Smart composite antibacterial coatings with active corrosion protection of magnesium alloys

A.S.Gnedenkov,S.L.Sinebryukhov,V.S.Filonina,N.G.Plekhova,S.V.Gnedenkov

Institute of Chemistry of FEB RAS,159 Pr.100-letiya Vladivostoka,Vladivostok,690022,Russia

Abstract A new method of the formation of composite coatings with the function of active corrosion protection of magnesium alloys was developed using the plasma electrolytic oxidation(PEO)method.Susceptibility of PEO-layers to pitting formation was evaluated using localized electrochemical methods(SVET/SIET).The morphological features and electrochemical properties of composite coatings were studied using SEM/EDX,XRD,micro-Raman spectroscopy and EIS/PDP measurements,respectively.The effect of surface layers impregnation with corrosion inhibitor on their protective properties in a corrosive environment was established.Additional protection was achieved using controllable coating pore sealing with polymer.It was found that the polymer treatment of the PEO-layer does not reduce the inhibitor’s efficien y.The formed protective composite inhibitor-and-polymer-containing layers decrease the corrosion current density of a magnesium alloy in a 3 wt.% NaCl solution to three orders of magnitude.This predetermines the prospect of new smart coatings formation that significantl expand the fiel of application of electrochemically active materials.The mechanism of smart composite coating corrosion degradation was established.The antibacterial activity of the inhibitor-containing coatings against S.aureus methicillin-resistant strain was proved using the in vitro model.These protective layers are promising for reducing the incidence of implant-associated infections.? 2022 Chongqing University.Publishing services provided by Elsevier B.V.on behalf of KeAi Communications Co.Ltd.This is an open access article under the CC BY-NC-ND license(http://creativecommons.org/licenses/by-nc-nd/4.0/)Peer review under responsibility of Chongqing University

Keywords:Magnesium alloy;Plasma electrolytic oxidation;Corrosion inhibitor;Self-healing;Protective coatings.

1.Introduction

Corrosion is a major problem in a variety of industries where metals and alloys are used as structural and functional materials.This is one of the main reasons for energy and material costs arising during the service life of many parts and devices in various areas of industrial application[1-3].To reduce the amount of damage from corrosive destruction of materials,it is necessary to develop technologies for their anticorrosive protection,which will not only improve the operational characteristics of the processed product(including ensuring stable operation in aggressive environments:in sea water,salt fog,high humidity,etc.),but also expand the area of practical application of materials.

One of the common strategies for protecting metal parts from corrosion is the use of protective coatings.The method of plasma electrolytic oxidation(PEO)has a leading position in this fiel[4-10].The layers formed using this method have high chemical resistance and reduce the access of the aggressive medium species to the protected material[11-13].The PEO-coating has a convoluted morphological structure,as well as a large number of pores of various sizes,which can serve as a reservoir for the various functional substances,for example,corrosion inhibitors or bioactive,bioinert compounds[14,15].It was found that sealing the porous part of the PEO-layer with a polymer material makes it possible to form composite polymer-containing coatings with high anticorrosive and antifriction properties[16-19].However,such layers provide only passive protection of the material,and,in the event of a breach of integrity,the corrosion process will proceed under the coating,gradually destroying the substrate.One of the solution of this problem is the formation of coatings with an active corrosion protection.When a defect forms on the surface of such a coating,an additional barrier layer is formed in the damaged area,which prevents further development of corrosion.Such anticorrosive layers in the world scientifi practice are called self-healing coatings[14,20-23].

Based on the analysis of literature data,two main mechanisms of self-healing of protective layers in the case of a corrosion degradation of material can be characterized:a)the firs one,due to the formation of corrosion products that block the access of an aggressive medium to the active centers of the sample,b)the second one,due to the formation of protective layers as a result of activation of organic or inorganic corrosion inhibitors present in the composition of the initial coating and reacting with components of the environment[24].

It should be noted that pores of the PEO-coating satisfy the requirements for reservoirs for impregnation with corrosion inhibitors.These pores are characterized by:1)chemical and mechanical stability,2)compatibility with the coating matrix,3)sufficien capacity,4)efficien y as storage of an inhibitor,5)ability to release an inhibitor in the case of corrosion process[25].

In our previous works[5,14],a self-healing coating was obtained on the MA8 magnesium alloy(Mg-Mn-Ce system)in a silicate-fluorid electrolyte using the PEO method.An increase in the protective properties of the PEO-coating was established due to its impregnation with corrosion inhibitor-8-hydroxyquinoline(8-HQ).The self-healing mechanism was proposed.

In order to improve the corrosion characteristics of the obtained coating and reduce the uncontrolled premature release of the inhibitor,an additional surface modificatio is proposed in this work.The polymer layer was formed atop the PEO-coating containing 8-HQ.Such protective layer,as a result of polymer sealing the pores of surface layer containing the inhibitor,will ensure to increase the corrosion resistance,as well as to provide a targeted action of the inhibitor in case of coating degradation or damage/breach of its integrity.To prove this hypothesis,it is necessary to establish the effect of the stages composite layer design,namely,the impregnation of the coating with inhibitor and the subsequent formation of the polymer layer,on the corrosion properties of the protected material.

The formation of a layer that reliably protects the material from corrosion degradation,taking into account the unique functional properties of magnesium alloys(high strength characteristics and low specifi weight[26-31]),can significantl advance their use in various industries,from aircraft and rocket production to medicine.

