Binding of Polycitydylic Acid to Graphene Oxide: Spectroscopic Study

Computer Modeling. Maksym V. Karachevtsev1*, Stepan G. Stepanian1, Alexander Yu. Ivanov1, Victor S. Leontiev1,. Vladimir A. Valeev1, Oksana S. Lytvyn2...
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Binding of Polycitydylic Acid to Graphene Oxide: Spectroscopic Study and Computer Modeling Maksym V. Karachevtsev,*,† Stepan G. Stepanian,† Alexander Yu. Ivanov,† Victor S. Leontiev,† Vladimir A. Valeev,† Oksana S. Lytvyn,‡,§ Ludwik Adamowicz,∥ and Victor A. Karachevtsev*,† †

B. Verkin Institute for Low Temperature Physics and Engineering, National Academy of Sciences of Ukraine, 47, Nauky Ave., Kharkiv, 61103, Ukraine ‡ V. Lashkaryov Institute of Semiconductor Physics, National Academy of Sciences of Ukraine, Kyiv, 03028, Ukraine § Borys Grinchenko Kyiv University, 18/2 Bulvarno-Kudriavska Str., Kyiv, 04053, Ukraine ∥ Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721, United States S Supporting Information *

ABSTRACT: Hybridization of nucleic acids with graphene nanomaterials is of great interest due to its potential application in genosensing and nanomedicine. In this work we study the interaction between polyribocytidylic acid (poly(rC)) and graphene oxide (GO). The study involves comparing the UV absorption spectra of the free polymer and the polymer bonded to graphene oxide and analyzing the vibrational structure of the systems and their components using FTIR spectroscopy. Spectral shifts of the electronic and vibrational bands of the poly(rC) and changes of their thermostability due to the adsorption on GO are observed. Molecular dynamics simulation of the adsorption process of the r(C)10 and r(C)30 oligomers on graphene demonstrates their disordering due to the π−π stacking of cytosines on graphene and shows that the longer oligomer adsorbs slower. The binding energies of a single cytosine stacked with graphene in water and in vacuum were determined. The calculated IR lines of the stacked cytosine with graphene are red-shifted by up to 20 cm−1 compared to free cytosine. A strong decrease of the intensities of the cytosine vibrations in the 1800−1400 cm−1 range resulting from the interaction with graphene is revealed in the spectra. When cytosine is adsorbed to graphene oxide, their complex is additionally stabilized by H-bonding. It leads to an increase of the red shifting of the cytosine lines.

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INTRODUCTION Graphene oxide (GO) is a water-soluble derivative of graphene, which has unique characteristics such as good water dispersibility, facile surface functionalization, and high mechanical strength. These properties make this nanomaterial very attractive in many applications including biosensing, designing drug delivery, photothermal and photodynamic therapy, tissue engineering, and imaging of different biological processes in vitro (in cells) and in vivo (animal studies).1−3 GO bears some oxygen functional groups on the basal planes (hydroxyl and epoxy groups) and edges (ester, carboxyl groups) resulting in mixed sp2/sp3 hybridized carbon domains.4 Existence of flat sp2-hybridized sections surrounded with different oxygen-containing groups facilitates efficient interaction of GO with biomolecules of different types, including small biologically active molecules and large structures like proteins and nucleic acids. Among them, adsorption of DNA/ RNA on GO has received special attention because of promising applications in genosensing and gene delivery.5−7 The π−π stacking of the nitrogen bases onto π-system of electrons of carbon nanomaterials such as nanotubes or graphene provides relatively strong binding of these systems to DNA/RNA.7−10 In the case of GO deprotonation of the oxygen-containing groups, especially carboxyl groups, at the © 2017 American Chemical Society

sheet edges renders negatively charged GO, which is electrostatically stabilized as a colloidal suspension in water. It is wellknown that DNA is a negatively charged polymer. Thus, these nanosized structures repel each other in aqueous solution. To compensate the negative charges on these systems high concentrations of salt (NaCl) that can screen the electrostatic repulsion of DNA and GO are used in the experiment.10 Divalent metal ions (Mg2+ or Ca2+) can also enable the DNAGO binding and adsorption of double-stranded DNA onto GO.11 The main mechanism of the ssDNA adsorption on GO is based on the hydrophobic and π−π stacking interactions between the sp2-hybridized domains of GO and the DNA nucleobases (the same mechanism as that involved in ssDNA adsorption on nanotube or graphene). However, in contrast to nonfunctionalized carbon nanomaterials, in the case of GO and ssDNA an additional enhancement of the binding appears due to hydrogen bonding (H-bonding) between positively partly charged groups of nucleobases and oxygen-containing functional groups of GO.12,13 Note that adsorption efficiency of Received: May 18, 2017 Revised: July 26, 2017 Published: July 28, 2017 18221

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chemical calculations are employed. The homopolynucleotide poly(rC) model system has been used before to study the DNA/RNA structural transformations occurring upon adsorption of these systems on different nanostructures including carbon nanomaterials.23 Another reason for selecting this system as a model in the present study is due to its being a component of the double-stranded poly(rI)·poly(rC) duplex which plays an important biological role in the activation of the human innate immune system and in the adaptive immune responses. The consideration of the poly(rC) adsorption on GO is also related to the possible use of GO as a scaffold for the delivery of the duplex to cells.

ordered dsDNA on GO is much lower than that of ssDNAs because the nucleobases are hidden inside the duplex due to Hbonding. As a result, π−π stacking of the bases with the πsystem of electrons of GO is possible only at the polymer ends.5,7 Additionally, the two negative strands of the sugar− phosphate backbone of the duplex arranged closely one to the another weaken the dsDNA adsorption onto GO.11 As a result, ds-DNA adsorption onto GO is possible only in the presence of a high salt concentration.11,14 GO is an efficient quencher of the adsorbed chromophores and this property is often exploited to study the adsorption of fluorescently labeled DNAs.1,5,6 Such DNAs (probes) are nearly fully quenched upon adsorption onto GO. The weaker binding of dsDNA to GO in comparison with the probe is proposed as a property enabling luminescent sensing of molecular recognition events. It can be also used for detection of a variety of analytes.1,2 In that approach, the addition of the complementary DNA (the target) results in probe desorption that leads to the emission enhancement.1,5,6 It should be added that ssDNA can bind strongly to the GO surface and be effectively protected from nuclease digestion. This effect confirms the stability of ssDNA adsorption on GO. The absorption of ssDNA on GO can be exploited for preventing the cleavage of these biopolymers from enzymatic cleavage by deoxyribonuclease.14−16 The elaboration of modern DNA-based optical sensors based on GO and promising perspectives of the use of DNA-GO hybrids in nanomedicine stimulate growing interest in acquiring comprehensive information concerning the DNA adsorption (desorption) on (from) graphene nanomaterials with the purpose to control these processes. Taking into account the wide spectrum of structures and conformations of DNA/RNA one can expect more investigations in this direction to appear in the near future. It was recently demonstrated that DNA adsorption onto and desorption from GO depends on the DNA length12,17−20 and on the degree of oxidation of GO21 and on its size.18 It should be noted that the information obtained about the binding ability of ss-DNAs with different lengths is often contradictory. Zhao et al. showed that short ssDNAs have weaker affinity to GO than long ones.22 Wu et al. compared the adsorption of 12-, 18-, 24-, and 36-mer fluorescent-labeled ssDNAs on GO and found that the quenching efficiency of dye was lower for longer ssDNAs and suggested that the binding of such polymers to GO is weaker.10 In a study of the adsorption kinetics of different oligomers it was shown that shorter DNAs were adsorbed more quickly than longer ones (FAM-labeled ss-DNA 12, 24, and 44mers).20 Other researchers compared three types of fluoresceinlabeled DNA oligomers (5, 20, and 43-mer oligonucleotides) and found that shorter DNA was more effectively loaded on GO regardless of the GO size18 and they also showed faster adsorption kinetics for GO with smaller sizes (called NGO).17 They concluded that the shorter fluorescein-labeled DNAs bind to GO less tightly while their association/dissociation with GO is faster. In this work, we study the interaction between a relatively long homopolynucleotide polyribocitydylic acid (poly(rC)) and GO. The UV absorption spectra of the free and bound polymers are recorded and an analysis of their vibrational structures is performed based on the date obtained using the FTIR absorption spectroscopy. To explain the observed spectral manifestation of the interaction between poly(rC) and GO, the molecular dynamics simulation and quantum-

