analysisoftitaniananosheetadsorptionbehaviorusinga...

Research ArticleAnalysis of Titania Nanosheet Adsorption Behavior Using aQuartz Crystal Microbalance Sensor

Yuichiro Tashiro1 Satoshi Komasa 1 Akiko Miyake 2 Hiroshi Nishizaki 1

and Joji Okazaki 1

1Department of Removable Prosthodontics and Occlusion Osaka Dental University 8-1 Kuzuhahanazono-cho Hirakata-shiOsaka Japan2Department of Oral Health Engineering Faculty of Health Sciences Osaka Dental University 1-4-4 Makino-honmachiHirakata-shi Osaka Japan

Correspondence should be addressed to Satoshi Komasa komasa-sccosaka-dentacjp

Received 17 August 2017 Accepted 29 November 2017 Published 19 February 2018

Academic Editor Marco Cannas

Copyright copy 2018 Yuichiro Tashiro et al is is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work isproperly cited

We investigated the adsorption of albumin and fibronectin on a titania nanosheet- (TNS-) modified quartz crystal microbalance(QCM) sensor A Ti QCM sensor was fabricated by reactive magnetron sputtering A thin layer of Ti was deposited on the QCMsensor is sensor was then alkali-modified by treatment with NaOH at room temperature to fabricate the titania nanosheetsScanning probe microscopy X-ray photoelectron spectroscopy and scanning electron microscopy were performed to investigatethe surface topology and chemical components of each sensor e TNS had a titanium oxide film exhibiting a nodular structureand a thickness of 13 nm on the QCM sensor Furthermore QCMmeasurements showed significantly greater amounts of albuminand fibronectin adsorbed on the TNS than on titanium e NaOH treatment of titanium modified the sensor surface andimproved the adsorption behaviors of proteins related to the initial adhesion of bone marrow cells erefore we concluded thatTNS improves the initial adhesion between the implant materials and the surrounding tissues

1 Introduction

Titanium is a biocompatible material that is commonlyused in dentistry and orthopedic reconstruction esurface features of implant materials have importantfunctions in cell or extracellular matrix interrelationshipsand eventual osseointegration erefore interrelation-ships between cells and microtopography have been in-tensively studied

An important contemporary advance in dental implantresearch is the ability to modify implant surface materials at thenanoscale [1 2] Materials with an expanded surface region anda better surface roughness may yield better mechanical inter-locking between tissues and titanium [3] However moreimportantly such nanoscale features are also believed to directlyaffect osteogenic cell behaviors around implant fixtures withnonconventional surfaces creating a biomimetic relationshipbetween alloplastic surfaces and host tissues by the replication ofthe natural cellular environment at the nanometer level [1 2 4]

Low-dimensional TiO2 nanostructures have attractedrecent attention because these materials can take the formsof nanotubes [5] nanofibers [6] and nanowires [7] Com-pared with mass materials or nanoparticles TiO2 nanotubeshave high particular surface zones accessible for the ad-sorption of color sensitizers and they provide channels toimprove electron exchange thus expanding the effectivenessof solar cells [5] Titania nanosheets (TNSs) are similar toTiO2 nanotubes created by titania deposition using thesputtering process [5]

In our previous study [8] we showed that TNSs createdby compound processing enhance the osteogenic separationof rodent bone marrow (RBM) cells e surface propertiesand structural characteristics of materials play an importantrole in protein and cell adsorption behaviors e initialamount adsorbed and the conformation of proteins con-tained in the serum could alter the bioactivity of stem cellson TNSs However the bioactivities of TNS materials in-cluding their roles in osteogenic differentiation and the

HindawiAdvances in Materials Science and EngineeringVolume 2018 Article ID 7461245 10 pageshttpsdoiorg10115520187461245

biointegration of dental implants into the alveolar bonehave not been elucidated

A quartz crystal microbalance (QCM) sensor is a pro-foundly delicate and handy device that is used to observeprotein adsorption and cell behavior in situ A QCM-basedsensor comprises a quartz crystal and a detection material A27MHz QCM can provide highly sensitive measurements ofmass in aqueous solutions the resonance frequency de-creases in relation to the mass of the protein bound on theQCM electrode surface We previously fabricated severaltypes of QCM sensors by coating the gold electrode of thequartz crystal with a thin film of a biomaterial [9] this QCMsensor achieved an increase in sensitivity approximately 24times that of a conventional 5MHz QCM PMMA (poly(methyl methacrylate)) Au and Ti have been used as QCMelectrode surface materials to imitate denture materials andevaluate the adsorption behaviors of various bovine salivaryproteins [10] ese previous findings support the potentialutility of the QCM method for the evaluation of proteinadsorption behaviors on implant surfaces

In this study we evaluated the effects of modified sur-faces on the adsorption of albumin and fibronectin in RBMcells and simulated body fluid (SBF) in QCM analyses

2 Materials and Methods

21 Sputtering Procedure A thin layer of Ti was depositedon quartz discs (diameter 8mm area 49mm2) by reactivemagnetron sputtering using a radio-frequency magnetronsputtering system (CFS-4ES-231 Shibaura MechatronicsCo Ltd Kanagawa Japan) e QCM crystal was cleanedusing piranha solution (H2SO430H2O2 (vv) 7 3) Beforedeposition was conducted the quartz surfaces were ultra-sonically cleaned in high-purity acetone (99999) Pure Tipowder was used to prepare the target which was formed bypressing the powder into a disc with a diameter of 75mme quartz substrates were positioned 85mm above thetarget and themagnetron sputtering chamber was evacuated toa pressure of 3times10minus3 Pa Argon was used as the working gasand its pressure was kept constant at 67times10minus1 Pa All of thefilms were fabricated using a constant radio-frequency dis-charge power of 480W and the Ti thin films were deposited atroom temperature at a deposition rate of 200 nmmin yieldinga film thickness of approximately 240nm e crystals werewashed and cleaned with both sodium dodecyl sulfate and UV-Ozone Cleaner (PC450 Meiwafosis Co Ltd Osaka Japan)prior to QCM measurements

22 Sample Preparation In the TNS group Ti sensors weretreated to produce TNS on their surfaces An unprocessedQCM sensor was used as the Ti sensor ese sensors wereimmersed in 10MNaOH (aq) and were then placed in an oilbath which was kept at a temperature of 30degC for 24 h esolution in each flask was replaced and treated with distilledwater (200mL) and this procedure was repeated until thepoint that a conductivity of 5 μScm3 was reached especimens were then dried at room temperature

