Donnan Potential Caused by Polyelectrolyte Monolayers - American

Aug 1, 2014 - With the salt concentration grown up to 1 M, the stretched DNA chains in low salt concentration are found to experience a gradual confor...
5 downloads 0 Views 1MB Size
Download PDF
Article pubs.acs.org/Langmuir

Donnan Potential Caused by Polyelectrolyte Monolayers Jing Zhang,† Yun Zhao,† Chun-Ge Yuan,† Li-Na Ji,‡ Xiao-Dong Yu,† Feng-Bin Wang,† Kang Wang,*,† and Xing-Hua Xia† †

State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering and ‡State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, 22 Hankou Road, Nanjing 210093, P. R. China S Supporting Information *

ABSTRACT: The Donnan potential is successfully isolated from ion pair potential on a ferrocene-labeled polyelectrolyte (DNA) monolayer. The isolated Donnan potential shifts negatively upon the increase in NaClO4 concentration with a slope of −58.8 mV/decade. With the salt concentration grown up to 1 M, the stretched DNA chains in low salt concentration are found to experience a gradual conformation relaxing process. At salt concentrations higher than 2 M, Donnan breakdown occurs where only the ion pair effect modulates the apparent potential. The apparent formal potential also shows strong dependence on solution pH, which reveals that the charge density in the polyelectrolyte monolayer plays an important role in the establishment of Donnan equilibrium.

1. INTRODUCTION Donnan potential appears at permselective membranes or boundaries where ion species in two separated ionic solutions maintain an unequal distribution. The pioneer work of Calvo group has provided an universal approach for measuring the Donnan potential in ultrathin layers using cyclic voltammetry.1 Lots of studies show that the establishment of Donnan equilibrium is a complex process and can be affected by various interactions.2−4 Although the concept of Donnan permselectivity has been successfully used in developing potentiometric sensors,5−7 the molecular mechanism of the Donnan equilibrium is not fully understood. For the study of electrochemical process and establishment of biosensors, self-assembled monolayer (SAM) is an ideal system because of its ordered structure, low mass transfer resistance, and consequently quick response upon chemical stimulation. By constructing SAMs of neutral alkylthiols with a redox species labeled on the top, the shift of apparent formal potential caused by the formation of ion pairs on top of the monolayer has been explored.8−12 The ion pair theory greatly improves our understanding of the composition of apparent formal potential. However, most biomolecules used for constructing monolayer, such as DNA and polypeptide, are charged and unlabeled polyelectrolyte.13,14 A remarkable difference of polyelectrolyte monolayer from that of alkylthiol is that the electrolyte ions from the solvent phase can selectively penetrate into the film, which form unequal distribution of ion within and outside of the monolayer. Therefore, the apparent formal potential is also affected by the Donnan potential raised from the polyelectrolyte monolayer. Exploring this new type of Donnan potential and its effect on © 2014 American Chemical Society

the electrochemical behavior of redox species on the polyelectrolyte monolayer can provide fundamental knowledge for understanding the biomimetic interfaces. Some research has discussed the effect of multicharged backbone of the polyelectrolyte monolayer on Donnan potential. Barton and collaborators analyzed the electrochemical behavior of methylene blue intercalated in DNA monolayer.15 The Levicky group simulated the distribution of ions in DNA monolayer and presented the relationship between the membrane potential and structural response of DNA monolayer to salt concentration.16 To the best of our knowledge, no research ever systematically analyzed or measured the Donnan potential caused by the polyelectrolyte monolayer. In the present study, we take DNA monolayer as a model system and use cyclic voltammetry to study the establishment of Donnan potential on a DNA polyelectrolyte monolayer. We build two monolayers using ferrocene-terminated DNA and morpholino (MO), respectively. MO is a neutral analogue of DNA with the backbone replaced by uncharged morpholine rings.17 Through comparing the formal potentials of ferrocene/ ferrocenium (Fc/Fc+) couple measured on both monolayers, the contribution of Donnan potential raised by the DNA monolayer is isolated from that caused by the ion pair effect, and the corresponding molecular mechanism is explored. Received: June 13, 2014 Revised: July 28, 2014 Published: August 1, 2014 10127

