]Available online at w

journal homepage: www.elsevier.com/locate/nanoenergy

ww.sciencedirect.com

Nano Energy (2015) 17, 323–329

http://dx.doi.org/12211-2855/& 2015 E

nCorresponding auE-mail address: y

RAPID COMMUNICATION

Epitaxial growth of vertically alignedpiezoelectric diphenylalanine peptidemicrorods with uniform polarization

Vu Nguyen, Kory Jenkins, Rusen Yangn

Department of Mechanical Engineering, University of Minnesota, Minneapolis, MN 55414, USA

Received 11 June 2015; received in revised form 6 August 2015; accepted 27 August 2015Available online 6 September 2015

KEYWORDSPiezoelectricity;Peptide;Polarization;Diphenylalanine;Microrods;Nanochannels

0.1016/j.nanoen.2lsevier Ltd. All rig

thor.angr@umn.edu (R

AbstractEnergy harvesting with piezoelectric nanomaterials spurred the development of self-powerednanosystems, and piezoelectric biomaterials are expected to play an important role in thebiomedical field. Bio-inspired piezoelectric diphenylalanine (FF) peptide microstructures werefabricated on various substrates through a novel epitaxial growth approach. The low-temperature process produced vertically aligned FF peptide microrods with hexagonallyarranged nanochannels and uniform polarization. Direct measurement of the piezoelectricitywas achieved for the first time from a solid FF peptide single crystal and yielded an effectivepiezoelectric coefficient d33 at 9.9 pm/V. The dense and aligned FF peptide microrods areadvantageous for energy and sensing applications.& 2015 Elsevier Ltd. All rights reserved.

Introduction

Molecular self-assembly of bio-inspired materials has attractedmuch research effort in recent years due to its potential tofabricate novel bottom-up structures for new applications.Among them diphenylalanine (FF) peptide, which is highlybiocompatible with low to no cytotoxicity [1–3], has beenwidely explored recently as a potential candidate for variousapplications such as supercapacitors [4–6], electromechanicalsensing and actuation [7], optics [8,9], nanostructure

015.08.020hts reserved.

. Yang).

fabrication [10], and drug delivery [2]. Many of those applica-tions aim to take advantage of the hydrophilic nanochannelsand charge polarization which are inherent in the crystal self-assembled by linear FF molecules [11,12]. Strong piezoelec-tricity has been demonstrated in FF peptide nanotubes. A higheffective piezoelectric coefficient d15 at 60 pm/V was mea-sured from the shear response of FF peptide hollow tubes [13]and d33 was estimated in the range of 5–50 pm/V [13–15].However, the current long and slender micro/nano tubularstructures lacking in large-scale alignment may prevent effec-tive access to the nanochannels. In addition, the coercive fieldwas estimated on the order of �30 MV/cm which madepolarization switching practically impossible [15]. The currentFF peptide nanotubes show random polarizations from the

V. Nguyen et al.324

growth. The growth of FF peptide nano and microstructureswith uniform polarization is needed in order to observe andtake advantage of the piezoelectricity at large-scale.

Here we report a new and scalable approach to thecontrolled fabrication on various substrates of verticallyaligned FF peptide microrods with uniform polarization. Westart by engineering a textured seed layer with preferentialvertical orientation, and then grow the FF peptide micro-rods epitaxially from this seed layer. Unlike high tempera-ture processes which yield cyclic-FF structure withoutnanochannels and charge polarization [16–18], our simpleprocess is mostly done at room-temperature and theproduced linear-FF peptide is of hexagonal crystal structurewith nanochannels and spontaneous polarization. This novelapproach provides for good alignment of the material incases where it will be used for structural applications. Inapplications where large-scale, uniform piezoelectricity ofthe material is required, this can be achieved by theapplication of an electric field during growth. This polariza-tion under electric field is independent of the good struc-tural alignment achieved by our method. Characterizationtechniques such as X-ray Diffraction (XRD) and Piezore-sponse Force Microscopy (PFM) consistently suggest that thehexagonal crystal structure is maintained. The measuredeffective piezoelectric coefficient d33 of the FF peptidemicrorods is comparable to that of ZnO nanowires [19]. Ourwork can enable new applications of bio-inspired materialsin areas such as energy harvesting and storage, electro-mechanical sensing and actuation, drug delivery, as well asfundamental studies of FF-based structures.