23 Characterization of Materials Scanning electron mi-croscopy (S-4800 Shimadzu Kyoto Japan) and scanningprobe microscopy (SPM-9600 Shimadzu) over a surfacearea of 20 μmtimes 20 μm were conducted to observe thesurface topology and roughness of the fabricated TNS and Tisensors e composition of the coating was analyzed byX-ray photoelectron spectroscopy (XPS ESCA 5600 Ulvac-Phi Inc Kanagawa Japan) using surface etching withionized argon In addition the surfaces of the fabricated Tisensors were subjected to XPS analysis with an Al Kα line(15 kV 300W) as an X-ray source During XPS argon ionsputtering was applied to determine the thickness andstructure of the surface layers

24 Contact Angle Measurements Contact angles weremeasured for the TNS and Ti sensors using a video contactangle measurement system (model VSA 2500 XE ASTProducts Inc Billerica MA USA) A small droplet ofa deionized water solution with Hanksrsquo Balanced Salt So-lution and bovine serum albumin (BSA approximately3mg) was placed on the TNS to measure the static contactangle Estimation of the contact edge is a straightforwardstrategy for breaking down the vitality and hydrophilicnature of a surface

25 Proteins BSA (Wako Pure Chemical Industries LtdOsaka Japan) was dissolved in phosphate-buffered saline(PBS pH 74) at 200 μgmL Human plasma fibronectin(HFN Nacalai Tesque Inc Kyoto Japan) was dissolved inPBS (pH 74) at 500 μgmL

26 Cell Culture Since most bone embed materials areembedded in adult bone that is in direct contact with bonemarrow tissue the effects and success of new embed ma-terials can be investigated by examining bone marrow cellcultures from adult rats RBM cells multiply and separateinto a phenotype that expresses bone cell markers in vitroRBM cells were extracted from the femurs of 7-week-oldSpraguendashDawley rats e rats were humanely sacrificedutilizing 4 isoflurane and the bones were asepticallyextracted from the hind limbs e external soft tissues werediscarded and the extracted bone samples were immersed in50mL of Eaglersquos minimal essential medium (EMEM Wako)supplemented with 20 fetal bovine serum (lot number1412447 Invitrogen Life Technologies Corp CarlsbadCA USA) and penicillin (850UmL) for approximately15min

e proximal end of the femur and the distal end of thetibia were cut An 18-gauge needle (TERMO Japan) wasintroduced into the opening at the knee-joint end of eachbone and the marrow was washed out of the bone shaft byEMEM e obtained marrow pellet was separated bytrituration and the cell suspensions obtained from all thebones were combined by centrifugation RBM cells werecultured in 75 cm2 culture flasks (TD75 Falcon) in EMEMAt confluence the cells were removed by trypsinizationwashed twice in EMEM resuspended in culture medium

2 Advances in Materials Science and Engineering

and seeded on test and control titanium disks at a concen-tration of 4times104 cellscm2 in 24-well tissue culture platese cells were incubated for 3 days in a CO2 incubator at37degC is investigation was conducted in accordance withthe Guidelines for Animal Experimentation of Osaka DentalUniversity (Approval no 16-08001)

27QCMMeasurements e amounts of proteins (BSA andHFN) and RBM cells were determined by QCM measure-ments (Affinix QN μ Initium Co Ltd Tokyo Japan)Affinix QN μ had a 550 microL cell outfitted with a 27MHz QCMplate at the base of the cell e diameter of the quartz platewas 8mm and the area of the gold-plated quartz was49mm2 e unit also included a mixing bar and a tem-perature controller e adjustment in recurrence waschecked utilizing a universal frequency counter connectedto a microcomputer

e Ti QCM sensors and TNS were immersed in 500 μLof PBS (001M PBS at pH 74) Changes in the QCM fre-quency were measured as a function of time recordingstarted immediately after the infusion of 5 μL (20 μgmL) ofBSA HFN and RBM cells e solution was mixed to avoidany influence of protein dispersion on the measured resultsStirring did not influence the soundness of the frequency orthe degree of frequency adjustment e frequency changerelied upon the adsorbed mass in accordance with theSauerbrey equation

ΔF minus2F2

0ΔmA

ρqμq

1113968 (1)

As per this condition at 27MHz a frequency shift of1Hz relates to a mass difference in roughly 062 ngmiddotcmminus2 Inthe Sauerbrey equation F0 is the fundamental frequency ofthe quartz crystal (27times106Hz) ΔF is the measured fre-quency shift (Hz) ρq is the density of quartz (265 gmiddotcmminus3)Δm is the mass change (g) A is the electrode area(0049 cm2) and μq is the shear modulus of quartz(295times1011 dynmiddotcmminus2) QCM observation was performed at25degC and the test was repeated four times Results are depictedas meanplusmn standard deviation

28 XPS Analysis after Measuring the Adsorption of BSA andHFN Using Ti and TNS QCM Sensors e biochemicalconstituents of the adsorbed protein films on the QCMsensors were investigated by XPS on an AXIS Ultra DLDspectrometer (Kratos Instruments Manchester UK)equipped with a monochromated Al Kα X-ray source(hv 14866 eV) operated at 75W XPS was utilized to in-vestigate the proteinaceous carbon (C1s) and nitrogen (N1s)signals produced by the protein Evaluation of the C1s andN1s signals emerging from the peptide bonds of the proteinwas conducted to determine the relative amount of proteinadsorbed on various surfaces

29 Preparation of SBF Since SBF is supersaturated withrespect to apatite improper planning can prompt the pre-cipitation of apatite in the solution erefore the solution

should remain colorless and transparent and there shouldnot be any deposition on the inner surface of the containerIf precipitation is observed the preparation of SBF should behalted and the procedure should be restarted at the step ofwashing the apparatus

For preparation of 1 L of SBF 700mL of ion-exchangedand distilled water was added to a 1 L plastic beaker witha stir bar e beaker was covered with plastic wrap and thewater was heated to 365plusmn 15degC under stirring Reagents 1ndash8were then dissolved into the solution in order (given in Table 1)at 365plusmn 15degC and reagents 9 (Tris) and 10 (HCl) were addedafter pH adjustment

Note that during this procedure instead of glass con-tainers plastic containers with smooth unscratched surfaceswere used because apatite nucleation can be induced on thesurfaces of glass containers or the edges of scratches Ad-ditionally reagents were dissolved completely before addi-tion of the next reagent e volume of 1M HCl wasmeasured using a cylinder after the cylinder had beenwashed with 1M HCl Finally hygroscopic reagents such asKCl K2HPO4middot3H2O MgCl2middot6H2O CaCl2 and Na2SO4 weremeasured as quickly as possible

e temperature of the solution was set to 365plusmn 15degC Ifthe amount of solution was less than 900mL ion-exchangedand distilled water was added to increase the volume to900mLe pH of the solution was then determined BeforeTris was added to the solution and dissolved the pH of thesolution was 20plusmn 10 e solution temperature was keptwithin the range of 35ndash38degC (optimal 365plusmn 05degC) and theadded Tris was dissolved slowly while changes in pH werenoted Tris was added until the pH reached approximately 745

