surfactants in accordance to their structural functionalities into aromatic, non-aromatic, ionic liquids and polymers. Innumerable surfactants have been studied in this regard and many of them appeared highly promising LPE results.For the further progress of surfactant assisted LPE, several shortcomings must be overcome: (1) The overall yield of LPE is still low; (2) Good exfoliating solvents are expensive and harmful; (3) Sonication/Shearing commonly lead to the drastic reduction in the size of exfoliated graphene sheets; (4) Residual surfactants are difficult to remove; (5) Typical surfactants are electrically insulators, which may significantly deteriorate the electrical connectivity among graphene layers; (6) All LPE methods produce graphene sheet with a high polydispersity in terms of lateral size as well as thickness. The future of real-life graphene applications strongly depends on how materials scientists address these formidable challenges and establish ideal large-scale LPE process for high quality graphene sheets. It is also highly required to attain more fundamental and systematic understanding of the exfoliation mechanism for innovative design of LPE schemes.


Cordyceps belongs to a genus of acormycete fungi and is known to exhibit various pharmacological effects. The aim of this study was to investigate the effect of Cordyceps species on the proliferation of vascular smooth muscle cells (VSMC) and their underlying molecular mechanism. A cell proliferation assay showed that Cordyceps bassiana ethanol extract (CBEE) significantly inhibited VSMC proliferation. In addition, neointimal formation was significantly reduced by treatment with CBEE in the carotid artery of balloon-injured rats. We also investigated the effects of CBEE on the extracellular signal-regulated kinase (ERK) signal pathway. Western blot analysis revealed increased ERK 1/2 phosphorylation in VSMCs treated with CBEE. Pretreatment with U0126 completely abrogated CBEE-induced ERK 1/2 phosphorylation. In conclusion, CBEE exhibited anti-proliferative properties that affected VSMCs through the ERK1/2 MAPK signaling pathway. Our data may elucidate the inhibitory mechanism of this natural product. Atherosclerosis is a disease characterized by thickening of the inner portion of the artery walls . One of Cordyceps belongs to a genus of acormycete fungi that includes about 400 species [Cordyceps species are C. sinensis and C. militaris, which are have been used as traditional medicine for various diseases in Asian countries. These exhibit various pharmacological effects including anti-inflammatory, antioxidative, antidiabetic, and anticancer properties [ species . RepreseCordyceps species have yet to be addressed. We investigated the anti-proliferative activity of Cordyceps species extracts in VSMC and studied the underlying signal transduction mechanism. Therefore, our study investigates that the molecular mechanism underlying the anti-atherosclerotic effects of Cordyceps species extracts. This pathway might highlight a potential therapeutic intervention for cardiovascular disease.Cordycepin inhibits VSMC proliferation . HoweverCordyceps bassiana ethanol extract (CBEE), Cordyceps militaris butanol extract (CMBE), and Cordyceps pruinosa methanol extract (CPME) were extracted using conventional extraction methods [ methods . The sigCordyceps (175 g) were powdered and extracted at 70 ~ 80 °C with 80 % ethanol, methanol and butanol. The extract was filtered, taken to dryness under reduced pressure and concentrated by a rotary vacuum evaporator . The lyophilized crude extracts were sequentially fractionated with n-hexane (Hexane), ethyl acetate (EtOAc) and n-butanol (BuOH). Each extract was dissolved in dimethyl sulphoxide (DMSO) as a concentration.Solvent extraction was performed as described previously , 7. In b2 flasks in a 37 °C incubator at 5 % CO2.Rat aortic smooth muscle cells (RaoSMCs) were cultured in DMEM supplemented with fetal bovine serum  in 75 cm3 cells per well. The cells were starved with 0.1 % FBS for 24 h and then treated with three kinds of Cordyceps extracts or 10 % serum. After treatment, cell proliferation was measured using a CCK-8 assay kit . The CCK-8 assay kit was treated with DMEM, and then 100 μl was added to each well and incubated for 2 h at 37 °C. The absorbance was measured at 450 nm with a spectrometer.RaoSMCs were plated in triplicate in 96-well plates at 2 × 10Animal experiment conforms to the Animal Care and NIH guidelines were approved by the local animal care committee. Balloon denudation of the common carotid artery endothelium was evoked in four-week old 250 g Male Sprague–Dawley rats  as previously reported . All ratSprague–Dawley rats were sacrificed 3 weeks after the balloon injury. And their carotid arteries were harvested and fixed with 4 % paraformaldehyde. Within 24 h after fixation, each section was embedded in OCT compound , snap frozen in liquid nitrogen, and sectioned in thickness increments of 10-μm carotid artery sections were cut with a microtome and mounted on slides. Sections were stained with hematoxylin and eosin (H&E) and evaluated with light microscopy to assess the histological effects. Normal and neointimal areas were measured using NIH images.The western blot analysis was performed as previously described , 10. Celt-test for unpaired samples between two groups. p < 0.05 was considered statistically significant.All data are presented as mean ± standard deviation (SD) from at least three independent experiments. The statistical significance was determined using the Student’s To confirm the effect of CBEE on VSMC anti-proliferation, cells were cultured for 24 h in complete medium containing 10 % FBS at various concentrations of CBEE (0 ~ 4 μg/ml). As shown in Fig. Cordyceps species, VSMC were cultured for 3 days in complete medium containing 10 % FBS with three Cordyceps species: CBEE, CMBF, and CPME. A cell proliferation assay showed that CBEE significantly inhibited VSMC proliferation   treated group exhibited a significant reduction of neointima formation compare with control (vehicle) group (19) Fig. .Fig. 2EfTo better understand the molecular mechanisms involved in the VSMC anti-proliferative effects of CBEE, we investigated the possible involvement of the MAPK signaling pathway. To determine whether extracellular signal–regulated kinase (ERK) 1/2 activation is involved in CBEE-induced VSMC proliferation, we treated cells with the MEK/ERK inhibitor U0126 for 1h and then stimulated with CBEE for 72 h. Proliferation assays were then performed with the CCK-8 assay. The ERK1/2 inhibitor U0126 significantly blocked CBEE-mediated VSMC anti-proliferation in a dose-dependent manner, as examined by cell counting Fig. .Fig. 3EfOur previous results prompted us to further investigate whether CBEE activates the ERK 1/2 signaling pathway. To address this issue, cell lysates were probed for ERK 1/2 phosphorylation and protein levels. Western blot analysis revealed increased ERK 1/2 phosphorylation in VSMCs treated with 4 μg/mL CBEE after 3, 6, and 24 h, with the maximal increase occurring after 3 h of treatment  ERK 1/2 phosphorylation Fig. .Fig. 5TiOur results clearly demonstrate that CBEE inhibits VSMC proliferation. Notably, CBEE inhibited VSMC proliferation by modulating the ERK1/2 MAPK signaling pathway. To the best of our knowledge, this is the first report connecting molecular mechanisms of CBEE to its anti-proliferative effects on VSMC.Endothelial injury initiates the pathogenesis of various vascular diseases such as atherosclerosis . ChronicCordyceps species on VSMC are not fully understood. We found CBEE, which contained a yellowish colored compound, was reported to have significant anti-proliferative effects on VSMC. However, data on the biological activities of CBEE are limited. One recent report demonstrated that CBEE suppressed the expression of interleukin-12 and phosphorylation of p38 in lipopolysaccharide-activated macrophages without altering cell viability [Cordyceps species extracts (CMBF or CPME) could exhibit inhibitory effect on VSMC. However, major active compounds derived from CBEE and their anti-proliferative or anti-inflammatory properties compare with CMBF or CPME should be clarified in further studies.Recent reports indicate that cordycepin inhibits VSMC proliferation  and atteiability . InteresAccording previous reports, ERK1/2 has distinct functions in the inhibition of cell growth . Jung etCordyceps bassiana, and helps us to understand its protective effects in cardiovascular disease-associated pathophysiology. However, further studies are necessary to identify the major active components of CBEE.In conclusion, CBEE exhibited anti-proliferative properties that affected VSMC through the ERK1/2 MAPK signaling pathway. Our data may elucidate the inhibitory mechanism of this natural product,