Nowadays the research and development of magnesium based implants are of significan interest[28,32-40].This metal has great potential for use in the fiel of temporary orthopedic implants and cardiovascular stands due to its good biocompatibility and unique biodegradability[41-43].The antibacterial potential of Mg is provided by its ability to raise the pH in liquids[44,45],which is important for its application in the clinic,since infections associated with surgical implants are becoming a serious problem nowadays.In orthopedic infection of the implant,the main etiological agents of bone tissue infecting areS.aureusandS.epidermidisand other staphylococci.Moreover andS.aureuscauses 80% of types of human osteomyelitis[46,47].In the area of acute osteomyelitis,damage to the bone matrix,compression and obliteration of the blood vessel as a result of the spread of infection lead to bone necrosis.In the presence of dead bone(sequestration),antibiotics and immune cells do not reach the infected area,and the treatment of osteomyelitis is ineffective[47].In this case,the use of metal implants with coatings,including targeted drug delivery,is relevant.Moreover,due to the high electrochemical activity of magnesium,its application in surgery is limited since the possible premature dissolution of Mg implant in the body can occurred.To effectively reduce the rate of degradation of Mg-based implants in body fluids their surface modificatio is used,for example,by applying an oxide ceramic-like PEO-coatings[16,33,48-50]or a hydroxyapatite coating using electrochemical deposition[33],etc.Since 8-HQ possesses a wide range of beneficia features including antibacterial activity[51]one of the purposes of this work is to combine such topics as the study of active corrosion protection of the formed protective inhibitor-containing layers and comparative evaluation of the antibacterial properties of various coatings on the MA8 alloy against methicillinresistantS.aureus(MRSA).

The literature analysis showed works,where plasma electrolytic oxidation was used to form an antibacterial protective layer on the surface of different alloys(Mg,Al,Ti).In the recent review[52]Fattah-alhosseini et al.illustrated the challenges in formation of antibacterial bioceramic PEO-coatings on magnesium and its alloys.Zhang et al.[53]grew ferric oxyhydroxide nanosheet film on the PEO-pretreated Mg alloy,which provided the antibacterial activity of the material.The composite coatings with antibacterial ability were fabricated on pure magnesium using plasma electrolytic oxidation and CuO particles addition[54].Han et al.[55]formed antibacterial PEO-coatings with incorporated Ag particles on pure Mg.The effect of particles addition in PEO electrolytes on the antibacterial activity of the produced coating was described in[56].The high antibacterial activity of PEO/nano-ZrO2coating formed on Mg alloy was established in[57].Pezzato et al.[58]formed PEO-layer with fungicidal properties on the Al alloy surface.Cerchier et al.[59]found the antimicrobial effect of the PEO-coating formed in alkaline solution containing silicates compounds and silver micrometric particles on the AA7075.The effect of PEO on the antibacterial ability of titanium implants was discussed in the review[60].In[61]authors established that PEO-coating with incorporated graphene oxide provides antibacterial activity to titanium substrates.However,the above-mentioned studies dealt with the evaluation of the bactericidal properties of modifie PEO-layers and the active corrosion protection was not achieved in the produced coatings.

The novelty of this work is based on the formation of the new smart composite antibacterial coating with prolonged and effective self-healing properties to targeted expand the fiel of practical use the magnesium-based materials in various field of industries including automobile,aerospace and medicine.For this reason,the different coating systems were designed and analysed to meet the requirements of the various possible area of material application.

2.Experimental

2.1.Materials

MA8 magnesium alloy(wt.%:1.5 to 2.5 Mn;0.15 to 0.35 Ce;Mg-balance)was used as a substrate for the coating formation.After grinding,the samples were degreased with ethanol and dried in air.The size of the investigated samples was 20×20×1 mm.

2.2.Base PEO-coating formation

In this work,two groups of samples with PEO-coatings were prepared.

The firs group was obtained by oxidation using a pulsed bipolar mode in a silicate-fluorid(SiF)electrolyte(Na2SiO3·5H2O,15 g L-1and NaF,5 g L-1),according to the method described in[14].The anodic component was maintained in a potentiodynamic mode,in which the voltage increased from 30 to 300 V with a sweep rate of 0.45 V s-1.The cathodic one was maintained potentiostatically at 30 V.The duration of oxidation was 10 min.

The second group was obtained in an electrolyte containing calcium glycerophosphate,sodium fluorid and sodium silicate(GP)(C3H7O6P)Ca2·H2O,25 g L-1;NaF,5 g L-1;Na2SiO3·5H2O,8 g L-1),using the pulsed bipolar PEO mode[44].The anodic component was maintained potentiostatically at 380 V,and the cathodic one was maintained in the galvanodynamic mode,in which the current density varied from 1.3 to 0.8 A cm-2at a sweep rate of 4.5 mA cm-2s-1).The oxidation time was 110 s.

The frequency of the polarizing signal for these PEO modes was 300 Hz(pulse duration 3.3 ms,no pauses).The ratio of the duration of the anodic and cathodic pulses was equal to 1.The duty cycle was 50%.The rms voltage values were recorded during the PEO.The electrolyte temperature was kept at 15 °C.

2.3.Composite coating formation

After washing with deionized water and drying in air,magnesium alloys with PEO-coating were treated with an aqueous solution of 8-hydroxyquinoline(3 g L-1)by dip-coating,C9H7NO,which was prepared by dissolving it in distilled water at a temperature of 90 °C with the addition of sodium hydroxide NaOH(pH=11)according to the procedure described in[5,14].

After coating impregnation with corrosion inhibitor,the pores of the PEO-layer were sealed with a polymer.Two types of polymer-containing coating were obtained.The firs one is formed using 15 wt.% suspension of superdispersed polytetrafluoroet ylene(SPTFE)(Forum,Russia)in isopropanol.The pores of PEO-coating were impregnated with polymer using dip-coating method.The samples were kept in suspension for 15 s,dried at room temperature for 15 min,and heated at 270 °C for 15 min.Then the samples were cooled down to 25 °C.In this work,we used a single(1x)and triple polymer treatment(3x).The second type of coating was also obtained by the dip-coating method using a 6 wt.%solution of polyvinylidene fluorid(PVDF)(MTI Corporation,USA)in N-methyl-2-pyrrolidone(C5H9NO).The samples were kept in solution for 10 s,then dried at 70 °C for 120 min and cooled to 25 °C.

