Mariana P Figueiredo

Géraldine Layrac

Anne Hébraud

Lionel Limousy

Jocelyne Brendle

Guy Schlatter
email: schlatter@unistra.fr

Vera R L Constantino

Design of 3D multi-layered electrospun membranes embedding iron-based layered double hydroxide for drug storage and control of sustained release

Keywords: electrospinning, electrospraying, layered double hydroxides, drug release modulation, functional membranes

ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

1

ABSTRACT

Nowadays polymer dressings are expected to possess multiply functions. Besides acting as physical barriers, dressings may provide for the injured tissue species able to turn wound healing process faster and painless. In this way, dressings can be designed aiming to enable the release of drugs. The possibility to modulate drug release kinetics is a desired characteristic to be achieved in order to turn drug delivery systems adequate to specific treatments. However, hydrophilic drugs and hydrophobic polymers incompatibility hinders such modulation and a long-term release cannot be achieved efficiently. Here we present the design of poly(lactic acid) (PLA) membranes containing iron-based Layered Double Hydroxide (LDH) particles able to storage a hydrophilic anionic drug (derived from the non-steroidal anti-inflammatory naproxen). LDH particles are excellent candidates to compose multifunctional composites. They may present diverse biocompatible compositions, possess an elevated encapsulation capacity and tends to promote drugs sustained release by its own, besides assisting tissues regeneration process. Nanofibrous membranes were prepared by the combination of electrospun PLA and electrosprayed LDH as alternated layers (approach A) and also by both technics performed at the same time (approach B). In approach A, by varying the thickness of the PLA fibrous layers, it was possible to easily modulate the drug release rate. Half of drug content was released after 1, 4 and 17 days for the membranes containing the thinnest, the intermediate and the thicker PLA layers, respectively, and after 56 hours for the membrane prepared by the approach B. Naproxen release was kept for 18 days for the thinnest membrane, 59 days for the membrane prepared by the approach B and 66 days for the thicker membranes. We believe that this work can inspire the development of new functional membranes with tunable drug release profile thanks to the versatile electrospinning and electrospraying techniques.

INTRODUCTION

The design of composite membranes allowing the precise control of the release kinetics of drugs emerged this last decade for advanced applications in wound dressing, tissue engineering and nanomedicine. [START_REF] Merino | Nanocomposite Hydrogels: 3D Polymer-Nanoparticle Synergies for On-Demand Drug Delivery[END_REF][START_REF] Mertz | Nanocomposite Polymer Scaffolds Responding under External Stimuli for Drug Delivery and Tissue Engineering Applications[END_REF] Especially, in the case of hydrophilic drugs, the controlled delivery over time is an important issue which has to be taken into account in order to deliver the optimal amount of therapeutics to a targeted diseased area while avoiding burst release and undesirable side effects due to overdose. Furthermore, when hydrophilic drugs are embedded in a hydrophobic insoluble polymer membrane, long-term release is more difficult to control due to drug-polymer incompatibility. [START_REF] Chou | Current strategies for sustaining drug release from electrospun nanofibers[END_REF] In this case, partitioning of the drug and the polymer matrix might occur, leading to either burst release or encapsulation of the drug within the matrix.

Nanofibrous membranes embedding drugs elaborated from the process of electrospinning have attracted researchers for such applications. [START_REF] Cheng | Biomedical application and controlled drug release of electrospun fibrous materials[END_REF][START_REF] Sun | Electrospun fibers and their application in drug controlled release, biological dressings, tissue repair, and enzyme immobilization[END_REF] Indeed, electrospun nanofibrous membranes mimic the extracellular matrix components, enabling cells growth and differentiation. Furthermore, reports have shown that cellfibrous interactions lead to a normal cells phenotypes and gene expression; and fibers possess high surface area/volume ratio and high porosity, which creates nutrient gradients. [START_REF] Bhardwaj | Electrospinning: A fascinating fiber fabrication technique[END_REF] Among the drug vectors which could be embedded in polymeric membranes, Layered Double Hydroxides (LDHs) have been identified as excellent candidates. [START_REF]Poly(L-lactic acid) nanocomposites with layered double hydroxides functionalized with ibuprofen -Dagnon[END_REF][START_REF] Mao | Poly(ε-caprolactone) filled with polydopaminecoated high aspect ratio layered double hydroxide: Simultaneous enhancement of mechanical and barrier properties[END_REF] LDHs are structured bidimensional materials with general formula [M 2+ (1-x)M 3+ x(OH)2](A n-)x/n•zH2O, formed by the stacking of positively charged layers composed by di (M 2+ ) and trivalent (M 3+ ) metal cations. [START_REF] Cavani | Hydrotalcite-type anionic clays: Preparation, properties and applications[END_REF] The M 2+ and M 3+ ions are distributed in the centre of octahedrons which are formed by six coordinated hydroxide anions, as shown in Figure 1. Structure neutrality is guaranteed by the presence of hydrated anions (A n-) between the layers. These materials are commonly abbreviated by M 2+ RM 3+ -A n-, where R is the molar ratio between M 2+ and M 3+ . LDH matrix by its own is commercialized as the antacid TALCID™, composed of magnesium and aluminium cations and intercalated with carbonate anions. [START_REF]Antacid talcid activates in gastric mucosa genes encoding for EGF and its receptor. The molecular basis for its ulcer healing action[END_REF] Thanks to the biocompatibility of LDHs compositions (verified in vitro and in vivo [START_REF] Choi | Biocompatible ceramic nanocarrier for drug delivery with high efficiency[END_REF][START_REF] Cunha | Delivery system for mefenamic acid based on the nanocarrier layered double hydroxide: Physicochemical characterization and evaluation of anti-inflammatory and antinociceptive potential[END_REF][START_REF] Kuthati | Layered double hydroxide nanoparticles for biomedical applications: Current status and recent prospects[END_REF][START_REF] Yan | Systemic toxicity induced by aggregated layered double hydroxide nanoparticles[END_REF][START_REF] Kuo | Layered double hydroxide nanoparticles to enhance organ-specific targeting and the anti-proliferative effect of cisplatin[END_REF][START_REF] Cunha | Accessing the biocompatibility of layered double hydroxide by intramuscular implantation: histological and microcirculation evaluation[END_REF][START_REF] Figueiredo | Iron-based layered double hydroxide implants: potential drug delivery carriers with tissue biointegration promotion and blood microcirculation preservation[END_REF]), their effective interactions with biological membranes, high encapsulation efficiency, and capacity to prolong drug release, these materials are promising to be applied as vehicles for drug storage and delivery. [START_REF] Gu | Hierarchical layered double hydroxide nanocomposites: structure, synthesis and applications[END_REF][START_REF] Choi | Layered double hydroxide nanoparticles as target-specific delivery carriers: uptake mechanism and toxicity[END_REF][START_REF] Oh | Cellular Uptake Mechanism of an Inorganic Nanovehicle and Its Drug Conjugates: Enhanced Efficacy Due To Clathrin-Mediated Endocytosis[END_REF] In this way, many species such as aminoacids, [START_REF] Nakayama | Intercalation of amino acids and peptides into Mg-Al layered double hydroxide by reconstruction method[END_REF] anti-inflammatories [START_REF] Rives | Layered double hydroxides as drug carriers and for controlled release of non-steroidal antiinflammatory drugs (NSAIDs): A review[END_REF] and cancer drugs [START_REF] Li | Co-delivery of siRNAs and anti-cancer drugs using layered double hydroxide nanoparticles[END_REF] have been intercalated between the LDHs layers. Recently our group showed that LDHs may interfere positively in tissue regeneration, being able to conduct the formation of different types of collagen according with the layer composition. [START_REF] Cunha | Accessing the biocompatibility of layered double hydroxide by intramuscular implantation: histological and microcirculation evaluation[END_REF][START_REF] Figueiredo | Iron-based layered double hydroxide implants: potential drug delivery carriers with tissue biointegration promotion and blood microcirculation preservation[END_REF] Furthermore, the combination of the LDH matrix with polymers has been used to prepare mimetic bone substituents, conferring mechanical strength and remodeling facilities by stress shielding. [START_REF] Kapusetti | Bone cement/layered double hydroxide nanocomposites as potential biomaterials for joint implant: Bone Cement and LDH Nanocomposite[END_REF][START_REF] Fayyazbakhsh | Novel layered double hydroxides-hydroxyapatite/gelatin bone tissue engineering scaffolds: Fabrication, characterization, and in vivo study[END_REF] The release of species such as healing cells activator from LDHs layers has improved even more bone regeneration process. [START_REF] Kang | Nanolayered hybrid mediates synergistic co-delivery of ligand and ligation activator for inducing stem cell differentiation and tissue healing[END_REF] Therefore, electrospun membranes embedding LDHs may form interesting implantable functional composites with drug release properties, acting as supports for damaged tissues due to surgery interferences and improving tissues regeneration. To the best of our knowledge, the only approach proposed in the literature consists in dispersing LDHs in the polymer solution which is further electrospun leading consequently to a membrane in which LDH nanoparticles are inside the matrix of fibers. [START_REF] Valarezo | Fabrication and sustained release properties of poly(ε-caprolactone) electrospun fibers loaded with layered double hydroxide nanoparticles intercalated with amoxicillin[END_REF] Besides, in this case, LDHs compositions presented only Al 3+ as trivalent cations, an exogenous metal. Valarezo et al. [START_REF] Valarezo | Fabrication and sustained release properties of poly(ε-caprolactone) electrospun fibers loaded with layered double hydroxide nanoparticles intercalated with amoxicillin[END_REF] reported the release of amoxicillin preencapsulated into the LDH layers of composition Zn2Al, itself encapsulated into poly(ɛpolycaprolactone) (PCL) fibers. While homogeneous fibers were obtained for LDH content of 3 % wt, the formation of beads was observed for a higher LDH content of 5 and 7 % wt, showing the limitation of the possible amount to be loaded in the polymer composites. The slowest drug release rate was observed for the membrane presenting 3 % wt of LDH amoxicillin, extending for more than 20 days. Other works reported the intercalation of non-steroidal anti-inflammatory drugs into LDHs of composition Mg2Al or Mg3Al. Yang et al. [START_REF] Yang | Synthesis and characterization of flurbiprofen axetil-loaded electrospun MgAl-LDHs/poly(lactic-co-glycolic acid) composite nanofibers[END_REF] applied flurbiprofen acetyl as model drug, poly(lactic-coglycolic acid) (PLGA) as polymeric matrix and 5 % of LDH-drug concentration relative to the polymer. Heterogeneous fibers diameter due to the presence of beads were observed for the LDH-PLGA nanofibers. After 5 days, less than 40 % of the drug was released. The work of Tammaro et al. [START_REF] Tammaro | Encapsulation of Diclofenac Molecules into Poly(-Caprolactone) Electrospun Fibers for Delivery Protection[END_REF] showed that PCL fibers containing Mg3Al