To determine the optimal extraction conditions, a number of experimental factors  were studied. The compounds of interest were separated and detected by an HPLC-UV employing a Kinetex XB-C18 column and a mobile phase consisting of acetonitrile and water flowing in a gradient mode. The optimized extraction conditions were: the amount of graphene 15 mg, desorbing solvent (acetonitrile) 5 mL, time desorption 10 min at 40°C, and amount of NaCl 1 g. The detection limit for metazachlor, tebuconazole, λ-cyhalothrin, and chlorpyrifos was 62.5 ng·g−1, and for deltamethrin, it was 500 ng·g−1. The obtained results lead to the conclusion that graphene may be successfully used for the isolation of the five pesticides from rape oil. However, their determination at low concentration levels, as they occur in real oil samples, requires the employment of appropriately highly sensitive analytical methods, as well as a more suitable graphene form .Isolation conditions for five pesticides (metazachlor, tebuconazole,  Due to the potential toxicity, bioaccumulating properties, and wide use of pesticides for supporting and intensification of crops almost all over the world, these substances belong to the most often controlled ones in the environment and in food. Every year, many publications devoted to the development of new methods for pesticide determination in various food matrices appear. However, pesticide residue analysis in high-fat samples is still a challenging analytical task. Edible oil samples belong to such analytically difficult matrices. Due to complex matrices, as well as low concentrations of pesticide residues, frequently originated from various classes, these samples are analyzed using such advanced coupled techniques as LC-MS/MS , 2, LC-Tπ-π interactions between its large delocalized electron system and the aromatic rings of the target molecules In most cases of pesticide analysis involving graphene as an adsorbent, magnetic solid-phase extraction (MSPE) was applied where magnetically modified graphene composites as adsorbents were used. Only few works reported the employment of nonmodified graphene alone or in combination with other sorbents (in the modified QuEChERS method) using dispersive solid-phase extraction. This is the first time when graphene in its nonmodified form, using d-SPE, was studied as the adsorbent for the pesticides from four different groups present in such analytically difficult matrices as edible oil. Graphene may be successfully exploited in the preparation of edible oil samples for determination of pesticides from various groups. However, some improvements should be introduced in such analyses. For example, a more sensitive analytical method  may be used. Additionally, a more suitable modified graphene form, for example, magnetically modified graphene, which will prevent the nanomaterial aggregation, increase its dispersion in solvents, and improve its adsorption properties, would better enable separation of graphene from the isolated analytes in the supernatant using an external magnet.


Botrytis cinerea in Arabidopsis thaliana and Camelina sativa overexpressing DEWAX and in Arabidopsis dewax mutant. Compared to wild type (WT) leaves, Arabidopsis DEWAX OX and dewax leaves were more and less permeable to toluidine blue dye, respectively. The ROS levels increased in DEWAX OX leaves, but decreased in dewax relative to WT leaves. Compared to WT, DEWAX OX was more resistant, while dewax was more sensitive to B. cinerea; however, defense responses to Pseudomonas syringae pv. tomato DC3000:GFP were inversely modulated. Microarray and RT-PCR analyses indicated that the expression of defense-related genes was upregulated in DEWAX OX, but downregulated in dewax relative to WT. Transactivation assay showed that DEWAX upregulated the expression of PDF1.2a, IGMT1, and PRX37. Chromatin immunoprecipitation assay revealed that DEWAX directly interacts with the GCC-box motifs of PDF1.2a promoter. In addition, ectopic expression of DEWAX increased the tolerance to B. cinerea in C. sativa. Taken together, we suggest that increased ROS accumulation and DEWAX-mediated upregulation of defense-related genes are closely associated with enhanced resistance to B. cinerea in Arabidopsis and C. sativa.The cuticle of land plants is the first physical barrier to protect their aerial parts from biotic and abiotic stresses. DEWAX, an AP2/ERF-type transcription factor, negatively regulates cuticular wax biosynthesis. In this study, we investigated the resistance to  Among tynthesis . The APE and ECR . HoweverCamelina sativa, which belongs to the Brassicaceae family, has recently gained increasing interest for its potential non-food usages, including in cosmetics, lubricants, and as biofuels (C. sativa has a short life cycle (100–120 days), and can be grown on marginal agricultural lands owing to its characteristics, such as tolerance to cold stress, relatively good growth under nutrient-poor soil conditions, and high levels of polyunsaturated fatty acids in the seed oils . To understand the molecular mechanisms underlying the defense responses of Arabidopsis and C. sativa plants overexpressing DEWAX to B. cinerea, further analyses were carried out. Finally, we revealed that DEWAX plays as a transcriptional activator that upregulates the expression of the defense-related genes such as PDF1.2a, IGMT1, and PRX37 via binding to their gene promoters. The results suggest that DEWAX-mediated upregulation of defense-related genes in addition to an increase in ROS levels is important in the enhanced tolerance to B. cinerea in Arabidopsis and C. sativa overexpressing DEWAX.In this study, we observed that DEWAX overexpression lines (OX1 and OX2) and dewax knockout mutant were described in A. thaliana (ecotype Columbia-0) seeds were surface-sterilized with 75% (v/v) ethanol and 0.05% (v/v) Triton X-100, washed with 100% ethanol, dried on filter paper, and germinated on half MS  at 25 ± 3°C under a 16-h-light/8-h-dark cycle , and 6-week-old transgenic C. sativa lines (T1 generation) using the NucleoSpin RNA Plant Extraction Kit (Macherey-Nagel) according to the instructions of the manufacturer. The total RNA was reverse-transcribed with GoScriptTM Reverse Transcription System (Promega), and the cDNA was used in RT-PCR analysis. RT-PCR was performed using the Access QuickTM RT-PCR system  with gene-specific primers (Supplementary Table Arabidopsis EIF4 (At3g13920) and C. sativa ACTIN11 genes  in a final volume of 20 μL. PCR was performed according to the manufacturer’s protocols using gene specific primers described in Supplementary Table PP2A (At1g13320) and CsACT11 genes were used to determine the RNA quality and quantity.Total RNA was isolated from 4-week-old Col-0, 11 genes  were useArabidopsis rosette leaves were stained at room temperature for 8 min without shaking using freshly prepared staining solution containing 0.05% toluidine blue and 0.01% Tween-20. After staining, leaves were rinsed five times with distilled water and photographed.Four-week-old in situ detection of hydrogen peroxide, Arabidopsis leaves were immersed in 10 mM MES (pH 6.5) containing 0.1% 3,3′-diaminobenzidine tetrahydrochloride , gently vacuum-infiltrated, and incubated for 30 min as described in Arabidopsis leaves were incubated in 10% MS salt solution containing 20 μM 2′,7′-dichlorofluorescein diacetate  and 0.1% Tween-20 in the dark for 30 min. DCF fluorescence in aliquots taken from the incubation solution was measured using a Synergy H1 Hybrid Multi-Mode Microplate Reader (BioTek) at 488 and 525 nm. Leaf fresh weights were used to normalize the values (2O2) content, 5-week-old leaves (200 mg) were incubated in 1.5 mL of 5 mM morpholineethanesulfonic acid (MES)-KOH buffer (pH 6.7) for 7 h  and was cultured on 2× V8 tomato agar . Harvesting and inoculation of conidia were carried out according to Arabidopsis leaves were laid out on 3MM paper with water and inoculated with 5-μl droplets of conidial suspension (5 × 105 spores/mL), kept under a sealed transparent cover to maintain high humidity and transferred to a growth chamber . C. sativa plants were sprayed with B. cinerea conidial suspension (2 × 105 spores/mL). After inoculation, the plants were transferred to growth chambers  and covered with a sealed clear lid to maintain low light intensity and high