The studied coatings are designated in this work as PEO+8-HQ,PEO+8-HQ+SPTFE(1x,3x),PEO+8-HQ+PVDF.To establish the effectiveness of the corrosion inhibitor,samples with 8-HQ-free coatings were also obtained.Such protective layers are designated as PEO,PEO+SPTFE(1x,3x),PEO+PVDF.The summarized information about the formation procedure of the abovementioned coatings is presented in Table 1.

2.4.Electrochemical tests

Electrochemical properties of samples were studied using potentiodynamic polarization(PDP)and electrochemical impedance spectroscopy(EIS)techniques.12558WB system(Solartron Analytical,UK),consisting of an SI 1287 electrochemical interface and an FRA 1255B frequency response analyzer connected to a computer was used.The measurements were carried out in a Model K0235 Flat Cell three-electrode cell(PAR,USA).Samples were studied in a 3 wt.% sodium chloride solution.The surface area in contact with the solution was 1 cm2.The saturated calomel electrode(SCE)served as a reference electrode.To stabilize the electrode potential,the samples were kept in the solution for 15 min.The potential sweep rate during potentiodynamic polarization was 1 mV s-1.The samples were polarized from the potentialE=Ec-300 mV toEc+500 mV.Corrosion potentialEc,corrosion current densityIc,anodic and cathodic Tafel slopes of the polarization curve,βaandβc,respectively,were calculated using the Levenberg-Marquardt method.For EIS measurements a sinusoidal disturbance signal with an amplitude of 10 mV was used.Impedance spectra were recorded in the frequency range from 0.01 Hz to 1 MHz with a logarithmic sweep of 7-10 points per decade.Measurements and experimental data processing were performed using CorrWare/ZPlot and CorrView/ZView software,respectively.To ensure the reliability and reproducibility of the experimental data obtained,three samples of each type were tested.

Heterogeneity and electrochemical activity of MA8 magnesium alloy sample with PEO-coating were studied using localized electrochemical methods(scanning vibrating electrode technique,SVET;scanning ion-selective electrode technique,SIET)for 30 h during exposure to a 0.05 M sodium chloride solution.Specimens were fi ed on the epoxy resin pellet.The scanned area was 1.5 mm×2.5 mm(for SiF-coating)and 1.5 mm×1.6 mm(for GP-coating)after beeswax isolation.Local current density was measured by SVET using a Pt-Ir-electrode(Microprobe Inc.,USA).Local concentration of

H+was measured by SIET using a pH-selective glass capillary electrode(2μm tip diameter).The detailed information about electrode preparation,calibration and methodology of SVET/SIET measuring are described elsewhere[16,62,63].Local pH and local current density were measured quasisimultaneously[64],using custom made dual-head stage.SVET/SIET measurements were performed using commercial system from Applicable Electronics(USA).

The summarized information about different protective coatings formed on MA8 Mg alloys.Table 1

Fig.1.SEM image and corresponding element distribution over the surface of MA8 magnesium alloy.

2.5.Coatings’characterization

X-ray diffraction analysis(XRD)of the samples was carried out on a SmartLab diffractometer(Rigaku,Japan)using a CuKβradiation source.Scanning was performed at 140 mA and 42 kV in the 2θrange from 4 to 85°at a speed of 0.01°/s.

Scanning electron microscopy(SEM)and energy dispersive X-Ray spectroscopy(EDX)were performed for surface characterization using EVO 40 microscope(Carl Zeiss,Germany)equipped with INCA X-act instrument(Oxford Instruments,UK).

Micro-Raman spectroscopy was applied to study the composition of 8-HQ-containing coatings.Tests were performed using a Raman spectrometer alpha 500 coupled with the confocal Raman microscope(WITec,Germany).Micro-Raman spectra were collected in the wavenumber range from 100 up to 1200 cm-1during 15 min(1 s exposure time,900 accumulated spectra).The measurements were carried out by means of 532 nm excitation laser and WITec Control software.The laser power was decreased down to 25 mW to prevent the local heating and probable degradation of inhibitor.2D maps of 8-HQ intensity distribution in the PEO-coating were obtained using the scanning mode.The size of analyzed surfaces was 200×200μm and 320×280μm.These scanned zones contain 50×50 micro-Raman spectra.All collected micro-Raman spectra were acquired using 1 s integration time.

Fig.2.SEM/EDX data of the morphology and elemental composition of the PEO-coatings formed in SiF and GP electrolytes.

2.6.Cross-section preparation

Cross-sections of specimens with base PEO-coating,inhibitor-containing coating(PEO+8-HQ)and composite inhibitor-and-polymer-containing coatings(PEO+8-HQ+SPTFE(3x),PEO+8-HQ+PVDF)obtained in SiF and GP electrolytes were prepared for SEM/EDX characterization.All steps of surface preparation were performed using Tegramin-25 grinding and polishing machine(Struers A/S,Denmark).Samples were embedded into SpeciFix epoxy resin.After preliminary grinding with SiC sandpapers,the specimens were degreased with ethyl alcohol.After that samples were polished with MD-Mol and MD-Nap disks(Struers,Denmark)using 3 and 1μm diamond suspensions(DPSuspension,Struers,Denmark),respectively.After the polishing step,the specimens were washed with deionized water,degreased with ethyl alcohol and air-dried.

2.7.Antibacterial assay

The antibacterial tests were performed using the direct contact method[65,66]on the following samples in the work:magnesium alloy without coating,magnesium samples with GP PEO-coating and GP PEO-coating impregnated with 8-hydroxyquinoline.Before tests,the obtained samples were sterilized in a dry oven(Thermo Fisher Scientific USA)at 180 °C for 2 h.Then the samples were rinsed with deionized water and incubated under ultraviolet light for 15 min on each side prior to inoculation with bacteria.Additionally the 8-hydroxyquinoline solution(3 g L-1)without Mg samples was tested for comparison of the antibacterial properties.