Experimental

Preparation of seed layer

Diphenylalanine lyophilized powder H–Phe–Phe–OH (FF) waspurchased from Bachem and stored in an enclosed dry

Figure 1 Schematic illustration of the fabrication process (a–c)(a) Formation of an amorphous FF peptide layer on the substracrystallization of the amorphous layer from (a). (c) Self-assemblymicrorods. Side SEM image of the seed layer (d) and FF peptide micrpeptide microrods with hexagonally arranged nanochannels. The enlmolecules.

container at 0 1C. 1,1,1,3,3,3-hexafluoro-2-propanol (HFP)was purchased from Sigma Aldrich and stored in a drycontainer under ambient conditions. The schematic of thefabrication process is shown in Figure 1a–c. First, the FFstock solution was prepared by dissolving lyophilized FFpowder in HFP to a concentration of 50 mg/mL in a glovebox (water contento1000 ppm) to minimize water absorp-tion to the source materials. 25 mL of the stock solution wasthen dropped on a substrate placed in a desiccator. Thedrop was quickly vaporized as the desiccator was pumpeddown to 200 Torr in 10 seconds and then vented withcompressed dry air (water content o6 ppm). A transparentamorphous film was formed on the substrate after theevaporation completed, Figure 1a. The low water vaporcontent in the growth environment with relative humidity(RH) lower than 50% is essential to prevent unintentionalcrystallization of the amorphous film [20]. This amorphousfilm was then transferred to an enclosed box which wasconnected in a closed loop of flowing moist air with RH�100% (Figure 1b). The amorphous film was intentionallycrystallized into a seed layer by vigorously circulating thehumid air through the enclosed box for 90 s.

Epitaxial growth of FF peptide microrods

The epitaxial growth from the seed layer was achieved by theprecipitation of a saturated FF aqueous solution, Figure 1c. Firsta concentrated FF aqueous solution was prepared by mixing75 mg FF with 50 mL deionized (DI) water and kept in an ovenat 65 1C until the FF powder was fully dissolved. The substratewith the prepared seed layer was placed floating upside downon the surface of concentrated FF solution right after thesolution was taken from the oven. Ventilation from a small fanwas used to facilitate cooling and evaporation of water in theconcentrated FF solution. Most of the water had evaporatedafter about 6 h. The substrate with FF peptide microrods wasthen taken out, briefly cleaned with DI water, and dried withcompressed air. The growth process is not limited by the

and SEM images and model of FF peptide microrods (d, f).te. (b) Formation of the FF peptide seed layer through theof FF peptide molecules for epitaxial growth of FF peptide

orods grown in the absence of electric field. (f) Illustration of FFarged circle illustrates a nanochannel enclosed by six FF peptide

Figure 2 X-ray powder diffraction patterns of FF peptideseeds and FF peptide microrods. The spectrum of FF peptidemicrorod is shifted upward for clarity.

325Epitaxial growth of vertically aligned piezoelectric diphenylalanine peptide microrods

substrate and FF peptide microrods have been successfullygrown on silicon, indium tin oxide (ITO), platinum and gold (seeFigure S11c–f). Samples on ITO-coated silicon substrate wereused for the characterization demonstrated in this paper.

Morphology and structure characterization

Crystallized seed layers and as-grown FF peptide microrodswere first examined using a Nikon Eclipse LV150 opticalmicroscope. All samples were coated with either gold orplatinum and analyzed using a field emission JOEL 6500scanning electron microscope. Both top view images andside view images were taken from the FF peptide microrods.The crystalline structure of both seed layers and FF peptidemicrorods were subject to XRD analysis after they werecrushed into powders and pressed on a o1004 siliconsubstrate. XRD measurement were made using Bruker D8system with Cobalt anode (λ=1.78899 Å).