humidity. Quantification of B. cinerea biomass in C. sativa was carried out according to Pto DC3000 strain harboring pDSK-GFPuv  containing 0.05% Silwet L-77  for few seconds. After inoculation, the plants were covered with a clear lid and transferred to a growth room . Inoculated plants were photographed under ultraviolet light to determine PtoDC3000:GFP proliferation using GFP signals and bacterial populations were measured by serial dilution assays (A n assays . Colony DEWAX-F (603 bp for full-length), DEWAX-N , and DEWAX-C  DNA fragments were amplified via PCR using DEWAX cDNA and DEWAX-specific primers (Supplementary Table SmaI and SacI-digested PCR fragments were cloned into the pGBKT7 vector (BD Biosciences Clontech). The recombinant vector  were transformed into yeast strain Y190  according to the manufacturer’s instructions (BD Biosciences Clontech). Transformants were selected on selective medium (SD-Trp) supplemented with 25 mM 3-amino-1, 2, 4-aminotriazole (3-AT). Transformants were cultured on selective medium (SD-Trp-His) for 1 day at 30°C and filter-lift assays were performed for 16 h at 37°C, as described in For the transcactivation assay in yeast, SalI and SpeI sites of the GAL4-LUC binary vector. The recombinant pPZP212 vector , IGMT (∼1.5 kb), PRX37 (∼1.2 kb) promoter regions were amplified using gene-specific primers and cloned into the 2 vector  harborinThe reporter and effector constructs were isolated using a Qiagen Plasmid Midi Kit and co-introduced into tobacco protoplasts . After iArabidopsis actin7 (At5g09810) was used to control cDNA quantity and quality.Chromatin immunoprecipitation (ChIP) assay was performed using 35S:MYC-DEWAX transgenic plants, which were previously reported . GenomicC. sativa plants overexpressing Arabidopsis DEWAX, the full-length DEWAX gene was amplified from cDNA converted from RNA isolated from 3-week-old leaves with DEWAX F1 and DEWAX R1 primers  BASTA herbicide.onstruct  was inocp method . T1 tranC. sativa plants (T1 generation) using extraction buffer  and subjected to genomic DNA PCR analysis with CaMV35S promoter F and DEWAX CR1 primers . When hydrogen peroxide content was spectrophotometrically measured after leaves of WT, dewax, and DEWAX OX lines were preadapted in incubation buffer, the levels of hydrogen peroxide were approximately twofold higher in DEWAX OX1 and OX2 leaves than in WT leaves, but no remarkable differences were detected between WT and dewax leaves .To investigate the cuticle permeability of WT, S levels , ROS assdewax, and two DEWAX OX lines to the necrotrophic pathogen B. cinerea and the hemibiotrophic bacterial pathogen Pto DC3000, first, we examined visible disease symptoms 72 h after inoculation of B. cinerea on the adaxial surface of the leaves. In a spot inoculation assay, significantly larger lesions were observed on the leaves of dewax mutant, while smaller lesions were formed on DEWAX OX as compared with WT leaves , indicating that DEWAX OX shows enhanced resistance, and dewax enhanced susceptibility, to B. cinerea. On the other hand, compared to WT, DEWAX OX was more susceptible to Pto DC3000:GFP, while dewax was more resistant to this pathogen . These results indicate that DEWAX is involved in the disease resistance responses of Arabidopsis and plays a positive role in the defense to fungal pathogens, but a negative role in the defense to bacterial pathogens.To investigate the response of WT, Arabidopsis, comparative transcriptome analysis of WT and DEWAX OX was conducted (http://www.ebi.ac.uk/arrayexpress). Interestingly, the expression of genes involved in pathogen defense and ROS production was clearly upregulated in the stems of DEWAX OX as compared to those of WT  and peroxidase C2 precursor (PRX37 and PRX38) were increased in DEWAX OX1 and OX2 leaves, while no significant downregulation was observed in dewax relative to WT. No noticeable differences in the expression of At1g67810 and At4g16260, encoding SULFUR (Stimulate CpNifS Cysteine Desulfurase Activity) and a β-1,3-glucanase precursor, respectively, were observed in dewax, and DEWAX OX1 and OX2 leaves .DEWAX has been reported to function as a transcriptional repressor in cuticular wax biosynthesis . However1 vector  and usedDEWAX were cloned downstream of the GAL4-binding domain in the pGBKT7 vector. Yeast Y190 cells, which possess the HIS3 and lacZ genes driven by the promoter including GAL4-responsive elements, were transformed with the constructed vectors. Yeast cells that were able to survived on tryptophan-deficient selective medium (SD/-Trp) were further selected on tryptophan- and histidine-deficient medium (SD/-Trp-His) supplemented with 10 mM 3-amino-1, 2, 4-aminotriazole (3-AT) and subsequently subjected to an X-gal filter-lifting assay. Yeast cells transformed with the F or N construct survived on SD/-Trp-His medium supplemented with 10 mM 3-AT, and generated blue colonies by the β-galactosidase activities in X-gal lift assay. However, yeast cells transformed with the pGBKT7 vector or the C construct did not survive on the selective medium, and showed no β-galactosidase activity. These results indicate that DEWAX is able to act as a transcriptional activator, and amino acid residues 1 to 106 of DEWAX are involved in transcriptional activation .In addition, the full-length , N-terminal , and C-terminal  regions of PDF1.2a, IGMT1, and PRX37, their promoter regions were transcriptionally ligated to the luciferase gene (LUC) in the Gal4-LUC binary vector, and used as reporter constructs . p35S and p35S-DEWAX as effector constructs were used in transactivation assay of tobacco protoplasts  and β-glucuronidase (GUS) activities were measured, and the LUC activity was normalized to the GUS activity. When DEWAX was expressed, LUC activity was upregulated in the protoplasts transformed with PDF1.2a, IGMT1, and PRX37 reporter constructs by approximately 2.3, 4.2, and 6.3-fold, respectively, but was downregulated in the protoplasts transformed with CER1 by approximately twofold relative to the control co-transformed with p35S .To examine whether DEWAX is able to activate the expression of the putative target genes, toplasts . After tPDF1.2a promoter region using 35S:MYC-DEWAX transgenic plants . This observation indicates that DEWAX activates the expression of PDF1.2a gene by direct interaction with its promoter region.We further investigated whether DEWAX directly interacts with the consensus GCC-box motifs of c plants . QuantitDEWAX overexpression in Arabidopsis led to strong resistance to B. cinerea, we next evaluated how overexpression of DEWAX in a Brassica crop, C. sativa, affects disease resistance. The 35SP-DEWAX construct harboring DEWAX gene under the control of CaMV 35S promoter was used in the transformation of C. sativa . Transgenic plants were screened through spraying of 0.03% BASTA herbicide . Genomic DNA PCR analysis of non-transgenic (NT) and 16 transgenic plants (TO) using gene-specific primers . To measure B. cinerea biomass, genomic DNA was isolated from the leaves 4 days after inoculation with B. cinerea and subjected to real-time quantitative PCR using B. cinerea-specific cutinase A-specific gene primers (Supplementary Table C. sativa GSK3/shaggy-like (iASK) gene was used for normalization. Compared to NT plants, all TO lines tested had lower B. cinerea biomass . Subsequently quantitative RT-PCR analysis showed that the levels of C. sativa PDF increased by 2- to 3.5-fold in DEWAX OX lines compared with NT . These results indicated that overexpression of Arabidopsis DEWAX confers tolerance to B. cinerea in transgenic C. sativa.Next, we examined the defense responses of NT and TO plants to DEWAX increased cuticle permeability and ROS accumulation in Arabidopsis. DEWAX is a transcriptional activator that upregulates the expression of the defense-related genes, PDF1.2a, IGMT1, and PRX37, by binding to their gene promoters. In particular, DEWAX directly interacts with the GCC motifs in the PDF1.2a promoter. Overexpression of DEWAX conferred enhanced tolerance to necrotrophic fungal pathogen, B. cinerea in Arabidopsis and C. sativa. Therefore, convergent line of evidence supports that increased ROS levels as well as upregulation of defense-related genes might be related to the resistance to B. cinerea in Arabidopsis and C. sativa overexpressing DEWAX.Plant immune responses are essential for not only growth and development, but also productivity and yields of crops. In this study, we investigated a plant defense mechanism related to the cuticle, which is the first defensive barrier of the aerial parts of plants. We revealed that