dx.doi.org/10.1021/la502320m | Langmuir 2014, 30, 10127−10132

Langmuir

Article

potential of Fc/Fc+ on electrolyte concentration and pH. The solution pH was adjusted from 2 to 6 by adding HClO4 in NaClO4 solution. (Under these conditions, the ionic strength of solution does not change appreciably with pH.) The liquid junction potential was almost the same during the measurements, and thus the small correction for liquid junction potential was neglected. For hybridization, the electrode that was modified with MO was immersed in 50 mM or 1 M NaClO4 solution, containing 100 nM FcDNA-target at room temperature without stirring. The time for surface hybridization is ∼1 h.

2. EXPERIMENTAL SECTION 2.1. Material. Oligonucleotides and Chemicals: Ferrocenemodified DNA was purchased from Takara Biotechnology (Dalian). MO was obtained from Gene Tools. The base sequence of thiolmodified DNA/MO and ferrocene-modified target DNA are as follows: ferrocene-AGCCTGATGTCGTCATAACTGA-SH (FcDNA), ferrocene-AGTTATGACGACATCAGGCT (Fc-DNA-target), and NH2-AGCCTGATGTCGTCATAACTGA-SH (MO). Ferrocene-modified peptide (Fc-LRRASLGGGGC) was purchased from Shanghai Apeptide (Shanghai, China). Protein Kinase A (PKA, catalytic subunit from bovine heart) and adenosine triphosphate (ATP) were purchased from Sigma-Aldrich. 6-mercapto-1-hexanol (MCH) and 1-hexanethiol (HT) were purchased from Aldrich. Sodium perchlorate monohydrate (NaClO 4·H2O), magnesium chloride hexahydrate (MgCl2·6H2O), perchloric acid, sodium fluoride, sodium nitrate, and sodium chloride were of analytical grade and used without further purification. All solutions were prepared with 18 MΩ deionized water (Millipore). 2.2. Instrumentation. All electrochemical measurements were performed with a CHI 660 E electrochemical system (CH Instruments, Shanghai) in a conventional three-electrode electrochemical cell. A platinum wire served as the auxiliary electrode, the modified gold electrode served as the working electrode, and an Ag/AgCl electrode served as the reference electrode. All tests were carried out at room temperature. 2.3. Methods. Preparation of ferrocene-modified morpholino: Ferrocene was covalently linked at the 5′ end of morpholino as an electroactive reporter. Both the synthesis process of N-hydroxysuccinimide (NHS) ester of ferrocene and the linkage reaction of ferroceneester with MO were previously described.18,19 The base sequence of ferrocene-modified MO is as follows: ferrocene-AGCCTGATGTCGTCATAACTGA-SH (Fc-MO). 2.4. Preparation of DNA/MO/Peptide-Modified Electrodes. Gold electrode was polished with 0.05 μm alumina slurry, rinsed with deionized water and then cleaned ultrasonically in ethanol and deionized water, respectively. Consequently, electrochemical cleaning with 0.5 M H2SO4 as the electrolyte was employed where the potential was swept from 0 V to +1.6 V (versus Ag/AgCl) for 25 cycles. Cleaned gold electrode was washed with deionized water and then immersed in 1 μM Fc-DNA, MO, or Fc-MO for 1 h with the presence of 1 M MgCl2 to eliminate the electrostatic repulsion among DNA strands. MO was prepared directly in pure water because no repulsion force existed among MO strands.18 The modified electrode was subsequently dipped in aqueous solution of 1 mM MCH for 0.5 h to remove DNA and MO physically absorbed on the electrode surface.19 After the modification steps, the electrode was rinsed with deionized water and electrolyte solution, respectively. The cleaning steps of peptide-modified electrode are the same with the DNA/MO-modified electrode. After thoroughly rinsing with deionized water, the cleaned electrode was immediately coated with 4 μL of 0.2 mM Fc-peptide solution and kept at 4 °C for 16 h to allow the self-assembly of Fc-peptide on the gold electrode. The modified electrode was subsequently dipped in 10 mM HT in absolute ethanol for 2 h to remove Fc-peptide physically absorbed on the electrode surface. After that, the electrode was rinsed with deionized water and electrolyte solution, respectively. 2.5. Peptide Phosphorylation on Modified Gold Electrodes. The PKA storing solutions were composed of 50 mM NaCl, 20 mM Tris-HCl buffer (pH 7.5, 25 °C), 1 mM EDTA, 2 mM DTT, and 50% glycerol. PKA reaction mixture contained 10 U mL−1 PKA, 0.2 mM ATP, and 10 mM MgCl2 in 10 mM Tris-HCl buffer (pH 7.5, 25 25 °C). The Fc-peptide-modified electrode was incubated in the previously described reaction mixture at 30 °C for 2 h. Then, the modified electrode was washed multiple times using 10 mM Tris-HCl buffer and deionized water, respectively. 2.6. Electrochemical Measurements. The electrolyte was NaClO4 or NaF solution containing 10 mM NaCl, which was used to stabilize the potential of Ag/AgCl electrode. Cyclic voltammograms (CVs) were recorded to analyze the dependence of formal redox