Piezoelectricity measurement

Piezoelectricity measurements of the FF peptide seed layerand aligned microrods were conducted with an Asylum MFP3D system. The probe used was an Asylum ResearchASYELEC-01 with Ti/Ir conductive coating, nominal stiffnessof 2 N/m, and nominal resonance frequency of 70 kHz. Forcharacterization of effective piezoelectric coefficient d33, asmall area (0.5� 0.5 mm2) was scanned in Single FrequencyMode with very low frequency (10 kHz) to avoid contactresonance (typically around 300 kHz). Drive voltage ampli-tude on the tip varied from 2 V to 14 V and the correspond-ing vibration amplitude of the tip was recorded andaveraged over the scanned small area to calculate d33. Torecord the phase map of the samples, Dual AC ResonanceTracking (DART) mode was used to enhance the vibrationsignal as well as to eliminate topography cross-talk on theresonance frequency for a large scan size (90� 90 mm2).

Results and discussions

Figure 1d and e shows the cross section of the crystallineseed layer and FF microrods after 6 h room-temperature

epitaxial growth in the absence of electric field. Thevertical microrods are dense and were grown uniformlyover the entire 1.25� 1.25 cm2 substrate (see Figure S1b forlow magnification top view of the array). Their height isapproximately 10 mm from the seed layer, or 15 mm from thesubstrate. The microrods were solid with the diametermostly ranging from 10 mm to 20 mm, which forms thelargest reported solid microrod array to our best knowledge[9]. No branches were observed in FF peptide microrodseven after 12 h of growth, as shown in Figure S1c–f. Thegrowth of the microrods is the result of the self-assembly ofFF molecules on the seed layer as the saturated FF aqueoussolution evaporates. The microrod was expected to have ahexagonal crystal structure [11,21]. As shown in Figure 1f,the FF peptide microrod can be considered as a bundle ofidealized close-packing of hydrophobic nanotubes withhydrophilic hollow channels. The channel is of a van derWaals' diameter of �10 Å and filled with water molecules[21]. Six FF peptide molecules constitute the circumferenceand the translation of those molecules along the axialdirection creates each tube. The side chains with inertphenyl group emanate from the channel cores and areexposed from the side wall. The more active carboxyl groupand amine group forms a hydrogen bond and connects theFF peptide molecules along the axial direction. Thus, unlikethe neutral phenyl groups on the wall, the positivelycharged amine groups or negatively charged carboxyl groupsat the end of the nanochannel assist the self-assembly of FFpeptide molecules for the fast growth along axial direction.Consequently, a highly textured seed layer with nanochan-nels preferentially orientated normal to the substrate iscritical for the epitaxial growth of high quality FF peptidenano or microstructures with desired alignment.

The crystal structure of FF peptide seed layers and FFpeptide microrods were confirmed with XRD analysis.Figure 2 shows typical XRD spectra of the seed layer andthe FF peptide microrod. The microrod structure results inan almost identical XRD pattern to that of the seed layer.The hexagonal structure (P61 spacegroup) proposed byGorbitz and the resultant crystallographic data in the Cam-bridge Crystallographic Data Center (CCDC) file number163340 are in agreement with our XRD spectra [21]. TheXRD data indicates the FF peptide crystals are a close-packing of nanotubes with a diameter of 24 Å. The hydro-philic nanochannels within FF hexagonal structures aredeemed able to host different ions or molecules [2–4,6]for various applications such as energy storage or drugdelivery. Consequently, the vertical FF peptide microrodwith well-aligned nanochannels can increase the accessi-bility and transport of foreign ions or molecules.