overexpression of OCTADECANOID-RESPONSIVE ARABIDOPSIS (ORA)59 and ERF96 with the “EDLL” motif to the two GCC motifs in the PDF1.2 promoter region, is essential for the expression of PDF1.2 , which is consistent with the results in the Arabidopsis mutants, bdg and lacs2  and ethephon, a form of liquid ethylene (PDF1.2 depended on ERF1 and ORA59 transcription factors, which act as integrators of JA and ethylene signaling pathways. Overexpression of ERF1 and ORA59 caused increased resistance, and ora59 mutant enhanced susceptibility to B. cinerea (DEWAX were elevated neither by JA nor by ethephon1, although DEWAX is classified with ERF1 and ORA59 into the group IX of Arabidopsis AP2/ERF gene family (Arabidopsis defense responses to herbivores and pathogens are regulated by circadian rhythms (PDF1.2 by the diurnally controlled DEWAX may be important in circadian clock-mediated plant innate immunity. In addition, JA and salicylic acid are known to play a role as an antagonist in plant innate immunity (PDF1.2 gene in DEWAX-overexpressing Arabidopsis indicates that JA-associated defense responses are activated, and this may cause the suppression of SA-related signaling. This hypothesis supports that DEWAX-overexpressing plants are more resistant to the necrotropic fungal pathogen, B. cinerea, but are more susceptible to the semi-biotropic bacterial pathogen, Pto DC3000 compared with WT.The expression of ethylene . The JA- cinerea . Howevere family . Based o rhythms , it woulimmunity . The indC. sativa is considered to be resistant to many diseases, it is reportedly sensitive to some fungal and bacterial pathogens causing wilting, root rot, clubroot, white rust, and downy mildew diseases (C. sativa overexpressing the antimicrobial peptide cecropin P1 exhibits enhanced resistance to Erwinia carotovora and Fusarium sporotrichioides. In this study, the Arabidopsis dewax mutant showed increased resistance to Pto DC3000, whereas overexpression of DEWAX conferred elevated resistance to B. cinerea in Arabidopsis and C. sativa, suggesting the potential for developing Brassica lines resistant to various diseases. In conclusion, our study revealed that DEWAX is a transcriptional activator that upregulates the expression of defense-related genes, and overexpression of DEWAX enhances the resistance to B. cinerea in Arabidopsis and C. sativa. We provide that DEWAX-mediated defense mechanisms may be applicable in the improvement of crops with enhanced resistance to fungal and bacterial pathogens.Although diseases . ZakharcSJ, YG, and MS conceived and designed research. SJ, YG, and HC conducted experiments. SJ, JP, and MS analyzed data. SJ, JP, and MS wrote the manuscript. All authors read and approved the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Graphene layers were grown by chemical vapor deposition (CVD) on copper substrates. Single and layer-by-layer graphene stacks were fabricated combining graphene transfer techniques and metal oxides grown by magnetron sputtering. The electrochemical properties of the samples were analyzed and the results suggest an improvement in the performance of the device with the increase in the number of graphene layers. Furthermore, deposition of transition metal oxides within the stack of graphene layers further improves the areal capacitance of the device up to 4.55 mF/cm2, for the case of a three-layer stack. Such high values are interpreted as a result of the copper oxide grown between the copper substrate and the graphene layer. The electrodes present good stability for the first 850 cycles before degradation.The combination of graphene with transition metal oxides can result in very promising hybrid materials for use in energy storage applications thanks to its intriguing properties, i.e., highly tunable surface area, outstanding electrical conductivity, good chemical stability, and excellent mechanical behavior. In the present work, we evaluate the performance of graphene/metal oxide (WOThe online version of this article (10.1186/s11671-017-2385-1) contains supplementary material, which is available to authorized users. Recently, electrochemical energy storage devices such as supercapacitors are becoming the most popular apparatus as power supplies in a wide variety of applications, from portable electronic devices, such as cell phones and laptops, to hybrid electric vehicles . SupercaThis is a result of the different energy storage mechanism between the two devices. In contrast to batteries, where ions are stored through chemical bonding to the electrode materials, in supercapacitors, an electrostatic storage of the energy takes place through the separation of charge in a Helmholtz double layer . AdditioHence, it is considered that supercapacitors have the potential to replace or complement batteries in energy storage applications. Research in this direction focuses in the development of novel electrodes that can exhibit superior characteristics. Similar to Li ion batteries, carbon-based materials are preferred due to their low environmental impact, chemical stability, high conductivity, and low cost .2-hybridized carbon atoms, has some very exciting properties which make it very attractive for being used as an electrode in this kind of applications. We highlight its light weight, high electrical and thermal conductivity, highly tunable surface area (up to 2675 m2/g), strong mechanical strength (~ 1 TPa), and chemical stability [2 and a corresponding specific capacitance of around 550 F/g when the entire surface area is fully utilized. At present, it is given much attention to three-dimensional graphene materials, such as graphene nanowalls and nanofoams, which can deliver high energy density and power density, in the order of 13 Wh kg−1and 8 kW kg−1, respectively [Graphene, an emerging nanomaterial which consists of all spup to 267 m2/g, stectively .Further, planar graphene films present the benefit of a homogeneous growth and good coupling to the metallic substrate, resultant of the mixture of covalent and ionic bonding on the graphene/copper interface , which sThe recent advances in the design and optimization of higher efficiency electrodes has promoted the combination of graphene and graphene oxide films with different metal and metal oxide composites –19, likeOn this design, graphene contributes, apart from its storage capacity, as a platform which permits the strong coupling and a good electric contact between the metallic nanoparticles and the current collector. Previous studies have revealed the beneficial role of graphene as a coupler between the current collector and carbon nanotubes .2O3 and MnO2, they show capacitances up to 380 F/g [On other research work, single-layer graphene electrodes have been measured to exhibit a specific double-layer capacitance of up to 135 F/g, while when combined with other compounds such as Fe 380 F/g , 22.3 and CeOx were sputtered.In the present work, we have manufactured graphene/metal oxide nanocomposites made from a single layer or three stacked layers of graphene/metal oxide nanocomposites, combining graphene transfer and magnetron sputtering techniques. On top of each graphene layer, different metal oxide particles of WOx in the whole manuscript as we have not characterized the grown particles. Although the sputtering process was performed with a CeO2 target, the formed particles should appear suboxidized because of the possible loss of oxygen during the sputtering process, but they are mainly formed by CeO2, which is the most stable form of cerium oxide. Compared to monolayer graphene, stacks of graphene films have more electrode/electrolyte interfaces, which is beneficial to the absorption/desorption of electrolyte ions and provide more electrical pathways for electrolyte ions during charging and discharging processes. The deposition of metal oxide particles enhances the specific capacitance of ultrathin layers at a relatively low mass loading [x particles have demonstrated high capacitance, in the order of 119 mF/cm2, when combined with nickel foam [3 films, a recent work has reported electrodes made with WO3 rods presenting a capacitance of 266 F/g [Cerium oxide is mentioned as CeO loading . In prevkel foam . Conside 266 F/g . Both naThe use of the same experimental conditions in the preparation of the two hybrid materials provides us with the opportunity to directly compare the electrochemical performance of the electrodes.To better interpret our results, we take into consideration the contribution of the native copper oxide layer in the overall capacitance of the electrode.2 pieces, cleaned in ultrasound