S.aureusmethicillin-resistant(MRSA,SCPM-Obolensk,Russia)andS.aureusPS84(SA,Public Health England,UK)strains were incubated with rotation in a thermostat at 37 °C for 2 h.Then strains were diluted with 1% tryptic soy broth(TSB,Corning Incorporated,USA)in 10 mM phosphate buffer solution to 2·105colony forming units(CFU)per mL.20μL of this suspension was applied on sterile nitrocellulose filte discs with a diameter of 10 mm,which were placed on the surface of blood agar(Oxoid,Thermo Fisher Scientific USA).The liquid was absorbed by the agar,and bacteria remained on the disk.

Separately,20μL which contains 1% TSB in 10 mM phosphate buffer solution and the same solution with addition of 10% blood serum was applied to the surface of each sample.Then discs inoculated withS.aureuswere placed on top of this liquid,while the bacteria were in contact with the surface of the samples.The samples together with the discs were placed in Petri dishes and incubated at 37 °C for 24 h in a humid atmosphere.After incubation,the samples and the corresponding discs were placed separately in 5 mL of 1% TSB,sonicated for 30 s,and shaken for 1 min to rinse off adhered bacteria.

Aliquots of 10μL of the undiluted suspension and its seven dilutions(from 101to 107)were dispensed into Petri dishes with blood agar,incubated for 18 h at 37 °C,and the number of colonies was counted.Three parallel experiments were performed in three replicates to provide the reliable results.

Statistical analysis of the results was performed using Statistica 8.0(StatSoft,USA).All values are presented as arithmetic mean±standard deviation(M±m(xù)).Data for groups were analyzed using Student’st-test with a confidenc level of 95% or higher(p＜0.05).

3.Results and discussion

3.1.Composition and morphology analysis

Fig.1 shows the results of SEM-EDX analysis of the microstructure of MA8 alloy.Presence of Mg-Ce round phases was detected in composition of the material.This secondary phase,namely Mg9Ce,is cathodic to the Mg matrix and responsible for heterogeneous corrosion of the alloy in accordance with our previous works[67,68],which show the necessity of the protective coating formation.

The morphology and elemental distribution on the surface and cross-section of the SiF-and GP-coatings were studied using SEM/EDX analysis.Fig.2 indicates that SiF-coating has more compact structure with less amounts of pores of smaller size as compared to GP-coating.The average pore size was 9±3μm and 18±4μm for SiF-and GP-coatings,respectively.The average coating thickness was 32±4μm and 81±14μm for SiF-and GP-coatings,respectively.Protective oxide layer obtained in SiF electrolyte characterizes with high amount silicon,sodium and magnesium.These elements uniformly distributed over the surface and within the thickness of the coating.The presence of thin strip of fluo rine near the substrate can be the result of magnesium alloy surface activation by F-ions during the oxidation process and formation of MgF2.Calcium,phosphorous,magnesium,silicon and sodium form the basis of the GP-coating.These elements spread throughout the entire thickness of the protective layer.

8-HQ treatment of the PEO-coating changes the morphology of the upper layer.The appearance of the fl wer-like structures on the surface is shown in Fig.3.Presence of carbon on the surface and in the pores indicates the presence of 8-HQ at the topmost part of the coating as well as its penetration as a result of continuous material immersion during impregnation.SEM/EDX analysis also shows that 8-HQ does not adversely affect on the microstructure of the PEO-coating and does not lead to its degradation.

To show the effect of polymer treatment on the morphology of the PEO-coating PEO+8-HQ+SPTFE(3x)was used as sample for SEM/EDX analysis.It is evident that the surface of the specimens with protective layers of the both types of base PEO-coatings become smoother as a result of polymer processing(Fig.4).Most part of the coating pores are sealed with SPTFE which is confirme by presence of high amount of fluorin and carbon on the surface and in porous part of the PEO-coating.However,the presence of calcium as a round areas on the surface of the GP-fil indicates that as a result of complex morphology of the PEOcoating the SPTFE layer is not continuous and uniform.This can promote more rapid emission of the 8-HQ and probably the better protective properties of the PEO+8-HQ+SPTFE GP-coating on the initial stage of the corrosion as compared to such type of protective SiF-layer.

Sealing pores with PVDF also increases the surface homogeneity(Fig.5).Similar to SPTFE,PVDF treatment also gives the pathway of the corrosive medium to the oxide GPcoating,which is shown by some holes in the polymer layer.However,such surface processing substantially smoothes the surface in the case of SiF-coating.

3.2.Raman microspectroscopy analysis

In order to prove the coating impregnation with 8-HQ the micro-Raman measurements were carried out.Inhibitorcontaining PEO-coating obtained in GP electrolyte was used as the test sample.Fig.6 shows two micro-Raman spectrum acquired from the crystal of 8-HQ powder(black curve)and from the surface of 8-HQ-containing PEO-coating(red curve).The spectrum of the 8-HQ illustrates peaks at 110,483 and 717 cm-1that are assigned to the torsion of quinoline ring[69].Raman bands at 1068 and 1141 cm-1correspond to in plane bending of the quinoline ring[69].The micro-Raman spectrum of the sample with inhibitor-containing coating depicts the same peaks at 110,717 and 1141 cm-1indicating the successful PEO-layer impregnation.However,this spectrum exhibits additional bands compared to the spectrum of the 8-HQ powder.This spectrum possesses peaks at 401,823 and 962 cm-1.The firs one at 401 cm-1is ascribed to the bending vibration(ν2)of the phosphate groups(HPO42-и PO43-)[70].The peak at 823 cm-1can be related to silicates in tetrahedral configuratio[71,72].The broad band at 962 cm-1is associated with symmetric stretching vibrations(ν1)of the PO43-group of the apatite phase[73].These three bands are related to the composition of the PEO-coating due to presence of magnesium orthosilicate(Mg2SiO4)and hydroxyapatite(Ca10(PO4)6(OH)2),which were established in the previous work[16].