3 Results

31 Scanning Electron Microscopy (SEM) and SPMAnalysis SEM images are shown in Figure 1 After modi-fication in NaOH at 30degC the TNS sensor surfaces showeda nanoscale network structure SPMwas utilized to gauge thedepth of the surface characteristics of the specimens and thesurface morphologies of the TNS and Ti sensors are shownin Figure 2 Many nanonodules were detected on the TNSsurface these formations had horizontal dimensions ofapproximately 300 nm e surface roughness (Ra) valueswere 39 and 189 nm for the Ti and TNS sensors respectively

32XPSAnalysis Figure 3 shows the results of wide-scanXPSsurface chemical analyses of Ti and TNS QCM sensors epresence of Ti O C andNwas confirmed on the surfaces of Tiand TNS QCM sensors In addition the presence of Na wasconfirmed on the surface of the TNS QCM sensor Moreoverthe Ti and O concentrations of TNS QCM sensors were higherthan those on the surface of TNS QCM sensors

33 Surface Wettability e contact angle for a water dropon the Ti sensor was 425deg However the water drops spreadvery rapidly as they reached the test sensor and the contactangle could not be determined However the behavior on

Advances in Materials Science and Engineering 3

the TNS sensor indicated that the sensor was super-hydrophilic (contact angle of less than 5deg)

34 QCM Measurement of Proteins Figure 4 shows theadsorption of albumin and fibronectin based on QCMmeasurements An immediate decrease in frequency wasobserved after the injection of albumin and fibronectin is

decrease in frequency was identified with the adsorption ofalbumin and fibronectin e adsorption of albumin andfibronectin on the TNS sensor produced a decrease infrequency that was greater than that measured for the Tisensor According to Sauerbreyrsquos equation at 27MHza frequency decrease of 1Hz compares to a mass differenceof around 062 ngcm2 [10] After 30min the amounts ofalbumin adsorbed on the TNS and Ti sensors were 1878 and

Table 1 Order amounts weighing containers purities and formula weights of reagents for the preparation of 1 L of SBF

Order Reagent Amount Container Purity () Formula weight1 NaCl 8035 g Weighing paper 995 5844302 NaHCO3 0355 g Weighing paper 995 8400683 KCl 0225 g Weighing bottle 995 7455154 K2HPO4middot3H2O 0231 g Weighing bottle 990 22822205 MgCl2middot6H2O 0311 g Weighing bottle 980 20330346 10M HCl 39mL Graduated cylinder mdash mdash7 CaCl2 0292 g Weighing bottle 950 11098488 Na2SO4 0072 g Weighing bottle 990 14204289 Tris 6118 g Weighing paper 990 121135610 10M HCl 0ndash5mL Syringe mdash mdash

(a) (b)

Figure 1 SEM images of (a) Ti and (b) TNS QCM sensors

(a) (b)

Figure 2 SPM images of (a) Ti and (b) TNS QCM sensors


1175 ngcm2 respectively and the amounts of bronectinadsorbed on the TNS and Ti sensors were 8276 and5337 ngcm2 respectively

35 QCM Measurement of RBM Cells Figure 5 shows theadsorption of rat bone marrow cells based on QCM mea-surements An immediate decrease in frequency was observedafter the injection of RBM cellse adsorption of RBM cells onthe TNS sensor produced a decrease in frequency that wasgreater than that measured for the Ti sensor Using Sauerbreyrsquosequation after 30min the amounts of albumin adsorbed on theTNS and Ti sensors were 14911 and 8725ngcm2 respectively

36 XPS Analysis after Adsorption of Proteins e XPSspectra of the Ti and TNS QCM sensors after immersion inBSA are summarized in Figures 6 and 7 and those of the Tiand TNS QCM sensors after immersion in HFN are shownin Figures 8 and 9 For XPS analysis the N1s and C1s spectrawere measurede coupling energies (BEs) of the C1s rangefor adsorbed BSA were 2848 eV (C-CC-H) 2864 eV(C-OC-N) and 2882 eV (OC-O) for the Ti QCM sensorsand 2848 eV (C-CC-H) 2864 eV (C-OC-N) and 2886 eV(OC-O) for the TNS QCM sensors ere were N1s peaks

for the Ti and TNS QCM sensors after the adsorption ofBSA e BEs of the C1s spectrum for adsorbed HFN were2848 eV (C-CC-H) 2864 eV (C-OC-N) and 2880 eV(OC-O) in the Ti QCM sensors and 2847 eV (C-CC-H)2864 eV (C-OC-N) and 2880 eV (OC-O) for the TNSQCM sensors After adsorption of the HFN there were alsoN1s center-level spectra for the Ti and TNS QCM sensors

N1s

Na1s

OlsCls

Ti2p

(a)

Na1sN1s

O1s

C1sTi2p

(b)

Figure 3 XPS analysis of (a) Ti and (b) TNS QCM sensors

0

50

100

150

200

250

Ti TNS

ltBSAgt lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

plusmn1 SDlowastp lt 005

(a)

lowast

0

200

400

600

800

1000

1200

Ti TNS

ltHFNgt

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(b)

Figure 4 Adsorption of two proteins on Ti and TNS QCM sensors (a) BSA (b) HFN


0200400600800

10001200140016001800

lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS

Figure 5 Adsorption of rat bone marrow cells on Ti and TNSQCM sensors


37Characterizationafter Immersion inSBF Figure 10 showsthe adsorption of apatite after exposure to the SBF solutionbased on QCMmeasurements A quick frequency reduction

was seen after the infusion of SBF e adsorption of SBF onthe TNS sensor produced a decrease in the frequency thatwas greater than that measured for the Ti sensor

275280285290295300

C=O

Binding energy (eV)

C-H C-C

C-O C-N

After adsorption of BSA

Nontreatment

(a)

C=O

275280285290295300Binding energy (eV)

C-H C-C

C-O C-N


Nontreatment

(b)

Figure 6 C1s XPS spectra of (a) Ti and (b) TNS QCM sensors after immersion in BSA

385395405415 410 400 390Binding energy (eV)


Nontreatment

(a)



Nontreatment

(b)

Figure 7 N1s XPS spectra of (a) Ti and (b) TNS QCM sensors after immersion in BSA

C=O

Binding energy (eV)