3. RESULTS AND DISCUSSION 3.1. Theory. Scheme 1 shows the ion and corresponding potential distribution in the electrode/monolayer/electrolyte Scheme 1. Ion and Corresponding Potential Distribution at (a) Fc-MO/Solution and (b) Fc-DNA/Solution Interface

system. It is assumed that all ferrocene covalently attached on the top of the MO/DNA monolayers lies in a common plane that we refer to as “plane of electroactive ions” (PEIs). The PEI bears positive charge only when ferrocene tags are electrochemically oxidized to ferrocenium. Inside this PEI, the potential drop depends on the charge property of the monolayer. On the electrolyte side of the PEI, both the ion pair of ferrocenium-counterion and diffuse layer dominate the potential drop. For a ferrocene-modified neutral MO monolayer, the distribution of ions at the Fc-MO/solution interface is relatively simple. Because the MOs are slight soluble in water (about millimolar solubility limit), the formed MO monolayers exist as desolvated films, which can hardly be charged. Therefore, one would not expect a significant Donnan potential at the Fc-MO/solution interface.18 The ion pair theory can be directly used to depict the redox potential shift of ferrocene at the interface.8−12 As is shown in Scheme 1a, once ferrocene (Fc0) is oxidized to ferrocenium (Fc+), the concentration of counteranions (X−) at the interface increases owing to the formation of ion pairs. The oxidization halfreaction of ferrocene and the formation of ion pairs (Fc+···X−) at the interface are written as eq 1. The apparent redox potential of Fc/Fc+ couple, φapp(MO), which is affected by the formation of ion pairs, is given by eq 2, where φ0 is the standard redox potential of the SAM-bound ferrocenes, Γ0FC and ΓFC+X− are the surface density of ferrocene and the formed Fc+···X− ion pairs, respectively, and αXsol− is the activity of anions in solution. K represents the formation constant of Fc+···X− ion pairs. For the negatively charged DNA monolayer, the concentration of cations inside the monolayer is significantly different from that in the bulk solution.16 As is shown in Scheme 1b, 10128

dx.doi.org/10.1021/la502320m | Langmuir 2014, 30, 10127−10132

Langmuir

Article

Figure 1. (a) Dependence of Ea of Fc-MO (squares) and Fc-DNA (circles) monolayers on the concentrations of NaClO4. Each point is an average of three time measurements. (b) Dependence of the isolated Donnan potential on the concentrations of NaClO4. The corresponding cyclic voltammograms for obtaining the Ea values were recorded in the potential window from 0 to 0.4 V for Fc-DNA monolayer and from 0 to 0.45 V for Fc-MO monolayer with a scan rate of 1 V/s. Concentration of NaClO4 from low to high: 0.05, 0.1, 0.2, 0.4, 0.6, 1, 2, 4, 6, 8, and 10 M.

