This review article looks at the use of lead-free halide perovskite materials for humidity sensing. A complete description of the underlying mechanisms and charge transport characteristics that are necessary for a thorough comprehension of the sensing performance will be provided. An overview of considerations and potential recommendations for the creation of new lead-free perovskite nanostructure-based sensors is presented.Over the past decade, perovskite-based nanomaterials have gained notoriety within the scientific community and have been used for a variety of viable applications. The unique structural properties of these materials, namely good direct bandgap, low density of defects, large absorption coefficient, high sensitivity, long charge carrier lifetime, good selectivity, acceptable stability at room temperature, and good diffusion length have prompted researchers to explore their potential applications in photovoltaics, light-emitting devices, transistors, sensors, and other areas. Perovskite-based devices have shown very excellent sensing performances to numerous chemical and biological compounds in both solid and liquid mediums. When used in sensing devices, Perovskite nanomaterials are for the most part able to detect O Double perovskite is the resultant substance, having the formula A2B’B”X6 and the cubic space group Fm3 m . A. A3) typup Fm3 m . The geoup Fm3 m . The thee formed . Lead-fr2BiAgCl6, Cs2BiAgBr6, Cs2SbAgCl6, and Cs2SbAgBr6  octahedra, even though researchers would want to name them, perovskites. Due to the isolated [BX6] octahedra in A2BX6 compounds, the optoelectronic properties of A2BX6-type perovskites deviate from those of 3D ABX3  perovskites. A2SnI6  [2TiBr6 [2BX6 compounds [Previous research has shown that tetravalent B(IV) replacements may be used to replace Pb in lead halide perovskites. To accommodate these heterovalent replacements, the chemical formula must be changed to Aeen them ,74,75,76 Cs, MA) ,78 and C [2TiBr6  have beeompounds ,77,78,792BiAgBr6 and Cs2PdBr6 are both recently developed lead-free perovskite with good photoelectric properties and high stability. Zhan Yiqiang et al. and Ye et al. both found that these materials reported humidity-dependent electrical properties and good stability and therefore giving them excellent humidity sensitivity [Cssitivity ,38. By aThere are two types of humidity measurements, i.e., absolute humidity, and relative humidity related to qualitative assessment. The proportion of vapor (moisture) in a unit volume of air, independent of temperature, is referred to as absolute humidity. The proportion of water vapor present in a unit volume of air is also referred to as relative humidity, but this time in connection to the temperature of the air.Most humidity sensors work by measuring the change in capacitance or resistance of a certain conductive substance as a function of relative humidity. Several fundamental parameters determine the good functioning of a humidity sensor. The proper operation of a humidity sensor is determined by several essential parameters. The key parameters to consider when it comes to evaluating the functioning of a humidity sensor are sensitivity, accuracy, and response/recovery time.Humidity sensors are categorized into several classes of sensors. These groups include electrical, mechanical, optical, and integrated sensors. Electrical sensors are types of sensors whose functioning is based on impedance and capacitance , strain The general humidity sensing mechanism of ceramic oxides or perovskites as a whole relies on the superficial water vapor absorption from chemisorption, physisorption, and capillary condensation process as can be seen in 2BiAgBr6 humidity sensor. When there are no water molecules adsorbed on the film surface, the conductivity comes from the carriers inside the p-type Cs2BiAgBr6 material, which may principally be the Ag vacancy [+ protons are transported to nearby water molecules, H3O+ are generated. In the presence of external electric fields, the protons can then be moved from one site to another across energy barriers. As a result, when the humidity level is low, the sensor’s electrical conductivity rises.In general, the adsorption of water molecules on the sensor surface during the humidity sensing process is divided into different phases, chemisorption, and physisorption ,83, as i vacancy . Water m vacancy . Meanwhi vacancy ,85, in a2BiAgBr6 thin film  on the flat (or porous) surface, these different processes can be involved in explaining the resistance or capacitance changes. Under applied voltage, singly bonded water vapor molecules, for example, become mobile and capable of forming dipoles and electrolyte layers, leading to an enhanced dielectric constant and bulk conductivity. As a result, the surface water protonation, as well as protonic conduction mechanisms, might cause a minor fluctuation in conductivity with humidity adsorption.2AgBiX6 and Cs2InBr5H2O  in their stable state ranges from t = 0.8 to t = 1 [3 configuration, inorganic perovskites based on Cesium exhibit low t-factor values, indicating poor phase stability. As can be observed from One of the most important ways in which scientists have been able to improve not just the structural characteristics of perovskite materials, but also their optical characteristics, has been to modify their compositional structure. These modifications have proven to have an impact on the material’s performance and stability in device applications. The capacity that perovskite materials have, to tune their bandgap by using mixed-cation or mixed-halide compositions, enables their optical absorption to be expanded spanning a broader spectrum of wavelengths ,89. The to t = 1 ,90,91. WHumidity sensors typically use two methods to measure humidity. One of those is resistive sensing, in which water molecules affect the bulk material’s resistance due to chemisorption or physisorption. The second is capacitive sensing, in which the sensor’s capacitance changes when it interacts with water vapor . The typThe various perovskite materials utilized in the production of the capacitive and resistive sensors reported in Typically, noble precious metal electrodes are fabricated on a glass or ceramic substrate via thick film printing techniques. Ref.  or thin-The key active materials used for the fabrication of humidity sensors include metal oxides, with perovskites being a subpart , carbon Studies conducted on polymeric humidity sensors have progressed and been incorporated into the industry over the past four decades. The majority of these sensors are made from porous polymer thin films  and emplOrganic polymer thin film humidity sensors with their applications, on the other hand, have a lower degree of satisfaction and significance when compared to metal-oxide thick or thin-film ceramic sensors , yet, thElectroceramic materials having single/polycrystalline structures, as single species or composites, could be great contenders for humidity sensing applications. The use of these innovative materials with novel humidity sensing properties is being investigated to overcome some of the limitations of many of the conventional materials, such as insufficient sensitivity or selectivity, low catalytic grade, insufficiency of cavities, surface degradation due to harsh contaminants in extreme conditions, and failure to operate in extremely dry or moistened environments. The addition of suitable nanomaterials with variable particle sizes/morphologies, hybridization of materials by replacement or doping of new atoms in the lattice, and particle size reduction to sub-nano scales can all be used to overcome most of the flaws.2O3) [With techniques such as thick film screen printing, porous ceramic or nanorod-based ceramic humidity sensors have also been manufactured and produced ,130, whe2O3) . In this2O4-TiO2 [2O4-LiZnVO4 [2-K2Ti6O3 [l, Fe)2O4-TiO2 [2O4 [2 [2O4-TiO2 porous ceramic humidity sensor for microwave oven applications [Ionic and electrical conduction sensors, as indicated previously, are two types of resistive sensors. Surface chemisorption and physisorption are used by ionic conduction types of humidity sensors to measure ambient air relative humidity, and they include MgCr2O4-TiO2 , ZnCr2O4-LiZnVO4 , TiO2-K2-K2Ti6O3 , . [3 [3 [3 [The research and analysis of perovskite films and bulk materials’ humidity sensing behavior and morphological structure continue to generate unique preliminary and novel findings ,141,142.oxygen). , and forgen). [3 , ZnSnO3 ). [3 [3 , and NaT[3 [3 [3 , were cl3 nanocrystalline films with great sensitivity, strong linearity with impedance values ranging in more than three times the normal magnitude, limited hysteresis, and quick response time at 100 Hz, AC 1 V from 33 to 95 percent relative humidity. The complex impedance was also used to investigate the humidity sensing mechanism. The findings suggested that NaTaO3 may be used in humidity sensors. In the example of ZnSnO3, Bauskar et al. [3 cubic crystallites produced using a hydrothermal technique to create a quick and stable humidity sensor. Linearity, rapid response and recovery behavior, hysteresis within 3.5 percent, outstanding repeatability, stability, and a wide range of operation (11–97% RH) were all excellent humidity sensing properties of the sensor. Finally, Mahmoud et al. [3 powder for moisture sensing measurements. The solid-state reaction approach was used to make perovskite PbTiO3 (PT) powder and ceramics in a cost-effective manner. It made it possible to create a perovskite tetragonal single phase. At an ambient temperature, however, pure PbTiO3 did not demonstrate a satisfactory reaction to relative humidity.Zhang et al.  were ablr et al.  conducteation 11–% RH were3NH3PbI3-xClx films and investigated its sensitivity in RH ranges of 32 to 97 percent with a recovery time of 74 s. Xu et al. [3NH3PbBr3 humidity sensor with a quicker response time (250 s) and quicker recovery (30–70 s) in a broader range of humidity detection (7–98%). A CH3NH3PbI3-xClx-based humidity sensor with a reaction time of 21 s, quicker than the quickest commonly available psychrometer on the market, was recently produced by Ren et al. [2+, such as double, triple and vacancy ordered perovskites, have gotten a lot of interest because of their flexibility in photoelectric devices. The produced metal-based halide perovskites such as (NH4)3Bi2I9 [2AgBiBr6 [3Bi2Br9 [2PdBr6 [2InBr5·H2O [3+, Pd2+, In+3 with a comparable electron configuration to Pb2+.Hu et al.  developeu et al.  have devn et al.  with vern et al. ,148,149.4)3Bi2I9 , Cs2AgBi2AgBiBr6 , and Cs3[3Bi2Br9 , Cs2PdBr [2PdBr6  and Cs2InBr5·H2O  provide Capacitive humidity sensors are often designed with a layered structure with two electrode interfaces, or as an interdigitated structure with comb electrodes, analogous to resistive RH sensors, with the insulating polymer film sandwiched in between ,127. SevPolymeric capacitive humidity sensors have been widely employed in the industrial and automation industries due to their simplicity in terms of coating, mass production, long-term stability, and a wide variety of potential sensing polymers such as polyimides ,155. On-Variations in the sensing film dielectric constant as a result of water absorption have a big impact on the capacitive RH sensors’ architecture, which alters the overall capacitance of the system. Differences