After adsorption of HFN

Nontreatment

C-H C-C

C-O C-N

280290300 295 285 275

(a)

C=O



Nontreatment

C-H C-C

C-O C-N

(b)

Figure 8 C1s XPS spectra of (a) Ti and (b) TNS QCM sensors after immersion in HFN


Figure 11 shows the surface morphology of the Ti andTNS QCM sensors after immersion in SBF for 24 h Afterimmersion a recently framed layer was seen on the Ti andTNS surfacese surface was covered by single and groupedball-like particles with a size of around 15 μm e numberof particles on the surface of the TNS QCM sensor wasclearly greater than that on the surface of the Ti QCM sensor

4 Discussion

In this study Ti sensors were realized by depositing a thin Tilm on a QCM electrode using a reactive DC magnetronsputtering technique this sensor was further alkali modiedusing NaOH treatment to fabricate the TNS SPM and XPSanalyses were carried out to characterize the nanostructureof the TNSs We evaluated the quality of the TNSs and thepotential application of the high-frequency TNS sensors byinvestigating the in situ binding behaviors for TNS sensorstwo proteins RBM cells and SBF to determine the ecentects ofthese parameters on biological reactions in solution

Several studies have demonstrated that implant surfacesacentect nanoscale topography and thereby alter cell behaviorsor change the nanofeatures of structures to improve theosseointegration process [4 11 12] e embed surface canbe adjusted by various approaches to add nanoscale featuresto the surfaces in specic combinations e most well-known techniques are chemical processes such as alkalinehydrothermal [13 14] or acid [15 16] oxidation on titaniumsurfaces to produce diverse nanoscale topographies Kasugaet al [5] demonstrated that TiO2 nanotubes with a diameterof about 8 nm and a length of about 100 nm could be formedby Ti treatment with a 10MNaOH aqueous solution for 20 hat 110degC without the need for templates or replicationEssential factors in regulating cell reactions at the implant-tissue interface can dramatically acentect tissue coordination[17] In a recent work we demonstrated that TiO2 nanotubesand TNSs could be formed on titanium metal surfaces bytreatment with a 10M NaOH aqueous solution at 30degC andwe used this method to prepare TNS-modied disksKomasa et al [8] suggested that TNSs on titanium surfacescan be applied to control the osteogenic dicenterentiation ofbone marrow cells and enhance mineralization Our results

demonstrated that TNS-modied titanium disks were morehydrophilic and showed uniquely enhanced wettability incomparison with unmodied disks Further studies of thesurface roughness and topography of modied titaniumalloy surfaces are needed to assess their wettability Ra isa commonly used height parameter to describe implantsurface roughness Ra of the TNS-modied titanium surfacewas 19 nm which was greater than that of the untreatedtitanium surface e contact angles of the alkali-treatedtitanium disks gradually decreased in comparison with thoseof the control group indicating that the wettability of thesurface of the test group was reduced by NaOH treatment Inprevious studies a surface roughness of between 13 and16 nm was found to be optimal for RBM cell culture [18 19]e nanonetwork structure framed on the titanium diskshere was like the hierarchical structure outlined by Zhaoet al [12] In their work hierarchical nanotextured titaniumsurface topographies with TNS structures that mirrored thehierarchical structures of bone tissues were created byetching followed by anodization Natural tissues are hier-archical structures of nanoscale building blocks organized ina structured way Hierarchical structures composed ofnanocomponents may give a more reasonable surface to-pography for bone marrow cell functions than simplerstructures because they can better copy the structures ofnatural tissues Our research revealed that NaOH treatmentprompts the development of a Ti-O-Na titanate layer on thetitanium surface us we expect that NaOH treatment


Nontreatment


(a)

385390395400405410415

Nontreatment


Binding energy (eV)

(b)

Figure 9 N1s XPS spectra of (a) Ti and (b) TNS QCM sensors after immersion in HFN


01000020000300004000050000600007000080000

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS

lowast

Figure 10 Adsorption of apatite on Ti and TNS QCM sensors


results in the formation of a thick oxide film on the TiO2layer on the titanium surface e deconvolution proceduresuggested that this may have been due to surface contam-ination resulting from the binding of O to C [20]

Before argon ion etching was performed the XPS depthprofiles perpendicular to the surface of the sample showeda carbon contamination overlayer on the surface iscarbon contaminant was removed after the initial etchingcycles e oxygen and Ti concentrations gradually de-creased and increased respectively as the number of etchingcycles increased Kim et al [21] demonstrated that NaOHand heat treatment resulted in the covering of the Ti sub-strate with a titanium oxide layer with a thickness of ap-proximately 1000 nm AES depth profiles of the Ti oxidelayer indicated the presence of an amorphous sodium ti-tanate hydrogel layer Kasuga et al [5] likewise showed thatNaOH treatment prompted the formation of a Ti-O-Natitanate layer on the Ti surface At a greater depth of ap-proximately 250 nm the oxygen concentrations remainedalmost constant at 5 atom In addition the remainingoxygen content was approximately 5 atom inside the ti-tanium film which was attributed to the presence of residualoxygen in the chamber after preparation of the Ti sample

All embed surfaces are promptly covered with a layer ofprotein from the in vitro culture medium or in vivo bi-ological fluids and this interface regulates the course of cellreactions and behaviors [22] To elucidate the connectionbetween the Ti implant surface properties opsonization andphagocytosis under in vivo conditions phagocytic experi-ments were previously conducted using a cell culture me-dium supplemented with serum albumin and humanopsonizing serum factors [23] Fibronectin plays a crucialrole in the progressive differentiation of osteoblasts [24]Additionally fibronectin has RGD sequences and is a largeextracellular matrix dimer glycoprotein [25] with a molec-ular weight of approximately 440 kDa [26] Albumin is themost abundant plasma protein it suppresses the adsorptionof other proteins that may empower aggravation and bac-terial colonization [27] e molecular weight of BSA isapproximately 6-7 kDa [28] In this study the amount offibronectin adsorbed was greater than the amount of al-bumin adsorbed suggesting that adsorption quantity wasrelated to the molecular weight of the injection solutions

After adsorption of BSA and BSF N1s core-level spectrawere obtained for both QCM sensors Endo [29] detected CO and N obtained from the organic material or metal oxideon the titanium surface by XPS investigation In the presentstudy based on XPS the C1s and N1s peaks derived fromBSA and BSF were detected on both QCM sensor surfacestested Evaluation of the N1s peak emerging from thepeptide bonds of the implant-bone bonding protein may bea successful measure of the relative amount of proteinadsorbed onto implant surfaces on the TNS QCM sensor