both the Fc+···X− ion pairs and the distribution of Na+ contribute to the apparent redox potential of Fc/Fc+ couple. Donnan potential that was produced by the distribution of Na+ + + (ΔED) satisfies eq 3, where αNa(sol) and αNa(DNA) represent the ion activities of Na+ in solution and DNA monolayer, respectively. As a result, the apparent redox potential of Fc/Fc+ couple on the DNA monolayer (φapp,DNA) can be written as eq 4. Fc 0 ⇌ Fc+ + e−

φ0

Fc 0 + X − ⇌ Fc+X − + e−

φapp(MO) = φ0 + ΔE D =

K = ΓFc+X −/(ΓFc+α Xsol− )

Γ + − RT RT − ln Fc X − ln a X(sol) F F ΓFc0K

(1)

(2)

+ aNa(DNA) RT ln + F aNa(sol)

φapp(DNA) = φ0 +

represents two linear regions. From 2 to 10 M of NaClO4, the Ea shifts negatively with a slope of −55.1 mV/decade, which is close to the Nernstian value of −59 mV/decade predicted from eq 4. Such potential shift can be attributed to the formation of ion pairs between oxidized ferrocenium and ClO4−. Similar phenomenon has been previously reported.8,9,12 At lower concentrations from 0.05 to 1 M, the slope decreases to −47.1 mV/decade. Repeated measurements have assured that the observed slope change is not caused by experimental error. We attribute this decrease to possible structural stretch of MO chains in low salt concentration solution, which changes the dielectric constant on electrode surface and eventually partially offsets the effect of ion pairs. For Fc-DNA monolayer, the Ea also shifts with the variation of NaClO4 concentration, which gives two linear regions (labeled as circles in Figure 1a). From 2 to 10 M of NaClO4, the Ea shifts negatively with a slope of −61.3 mV/decade. Such slope shows no obvious difference from the Fc-MO monolayer. At NaClO4 concentrations lower than 1 M, the Ea gives a completely different slope of +11.7 mV/decade from the FcMO monolayer. As previously discussed, both the ion pairs effect and the Donnan potential contribute to the Ea in FcDNA monolayer. To isolate the effect of Donnan potential on Ea, we subtract the corresponding Ea of Fc-MO by the ones of Fc-DNA and plot the results in Figure 1b. It can be seen that the obtained ΔEa keeps constant at concentrations higher than + + 2 M, which reveals that the ratio of αNa(DNA)/αNa(sol) in eq 3 does not change in high concentrations of NaClO4. In other words, the DNA monolayer loses its permselectivity to ions when the charges on its backbones are screened by high salt concentration. This result falls in line with the theoretical prediction that DNA monolayer “collapses” at high salt concentration and the Donnan equilibrium breaks down simultaneously.1,16 It is worth noticing that ΔEa is not zero in high salt concentrations, which may be caused by the difference of desolvation ability for MO and DNA monolayers + + that then resulted in a different ratio of αNa(DNA)/αNa(sol). At NaClO4 concentrations lower than 1 M, the ΔEa shows a slope of −58.8 mV/decade, which perfectly matches the Nernstian estimation in eq 3. For now, it seems that the formation of Donnan potential in charged DNA monolayer has been clarified. However, a comparison of eqs 3 and 4 with Figure 1a,b gives a self-contradictory conclusion: The Nernst response of DNA monolayer shown in Figure 1b suggests a constant + αNa(DNA) in the DNA monolayer at low salt concentrations. The

(3)

aNa+ Γ + − RT RT RT (DNA) ln Fc X − ln a X(sol)− − ln F F F aNa+ ΓFc0K (sol)