LDHs intercalated with diclofenac anions also exhibited a slow release of the drug (50 % after 240 days). Gao et al. [START_REF] Gao | Electrospun organic-inorganic nanohybrids as sustained release drug delivery systems[END_REF] reported the intercalation of ibuprofen or ketoprofen into the LDH of composition Mg2Al and composites formed with PCL polymer. It was verified that fibers diameter tends to increase when the LDH amount in the composite increases. After 5 days, 44 and 48 % of ibuprofen were released from composite membranes containing 5 and 10 % wt of LDH, respectively. However, for the same period of time and same LDH content, only 25 and 20 % of ketoprofen were released.

Miao et al. [START_REF] Miao | Electrospun fibers of layered double hydroxide/biopolymer nanocomposites as effective drug delivery systems[END_REF] also reported the pre-encapsulation of anions derived from ibuprofen into the Mg2Al LDH for preparation of composites based on poly(lactic acid) (PLA) and PCL electrospun fibers containing 5 % wt of LDH. Despite the hydrophobicity of both polymers, ibuprofen release from PCL was faster than from PLA fibers: approximately 40 % and less than 10 % after 2 h, respectively. This behavior was attributed to the poor chain flexibility of PLA. Although a slow release is desired most of the time in medicine's formulation, it is interesting to find mechanisms to modulate the drug release rate from the carrier, in order to adjust the formulation according with the treatment of different diseases, with different bioactive species and administration routes. Examples of a desired faster drug release include the administration of extra dose insulin during meals [START_REF] Raskin | Use of insulin aspart, a fast-acting insulin analog, as the mealtime insulin in the management of patients with type 1 diabetes[END_REF][START_REF] Morello | Pharmacokinetics and pharmacodynamics of insulin analogs in special populations with type 2 diabetes mellitus[END_REF] and anti-inflammatories to immediate relief of low or moderate pain. [START_REF] Christensen | Analgesic Efficacy of a New Immediate-Release/Extended-Release Formulation of Ibuprofen: Results From Single-and Multiple-Dose Postsurgical Dental Pain Studies[END_REF] On the other hand, a sustained drug release is intended when longterm drug circulation and treatments are necessary, such as in the administration of basal doses insulin unrelated to meals [START_REF] Morello | Pharmacokinetics and pharmacodynamics of insulin analogs in special populations with type 2 diabetes mellitus[END_REF][START_REF] Eliaschewitz | Concepts and clinical use of ultra-long basal insulin[END_REF] and postoperative antibiotics, [START_REF] Gao | Recent advances in materials for extendedrelease antibiotic delivery system[END_REF] anaesthetics [START_REF] Weiniger | Review of prolonged local anesthetic action[END_REF] and anti-inflammatories, [START_REF] Christensen | Analgesic Efficacy of a New Immediate-Release/Extended-Release Formulation of Ibuprofen: Results From Single-and Multiple-Dose Postsurgical Dental Pain Studies[END_REF] to avoid infections and severe chronic pains. In this way, the same work presented a strategy in order to increase and modulate the hydrophilicity of the PLA fibers based on the addition of poly(oxyethylene-boxypropylene-b-oxyethylene) (Pluronic) into the processed formulation, leading to a faster drug release. The studies above mentioned showed that the LDH concentration in the polymer solution is limiting, since its increase tends to the formation of beads and, consequently, non-homogeneous fibers. Furthermore, hydrophobicity of most biocompatible polymers such as PLA, PLGA and PCL, which can be easily processed by electrospinning, hinders drug release in aqueous media. [START_REF] Boda | Electrospraying an enabling technology for pharmaceutical and biomedical applications: A review[END_REF] Therefore, the development of strategies in order to overcome the polymers hydrophobicity and modulate drug release rates is desirable to expand the possible administration routes of electrospun membranes.

The present work is inspired by our previous results for which composite scaffolds elaborated from the combination of electrospinning and electrospraying processes show interesting properties due to the presence of both nanofibers and microparticles components. For example, when electrosprayed particles are simultaneously or alternately deposited with electrospun fibers, scaffolds with various patterns can be obtained. [START_REF] Nedjari | Electrostatic template-assisted deposition of microparticles on electrospun nanofibers: towards microstructured functional biochips for screening applications[END_REF][START_REF] Wittmer | Well-organized 3D nanofibrous composite constructs using cooperative effects between electrospinning and electrospraying[END_REF] Hybrid scaffolds with PCL fibers and hydroxyapatite particles can also be envisaged for bone regeneration application. [START_REF] Garcia | Poly(ε-caprolactone)/Hydroxyapatite 3D Honeycomb Scaffolds for a Cellular Microenvironment Adapted to Maxillofacial Bone Reconstruction[END_REF] Furthermore, it was also demonstrated that scaffolds entrapping hydrophilic particles in a hydrophobic fibrous mesh allow the modulation of the hydrophilicity/hydrophobicity balance. Such property was exploited to fabricate fibrous membranes allowing the controlled release of a drug. [START_REF] Lavielle | 3D Composite Assemblies of Microparticles and Nanofibers for Tailored Wettability and Controlled Drug Delivery[END_REF] Here, two kinds of membranes having different structures and made of PLA electrospun nanofibers and drug-loaded-LDH electrosprayed microparticles were elaborated in order to modulate the kinetic of drug release while avoiding the use of any hydrophilic adjuvant. Multilayer membranes with LDH particles in sandwich between PLA fibrous layers of various thicknesses were studied and compared with membranes in which the LDH particles were entrapped in the fibrous PLA mesh. PLA was chosen as polymer due to its biocompatibility, good mechanical properties and performance for fibers formation. Aluminum accumulation has been verified in different tissues and has being associated to neurodegenerative diseases. [START_REF] Mold | Insight into the cellular fate and toxicity of aluminium adjuvants used in clinically approved human vaccinations[END_REF][START_REF] Krewski | Human health risk assessment for aluminium, aluminium oxide, and aluminium hydroxide[END_REF][START_REF] Kumar | Aluminium neurotoxicity: Neurobehavioural and oxidative aspects[END_REF] Therefore, in order to turn the drug delivery devices more promising for biological applications, half of the Al 3+ content was substituted by Fe 3+ cations (endogenous) in the LDHs composition.

Naproxenate was applied as a model hydrophilic drug and was pre-encapsulated into LDHs layers. In order to compare the effect of the organic anions regarding the interaction between LDHs particles and fibers, similar membranes were prepared with LDH precursors intercalated with Cl -anions.

Experimental section

Poly(D,L-lactide) (PLA) (MW 180 kDa, L = 95%, NatureWorks 7000D). N,Ndimethylformamide (≥ 99.8%), dichloromethane (≥ 99.8%), magnesium chloride hexahydrate (MgCl2•6H2O) (99 %), aluminum chloride hexahydrate (AlCl3•6H2O) (99 %), iron(III) chloride hexahydrate (FeCl3•6H2O) (98 %), sodium chloride (NaCl) (99.5 %), sodium hydroxide (NaOH) and sodium naproxenate (NaNAP) (NaC14H13O3) with purity ≥ 98 %, were purchased from Sigma-Aldrich. Ethanol absolute was purchased from Carlo Erba. All products were used as received.

LDH preparation

LDHs, intercalated with NAP and Cl -anions, were prepared following the procedure previously reported, [START_REF] Figueiredo | Iron-based layered double hydroxide implants: potential drug delivery carriers with tissue biointegration promotion and blood microcirculation preservation[END_REF] with some changes and adapted as follows.

Synthesis of the Mg4FeAl-Cl sample

The LDH intercalated with Cl -anions, abbreviated by Mg4FeAl-Cl LDH, was prepared under N2 atmosphere by the co-precipitation method at constant pH equal to 10.5, by adding under mechanical stirring at 900 rpm and 50°C concomitantly a mixture of a 0.05 mol•L -1 solution of MgCl2•6H2O, FeCl3•6H2O and AlCl3•6H2O (to obtain a Fe 3+ /Al 3+ molar ratio equal to 1) and a 0.2 mol L -1 NaOH solution into 200 mL deionized water. After addition, the mixture was aged under stirring for 1 h at the same synthesis temperature. The obtained solid was recovered by filtration, washed with 2 L of deionized water and lyophilized for 24 h at 200 mbar and -50 ° C (in a Thermo Savant ModulyoD equipment). The Mg4FeAl-Cl LDHs used to test the stability of the particles suspension were prepared by a similar procedure, with exception of the mechanical stirring rate, kept at 400 rpm.