in capacitance can be used to detect humidity changes because polymer dielectrics’ physical properties, such as permittivity, alter proportionally with the high dipole moments of water molecules. At ambient temperature, capacitive polymer sensors have a relative dielectric permittivity of roughly 5, whereas pure water has a permittivity of about 80. The dielectric permittivity increases the magnitude of the adsorption of water vapor by polymers (78.54), resulting in a sensitive linear change in capacitance. Polyimides (PI) and cellulose acetates are examples of such polymers, with relative dielectric permittivity values in the range from 3 to 6.Based on established observations, the porosity of the humidity-sensitive polymer sheet may be changed to boost sensor responses. In 1983, Delapierre et al.  proposedYang et al.  made a cBi et al.  develope2O3) with a microporous structure is the best material for these types of sensors [2O3 with a small pore radius is extremely prone to very low humidity levels, according to the electron tunneling mechanism effect inside the compacted immobile layers of water [2O3 (amorphous) and Al2O3 (corundum) are often used; however, the former is somewhat more sensitive than the latter due to its higher porosity [2O3 humidity sensors for their low cost and convenience of usage. As per the electron tunneling mechanism effect within the condensed immobile layers of water, Al2O3 with a tiny pore radius is particularly sensitive to very low humidity levels. Aluminum oxide (Al2O3) with a porous structure is the most suited material for these types of sensors. Only the two phases γ-Al2O3 (amorphous) and α-Al2O3 (corundum) are typically utilized in humidity sensing applications; however, the former is more sensitive than the latter due to its greater porosity [2O3 humidity sensors. McGhee et al. [Aluminum oxide  shells were placed over hydrothermally generated zinc oxide (ZnO) nanorod cores. Per the morphological study, the initial zinc oxide nanorods were coated by anatase titanium oxide shells as a second layer. The ZnO/TiO2 nanocomposite (ZTNA) sensors have substantially better sensitivity at 95 percent RH than separate ZnO and TiO2 sensors . Moreover, at ambient temperature, the capacitance of the core/shell arrays varied from 101 to 106 pF throughout the whole humidity range of 11 to 95 percent RH [On glass substrates, Gu et al. built ZnO/TiOrcent RH . The design configurations of impedance- (resistive) and capacitive-based humidity sensors that have proven to be the best suited and most prevalent in advanced applications such as laboratory research or automated industries were examined. An interesting fact to note was that resistive sensors operate on the same principle as capacitive sensors, measuring electrical changes to provide a relative humidity value. Although resistive sensors use hygroscopic materials similar to capacitive sensors, the humidity measurement processes in these systems differ. The distinction made was that the resistance change in the material is measured instead of the capacitance, and capacitive measurement, which is based on electrical capacitance, is used by a capacitive humidity sensor. In comparison to resistive measurements, capacitive measurements are widely utilized in the literature because they are more resilient and stable with temperature and humidity changes. Although both capacitive and resistive sensors have the same goal, which is that of measuring humidity, their methodologies are different. Capacitive sensors have also proven to be more accurate and stable than resistive sensors while giving reproducible findings, making them more ideal for medical applications where precision is critical. However, they are more expensive than resistive sensors. The less expensive resistive sensors are more commonly utilized in circumstances where frequent measurements are required but precision is not required.The success of these configurations moreover is linked to their ability to address common demands such as streamlined construction, free selection of sensing materials from several accessible kinds, cost, circuit adaptability, ease of manufacture, and measurement setup. Due to advantages such as low cost, proper gasketing, design flexibility, and quick deposition rate, film-based humidity sensors are more frequently employed. Semiconducting metal oxide and metal oxide/polymer-based sensors, which are primarily manufactured using thick film and thin film deposition techniques, stand out among the various types of humidity sensors due to their wide range of sensitive element options, post-processing capabilities, and superior response characteristics. 2BX6 or Cs2B”B’X6 and Cs2InX5.H2O, should be prioritized as a viable alternative to the endless difficulties that plagued other metal halides and oxides perovskites with lower stabilities and more efficient performances. Their nanoscale qualities may boost performance, having a significant impact on accuracy, reliability, and cost. In the future, nanocrystalline lead-free perovskite composites with ceramics, polymers, or ceramics/polymers, which are among the most promising materials for humidity sensors, may yield significant improvements in terms of humidity sensing performances.Exploration of the synthesis and use of lead-free metal halide perovskites, such as Cs


Spinal cord injuries and neurodegenerative diseases, including Parkinson’s, Alzheimer’s, and traumatic brain injuries, remain challenging to treat. Nowadays, neural stem cell therapies excite high expectations within academia. The increasing demand for innovative solutions in regenerative medicine has drawn considerable attention to graphene materials. Due to unique properties, carbon materials are increasingly used as cellular scaffolds. They provide a biological microenvironment supporting cell adhesion and proliferation. The topography and mechanical properties of the graphene culture surface influence the forces exerted by the cells on their extracellular matrix. Which consequently affects the cell proliferation and differentiation. As a result, material properties such as stiffness, elasticity and mechanical strength play an important role in stem cells’ growth and life. The ink unification process is crucial while the layer homogeneity is essential for obtaining suitable surface for specific cell growth. Different ink unification processes were tested to achieve appropriate layer homogeneity and resistivity to successfully applied the GNPs layers in neural cell electrostimulation. The GNP coatings were then used to electrostimulate mouse NE-4C neural stem cells. In this study, the authors investigated how the stimulation voltage amplitude’s value affects cell behaviour, particularly the number of cells. Sinusoidal alternating current was used for stimulation. Three different values of stimulation voltage amplitude were investigated: 5, 10, and 15 V. It was noticed that a lower stimulation voltage amplitude had the most favourable effect on the stem cell count. Stem cell therapies are gaining popularity due to their versatility, self-renewal, and multipotency . NervousThe ideal material for neuronal stimulation should be characterised by low electrical impedance, sizeable electroactive surface area, and appropriate biomimetics . Hence, Creating a scaffold and a microenvironment similar to the ECM enables cells to adhere, proliferate and differentiate appropriately , which aThe existing solutions do not achieve the desired reproducible therapeutic effect, and there is a significant discrepancy between scientific reports . SimultaModifying the particle surface by attaching surfactants is a standard procedure to improve the homogeneity of solutions and counteract the agglomeration of hydrophobic particles. The homogeneous dispersion of graphene particles in solution and their efficient distribution in the polymer matrix can be achieved using ultrasound . The larDue to the unique electrical properties of graphene, it is possible to combine two types of stimulation: material and electrical . The eluin vitro and in vivo, studying the degree of particle accumulation in tissues or blood vessels  and a Sonics Vibra Cell VC505 sonotrode homogeniser (maximum power 500 W and 20 kHz frequency). The baseline sonication process using the homogeniser lasted 1 min, while sonication using the scrubber lasted 30, 60, 90, or 120 min . The beaAfter combining the components with the pre-prepared base, the mix was sonicated again. In the case of inks homogenised using a sonotrode, the re-sonication time was 1 min, while the sonication time for inks in an ultrasonic washer was 15 min.® polyamide film substrate (from Dupont™) with screen-printed silver contacts and polystyrene 6-well cell culture plates from Nest Scientific Biotechnology. The printed silver contacts allowed for a measurement of the resistivity of the layers. The dimensions of the tested layers were 24 mm × 24 mm in width and length. As in the previous study, coatings were applied via spraying. A manual Infinity Airbrush CR Plus Solo spray gun from Harder & Steenbeck was used. For Kapton film substrates, the produced layers were dried in a laboratory dryer at 120°C for 60 min . A UNI-T laboratory multimeter, model UT804, was used to measure the resistance of the fabricated coatings. The thickness of the layers was measured with the DektakXT Bruker BNS stylus profilometer. Microscopic observations of layers were made using a Hitachi SU8230 scanning electron microscope (SEM) with an accelerating voltage of 5.0 kV. Images were taken with SEM at magnifications of 1 × 100 and 1 × 500.RRID: CVCL_B063) were used to perform electrostimulation on graphene substrates. NE-4C neuroepithelial cell lines were obtained from brain follicles of 9-day-old mouse embryos. In this study, neural stem cells from 8 to 10 passages were used. The cells medium and culture substrate had been prepared, as described in detail in previous work (cm2), which were then incubated at 37°C in an atmosphere of 5% CO2. Cells from the control group and stimulated with an electric field were cultured for 3 days.Mouse NE-4C neural stem cells of neuroectodermal origin  with a SE secondary electron detector at an accelerating voltage of 5 kV. Specimens are observed in high vacuum and high resolution. Images were taken with an SEM microscope at 1 × 1,000 magnification.Inadequate ink viscosity results in uncontrolled spreading over the coated surface, resulting in a layer of non-uniform thickness. The composite’s viscosity depends on several factors, including the content and type of functional phase, the matrix material used and the solvent. Inappropriate viscosity degrades the mechanical and electrical properties of the graphene layer. Previous studies showed the particle size’s effect on solution viscosity and the effect of type and surfactant content on the rheological properties of inks. It also demonstrates that depending on the surfactant content, the viscosity values of the inks were in the range of 1.3–2.0 mPas. The viscosity dependence on shear rate for graphene inks with different sonication times and methods are shown in The viscosity curves of all the tested inks have the desired shape for spray-coating techniques, characteristic of shear-thinning non-Newtonian fluids. The curves represent a decrease in viscosity values as the shear rate increases, results