2. MATERIALS AND METHODS Materials. Potassium salt of polyribocytidylic acid (poly(rC)) is purchased from Sigma-Aldrich (Europe) and used as received. Poly(rC) is dissolved in aqueous buffer solution consisting of 1 × 10−3 M Na+ cacodylate (pH 7.4) (Serva, Germany) and 9.9 × 10−2 M NaCl (pH7.4). Deionized water with 18 MΩ resistance obtained with a Millipore Super-Q system (Millipore Co., Billerica, MA) is used in the experiment. The concentration of the polynucleotide is determined optically (see, for example, ref 23). The initial poly(rC) concentration in solution is 80 μg/mL. Water-soluble graphene oxide is prepared using the modified Hummers method.24,25 Briefly, 3 g of pure graphite flakes (Sigma-Aldrich) is added to a mixture solution containing concentrated H2SO4 (75 mL) while stirring in an ice bath for 30 min. Under vigorous agitation, KMnO4 (9.0 g) is added slowly to keep the temperature of the suspension lower than 20 °C. Successively, the reaction system is transferred to a 40 °C oil bath and vigorously stirred for about 40 min. Then 200 mL of water is added, and the solution is stirred for 15 min at 95 °C. An additional 700 mL of water is added and followed by a slow addition of 75 mL of H2O2 (35%). To remove the unexfoliated graphite and the remaining metal species, the GO dispersion is centrifuged at 6000 rpm for 20 min. Next the selected supernatant is centrifuged at 8000 rpm for 40 min. The brown sediment is washed with water by performing five cycles each involving suspension and subsequent centrifugation to remove the remaining unexfoliated graphite. To calculate the concentration of the GO solution, 3 mL of stock solution is taken for drying on a substrate by warm air and then the obtained dry GO is weighed. The procedure gives the GO concentration in stock of 300 μg/mL. The resulting homogeneous brown dispersion is stable for a few months. Next GO is analyzed by the UV−visible absorption spectroscopy and by the FTIR technique. Both show the presence of GO. Additionally the morphology of GO nanosheets on mica is analyzed by atomic force microscopy (AFM). The AFM observation shows that the supernatant suspension of the final product contains GO flakes with various thicknesses (ranging from a monolayer to a few layers). Methods. Absorption Spectroscopy. UV−visible absorbance measurements are carried out with Specord M40 (Carl Zeiss, Jena, Germany) equipped with a cuvette holder containing a thermoelectric converter for temperatures stabilization. Quartz cuvettes with 10 mm path length are used in the experiments. The absorption spectra are recorded in the range of 200−700 nm at room temperature. The temperature dependence of the changes in the optical density (ΔA(T)) of polynucleotide (the melting curve) is measured at the heating rate of 0.25 °C/min at λ = 268 nm (the 18222

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(with 1 fs step size). During the simulation, the usual periodical boundary conditions are applied. In the periodic box, the modeling temperature is 303 K and the pressure is 1 atm. The long-range electrostatic interactions are calculated with the Particle Mesh Ewald method.29 Quantum-Chemical Calculations. Structures, interaction energies, and vibrational spectra of the graphene-cytosine (GRCYT) and graphene oxide-cytosine (GO-CYT) complexes are calculated using the B3LYP density-functional method30−32 with an empirical dispersion correction. The D3 version of Grimme’s dispersion with the Becke-Johnson damping33,34 invoked via the “GD3BJ” keyword is employed. This particular computational method is used because the calculations concern vibrational spectra of complexes that involve stacking interaction between cytosine and graphene. The B3LYP density-functional method is the standard tool for predicting vibrational spectra. Including the dispersion correction allows one to apply the method to stacked complexes. The standard 631++G(d,p) basis set is used in all calculations. The use of the “5d” keyword reduces the number of orbitals in each d-shell to five. A hexagonal 96 atom fragment of the carbon surface with terminal hydrogen atoms is used in the calculations as a model of graphene. The models of graphene oxide surface are built by addition of oxygen containing groups to the graphene model. For the graphene oxide models we used notation GO(XYZ), where X, Y, and Z correspond to the number of peripheral carboxylic groups (−COOH), number of basal epoxy oxygen atoms (−O−), and the number of basal hydroxyl groups (−OH), respectively. The structures of the graphene models used in this work are shown in Figure S1. As can be seen, in the models the basal groups appear on both sides of the graphene sheet. The energy of the interaction between the fragments of each complex is calculated with the inclusion of the ZPVE and BSSE corrections. All calculations are performed using the Gaussian 09 program package.35