The highly textured and crystallized seed layer is criticalfor the successful epitaxial growth of FF peptide microrods,and we explore systematically the formation of FF seedlayers shown in Figure 3. The scattered dark spots in Figure 3are the dark ITO-coated silicon substrate revealed throughvertical FF peptide domains which are good waveguides withlow longitudinal optical loss [9,22], while the bright parts aremainly horizontal FF peptide strips. We found that both thethickness of the initial FF amorphous layer and the supply ofwater molecules during the crystallization control the orien-tation of the seed layer. The thickness of the amorphous layeris proportional to the concentration and the amount of FF-

Figure 3 Optical images of the crystalline seed layer formed from different FF concentrations and moist air. (a) From 100 mg/mLFF solution under stationary moist air. (b) From 100 mg/mL FF solution under circulating moist air. (c) From 50 mg/mL FF solutionunder stationary moist air. (d) From 50 mg/mL FF solution under circulating moist air. All the amorphous layers used to form thecrystalline seed layer were obtained by dropping 25 mL of FF solution on a 1.25� 1.25 cm2 substrate.

V. Nguyen et al.326

HFP stock solution dropped onto the substrate [18, 20] (alsoshown in Figure S2). Figure 3 shows the effect of thethickness of the amorphous layer and the supply of watermolecules. 25 mL FF solution of concentration either50 mg/mL or 100 mg/mL was dropped and dried on a1.25� 1.25 cm2 ITO-coated silicon substrate to form anamorphous FF layer. Crystallization of these layers wasachieved with either stationary or circulating moist air.Comparing Figure 3a and b and Figure 3c and d, it is clearthat lower concentration, corresponding to thinner amor-phous layer, yields higher portion of vertical orientation. For100 mg/mL concentration, the FF seed layer was dominatedby bright horizontal FF stripes in random directions with afew transparent vertical domains which are shown as darkspots. Circulating moist air was also proved to be critical toproduce a seed layer with dominant vertical domains, asshown in Figure 3c and d. The optimal seed layer with amajority of vertical domain is obtained with 50 mg/mL FFsolution and circulating moist air. The highly reflectivehorizontal stripes become the minority in the crystallizedseed layer and most of the dark silicon substrate is visiblethrough the dense vertical FF peptide microrods.

This observation indicates that the crystallization processis driven by the diffusion of water molecules into theamorphous FF peptide layer. Fick's first law of diffusionsuggests that the diffusion flux of water molecules in thevertical direction is proportional to the gradient of theconcentration of water molecules, which in turn is propor-tional to the water vapor concentration at the surface andinversely proportional to the layer thickness. When a thickamorphous FF peptide layer is used, the water molecule

diffusion normal to the substrate is limited and horizontal FFpeptide strips are preferentially formed. In comparison, themuch enhanced vertical diffusion in a thin amorphous layerwith continuous supply of water molecules from circulatingmoist air significantly facilitates the creation of vertical FFpeptide microrods. The Scanning Electron Microscope (SEM)images in Figure 4a and b show a vertical area in the seedlayer, which is approximately a hexagon and consistent withits crystal structure and orientation. The zoom-in image inFigure 4b confirms this vertical seed is solid, and thescattered nanowires imply that the top surface has activesites for the self-assemble of FF molecules. The vertical seedparticles are expected to initiate epitaxial growth of FFpeptide crystals. An FF peptide microrod grown from theseed layer in Figure 3d is shown in Figure 4c, along with thetop view of the micro rod tip in Figure 4d and the side wallwith vertical strips in Figure 4e. The rough tip and smoothside wall of the micro rod is a result of the fast growth alongthe axial direction and consistent with the observation thatthere is no branch formed.

The highly-textured seed layer and the verticallyoriented microstructures enable us to conveniently char-acterize their piezoelectric property using PFM, which hasproved challenging to perform on free standing FF peptidenanotubes [13]. The amplitude responses of the PFMmeasurement on the peptide seed and on the FF peptidemicrorod are shown in Figure 5a. The amplitude increaseslinearly with the applied voltage, and the slope gives theeffective piezoelectric coefficient 9.4 pm/V for the FFpeptide seed and d33 ¼ 9.9 pm/V for the FF peptide micro-rod. This result indicates that the longitudinal piezoelectric

Figure 4 (a) Top-view SEM image of a solid FF peptide seed with a roughly hexagonal shape. (b) Magnified SEM image from the circledarea in a) showing nanowires leading the later growth of a FF peptide microrod. (c) Side-view SEM image of a FF peptide microrod.(d) Top view magnified image of the tip of a microrod and (e) side view magnified image of the side-wall of the micro rod in (c).