Our results are the first comparison of RBM cell pro-liferation on TNS-modified titanium surfaces and un-processed controls Surfaces with nanostructures havehigher surface areas than those of surfaces without suchfeatures [11 30] is expanded surface territory permitsincreased adhesion of cells such as osteoblasts and fibro-blasts us the altered surface energies of materials withnanostructures may promote tissue growth via increasedadsorption of select proteins compared with materials withmicroscale features [31] e adsorption of select proteinscan in this manner guide the adhesion of cells on the implantmaterial surface among other capabilities Various in-vestigations have exhibited improved cell adhesion andmultiplication on nanostructured surfaces with variouspotential tissue applications including applications in thebladder bone vasculature and nervous system [32ndash34] Aprior study suggested that TNSs on titanium surfaces fa-cilitate the regulation of osteoblastic differentiation of bonemarrow cells and enhance mineralization e current studyshowed that the TNSs formed nanonodules with a diameterof about 19 nm on the titanium surfaces and these structurespromoted the adhesion andor multiplication of cells enetwork structure of TNSs on titanium alloy facilitated rapidcell adhesion spreading and multiplication due to themechanics of the TNS structure and chemical nature of theTi-O-Na layer

To enhance bone-titanium bonding Kokubo et al re-cently demonstrated that a blend of alkalis resulted in theformation of bone-like apatite on the surface of titanium inSBF with an ion concentration almost equivalent to that ofhuman blood [35] Apatite development on the materialsurface is accepted to be essential for bioactivity that isdirect bone bonding In our study there was an apatite layer

(a) (b)

Figure 11 SEM images of exposure to SBF solution on (a) Ti and (b) TNS QCM sensors


on the TNSQCMsensor after 24hus the increased adhesionof RBM cells and SBF on the TNS QCM sensor suggested thatTNS induced bone differentiation

In the TNS sensor the concentration of Ti exceeded thatof oxygen at a depth of approximately 170 nm Subsequentlythe NaOH treatment was thought to produce a thick oxidefilm Nonetheless the chemical structure of the treatedsurfaces did not vary fundamentally Ti oxides (mostly TiO2)formed at the surface Albumin and fibronectin were readilyadsorbed to a greater extent on the TNS sensor than on thereference Ti sensor e TNS surface seemed to adsorb moreprotein for a given geometric surface area than the referenceTi sensor this may have been a result of its rough mor-phology as evidenced by the SEM and SPM results Severalstudies [36 37] have shown that nanostructured topogra-phies can act as good mimics of natural extracellular ma-trixes Advancement of the surface topography could beindirect the adsorption of proteins or ions may function asan extension between the nanosurface structure and cells[38] Webster et al [39] observed increases in the adsorptionof vitronectin on nanostructured surfaces (compared withconventional surfaces) which resulted in preferable adhe-sion of osteoblasts In addition here the protein adsorptionrate on the TNSs was correlated with the contact anglesuggesting that the hydrophilicity of titanium greatly af-fected its protein adsorption ability e contact angle of theTNS and Ti sensors fluctuated in the hydrophilic range thehydrophilicity of the surfaces expanded after treatmentcorresponding to the formation of the TNSs us thegrowth of the titanium oxide layer increased the surfaceenergy resulting in a more hydrophilic surface [40]

5 Conclusions

In conclusion TNS structures were obtained on a titaniumsurface via treatment with a NaOH aqueous solution at roomtemperature Nanoscale network structures and a largenumber of nanoscale nodules were observed by SEM andSPM In addition the chemical composition of the TNSstructures was estimated by XPS e results confirmed thepresence of a combined titanium and oxide titanate layerwhich induced the adsorption of albumin and fibronectin Inthe fields of tissue engineering and biomaterials nano-structuring technologies are expected to yield novel bi-ologically optimized surfaces

Conflicts of Interest

e authors declare that they have no conflicts of interestregarding the publication of this article

Authorsrsquo Contributions

Satoshi Komasa conceived and designed the experimentsYuichiro Tashiro performed the experiments YuichiroTashiro Satoshi Komasa and Akiko Miyake analyzed thedata Hiroshi Nishizaki and Joji Okazaki contributed re-agents materials and analysis tools Yuichiro Tashiro andSatoshi Komasa wrote the paper

Acknowledgments

e authors wish to express their thanks to Tohru Sekinofrom Osaka University for setting up the nanosheets and forhis supportive recommendations e authors are alsograteful to the members of the Department of RemovableProsthodontics and Occlusion and the Department of OralHealth Engineering for their advice and assistance eauthors also thank Toshio Tamaki and Hirokazu Hojyoiswork was supported by grants from the Japan Society for thePromotion of Science (16K20524)

References

[1] M Annunziata A Oliva A Buosciolo M Giordano A Guidaand L Guida ldquoBone marrowmesenchymal stem cell response tonano-structured oxidized and turned titanium surfacesrdquo ClinicalOral Implants Research vol 23 no 6 pp 733ndash740 2012

[2] G Mendonca D Mendonca F J L Aragao and L F CooperldquoAdvancing dental implant surface technologyndashFrommicron-to nanotopographyrdquo Biomaterials vol 29 no 28 pp 3822ndash3835 2008

[3] L Meirelles F Currie M Jacobsson T Albrektsson andAWennerberg ldquoe effect of chemical and nanotopographicalmodifications on the early stages of osseointegrationrdquo In-ternational Journal of Oral and Maxillofacial Implants vol 23no 4 p 641 2008

[4] K Kubo N Tsukimura F Iwasa et al ldquoCellular behavior onTiO2 nanonodular structures in amicro-to-nanoscale hierarchymodelrdquo Biomaterials vol 30 no 29 pp 5319ndash5329 2009

[5] T Kasuga M Hiramatsu A Hoson T Sekino and K NiiharaldquoTitania nanotubes prepared by chemical processingrdquoAdvancedMaterials vol 11 no 15 pp 1307ndash1311 1999

[6] X Gao H Zhu G Pan et al ldquoPreparation and electro-chemical characterization of anatase nanorods for lithium-inserting electrode materialrdquo Journal of Physical Chemistry Bvol 108 no 9 pp 2868ndash2872 2004

[7] A R Armstrong G Armstrong J Canales and P G BruceldquoTiO2 nanowiresrdquo Angewandte Chemie International vol 43no 17 pp 2286ndash2288 2004

[8] S Komasa Y Taguchi H Nishida M Tanaka andT Kawazoe ldquoBioactivity of nanostructure on titanium surfacemodified by chemical processing at room temperaturerdquoJournal of Prosthodontic Research vol 56 no 3 pp 170ndash1772012