Intercalation of NAP into the Mg4FeAl LDH by ion exchange method

Starting from the remaining slurry of the Mg4FeAl-Cl LDH synthesis, not submitted to washing procedure, the excess of Cl -ions was removed by centrifugation with 4 cycles of 3 min each at 10000 rpm. For the preparation of approximately 0.5 g of the LDH intercalated with NAP, abbreviated by Mg4FeAl-NAP, the precursor was resuspended in 20 mL of an aqueous solution of NaNAP in a molar ratio NAP / (Fe 3+ + Al 3+ ) equal to 3. The suspension was kept under N2 atmosphere and vigorously stirring with stir bar at 200 rpm for 24 h at 80 °C. The solid was washed with deionized water by filtration under reduced pressure and then dried by freeze drying in the same conditions applied for the Mg4FeAl-Cl LDH.

Preparation of suspensions and solution

Preparation of LDH suspension to be electrosprayed

The LDH suspension used to prepare the LDH-PLA membranes has been optimized for efficient electrospraying of the smallest aggregates (see Supporting Information and figures S1-S3). LDHs-ethanol suspensions (1% wt) were previously submitted to ultrasonication (Branson Digital Sonifier equipment) at 40 % amplitude for 30 min (2 seconds on / 2 seconds off) just before the electrospraying procedure.

Preparation of PLA solution to be electrospun

A 10 % wt PLA solution was prepared by dissolving the beads in a mixture of dichloromethane and dimethylformamide in weight ratio equal to 3:2 overnight under slow stirring with a magnetic stirring bar.

2.3.Preparation of the composite membranes

Two approaches are proposed for the preparation of composite membranes. The approach A consists in the fabrication of multi-layered membranes by alternating electrospinning PLA fibers and electrospraying LDHs particles. In approach B, LDHs

Electrospun PLA and electrosprayed LDH: building up a delivery membrane (Approach A)

Electrospinning and electrospraying experiments were conducted with a homemade setup previously described [START_REF] Lavielle | 3D Composite Assemblies of Microparticles and Nanofibers for Tailored Wettability and Controlled Drug Delivery[END_REF] and allowing the deposition on a drum collector rotating at 125 rpm. All solutions or suspensions were electrospun or electrosprayed at a temperature T = 22 +/-1°C and a relative humidity RH = 40 +/-5%. Working distance between needle and collector was equal to 16 cm for all samples. For all electrospinning and electrospraying steps, an alternating translation movement (see Figure 2, amplitude of the movement = 7 cm, average velocity of the movement = 2 cm/s) of the needle was imposed in order to enlarge the deposition leading to the fabrication of membranes having a width of 11 cm.

In approach A, the membranes contain 7 layers made from the deposition of 4

PLA electrospun fibers taking in sandwich 3 LDHs electrosprayed particles (Figure 2a). Three different PLA layer thicknesses were prepared by changing the production time of the electrospun layers. Applied voltages, flow rates and production times are given in Table 1. Membranes containing electrosprayed Mg4FeAl-NAP particles are called NAP-Athin, NAP-Amed, NAP-Athick and the membranes with electrosprayed Mg4FeAl-Cl are called Cl-Athin, Cl-Amed, Cl-Athick. In order to compare the samples, the total electrospraying time was the same for each membrane and equal to 78 minutes. In approach B (Figure 2b, Table 1), the membranes were prepared by simultaneous deposition of electrospun PLA and electrosprayed LDH aiming to contain the same total amount of LDH as applied in approaches A (total electrospraying time equal to 78 min). For both needles an alternating translation movement (amplitude of the movement = 7 cm, average velocity of the movement = 2 cm/s) was imposed in order to enlarge the deposition and allowing thus the fabrication of membranes having a width of 11 cm. Samples were abbreviated as Cl-B and NAP-B, indicating the application of B approach and LDH particles containing Cl -and NAP anions, respectively.

Pristine PLA membrane

As a control, pristine PLA membrane was electrospun for 2 hours. Therefore, it contained the same polymer amount as in the membranes prepared according with approach Athick.

NAP release tests

Naproxen release experiments were conducted in triplicate from 3 cm x 10 cm membranes. The membranes were accommodated in closed glass flasks containing 70 mL of saline aqueous solution (0.9 % wt NaCl); the pH value was equal to 5.55 ± 0.01.

The release media was magnetically stirred at 130 rpm and the assays were performed Inductively coupled plasma atomic emission spectroscopy (ICP-AES) analyses of metals were performed in duplicate on Spectro Analytical Instrument.

Images by scanning electron microscopy (SEM) of the membranes were obtained in a VEGA3SEM equipment using samples deposited on a copper tape and coated with gold using a Q150RS Quorum equipment. The LDHs micrographs were obtained in a XL30 FEG Philips equipment depositing the powders in a copper tape and covering samples with carbon.

Mechanical tensile measurements were performed with a TA Instruments Discovery HR-3 hybrid rheometer at room temperature and applying a velocity of 50 μm s -1 . The normal force sensor of the rheometer was used to record the tensile force. Samples were analysed in quintuplicate and were prepared cutting 3 x 1.5 cm rectangles. The mechanical tests were carried out at room temperature (T = 21 ± 2°C) and humidity (RH = 50 ± 20%) without environmental control.

Thermogravimetric analysis (TGA) data were recorded on TA instrument, Q5000 model, using platinum crucible and a heating rate of 10 °C min -1 , from room temperature to 1000 °C, under synthetic air flow of 25 mL min -1 .

Membranes thicknesses were determined using a Schut digital micrometre. Six measurements in different points were performed in samples with 3 cm x 10 cm area.

Contact angle measurements were performed with static deionized water drop on the membranes using a DSA 25 tensiometer.

Absorbance in the ultraviolet-visible spectral region was measured on a Shimadzu UV-2600 spectrometer.

Results and discussion

Mg4FeAl-Cl and Mg4FeAl-NAP LDHs characterization

LDH samples intercalated with Cl -anions or NAP were first fully characterized.

Figure S4 (Supplementary Information)

presents the XRD patterns of Mg4FeAl-Cl (precursor) and Mg4FeAl-NAP samples. Precursor LDH diffractogram displays the characteristics peaks (narrow and well resolved) related to the LDH phase. [START_REF] Cavani | Hydrotalcite-type anionic clays: Preparation, properties and applications[END_REF] On both diffractograms peaks are narrow revealing a structural organization. After NAP intercalation, the shift of the (00l) peaks to small angles region is observed, which indicates that the interplanar distances are higher, endorsing the organic anion intercalation. Moreover many (00l) harmonic peaks are visualized for the hybrid LDH.

The presence of the residual Mg4FelAl-Cl phase in the Mg4FeAl-NAP sample is evidenced by the permanence of the (113) reflection, normally displaced to small angles region and not visualized for single phases LDHs intercalated with bulky anions, once it is dependent of the c crystallographic parameter. Mixture of phases is commonly observed for NAP intercalation by ion-exchange reaction in MgAl LDHs and also by coprecipitation method. [START_REF] Figueiredo | Iron-based layered double hydroxide implants: potential drug delivery carriers with tissue biointegration promotion and blood microcirculation preservation[END_REF][START_REF] Delarco | Release studies of different NSAIDs encapsulated in Mg,Al,Fe-hydrotalcites[END_REF][START_REF] Del Arco | Synthesis and characterization of layered double hydroxides (LDH) intercalated with non-steroidal anti-inflammatory drugs (NSAID)[END_REF][START_REF] Wei | Studies on the intercalation of naproxen into layered double hydroxide and its thermal decomposition by in situ FT-IR and in situ HT-XRD[END_REF] FT-IR spectra of the LDH materials are present in Figure S5. The band at 1361 cm -1 is assigned to the antisymmetric stretching of the carbonate anions (contaminant), while the broad bands below 700 cm -1 are assigned to the metal-OH translation. [START_REF] Cavani | Hydrotalcite-type anionic clays: Preparation, properties and applications[END_REF] For the Mg4FeAl-NAP sample, the band at 1230 cm -1 is attributed to the in plane bending of the C-H bounds in the naphthalene ring. The bands at 1392 and 1545 cm -1 can be assigned to the stretching modes of the COO -group. [START_REF] Delarco | Release studies of different NSAIDs encapsulated in Mg,Al,Fe-hydrotalcites[END_REF] FT-Raman spectra of the LDHs are presented in Figure S6. For the precursor phase, the less intense band at 1062 cm -1 is assigned to the symmetric stretching of adsorbed carbonate anions. [START_REF] De Faria | Raman microspectroscopy of phthalocyanine intercalates: tetrasulphonated cobalt and nickel phthalocyanines in layered double hydroxide[END_REF] The band at 540 cm -1 is assigned to metal-OH translation modes. For the Mg4FeAl-NAP sample, the most intense bands at 1632 and 1416 cm -1 are attributed to C-C ring vibrations while the band at 1386 cm -1 is assigned to CH3 deformation. [START_REF] Figueiredo | Iron-based layered double hydroxide implants: potential drug delivery carriers with tissue biointegration promotion and blood microcirculation preservation[END_REF] TGA curves of the Mg4FeAl-Cl and Mg4FeAl-NAP materials are shown in Indeed, in the approach A, a large number of particles is electrosprayed at the same time, favoring the formation of aggregates on the mat, as was seen when electrospraying on a flat collector (Figure S1 in SI) whereas in approach B, only few particles are electrosprayed before new fibers land on the mat and hide them, hindering the formation of aggregates. FT-IR spectra of the membranes containing Cl-LDH (Figure 4a) and NAP-LDH (Figure 4b) particles have been obtained. All bands observed for the pristine PLA (Figure S10) are present in the spectra related to the composites, whose assignment was made as follows. [START_REF] Cai | Spectral Characterization of Four Kinds of Biodegradable Plastics: Poly (Lactic Acid), Poly (Butylenes Adipate-Co-Terephthalate), Poly (Hydroxybutyrate-Co-Hydroxyvalerate) and Poly (Butylenes Succinate) with FTIR and Raman Spectroscopy[END_REF] The bands at 2994 and 2945 cm -1 are attributed to the antisymmetric and symmetric stretching of the -CH3 group, respectively. The band at 1751 cm -1 is assigned to the stretching of the -C=O group. Antisymmetric -CH3 bending frequency is observed at 1452 cm -1 , while the corresponding symmetrical mode appears at 1361 cm -1 . The bands at 1261, 1184, 1129, 1084 and 1043 cm -1 are related to vibrational modes of the C-O-C group. Finally, the band at 867 cm -1 is related to the stretching of the O-CH-CH3 portion and the band at 753 cm -1 to the wagging of the α-CH3 group. For the membranes whose SEM images show higher levels of exposed particles, the bands at 3430 cm -1 , in the spectra for Cl-AthickL and Cl-B, and at 3433 cm - When NAP-LDH particles are present in the composites, the decrease in hydrophobicity compared to the pristine electrospun PLA membrane is less pronounced, showing the property of NAP anions in conferring higher hydrophobicity to the LDHs. Cl-AthickL --------121 ± 5 --------------------------------------