(4)

In practical cyclic voltammetric measurements, it is hard to obtain the Donnan potential merely by recording the formal potential shift of ferrocene because of the coexistence of ion pairs on the DNA monolayer. A practical way is to measure the apparent potential of ferrocene on both DNA and MO monolayers with similar surface density, then deduct the terms of ion pair and formal potential. 3.2. Effect of Electrolyte Concentration on Donnan Potential of Charged Monolayer. Cyclic voltammetries of ferrocene attached on both DNA and MO were recorded in NaClO4 solution with a series of concentrations. (See Figure S1 in the Supporting Information.) Because of the good redox reversibility of Fc in all measurements, the anodic peak potentials (Ea) are used to represent the formal potential for convenience. The surface density (Γ) of the Fc-MO and FcDNA is estimated to be 5 ± 0.5 × 10−12 mol cm−2, corresponding to 33 ± 3 nm2/molecular. This coverage is much lower than that expected for a closely packed monolayer of ferrocene units (0.36 nm2/molecular), which indicates that the interaction between ferrocenes is much lower. As is shown in Figure 1a, Ea of Fc-MO and Fc-DNA monolayer is plotted versus the logarithm of electrolyte concentrations. For Fc-MO monolayer (labeled as squares in Figure 1a), the Ea shifts negatively with the increase in NaClO4 concentration and 10129

dx.doi.org/10.1021/la502320m | Langmuir 2014, 30, 10127−10132

Langmuir

Article

Figure 2. (a) Dependence of the Ea of Fc-DNA (circles) and Fc-MO (squares) on the concentrations of NaF. Each point is an average of three time measurements. (b) Dependence of the isolated Donnan potential on the concentrations of NaF. The measurement conditions were the same as those in Figure 1

Figure 3. Dependence of the Ea of (a) Fc-MO and (b) Fc-DNA monolayer on pH in NaClO4 solution with different concentrations. Concentrations from low to high: 0.05 (square), 0.2 (circle), and 1 M (triangle). The measurement conditions were the same as those in Figure 1 +

constant αNa(DNA) then results in a constant φapp,DNA in low salt concentrations according to eq 4, which conflicts with the nonzero slope measured on Fc-DNA, as is shown in Figure 1a. A possible explanation is that the stretched DNA chains in low salt concentration gradually collapses with the addition of

NaClO4, the isolated Donnan potential is still in accordance with the result obtained in NaClO4. This observation also shows that Donnan potential does exist across the charged monolayer and is independent of the ion pairs effect. 3.3. Effect of pH on Donnan Potential of Charged Monolayer. To explore the establishment of Donnan potential in the charged monolayer, a direct way is to modulate the charge density by controlling the solution pH. The charge density in a neutral MO monolayer should keep as zero under all pH values. As is shown in Figure 3, the dependence of Ea on pH is recorded in a series of NaClO4 solutions with different concentrations. For the Fc-MO monolayer, the Ea keeps constant within the pH range from 6 to 4.3, which is in accordance with our estimation(Figure 3a). At pH lower than 4, a positive shift of Ea is observed. Such a potential shift can only be attributed to the protonation of bases on MO chains under very low pH. It is known that the isoelectric points of four different types of DNA bases are around pH 2.3 to 4.2,22 which makes the MO monolayer positively charged at low pH, and thus the ion activities of ClO4− in the monolayer are larger than those in solution, which produces Donnan potential at the interface of the monolayer and solution, and the corresponding formal potential of Fc/Fc+ couple can be expressed as eq 5. Therefore, the protonation of bases on MO chains under low