maximum of the poly(rC) UV-absorption band). In these measurements, a two-cuvette arrangement is used: one cell containing the GO suspension (or polymer solution) is placed in the working channel of the spectrophotometer and heated, while the second cell (the reference cuvette) containing a buffer solution is thermostated within 20 ± 0.5 °C. Melting curves of poly(rC) (free and adsorbed onto GO) are measured and recorded as h(T)= (A(T) − A0)/A0, where A0 is the absorbance of the polymer at 20 °C, A(T) is the absorbance of the polymer at the temperature indicated, and h(T) is the hyperchromic coefficient. The influence of doping of the GO suspension on the absorption spectrum of poly(rC) is studied by employing the differential-absorption spectroscopy method. This technique has been often used to analyze the structural changes in polynucleotides caused by their interaction with nanostructures. The measurement involves the use of a four-cuvette arrangement (see the Supporting Information, Scheme S1) which includes a cuvette of poly(rC) solution placed in each of the two channels and two cuvettes with buffer solution. GO doping of poly(rC) solution in a working channel is compensated with (a) the same volume of GO added to the buffer cuvette and (b) the same volume of buffer solution added in the other cuvette of poly(rC) in the reference channel. The purpose of the latter addition is to compensate the dissolution of poly(rC) by GO suspension. AFM Measurements. The aqueous suspensions of GO are used to prepare samples for the AFM measurements. A volume of 7 μL of a dilute (1:100) suspension is placed onto a freshly cleaved mica surface by the spin coating method. The sample is then air-dried and analyzed by AFM. AFM images of GO are obtained using a Nanoscope III D3000 AFM (Digital Instruments, Santa Barbara, CA) operating in the tapping mode. FTIR Spectroscopy. FTIR absorption spectra (taken in the 3800−500 cm−1 range) of poly(rC), GO, and poly(rC):GO films are obtained by employing a FTIR spectrometer with the 0.3 cm−1 apodized spectral resolution (for details on the setup see, for example, ref 26). Films for the measurements are prepared by the deposition of the corresponding compounds from aqueous suspensions on ZnSe substrates by the dropcasting method. In order to obtain a poly(rC):GO film for the FTIR measurement, GO in the aqueous suspension is mixed with the aqueous solution of polynucleotide in the 1:1 weight ratio. All FTIR absorption spectra are obtained at room temperature. Molecular Dynamics Simulation Protocol. The formation of graphene (GR):oligonucleotide hybrid is simulated by the molecular dynamics method. In the simulations, the program package NAMD27 with Charmm27 force field parameter set28 is used. The calculations are performed for two oligonucleotides with 10 and 30 nucleotides in length. Each system is embedded in water (number of water molecules varies from 28221 to 30284). The dimensions of the simulation boxes are 148 Å × 145 Å × 45 and 148 Å × 145 Å × 49 Å for GR:r(C)10 and GR:r(C)30, respectively. In both cases the graphene sheet consists of 5682 C and 212 H atoms. Before modeling, r(C)10 or r(C)30 (A-form) is placed near graphene. To neutralize the charge of the sugar−phosphate backbone for GR:r(C)10, 10 Na+ ions are added to the box (30 ions for GR:r(C)30). During the molecular dynamics simulation, the coordinates of the graphene atoms are frozen. The energies of such systems are first minimized during 500 steps and then modeled for 30 ns

3. RESULTS AND DISCUSSION 3.1. Characterization of GO. 3.1.1. Atomic Force Microscopy Experiments. AFM is a powerful tool to measure the heights of carbon nanostructures deposited on a substrate and to estimate their sizes. Figure 1 shows the AFM image of GO nanosheets produced in the present experiments and adsorbed to freshly cleaved mica. The average thickness of a GO nanosheet is about 0.83 nm. This value indicates that on mica mainly a single layer of GO is adsorbed and this finding agrees with the previous reports.36−38 The height measurements performed at different locations indicate that the GO sheet has a uniform thickness (without any noticeable wrinkles). The thickness value of about 1.64 nm shown in Figure 1 indicates overlapping of two single GO layers. On the basis of an analysis of different images of GO, the linear sizes of the GO sheets in the suspension can be estimated as ranging from 0.5 to 2 μm. 3.1.2. FTIR Spectroscopy. FTIR spectroscopy is another useful method for the GO characterization, as the FTIR spectrum provides information about the presence of different functional groups in the GO structure. The FTIR spectrum of the GO film is obtained in the 3700−700 cm−1 spectral range (Figure S2A). At high frequencies (3600−3300 cm−1), an intensive broad band appears that is attributed to the O−H stretching vibrations of the hydroxyl and carboxyl groups of 18223

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spectrum we consider the spectrum of the poly(rC) film. Similarly to the GO spectrum, the spectrum of homopolynucleotide is characterized by intensive bands in the 3700−700 cm−1 range. The broad intensive band at 3357 cm−1 is assigned to the stretching modes of the O−H group of the ribose ring and of the NH2 group of cytosine (Figure S2B).45,46 Note that the width of this band practically coincides with the absorption band of GO in this spectral region (Figure S2A). However, in the fingerprint range, the widths of the bands of poly(rC) are noticeable narrower compared to the GO bands (Figure 2B). Before we consider each band in detail, two spectral regions should be identified, namely, the 1700−1500 and 1100−900 cm−1 regions which correspond to the cytosine and sugar− phosphate moiety vibrations, respectively. Detailed assignments of the bands in these two regions can be found elsewhere.45−49 The most intensive band of the first spectral region (at 1654 cm−1) is likely related to the stretching vibration of the CO group of cytosine.45,46 The frequency of this band decreases compared to the corresponding band of the GO hexagon due to the presence of nitrogen in the pyrimidine ring of cytosine.46 It should be noted that the intensity of this band is increased due to a contribution from the band corresponding to the scissors mode of the closely located NH2 group.45,46 The stretching mode of the CC bond of pyrimidine ring of cytosine gives rise to the band at 1611 cm−1 (in GO this mode is responsible for the band at 1587 cm−1). Two bands at 1527 and 1497 cm−1 arise from the in-plane vibrations of the base residue. The band at 1235 cm −1 is assigned to the antisymmetric stretching vibration of the phosphate groups. The symmetric stretching vibration of these groups appears at 1078 cm−1. The stretching vibration of the C−O bond also contributes to the intensity of the latter band. At lower frequencies in the spectrum of poly(rC), weak bands attributed to the ribose-phosphate linkage are observed.44,49 In the FTIR spectrum of the poly(rC):GO film (Figure S2C) there are some noticeable spectral differences compared to the spectra of the constituents (Figure 3A). By comparing the

Figure 1. AFM image of GO nanosheets adsorbed on freshly cleaved mica. The cross-session profile corresponds to the black line drawn in the topographic image.

GO and, partly, to residual water located between the GO sheets. However, more detailed information about oxygencontaining groups in the GO structure can be extracted from the fingerprint range (1800−700 cm−1, Figure 2). The

Figure 2. FTIR spectra of GO (A) and poly(rC) (B) films in the spectral range 1800−700 cm−1. Numbers indicate the peak position in cm−1.

characteristic features in the 1780−1650 cm−1 range of the spectrum are the absorption bands at 1720 cm−1, which corresponds to the CO stretching vibrations of the carbonyl and carboxyl groups and the band at 1626 cm−1 assigned to the carboxyl group.4,39−43 In this range we also observe an intensive band which peaks at 1587 cm−1. This band can be attributed to the CC vibrations involving fragments of the sp2-hybridized domains. The epoxide (C−O−C) stretching vibration leads to appearance of the bands at 1235 and 925 cm−1.43 The band at 1163 cm−1 is mainly due to the C−O vibrations of furan-like ether and hydroxyl groups.44 3.2. Binding of Poly(rC) with GO. 3.2.1. FTIR Study. Before making an assignment of the poly(rC):GO FTIR