Fig.3.SEM/EDX data of the morphology and elemental composition of the PEO-coatings formed in SiF and GP electrolytes after impregnation with 8-HQ.

The scanning mode of the confocal Raman microspectroscopy was applied to analyze the 8-HQ distribution in the coating.The 2500 micro-Raman spectra were collected from two different zones of the protective coating presented in optical images(Figs.7a,d).2D intensity distribution maps were designed using the filte,which was applied to all spectra in the range from 690 to 750 cm-1,which is corresponded to the intensive band of the 8-HQ at 717 cm-1(Fig.6).Both obtained 2D maps(Figs.7b,e)show the high intensity of 8-HQ distribution over the surface of PEO-coating(yellow-orange regions),which can also be seen from the optical images.However,the complementary information can be seen when overlaying 2D intensity map on the optical images(Figs.7c,f).These data show that most of the 8-HQ is adsorbed on the surface of the PEO-layer(Fig.7c).However,some part of the inhibitor also penetrated to the pores of the protective coating(Fig.7f).Symbol“+”was used to show the point,where the micro-Raman spectrum presented in Fig.6(red curve)was acquired.

3.3.Electrochemical properties

To show the moment of PEO-coating breakdown SVET/SIET were used(Fig.8).Analysis of SVET data indicates low electrochemical activity of the sample with SiFcoating.The local anodic current density did not exceed 1 μA cm-2(at the firs hour of sample exposure),which indicates the high anticorrosion properties achieved by treating the alloy by the PEO method.In spite of the absence of intensive corrosion processes of the material detected by SVET,the local pH values(SIET map,1 h)over the surface of the test material were in the range of 8.5-9.0,indicating the penetration of the aggressive medium through the PEOlayer to the Mg alloy substrate and its further degradation,which results in alkalization of the medium(equations 1,2).The breakdown of the PEO-layer was occurred after 20 h of sample exposure when SVET method starts to record at the microlevel the formation of electrochemically inhomogeneous regions on the surface of the material.SIET also shows zones with high alkalinity at these places of the coated Mg sample.These results indicate higher sensitivity of the SIET over SVET.However,both electrochemical techniques identifie the electrochemically active zones of the sample before pitting was indicated in the optical image.

Fig.4.SEM/EDX data of the morphology and elemental composition of the composite coatings obtained on the base of PEO-layers formed in SiF and GP electrolytes after 8-HQ impregnation and treatment with SPTFE.

The intensive change in the electrochemical activity of the sample was observed after 24 h of exposure.Local scanning electrochemical methods recorded the formation of microgalvanic couples on different parts of the surface under study.The anodic current density in the active zones reached 180 μA cm-2and the local pH values over practically all the investigated surface shifted to the alkaline range and increased up to 10.4 after 48 h of immersion.The electrochemical heterogeneity established by SVET/SIET was confirme by optical images of the studied area.The concentration of dark areas on the surface of the PEO-coating increased with the time of exposure of the sample to a corrosive environment.Moreover,pittings were observed in the protective layer on electrochemically active areas registered by SVET/SIET.

For the sample with GP-coating the same corrosion behavior was revealed(Fig.9).However,the coating breakdown was established after 12 h of material exposure(20 h for SiFcoating)as a result of lower compactness and higher porosity of this coating as compared to SiF-protective layer established by SEM/EDX analysis.The initial stage of the protective PEO-layer degradation was also detected using SIET after the 1 h of sample exposure.The firs signs of the electrochemical activity(alkaline area)of the Mg alloy sample were established in the right part of the sample.With the increase in immersion time,the corrosion activity of this area intensified which is evident from the current density and pH growth on SVET/SIET maps.The pitting formation was also approved by analysis of the optical images(Fig.9).

Fig.5.SEM images of the morphology of the surface PVDF-treated PEO-coatings(impregnated with 8-HQ)formed in SiF and GP electrolytes.

For inhibitor-and-polymer-containing coatings the local-ized electrochemical methods were not applied since the long period of measurements performing required to achieve the moment of the coating degradation.To show the substantial increase in protective properties of these composite coatings the conventional electrochemical methods were used.Using electrochemical impedance spectroscopy and potentiodynamic polarization,the mutual influenc of the inhibitor and the polymer on the protective properties of the coatings formed on MA8 magnesium alloy was studied.3 wt.% sodium chloride solution was used as a medium for studying the corrosion behavior of functional materials based on magnesium alloys under severe operating conditions,as well as for comparing the level of protective properties of the formed anticorrosion coatings.The use of this aggressive environment is appropriate for accurate and accelerated determination of the protective properties of coatings.The idea is simple-if a sample with a protective coating shows good anticorrosion properties in 3 wt.% NaCl solution,then it will also be stable in physiological saline(0.9 wt.%)in case of its further use in the biomedical field However,it is better to study the inhibitor efficien y of the formed coating firs in a harsh environment in order to reveal all possible areas of practical use of such a material with a smart coating.