NAP-AthickL -------- 127 ± 5 -------- -------- -------- -------- --------
Table 3 presents mechanical properties (Young modulus, yield stress and strain and stress and strain at break) of the membranes obtained by tensile measurements.

Stress-strain curves can also be seen in Figure S12. Although LDH-fibers interactions might change the mechanical behaviour, it is shown that the amount of incorporated LDH is not enough to significantly affect the tensile properties.

Technological medical devices, such as multifunctional dressings, require a deep understanding of the materials structure in order to tune materials properties and explain their behaviour once applied. LDHs were previously fully characterized and a high drug load was verified. Physicochemical properties of the nanofibers membranes were investigated. The interaction degree among PLA nanofibers and LDHs particles responded to the applied approach and resulted in mechanical properties improvements.

The different membranes presented a particular superficial morphology, in which modifications in the thickness of the polymeric diffusional barrier are expected to affect the drug release profile, discussed below. In order to get more insight in the release mechanisms, the naproxen release profiles of the different membranes were fitted by the power law Ritger-Peppas model [START_REF] Ritger | A simple equation for description of solute release I. Fickian and non-fickian release from non-swellable devices in the form of slabs, spheres, cylinders or discs[END_REF] (equation 2) which is currently used to describe the drug release process from matrices in general and polymeric systems:

= Kt (2)
Where Mt is the drug amount released after a time t and M∞ is the drug amount at the equilibrium state. In order to better understand the dissimilarity among the dissolution profiles for the membranes prepared by the different approaches, the "difference factor" ( ) and the "similarity factor" ( ), described by equations 3 and 4, respectively, were calculated and are indicated by the Agência Nacional de Vigilância Sanitária (The Brazilian Health Regulatory Agency) [START_REF] Sparks | Kinetics and Mechanisms of Chemical Reactions at the Soil Mineral/Water Interface[END_REF] and by the FDA [START_REF] O'hara | A review of methods used to compare dissolution profile data[END_REF] to compare tested and reference formulations. Where, N is the number of dissolution time points and Rt and Tt are the dissolution values (percentages in our case) for the reference and the tested system at a time t, respectively. The factor values are shown in

Figure 1 .

 1 Figure 1. Schematic representation of the LDH structure.

Figure 2 .

 2 Figure 2. Preparation of the composite membranes in the case of a) approach A with variable and adjustable thickness of the PLA fibrous layers and b) approach B with simultaneous PLA electrospinning and LDH electrospraying.

in a climatized room at 22

 22 ºC. At specified time, 3 mL of saline solution were collected and replaced with an equivalent amount of dissolution medium. NAP concentration was determined by UV-visible absorption spectrophotometry at a maximum absorption (λmax) equal to 230 nm. After the first released NAP aliquot measured, accumulative NAP weight (m NAP i) released was corrected through the equation (1), where Ci is the NAP concentration (mL L -1 ) in such a time i, considering the removed aliquots(3 mL) for measurement in anterior times k and then replaced by the same volume of saline solution in order to maintain the flasks volume equal to 70 mL. diffraction (XRD) patterns of powdered LDHs were obtained on a Bruker D8 Advance A25 equipment with variable slits mode, using Cu anode (λ = 1.518 Å), scan range 1.5-70°(2θ) and scan step of 0.02° (2θ) / s. Fourier transform infrared (FT-IR) spectra of powdered LDH samples and of the composites were recorded in the 4000-400 cm -1 range on a Bruker spectrophotometer, model α, by attenuation total reflection (ATR) mode with acquisition step of 4 cm -1 and 512 scans.Fourier transform Raman (FT-Raman) spectra of the LDH samples were recorded in an FTRaman Bruker FRS-100/S spectrometer using 1064 nm exciting radiation (Nd:YAG laser Coherent Compass 1064-500 N), a Ge detector, laser power of 100 mW, acquisition step of 4 cm -1 , and 2048 scans.Chemical elemental analyses of carbon, hydrogen, and nitrogen were recorded on Perkin Elmer-CHN 2400.

FiguresFigure S9 .

 S9 Figures S7and S8, respectively. Three and four main mass loss events are observed, for the precursor and the hybrid LDH sample, respectively. For both compounds, the first and second events, occurring from room temperature up to 180 °C and 230 °C for the precursor LDH and for the hybrid material, respectively, are associated to the release of adsorbed and intercalated water molecules. For the Mg4FeAl-Cl sample, the third event, that occurs between 180 -600 °C, is assigned to the matrix dehydroxylation. The residual mass corresponds to 59.5 % wt. For the Mg4FeAl-NAP LDH, the third and fourth mass loss events correspond to the overlapping matrix dehydroxylation and NAP decomposition. The smaller residual mass (46.4 % wt) observed, in comparison with the precursor material, agrees with NAP presence. SEM micrographs of the Mg4FeAl-Cl and Mg4FeAl-NAP samples are present in Figure S9. For both samples, a sand rose morphology and many sets of 2D plates are visualized. The great similarity between the sample's morphology suggests that the chloride-exchange is a topotactic process, in which the final lattice is determined by the lattice of the pristine LDH. The elemental chemical composition of the Mg4FelAl-Cl and Mg4FeAl-NAP LDHs, as well as the amount of water obtained by TGA experiments, are gathered in

3. 2 .

 2 3D Composite membranes by PLA electrospinning and LDH electrospraying Membranes micrographs are presented in Figure 3. Regarding the multi-layered membranes obtained from approach A, some LDH particles are exposed on the surface of the membranes containing the thinnest PLA layers (Cl-Athin and NAP-Athin). Increasing the PLA thickness, fewer particles are visualized on Cl-Amed and NAP-Amed surfaces and no particles are observed for the Cl-Athick and NAP-Athick samples. The surface aspects of the Cl-Athick and NAP-Athick samples are similar to the one of the pristine PLA membrane, indicating that 30 min of PLA electrospinning is enough to completely cover the last layers containing LDHs particles. Less particles aggregation is observed for the membranes prepared from approach B by simultaneous PLA electrospinning and LDH electrospraying (Cl-B and NAP-B samples) compared to the particles distribution on surface of the thinnest membranes and to the surfaces aspect of the last deposited LDH layers (L), abbreviated by Cl-AthickL and NAP-AthickL samples.

Figure 3 .

 3 Figure 3. SEM micrographs of pristine PLA membrane and PLA membranes containing Mg4FeAl LDHs intercalated with Cl -and NAP anions, prepared by alternating 4 electrospun PLA layers with 3 electrosprayed LDH layers (Approach A) with different PLA thickness (Athin -electrospinning of each layer performed for 15 min, Amed-22 min and Athick -30 min) and prepared by both electrospinning and electrospraying at the same time (Approach B). Samples were analyzed applying BSE detector, 5 kV and work distance around 10 mm.

1 ,Figure 4 .

 14 Figure 4. FT-IR spectra of membranes containing a) Cl-LDHs particles and b) NAP-LDH particles.

3. 3 .Figure 5 .