from the gradual stacking of functional phase particles along the flow line. The viscosity decreases until no further increase in system ordering is possible – which corresponds to the Newtonian behaviour of systems at high shear rates . FurtherGraphene coatings, made using inks with different sonication times, were applied to flexible Kapton film substrates to determine the layer micro-geometry. The example images of graphene coatings on polystyrene substrates made using inks sonicated with a sonotrode are presented in Electrical conductivity is a crucial factor in using graphene substrates in the electrostimulation of nerve cells. As-low-as-possible resistance of conductive culture substrates is sought to increase the efficiency of stimulation and allow the use of a lower current signal to reduce potential cell weakness caused by the flow of an extensively strong electrical signal . AchieviTo determine resistivity, the thickness of layers was measured using a profilometer. The average thickness of the coatings was 37.46 μm.The measurements show a significant scattering of resistivity values for different sonication times. As the sonication time increases in the ultrasonic bath, a visible rise in coating resistance is present. The highest resistance values were obtained for inks sonicated for 90 and 120 min using the ultrasonic bath. On average, the resistance values of the coatings made with the inks sonicated for 120 min were twice as high as the resistance values achieved for the inks sonicated with the sonotrode. The lowest resistance values were obtained for the coatings made with the inks sonicated using a sonotrode and ultrasonic bath for 30 min; therefore, both were used for further electrostimulation studies.cm2) is also included in the graph (red column – seeded cells).To evaluate the effect of electrostimulation on NE-4C cells, the cell counts were examined on substrates without graphene coating, on graphene substrates without stimulation and with stimulation . To faciFirstly, it can be seen that the use of graphene substrates significantly increases cell counts. The use of graphene substrates alone increases cell counts more than threefold. When applying graphene substrates, cell counts approximating those on polystyrene control substrate without graphene coating were obtained. A high increase in cell number is observable for the graphene substrate prepared with ink sonicated using a sonotrode.For both types of substrates, the highest population size was obtained for cells stimulated with the lowest voltage amplitude of 5 V. More than the sevenfold increase of the cell count (compared to seeded cells) was obtained using 5 V electrostimulation and graphene substrate prepared using the ink sonicated in the ultrasonic bath. In the case of the other substrate, an almost fourfold increase in cell count was obtained. Slightly lower cell counts were obtained with a stimulation voltage of 10 V. The weakest stimulation effects, for both types of substrates, were obtained using the highest voltage amplitude of 15 V. The 15 V sonotrode substrate was the only one in which the decrease in cell count was observed. In the 15 V US bath substrate, the cell count only slightly higher than the number of seeded cells was observed.Observations of selected culture substrates were also made using SEM microscopy to assess the effect of electrostimulation. The cell count study and their microscopic observations confirmed the positive effect of graphene substrates and electrostimulation on neural stem cell behaviour. Using graphene substrates increases cell counts by 300% (without electrostimulation) and over 700% by using 5 V stimulation. Graphene layers improve the shape and adhesion of cells to the substrate. It is clear that lower values of stimulation voltage amplitude improve the cell behaviour. The highest cell count for both types of substrates was obtained when electrostimulation with a voltage amplitude of 5 V was applied. Therefore, these substrates were selected for further observations using SEM microscopy.The literature analysis of graphene surface application evidences a high demand for innovative solutions in the field of neural tissue engineering. The production of a biocompatible layer supporting the development of nerve cells, promoting their adhesion and proliferation will allow for significant improvements in the treatment of many such neurodegenerative diseases as neural tube defects, spinal cord injuries, or neuropathies. Achieving the desired homogeneity of the produced composite is crucial for the substrate’s conductivity, which makes them useful in the electrostimulation of nerve cells. The properties of graphene layers are also affected by the composite fabrication process itself particularly the time and method of solution homogenisation.The studies showed a significant influence of the graphene ink preparation method on the properties of the graphene coatings. Here, two alternative ultrasound sonication methods were used, which allowed for high dispersion of graphene particles, resulting in homogeneous, highly conductive graphene layers successfully used in the electrostimulation of neural stem cells.While comparable ink viscosity values decreased with the increasing homogenisation time. Many factors can influence the rheological properties of fluids, including the particle’s shape and size, the molecular weight of the functional phase or particle size distribution in solution . The visDespite the lack of differences in ink viscosity, apparent differences were observed in the conductivity of layers produced at different sonication times. As the viscosity decreased (with longer sonication times), the conductivity of the layer decreased. This is presumably related to the degree of graphene particle dispersion in the produced inks. The sonication time probably affected the fragmentation of graphene nanoplatelets and also changed the degree of the graphene particles’ surroundings by the surfactant, thus influencing the formation of particle agglomerates. According to Nevertheless, each manufactured ink fulfils the rheological requirements enabling its application in the spray coating technology. The sonication time and method did not cause significant discrepancies in the viscosity values of the inks. It can be concluded that both sonication methods are effective ways to homogenise solutions. The relationship between viscosity and conductivity of the layers requires further investigation and additional studies – including the size of graphene nanoplatelets obtained in ink after the sonication process.Observations of the micro and macro geometries of the substrates conclude that both sonication methods can be successfully used in the homogenisation of graphene inks. However, the taken photographs do not allow an assessment of the effect of sonication time on the dimensions of the graphene particles themselves. Particle dimensions are an essential parameter for coating applications in tissue engineering, as they could strongly influence the occurrence of cytotoxic phenomena. Therefore, in future work, it is necessary to perform SEM photography with higher accuracy to measure the size of the flakes.–3 Ω⋅m and 2,154 ⋅ 10–3Ω ⋅m, respectively, for sonotrode and ultrasonic bath. The results show that a short, 1-min sonication process using a homogeniser or extending the process to 30 min when using an ultrasonic bath is optimal for the accurate combination of surfactant particles with graphene flakes. The study showed that the obtained resistance values strongly depend on the graphene ink’s method and sonication time. With an extension of the sonication time, apparent increase in the resistance of the coatings can be seen. This phenomenon may be due to the excessive fragmentation of graphene nanoplatelets, as flakes with smaller dimensions are weaker conductors.The lowest resistivity values were obtained for the coatings made with the ink sonicated using a sonotrode for 1 min and an ultrasonic bath for 30 min. The obtained resistivity of the films were 1,623 ⋅ 10Through the successful use of a sonotrode, the efficiency of the fabrication process of graphene heterophasic inks was increased by shortening the sonication time from 90 to 2 min while increasing the conductivity of the layers. Comparing the fabrication times of the inks made in our previous work (90 + 15 min sonication) , one canUpon analysing the results, graphene substrates and electrostimulation significantly increase the cell counts. The simulations carried out in this work, using different stimulating voltages with amplitudes, showed over a 700%-cell-count increase when 5 V was applied. The produced substrates improved cell adhesion and morphology, as shown in the SEM photographs in the section below. The fabricated graphene substrates had a beneficial effect not only on cell adhesion but also on cell proliferation. Using graphene substrates enabled to increase cell count by 300% (without electrostimulation).The occurrence of functional groups on graphene surfaces could influence the types of cell surface receptors and proteins, such as the cellular adhesion molecules (CAM), which cells use to bind to graphene surfaces . Lee et Cell adhesion plays a significant role in regulating such processes as growth, proliferation and migration. The increase in the number of cells on graphene illustrates a positive effect on cell adhesion, also confirmed by For both types of substrates, a decrease in cell count is observed as the stimulation voltage amplitude increases. This observation indicates that the stimulation voltage amplitude of 15 V is too high and probably hinders the cells from adhering correctly to the substrate, thus preventing their proliferation, which is also stated by In the study, the 5 V voltage amplitude was used. However, the literature reports confirm the efficiency of even lower stimulating voltages . This isIn conclusion, we can state that using graphene substrates as cellular scaffolds enhance and improve neural tissue regeneration protocols. The preliminary data showed the efficiency of the proposed simulation method and the beneficial effects of graphene substrates and electrical stimulation. We propose an alternative tissue stimulation method, which could be a starting point for new, innovative treatment routes. The high homogeneity and conductivity of the layers allowed them to be effectively used in the electrostimulation of neuronal stem cells. The increased cell count of cells cultured of the graphene substrates indicates that our substrates provide an environment suitable for cells’ development, mainly supporting cell adhesion, which enables their proper growth and proliferation. Graphene substrates show significant potential in cellular scaffolds and regenerative medicine, which in the future may contribute to the development of effective clinical therapy for peripheral nerve injuries.The original contributions presented in this study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.ŁD-S: conceptualisation and methodology. WS: writing—first draft, experiments—materials preparation and spray coating, and figures preparation. AG: experiments—cell culture. JK: formal analysis. WS and JK: writing—review and editing. ŁD-S and MJ: supervision and funding acquisition. All authors contributed to the article and approved the submitted version.