The study of corrosion properties evolution carried out using EIS indicates an increase in the values of the impedance modulus|Z|in the full frequency range after incorporation of 8-hydroxyquinoline into the PEO-coating formed in a SiF electrolyte and subsequent polymer treatment(Figs.10a,11).After protective layer impregnation with 8-HQ,the impedance modulus at a frequency of 0.01 Hz increased up to 9 times for the base PEO-coating(Fig.10a)and 3-8 times for the polymer containing samples as compared to inhibitor free coating systems(Fig.11).Different increases of the|Z|f=0.01Hzdepended on the initial condition of the treated surface and the type of polymer processing of the base PEO-coating(Table 2).The dependence of the phase angle(Theta)on frequency indicates a change in the structure and properties of the coating upon treatment of the samples with an inhibitor and polymer.The form of the EIS spectra indicates the presence of two time constants responsible for the inner non-porous and the outer porous layers.In this work,we used a series-parallel equivalent electrical circuit(EEC),consisting of twoR-CPEchains.A detailed description of the EEC is presented in[14,16].Analysis of the calculated parameters of equivalent electrical circuits(Table 2)indicates a significan improve of the corrosion properties of composite coatings with the 8-HQ.The resistance of the outer(R1)and inner(R2)sublayers of the coating increased,while the capacities of these layers(Q1andQ2)decreased due to an increase in their thickness when the polymer and inhibitor were applied.The total resistance(R1+R2)for the investigated composite layers with 8-HQ increases from 1.5 to 27 times,depending on the type of applied polymer and the multiplicity of its processing.

Fig.6.Micro-Raman spectra acquired from the 8-HQ powder(upper curve)and from the surface of 8-HQ-containing PEO-coating(lower curve).(For interpretation of the references to colour in this figur legend,the reader is referred to the web version of this article.)

Table 2Calculated parameters of the EEC elements for MA8 magnesium alloy samples with various types of coatings formed on the basis of PEO-layer in the SiF electrolyte.

Fig.7.The optical image of the studied area of the inhibitor-containing PEO-layer(a,d);the corresponded 2D maps of the intensity distribution of 8-HQ over the surface of coating(b,e);overlaid optical images with 2D maps(c,f).Symbol“+”shows the point,in which the micro-Raman spectrum presented in Fig.6 was acquired.(For interpretation of the references to colour in this figur legend,the reader is referred to the web version of this article.)

Table 3Calculated parameters of the EEC elements for MA8 magnesium alloy samples with coatings formed on the basis of the PEO-layer obtained in the GP electrolyte.

The impedance spectra recorded for the coating formed in the GP electrolyte(Figs.10b,11)also indicate the positive combined effect of the inhibitor and polymer application,which improves the protective properties of the anticorrosive layers.8-HQ increases the corrosion resistance of the base PEO-coating by 4 times(before treatment:|Z|f=0.01Hz=1.3·104Ω·cm2;after treatment:4.7·104Ω·cm2),while the value of this parameter for polymercontaining layers increases from 6 to 1940 times(the values|Z|f=0.01Hzfor the corresponding coating without 8-HQ and impregnated with corrosion inhibitor are indicated in Table 3).It should be noted that|Z|f=0.01Hzfor a GP-coating with an inhibitor and a single SPTFE treatment is higher than the value of this parameter for an inhibitor free GP-coating with three times SPTFE treatment(Fig.11).This result was not established for the corresponding SiF-coatings,which is due to the high porosity of the protective layers formed in the GP electrolyte,which ensure more intensive coating impregnation with inhibitor.The morphology of these PEO-layers estimated using SEM/EDX analysis(Figs.2,3)confirme this suggestion.Considering that pores are the container for the inhibitor,the amount of the inhibitor in the protective layer after impregnation depends on the number and size of pores in the coating.

The EIS spectra for the samples with GP-coatings were described using the EEC indicated above.The result of impedance spectra modeling is presented in Table 3.The change in the main electrochemical parameters of the protective layers upon 8-HQ addition is consistent with the trend to anticorrosive properties increase and morphology evolution of the coating,which was previously established for the composite layers obtained on the basis of PEO-coating formed in the SiF electrolyte(Table 2).The addition of 8-HQ into the PEO matrix leads to an increase in the total resistance of the inner and outer layers of the coatings(R1+R2)from 4 to 2800 times.

Fig.8.The optical images of the scanned area as well as SVET/SIET maps of the local current density and local pH distribution over the surface of the SiF-coating after 1 h,20 h,24 h,and 48 h of sample exposure to 0.05 M NaCl solution.

The results of analysis of PDP curves(Figs.12,13)agree with the EIS measurements.The polarization curves of the samples with 8-HQ are in the zone of lower currents in comparison with curves of the samples without the inhibitor addition.Corrosion current density for coatings formed in SiF(Table 4)and GP(Table 5)electrolytes decrease 2-11 times and 4-4500 times,respectively.Similar to the results of electrochemical impedance spectroscopy,the most intense effect of the inhibitor is observed for coatings formed in the GP electrolyte due to the higher pore capacity of the layer.

According to the analysis of the obtained experimental data the composite polymer-containing coatings formed using PEO with subsequent impregnation with 8-HQ and three-fold treatment with SPTFE possessed the best protective properties among the studied samples.

XRD analysis of the samples was carried out before and after potentiodynamic polarization(Fig.14).The XRD diagram of samples before PDP test shows the presence of 8-HQ as one of the components of SiF-and GP-coatings,which confirm the successful protective layer impregnation with inhibitor.After PDP measurements the formation of magnesium 8-hydroxyquinolinate(MgQ2)was established in the composition of coatings obtained in SiF and GP electrolytes and treated with a corrosion inhibitor.This indicating the interaction of magnesium ions with 8-HQ,that improves the protective properties of the coatings in comparison with the samples without 8-HQ.The other compounds(MgO,Mg2SiO4,Na2MgSiO4,Ca10(PO4)6(OH)2))shown in the XRD diagram are responsible for the composition of the base PEO-coating,that is in agreement with SEM/EDX data(Figs.2-4).

Fig.9.The optical images of the scanned area as well as SVET/SIET maps of the local current density and local pH distribution over the surface of the GP-coating after 1 h,12 h,24 h,and 48 h of sample exposure to 0.05 M NaCl solution.