+

NaClO4. The αNa(DNA) then slightly increases and leads to the deviation of Ea shift from the theoretical deduce. Several papers have proved the conformational change of DNA in different salt concentrations.20,21 It happens that MO monolayer also undergoes a similar collapsing process at low salt concentrations. By subtraction of eq 2 by eq 4, these two conformational change-induced fluctuation of activity of Na+ in monolayer are compensated. The previously described analysis also convinces us that the establishment of Donnan equilibrium on charged monolayer is a complex process and may include the conformational information on the monolayer. To confirm the previously described experimental results and theoretical hypothesis, we measured similar CVs in a solution of NaF. It is known that F− has the weak association ability with Fc+,11,12 which will greatly decrease the shift of Ea caused by the formation of Fc+···F− ion pairs. Figure 2a shows the dependence of Ea of Fc-MO and Fc-DNA monolayers on the concentrations of NaF. Because of the limited solubility of NaF in water, the highest salt concentration employed in these measurements was 1 M. It can be seen that the Ea of both the Fc-MO and Fc-DNA monolayers changes nonlinearly with the addition of NaF. After subtraction of the Ea of Fc-MO by the Ea of Fc-DNA, the obtained ΔEa shows a slope of −56.2 mV/ decade in concentration of NaF lower than 1 M (Figure 2b). In this case, although the existence of weak Fc+···F− ion pairs results in a distinct different slope from the result obtained in

−

pH that makes αClO4(MO) larger results in the positive shift of Ea. ΓFc+ClO4− RT RT ln ln aClO4(sol)− − F F ΓFcK aClO4(MO)− RT ln + F aClO4(sol)−

φapp(MO) = φ0 +

10130

(5)

dx.doi.org/10.1021/la502320m | Langmuir 2014, 30, 10127−10132

Langmuir

Article

Figure 4. (a) Cyclic voltammograms of MO-modified electrode upon hybridization with Fc-DNA-target in 0.05 M NaClO4 solution. (b) Relationship between Ea from Figure 4a and the coverage of surface-hybridized target DNA.

detection of DNA or other charged biomolecules along with the change of the charge density in the monolayer. 3.5. Application of Donnan Potential on Peptide Phosphorylation. On the basis of the molecular mechanism for Donnan equilibrium, we design a new potentiometric biosensor for the detection PKA. Scheme 2 shows the peptide

For the Fc-DNA monolayer (Figure 3b), the positive shift of Ea can be observed in the whole pH range in low salt concentrations (0.05 and 0.2 M). At high salt concentration (1 M), the positive shift of Ea becomes nonsensitive to the decrease in pH. In solutions with low salt concentrations, the positive shift of Ea starts from pH 6 can be attributed to the +

protonation of DNA backbones,23 which makes αNa(DNA) smaller. With the decrease in pH, the protonation of bases also contributes to the total potential shift. Therefore, the curves measured on the Fc-DNA monolayer in low salt concentrations have a different shape from the ones of the Fc-MO monolayer, and the extent of total potential shift is much larger than that in Fc-MO monolayer. With the increase in salt concentration, the negative charges on the DNA backbones become screened, which results in the smaller of the total potential shifts in the higher concentration. 3.4. Donnan Potential Caused by Surface Hybridization. On the basis of the previously described analysis, the charge density in the monolayer determines the formal potential of the redox species on the monolayer-modified electrode. We then employed the negatively charged complementary target DNA end-labeled with an Fc molecule (Fc-DNA-target) to hybrid with the neutral MO monolayer on an Au electrode to monitor the Donnan potential during the surface hybridization process. Figure 4a shows the CVs of MOmodified electrode during the hybridization with Fc-DNAtarget in 0.05 M NaClO4 solution. It is observed that Ea shifts negatively with the increase in the oxidization peak. On one hand, the increase in the peak current implies the increase in the coverage of surface-hybridized target DNA with hybridization time. On the other hand, Ea shifts negatively with hybridization time. When time is larger than 1 h, the hybridization reaches equilibrium, and thus the peak current and Ea keep constant. Direct plot of Ea versus the coverage of surface-hybridized target DNA (Figure 4b) gives a linear relationship. The peak shift reveals that the Fc-DNA-target gradually brings negative charges into the neutral MO monolayer during the hybridization process. As a result, the

Scheme 2. Peptide Phosphorylation on the Modified Gold Electrodes

phosphorylation on the modified gold electrodes. After the peptide phosphorylation (Fc-P-peptide), the monolayer becomes negatively charged because of the introduction of a phosphate group with two negative charges.24 Figure 5 shows

Figure 5. Cyclic voltammograms before (a) and after (b) the peptide phosphorylation.