In the design, graphene is served as excellent carrier to improve the catalysis of Mn3(PO4)2 nanoparticles; and DNA adsorbed on graphene acts as medium to assist the growth of Mn3(PO4)2 on graphene. The fabricated graphene/DNA/Mn3(PO4)2 composites exhibit excellently electrochemical activity, significantly decrease the response time and increase the sensitivity of the sensor towards O2˙−. The successful detection of O2˙− released from cancer cell demonstrated its potential applications in biology and medicine.In this work, we successfully fabricate a rapid, sensitive sensor for the detection of superoxide anions O 2˙− based on graphene/DNA/Mn3(PO4)2 biomimetic enzyme.In this work, we successfully fabricate a rapid, sensitive sensor for the detection of superoxide anions O Under normal conditions, O2˙− is highly reactive and unstable and its metabolism is a rapid and spontaneous process. Overproduction of O2˙− can cause various disease, such as aging, asthma, ulcer disease, cancer, atherosclerosis, neurodegenerative diseases and other diseases.1–4 Thus the detection of O2˙− is very important in various biological systems. Many methods have been developed for the detection of O2˙− including chemiluminescent, spectrophotometric, fluorometric, and electrochemical technique, etc.5–8 Among all these techniques, electrochemical method receives most attention due to its advantages of real-time assay, high sensitivity and selectivity. And most of the reported electrochemical methods for O2˙− detection are based on enzyme catalysts such as cytochrome c and superoxide dismutase (SOD).2,9–11 Though enzyme improves the assay performance of sensors, but its high cost and poor long-term stability increase difficulty of direct monitoring of O2˙− in biological samples. Therefore, it is a serious challenge for us to establish a fast, reliable, and sensitive non-enzyme approach for O2˙− monitoring in physiological and pathological processes.Superoxide anions 2) has the ability to catalyze the dismutation of O2˙− compared with free Mn2+ ion that only stoichiometrically reacts with O2˙−.13 Therefore Mn3(PO4)2 is usually selected as a substitute for enzyme to detect O2˙−.14 Mao et al. synthesize SiO2-Mn3(PO4)2 nanoparticles derived from phytic acid and utilize for O2˙− detection.15 Lan and co-workers deposit Mn3(PO4)2 nanoparticles on carbon nanomaterials directly for the fabrication of O2˙− sensor.16 These works verify the catalysis of Mn-superoxide dismutase mimics towards O2˙−. But the synthesis and property of nanoparticles need further study to improve the catalytic performance of sensor. In this work, we synthesize Mn3(PO4)2 nanoparticles by DNA induction, which results in even dispersion and excellent catalytic activity towards O2˙− disproportionation reaction.As early as 1982, manganese was reported possessing an effective catalytic effect 17–20 However, the two dimensional structure is easy to aggregate due to the single-atom-thickness, and hydrophobic aromatic structure. To solve this problem, we functionalize graphene with deoxyribonucleic acid (DNA) which can be adsorbed on graphene through π–π stacking and don't destroy the intact structure of the carbon materials.20 Under the induction of ssDNA, Mn3(PO4)2 nanoparticles was evenly deposited on the surface of graphene. The obtained graphene/DNA/Mn3(PO4)2 nanocomposites displayed significant biomimetic enzyme activity, rapid and sensitive response towards O2˙−. This approach holds a great promise for broad applications in biomedical research and clinical test.Graphene is an increasingly important nanomaterials due to its excellent electronic conductivity, good stability, and promising catalytic performance in sensing.4), potassium phosphate tribasic (K3PO4) and Nafion were purchased from East Sichuan Chemical Industry (Group) Co., Ltd.  and used as received. Graphene was obtained from Sinocarbon Materials Technology Co., Ltd., China. Zymosan A  SOD, DNA , and potassium superoxide were purchased from Sigma-Aldrich and used without further purification. The O2˙− solutions were prepared by dissolving KO2 in PBS solution . The concentration of O2˙− was determined by the reduction of ferri cytochrome c spectrophotometrically.21 All the other solutions were prepared using deionized water (18 MΩ cm) and were degassed with high purity nitrogen before experiments. All electrochemical experiments were carried out at room temperature.MCF-7 was bought from Cell bank of the representative culture preservation committee of the Chinese Academy of Sciences, China. Manganese sulfate (MnSO3(PO4)2 were characterized by transmission electron microscopy . Fourier transform infrared spectroscopy (FTIR) spectra were determined using a Thermo Nicolet 6700 FTIR spectrometer. Electrochemical performances were characterized using a CHI 660 system . Graphene/DNA/Mn3(PO4)2 modified glassy carbon electrode (graphene/DNA/Mn3(PO4)2/GCE) served as working electrodes, platinum sheet as counter electrode and Ag/AgCl (in 3 M KCl) acted as reference electrode. All O2˙− measurements were performed in 10 mL 0.01 M PBS solution (pH = 7.4).Scanning electron microscopy (SEM) images were taken by JSM-6510LV, Japan. Field emission scanning electron microscopy (FESEM) images were taken by JSM-7800F, Japan. X-Ray diffraction (XRD) measurements were performed on a Shimadzu diffractometer  operating in reflection mode with Cu Kα radiation at a step size of 0.06 per second. The nanosheets of graphene/DNA/Mn3(PO4)2 nanosheets were prepared by the previously reported method.22 Thirty milligrams of DNA was dissolved in 30 mL de-ionized water under stirring, after which the solution was annealed at 95 °C for 15 min to produce single-stranded DNA (ssDNA). Then the obtained ssDNA was mixed with 15 mL of 1 mg mL−1 graphene by mildly sonicated at 4 °C for 2 h, thereafter filtration and washing were performed to remove excess ssDNA. Subsequently, the mixture was dispersed into 0.1 M MnSO4 solution, then 0.1 M K3PO4 solution were dropwisely added under stirring and kept at room temperature for 30 min. Then graphene/DNA/Mn3(PO4)2 nanosheets were obtained by centrifugation at 8000 rpm for 10 minutes and washing with deionized water for 3 times.Graphene/DNA/Mnd = 3 mm) was polished with alumina slurry to a smooth and bright surface, washed by sonication for 30 s and dried under nitrogen. Then 5 μL of the obtained graphene/DNA/Mn3(PO4)2 suspension was dropped on the electrode surface and dried at room temperature. Finally, 5% Nafion solution was coated to stabilize the fabricated electrode.Glassy carbon electrode  at 37 °C. The cells were cultured in Dulbecco's Modified Eagle's Medium (DMEM)  supplemented with 1 mol L−1 glutamine, 50 U mL−1 penicillin/streptomycin and 10% heat inactivated fetal bovine serum.14 For the detection of O2˙− released from living cell, the incubation solution was removed and washed with PBS solution (pH 7.4) for three times. Before electrochemical measurement, zymosan (Zym) was added to motivate cells generation of O2˙−. Amperometric response was recorded by CHI-660B electrochemical station at applied potential of 700 mV (versus Hg/Hg2Cl2).MCF-7 cells were cultured in a humidified incubator (95% air with 5% CO3(PO4)2 enzyme mimics by growth of Mn3(PO4)2 on graphene under the induction of DNA. As we all know, single-strand DNA (ssDNA) can be adsorbed on graphene by π–π interaction.23,24 Here ssDNA was prepared by annealing double-strand DNA (dsDNA) and applied for graphene modification. In the presence of Mn2+, the divalent cations accumulate along the ssDNA backbone by the electrostatic interaction, which facilitate the formation of Mn3(PO4)2 crystal nucleus on graphene and growth of Mn3(PO4)2 nanoparticles upon the addition of negatively charged PO43− groups.3(PO4)2 nanosheets is characterized with scanning electron microscope (SEM). 4 and K3PO4, typical wrinkle of graphene can be clearly observed and no evident Mn3(PO4)2 nanoparticles is found in 3(PO4)2 nanoparticles on graphene surface without DNA. However, when ssDNA is modified on graphene (3(PO4)2 nanoparticles can be evidently observed on the surface of graphene. The phenomena reveal the crucial role of ssDNA on the growth of Mn3(PO4)2 on graphene. The TEM images in 3(PO4)2 nanoparticles due to the overlapping of particles. In order to verify the results, energy disperse spectrometer (EDS) was performed to explore chemical composition of the materials. The EDS results in Fig. S23(PO4)2 nanoparticles. The results further confirm the formation of well-defined thin-layer structure of the nanocomposites of Mn3(PO4)2 nanoparticles on graphene surface.The morphology of the as-prepared graphene/DNA/Mngraphene , even-di3(PO4)2 nanosheets. As shown in −1 can be assigned to the aliphatic carboxylic acid salts, the peaks at 996.0 cm−1 and 3196.3 cm−1 were attributed to inorganic phosphates. The results suggest that DNA and Mn3(PO4)2 have been successfully modified on the graphene. In addition, the X-ray diffraction pattern (XRD) of graphene/Mn3(PO4)2 in 3(PO4)2. The results further confirm that Mn3(PO4)2 nanoparticles are successfully modified on the surface of graphene. Then the zeta potential at various stages of the synthesis were measured to further characterize the formation of the nanocomposites. The results in Fig. S13(PO4)2 was deposited, the zeta potential increase to −15.6 mV, indicating the formation of graphene/DNA/Mn3(PO4)2 nanocomposites.Then FTIR spectra are determined to further verify the formation of graphene/DNA/Mn2˙− and the mixture of O2˙− and SOD (red column) in PBS. When graphene/DNA/Mn3(PO4)2 nanosheets were modified on GCE, the obtained modified electrode displayed evident redox peaks around 0.7 V and 0.5 V in the absence of O2˙− 2 nanosheets towards O2˙−. These results verify the biomimetic enzyme activity of graphene/DNA/Mn3(PO4)2 to catalyze the dismutation of O2˙−.253(PO4)2 nanosheets, graphene/Mn3(PO4)2 were synthesized by direct depositing Mn3(PO4)2 nanoparticles on graphene in the absence of DNA, DNA/Mn3(PO4)2 nanoparticles were by prepared depositing Mn3(PO4)2 on dsDNA template and graphene/DNA/Mn2+ was synthesized by adsorbing DNA on graphene and subsequent depositing Mn2+ on DNA. Electrochemical performance of these materials were explored with electrodes modified with the above three nanomaterials (3(PO4)2 (graphene/Mn3(PO4)2/GCE) possesses excellent electrical conductivity but the current response towards O2˙− is weak, verifying the good conductivity of graphene and poor modification of Mn3(PO4)2 (3(PO4)2/DNA and graphene/DNA/Mn2+ display evident current response to O2˙−, but the current response is lower than that on graphene/DNA/Mn3(PO4)2/GCE (as shown in 3(PO4)2 nanosheets. In addition, in comparison with the Mn3(PO4)2/DNA/GCE (3(PO4)2/GCE 2 . Similar/DNA/GCE , we can O4)2/GCE  displays3(PO4)2 nanoparticles on graphene can be ascribed to two factors. First, the π–π interaction between DNA and graphene induces ordered assembly of DNA on the surface of graphene, which is a critical factor to the synthesis of Mn3(PO4)2 nanoparticles. Next, Mn2+ is adsorbed on DNA backbone to assist the nucleation of Mn3(PO4)2 nanoparticles. In addition, in comparison with other conventional method, the DNA induced synthesis is easier to control the morphology and dispersion of products by adjusting temperature, pH, or DNA concentration.The reason for DNA facilitating the growth of Mn3(PO4)2/GCE towards O2˙− in 10 mM PBS (pH = 7.4). 