 35 Figure 5b, shows a considerable release rate difference among the samples. Half of NAP amount was released after 24, 56, 96 and 408 h (17 days) of experiment for the NAP-Athin, NAP-B, NAP-Amed and NAP-Athick composites, respectively. The release rate can be correlated with the degree of LDH particles exposure to the saline solution media. Besides, in order to reach the LDH particles inside the structure, the aqueous media has to penetrate through the PLA layers, which are hydrophobic. From Athin to Athick, the thickness of the PLA layer increases, increasing the penetration time inside the membrane and slowing the drug release. Naproxen was released over 18 days for the

Figure 6

 6 presents the curves of the model as well as the obtained kinetic parameters (constant K and release exponent n) and linear correlation coefficients (R 2 ). Drug release from all devices was well fitted by the Power Law model, with R 2 values greater than 0.98. The low n values obtained for NAP-Athin, NAP-Amed and NAP-B membranes (respectively 0.28, 0.23 and 0.26), smaller than 0.45, suggests a Fickian type release with dominant diffusion process.[START_REF] Ritger | A simple equation for description of solute release I. Fickian and non-fickian release from non-swellable devices in the form of slabs, spheres, cylinders or discs[END_REF][START_REF] Haroosh | Synthesis, morphological structures, and material characterization of electrospun PLA:PCL/magnetic nanoparticle composites for drug delivery[END_REF][START_REF] Fahami | Development of cress seed mucilage/PVA nanofibers as a novel carrier for vitamin A delivery[END_REF] In the case of NAP-Athick membranes, n value is larger and equal to 0.59, characterizing an anomalous transport. It must be mentioned that such model gives only an overall behavior of the membrane rendering difficult an exact description of the drug release mechanisms and especially the lag-time observed for the NAP-Athick membranes. The release behavior of LDH materials (i.e. no associated to a polymer) is often related to a Fickian diffusion, which is based on ion exchange process.[START_REF] Chakraborty | A facile synthetic strategy for Mg-Al layered double hydroxide material as nanocarrier for methotrexate[END_REF][START_REF] Zhang | Methotrexate intercalated layered double hydroxides with different particle sizes: Structural study and Controlled release properties[END_REF] However, in our case, the multilayer structure of the membranes induces a complex sequential flow through the PLA nanofibrous layers and then, through the LDH electrosprayed layers. The latter being the domains where the drug is initially released by intra-LDH-particles and inter-LDHparticles diffusion. Considering the architecture of the NAP-Athick membranes, it is possible to propose that initially the release medium permeates the pores of the PLA hydrophobic mesh of the external layer. Then, after this first step, exchange of NAP by Cl -saturating the LDH layers allows the release of the drug through the porous layer of PLA towards the external medium. The drug release delay was not evidenced for all other membranes because some LDH particles were immediately exposed to the release medium as shown in Figure3for the membranes containing the thinnest PLA layers (i.e. NAP-Athin, NAP-Amed, NAP-B).

Figure 6 .

 6 Figure 6. Experimental points and Ritger-Peppas model (solid lines) of Naproxen release percentage as a function of time for the different composites in log-log scale.

Table 1 .

 1 Applied parameters for preparation of the membranes

			Voltage on	Voltage on		Production
					Flow rates
	Approach	Method	the needle	the collector		Time
					(mL.h -1 )
			(kV)	(kV)		(min)
	Athin, Amed,	Electrospraying	25	0	6	3 layers
	Athick	Layers 2, 4, 6				26 min/layer
		Electrospinning	16	-5	2.8	4 layers
	Athin
		Layers 1, 3, 5, 7				15 min/layer
		Electrospinning	16	-5	2.8	4 layers
	Amed
		Layers 1, 3, 5, 7				22 min/layer
		Electrospinning	16	-5	2.8	4 layers
	Athick
		Layers 1, 3, 5, 7				30 min/layer
		Electrospinning	16		2.8
	B
		Electrospraying	28	0	6	78 min

Table 2 .

 2 Molar ratio between the metals are close to the nominal values. Naproxen

anions were intercalated in appreciable amounts (36.17 g of NAP for 100 g of hybrid).

Table 2 .

 2 Elemental chemical composition of Mg4FeAl-Cl and Mg4FeAl-NAP LDHs.

					Mg /
	Sample	Mg/Al Mg/Fe Fe/Al		% H2O	wt % C	wt % NAP
					(Fe + Al)
	Mg4FeAl-Cl	4.32	4.08	1.06	2.10	13.0	-----	-----
	Mg4FeAl-NAP	4.36	4.18	1.04	2.14	14.4	26.51	36.17

Table 3 .

 3 Membranes average thickness, static water contact angle and mechanical properties obtained by tensile tests for PLA membrane and PLA_LDH composites.

		Membranes	Contact	Yield	Yield	Stress at	Strain at	Young
	Sample	Thickness	Angle	stress	strain	break	break	modulus
		(µm)	(º)	(MPa)	(%)	(MPa)	(%)	(MPa)
	PLA	148 ± 22	129 ± 5	1.9 ± 0.1	2.9 ± 0.1	3.1 ± 0.2	70 ± 1	98 ± 5
	Cl-Athin	61 ± 24	121± 4	1.5 ± 0.1	3.3 ± 0.3	2.1 ± 0.2	82 ± 4	74 ± 6
	Cl-Amed	102 ± 22	121 ± 5	1.5 ± 0.2	2.8 ± 0.6	2.5 ± 0.2	82 ± 6	82 ± 7
	Cl-Athick	116 ± 14	124 ± 6	2.1 ± 0.1	3.0 ± 0.3	3.9 ± 0.2	100 ± 3	109 ± 6
	Cl-B	86 ± 26	121 ± 5	1.5 ± 0.2	2.8 ± 0.9	2.8 ± 0.3	79 ± 6	76 ± 12

Table 4 .

 4 Dissolutionsprofiles are considered equivalents if is between 0 and 15 and if is between 50 and 100, as observed among the NAP-Amed and NAP-B membranes, while all other membranes show a different profile, compared two by two.

	= 100	∑ | ! " ! | # !$% ∑ ! # !$% &

Table 4 .

 4 "Difference" ( ) and "similarity" ( ) factors calculated for the composites compared two by two.The development of 3D multi-layered electrospun membranes containing drug loaded iron-based LDH particles was achieved with the aim of forming multifunctional dressings by different approaches. In approach A, a faster, an intermediate and a slower NAP release were observed for the thin, medium and thick PLA layer thickness, respectively. Therefore, drug release could be modulated by simply varying the preparation parameter. Compared to the membranes prepared by the approach A, approach B involved only one step preparation and the membrane presented an intermediate NAP release profile. The developed membranes fulfilled important characteristics expected for a technological drug delivery device, since the prepared drug delivery composites were able to modulate and keep NAP release for more than two months. Furthermore, membranes architecture can inspire the development of even more interesting and functional devices based on electrospinning and electrospraying techniques. Paulo (Research Internships Abroad, BEPE-FAPESP 2018/22726-7) and Conselho Nacional de Desenvolvimento Cientifı ćo e Tecnolo ́gico (CNPq, 305446/2017-7), and is part of the Research Academic Cooperation Agreement PRC CNRS-FAPESP (PRC-Projet de recherche Conjoint 1688 and SPRINT-Sa õ Paulo Researchers in International Collaboration 2016/50317-9). We are thankful to Céline Piras for the technical support. Laborato ́rio de Espectroscopia Molecular Hans Stammreich (LEM, Instituto de Química-USP) is also acknowledged for the Raman and FT-IR spectra recording.

	Rt	NAP-Athin	NAP-Amed	NAP-B	NAP-B	NAP-B
	Tt	NAP-Amed	NAP-Athick	NAP-Athin	NAP-Amed	NAP-Athick
		28.4	44.7	27.7	10.0	49.5
		33.2	32.1	39.9	60.4	27.7
		4. Conclusions

ACKNOWLEGEMENTS

This work was supported by Fundaça õ de Amparo a ̀ Pesquisa do Estado de Sa õ


Miltiadis Alepidis

Licinius Benea

Davide Bucci

Xavier Mescot

Maryline Bawedin

Irina Ionica

Experimental and simulation investigation of the out-of-equilibrium phenomena on the pseudo-MOSFET configuration under transient linear voltage ramps

Keywords: Pseudo-MOSFET configuration, transient measurements, simulation, out-of-equilibrium body potential, out-of-equilibrium phenomena

ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

The pseudo-MOSFET configuration measurements are performed by the placement of two metal probes with controlled pressure on top of bare SOI wafers [START_REF] Cristoloveanu | A review of the pseudo-MOS transistor in SOI wafers: operation, parameter extraction, and applications[END_REF]. The metal probes mimic the source and drain contacts. The buried oxide (BOX) underneath the silicon film can be biased through the substrate and consequently acts like a gate dielectric. The combination of those elements creates a structure similar to an upside-down MOSFET. The main utilization of this pseudo-transistor is the extraction of important parameters of the top silicon film, by exploiting different measurement methods: current-voltage characteristics for extraction of the mobility of electrons (e -), holes (h + ) and threshold/flatband voltages [START_REF] Cristoloveanu | A review of the pseudo-MOS transistor in SOI wafers: operation, parameter extraction, and applications[END_REF][START_REF] Fernandez | On the effective mobility extraction by point-contact techniques on silicon-on-insulator substrates[END_REF], capacitance [START_REF] Pirro | Interface trap density evaluation on bare silicon-on-insulator wafers using the quasi-static capacitance technique[END_REF] and noise measurements [START_REF] Hajj Diab | Low-frequency noise in SOI pseudo-MOSFET with pressure probes[END_REF][START_REF] Cristoloveanu | A review of electrical characterization techniques for ultrathin FDSOI materials and devices[END_REF] for estimation of interface state density. Carrier life times were also estimated by the pseudo-MOSFET configuration [START_REF] Cristoloveanu | Model for carrier lifetime extraction from pseudo-MOSFET transients[END_REF][START_REF] Kang | SOI bulk and surface generation properties measured with the pseudo-MOSFET[END_REF]. Recent advancements showed that out-of-equilibrium phenomena, such as the out-of-equilibrium body potential, manifest themselves in the pseudo-MOSFET configuration [START_REF] Benea | Out-of-equilibrium body potential measurements in pseudo-MOSFET for sensing applications[END_REF]. Similar to the fully depleted SOI MOSFETs, in which it was firstly reported [START_REF] Bawedin | Dynamic body potential variation in FD SOI MOSFETs operated in deep non-equilibrium regime: Model and applications[END_REF], the out-of-equilibrium body potential variation arises when the transition between depletion and inversion/accumulation occurs and the carriers are not provided by the source and drain probes instantaneously. In this case, during the variation of the gate voltage, the lack of carriers needed to reach equilibrium drives the body potential into an out-of-equilibrium state which will simply follow the VG variation. Obviously, one of the key elements to understand and exploit this phenomenon is related to the contacts that provide (or not) the free carriers for the channel formation. In the pseudo-MOSFET, this aspect is very complex to consider since, theoretically, metal probes on the low doped silicon film should behave as Schottky contacts. However, the metal probes used in the pseudo-MOSFET show ohmic behaviour and provide both types of carriers into the film [START_REF] Cristoloveanu | A review of the pseudo-MOS transistor in SOI wafers: operation, parameter extraction, and applications[END_REF][START_REF] Cristoloveanu | A review of electrical characterization techniques for ultrathin FDSOI materials and devices[END_REF][START_REF] Benea | Nanoindentation effects on the electrical caracterizaron in Ψ-MOSFET configuration[END_REF][START_REF] Ionica | Characterization of silicon-on-insulator films with pseudo-metal-oxide-semiconductor field-effect transistor: Correlation between contact pressure, crater morphology, and series resistance[END_REF][START_REF] Williams | Point contact pseudometal/oxide/semiconductor transistor in as-grown silicon on insulator wafers[END_REF][START_REF] Munteanu | Numerical simulation of the pseudo-MOSFET characterization technique[END_REF][START_REF] Van Den Daele | Adaptation of the pseudo-metal-oxide-semiconductor field effect transistor technique to ultrathin silicon-on-insulator wafers characterization: Improved set-up, measurement procedure, parameter extraction, and modeling[END_REF]. The origin of this behaviour is not fully-understood and can be explained by defects under the metal contacts [START_REF] Ionica | Characterization of silicon-on-insulator films with pseudo-metal-oxide-semiconductor field-effect transistor: Correlation between contact pressure, crater morphology, and series resistance[END_REF][START_REF] Van Den Daele | Adaptation of the pseudo-metal-oxide-semiconductor field effect transistor technique to ultrathin silicon-on-insulator wafers characterization: Improved set-up, measurement procedure, parameter extraction, and modeling[END_REF] or phase transformation in the silicon film [START_REF] Benea | Nanoindentation effects on the electrical caracterizaron in Ψ-MOSFET configuration[END_REF]. Therefore, an implementation of ohmic contacts allowing both electron and hole transport with pressure controlled probes in the simulation software has not yet been developed.

The first aim of this work is to develop a simulation strategy for technology computer-aided design (TCAD) software that allows an accurate reproducibility of the measured "static" electrical behaviour of the pseudo-MOSFET with metal contacts. The simulations will be used in the second part of the paper (section 3) to reproduce the electrical measurements of the configuration under fast transient conditions.

2.Pseudo-MOSFET contact emulation for the simulations

Pseudo MOSFET configuration characteristics

In order to understand the challenges for the pseudo-MOSFET simulation, we will describe the typical experimental aspects. The wafer used for this work has mesas of 5x5mm 2 top silicon film defined by lithography and reactive ion etching from SOI. This produces a defined area of the silicon and restricts systematic current leakage that may appear if the BOX has defects on a side of the full wafer. The SOI has a silicon thickness tSi=88nm and a BOX thickness tox=145nm. The film is lightly P-type doped (~10 15 cm -3 ). The substrate underneath the BOX has the same doping concentration as the film. The source and drain contacts are two tungsten pressure-controlled probes installed in a Jandel probe station. The wafer is placed on a metal chuck onto which the gate voltage (VG) is applied (Fig. 1a).

Typical drain current (ID) with respect to gate voltage (VG) characteristics for low drain voltage (VD), obtained with an Agilent B1500 semiconductor analyser, are presented in Fig. 1b. In the characteristics, we observe the ID associated to the electron channel in strong inversion (VG>0) and the hole current in accumulation (VG<0). The subthreshold slopes are comparable for the electrons and holes channels but the maximum current for holes is significantly lower due to different carrier mobilities and different series resistances [START_REF] Ionica | Characterization of silicon-on-insulator films with pseudo-metal-oxide-semiconductor field-effect transistor: Correlation between contact pressure, crater morphology, and series resistance[END_REF]. As previously seen [START_REF] Williams | Point contact pseudometal/oxide/semiconductor transistor in as-grown silicon on insulator wafers[END_REF], low VD guarantees ohmic regime and the metallic probes are able to provide both e -and h + according to the VG sign.

Pseudo-MOSFET simulation strategy

The challenge in the simulation is that the contacts should be defined to match the experimental configuration and to allow the injection of both types of carriers into the channel. We start by defining a structure (tsi=88nm and tBOX=145nm) in Sentaurus with the same layer thicknesses as the experimental one (Fig. 1a). To consider substrate effects, a tsub=1.2μm bulk silicon region was included under the BOX in the simulation (the maximum space charge region was calculated at about 880nm for the doping of the substrate). The injection of both carrier types was demonstrated by defining metal regions as contacts with variable metal workfunctions [START_REF] Munteanu | Numerical simulation of the pseudo-MOSFET characterization technique[END_REF]. However, this strategy was not very successful to reproduce the out-of-equilibrium phenomena. An alternative way to inject the two carrier types is to use highly doped P and N type regions as source and drain. If only one type of dopants is chosen, the electrical simulations will only show the channel of e -or h + .

Consequently, we would need to combine the results produced from two different structures, one for VG>0 and one for VG<0, to reproduce the complete ID-VG characteristics. For the study of the out-of-equilibrium phenomena this is not an option since they occur during the transition between the accumulation and the inversion regimes. In order to reproduce the full pseudo-MOSFET behaviour with a single structure, the contacts were defined as two regions of silicon with different doping types and a concentration of 10 20 cm -3 . The source and drain in simulations both have a P + -type and N + -type regions for the injection of electrons and holes, as shown in Fig. 2. The total length of the device is made of the contacts length Lcon, the distance between their inner edges L and 10% of L added on each side of the structure.

This extension was included to the simulation structure for improved correlation to the experimental configuration. For all the simulations, we kept the L constant at 1μm. Different simulated combinations of doped areas were examined in order to emulate the contacts. The selection of the proper structure emerges from the benchmarking of the ID-VG simulated curves versus the real measurements. The drain voltage was set to 0.1V for all the simulations as in experiments and the source contact was grounded. Default models (ohmic boundary conditions between electrodes and highly doped regions) were applied for the electrode definition which was placed on top of highly doped regions. For the carrier mobility, the Lombardi model and doping dependence were used. In Fig. 3a, the doping regions P + or N + on the contacts are mirrored with respect to the centre of the device. The simulated ID-VG characteristics shown in Fig. 3b reveal that one carrier is favoured over the other. We evidence a really steep subthreshold slope with higher ON-current in PN-NP for electrons and in NP-PN structure for holes. Positioning a highly doped region on a lightly doped silicon layer creates a barrier for the opposite carrier type. For instance, the space charge region under the N + region is a barrier for the hole flow through that side of the contact. To avoid this effect and to emulate the contacts properly, we studied the alternating doping regions (PN-PN and NP-NP, Fig. 4a). In this case, we reproduce relatively similar subthreshold slopes in the ID-VG curves (Fig. 4b) on both electrons and holes sides and a comparable ON-current for both configurations. The ON-current is slightly stronger in the electrons side compared to the holes side similarly to the experiments. While the difference in series resistance is mostly responsible for the higher electron ON current for the experiments, The dynamic character of any phenomenon originates from its dependence on time.

To proceed with the time dependent experimental study, we apply a large triangular voltage ramp (AC), as illustrated in Fig. 5, to the gate of the device in order to mimic the quasi-static (DC) VG sweep in the forward and reverse scanning directions. The VG scanning range of the measurement corresponds to the peak to peak of the triangular signal. We used a constant range from -5V to 5V (bias range allowing to cover the transient behaviour of interest) in all the transient experiments. The scanning speed of the back gate was varied by changing the triangular pattern frequency covering scanning speeds (SS) from 10V/s up to 400V/s. In all cases, the source contact was grounded and the drain was biased at VD=0.1V. To perform with precision the above study, we used an Agilent B1500 analyser with a B1530A fast I-V module. Additionally, a slow transient measurement with a scanning speed of SS≈1.667 V/s was included for a direct comparison between the large signals and "quasi-static" sweep measurements. The scanning speed was chosen to correspond to a DC sweep with a 100mV gate step, 20ms integration time and 40ms delay time.

Figure 5: Schematic of the triangular large signal, applied to the gate of the device.