bath of isopropanol and acetone, 10 min each, to remove impurities and loaded in the chamber. First, we apply a hydrogen plasma etching to remove the native copper oxide from the copper surface. Radio frequency (RF) plasma is generated by applying 100 W at 20 Pa pressure, under 20 sccm hydrogen flow. The plasma etching lasts 10 min. Then, the sample is forwarded in a quartz tube (coupled to the plasma chamber) surrounded by an oven. The oven is heated at 1040 °C and the gases are introduced in the tube. A mixture of methane and hydrogen is introduced (5/20 sccm of methane/hydrogen) during 20 min at 15 Pa, resulting in the complete coverage of the copper foil by single-layer graphene. Then, the sample is let to cool down in room temperature in high vacuum (3 × 10−4 Pa) before being removed from the chamber. Then, the sample is placed in another reactor to deposit the metal oxide particles. The metal oxide particles were deposited on the graphene layer by pulsed reactive magnetron sputtering , using, each time, the corresponding target (W or Ce). To prepare three stacked layers of graphene/metal oxide nanocomposites, we used graphene transfer method with a polymer coating support [3 to etch away the copper. The remaining graphene/metal oxide layer was then transferred over another layer of the same nanocomposite enabling the preparation of the stacked material. After the transfer process, the PMMA was removed by rinsing with acetone. The preparation process of the composite is presented in the schematic drawing of Fig. Continuous graphene films were grown by CVD following the growth recipes described in our previous work . We repo support . Polymetα line of 1486.6 eV energy and 350 W). Depth profile measurements of chemical composition by XPS were obtained by sputtering the surface with an Ar+ ion source (4 keV energy). All these measurements were made at ultra high vacuum (UHV) conditions, between 7 × 10−7 and 3 × 10−6 Pa.Samples were characterized by Raman spectroscopy (Jobin-Yvon LabRam HR 800), scanning electron microscopy (SEM) (JEOL JSM7100F), and transmission electron microscopy (TEM) (Bioscan Gatan JEOL 1010). X-ray photoelectron spectroscopy (XPS) measurements were performed in a PHI 5500 Multitechnique System  with a monochromatic X-ray source (Al K4 solved in ethylene carbonate (EC) and diethyl-carbonate (DEC) mixed in 1:1 volumetric proportions) electrolytes. A glass fiber filter served as separator (Whatman glassy-fiber GF/A). Figure 2 glove box (< 1 ppm O2 and < 1 ppm H2O) by sandwiching an organic electrolyte-soaked separator between two graphene/MeO electrodes. To study the supercapacitance behavior of the devices, we first performed cyclic voltammetry (CV) measurements at different scan rates and with a voltage window of 1.8 V.The electrochemical properties of the samples were analyzed using a Swagelok cell and organic  and the 2D peak FWHM (~ 40 cm−1) confirm that graphene is a single layer. The spectrum was obtained after transferring the graphene film over a SiO2 substrate in order to eliminate the noise resulting from the copper foil luminescence [The short sputtering deposition of the MeO nanoparticles aims in avoiding the damage of the graphene layer. Longer sputtering periods could result in the damaging of the graphene, since the sputtering is performed in an argon/oxygen plasma. Figure ilm Fig. . We obseilm Fig. , the I2nescence .Fig. 2MoXPS provided information considering the copper oxide formation after plasma annealing and graphene growth. The measurements were performed on copper substrates with and without graphene grown on top in order to show that the graphene presence favors the formation of the copper oxide layer. The native copper oxide layer was reduced by plasma annealing in all samples . After each pickling process, we obtain information on the chemical composition at the most depth. The first two spectra (black and red line) have the same intensity. The rest of the spectra have a lower intensity. Defining the x. The specific capacitance, Cs, was calculated by the equation,Cs is the specific capacitance in farad per gram, m is the mass of the active material in grams, ΔV is the voltage window in volts, and qa and qc are the anodic and cathodic charges in coulomb, respectively.In Fig. Ci, was calculated using the relation,A is the area of active material dipped in the electrolyte . A similar behavior, resulting although in a smaller capacitance increase, is observed for the Gr/WO3 electrodes. Their capacity increases from 2.69 to 4.15 mF/cm2 when two more layers of Gr/WO3 are added over the first layer (+ 54%).The as-grown graphene film presents an interfacial capacitance 3 electrodes, reveal an agreement in the percentage increase with respect to the Gr/MnO2 hybrid structure.Similar percentage increase is expected when more graphene/metal oxide layers are added, as the surface area will increase proportionally while the interlayer distance may also allow multilayer ion absorption. In Fig. −4 W/cm2 in the case of three layers of Gr/CeOx electrodes, a value of the same order of magnitude as that of other electrodes, with similar architecture, combining graphene with MnO2 particles [−8 W-h/cm2, a value which is two orders of magnitude lower than the values given for the case of Gr/MnO2-based electrodes.To demonstrate the overall performance of the supercapacitors, we illustrate a Ragone plot with the energy density and power density of the various electrodes Fig. . We obsearticles . Althoug2, while in our work, it is measured to be 0.87 mF/cm2. In our work, the graphene layer was deposited on a copper foil, which was used as a current collector, making transfer of graphene unnecessary. We consider that the formation of copper oxides in the graphene/copper interface, resulting from the copper oxidation, affects the total capacitance of the system. Additionally, we know that the presence of graphene favors the growth of a copper oxide layer of some tenths of nanometer, as it has been observed by us and also reported by other authors [2O) layers. Their “brighter” appearance is the result of the higher backscattering of electrons on copper oxide than in the case of naked copper.We observe that the capacitance of the sample with single-layer graphene is much higher, about nine times, than those mentioned elsewhere . In the  authors , 29. Alt authors . By delact is the total capacitance that we measure, cox the capacitance of the copper oxide, and cg the graphene quantum capacitance. Although, as it has been evaluated by experimental observations, graphene presents a negative capacitance when it is decorated with metallic adatoms. These adatoms act as resonant impurities and form nearly dispersionless resonant impurity bands near the charge neutrality point (CNP). Resonant impurities quench the kinetic energy and drive the electrons to the regime dominated by the Coulomb energy with negative compressibility. If we consider a negative quantum capacitance of the graphene [2), we will be able calculate the copper oxide capacitance (11.1 mF/cm2) that corresponds to a copper oxide thickness of approximately tenths of a nanometer [Therefore, to better interpret our results, we should consider that each electrode consists of two capacitors, the graphene film and the copper oxide film, in series, contributing to the total capacitance, asgraphene  with a vgraphene  (0.1 mF/anometer , in agreanometer , conside3 and three-layer Gr/CeOx. The electrodes consisting of Gr/CeOx have a better charge/discharge efficiency during more cycles as shown in Fig. Finally, we present results considering the stability in the device performance. All electrodes present a capacitance retention of between 70 and 90% during the first 850 cycles, as we can see in Fig. x hybrid and in Fig. 3 hybrid. One-layer Gr/CeOx needs 1.7 s approximately for a charge/discharge cycle when charged by 400 mA/cm2. When two more layers were added over the first one, this period increased to ~ 4.7 s. Measurements performed on a single-layer graphene showed similar charge/discharge time as in the case of the single-layer Gr/CeOx electrodes. Similar results were obtained in the case of WO3 particles, where charge-discharge time was 1.9 s for one layer and 5.5 s for three layers. This demonstrates the higher power density that the CeOx hybrid is performing. The electrochemical results of the study are listed in Table The galvanostatic charge/discharge curves reveal that when more layers of metal oxide/graphene are added, more time is needed for the charging and discharging process. This is visualized in Fig. x have a slightly higher charge/discharge efficiency than those in which Gr is combined with WO3. Considering stability, all devices maintain their initial performance for more than 800 cycles. The charge/discharge period increases about 2.5 times with the addition of two more layers over the first one.Layer-by-layer evaluation of graphene electrodes combined with different metal oxides has been performed. The deposition of metal oxide particles over the graphene increases the total capacitance of the hybrid material, as metal oxide particles contribute with an additional pseudocapacitance. An increase has been also observed when more layers of Gr/metal oxide were added over the first layer. The devices in which Gr is combined with CeO