[9] Y Hashimoto S Minoura A Nishiura et al ldquoDevelopmentof titanium quartz crystal microbalance sensor by magnetronsputteringrdquo Journal of Oral Tissue Engineering vol 8pp 52ndash59 2010

[10] A Miyake S Komasa Y Hashimoto Y Komasa andJ Okazaki ldquoAdsorption of saliva related protein on denturematerials an X-ray photoelectron spectroscopy and quartzcrystal microbalance studyrdquo Advances in Materials Scienceand Engineering vol 2016 Article ID 5478326 9 pages 2016

[11] G Mendonca D B Mendonca F J Aragao and L F Cooperldquoe combination of micron and nanotopography byH(2)SO(4)H(2)O(2) treatment and its effects on osteoblast-specificgene expression of hMSCsrdquo Journal of Biomedical MaterialsResearch A vol 94 no 1 pp 169ndash179 2010

[12] L Zhao S Mei P K Chu Y Zhang and Z Wu ldquoe in-fluence of hierarchical hybrid micronano-textured titaniumsurface with titania nanotubes on osteoblast functionsrdquoBiomaterials vol 31 no 19 pp 5072ndash5082 2010


[13] W Zhang Z Li Y Liu et al ldquoBiofunctionalization of a titaniumsurface with a nano-sawtooth structure regulates the behavior ofrat bonemarrowmesenchymal stem cellsrdquo International Journalof Nanomedicine vol 7 pp 4459ndash4472 2012

[14] W Dong T Zhang J Epstein et al ldquoMultifunctionalnanowire bioscaffolds on titaniumrdquo Chemistry of Materialsvol 19 no 18 pp 4454ndash4459 2007

[15] S Bauer J Park K von derMark and P Schmuki ldquoImprovedattachment of mesenchymal stem cells on super-hydrophobicTiO2 nanotubesrdquo Acta Biomaterials vol 4 no 5 pp 1576ndash1582 2008

[16] E Hosono H Matsuda I Honma M Ichihara and H ZhouldquoSynthesis of a perpendicular TiO2 nanosheet film with thesuperhydrophilic property without UV irradiationrdquo Langmuirvol 23 no 14 pp 7447ndash7450 2007

[17] T Albrektsson P I BranemarkHAHansson and J LindstromldquoOsseointegrated titanium implants requirements for ensuringa long-lasting direct bone-to-implant anchorage in manrdquo ActaOrthopaedica vol 52 no 2 pp 155ndash170 1981

[18] H Xing S Komasa Y Taguchi T Sekino and J OkazakildquoOsteogenic activity of titanium surfaces with nanonetworkstructuresrdquo International Journal of Nanomedicine vol 9p 1741 2014

[19] T Fujino Y Taguchi S Komasa T Sekino and M TanakaldquoCell differentiation on nanoscale features of a titaniumsurface effects of deposition time in NaOH solutionrdquo Journalof Hard Tissue Biology vol 23 no 1 pp 63ndash70 2014

[20] M Pisarek A Roguska M Andrzejczuk et al ldquoEffect of two-step functionalization of Ti by chemical processes on proteinadsorptionrdquo Applied Surface Science vol 257 no 19pp 8196ndash8204 2011

[21] H M Kim F Miyaji T Kokubo S Nishiguchi andT Nakamura ldquoGraded surface structure of bioactive titaniumprepared by chemical treatmentrdquo Journal of BiomedicalMaterials Research vol 45 pp 100ndash107 1999

[22] M Roser D Fischer and T Kissel ldquoSurface-modified bio-degradable albumin nano-and microspheres II effect ofsurface charges on in vitro phagocytosis and biodistribution inratsrdquo European Journal of Pharmaceutics and Biopharmaceuticsvol 46 no 3 pp 255ndash263 1998

[23] P Roach D Farrar and C C Perry ldquoInterpretation of proteinadsorption surface-induced conformational changesrdquo Jour-nal of the American Chemical Society vol 127 no 22pp 8168ndash8173 2005

[24] A M Moursi C H Damsky J Lull et al ldquoFibronectinregulates calvarial osteoblast differentiationrdquo Journal of CellScience vol 109 pp 1369ndash1380 1996

[25] J P Quigley L I Gold R Schwimmer and L M SullivanldquoLimited cleavage of cellular fibronectin by plasminogen activatorpurified from transformed cellsrdquo Proceedings of the NationalAcademy of Sciences U S A vol 84 no 9 pp 2776ndash27801987

[26] D Khang S Y Kim P Liu-Snyder G T R PalmoreS M Durbin and T J Webster ldquoEnhanced fibronectinadsorption on carbon nanotubepoly (carbonate) urethaneindependent role of surface nano-roughness and associatedsurface energyrdquo Biomaterials vol 28 no 32 pp 4756ndash47682007

[27] C McFarland C De Filippis M Jenkins et al ldquoAlbumin-binding surfaces in vitro activityrdquo Journal of BiomaterialsScience Polymer Edition vol 9 no 11 pp 1227ndash1239 1998

[28] A Amaral N Alvarado I Marigomez R Cunha K Hyllandand M Soto ldquoAutometallography and metallothionein im-munohistochemistry in hepatocytes of turbot (Scophthalmus

maximus L) after exposure to cadmium and depurationtreatmentrdquo Biomarkers vol 7 no 6 pp 491ndash500 2002

[29] K Endo Y Araki H Ohno and K Matsuda ldquoESCA analysisof tarnish films on dental alloys removed from the oral cavities(Part 1) Ag-In alloysrdquo Journal of Dental Materials vol 7pp 184ndash191 1988

[30] K Anselme ldquoOsteoblast adhesion on biomaterialsrdquo Bio-materials vol 21 no 7 pp 667ndash681 2000

[31] G Balasundaram and T J Webster ldquoAn overview of nano-polymers for orthopedic applicationsrdquo Macromolecular Bio-science vol 7 no 5 pp 635ndash642 2007

[32] R L Price K Ellison K M Haberstroh and T J WebsterldquoNanometer surface roughness increases select osteoblastadhesion on carbon nanofiber compactsrdquo Journal of Bio-medical Materials Research Part A vol 70 no 1 pp 129ndash1382004

[33] R Langer and D A Tirrell ldquoDesigning materials for biologyand medicinerdquo Nature vol 428 no 6982 pp 487ndash492 2004

[34] L Zhang and T J Webster ldquoNanotechnology and nano-materials promises for improved tissue regenerationrdquo NanoToday vol 4 no 1 pp 66ndash80 2009

[35] T Kokubo F Miyaji H M Kim and T NakamuraldquoSpontaneous apatite formation on chemically surface treatedTirdquo Journal of the American Ceramic Society vol 79 no 4pp 1127ndash1129 1996