Large signal versus sweep voltage measurements

The ID-VG and IG-VG curves for the static sweep and transient measurements are illustrated in Fig. 6. Although, the produced drain current curves have similar response for strong inversion or accumulation, a deviation on the subthreshold slope on the holes current can be observed. This behaviour can be attributed to the charging/discharging of slow traps which might be different in the two experiments because the evolution of the voltage is not done the same way, even if the total scan time is the same. For the sweep measurements, the voltage variation is done step by step while VG is ramped with shorter voltage intervals for the transient ones. Also, the hysteresis effect on the electrons side between forward and reverse directions vanishes. On the other hand, gate current in IG-VG curves has significant

Transient effects on the drain current

To investigate the limit for which the transient phenomena do not affect drastically the drain current characteristics, we traced them for various scanning speeds. The respective curves are illustrated in Fig. 7a for the forward scanning direction and in Fig. 7b for the reverse. The effect of scanning speed in the ID-VG characteristics has a significant impact for scanning speeds greater than SS=20V/s. Additionally, hysteresis effects are noticed between the two scanning directions. A similar hysteresis has been reported for silicon [START_REF] Shen | A fast measurement technique of MOSFETI/sub d/-V/sub g/ characteristics[END_REF] and SiC MOSFETs [START_REF] Gurfinkel | Characterization of Transient Gate Oxide Trapping in SiC MOSFETs Using Fast I-V Techniques[END_REF] and was associated with the threshold voltage variation due to the charging of traps during VG scans. In order to check if this explanation applies here, we simulated the ID-VG curves in a NP-NP structure with surface traps at the Si/BOX interface (Fig. 8). The traps density was Dit=2•10 11 eV -1 cm -2 and they follow a Gaussian distributions located at 0.35eV from the valance/conduction band for donors/acceptors, as used [START_REF] Van Den Daele | Adaptation of the pseudo-metal-oxide-semiconductor field effect transistor technique to ultrathin silicon-on-insulator wafers characterization: Improved set-up, measurement procedure, parameter extraction, and modeling[END_REF] or recommended in the literature [START_REF] Broqvist | Defect levels of dangling bonds in silicon and germanium through hybrid functionals[END_REF][START_REF] Tsipas | Modeling of negatively charged states at the Ge surface and interfaces[END_REF]. The simulated results show a shift in the subthreshold slope for increasing scanning speed. However, we cannot explain the experimental curve behaviour only with the appears in strong inversion and accumulation and decreases in the depletion region. This is related to the total capacitance that drops significantly in the depletion region [START_REF] Diab | RC Model for Frequency Dependence of Split C-V Measurements on Bare SOI Wafers[END_REF]. The peak in Fig. 9 is the result of a displacement current which is imposed by the lack of carriers in the transition from depletion to strong inversion/accumulation. Due to the opposite carrier types needed in the two different scanning directions, the peak is positive for a forward scan and negative for the reverse.

Comparing to "static-like" measurements, a significant increase in the maximum value of IG peak value is noticed. Additionally, the peak shifts to higher VG values for increasing scanning speed. Considering the dynamic character of this phenomenon, the increment in IG peak value was foreseen. In the silicon film, during the transition from depletion to inversion (or accumulation), the total device capacitance increases and so does the need for carriers to charge/discharge the BOX capacitance. Since the total carrier density to reach equilibrium is constant [START_REF] Benea | Out-of-equilibrium body potential measurements in pseudo-MOSFET for sensing applications[END_REF], the increase in scanning speed induces a higher IG.

Indeed, as the scanning speed increases, we notice an increase in the gate current value. This evolution of the plateau current with the scanning speed can be explained by the displacement current which is equal to the product of a capacitance and the derivative of the voltage with respect to time. In strong inversion/accumulation the capacitance is constant and given by the BOX capacitance. As a result, IG plateau current increases linearly with the scanning speed. Simulations with the NP-NP structure have also been performed in similar conditions for the forward scan direction. In simulations, the scanning range was set from -10V to 10V, in order to confirm the area of interest. The scanning time was adjusted accordingly to maintain the same scanning speeds as in the experiments. The contact length for those simulations was set to Lcon=0.5μm. It is worth noting that since no VD is applied here and the effects are related to charging of the BOX capacitance, Lcon was not kept at 0.1μm as in the static simulations but was chosen to give strong out-of-equilibrium signatures. The results are presented in Fig. 10a. Although they follow the same trends as the experimental curves in Fig. 9, the current values are significantly lower. Considering that all the above mentioned phenomena are related to the total BOX capacitance, they will be impacted by the area of the simulated structure which is different from the actual size of the configuration. To verify this assumption, we proceed to a normalization of the current with respect to the total area of the However, the total area of the device is not the only difference between the experiments and the simulations. The length of the contacts with respect to the channel length is also different and might affect the simulated results. To further investigate its possible role on the maximum IG value, simulations of various contact lengths and areas have been performed. The total area of the simulated device equals to the product of its total length by its width (set at 1μm in the simulator). The results are shown in Fig. 11 and reveal that the increase of the IG maximum value and position has a strong dependence on the contact size.

Larger contacts result in higher IG maximum values. This is an indication that the maximum value might depend on the ability of the carriers to be injected in the film. In the case of Lcon=0.1μm, the highly doped silicon regions cannot provide the complete amount of carriers needed for the capacitance charge or create the necessary barrier, hence the IG peak is not present. For larger contacts, the IG peak occurs as in the experiments. When the contact size increases, the interface between the highly doped regions and the silicon film increases. If the area is sufficient, the simulated contacts are able to provide the carriers or create the barrier and the peak increases. The capacitive effects are connected to the geometry of the device. The large area of the SOI mesas used experimentally induces capacitive effects that affect significantly the device performance when the scanning speed is sufficiently high. This is evidenced in Fig. 7 with the deformation of the ID-VG curves which, in strong inversion/accumulation, have comparable values as the capacitive induced current. Moreover, the dynamic character of the IG current indicates analogous behaviour to other out-of-equilibrium phenomena which are related to the body potential response, that are shown in the next section.

Transient out-of-equilibrium body potential measurements

The connection of the IG displacement term with the out-of-equilibrium body potential (VB) is well defined in FD MOSFETs [START_REF] Bawedin | Dynamic body potential variation in FD SOI MOSFETs operated in deep non-equilibrium regime: Model and applications[END_REF]:

( ) ∆ - = ∆ G B G V V I C t ∆ ∆ = - ∆ ∆ B G G V V I t t C ∆ ∝ -∆ B G V I t ( 1
)
where C is an adjusting capacitive coefficient. In the pseudo-MOSFET configuration, the VB response under quasi-static measurements is documented [START_REF] Benea | Out-of-equilibrium body potential measurements in pseudo-MOSFET for sensing applications[END_REF] and was monitored through a third probe placed halfway between the source and drain probes. In accumulation and inversion regimes, the body voltage is simply halfway between the source and drain voltage, hence VB=VD/2 since source is grounded. In the depletion region, if VG is scanned fast between accumulation and inversion, the carriers are lacking and VB varies.

The measurement of the body potential in the depletion region is complex because the experimental configuration has a high impedance value of Z≈10 9 Ω at 40Hz [START_REF] Sato | Impact of contact and channel resistance on the frequency-dependent capacitance and conductance of pseudo-MOSFET[END_REF].

Additionally, the RSU (Remote-sense and Switch Unit) of the B1530A cannot be used as a current source and impose a zero current to the body probe which would be necessary for an accurate VB measurement. The modification of the experimental set-up was a necessary step to overcome the above drawbacks (see Fig. 12). An arbitrary waveform pulse generator (Agilent 33250A) was used to impose the large triangular signal. As demonstrated in [START_REF] Benea | Out-of-equilibrium body potential measurements in pseudo-MOSFET for sensing applications[END_REF], the drain electrode is not mandatory for the observation of VB, but in order to mimic the static measurements, an external power supply (Agilent E3642A) was biasing the drain at VD=0.1V. The VB probe was connected to the input of a voltage follower, which is a unitygain amplifying circuit with a high input impedance. The circuit was built using a TL081 operational amplifier with its inverting input connected to its output. The monitoring of the signals was performed with an oscilloscope (LeCroy Wafesurfer 424). The same scanning speed signals as in transient measurements were applied and a slow measurement was included to benchmark the new set-up with a quasi-static measurement. the accumulation/inversion regimes, with a value of VD/2. For the forward scan, there is a small shoulder appearing in the new configuration. Additionally, the reverse VB peak increases slightly. The body potential drop also shifts towards inversion or accumulation.

That was actually expected because as shown by eq.( 1), the body potential drop is linked to the maximum value of IG. In Fig. 6b the measured IG peak is shifted also towards accumulation/inversion, when moving from static to transient measurements. The shift in the VB drop is consistent to the above mentioned result and is an additional argument for the validation of the experimental set-up. VB and IG peaks in the reverse scan can be reproduced by the simulations as well (Fig. 17) but, it is the PN-PN structure that must be used in this case. Indeed, the forward behaviour is well reproduced by the NP-NP and the reverse needs the PN-PN structure (Fig. 17). VB is probed on the right electrode in both cases. This outcome is connected to the nature of the simulation. Different carrier types would be injected from different probes according to the device conditions. By setting the boundary condition of zero current to one of the probes in the simulation, the carrier injection is blocked through the probe (due to electrode conditions in the simulation). Thus, only one scanning direction can be reproduced per structure.