Thus,it was found that 8-HQ provides additional corrosion protection to MA8 magnesium alloy treated using PEO method.8-hydroxyquinoline forms a thin layer on the surface and in the pores of the PEO-coating,which are subsequently sealed with a polymer material.In case of mechanical damage or natural degradation of this composite layer the activation of the corrosion process is occurred.After that,due to the contact of the material substrate with an aggressive medium,a poorly soluble complex of magnesium 8-hydroxyquinolinate is formed(Fig.14).This confirm the results of works[5,14].The inhibitory effect is based on the suppression of the Mg dissolution in corrosive areas of the coated surface.The 8-HQ action is also associated with the adsorption of its molecules on the surface of the material,which prevents the adsorption of such aggressive ions as Cl-and OH-.

Fig.10.Impedance spectra(Nyquist and Bode plots)recorded in a 3 wt.% NaCl solution for MA8 magnesium alloy samples with PEO-coatings formed in the SiF(a)and GP(b)electrolytes:1-without 8-HQ,2-with 8-HQ.

Table 4Main electrochemical parameters of MA8 magnesium alloy samples with coatings formed on the basis of PEO-layer obtained in the SiF electrolyte.

Table 5Main electrochemical parameters of MA8 magnesium alloy samples with coatings formed on the basis of PEO-layer obtained in the GP electrolyte.

For an additional assessment of the active corrosion protection of the alloy the inhibitor efficien y(IE)was calculated according to the Eq.(3).

whereIc0andIcare the corrosion current density(A cm-2)obtained from the analysis of PDP curves for the coated samples without and with inhibitor,respectively.TheIEvalue is presented on the PDP diagrams,it is in the range of 57-99%for various coatings.The maximumIE,equal to 97% and 99%,was found for the coatings formed in a GP electrolyte and with subsequent one and threefold SPTFE treatment,respectively.The results obtained show the positive effect of the multiplicity of polymer treatment on the efficien y of the corrosion inhibitor for the coatings obtained on the base PEOlayer formed in GP electrolyte.As compared to the coatings formed in SiF electrolyte for which polymer treatment reduces the inhibitor efficien y,IEfor GP-coatings increases after polymer addition.This is the result of higher inhibitor amount in the GP-coating due to higher porosity of these layers as compared to SiF-ones.Therefore,the greater 8-HQ amount is presented both inside pores and in the polymer matrix due to the partial inhibitor dissolution during polymer treatment.However,for the SiF-coatings theIEvalues were still high after SPTFE and PVDF treatment.Table 6 shows the summarized data of the inhibitor efficien y for all systems of the studied samples.

Fig.11.Impedance spectra(Nyquist and Bode plots),recorded in a 3 wt.% NaCl solution for MA8 magnesium alloy samples with composite coatings formed on the basis of a PEO-layer obtained in SiF and GP electrolytes:1-single SPTFE treatment,2-coating impregnation with 8-HQ+single SPTFE treatment,3-triple SPTFE treatment,4-coating impregnation with 8-HQ+triple SPTFE treatment,5-PVDF treatment,6-coating impregnation with 8-HQ+PVDF treatment.Symbols indicate the experimental data,the line shows the fittin curve.

Table 6The values of the inhibitor efficien y(IE)calculated for MA8 magnesium alloy samples with protective coatings on the base of the data of PDP measurements in the 3 wt.% NaCl solution.

3.4.Mechanism of the composite coating degradation

The formed polymer-containing coatings with inhibitor addition provide the following mechanism of active corrosion protection of MA8 magnesium alloy at the moment of coating degradation when the electrolyte reaches the substrate surface and the corrosion process begins(Fig.15):

I-as a result of the cathodic half-reaction(2),the local pH changes(alkalization in the cathodic region due to the emission of OH-ions);

II-these changes increase the solubility of 8-HQ impregnated in the porous part of the PEO-layer,which leads to the release of the inhibitor from the reservoir-pore;

Fig.12.PDP curves recorded in a 3 wt.% NaCl solution for MA8 magnesium alloy samples with coatings formed on the basis of a PEO-layer in SiF electrolyte:a-base PEO-coating,b-PVDF treatment,c-single SPTFE treatment,d-triple SPTFE treatment;1-without 8-HQ addition,2-with 8-HQ.IE is the inhibitor efficien y.

Fig.13.PDP curves recorded in a 3 wt.% NaCl solution for MA8 magnesium alloy samples with coatings formed on the basis of a PEO-layer in GP electrolyte:a-base PEO-coating,b-PVDF treatment,c-single SPTFE treatment,d-triple SPTFE treatment;1-without 8-HQ addition,2-with 8-HQ.IE is the inhibitor efficien y.

Fig.14.XRD pattern of MA8 magnesium alloy samples with an inhibitor-containing coating obtained using the PEO method in SiF(a,c)and GP(b,d)electrolytes.The data were obtained before(a,b)and after(c,d)carrying out the potentiodynamic polarization of the samples in a 3 wt.% NaCl solution.

III-the inhibitor diffuses into the active area and forms a protective film

IV-the passive layer reduces corrosion activity,while the pH returns to neutral values(as before the corrosion process);

V-the solubility of the unreacted inhibitor decreases,which indicates its ability to further participate in the formation of the protective layer in case of repeated coating damage.

3.5.Evaluation of the antibacterial activity of inhibitor-containing coatings on MA8 alloy against the staphylococcus aureus strains

The antibacterial test of the inhibitor-containing coating was carried outin vitro.Two different experimental conditions of co-cultivation of samples with a suspension of bacteria in 1% TSB in 10 mM phosphate buffer solution and in the same solution with 10% blood serum addition were used.Blood serum was included in order to mimic theinvivocondition.Fig.16 shows the number of colonies(calculated according to the data of three experiments)after contact with the samples.Quantitative analysis of CFU of SA and MRSA bacteria after 24 h of incubation showed the antibacterial activity of the coating containing 8-hydroxyquinoline(PEO+8-HQ,Fig.16).After contact with PEO+8-HQ samples on agar,the minimum number of colonies was determined at all dilutions.Almost all SA and MRSA bacteria died on the inhibitor-containing coating of the PEO+8-HQ sample within 24 h,while upon contact with the bare Mg alloy(Mg)and alloy with PEO-coating(PEO)samples,a 100-fold increase in the CFU number was registered(Fig.17).Similar results were obtained when samples were co-cultured with a bacterial suspension in both abovementioned 1% TSB solutions.Therefore,we presented the experimental data only for a suspension of bacteria with 10% blood serum addition.