2˙− concentration from 5 nM to 400 nM. From the results in 2˙− concentrations in the range of 5 nM to 400 nM with a regression equation expressed as I (μA) = 0.00354 c (nM) + 0.1504 (R2 = 0.999). The detection limit of the assay is calculated to be about 1.67 nM (S/N = 3) with a sensitivity of 3.54 μA μM−1, which is much more superior to most of previous reports.14,23,26–28 The response time of the electrode to O2˙− is as short as 5 s, which is an advantage for the detection of O2˙− due to its short lifetime. These results demonstrate that the present sensor may meet the requirement of O2˙− detection in normal physiological conditions.29Chronoamperometry is utilized to investigate the electrochemical response of graphene/DNA/Mn+, Na+, Cl−, NO3−, SO42−, hydrogen peroxide (H2O2), glucose (Glu), glutathione (GSH) uric acid (UA) and ascorbic acid (AA). Among these interferences, H2O2, UA and AA possess good electrochemical activity and perplex the detection of O2˙− due to their wide existence in biological systems. As shown in 2˙−, the presence of 10 μM K+, Na+, Cl−, SO42−, NO3−, Glu. GSH, AA, UA and H2O2 did not cause any noticeable current response. The result suggests that the proposed sensor possesses excellent specificity to O2˙− detection in biological systems. In addition, the reproducibility and stability of the electrode were explored. The electrochemical response of the sensor was performed using five different electrodes fabricated at the same time, the standard derivations of current response was lower than 5.2%. When the electrode was stored at 4 °C for one month, the current response towards O2˙− decrease 6.8% of original value  of zymosan , a significant current response was observed (corresponding to O2˙− oxidation). According to the calibration curve in 2˙− released from MCF-7 cells is calculated to be around 156.38 nM, 315.71 nM and 631.81 nM, respectively 2 nanosheets is successfully developed. The synthesized nanosheets display strong electrocatalytic activity towards O2˙− with high sensitivity, excellent selectivity and fast current response. The successful determination of O2˙− released from cancer cells demonstrates the great potential for application in biological system. This work provides a new method for the fabrication of biomimetic enzyme sensor and a great promise for biosensing and biomedical application.A novel OThere are no conflicts to declareRA-008-C7RA12962A-s001


Chronic cough is experienced by most patients with idiopathic pulmonary fibrosis (IPF). It is often the first symptom and is associated with reduced quality of life, increased rates of depression and anxiety, more severe physiological impairment, and disease progression. Although not fully understood, recent gains in understanding the pathophysiology of chronic cough in IPF have been made. The pathophysiology is likely multifactorial and includes alterations in mucous production and clearance, architectural distortion, and increased cough reflex sensitivity, suggesting a role for targeted therapies and multidisciplinary treatment. Modifiable comorbidities can also induce cough in patients with IPF. There is a renewed emphasis on measuring cough in IPF, with clinical trials of novel and repurposed therapies for chronic cough emerging in this population. This review provides an update on the clinical characteristics, pathophysiology, and measurement of chronic cough in patients with IPF and summarizes recent developments in non-pharmacological and pharmacological therapies. Idiopathic pulmonary fibrosis (IPF) is a chronic fibrosing idiopathic interstitial pneumonia, most prevalent after the 6th decade of life. It is associated with significant morbidity and mortality, with progressive worsening of respiratory symptoms over time. Cough is a common symptom, often occurring years prior to diagnosis, contributing to poor prognosis and reduced quality of life. The severity of cough encompasses its frequency, intensity, and impact to patients' lives . Cough fCough and dyspnoea are the two most common symptoms described in patients with IPF, with up to 80% experiencing chronic cough . Cough iCough is more prevalent in patients with advanced IPF . IncreasCough due to IPF is typically described as dry or non-productive and refractory to antitussive therapy. A minority of patients have a chesty cough from the onset of disease. The severity of cough in IPF is greater than in other types of interstitial lung disease (ILD) –8. The iPatients identify cough as a key symptom associated with physical and emotional distress, with a more limited appreciation by health care professionals . Physicavia rapidly adapting receptors, slowly adapting stretch receptors, and c-fibers in the afferent pathway that conduct mechanoreceptor and chemoreceptor stimuli to the medullary control center by branches of the vagus nerve  gene, the most well-validated genetic risk factor for IPF –35, incrIPF-related comorbidities and other health conditions can be important contributors to cough in this population and are therefore key targets for treatment. The role of GOR in IPF remains unsettled, with inconclusive results on therapeutic effects of antacid medication and anti-reflux surgery on progression-free survival, quality of life, and all-cause mortality in this population , 52. As The prevalence of GOR is higher in IPF than in other chronic lung diseases , 54, 55 P and mast cell-tryptase are both elevated in the sputum of patients with chronic cough and comorbid GOR, which inversely correlate with cough threshold  on lung function testing may prompt further investigation with computed tomography of the chest to assess for acute exacerbation, infection, or disease progression. Red flag symptoms for lung cancer include haemoptysis, dysphonia or dysphagia, and weight loss. Relevant comorbidities  and the Leicester cough monitor are two validated monitors , 71, whiPatient reported outcomes (PROs) have gained importance in recognition of their value to capture patients' perceptions of their symptoms and health status. They are measured using standardized questionnaires, which may be generic, disease-specific, or symptom-specific. PROMs for cough have been developed in other conditions such as lung cancer, while no cough-specific instruments have been developed and validated prospectively in the IPF population. The currently used cough questionnaires were developed in patients with chronic cough and other respiratory conditions such as chronic obstructive pulmonary disease. Instead of comprehensively reporting on all PROMs for cough that have been used in IPF, this review focuses on selected commonly used cough-specific instruments, including the VAS, LCQ , Cough QVAS subjectively assesses cough severity and is popular due to its simplicity. Patients are asked to place a vertical mark on a 0–100-mm linear scale reflecting their cough intensity or frequency, with higher scores indicating greater severity. It can be used to assess both the status and changes in cough severity. There is limited robust data on its reliability and validity in specific respiratory conditions, including asthma and chronic obstructive pulmonary disease . The VASThe LCQ is the most widely used validated cough instrument in clinical and research settings. Its advantages are that it is relatively easy to administer, translated into many languages, and used in many cultures. It is a self-administered questionnaire consisting of 19 items covering three health domains  using a 7-point Likert response scale for each item. The total LCQ score ranges from 3 to 21, with lower scores indicating greater impairment in health status because of cough. All domains of the LCQ inversely correlate with objective daytime cough frequency  in patieThe CQLQ consists of 28 items and six health domains . Each item is rated using a 4-point Likert scale. The CQLQ has good internal consistency in IPF but this does not apply to all domains . InternaThe CASA-Q comprises four domains: symptom and impact for cough and sputum. The score for each domain ranges from 0 to 100, and lower scores indicate worse symptoms. It has been used in landmark trials for antifibrotic therapies in IPF , 92 withClinical trial design in IPF has historically been hampered by a lack of consensus on appropriate outcome measures to reliably assess treatment response . For exaThere is no therapy for cough in IPF that has proved effective in clinical trials. Aside from treatment of relevant comorbidities, several therapies have been tried for chronic cough in IPF and may be useful in some individuals, which include both non-pharmacological and pharmacological interventions . It is sWhile the evidence is scarce in patients with IPF, multimodality speech therapy, an established therapy for unexplained chronic cough, may be a suitable treatment. Speech therapy has been reported to decrease cough frequency and improve quality of life in chronic cough . The PhyThe outlook for patients with IPF changed with the availability of two antifibrotic drug therapies; nintedanib and pirfenidone. Nintedanib alters cell signaling mediated by tyrosine kinases resulting in the inhibition of mechanisms fundamental to the development of fibrosis . PirfeniStudies evaluating antitussive therapies in IPF are limited. Opioids have been shown to decrease cough frequency and improve cough-related quality of life in patients with chronic cough in a study of slow-release oral morphine at a dose of 5 mg twice daily . Common via high-efficiency nebuliser reduced mean daytime cough frequency by 31% at 14 days in patients with IPF and was generally well-tolerated (In a randomized placebo-controlled trial of 24 patients with IPF, low-dose thalidomide was shown to improve cough severity and respiratory-related quality of life measured by the CQLQ and VAS scores . Howeverolerated . This beolerated . An oldeolerated . Howeverolerated , and itsolerated . Furtherolerated .Cough is a common and often under-recognized symptom of IPF, which is associated with disease progression. Identified as a central symptom by patients and their caregivers, cough impacts health-related quality of life. Careful and timely clinical evaluation and assessment using validated tools that objectively and subjectively measure cough in this specific patient population are recommended. Currently available measurement tools for chronic cough have been suboptimally validated in patients with IPF. The development of disease-specific PROMs for cough in the IPF population is required to accurately and reliably show the difference between competing therapies. Further clinical research into effective non-pharmacological and pharmacological interventions for cough is vital to improve symptom management and health outcomes for patients with this progressive disease. In light of the limited proven efficacy of single therapeutic agents for cough in IPF, future clinical trials may consider multimodal interventions or combination drug therapy in this patient population.YK and JM conceived the presented idea. JM performed the literature search and wrote the manuscript with support and critical feedback from YK. Critical feedback was also provided by NG and AH. All authors contributed to the final manuscript.YK reports grants and personal fees from Boehringer Ingelheim, personal fees from Roche, other from Air Liquide Healthcare, outside the submitted work. NG reports personal fees from Boehringer Ingelheim, Roche, AstraZenaca, and Novartis, other from Air Liquide Healthcare, outside the submitted work. AH reports non-financial support from BOC Ltd Australia and Air Liquide Healthcare, outside the submitted work. The remaining author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Realizing the neurological information processing by analyzing the complex data transferring behavior of populations and individual neurons is one of the fast-growing fields of neuroscience and bioelectronic technologies. This field is anticipated to cover a wide range of advanced applications, including neural dynamic monitoring, understanding the neurological disorders, human brain–machine communications and even ambitious mind-controlled prosthetic implant systems. To fulfill the requirements of high spatial and temporal resolution recording of neural activities, electrical, optical and biosensing technologies are combined to develop multifunctional bioelectronic and neuro-signal probes. Advanced two-dimensional (2D) layered materials such as graphene, graphene oxide, transition metal dichalcogenides and MXenes with their atomic-layer thickness and multifunctional capabilities show bio-stimulation and multiple sensing properties. These characteristics are beneficial factors for development of ultrathin-film electrodes for flexible neural interfacing with minimum invasive chronic interfaces to the brain cells and cortex. The combination of incredible properties of 2D nanostructure places them in a unique position, as the main materials of choice, for multifunctional reception of neural activities. The current review highlights the recent achievements in 2D-based bioelectronic systems for monitoring of biophysiological indicators and biosignals at neural interfaces.  