+

αNa(MO) grows and the apparent redox potential of Fc/Fc+ couple decreases, as described by eq 5. For comparison, the shift of Ea during hybridization can hardly be observed in 1 M NaClO4 electrolyte (see Figure S2 in the Supporting Information), which confirms that the Donnan potential breaks down in high salt concentration. On the basis of these findings, it is promising to design new potentiometric biosensors for the

the CVs before and after the peptide phosphorylation. It is observed that the Ea shifts negatively after peptide phosphorylation. It reveals that the concentration of Na+ in the peptide monolayer grows after the introduction of a phosphate group, which leads to the apparent redox potential of Fc/Fc+ couple decreasing, as described by eq 4. 10131

dx.doi.org/10.1021/la502320m | Langmuir 2014, 30, 10127−10132

Langmuir

Article

(8) Rowe, G. K.; Creager, S. E. Redox and ion-pairing thermodynamics in self-assembled monolayers. Langmuir 1991, 7 (10), 2307−2312. (9) Redepenning, J.; Flood, J. M. Influence of Electrolyte Activity on Formal Potentials Measured for Ferrocenylhexanethiol Monolayers on Gold: Indistinguishable Responses in Aqueous Solutions of HClO4 and NaClO4. Langmuir 1996, 12 (2), 508−512. (10) Ohtani, M.; Kuwabata, S.; Yoneyama, H. Voltammetric response accompanied by inclusion of ion pairs and triple ion formation of electrodes coated with an electroactive monolayer film. Anal. Chem. 1997, 69 (6), 1045−1053. (11) Valincius, G.; Niaura, G.; Kazakeviciene, B.; Talaikyte, Z.; Kazemekaite, M.; Butkus, E.; Razumas, V. Anion effect on mediated electron transfer through ferrocene-terminated self-assembled monolayers. Langmuir 2004, 20 (16), 6631−6638. (12) Ju, H.; Leech, D. Effect of electrolytes on the electrochemical behaviour of 11-(ferrocenylcarbonyloxy) undecanethiol SAMs on gold disk electrodes. Phys. Chem. Chem. Phys. 1999, 1 (7), 1549−1554. (13) Sakata, T.; Miyahara, Y. Potentiometric Detection of Single Nucleotide Polymorphism by Using a Genetic Field-effect transistor. ChemBioChem. 2005, 6 (4), 703−710. (14) Wang, M.; Wang, G.-X.; Xiao, F.-N.; Zhao, Y.; Wang, K.; Xia, X.-H. Sensitive label-free monitoring of protein kinase activity and inhibition using ferric ions coordinated to phosphorylated sites as electrocatalysts. Chem. Commun. 2013, 49 (78), 8788−8790. (15) Ceres, D. M.; Udit, A. K.; Hill, H. D.; Hill, M. G.; Barton, J. K. Differential ionic permeation of DNA-modified electrodes. J. Phys. Chem. B 2007, 111 (3), 663−668. (16) Wang, K.; Zangmeister, R. A.; Levicky, R. Equilibrium Electrostatics of Responsive Polyelectrolyte Monolayers. J. Am. Chem. Soc. 2009, 131 (1), 318−326. (17) Summerton, J.; Weller, D. Morpholino antisense oligomers: design, preparation, and properties. Antisense Nucleic Acid Drug Dev. 1997, 7 (3), 187−195. (18) Tercero, N.; Wang, K.; Gong, P.; Levicky, R. Morpholino monolayers: preparation and label-free DNA analysis by surface hybridization. J. Am. Chem. Soc. 2009, 131 (13), 4953−4961. (19) Cao, L.; Zhao, Y.; Ji, L. N.; Zhang, J.; Wang, K.; Xia, X. H. The Enhanced Enzymolysis Resistance of Surface-Immobilized DNA Caused by Hybridizing with Morpholino. Electroanalysis 2013, 25 (4), 1074−1079. (20) Kaiser, W.; Rant, U. Conformations of end-tethered DNA molecules on gold surfaces: influences of applied electric potential, electrolyte screening, and temperature. J. Am. Chem. Soc. 2010, 132 (23), 7935−7945. (21) Baumann, C. G.; Smith, S. B.; Bloomfield, V. A.; Bustamante, C. Ionic effects on the elasticity of single DNA molecules. Proc. Natl. Acad. Sci. U. S. A. 1997, 94 (12), 6185−6190. (22) Izatt, R. M.; Christen, Jj; Rytting, J. H. Sites and thermodynamic quantities associated with proton and metal ion interactuion with ribonucleic acid, deoxyribonucleic acid, and their constituent bases, nucleosides, and nucleotides. Chem. Rev. 1971, 71 (5), 439−481. (23) Puppels, G.; Otto, C.; Greve, J.; Robert-Nicoud, M.; ArndtJovin, D.; Jovin, T. Raman microspectroscopic study of low-pHinduced changes in DNA structure of polytene chromosomes. Biochemistry 1994, 33 (11), 3386−3395. (24) Xu, X.; Liu, X.; Nie, Z.; Pan, Y.; Guo, M.; Yao, S. Label-free fluorescent detection of protein kinase activity based on the aggregation behavior of unmodified quantum dots. Anal. Chem. 2010, 83 (1), 52−59.