Moreover, the changes in both the thermal and electrical conductivities with grain size change are greater than those of typical semiconducting thermoelectric materials.Understanding the influence of grain boundaries (GBs) on the electrical and thermal transport properties of graphene films is essentially important for electronic, optoelectronic and thermoelectric applications. Here we report a segregation–adsorption chemical vapour deposition method to grow well-stitched high-quality monolayer graphene films with a tunable uniform grain size from ∼200 nm to ∼1 μm, by using a Pt substrate with medium carbon solubility, which enables the determination of the scaling laws of thermal and electrical conductivities as a function of grain size. We found that the thermal conductivity of graphene films dramatically decreases with decreasing grain size by a small thermal boundary conductance of ∼3.8 × 10 Understanding the effect of grain boundaries on the electrical and thermal transport properties of graphene is of both fundamental and technological importance. Here, the authors fabricate graphene films with controlled grain size, and determine the scaling laws of thermal and electrical conductivities. Graphene has attracted increasing interest because of the extraordinary properties of its defect-free pristine form, such as the highest known carrier mobility, record thermal conductivity and extremely high mechanical strength1245678910111256812121314151617181920212223From the point of view of crystal growth, it is equally difficult to reduce and increase nucleation density on metals by CVD to fabricate large-size single-crystal graphene and polycrystalline graphene with nano-sized grains, respectively, while keeping monolayer growth of graphene. For surface adsorption growth on the commonly used Cu with a low carbon solubility, a high-concentration carbon source and/or defective substrates have usually been used to obtain a high domain density, however, these conditions led to the formation of multi-layer graphene domains102527285It has been theoretically predicted that electrical transport in graphene could be markedly altered by electron scattering at GBs14410111275209 W m−2 K−1, while the electrical conductivity slowly decreases with an extraordinarily small GB transport gap of ∼0.01 eV and GB resistivity of ∼0.3 kΩ μm. Moreover, both the thermal and electrical conductivities of graphene change more significantly with grain size change than that of typical thermoelectric materials3334Here we have developed a segregation–adsorption CVD (SACVD) method to achieve a great increase in the nucleation density of graphene (by segregation) and monolayer growth (by surface adsorption) simultaneously, by using a Pt substrate with medium carbon solubility. As a result, we can easily grow well-stitched high-quality monolayer graphene films with a tunable uniform grain size from ∼200 nm to ∼1 μm, which have never been achieved before by the present conventional CVD methods based on either surface adsorption45678910111213272728−2 were obtained 5We obtained histograms of grain sizes by measuring more than 100 grains for each sample . The mea2/Si holey substrates  to make sure that the suspended area is intact. The SEM image shows that most area of the substrate is covered by graphene without visible cracks  of the graphene films was calculated by κ=χ(1/2hπ)(δw/δP)−1, where δw is the shift of 2D peak position due to the change of heating power δP on the sample, χ is the 2D peak temperature coefficient, and h is the thickness of the graphene film.Confocal micro-Raman spectroscopy is an efficient method for measuring the thermal conductivity of suspended graphene. Its value is extracted from the dependence of the Raman G or 2D peak frequency on the excitation laser power37e cracks . Figure ID/IG increase sharply while the G peak intensity decreases dramatically when the laser power is larger than 1.2 mW  and the extracted temperature coefficient (0.039 cm−1 K−1), we estimated that the temperature at the GBs of the graphene film with ∼200 nm grains reached 650 K when the laser power was 1.2 mW. Combined with the Raman spectra evolution, the physical origins of the D and G peak12n 1.2 mW . The 2D n 1.2 mW . Howeverecreased . These pκ) as a function of grain size (lg). It is clear that the thermal conductivity increases exponentially from ∼610 to ∼5,230 W m−1 K−1 when the grain size is increased from ∼200 nm to ∼10 μm. In fact, the graphene films with grain size larger than ∼5 μm (the size of the suspended area) all show a similar thermal conductivity of ∼5,200 W m−1 K−1 , which is similar to the value reported for pristine graphene made by mechanical exfoliationleff−1= lph–ph−1+lg−1, where lph–ph denotes the phonon–phonon scattering length and lg is the scattering length due to the boundaries κ−1) versus the inverse of grain size (lg−1) can be well fit by κ−1=κg−1+(lgG)−1, where κg is the thermal conductivity within the grain  and G is the boundary conductance9 W m−2 K−1, which is consistent with the theoretical value obtained using non-equilibrium Green's functions (3–8 × 109 W m−2 K−1) ]} , R is the universal gas constant, T is the absolute temperature, lg is the grain size and c is the correction value. The fitting gives σ0≈2.85 × 106 S m−1 and Ea≈0.01 eV. Note that the GB transport gap extracted here is dramatically smaller than the theoretically predicted value for asymmetric GBs (0.3–1.4 eV) ]} , where σeV) ref. . Using tG+ρGB/lg 13, whereBs 0.3–1. eV (ref.lg lg−1). Note that the thermal conductivity change rate (Δκ κ−1) increases linearly with grain size change rate (σ σ−1) increases exponentially with grain size change rate 47483334nL, mL) and  that belong to the left and right crystalline domains, respectively. If only one matching vector fulfills the criterion (n–m)=3q , then the GB is of class-II type. Otherwise it belongs to class-I. For class-II GB, there is significant misalignment of the allowed momentum–energy manifolds corresponding to the two crystalline domains of graphene, which introduces a transport gap  that depends exclusively on the periodicity1414The GBs in graphene can be approximated as linear periodic arrays of dislocations9 W m−2 K−1, while the electrical conductivity slowly decreases with an extraordinarily small GB transport gap of ∼0.01 eV and GB resistivity of ∼0.3 kΩ μm. Moreover, the changes in both the thermal and electrical conductivities with grain size change are greater than those of typical semiconducting thermoelectric materials. These findings provide valuable information for tuning the thermal and electrical properties of graphene for electronic, optoelectronic and thermoelectric applications through grain size engineering.In conclusion, we report a SACVD method to grow well-stitched high-quality monolayer graphene films with a tunable uniform grain size from ∼200 nm to ∼1 μm, by using a Pt substrate with medium carbon solubility. Using these materials, we determined the scaling laws of the thermal and electrical conductivities of graphene films as a function of grain size. It was found that the thermal conductivity of polycrystalline graphene films dramatically decreases with decreasing grain size by a small thermal boundary conductance of ∼3.8 × 10A typical procedure for the SACVD growth of graphene films with grain sizes <1 μm includes four steps: surface growth, etching, segregation and surface growth. Before growth, a piece of Pt foil  was rinsed with acetone and ethanol in sequence for 1 h each, loaded into a fused-silica tube (inner diameter: 22 mm), heated to a certain temperature under the protection of hydrogen, and then annealed for 10 min to remove any residual carbon or organic substances. The first step: surface growth was started with the substrate being held for a certain time under a mixture of methane and