The rapid advancements in neuroscience and bioinspired neurotechnologies require the accurate monitoring, reception and decoding of neurological potential signals of the nervous system . In thisThe advent of the first generation of neural interfaces was considered as a key development toward the realization of electrical properties of cell and tissues that enabled in vitro neural signal acquisition. The voltage clamp technology, developed in 1976 by Erwin Neher, enabled us to record single ion channel signals . In 1981Neural signal recording can be divided into two main categories, i.e., intercellular and extracellular recordings. The intracellular technique records the electric signals generated across the cell membranes; therefore, the electrodes are placed inside and outside of the membranes. In a different configuration, extracellular recording is enabled through the employment of recording electrodes in the extracellular fluid for the reception of signals generated by nearby neurons. There are several main differences between intracellular and extracellular recording. The intracellular neural signals are more localized and feature more powerful amplitudes, while the extracellular signals are weaker and originate from a large area of the brain. The extracellular recording of neural signals can be divided into two main categories of invasive and non-invasive methods based on the location of electrodes. In the non-invasive neural interfacing, the signals are collected via electrodes assembled on the top surface of the scalp b to recoThe invasive neural interfaces can be categorized into penetrating and non-penetrating sensors c–e 27,2,29. The From a technical point of view, an efficient and reliable neural interfacing process can be achieved through the development of biocompatible sensors with high signal-to noise-ratio (SNR) characteristics and low and stable impedance values . Figure Studies on multifunctional biosensing probes have fueled a continuous need for new materials. These materials are expected to combine multiple sensing and stimulation functionalities and simultaneously create chronic biocompatible interfaces with neural cells and brain tissue. In this regard, 2D nanomaterials are among the main advanced atomically thin nanostructures that hold great promise for the development of highly active and miniaturized sensors ,50,51,52Graphene, the most famous carbon-based 2D structure, is recognized as a functional material for neural interfacing applications . Its excHigh-temporal-resolution recording of neuron cell activities is one of the main targets of electrophysiology, where optical and cellular imaging techniques are accompanied by electrophysiological signal recordings to complement the performance of each individual technique and then provide a clear image of the neurological behavior ,61,62. HFluorescence microscopy can be combined with invasive graphene-based microelectrodes to visually depict the precise active location of neural signals . Calcium−2 was recorded for a 3D graphene-based electrode that was employed for in vivo stimulation of motor cortex arrays of mouse leg ..45]. ThiIn parallel with microelectrodes, GFETs are being developed for a wide range of biosensing applications, where the transduction of biological signals is amplified locally due to the strong field effects induced ,72,73,74−2 at the 1 kHz frequency), while it showed considerably better charge storage capacity (361 ± 45 mC cm−2) compared with that of the conventional metallic-based filament microelectrodes used in neural interfacing systems. Similar research studies also included graphene as the main sensing component in microfibers [The availability of various types of graphene-based materials, including single-layer CVD films, multilayered exfoliated films, and chemically and mechanically synthesized films, has enabled the development of various types of graphene-based neural interfaces. Despite the fact that a large number of bioresearch activities have been devoted to graphene, the high hydrophobicity of pristine graphene has limited its applications. Functionalized graphene, graphene oxide (GO) and reduced graphene oxide (rGO) 2D structures can be valuable alternatives to graphene in neural interfaces. Two-dimensional GO is an atomically thin structure where oxidation enables the incorporation of oxygen functional groups into the surface of the graphene, thus decreasing the graphene aggregation in aqueous solutions as well as reducing its electrical conductivity. Recently, ultra-large graphene oxide flakes were deposited on binder-free microfibers to replace conventional filament-shaped metallic invasive neural electrodes ,65. In trofibers ,77.Despite its greater functionalization capabilities, GO has highly restricted electron mobility on its surface. GO is technically an insulator; therefore, reduction reactions are carried out to partially remove oxygen and then alter or recover the electrical conductivity of 2D GO films. Consequently, reduced graphene oxide (rGO) with higher conductivity is extensively employed for biosensing and especially non-invasive neural interfacing applications. Typically, 2D rGO nanostructures are developed or grown during the fabrication process for neural interfaces at the heterointerfaces between the sensing components, which in turn facilitate swift charge transfer and low contact impedance at the neural interfaces. Non-invasive graphene-based polymer wearable sensors are well-developed for the characterization and precision sensing of biosignals with high levels of accuracy ,21,22. O−2) [2O3) nanocomposite electrodes as implantable neural interfaces for applications in electrophysiology and neurochemistry. The rGO/Au2O3-modified electrodes displayed significantly improved activities [Due to the facile dispersion of GO and rGO, these structures can be feasibly used for the fabrication of fiber neural interfaces. In such development, the fibers  are dip-coated in GO or rGO dispersions, followed next by thermal treatment. These microfiber neural interfaces showed considerably improved conductivity and low impedance for wearable neural interfacing ,87,88,89−2) . Anothertivities . CurrentTwo-dimensional materials beyond graphene represent atomically thin nanostructures with remarkably novel semiconducting, insulating and metallic characteristics that put them in a distinguished position in comparison with their carbon-based counterparts. The family of 2D materials beyond graphene covers various type of nanostructures, transition-metal dichalcogenides (TMDCs) and MAX phase 2D materials (MXenes). These 2D structures recently found their way into neural interfacing technologies . The fol2 chemical formula, where M is one of the transition metals , and X is one of the chalcogen elements . The electronic characteristics of 2D TMDCs are of particular interest, since they differ dramatically from those of their bulk and multi-layered structures and even their single-layered 2D structures [TMDCs are the well-known and perhaps the most eminent 2D materials beyond graphene . TMDCs aructures ,98,99. Eructures ,101,102.2 and PtTe2, have attracted researchers’ attention due to their electronic characteristics. The ultra-thin few-layered PtTe2 has shown metallic properties attributable to the strong interactions between its Te interlayer chalcogen atoms [2 is generally a semiconductor material that undergoes semiconductor-to-metal transition when the number of its layers is increased [2 and PtTe2 ultra-thin films on polymer-based substrate at temperatures of less than 400 °C has allowed the development of flexible neural interfaces based on these 2D materials [2 and PtTe2 were initially grown by CVD on Si/SiO2 or Kapton substrate at 400 °C. The developed films on Si/SiO2 were afterwards transferred to 200 nm thick poly methyl methacrylate (PMMA) flexible substrate to fabricate wearable sensors  forces between the 2D layers of MoS2 films made it possible to fabricate free-standing 2D MoS2 devices. The weak vdW bonding forces enabled the rotational and sliding motion of 2D layers and, therefore, facilitated the high level of flexibility of thin films. The free-standing vdW 2D films also showed a well-behaved linear relationship and displayed a gain in tensile strength of 43% with a measured Young’s modulus of 47.3 MPa [2 films and underlayer films provided exceptional electronic functionalities for applications on living organisms [2 transistors attained a considerable SNR of 49.8 dB during human motion. Skin-gate transistor neural interfaces developed on the basis of 2D MoS2 films demonstrated versatile performance for biosensing, biopotential amplified signals and motion analyzing/processing targets [Recently, two types of Pt-based TMDCs, i.e., PtSeen atoms ,104,105.ncreased . Apart faterials ,109,110.aterials  develope sensors b. The ne sensors c. Au sof sensors c. The he sensors d. The no8.6 Ω·cm e were reectrodes f. It wasectrodes g of natudicators . In a reDC films . These 247.3 MPa . The vdWrganisms . The dev targets .n+1AXn, where M belongs to the group of early-transition metals , A is an element from group 12–16 of the periodic table, and X is either carbon or nitrogen, or both of them. The “n” in the atom indices can be 1, 2 or 3. While the bulk of MXenes represent metallic conductive characteristics, the 2D MXene films show various electronic properties, ranging from dielectric to semi-metal behavior. By implementing these tunable chemical characteristics, it is possible to orchestrate a combination of surface characteristics originated from the presence of surface-terminating functional groups on the 2D MXene films similar to hydroxyl (−OH), oxygen (−O), or fluorine (−F) groups. This prepares these 2D nanostructures for facile surface functionalization and modification. The hydrophilicity characteristics of 2D MXenes provide an excellent opportunity for solution-based processing techniques, and consequently, the need for functionalization treatments by super-acids and surfactants is eliminated [MXenes are novel 2D materials representing a broad range of atomically thin nanostructures composed of early-transition metal carbides, nitride and carbonatites. These 2D materials are synthesized from the selective etching of MAX phase materials . The geniminated ,119,120.iminated ,122.3C2 films demonstrated outstanding volumetric capacitance (1500 F/cm3 for Ti3C2Tx) [4 S/cm for Ti3C2Tx) [3C2 is by far higher than that of graphene (100 F/cm3) and graphene gel films (~260 F/cm3). A research study reported a fabrication approach for the development of high-resolution neural interfaces, where the electrode arrays were fabricated based on 2D Ti3C2. This solution-processing method was scalable and reliable for the fabrication of various types of non-invasive neural interfaces for the monitoring of EEG signals [3C2Tx acted as the active sensing material. The 2D Ti3C2Tx films were spray-coated and patterned on parylene C substrates. The measurement of on-skin sensing capabilities of the micro sensors showed impedance values of ~29.78 ± 8.1 kΩ.cm2 at the frequency of 1 kHz during the identification of muscle activation. The measured SNR of this sensor was in the range of 39–16 dB [Among the various types of MXenes, the Ti-based MXenes are the most investigated 2D nanostructures. In particular, 2D TiTi3C2Tx)  and elecTi3C2Tx) . The