4.Conclusion

We propose a novel approach to simulate the pseudo-MOS configuration with a single structure that works from accumulation to strong inversion. In transient measurements, the drain current in the pseudo-MOSFET configuration can be described as the sum of two components, static and transient. The transient starts to affect the static-like curves for scanning speeds higher than 20V/s. The effect on the drain We simulated ID-VD for the two alternating combinations of contacts NP-NP (Fig. A2) and PN-PN (Fig. A3) under two bias conditions: VG=5V (corresponding to the subthreshold regime) and VG=8V (corresponding to strong inversion). The contact length (Lcon) was added as a supplementary parameter.

The smallest contacts (Lcon=0.1μm), give a linear behaviour in ID-VD curves, in every case.

As the contact size increases the deviation from the linearity is more evident especially for lower gate voltage (VG=5V). Short contacts have similar response to the experiments. This can be related to the ratio between the total area of the contacts and of the silicon island.

Indeed, the size of the contact in the experimental configuration is several orders of magnitude smaller compared to the total size of the device (about 30μm diameter of contact for 5mm side of silicon). In the case of the large simulated contacts, the length of the contacts is almost half of the total device length. Additionally in the 2D simulation, the profile of the device is the same on the full width of the device but in experimental conditions, the current density surrounds the contact [START_REF] Munteanu | Numerical simulation of the pseudo-MOSFET characterization technique[END_REF]. A more appropriate approach could be the 3D simulations which are more complex. However, since the 2D simulation with these structures reproduce well the experimental results, we use them in this paper for the proof of concept of the transient signatures.

Although, the short contacts mimic well the ID-VD characteristics under quasi-static conditions, larger contacts enhance the transient response as shown in section 3.4. A fully quantitative study would require a fine tuning of the structure which is not the purpose here.

Consequently we studied the dynamic phenomena with higher Lcon in order to amplify the out-of-equilibrium responses. 14). As already mentioned, the "home-made" reading circuit is vulnerable to environmental noise and to the 50Hz harmonic. To illustrate this clearly, we present the magnitude of the FFT on the frequency domain (Fig. B1a, B2a, B3a). For the slow scanning speeds an isolation of the harmonic by blocking frequencies between 25Hz and 75Hz results in a clear imprint of the body potential in the depletion region. However, high frequency noise is still present. An increase in the range of frequencies that are blocked results in a clean representation of the monitored signal. The most interesting case correspond to the medium frequency, Fig. B2. In this situation, during the linear increase in VB, several periods of the 50Hz noise are present. This becomes clear from Fig. B2a. Since, the frequencies of the output are comparable to the harmonic, a proper isolation of it is not feasible. Lastly, when the scanning speed is sufficiently high, the monitored signal has a frequency close to 50Hz so, the 50Hz no longer appears independently in the Fig. B3a. To be consistent with the data treatment, a 6kHz low pass filtering was applied the raw experimental data with high scanning speed to exclude the high frequency noise.

Figure 1 :

 1 Figure 1: a) Schematic of the experimental configuration and b) Measured ID-VG characteristic curve for VD=0.1V.

Figure 2 :

 2 Figure 2: Pseudo-MOSFET simulation structure to emulate the source and drain contacts obtained experimentally with controlled pressure probes.

  in the simulations, this deviation is mainly caused by the different mobility values for electrons and holes in the silicon. This can be evidenced by the ratios between the maximum values of currents and the mobility values in the simulator which are almost the same (IONelectrons/ION-holes≈3.45, μelectrons/μholes≈3.11). The alternating doped contacts are more appropriate for the pseudo-MOSFET simulations and will be used for the next sections. A supplementary study based in the ID-VD curves is shown in the appendix A.

Figure 3 :

 3 Figure 3: a) Schematic of the simulated structures with mirrored doping regions with respect to the centre of the device b) Simulated ID-VG for mirrored contacts, VD=0.1V and Lcon=0.5μm.

Figure 4 :

 4 Figure 4: a) Schematic of the simulated structures with alternating doping for the contacts b)Simulated ID-VG for alternating doping contacts, VD=0.1V and Lcon=0.5μm.3. Transient Behaviour of the pseudo-MOSFET

  higher values for the whole VG scanning range in transient measurements and the peak of IG shifts towards inversion/accumulation. The stronger signature on the transient IG can be assigned to (i) the shorter voltage intervals that are applied to the gate in the transient measurements and (ii) the averaging of several measurement points in static (DC) experiments which reduce the dynamic response.

Figure 6 :

 6 Figure 6: a) ID-VG (red full and blue empty symbols) and b) IG-VG (black and blue lines) for static and transient measurements. Scanning speed SS≈1.667V/s and drain voltage VD=0.1V.

  effect of Dit. The transient behaviour of the traps is, most likely, screened by other transient effects which originate from capacitive phenomena evidenced by the gate current transient response in the next section.

Figure 7 :

 7 Figure 7: Measured absolute ID (empty symbols correspond to negative values) with respect to VG for various scanning speeds. a) scanning from accumulation to inversion and b) from inversion to accumulation.

Figure 8 :

 8 Figure 8: Simulated ID-VG curves for NP-NP structure with interface traps between the BOX and the film for various transient simulation times.

Figure 9 :

 9 Figure 9: Experimental transient gate current versus gate voltage characteristics for various scanning speeds.

  device in both experiments and simulations. The normalized IG current (IG density) is presented in Fig10b. The comparison between the measurements and simulations of IG are almost perfectly matched, besides the position of the IG maximum value.

Figure 10 :

 10 Figure 10: a) Simulated IG-VG curves for various scanning speeds using the NP-NP structure scanning from accumulation to inversion b) Comparison between experimental curves and simulated IG density.

Figure 11 :

 11 Figure 11: Simulated IG-VG curves for NP-NP structure for various contact sizes and areas.

Figure 12 :

 12 Figure 12: Experimental configuration for the measurement of the transient body potential.

Fig. 13 shows

 13 Fig.13 shows the comparison of the VB response between transient and sweep

Figure 13 :

 13 Figure 13: VB versus VG (red line for transient and blue line and symbols for static "static-like" measurements respectively) and ID versus VG (black symbols) characteristics. Transient SS=0.416V/s, equivalent sweep period T=24s.

Figure 14 :

 14 Figure 14: Measured transient VB versus VG curves for various scanning speeds. VD=0.1V.

Figure 15 :

 15 Figure 15: Simulated body potential with respect to gate voltage and for various scanning speeds.

Figure 16 :

 16 Figure 16: a) Simulated IG and derivative of VB b) Simulated IG and VB in log scale for SS=20V/s produced by NP-NP structure scanning from accumulation to inversion

Figure 17 :

 17 Figure 17: Simulated IG and of VB versus VG for a) NP-NP structure scanning from accumulation to inversion b) NP-NP structure scanning from inversion to accumulation c) PN-PN structure scanning from accumulation to inversion d) PN-PN structure scanning from inversion to accumulation. Simulation ramp time=1s.

  Its validity was highlighted by a direct comparison to sweep and transient measurements. The new structure emulates the experimental configuration with the controlled pressure probes by combining N + and P + regions for each source/drain electrodes. It is capable of reproducing the out-of-equilibrium phenomena with good agreement with the experiments, even if the actual experimental configuration is rather 3D than 2D as in the simulations. However, the order of the N + and P + regions has to be modified to successfully reproduce the results for scanning from accumulation to inversion or the opposite. All the observed phenomena in the different experiments are reproduced, starting from ID in ohmic regime (low VD), to IG and to the transient body potential.

Figure A1 :

 A1 Figure A1: Experimental ID-VD characteristics for three positive VG biases.

Figure A2 :

 A2 Figure A2: ID-VD characteristics reproduced with NP-NP structure for a) VG=5V and a) VG=8V for three contact lengths.

Figure A3 :

 A3 Figure A3: ID-VD characteristics reproduced with PN-PN structure for a) VG=5V and a) VG=8V for three contact lengths. Appendix B: Data treatment

Figure

  Figure B1: a) FFT magnitude in frequency domain. b) Measured VB versus time and two signals treatments. Signal period T=24s.

Figure

  Figure B2: a) FFT magnitude in frequency domain. b) Measured VB versus time and two signals treatments. Signal period T=2s.

Figure

  Figure B3: a) FFT magnitude in frequency domain. b) Measured VB versus time and two signals treatments. Signal period T=0.05s.

Acknowledgments

Authors would like to thank SOITEC for providing the wafers and inspiring conversations.

current versus gate voltage curves that is visible over this limit, is actually the result of the strong capacitive induced gate current, which in this configuration has a really high value due to the large device area. For a strong transient character (high scanning speeds), IG is comparable to the ON-current. Therefore, the deformation of ID is strong. The plateau gate current in equilibrium regimes is induced by the BOX capacitance and the respective peak originates from its charging with the appropriate type of carriers.

The out-of-equilibrium phenomena are enhanced by the dynamic character of the transient measurements although special equipment and treatment is needed to observe their behaviour, especially for the transient body potential. The high impedance of the configuration in the depletion region requires a high input impedance circuit to measure VB properly.

Finally, the transition from quasi-static to transient measurements was established. All the experimental results are reproduced by the simulation and the connection between the gate current and the body potential is validated.

Appendix A: Response of the emulated contacts through ID-VD characteristics

The simulated ID-VG curves in section 2.2 showed that the alternating-doping contacts (PN-PN or NP-NP) are the most appropriate ones. In order to further assess the behaviour of the emulated probes, drain current versus drain voltage ID-VD simulated curves are benchmarked with the experimental ones. In Fig. A1, the experimental ID-VD curves show a linear behaviour for low VD. For VG=5V, the appearance of the onset of the saturation is noticeable for high VD values. The increase of VG enhances the inversion charge and induces a higher ID.