Figure 3. FTIR spectra of poly(rC):GO (A) and the differential spectrum of poly(rC):GO-poly(rC) (B) (black) films in the spectral range 1800−700 cm−1. For comparison, the spectra of poly(rC) (A) and GO (B) (blue, dotted) are shown too. 18224

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The Journal of Physical Chemistry C spectra of GO and poly(rC) one can conclude that they are quite similar. Some of the bands are slightly shifted toward higher or lower frequencies due to the differences in the molecular surroundings of the groups responsible for the bands in the two systems. The similarities complicate the detection of spectral manifestations of the noncovalent interaction between the components of the poly(rC):GO complex. Comparing the FTIR spectra of composite and its components one notices the absence in the composite spectrum of the band at 1720 cm−1 assigned to the stretching vibration of the CO group in the GO sheet. In addition, the peak of the band at 1654 cm−1 corresponding to this group in poly(rC) is shifted to 1651 cm−1 in the composite. These observations suggest the involvement of the CO groups in the H-bonding with corresponding functional groups of poly(rC) and/or GO. Additional spectral information can be obtained from the differential spectrum generated by subtracting the poly(rC) spectrum from the spectrum of the composite (Figure 3B). As the direct subtraction of the spectra can lead to some errors, the spectra are normalized before they are subtracted. In the normalization, the band with the peak at 1497 cm−1 in the biopolymer spectrum is used. This band practically does not overlap with any band in the GO spectrum in this spectral range. We also assume that this mode changes little when poly(rC) is mixed with GO. The differential spectrum is shown in Figure 3B along with the GO spectrum for comparison. The differential spectrum shows differences between spectra of bound and unbound GO, especially for the bands attributed to the GO functional groups. Let us first consider the two absorption bands in the differential spectrum at 1646 and 1593 cm−1 (Figure 3B). The appearance of these bands and the disappearance of the band at 1720 cm−1 can be explained by the formation of hydrogen bonds between the oxygencontaining groups of the GO sheet (COOH and COH) and the exocyclic CO groups of cytosine. The hydrogen bonds also cause spectral shifts of the bands observed in the spectra of the starting materials. Obviously, the hydrogen bonds may also involve the NH2 group of cytosine and the OH group of the ribose ring. An involvement of the OH groups of GO is manifested by noticeable changes in the 1200−900 cm−1 spectral region (Figure 3B). In the differential spectrum, as compared with the spectrum of GO, the intensity of the absorption band at 1235 cm−1 decreases. This likely indicates the formation of the hydrogen bonds with the additional involvement of the epoxy groups of GO. 3.2.2. UV Absorption Spectroscopy of poly(rC):GO. The UV−visible spectrum of the GO aqueous dispersion demonstrates a significant increase of light absorption with decreasing wavelength. The UV spectrum of GO shows two features: absorption peak at 220−240 nm due to the π → π* electronic transition of the carbon atoms in the sp2-hybridization (CC bond) and a shoulder in the 260−320 nm region associated with the n → π* electronic transition of the CO bonds and related to the sp3 hybridization in GO (see, for example, 40). This shoulder in the UV spectrum of GO is observed in the spectrum shown in Figure 4A. DNA and synthetic homopolynucleotides show a broad band in the UV absorption spectra with a maximum at 250−280 nm. The position of this band depends on the nitrogen base or their sequence50,51 and is attributed to a π → π * electronic transition in the base. The absorption spectrum of poly(rC) at room temperature shows an absorption band at 268 nm (Figure 4A).51

Figure 4. (A) UV absorption spectra of the aqueous dispersion of GO (black) and of poly(rC) at room temperature (green, T = 20 °C) and 90 °C (red). (B) Differential spectra of poly(rC):GO-poly(rC) at different GO concentrations: 0.01 (black), 0.02 (red), 0.04 (green), 0.06 (blue), 0.08 (brawn), 0.1 (crimson), and 0.15 (cyan) mg/mL.

Since the absorption spectra of both poly(rC) and GO in the UV range overlap, the method of differential spectrum is employed to reveal the spectral manifestations of the interaction between the nanostructures. In the differential spectrum (obtained using four cuvettes in two channels; see Scheme S1), only changes in the poly(rC) spectrum that result from the addition of GO are showing. The differential spectrum does not show the effects of the poly(rC) dilution and of the increase in the absorption due to the addition of GO. As the poly(rC) aqueous solution is doped with GO suspension, a new band at 291 nm appears in the differential spectrum (Figure 4B). The intensity of this band is enhanced when the GO concentration increases. The appearance of the new band correlates with a decrease of the light absorption at 268 nm where the main band of nonbonded poly(rC) is located. Hence it is obvious that the observed spectral transformation is related to the poly(rC) adsorption on GO and to the poly(rC)−GO interaction. The red shift of the absorption band can be caused by hydrogen-bonding between the functional groups of cytosine and GO. If in the excited state an intermolecular hydrogen bonding is strengthened compared to the ground state, a red shift of the band in the absorption spectrum should be observed.52 Note that the band intensity in the differential spectrum is not more than 5% of the total 18225

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The Journal of Physical Chemistry C absorption band intensity of the poly(rC). There is some level of uncertainty in detecting such a small change in a broad band in the absorption spectrum. The small change of the total band intensity also indicates that only a small part of cytosine participate in the hydrogen bonding with GO in the aqueous suspension. When a single-stranded biopolymer is adsorbed to a singlewalled carbon nanotube (SWCNT), its helical structure becomes disordered because of the relatively strong interaction between the nitrogen bases (NBs) of the biopolymer and the πsystem of the carbon surface.8 Hughes et al.51 studied UV− visible absorption spectra of 30-base-long homooligonucleotides wrapped around SWCNTs in aqueous suspension and observed changes in the absorption spectra of the homopolymer in the UV range (from 200 to 300 nm). They explained the changes by anisotropic hypochromicity of the electronic transitions in the DNA bases. The absorption intensity of π-stacked compounds decreases upon electronic excitations because of the changes in the interactions between them in excited states50 (the so-called hypochromic effect53). Theory explains the hypochromicity as a result of weak dipole−dipole interactions between the dipole moment formed by an electronic transition in one molecule after light absorption and an induced dipole moment (its direction is opposite with respect to the formed dipole moment) in the stacked molecule.53 As a result of the interaction, the absorbance of the stacked dimer decreases in comparison with the combined absorbance of the two isolated systems forming the dimer. The well-known example is DNA. Its UV absorption decreases when the duplex structure is formed by two single DNA strands.50 For the DNA:SWCNT hybrid, a strong hypochromicity is observed for those electronic transitions in NBs whose induced dipolar moments align with the nanotube axis. For transitions whose transition moments are perpendicular to the nanotube axis, the hypochromic effect decreases.51 Note that the π−π stacking interaction between the nanotube and π-conjugated NBs was directly manifested not only in the DNA absorption spectrum but was also observed in the absorption spectrum of biopolymer-wrapped SWCNTs.54 It was shown that, at wavelengths longer than 300 nm, in the absorption spectrum of the polymer-wrapped SWCNTs the absorption intensity was lower than for nonbonded SWCNTs. The hypochromic effect is observed in the DNA absorption spectra when a NB self-ordering occurs, but when NB helical structure is disordered, the intensity of absorption bands increases (hyperochromic effect). The ordering/disordering of DNA is manifested with decreasing/increasing temperature of the biopolymer in solution (the so-called polymer melting effect). The dependence of the intensity of the poly(rC) absorption bands on the temperature is shown in Figure 5. The melting curve of poly(rC) shows ∼12% increase in the intensity of light absorption when the temperature rises from room temperature to 90 °C.23 At high temperature the band absorption is also red-shifted by ∼2 nm (Figure 4A). This effect is also caused by the NB disorder.55 The most significant changes in the melting curves are observed for duplexes, which has a S-like shape (the intensity can increase by 70%) while for single-stranded polymers the curve shows a monotonic growth with heating. The observations indicate an ordered structure of poly(rC) at room temperature. However, the same polymer shows very week hyperochromic effect in aqueous suspension with GO (the intensity increase with temperature is less than