vol signals ,123. The39–16 dB . Specifi39–16 dB .3C2 MXene nanosheets were recently developed for the high-resolution sensing of brain signals [3C2 nanosheets was used to fabricate the sensors on flexible parylene substrates  and perform faultlessly in the human body; therefore, it is necessary to satisfy various requirements during the design of neural interfaces and selection of materials . Due to 2) was tangibly higher than that on a glass substrate (178 2) due to the stronger adhesive characteristics of graphene compared with those of glass substrate [The biocompatibility of 2D materials for neural interfacing applications needs to be exploited extensively. So far, most researchers have studied the biocompatibility of graphene neural interfaces. Their studies mostly focused on cell-to-graphene direct interactions. In a typical study, the cellular survival rates on 2D graphene-based substrates were investigated and then compared with other common substrates for neural interface applications ,132,133.ubstrate . An inteubstrate .The biostability issues in electronic neurotechnologies are of less concern, since graphene 2D nanosheets are not freely in contact with bio-tissues and are mainly encapsulated by the polymer-based insulating structures. The main group of neural interfaces is composed of multilayer graphene 2D flakes that are randomly oriented into 3D scaffold structures. In this group of neural interfaces, biocompatibility has a vital role in functional applications ,137,138.3C2 films showed no significant difference after 7 days in an in vitro experiment. In a comparative measurement between Ti3C2 and polystyrene substrates, the widespread formation of neural networks on both substrates was confirmed by immunocytochemistry [3C2 and polystyrene substrates yielded equivalent neuronal viability. These early observations indicate the capacity of Ti3C2 substrate for the formation and growth of neuronal networks with negligible cytotoxicity effects. However, the mentioned study showed that the neuron cultures on Ti3C2 experienced a reduction in neuron density compared with that of the polystyrene substrate. This indicates that neural adhesion on the Ti3C2 substrate decreased during the 7 days experiment. It also suggests that further optimization processes are required to promote neuronal adhesion on the surface of Ti3C2 as an atypical MXene film [Regarding the hydrophilicity properties of 2D materials, recently introduced MXene-based neural interfaces have been found to be capable platforms for the adhesion of neural cells. Recent experiments showed promising biomedical applications for MXene sensors in cancer theranostics , diagnoshemistry . Overallene film .e-skins. Specifically, recently fabricated e-textrode neural sensors based on graphitized electro-spun fiber/monolayer graphene clearly demonstrated their stability after washing up to 10 times. Graphene-coated textiles are well-known flexible nanostructures for fabrication of on-skin sensor electrodes for long-term monitoring of EEG signals [Another concern is the durability of 2D nanostructures on various substrates. The materials in neural interfaces are prone to delamination, corrosion, breakage, and failure of interconnections. For instance, 2D graphene-based neural interfaces suffer from delamination of 2D graphene nanosheets from the flexible polymer substrates. The Si/SiC substrates were found to be the most stable for long-term performance in bioactive media. However, rigid substrates are not desired for functional neural interfacing applications in the human body. Soft substrates or mesh-like morphologies are possibly the most appropriate options for biosensing and neural interfacing applications. Water-soluble polymer supporting layers (similar to PVA) decorated with functional 2D materials for biosensing are one of the preferable options for the next generation of on-skin dry neural interfaces. Therefore, there is a great opportunity in the fabrication of polymer-based e-skins based on 2D materials for the long-term performance of neural interfaces on human skin. Reusability is another important detail for the desired applications of novel  signals ,87,88. T signals ,143. ThiGenerally speaking, the present article provided an overview of recent advancements in the fabrication and development of various types of neural interfaces based on 2D nanomaterials. Both non-invasive and invasive neural interfaces based on 2D materials were introduced, and their functionalities were demonstrated. The multi-functionalities of 2D materials and their physicochemical properties bring both unexpected properties and improved benefits to active nanointerfaces. Recent advances have enabled rapid progress in the fabrication of micron-sized sensors with bifunctional capabilities. Graphene and its family are by far the most employed 2D structures in neural interface technology. Recent advancements in applications of 2D carbon-based nanostructures have enabled the successful monitoring of neural activities and human motion. Apart from graphene-based neural interfaces, researchers have recently focused on other types of 2D materials beyond graphene. Currently, only a few research studies have reported on the application of TMDC and MXene 2D structures in neural interface technology. Therefore, this area represents a great opportunity for the further development and progress of bioelectronic technologies. For successful development in this area of bioelectronics, it will be necessary in the future to address the challenges of fabrication technology, durability and biocompatibility of neural interfaces.


As proof-of-concept, long-term wireless respiration monitoring is demonstrated during sleep to identify symptoms of sleep apnea in wearers.A flexible sensor that can be attached to the body to collect vital data wirelessly enables real-time human healthcare management. One potential application for home-use healthcare devices is monitoring of sleep conditions to diagnose sleep apnea syndrome. Such data are not readily gathered using conventional tools, owing to the bulk and cost of instrumentation. In order to monitor respiration at home, it is necessary to improve sensing performance and long-term stability of the sensors without sacrificing wearability and comfortability. To build a platform for wireless home-use respiration monitoring, this study develops a mask-borne flexible humidity sensor using ZnIn  •A highly stable humidity sensor•Wireless wearable respiration monitoring•Per-form for sleep apnea monitoring Clinical measurement in health technology; Bioelectronics; Materials in biotechnology Continuous, real-time vital information allows detection of abnormal vital sign changes that may be caused by a disease. If such signs can be detected at an early stage, there is a greater chance of successful treatment and of saving a life. However, monitoring must measure continuous vital data during daily life in order to identify small changes. Because conventional medical instrumentation is unsuitable for home monitoring owing to bulky and expensive sensors, wearable sensors have the potential to revolutionize vital monitoring. Moreover, to enhance their wearability and comfort, wearable flexible sensors have been widely reported for healthcare applications , 2021b. 2S4 (ZIS) nanosheet-based humidity sensor  film, laser-induced graphene (LIG) electrodes, a ZIS stacking layer, and a humidity pass filter A. A wire3O+ ions and water molecules ) is relatively stable even after 20 cycles A. To ens0, where ΔR is the difference between initial resistance, R0, at 30°C in the flat state and the set temperature or bending radius, respectively. The resistance change ratio of the ZIS humidity sensor remains essentially invariant at temperatures from 30°C to 50°C  is a breathing disorder that causes human breathing to slow or stop for brief periods during sleep. If this sensor system can monitor SAS during sleep at home, it will be useful to detect the syndrome in its early stage. A wearable wireless sensor system on a mask was employed to monitor breathing rate and movement during sleep as a proof-of-concept for home-use SAS diagnosis. Three volunteers were invited to monitor their respiration rates during sleeping. They wore the mask with the sensor system, and real-time and continuous breathing rate and body motion during sleep were recorded using a smartphone A. FigureNext, reproducibility of this measurement was confirmed by repeating the monitoring from this and two other volunteers. First, breathing rate and body motion during sleep could be repeatedly monitored during sleep regardless of the volunteer , althougIn this study, a wireless, flexible humidity sensor system for real-time monitoring of sleep respiration was developed by optimizing a humidity sensor design and successfully monitoring respiration during sleep. We found that a smaller LIG interdigital electrode distance and thinner ZIS nanosheets for humidity sensing show greater stability. After confirming the stability of the sensor, wireless, real-time breathing rate and movement were monitored in three volunteers during sleep, based upon wireless humidity sensor system on a mask. This wearable humidity sensor and a smartphone app successfully recorded respiration including normal, deep sleep, and identified symptoms of SAS in a volunteer with sleep apnea. Based on real-time BPM data from the humidity sensor, trends of REM, non-REM, and SAS symptoms were clearly monitored. Although many challenges remain to replace conventional medical SAS tools, this mask-based wireless sensor system may be a good platform to advance home-use medical and healthcare diagnostic electronics.In this study, we have discussed the stability factor of the stacked ZIS nanosheet film, concluding that interface between nanosheets affects the stability. However, we did not find the specific material or structure at the interface caused for an instability factor. For the physiological trial, we only showed the possibility to find sleep apnea syndrome from respiration monitoring. For the practical use, clinical trials compared to the conventional methods will be required.takei@pe.osakafu-u.ac.jp or takei@omu.ac.jp).Further information and requests for resources and reagents should be directed and will be fulfilled by the lead contact, Professor Kuniharu Takei , InCl3·4H2O , and thioacetamide . The detailed process has been previously reported9. In this work, a ZIS nanosheet was formed using a solution of 10 mg mL−1.The ZIS solution was prepared from ZnCl2 laser ablation  on a PI film (thickness: 50 μm). The width of the electrodes was 0.4 mm, and the distance between interdigitating electrodes was varied from 0.1 mm to 1.1 mm in order to find the optimal distance. Subsequently, 75 μL of the ZIS nanosheet solution was drop-cast and annealed at 100°C for 20 min. To connect this humidity sensor to a measurement system, silver (Ag) electrodes were printed with a dispensing machine and annealed at 70°C for 30 min. Finally, the surface of the humidity sensor was covered with a humidity pass filter  to protect the ZIS nanosheet film from contact with water.LIG electrodes of interdigitating pattern were formed using COFundamental experiments with the ZIS humidity sensor were performed in a humidity and temperature controllable oven . All were recorded with a multi-channel data logger . As controls, commercial humidity and temperature sensors were also used to check humidity and temperature in the oven.The humidity sensor with a wireless circuit board was placed in a mask pocket and worn by volunteers  during sleep. Humidity and acceleration sensor outputs were automatically recorded by a smartphone linked via Bluetooth. Human subject experiments were performed in compliance with a protocol approved by the ethics committee at Osaka Prefecture University. Informed consent was obtained from all volunteers to record and use all data.Our study does not include statistical analysis or quantification.