hydrogen. In the second step the methane and hydrogen flows were turned off, and pure argon (700 sccm) was introduced to the system for 20 min to etch the graphene grown on the Pt in the first step into the bulk. In the third step, a small amount of hydrogen was introduced into the system to mix the argon flow, initiating segregation of the carbon to form small graphene domains on the Pt surface. In the fourth step a low flow rate of methane (0.1 sccm) was introduced into the system while maintaining the hydrogen and argon flows, to cause surface growth of the graphene domains produced in step 3 to form a continuous polycrystalline film. The detailed experimental conditions were given in the main text and 2 (290 nm) substrates using a improved bubbling transfer methodTo investigate the structure of the graphene formed at different stages, the Pt foil was quickly pulled out of the high-temperature zone after SACVD growth. The furnace was then shut down and the methane flow was stopped after the furnace temperature had decreased to 600 °C. Finally, the Pt foil was taken out and characterized by SEM . The small graphene domains and polycrystalline films were transferred onto Si/SiO2/Si substrate and thermal conductivity of the suspended graphene in our experiments show the similar values with those of graphene with similar grain size reported in the literatures443 W m−1 K−1, close to the reported value (5.3 × 103 W m−1 K−1) , and the integration time at each position was 10 s. The temperature rise was determined from the shift of the Raman 2D peak. The sheet resistances of the graphene films were measured by a four-probe method (RTS-9) at room temperature. These two methods have been widely used in the literatures12133844The data that support the findings of this study are available from the corresponding author upon request.How to cite this article: Ma, T. et al. Tailoring the thermal and electrical transport properties of graphene films by grain size engineering. Nat. Commun.8, 14486 doi: 10.1038/ncomms14486 (2017).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.Supplementary Figures, Supplementary Notes and Supplementary References


It is found that the contact resistances between all metals and graphene are remarkably reduced after annealing, indicating that not only chemically adsorbed metal (Pd) but also physically adsorbed metals (Ag and Pt) readily form end-contacts at intrinsic defect locations in graphene. In order to further improve the contact properties between Ag, Pd, and Pt metals and graphene, a novel method in which self-aligned He+ bombardment to induce exotic defects in graphene and subsequent thermal annealing to form end-contacts was proposed. By using this method, the contact resistance is reduced significantly by 15.1% and 40.1% for Ag/graphene and Pd/graphene contacts with He+ bombardment compared to their counterparts without He+ bombardment. For the Pt/graphene contact, the contact resistance is, however, not reduced as anticipated with He+ bombardment and this might be ascribed to either inappropriate He+ bombardment dose, or inapplicable method of He+ bombardment in reducing contact resistance for Pt/graphene contact. The joint efforts of as-formed end-contacts and excess created defects in graphene are discussed as the cause responsible for the reduction of contact resistance.Low contact resistance between graphene and metals is of paramount importance to fabricate high performance graphene-based devices. In this paper, the impact of both defects induced by helium ion (He There has been plentiful studies on the properties of graphene-metal (G-M) contact [−7 Ω·cm2. Similar to this plasma treatment, ultraviolet/ozone (UVO) was used to clean the graphene surface after lithography which reduced contact resistance to 7 × 10−7 Ω·cm2 and 568 Ω·μm respectively [c over a wide range cannot be easily realized, or even the Rc will be degraded under some circumstances. Alternatively, the graphene/metals end-contacts, initially disclosed in a theoretical work [c dramatically. The utilization of end-contacts in reducing Rc is also confirmed by experimental efforts [c were demonstrated by introducing holes (~2.2 × 10−9 Ω·cm2) [c can be reduced using end-contacts, the approach to introducing various patterns in the contact area of graphene is too complicated to be implemented in practical applications, since in these cases the lithography and subsequent dry etching is usually needed in order to shape the holes, zigzag edges, or line cuts [c. In this regard, a simple while efficient method featuring the end-contacts concept to reduce Rc is of great importance and interest. In this work, a novel method to form end-contacts between graphene and various metals  is explored. In this method, defects are introduced to graphene by light He+ bombardment and thereafter end-contacts are formed between graphene and metals. Since the thermal annealing treatment is a common yet effective way to reduce Rc [c with and without He+ bombardment is also investigated in this work.In the past decade, graphene has attracted worldwide attention as a miracle material with unique electrical and physical properties ,2,3. Graformance ,16,17,18 contact ,20,21 an contact  and Robi contact  proposedectively ,25. Nevecal work , is clai efforts ,28. In t9 Ω·cm2) , or lineine cuts ,28. Moreeduce Rc ,30, the −1 respectively are observed and the ratio of 2D/G peaks is higher than 2 indicating that the transferred graphene is single layer.Thermal chemical vapor deposition (CVD) grown graphene ,32 was tx) isolation layer is deposited between PR and graphene as a hardmask by Atomic Layer Deposition (ALD) [x and underneath graphene (cf. 3PO4 (1:3) at 40 °C to remove the unwanted AlOx layer in these contact windows  were deposited directly on graphene by e-beam evaporation  is used to extract the contact resistance between graphene and metals. The fabrication of TLM test structures is schematically shown in on (ALD)   consists of graphene sheet resistance (RS), G-M contact resistance (RGM), metal wire resistance (RW), and probe-pad contact resistance (RPP) as seen in Equation (1).For the extraction of G-M contact resistance, four probe configuration illustrated in S relates to the length of graphene diffusion strip (lG) between two G-M contacts and the value of RW depends on the length of metal wires between pads and G-M contact windows (lW). Benefitting from the four point probe configuration in TLM method, RPP can be ignored. The slope in T which is the sum of RS, 2RGM, and RW. Normally, Rw is small which can be omitted in Equation (1) if metal wires are thick and/or short. However, since in this work the deposited metal wires are thin i.e., 50-nm-thick and long, Rw cannot be omitted which is also a disturbing factor for the linear plotting of RT vs. lG. In order to get a perfect linear plotting, it is a must to subtract Rw from RT. The sheet resistances of metal wires are extracted to be 0.96, 3.72 and 4 Ω/□ for Ag/Au, Pd/Au and Pt/Au, respectively. As a result, the values of Rw can be calculated according to lW and the width of metal wires and be subtracted from RT. After the Rw subtraction, perfect linear relationships between RT and lG for all G-M contacts are accomplished as shown in Among them, R+ bombardment, the linear plots of RT vs. lG before annealing (a–c) and after annealing (d–f) are shown in G, 10 data points are used and the linear fitting is performed using concatenate fit method. The perfect linear fittings for all G-M contacts are evident demonstrating the validity of employed method to extract Rc in this work. The interception of the fitted red line with RT (Y-axis) yields the value of 2RGM. Since the width of the graphene diffusion strip is 15 μm, Rc is therefore calculated by RGM × 15 μm which is also offered in each corresponding plot. For all three G-M contacts, it is clearly observed that Rc values are reduced significantly after annealing which confirms the effectiveness of the annealing in improving G-M contacts [For the contacts between Ag, Pd, Pt, and graphene without Hecontacts .c with different metal work functions [c data in c values for different metals  and filled red squares depict the mean Rc values with error bar. As seen, albeit the metal work function alters from ~4.26 to ~5.65 eV for different metals, the Rc value remains nearly invariable, manifesting that Rc is barely—or even not—relevant to metal work functions before annealing. This observation is also in accordance with previous finds of E. Watanabe’s et al. [Except the plot of Runctions ,36 in Fis et al.  in whichs