[36] KWoo GWei and P Ma ldquoEnhancement of fibronectin-andvitronectin-adsorption to polymerhydroxyapatite scaffoldssuppresses the apoptosis of osteoblastsrdquo Journal of BoneMineral Research vol 17 p 49 2002

[37] K Woo R Zhang H Deng and P Ma ldquoProtein-mediatedosteoblast survival and migration on biodegradablepolymerhydroxyapatite composite scaffoldsrdquo in Proceedingsof Transactions of the 28th Annual Meeting of the Society forBiomaterials Tampa FL USA April 2002

[38] V Bucci-Sabattini C Cassinelli P G Coelho A MinniciA Trani and D M D Ehrenfest ldquoEffect of titanium implantsurface nanoroughness and calcium phosphate low impreg-nation on bone cell activity in vitrordquo Oral Surgery OralMedicine Oral Pathology Oral Radiology Endodontologyvol 109 no 2 pp 217ndash224 2010

[39] T J Webster C Ergun R H Doremus R W Siegel andR Bizios ldquoSpecific proteins mediate enhanced osteoblastadhesion on nanophase ceramicsrdquo Journal of BiomedicalMaterials Research vol 51 no 3 pp 475ndash483 2000

[40] Y Shibata D Suzuki S Omori et al ldquoe characteristics of invitro biological activity of titanium surfaces anodically oxi-dized in chloride solutionsrdquo Biomaterials vol 31 no 33pp 8546ndash8555 2010


CorrosionInternational Journal of

Hindawiwwwhindawicom Volume 2018

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Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

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TribologyAdvances in



ChemistryAdvances in


Advances inPhysical Chemistry


BioMed Research InternationalMaterials

Journal of


Na

nom

ate

ria

ls


Journal ofNanomaterials

Submit your manuscripts atwwwhindawicom

biointegration of dental implants into the alveolar bonehave not been elucidated

A quartz crystal microbalance (QCM) sensor is a pro-foundly delicate and handy device that is used to observeprotein adsorption and cell behavior in situ A QCM-basedsensor comprises a quartz crystal and a detection material A27MHz QCM can provide highly sensitive measurements ofmass in aqueous solutions the resonance frequency de-creases in relation to the mass of the protein bound on theQCM electrode surface We previously fabricated severaltypes of QCM sensors by coating the gold electrode of thequartz crystal with a thin film of a biomaterial [9] this QCMsensor achieved an increase in sensitivity approximately 24times that of a conventional 5MHz QCM PMMA (poly(methyl methacrylate)) Au and Ti have been used as QCMelectrode surface materials to imitate denture materials andevaluate the adsorption behaviors of various bovine salivaryproteins [10] ese previous findings support the potentialutility of the QCM method for the evaluation of proteinadsorption behaviors on implant surfaces

In this study we evaluated the effects of modified sur-faces on the adsorption of albumin and fibronectin in RBMcells and simulated body fluid (SBF) in QCM analyses

2 Materials and Methods

21 Sputtering Procedure A thin layer of Ti was depositedon quartz discs (diameter 8mm area 49mm2) by reactivemagnetron sputtering using a radio-frequency magnetronsputtering system (CFS-4ES-231 Shibaura MechatronicsCo Ltd Kanagawa Japan) e QCM crystal was cleanedusing piranha solution (H2SO430H2O2 (vv) 7 3) Beforedeposition was conducted the quartz surfaces were ultra-sonically cleaned in high-purity acetone (99999) Pure Tipowder was used to prepare the target which was formed bypressing the powder into a disc with a diameter of 75mme quartz substrates were positioned 85mm above thetarget and themagnetron sputtering chamber was evacuated toa pressure of 3times10minus3 Pa Argon was used as the working gasand its pressure was kept constant at 67times10minus1 Pa All of thefilms were fabricated using a constant radio-frequency dis-charge power of 480W and the Ti thin films were deposited atroom temperature at a deposition rate of 200 nmmin yieldinga film thickness of approximately 240nm e crystals werewashed and cleaned with both sodium dodecyl sulfate and UV-Ozone Cleaner (PC450 Meiwafosis Co Ltd Osaka Japan)prior to QCM measurements

22 Sample Preparation In the TNS group Ti sensors weretreated to produce TNS on their surfaces An unprocessedQCM sensor was used as the Ti sensor ese sensors wereimmersed in 10MNaOH (aq) and were then placed in an oilbath which was kept at a temperature of 30degC for 24 h esolution in each flask was replaced and treated with distilledwater (200mL) and this procedure was repeated until thepoint that a conductivity of 5 μScm3 was reached especimens were then dried at room temperature

23 Characterization of Materials Scanning electron mi-croscopy (S-4800 Shimadzu Kyoto Japan) and scanningprobe microscopy (SPM-9600 Shimadzu) over a surfacearea of 20 μmtimes 20 μm were conducted to observe thesurface topology and roughness of the fabricated TNS and Tisensors e composition of the coating was analyzed byX-ray photoelectron spectroscopy (XPS ESCA 5600 Ulvac-Phi Inc Kanagawa Japan) using surface etching withionized argon In addition the surfaces of the fabricated Tisensors were subjected to XPS analysis with an Al Kα line(15 kV 300W) as an X-ray source During XPS argon ionsputtering was applied to determine the thickness andstructure of the surface layers

24 Contact Angle Measurements Contact angles weremeasured for the TNS and Ti sensors using a video contactangle measurement system (model VSA 2500 XE ASTProducts Inc Billerica MA USA) A small droplet ofa deionized water solution with Hanksrsquo Balanced Salt So-lution and bovine serum albumin (BSA approximately3mg) was placed on the TNS to measure the static contactangle Estimation of the contact edge is a straightforwardstrategy for breaking down the vitality and hydrophilicnature of a surface

25 Proteins BSA (Wako Pure Chemical Industries LtdOsaka Japan) was dissolved in phosphate-buffered saline(PBS pH 74) at 200 μgmL Human plasma fibronectin(HFN Nacalai Tesque Inc Kyoto Japan) was dissolved inPBS (pH 74) at 500 μgmL