The impact of phenolic acid on the antioxidant activity of SSPS was evaluated. Then, films were prepared by using phenolic acid-grafted SSPS. The physical, mechanical and biological performances of phenolic acid-grafted SSPS films were further investigated. The results indicated that an ester linkage was formed between the SSPS and phenolic acid. The grafting ratio of para-hydroxybenzoic acid, protocatechuic acid and gallic acid-grafted SSPS was 29.45, 31.76 and 30.74 mg/g, respectively. Phenolic acid endowed SSPS with improved antioxidant properties. Gallic acid (GA)-grafted SSPS possessed the best DPPH radical scavenging ability and reducing power, which may be related to the three phenolic hydroxyl groups in GA. Phenolic acid-grafted SSPS films showed increased moisture content and decreased water solubility compared to SSPS film. The phenolic acid-g-SSPS decreased the mechanical properties but enhanced the water vapor barrier property, and antioxidant and antibacterial properties of SSPS film. Meanwhile, the para-hydroxybenzoic acid-grafted SSPS film showed the lowest water vapor permeability (3.70 × 10−7 g mm/h cm2 Pa), and the GA-grafted SSPS film exhibited the best antioxidant and antibacterial activities.Three kinds of phenolic acid-grafted soluble soybean polysaccharide (SSPS) with similar grafting ratios were prepared, and their structure was characterized by FT-IR, UV-vis and  Petroleum-derived plastic materials are excessively consumed and are difficult to degrade, causing serious environmental pollution . Thus, eSoluble soybean polysaccharides having a structural resemblance to pectin is an acidic polysaccharide. SSPS possesses a galacturonan skeleton of rhamnogalacturonan and homogalacturonan  with a side chain of α-1,3- or α-1,5-arabinan and β-1,4-galactan . PreviouIn recent years, chitosan (CS) has been frequently grafted with various phenolic acids to enhance its water solubility and antioxidant and antibacterial activities ,11, whicIn this work, three kinds of phenolic acid-grafted SSPS (phenolic acid-g-SSPS) with similar grafting ratios were prepared. Structural characterization of phenolic acid-g-SSPS was carried out. Then, the phenolic acid-g-SSPS was further prepared into films. Finally, the physical and mechanical properties and biological performances of phenolic acid-g-SSPS films were studied.Soluble soybean polysaccharide , para-hydroxybenzoic acid (PPA), gallic acid (GA), protocatechuic acid (PA), 1,1-diphenyl-2-picrylhydrazyl (DPPH), gelatin, Folin-Ciocalteu-reagent, N-hydroxysuccinimide (NHS) and 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) were all bought from Yuanye Biological Co., Ltd., Shanghai, China.A series of phenolic acid-g-SSPSs were prepared, and the three kinds of phenolic acid-g-SSPSs with similar grafting ratios were selected for further evaluation. Details are as follows. v/v), followed by the addition of EDC . Then, NHS  was added to the reaction system and stirred for 2 h in the ice bath (4 °C). Afterward, the reactive system was mixed with the prepared SSPS solution, and the solution was stirred at room temperature for 48 h after stirring in the ice bath for 4 h. Then, the reaction solution was centrifuged  to remove residual GA. The final product was obtained by dialyzing against distilled water for a three-day period , lyophilized for 24 h and named GA-g-SSPS. Similarly, PPA-g-SSPS and PA-g-SSPS were synthesized according to the aforementioned method when the PPA  and PA  were substituted for GA. The yields of PPA-g-SSPS, PA-g-SSPS and GA-g-SSPS were 70.59%, 51.70% and 58.53%, respectively.Phenolic acid-g-SSPS was synthesized by the EDC/NHS reaction according to Shreiber’s method . SSPS  from 600–4000 cmf 4 cm−1 . UV-vis spectra of sample (1 mg/mL) were recorded (200–400 nm) using a U-3900 spectrophotometer (Hitachi (China) Ltd., Tokyo, Japan) .2O (0.55 mL). Then, 1 H NMR spectra were recorded at 25 °C at 400 MHz by using a Bruker Avance III HD spectrometer   reagent  and sodium phosphate buffer solution  were mixed; then the mixture was incubated in water  after evenly mixing in a vortex oscillator. Trichloroacetic acid (TCA)  was mixed with the reaction solution and centrifuged for 10 min. The supernatant of the mixture (2.5 mL), deionized water (2.5 mL) and FeCl3 solution  were mixed, and the mixture was allowed to stand for 0.5 h. At last, the absorbance was obtained at 700 nm.The reducing power was tested according to Oyaizu’s method . PhenoliPhenolic acid-g-SSPS films were obtained in accordance with Liu’s way with some changes . PhenoliA spiral micrometer  was used to test the thickness of films at any of 10 locations (as dispersed as possible) .W0), then the film was placed in an oven (105 °C) and dried to a constant weight (W1). Then, the film was soaked in 50 mL of deionized water and stirred slowly for 6 h. The remaining film was removed and placed into the oven again, and dried to a stable weight (W2) [The sample film was cut into 3 × 3 cm squares and weighed on an analytical balance (ght (W2) . The moiM indicates the increasing weight of the weighing bottle (g), d indicates the thickness (mm), t indicates the time period of weight measurement (h), A indicates the circular area covering the mouth of the weighing bottle (cm2) and The WVP of the film was measured by using the method of Gomez-Estaca . The filF was the stress at film break (N), d was the film thickness (mm), h was the width of the film (mm), 0L was the initial length of the film (mm).The mechanical performance of the film was evaluated on a DCP-KZ300 experimental machine . The rectangular film (150 × 15 mm) was placed on the measuring equipment with a spacing of 60 mm and a stretching speed of 50 mm/min . The tenThe released phenol content and DPPH radical scavenging ability were determined by Liu’s method . BrieflyStaphylococcus aureus and Escherichia coli was evaluated [6 CFU/mL) was evenly dispersed on an agar plate. The filter paper was removed from the bottle and put on the agar plate to incubate at 37 °C for 1 day. The diameter of the inhibition circle was then recorded with vernier calipers. Sterile water and potassium sorbate were considered the blank and positive control, respectively.The antibacterial activity of phenolic acid-g-SSPS films against valuated . First, p < 0.05.Three parallel tests were performed for each group, and the values were stated as mean ± standard deviation. The values were subjected to Duncan’s multiple range test, and analysis of variance (ANOVA) with SPSS 25.0 software , and a significant difference between the values was considered obtained if −1 were assigned to the O–H stretching vibrations, C–H and COO– stretching vibrations [−1 corresponded to the bending vibration of C–H. In addition, some typical peaks of polysaccharides were observed in the range of 1000–1200 cm−1, which are due to ring vibrations overlapped with C–OH stretching vibrations and C–O–C glycosidic bond vibrations [−1 is assigned to the β-glycosidic linkage in SSPS [−1 in phenolic acid-g-SSPS, which may be due to the ester groups (C=O stretching vibrations) formed between the hydroxyl group of SSPS and carboxyl groups of phenolic acid. A new absorption band at 1731 cm−1 was also noted in gallic acid-grafted chitosan, indicating the formation of an ester bond [As presented in brations . The peabrations . The abs in SSPS . The aboter bond .UV-vis spectra of SSPS and phenolic acid-g-SSPS are represented in 1 H NMR spectra can also provide structural support for phenolic acid-g-SSPS in The The grafting ratio of phenolic acid-g-SSPS was evaluated by the Folin-Ciocalteu method. The grafting ratio of PPA-g-SSPS, PA-g-SSPS and GA-g-SSPS was determined to be 29.45, 31.76 and 30.74 mg/g, respectively. Studies have shown that the biological activities of phenolic acid-grafted chitosan and the physical, mechanical and biological activities of phenolic acid-grafted chitosan films were related to the grafting ratio and the structure of phenolic acid . In our DPPH is extensively used to assess the antioxidant ability of samples because it is a stable free radical . As showThe reducing power of phenolic acid-g-SSPS is presented in As seen in The water resistance of a film can be assessed by two indicators, water solubility and moisture content. The moisture content of phenolic acid-g-SSPS films was higher than that of SSPS film . It migh−7 to 3.70 × 10−7 g mm/h cm2 Pa. The WVP of PA/GA-grafted CS films was lower than that of CS films [A key role of food packaging film is to prevent or reduce as much as possible, the transfer of moisture from the environment to the food . Thus, lCS films ,15. CovaCS films . In addiEnough mechanical strength can guarantee the integrity of the film and protect the film against small flaws, so TS and EB are two key indicators of the film’s mechanical properties . TS and Staphylococcus aureus and Escherichia coli than SSPS film. The order of antibacterial ability is as follows: GA-g-SSPS film > PA-g-SSPS film > PPA-g-SSPS film. The result exhibits that the antibacterial ability of phenolic acid-g-SSPS films is positively correlated with the released phenol content. The GA-g-SSPS film that has the highest released phenol content possesses the best antibacterial ability. In addition, due to differences in the structure of the bacterial outer membrane, all films exhibit stronger inhibitory ability against Staphylococcus aureus than that of Escherichia coli [As listed in hia coli . Comparehia coli .1 H NMR spectroscopy revealed that the reaction had successfully occurred. Phenolic acid endowed the SSPS with antioxidant properties. GA-g-SSPS possessed the best DPPH radical scavenging ability and reducing power, which could be linked to the three phenolic hydroxyl groups in GA. Meanwhile, phenolic acid-g-SSPS can decrease the water solubility and WVP and enhance the biological activities of the SSPS film. Since GA-g-SSPS film showed the best antioxidant and antibacterial performances, it can be expected to be a progressive material for food-active packaging. In subsequent studies, the influence of GA-g-SSPS film on the freshness of foods such as vegetables, fruits and meats should be assessed. The results in this study have important insights into the performance of active food packaging materials and their possible applications for the future.In this work, PPA, PA and GA were grafted onto SSPS by the EDC/NHS coupling method through ester bonding, respectively. FT-IR, UV-vis and