Figure 5. Melting curves of poly(rC) (1) and poly(rC):GO (2) (measured at λ = 268 nm).

4%) (Figure 5, curve 2). The weakening of the effect is evidence of the adsorption of the polymer on the GO surface as well as of its disorder. Simulations of (rC)10 and (rC)30 oligomer adsorption on the graphene surface (presented in the next section) performed with molecular dynamics show that oligomer disordering occurs upon adsorption on graphene. As the polymer becomes disordered at adsorption on GO, it can be assumed that a part of the observed 22 nm shift of the poly(rC) band can be assigned to the transformation of the biopolymer conformation caused by adsorption. The small differential-spectrum intensity of poly(rC) is explained by only a small part of the NBs being involved in the hydrogen bonding with GO in the aqueous suspension. This part in the composite film significantly increases upon drying due to increasing of the space confinements between the components of the composite. That is why H-bonds play a more essential role in the FTIR spectrum of the poly(rC):GO film. 3.3. Computer Simulation of Cytosine Oligomer Adsorption on Graphene in Water Environment. The main purpose of the computer simulation of the oligonucleotide adsorption on graphene is to analyze the structure transformations of short and long oligonucleotides that occur as a result of the binding with the carbon surface and to estimate the binding energies. Another aim of the study is to elucidate the influence of the oligomer length on the rate of its achieving the most energy-favorable conformation on graphene. Such conformation corresponds to the maximum magnitude of the binding energy with the surface. In the simulations we use two oligomers with different lengths: r(C)10 and r(C)30. At the beginning of the simulations, an oligomer in the self-ordering helical A-form is placed near the graphene surface (Figure S3). During the simulation, the structure of the oligomer and the interaction energy between graphene and the oligomer are monitored. The interaction of r(C)10 with the carbon surface leads to the destruction of the cytosine self-stacking and its replacement by cytosine stacking with graphene. After 5 ns simulation, oligomer r(C)10 already has four cytosines stacked with graphene (Table S1). The adsorption of the oligonucleotide is accompanied by a sharp increase of the interaction energy (up to −110 kcal/mol; see Figure 6). During the following 10 ns of the simulation, the oligomer finds the most energetically favored conformation on the graphene surface. Note that, a stable ordered self-stacked structure of the oligomer prevents a structural reorientation which needs for the oligomer to acquire 18226

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nucleotide and the nanotube obtained earlier in MD simulation was about 13−15 kcal/mol.57−59 We also estimated the binding energy of the central cytosine (N5) with the two neighbor cytosines (N4 and N6) on simulation time (Figure S4). The binding energy of cytosine5 with cytosine6 was very weak for all simulation time while between cytosine5 and cytosine4 the value of this energy reaches in some steps up to 10 kcal/mol. The main contribution to the total binding energy provides H-bonds between carbonyl or amine groups of one cytosine with corresponding groups of the neighbor one (Figure S5). The Hbonding between two cytosines occurs during all simulation steps as well as dissociation of this bond in some steps. In whole, the binding energy of cytosine with graphene exceeds the interaction between neighbor cytosines (Figure S4). We also considered the influence of the cytosine binding process with graphene on the interaction of this cytosine with water molecules (the role of the solvation effects) limiting this consideration only the first hydration shell (ex., including only the strong bounded water molecules). There have been numerous quantum chemical calculation on the interaction between cytosine and water (see, for example,ref 60). The number of water molecules may be up to seven water molecules that can bind directly to cytosine.60 We determined the number of H-bonds formed by cytosine5 with water molecules at each step and showed their variation with time (Figure S6). This number of H-bonds was varied from 2 until 15 with time and the mean value was about 6−7 bonds (depending on the stacked cytosine number). In according with MP2 calculation for such number of water molecules (6−7) bound with cytosine via H-bonds, each energy of the bond is about 3 kcal/mol.60 Note that we did not reveal the notable change in the number of H-bonds after cytosine stacking with graphene. For some stacked cytosines, the mean value of H-bonds decrease up to 0.5−0.6 while for others cytosines we did not observe any change in the magnitude. This means that in spite of removal of water molecules from cytosine plain after base stacking with graphene, the cytosine−water H-bonds are not broken. This phenomenon can be explained by the arrangement of H-bonds near polar groups on the periphery of the planar structure of cytosine, which are not dissociated after stacking with graphene.61 A different picture is observed in the case of the r(C)30 adsorption on graphene. During the simulation, an adsorption of the oligonucleotide onto the carbon surface occurs through stacking of cytosines with graphene. However, in contrast to the short oligomer, only two cytosines become stacked with graphene in the first 5 ns of the simulation and this number increases to 4 at 10 ns (Table S1). At this time, the interaction energy reaches ∼−280 kcal/mol (Figure S7). It is obvious that by 10 ns the contribution of the π−π-stacking energy to the total binding energy is not the dominant contribution. The oligomer adsorption is accompanied by a gradual destruction of the cytosine self-stacked structure which provides the polymer with a quiet strong ordering at room temperatures with short single-stranded helical domain structure.62 After 20 ns of simulation of r(C)30, seven cytosines are stacked with graphene but five domains with self-stacked cytosines can still be found (Table S1). At that point, the binding energy of r(C)30 with graphene is about −300 kcal/mol (Figure S7). The oligonucleotide conformation on the carbon surface observed between 20 and 30 ns of modeling changes little and the interaction energy in the complex GR:r(C)30

Figure 6. Interaction energy between graphene and r(C)10 as a function of the MD simulation time.

a stacked conformation on graphene. Nevertheless, with time the ordered starting helical structure of oligomer r(C)10 gradually gets destroyed. At 15 ns the binding energy becomes equal to about −135 kcal/mol with 6 cytosines stacked with graphene (Table S1). During the remaining time of the simulation the conformation of r(C)10 on graphene changes little. Only one additional cytosine stacks to graphene and the interaction energy reaches −147 kcal/mol at 20 ns. For the last 10 ns of the simulation, the oligomer remains in the stable conformation with graphene (Figure 7).