Figure 5 (a) PFM amplitude response of an FF peptide seed layer and FF peptide microrods. Phase map of an FF peptide seed layergrown (b) without electric field and (c) with electric field by turning on the voltage in Fig. 1b. (e) Histogram calculated from thephase map in (b). (f) Histogram calculated from the phase map in (c).

327Epitaxial growth of vertically aligned piezoelectric diphenylalanine peptide microrods

coefficient d33 of FF peptide microrods is comparable tothat of ZnO nanowires [19], and the vertically aligned arrayof FF peptide microrods can potentially be a good candidatefor mechanical energy conversion applications.

The PFM phase response shows that the domain patternof the microrod is determined by the seed layer. A seedlayer that had domains with random polarization resultedin the growth of microrods with random polarization,

V. Nguyen et al.328

while an array of microrods with uniform polarization wasachieved from a seed layer with uniform polarization. Thecontrol of the polarization was achieved using the electricfield applied during the growth, while the electrical fielddid not affect the alignment of the microrods from thisnovel growth process. When a seed layer was formed inthe absence of an electric field, the PFM phase image ofthe seed layer in Figure 5b shows many domains withopposite polarization. These antiparallel dipoles are stillmostly vertical instead of randomly pointed due to thestructural vertical alignment of the micro rod. The phasehistogram in Figure 5d shows that the seed layer hasroughly equal amount of domains which are 1801 out ofphase. The microrods from such seed layer showed similardomain distribution. The piezoelectric effects of indivi-dual domains will cancel out and the microrod array willnot exhibit macroscopic piezoelectric effect. Consideringthe difficulty in the polarization switching of FF peptideby an electric field [13, 14], which is also demonstrated inFigure S3f, we explored a novel approach for the fabrica-tion of microrods with uniform spontaneous polarization.The uniform polarization can be achieved through apply-ing an electric field during the growth. In the setup inFigure 1b the applied voltage can be switched from 0 V to5 kV, which produced an electric field of 1 MV/m betweena 5 mm electrode gap. The resultant seed layer showedmuch more uniform phase maps with much less domainswith the opposite phase, as shown in Figure 5c. The phasehistogram in Figure 5e shows that the seed layer wasdominated by domains with one polarization direction.Preliminary data indicates that most examined microrodsfrom this seed layer were approximately in phase, butmore statistical characterization for the array is neededand being investigated.

Conclusion

To summarize, in this paper we demonstrated the low-temperature fabrication of densely packed, verticallyaligned FF peptide microrods. The crystal structure of FFpeptide microrods with hexagonally arranged nanochannelswas confirmed with XRD. The effective longitudinal piezo-electric coefficient d33 of 9.9 pm/V was measured with PFM.Uniform spontaneous polarization was achieved by applyingan electric field during the FF peptide growth. The highquality piezoelectric biomaterial from this work provides anew platform for applications in sensing, energy harvesting,and medicine.

Acknowledgments

The authors are truly grateful for the financial support fromthe Department of Mechanical Engineering and the Collegeof Science and Engineering of the University of Minnesota.Research is also supported by in part by NSF (ECCS-1150147)and by NSF IGERT Grant DGE-1069104. The electron micro-scopy image was obtained in the Characterization Facility,University of Minnesota, which receives partial support fromNSF through the MRSEC program. The device fabrication was

performed in the Minnesota Nano Center, a part of the NSF-funded National Nanotechnology Infrastructure Network.

Appendix A. Supplementary material

Supplementary data associated with this article can befound in the online version at http://dx.doi.org/10.1016/j.nanoen.2015.08.020

References

[1] N. Santhanamoorthi, P. Kolandaivel, L. Adler-Abramovich, E. Gazit,S. Filipek, S. Viswanathan, A. Strzelczyk, V. Renugopalakrishnan,Adv. Mater. Lett. 2 (2011) 100.

[2] R.F. Silva, D.R. Araujo, E.R. Silva, R.A. Ando, W.A. Alves,Langmuir 29 (2013) 10205–10212.