26 Cell Culture Since most bone embed materials areembedded in adult bone that is in direct contact with bonemarrow tissue the effects and success of new embed ma-terials can be investigated by examining bone marrow cellcultures from adult rats RBM cells multiply and separateinto a phenotype that expresses bone cell markers in vitroRBM cells were extracted from the femurs of 7-week-oldSpraguendashDawley rats e rats were humanely sacrificedutilizing 4 isoflurane and the bones were asepticallyextracted from the hind limbs e external soft tissues werediscarded and the extracted bone samples were immersed in50mL of Eaglersquos minimal essential medium (EMEM Wako)supplemented with 20 fetal bovine serum (lot number1412447 Invitrogen Life Technologies Corp CarlsbadCA USA) and penicillin (850UmL) for approximately15min

e proximal end of the femur and the distal end of thetibia were cut An 18-gauge needle (TERMO Japan) wasintroduced into the opening at the knee-joint end of eachbone and the marrow was washed out of the bone shaft byEMEM e obtained marrow pellet was separated bytrituration and the cell suspensions obtained from all thebones were combined by centrifugation RBM cells werecultured in 75 cm2 culture flasks (TD75 Falcon) in EMEMAt confluence the cells were removed by trypsinizationwashed twice in EMEM resuspended in culture medium





ΔF minus2F2

0ΔmA

ρqμq

1113968 (1)








3 Results










(a) (b)


(a) (b)







N1s

Na1s

OlsCls

Ti2p

(a)

Na1sN1s

O1s

C1sTi2p

(b)


0

50

100

150

200

250

Ti TNS

ltBSAgt lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(a)

lowast

0

200

400

600

800

1000

1200

Ti TNS

ltHFNgt

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(b)



0200400600800

10001200140016001800

lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS





275280285290295300

C=O

Binding energy (eV)

C-H C-C

C-O C-N


Nontreatment

(a)

C=O


C-H C-C

C-O C-N


Nontreatment

(b)




Nontreatment

(a)



Nontreatment

(b)


C=O

Binding energy (eV)


Nontreatment

C-H C-C

C-O C-N

280290300 295 285 275

(a)

C=O



Nontreatment

C-H C-C

C-O C-N

(b)




4 Discussion





Nontreatment


(a)

385390395400405410415

Nontreatment


Binding energy (eV)

(b)



01000020000300004000050000600007000080000

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS

lowast









(a) (b)





5 Conclusions






Acknowledgments


References














































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls







ΔF minus2F2

0ΔmA

ρqμq

1113968 (1)








3 Results










(a) (b)


(a) (b)







N1s

Na1s

OlsCls

Ti2p

(a)

Na1sN1s

O1s

C1sTi2p

(b)


0

50

100

150

200

250

Ti TNS

ltBSAgt lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(a)

lowast

0

200

400

600

800

1000

1200

Ti TNS

ltHFNgt

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(b)



0200400600800

10001200140016001800

lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS





275280285290295300

C=O

Binding energy (eV)

C-H C-C

C-O C-N


Nontreatment

(a)

C=O


C-H C-C

C-O C-N


Nontreatment

(b)




Nontreatment

(a)



Nontreatment

(b)


C=O

Binding energy (eV)


Nontreatment

C-H C-C

C-O C-N

280290300 295 285 275

(a)

C=O



Nontreatment

C-H C-C

C-O C-N

(b)




4 Discussion





Nontreatment


(a)

385390395400405410415

Nontreatment


Binding energy (eV)

(b)



01000020000300004000050000600007000080000

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS

lowast









(a) (b)





5 Conclusions






Acknowledgments


References














































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls









(a) (b)


(a) (b)







N1s

Na1s

OlsCls

Ti2p

(a)

Na1sN1s

O1s

C1sTi2p

(b)


0

50

100

150

200

250

Ti TNS

ltBSAgt lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(a)

lowast

0

200

400

600

800

1000

1200

Ti TNS

ltHFNgt

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(b)



0200400600800

10001200140016001800

lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS





275280285290295300

C=O

Binding energy (eV)

C-H C-C

C-O C-N


Nontreatment

(a)

C=O


C-H C-C

C-O C-N


Nontreatment

(b)




Nontreatment

(a)



Nontreatment

(b)


C=O

Binding energy (eV)


Nontreatment

C-H C-C

C-O C-N

280290300 295 285 275

(a)

C=O



Nontreatment

C-H C-C

C-O C-N

(b)




4 Discussion





Nontreatment


(a)

385390395400405410415

Nontreatment


Binding energy (eV)

(b)



01000020000300004000050000600007000080000

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS

lowast









(a) (b)





5 Conclusions






Acknowledgments


References














































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls








N1s

Na1s

OlsCls

Ti2p

(a)

Na1sN1s

O1s

C1sTi2p

(b)


0

50

100

150

200

250

Ti TNS

ltBSAgt lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(a)

lowast

0

200

400

600

800

1000

1200

Ti TNS

ltHFNgt

Am

ount

of a

dsor

ptio

n(n

gcm

2 )


(b)



0200400600800

10001200140016001800

lowast

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS





275280285290295300

C=O

Binding energy (eV)

C-H C-C

C-O C-N


Nontreatment

(a)

C=O


C-H C-C

C-O C-N


Nontreatment

(b)




Nontreatment

(a)



Nontreatment

(b)


C=O

Binding energy (eV)


Nontreatment

C-H C-C

C-O C-N

280290300 295 285 275

(a)

C=O



Nontreatment

C-H C-C

C-O C-N

(b)




4 Discussion





Nontreatment


(a)

385390395400405410415

Nontreatment


Binding energy (eV)

(b)



01000020000300004000050000600007000080000

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS

lowast









(a) (b)





5 Conclusions






Acknowledgments


References














































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls






275280285290295300

C=O

Binding energy (eV)

C-H C-C

C-O C-N


Nontreatment

(a)

C=O


C-H C-C

C-O C-N


Nontreatment

(b)




Nontreatment

(a)



Nontreatment

(b)


C=O

Binding energy (eV)


Nontreatment

C-H C-C

C-O C-N

280290300 295 285 275

(a)

C=O



Nontreatment

C-H C-C

C-O C-N

(b)




4 Discussion





Nontreatment


(a)

385390395400405410415

Nontreatment


Binding energy (eV)

(b)



01000020000300004000050000600007000080000

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS

lowast









(a) (b)





5 Conclusions






Acknowledgments


References














































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls





4 Discussion





Nontreatment


(a)

385390395400405410415

Nontreatment


Binding energy (eV)

(b)



01000020000300004000050000600007000080000

Am

ount

of a

dsor

ptio

n(n

gcm

2 )

Ti TNS

lowast









(a) (b)





5 Conclusions






Acknowledgments


References














































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls










(a) (b)





5 Conclusions






Acknowledgments


References














































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls






5 Conclusions






Acknowledgments


References














































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls




































Advances in



Journal of

Chemistry















Journal of





Volume 2018









Journal of


Na

nom

ate

ria

ls




analysisoftitaniananosheetadsorptionbehaviorusinga...

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