Figure 7. Snapshots of the hybrid formed by oligonucleotides r(C)10 with graphene after 30 ns of the MD simulation.

We selected 7 nucleotides which have cytosines stacked with graphene and estimated the interaction energy of each selected nucleotide and cytosine with graphene in water surrounding. Our estimation shows that the average interaction energy of a single (selected) nucleotide with graphene in water is ∼−19.7 kcal/mol (closed to value calculated earlier for short purine oligomer adsorbed to graphene56), and the average binding energy of a single cytosine stacked with graphene in water surrounding is about −12 kcal/mol. Thus, we can conclude that the contribution of the π−π-stacking between cytosines and graphene to the total binding energy of r(C)10 is more than 56%. Note that the mean energy of the interaction between one 18227

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stable structures for the graphene-cytosine complex (called GRCYT A, B, C, and GR-CYT D in Figure S9) are identified. The lowest energy configuration (GR-CYT A) is shown in Figure 9

remains practically unchanged (Figure S7). Most likely, this is due to the fact that the number of bases that stack with the carbon surface also remains unchanged (Table S1). Note that the contribution of the π−π-stacking energy between cytosines and graphene to the total binding energy for this oligomer does not exceed 30% at 30 ns. Snapshots of the hybrid formed by oligonucleotides r(C)30 with graphene after 30 ns simulation are shown in Figure S8. In general, the adsorption of the longer oligonucleotide r(C)30 occurs slower compared to the adsorption of shorter r(C)10. This likely happens because of strong structure selfordering of longer oligomers, which prevents fast oligomer adsorption. In the case of r(C)10, there are no self-stacked cytosine at 10 ns while for r(C)30 five domains with self-stacked cytosines are observed even after 30 ns simulation (Table S1). Note that a larger number of atoms in the longer oligomer leads to a greater total binding energy with graphene than it was obtained for the short oligomer (Figure S7). It is interesting to compare the adsorption rates of the two oligomers different in length on graphene. To do that, the dependence of the averaged interaction energy per nucleotide for each oligomer is plotted in terms of the simulation time (Figure 8). This average energy increases slower with time for

Figure 9. Structure of the most stable configurations of graphenecytosine and graphene oxide-cytosine complexes calculated at the B3LYP(GD3BJ)/6-31++G(d,p) level of theory.

(left). The calculated interaction energies of the four GR-CYT configurations are shown in Table 1. In the calculations of the interaction energies, we used the standard counterpoise correction procedure to account for the BSSE correction.64 As it is seen from the table, the interaction energies obtained for the graphene-cytosine complexes are similar and vary from −19.5 to −17.1 kcal/mol. To test the accuracy of the B3LYP(GD3BJ) approach in prediction of interaction energies, additional calculations of the GR-CYT A complex for a smaller graphene fragment which contains 54 carbon atoms are carried out (Figure S10). These calculations are performed at the B3LYP(GD3BJ)/6-31++G(d,p) and MP2/6-31++G(d,p)// B3LYP(GD3BJ)/6-31++G(d,p) levels of theory. The resulting BSSE- and ZPVE-corrected interaction energies are −19.2 (B3LYP(GD3BJ)) and −17.9 kcal/mol (MP2). It should be noted that the ZPVE correction obtained in the B3LYP(GD3BJ) calculation is used to determine the MP2 interaction energy. The two interaction energies are close, but the B3LYP(GD3BJ) result perhaps slightly overestimates (in absolute value) the interaction energy. Total energies of all calculated complexes as well as ZPVE and BSSE corrections are collected in Table S2. For each graphene-cytosine complex, we also calculate harmonic frequencies and intensities. A graphical representation of the fingerprint region of the calculated IR spectra is shown in Figure S11. As it is seen, the calculated spectra of all complexes are similar. The shifts of the vibrational bands due to different stacking interactions are small and similar for all complexes. This demonstrates weak dependency of the spectra on the mutual orientation of graphene and cytosine molecules. In Table S3 (GR-Cyt A complex), the calculated shifts of the cytosine frequencies in the fingerprint region are shown. The maximal red-shift (−18 cm−1) is observed for the cytosine C O stretching vibration. For most of the cytosine vibrations, the shifts are within several wavenumbers. On the other hand, the interaction with graphene significantly changes the IR intensities of the cytosine vibrations. The calculated IR spectra of cytosine, graphene, and the GR-Cyt A complex are shown in Figure 10. The data demonstrates a strong decrease of the intensities of the cytosine vibrations in the 1800−1400 cm−1 region. For example, the intensity of the CO stretching vibration decreases from 774.1 km/mol in cytosine to 204.7 km/mol in GR-Cyt A. This can be explained by the high polarizability of graphene. The higly polarizable graphene πelectrons partially offset the changes in the dipole moment of cytosine associated with its in-plane vibrations and thus reduce

Figure 8. Dependences of the averaged interaction energy per a single nucleotide for r(C)10 (black) and r(C)30 bound with graphene on simulation time.

the long oligomer than for the shorter one. By 30 ns, the average interaction energy per nucleotide for r(C)30 is about −10 kcal/mol while for r(C)10 this energy is about −14.5 kcal/ mol. Different time dependences of the average interaction energy per nucleotide for short and long oligomers can be explained by the stronger self-ordering of the longer oligomer than of the shorter one. The stronger ordering hinders the acquisition of the energy-favorable conformation of oligomer on graphene and decreases the rate of oligomer adsorption. 3.4. Quantum-Chemical Calculations of the Graphene-Cytosine Complex. To search for the possible configurations of the graphene-cytosine complex ten initial structures of the complexes with different orientation of the cytosine molecule with respect to the graphene surface are selected. Here we took into account the results of previous calculations of the structure of cytosine complexes with graphene (see a review, ref 63 and the references cited therein). Selected structures are fully optimized at the B3LYP(GD3BJ)/6-31++G(d,p) level of theory. In total, four 18228

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Table 1. BSSE and ZPVE Corrected Interaction Energies (IE, kcal/mol) of Graphene-Cytosine and Graphene Oxide-Cytosine Complexes Calculated at the B3LYP(GD3BJ)/6-31++G(d,p) Level of Theory complex GR-Cyt GR-Cyt GR-Cyt GR-Cyt