[3] X. Yan, P. Zhu, J. Li, Chem. Soc. Rev. 39 (2010) 1877–1890.[4] J. Zhang, X. Wu, Z. Gan, X. Zhu, Y. Jin, ,, Nano Res. 7 (2014)

929–937.[5] L. Adler-Abramovich, D. Aronov, P. Beker, M. Yevnin,

S. Stempler, L. Buzhansky, G. Rosenman, E. Gazit, Nat.Nanotechnol. 4 (2009) 849–854.

[6] P. Beker, G. Rosenman, J. Mater. Res. 25 (2010) 1661–1666.[7] E. Bosne, A. Heredia, S. Kopyl, D. Karpinsky, A. Pinto,

A. Kholkin, Appl. Phys. Lett. 102 (2013) 073504.[8] X. Yan, J. Li, H. Möhwald, Adv. Mater. 23 (2011) 2796–2801.[9] Q. Li, Y. Jia, L.R. Dai, Y. Yang, J.B. Li, Acs Nano 9 (2015)

2689–2695.[10] M. Reches, E. Gazit, Science 300 (2003) 625–627.[11] C.H. Görbitz, Chem. Commun. (2006) 2332–2334.[12] M. Wang, L. Du, X. Wu, S. Xiong, P.K. Chu, ACS Nano 5 (2011)

4448–4454.[13] A. Kholkin, N. Amdursky, I. Bdikin, E. Gazit, G. Rosenman, ACS

Nano 4 (2010) 610–614.[14] I. Bdikin, V. Bystrov, I. Delgadillo, J. Gracio, S. Kopyl,

M. Wojtas, E. Mishina, A. Sigov, A.L. Kholkin, J. Appl. Phys.111 (2012).

[15] A. Heredia, I. Bdikin, S. Kopyl, E. Mishina, S. Semin, A. Sigov,K. German, V. Bystrov, J. Gracio, A.L. Kholkin, J. Phys. D:Appl. Phys. 43 (2010).

[16] B. Bank-Srour, P. Becker, L. Krasovitsky, A. Gladkikh,Y. Rosenberg, Z. Barkay, G. Rosenman, Polym. J. 45 (2013)494–503.

[17] A. Heredia, I. Bdikin, S. Kopyl, E. Mishina, S. Semin, A. Sigov,K. German, V. Bystrov, J. Gracio, A. Kholkin, J. Phys. D: Appl.Phys. 43 (2010) 462001.

[18] J. Ryu, C.B. Park, Adv. Mater. 20 (2008) 3754–3758.[19] Z.L. Wang, J. Song, Science 312 (2006) 242–246.[20] J. Ryu, C.B. Park, Chem. Mater. 20 (2008) 4284–4290.[21] C.H. Görbitz, Chem. Eur. J. 7 (2001) 5153–5159.[22] X.H. Yan, J.B. Li, H. Mowald, Adv. Mater. 23 (2011) 2796–2801.

Vu Nguyen received his B.S. degree inMechanical Engineering from WorcesterPolytechnic Institute, Worcester, Massachu-setts in 2012. He is currently pursuing Ph.D.degree at the University of Minnesota,Minneapolis, Minnesota. His research inter-ests are energy harvesting and self-powersystems at micro/nano scale.

329Epitaxial growth of vertically aligned piezoelectric diphenylalanine peptide microrods

Kory Jenkins received his Bachelor’s degreein Aerospace Engineering and Mechanics fromthe University of Minnesota in 2007. He iscurrently a PhD candidate at the University ofMinnesota. His research interests includefabrication of novel piezoelectric sensors forhaptics and human machine interaction.

Rusen Yang received his PhD degree inMaterials Science and Engineering fromGeorgia Institute of Technology in 2007,where he continued as Post-Doctoral Associ-ate. He joined Mechanical Engineering atthe University of Minnesota-Twin Cities asan assistant professor in 2010. He has dis-covered novel nanostructures, such as ZnO,SnO2, Zn3P2 and investigated their applica-tion potentials. His most recent work on

energy harvester based on piezoelectric nanomaterials made sig-nificant contribution in the field of renewable energy.