et al. .c values for all G-M contact before and after annealing shown in c value is reduced significantly after annealing since it is a chemically adsorbed metal. The intriguing point in this work is why physically adsorbed metals like Pt and Ag also show remarkably reduced Rc values after annealing, i.e., from 247.43 and 285.07 to 180.75 and 227.81 Ω·μm for Pt/graphene and Ag/graphene contacts, respectively. In order to clarify this point, elaborate characterizations of graphene in G-M contact windows before and after annealing are implemented by Raman scattering spectrum. The contact windows are opened using aqua regia (HCl:HNO3 = 3:1) to remove metals thus exposing the graphene under metals [−1 for the graphene after annealing in comparison to the graphene before annealing. Note that the Raman spectra of the graphene before annealing resemble those of as-transferred graphene on SiO2  [S and rA are length scales that determine the region where the D band scattering takes place. rS determines the radius of the structurally disordered area and rA is the radius of the area surrounding the point defects in which the D band scattering takes place. CA is a measure of the maximum possible value of the ID/IG ratio in graphene. CS is the value of the ID/IG ratio in the highly disordered limit [D is the mean defect distance and the defect density is proportional to 1/LD2. In accordance with the empirical data in [A = (4.2 ± 0.1), CS = (0.87 ± 0.05), rA = (3.00 ± 0.03) nm, and rS = (1.00 ± 0.04) nm, the ID/IG as a function of LD for the graphene with and without He+ bombardment is plotted in D/IG increases from 0.032 for the un-bombarded graphene to 0.143 for the bombarded one, the LD increases dramatically from 58.05 nm to 27.56 nm. This means that the defect density in the graphene is increased approximately 4.4 times using He+ bombardment.The graphene in G-M contact windows was bombarded by Hetion (2) :(2)IDIGed limit ,40. LD i data in , CA =  and after annealing (d–f) are shown in + bombardment, 10 data points are used and the linear fitting is performed using concatenate fit method. As seen, the effectiveness of annealing in improving the G-M contacts with He+ bombardment is also evident. However, how the He+ bombardment impacts the specific contact resistance is still not clear. In order to provide a direct comparison for the readers, the Rc values for the G-M contacts with/without He+ bombardment are summarized in + bombardment before annealing. It is obvious that except Pd, He+ bombardment leads to an increase in Rc value for the Ag/graphene and Pt/graphene contacts. The defects induced by He+ bombardment should be blamed for the increase of Rc value and this observation also agrees well with the G-M contacts formed by metal sputtering on graphene [c [+ bombardment after annealing. As seen, for Ag/graphene and Pd/graphene contacts with He+ bombardment, the Rc values are reduced dramatically by 15.1% and 40.1% compared to their counterparts without He+ bombardment. Note that the obtained Rc values in this work for all three G-M contacts, i.e., 193.33, 118.28 and 189.15 Ω·µm for Ag, Pd, Pt/graphene contacts respectively, are among the lowest specific contact resistances [+ bombardment after annealing, the Rc value is increased a little bit compared to its counterpart without He+ bombardment and this could be tentatively interpreted as what follows.For Ag, Pd, and Pt/graphene contacts with Hegraphene . The defphene [c . In Figuistances ,37. Nevec value is lower for Ag/graphene and Pd/graphene contacts whereas is higher for Pt/graphene with He+ bombardment after annealing. We think the discussion about the higher or lower of contact resistance after annealing should take both the capability of metals forming end-contacts with defects and carrier scattering induced by defects within graphene into account [optimum) of He+ bombardment where the lowest Rc value (Rcmin) takes place. Four cases happen to the dose of He+ bombardment. (1) If He+ dose is Doptimum, the created defects in graphene can be completely consumed by the formation of end-contacts between metals and graphene. As a result, Rcmin can be obtained; (2) If He+ dose is less than Doptimum, Rc value is reduced but not to Rcmin in that as-formed end-contacts are not saturated; (3) On the other hand, if He+ dose is more than Doptimum but less than Dmax, though the formation of saturated end-contacts leads to the reduction of Rc, excess created defects enhance the carriers scattering which unfortunately degrades Rc concurrently. The joint efforts of as-formed end-contacts and excess created defects render the Rc value not the lowest Rcmin; (4) If He+ dose is more than Dmax, the effort of excess created defects prevails over that of saturated end-contacts and this leads to drastically increased Rc value that is even larger than Rc0 i.e., the Rc value for G-M contacts without He+ bombardment after annealing. Specifically, for the Ag/graphene and Pd/graphene contacts, they belong to cases (2) and (3) where their Rc values are reduced but possibly not to the lowest values Rcmin. It is worth noting that for Pd/graphene contact, the employed dose 2 × 1013 cm−2 is even closer to its Doptimum since its reduction percentage 40.1% is much bigger than 15.1% for Ag/graphene contact. For Pt/graphene contact, it belongs to case (4) where the effort of excess created defects prevails over that of as-formed saturated end-contacts, resulting in even larger Rc value compared to its counterpart without He+ bombardment after annealing. The specific contact resistance as a function of ion bombardment dose is still under investigation and will be revealed as a continuation of this work in the near future.In  account ,42,43,44+ bombardment before and after annealing are also performed by Raman scattering spectrum. The Raman spectra of as-exposed graphene with He+ bombardment  are depicted in −1 for the graphene before annealing in comparison to as-transferred graphene in + bombardment. However, for the graphene with He+ bombardment after annealing, the D peak becomes startlingly conspicuous. If scrutinized, the intensities of D peak for the graphene with He+ bombardment in all G-M contact windows are higher than those for the graphene without He+ bombardment in + bombardment. We believe the key to achieve extremely low specific contact resistance is to introduce defects in graphene thus forming plenty of end-contacts after annealing like other methods to form end-contacts [+ bombardment, but also by the implantation of other ion species. It is a remarkable fact that the optimum dose to achieve the lowest specific contact resistance may vary depending on the ion species used or on the implantation energy.Similarly to the discussions in previous section, the elaborate characterizations of graphene in G-M contact windows with Hecontacts ,27,28. T+ bombardment to induce exotic defects in graphene and subsequent thermal annealing to form end-contacts, was proposed in order to further reduce the specific contact resistance. Achieved results show that the specific contact resistances are reduced significantly by 15.1% and 40.1% for Ag/graphene and Pd/graphene contacts with He+ bombardment compared to their counterparts without He+ bombardment, respectively. For the Pt/graphene contact, the contact resistance is, however, not reduced as anticipated with He+ bombardment and this might be ascribed to either inappropriate He+ bombardment dose, or inapplicable method of He+ bombardment in reducing contact resistance for Pt/graphene contact. The effort of as-formed end-contacts prevailing over that of excess created defects is attributed to the reduction in Rc values for G-M contacts with He+ bombardment after annealing. By manipulating the He+ bombardment dose in conjunction with Ag, Pd, and Pt metals, the lowest Rc value for each G-M contact could be possibly accomplished. It is worth noting that the proposed He+ bombardment and metal depositions share the same lithography mask and this processing simplicity demonstrates that our proposed method is very efficient in improving the contact properties for graphene-based devices in the future.To summarize, the specific contact resistance for three G-M contacts  are all reduced after annealing from 285.07, 267.60, and 247.43 Ω·µm to 227.81, 197.33, and 180.75 Ω·µm, respectively. This indicates that not only chemically adsorbed metal (Pd) but also physically adsorbed metals (Ag and Pt) readily form end-contacts at intrinsic defect locations in graphene. Along with this guideline, a novel method, in which self-aligned He