Among the number of tests available to measure the antibacterial activity of hard surfaces,only a few reflec the actual conditions under which bacteria can be in close contact with the potentially antibacterial surface of the implant.Testing conditions should mimic the scenario of infection during primary surgery or re-surgery after infection,when bacteria in the form of biofilm surround the implant[74].The analysis of the properties of the material should be carried out taking into account the number of bacteria initially adhered to the tested surface,as well as the decrease or increase in their number over time under conditions close to the tissue environment[75].In addition,the number of survivedS.aureuson the antibacterial surface should be assessed in comparison with samples without antibacterial activity.In our study,the antibacterial activity of coatings on magnesium alloy was evaluated using a model,which mimics the situation where infection occurs during primary surgery.In addition,the modifie assay allowed testing in the presence of biomatrix such as blood serum.The results showed that 8-hydroxyquinolinecontaining coating killed MRSA within 24 h.The antibacterial activity of this composite coating was not affected by the addition of 10% human blood serum to the culture medium,which suggests a similar potential effectin vivo.The antibacterial performance of such coatings should be attributed to the 8-HQ included in its composition,since other samples of bare Mg alloy and PEO coated alloy did not show antibacterial activity,as evidenced by a 100-fold increase in CFU after 24 h.

Fig.15.The mechanism of active corrosion protection of the MA8 magnesium alloy with polymer-inhibitor-containing coating at the moment of protective layer degradation.I,II,III,IV,V are the stages of the self-healing effect.

4.Conclusion

In the present study,the design and analysis of the smart composite antibacterial coatings with active corrosion protection of magnesium MA8 alloy(Mg-Mn-Ce system)were performed.Based on the obtained results,the following main conclusions can be drawn:

1.The relationship between the composition,structure,morphology,and properties of coatings on magnesium alloys with the composition of electrolytes and modes of plasma electrolytic oxidation was established using SEM/EDX analysis,optical microscopy,XRD,confocal micro-Raman spectroscopy,and electrochemical methods.

2.According to the data of local electrochemical methods(SVET/SIET)it was detected on a microscale the breakdown of the coating obtained by plasma electrolytic oxidation on the surface of Mg alloy,and the necessity of the protective layer modificatio was highlighted.

3.Based on the experimental results obtained using the conventional EIS and PDP techniques,a significan increase in the corrosion protection of Mg alloy was established as a result of composite coating formation.These layers were obtained on the basis of PEO method in two different electrolytes,followed by impregnation with corrosion inhibitor and subsequent polymer treatment.The presence of 8-HQ significantl retards the corrosion processes due to the penetration of the inhibitor to the pores of the PEOlayer and the formation of a thin adsorption protective fil on the surface of the material.The dissolution of the 8-HQ and release of the inhibitor occur as a result of local alkalization during the cathodic half-reaction of the magnesium alloy corrosion.The addition of an inhibitor to a coating system formed on the basis of a PEO-layer leads to a decrease in the corrosion current density from 2 to 4500 times as compared to the inhibitor-free coating system.

4.It was found that additional treatment of the coating with a polymer(SPTFE and PVDF)sufficientl improves the initial PEO-layer microstructure,while does not significantl change the activity of 8-HQ and leads to the formation of a reliable barrier preventing the rapid spontaneous release of the inhibitor that is not associated with the corrosion process.The inhibitor efficien y in the composition of such coatings can reach 99%.

5.A new method of active corrosion protection of magnesium alloys was developed.This method prevents the material from degradation even in the case of mechanical damage or destruction of the composite layer in an aggressive environment.Formed protective composite polymer-and inhibitor-containing coatings significantl reduce the corrosion current density of a magnesium alloy in a 3 wt.%NaCl solution(the maximum decrease is fi e orders of magnitude compared to the base PEO-layer).This indicates the prospect of formation a new smart coatings,which expand the fiel of application of electrochemically active materials.

6.PEO-layer impregnation with 8-HQ method results in the antibacterial coating formation on the Mg alloy.The antibacterial activity of the coatings againstS.aureusmethicillin-resistant strain was studiedin vitrousing the direct contact method,which was modifie to better reproduce the infection conditions during primary surgery.The developed inhibitor-containing coating shows the potential of magnesium alloys application in orthopedic surgery.These protective layers are also promising for reducing the incidence of implant-associated infections.

Fig.16.S.aureus colonies after the contact with Mg alloy with and without coatings after incubation for 24 h at different dilutions.Data for the sample without Mg alloy are presented for comparison.

Fig.17.The number of colony-forming units of S.aureus PS84(SA,a)and S.aureus methicillin-resistant(MRSA,b)strains at various dilutions after the contact with 8-HQ solution(8-HQ),with uncoated Mg alloy(Mg)and Mg alloy with protective layers:GP PEO-coating(PEO),GP PEO-coating impregnated with 8-hydroxyquinoline(PEO+8-HQ).Bacteria were removed from ultrasonicated nitrocellulose discs.The contact time was 24 h.

Acknowledgments

Local electrochemical measurements,coating formation and antibacterial assay were supported by the Grant of Russian Science Foundation,Russia(project no.21-73-10148,https://rscf.ru/en/project/21-73-10148/).The study of material‘s structure,composition,and corrosion processes kinetics was supported by the Grant of Russian Science Foundation,Russia(project no.20-13-00130,https://rscf.ru/en/project/20-13-00130/).Raman spectra were acquired under the government assignments from the Ministry of Science and Higher Education of the Russian Federation,Russia(project no.FWFN(0205)-2022-0003).