IE

complex

−19.5 −18.8 −18.4 −17.1

A B C D

GO(222)-Cyt GO(222)-Cyt GO(222)-Cyt GO(222)-Cyt GO(222)-Cyt GO(222)-Cyt

IE

complex

−28.0 −27.5 −26.9 −24.9 −24.6 −24.5

A B C D E F

IE

GO(022)-Cyt GO(022)-Cyt GO(022)-Cyt GO(022)-Cyt GO(022)-Cyt GO(022)-Cyt

A B C D E F

−27.7 −26.8 −26.5 −24.9 −24.6 −24.4

in Figure 9. All calculated GO-cytosine structures are presented in Figure S12 and Figure S13 for GO(222)-Cyt and GO(022)Cyt systems, respectively. For each system, six stable configurations are identified. As it is seen in Figure S1, the configurations differ in terms of the location of the epoxy and hygroxyl groups in the front and back sides of the graphene fragment. This results in different interaction energies with cytosine (Table 1 and Table S2). A comparison of the interaction energies of cytosine with two different graphene oxide models (GO(222) and GO(022)) demonstrates very weak influence of the peripheral carboxylic groups on the interaction energy. Additional calculations are performed to separate the contributions of the hydrogen bonds and the stacking interactions to the total interaction energy between GO and cytosine. In the calculations, the structure of graphene oxide is modified by removing oxygen-containing groups. The position of the cytosine molecule relative to the graphene surface is not changed. For each modified structure, the interaction energy between graphene and cytosine is calculated. The calculations are performed with accounting for the BSSE correction. Since the modified structures are not optimized, the accounting for the ZPVE correction is not possible. With the removal of the oxygen-containing groups, the obtained energies (Table 2) are only due to the stacking interaction (IEST). The interaction energy due to the H-bonding (IEHB) is estimated for each complex as the difference between the total interaction energy and the stacking interaction energy. In this case the total interaction energy is recalculated with only accounting for the BSSE correction. One should note that, as a result of neglecting of the ZPVE correction, the total energies (IEST + IEHB) are not identical to the values shown in Table 1. The results presented in Table 2 show that, for all GO-cytosine structures, the stacking interaction contributes more to the total energy than the H-bonding. However, the energy of the stacking interaction for the studied complexes varies within a narrow range and thus the difference in the relative stabilities of the complexes of GO with cytosine is mainly dependent on the strength of the hydrogen bonds. Interaction energies for the graphene-cytosine and GOcytosine complexes were also calculated at the B3LYP(GD3BJ)/6-31++G(d,p) level of theory with accounting for

Figure 10. Fingerprint region of the calculated IR spectra of the graphene, cytosine, and GR-Cyt A complexes. Frequencies are scaled down by 0.98. Bands are approximated by a 40 cm−1 full-width-at-halfmaximum Lorentzian line shape.

the vibrational intensities. To elucidate how the size of the graphene fragment used in the calculations (and thus its polarizability) affects the intensities of the cytosine vibrations, additional harmonic frequency calculations are peformed for the complexes of cytosine with graphene fragments with the following sizes: C24H12, C54H18, and C150H30. The orientation of the cytosine molecule in the complexes is similar as in the GR-CYT A complex. The calculations show strong dependency between the IR intensity of the cytosine CO stretching vibration and size of the graphene fragments. The internsities are 371.2 km/mol for C24H12, 293.1 km/mol for C54H18, 204.7 km/mol for C96H24, and 157.9 km/mol for C150H30. The increase in the size of the graphene fragment is accompanied by the rise of its polarizability, which as it is seen leads to a decrease in the intensity of the CO stretching vibration. A similar decrease in intensity is observed for other in-plane vibrations of cytosine. 3.5. Quantum-Chemical Calculations of Graphene Oxide-Cytosine Complexes. Complexes of graphene oxide with cytosine are stabilized by both H-bonding and π-stacking. The lowest energy configurations of the GO-cytosine (GO(022)-Cyt A and GO(222)-Cyt A) complex are shown

Table 2. Decomposition of the Interaction Energies in Graphene Oxide-Cytosine Complexes (in kcal/mol) complex GO(222)-Cyt GO(222)-Cyt GO(222)-Cyt GO(222)-Cyt GO(222)-Cyt GO(222)-Cyt

A B C D E F

IEST

IEHB

−16.4 −16.4 −15.4 −15.0 −16.4 −15.9

−11.1 −10.6 −10.9 −10.4 −8.3 −8.7

complex GO(022)-Cyt GO(022)-Cyt GO(022)-Cyt GO(022)-Cyt GO(022)-Cyt GO(022)-Cyt 18229

A B C D E F

IEST

IEHB

−16.4 −16.3 −15.2 −15.4 −16.2 −16.2

−10.9 −10.4 −10.6 −10.1 −8.5 −8.4

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Figure 11. Fingerprint region of the calculated IR spectra of the graphene oxide (022 and 222), cytosine and GO-cytosine complexes.

graphene-oxide fragment results in a significant decrease of the ratio of the terminal carbon atoms to the total number of carbon atoms. This means that in the graphene oxide systems used in the experiment the number of peripheral carboxyl groups is significantly smaller than the number of the basal oxygen-containing groups, and as a result, the intensity of the absorption bands of the peripheral groups in the IR spectra is greatly reduced. In the complexes of GO with cytosine considered in the calculations, several H-bonds between the oxygen-containing groups of graphene oxide and the polar groups of cytosine are present (Figures S12 and S13). As a result, the calculated IR spectra of the complexes with different structures are significantly different (Figure S14). The calculated interaction energies for the complexes lie in a relatively narrow range between −28 and −24 kcal/mol (Table 1). Therefore, one can assume that in the experiment all possible structures should be present and the experimental FTIR spectra should be a superposition of the spectra of these structures. However, the question about the quantitative ratios among these structures remains open. A calculated averaged IR spectrum of all calculated structures is shown in Figure S15. This spectrum is compared with the experimental spectrum in Figure 12 in the 1800−1300 cm−1 range (see the spectrum in the 1800−700 cm−1 range in Figure S15). The comparison shows a reasonably

BSSE corrections and for thermal corrections to Gibbs free energies at 298.15 K. They are presented in Table S4. These data demonstrated that accounting for the thermal corrections at room temperature resulted in decrease of the interaction energies as compared to ones calculated with accounting for the ZPVE corrections (Table 1). At the same time both calculations predicted identical most stable configurations of the graphene-cytosine and GO-cytosine complexes. The calculated maximal value of the binding energy of cytosine stacked with graphene (−7.3 kcal/mol) is lower than binding energy of a single cytosine stacked with graphene (∼−12 kcal/mol) estimated from MD simulation as about −4.7 kcal/mol. The calculated difference in the binding energy of cytosine with graphene can be caused by the influence of the solvation effects considered at MD simulation.61 The IR spectra for all GO-cytosine complexes are calculated at the B3LYP(GD3BJ)/6-31++G(d,p) level of theory. Graphical representations of the spectra of the most stable GO(022)Cyt A and GO(222)-Cyt A complexes are shown in Figure 11. All calculated spectra of the graphene oxide(022)-cytosine complexes are presented in Figure S14. This figure shows the noticeable transformation of IR spectrum in the 1800−1400 cm−1 range in the dependence on the H-bonding type. As it is seen in Figure 11, the interaction of cytosine with GO leads to more significant shifts of some absorption bands of cytosine due to the formation of H-bonds as compared to graphene. In addition, as in the case of graphene-cytosine complexes, a significant reduction of intensity of cytosine vibrations in complex with GO is observed in the 1800−1400 cm−1 range. Calculated IR spectra of the GO models agree well with results of recent modeling of the GO IR spectra.43,65 The spectral changes are more clearly visible in the IR spectrum of the GO(022)-cytosine complex. As already noted, there are no peripheral carboxyl groups in the GO(022) model. These groups have strong absorption bands in the fingerprint region which overlap with bands of cytosine. Vibrations of the peripheral carboxyl groups dominate in the calculated spectra. They are not sensitive to the interaction of cytosine with the surface of graphene and the oxygen-containing groups of the base. In the fingerprint region of the calculated IR spectrum of the GO(222)-Cyt A complex (Figure 11), intense absorption bands of the carboxyl groups dominate. In the experimental IR spectra of graphene oxide, the intensity of the bands attributed to the carboxyl groups is much lower. This is because the size of the graphene oxide fragment which is used in the calculations is much smaller than the actual graphene fragments for which the experimental IR spectra are obtained. Increasing the size of a

Figure 12. Calculated spectrum (red) of GO-cytosine and the experimental FTIR spectrum of GO-poly(rC) (black) in the 1800− 1300 cm−1 range. 18230

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vibrations in the 1800−1400 cm−1 region resulting from the interaction with graphene is revealed. The decrease can be explained by the graphene high polarizability associated with its π-electron system. When cytosine is adsorbed to graphene oxide, their complex is additionally stabilized by H-bonding between the oxygencontaining groups of GO and the carbonyl or amino groups of cytosine. It leads to an increase of the red shift of the cytosine lines. For the cytosine-GO complexes it is possible to separate the contribution of the hydrogen bonding and stacking interactions to the total energy in the calculations. It is found that the energy of the stacking interaction for different complexes varies within a small range, and the difference in the relative stabilities of the complexes of graphene oxide with cytosine mainly depends on the strength of the hydrogen bonding. The estimation shows that the contribution of the Hbonding (two or three bonds) to the total binding energy of cytosine with GO (25−28 kcal/mol) reaches 35−40%. The analysis of the calculated IR spectra of model structures of GO differing in the ratio of oxygen containing groups (carboxylic:epoxy:hydroxyl) shows that the IR spectrum GO strongly depends on this ratio and the size of the GO flakes. The obtained spectral information about the adsorption of relatively long DNA/RNA strands on graphene oxide and the results of the MD simulations and the quantum-chemical calculations can be useful in potential applications of graphene nanomaterial such as the development of novel genosensors, nanosized scaffolds for drug delivery, and other applications in nanomedicine.

close agreement between the two spectra. Good agreement between band positions in calculated and observed spectra in the range 1700−1300 cm −1 is observed. The strong experimental bands at 1235 and 1094 cm−1 are mainly due to the vibrations of the poly(rC) sugar−phosphate backbone and they are absent in the calculated spectrum of the GO-cytosine complex. The intense band at 900 cm−1 in the calculated spectrum is due to CH out-of-plane vibration. The corresponding band in the experimental spectrum is much weaker. This may be explained by the significant size difference of the graphene fragment used in the calculations and the graphene oxide fragments present in the experiment. With an increase of the size of graphene, the ratio of the number of peripheral hydrogen atoms and the number of basal carbon atoms decreases significantly. This leads to a decrease in the relative intensity of the CH out-of-plane vibration. The calculated intensities and positions of the vibrational bands of GO can be used to assign the experimental IR spectra of the GO-cytosine system. For the different structures of this system obtained in the calculations, the stacking interaction of cytosine with the GO surface results in a significant decrease of the intensity of the cytosine bands in the 1800−1400 cm−1 region. Thus, this decrease is common to both graphene and GO complexes with cytosine. The H-bonding between the oxygen-containing groups of GO and cytosine molecules leads to additional spectral changes. These changes are found to strongly depend on the type of the H-bonding formed in the GO-cytosine complexes.

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CONCLUSION As follows from comparing the UV absorption spectra of free and bound synthetic homopolynucleotide (poly(rC)) and from the analysis of the spectra of these systems obtained using the FTIR absorption spectroscopy, relatively long biopolymers can be adsorbed to graphene oxide. The polymer adsorption is accompanied by the spectral shift of the electronic and vibrational bands of biopolymer and a change of the (poly(rC)) thermostability due to the biopolymer adsorption on GO. The observed spectral manifestation of the interaction between poly(rC) and GO can be for most parts explained by the results of the molecular dynamics simulation and the quantum-chemical calculations. MD simulations of the r(C)10 and r(C)30 adsorption on graphene demonstrates structural disorder occurring for both oligomers due to the π−π stacking of cytosine with the graphene surface. The simulations also show slower adsorption of a longer oligomer on GO than a short one. The slower adsorption of r(C)30 is explained by the stronger structure ordering of this longer oligomer that takes a longer time to be replaced by the arrangement of cytosines on the GO surface occurring in the adsorption process. The cytosine self-stacking provides the polymer with a quiet strong ordering at room temperatures with a short single-stranded helical domain structure. The van der Waals interaction of cytosine with graphene due to the π−π stacking provides the main contribution to the total binding energy. The MD estimation gives the average binding energy per cytosine stacked with graphene in water surrounding of ∼−12 kcal/mol while for its nucleotide the energy is ∼−19.7 kcal/mol. Quantum chemical calculation gives the binding energy of stacked cytosine with graphene as about −19 kcal/mol in vacuum. The calculated IR lines of the stacked cytosine with graphene are red-shifted by up to 20 cm−1 compared with free cytosine. The strong decrease of the intensities of cytosine

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jpcc.7b04806. Models of graphene and graphene oxide used in quantum-chemical calculations; FTIR spectra of GO, poly(rC), and composite poly(rC):GO films; snapshots of hybrid formed by oligonucleotides r(C)30 with graphene; calculated structures of the graphene-cytosine and GO-cytosine complexes; and calculated IR spectra of the graphene-cytosine and graphene oxide-cytosine complexes (PDF)

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AUTHOR INFORMATION

Corresponding Authors

*Phone: (380) 57 340-1595. Fax: (380) 57 340-3370. E-mail: [email protected]. *Phone: (380) 57 340-1595. Fax: (380) 57 340-3370. E-mail: [email protected]. ORCID

Victor A. Karachevtsev: 0000-0003-4580-6465 Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS The authors are grateful to Dr. A. M. Plokhotnichenko for the helpful discussion and help in the UV spectra measurements. This work has been partially supported by National Academy of Sciences of Ukraine (Grant N 15/17-H within the program “Fundamental Problems of the creation of new Nanomaterials and Nanotechnology” and Grant N 0117U002287). This 18231

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research was also provided by the grant support of the State Fund for Fundamental Research of Ukraine (Grant No. 73/892017). An allocation of computer time from the computational facilities of the grid-cluster at the Institute for Low Temperature Physics and Engineering and from UA Research High Performance Computing (HPC) and High Throughput Computing (HTC) at the University of Arizona is gratefully acknowledged.

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