4. CONCLUSIONS Donnan equilibrium commonly exists across charged monolayers, and the resulting Donnan potential can be isolated from the contribution of ion pairs effect by comparing the apparent potential of a redox species on the charged monolayer with that on a neutral analog monolayer. Charge density of the monolayer and the salt concentration primarily modulate the Donnan equilibrium across the charged monolayer, but the effect of conformational change of the monolayer on Donnan potential also cannot be ignored. It is also found that the stretch of the negatively charged DNA monolayer produces a negative shift of the apparent potential. The molecular mechanism for Donnan equilibrium found in charged monolayer paves a way for developing new potentiometric biosensors.

■

ASSOCIATED CONTENT

S Supporting Information *

Cyclic voltammograms of Fc-MO modified electrode and FcDNA modified electrode in different concentrations of NaClO4 with a scan rate of 1 V/s. Cyclic voltammogrames of MOmodified electrode during the hybridization with Fc-DNAtarget in 1 M NaClO4 solution. This material is available free of charge via the Internet at http://pubs.acs.org.

■

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Tel: +86-25-83685947. Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS This work was supported by the grants from the National 973 Basic Research Program (2012CB933800), the National Natural Science Foundation of China (21190044, 21275071, 11179004, 31200583), the National Science Fund for Creative Research Groups (21121091), the Natural Science Foundation of Jiangsu Province (BK2011569), and the Program for New Century Excellent Talents in University (NCET-11-0237).

■

REFERENCES

(1) Calvo, E. J.; Wolosiuk, A. Donnan permselectivity in layer-bylayer self-assembled redox polyelectrolye thin films. J. Am. Chem. Soc. 2002, 124 (28), 8490−8497. (2) Xu, M.; Goponenko, A. V.; Asher, S. A. Polymerized polyHEMA photonic crystals: pH and ethanol sensor materials. J. Am. Chem. Soc. 2008, 130 (10), 3113−3119. (3) Vega, F. A.; Weng, L.; Temminghoff, E. J.; Riemsdijk, W. H. V. Donnan membrane technique (DMT) for anion measurement. Anal. Chem. 2010, 82 (7), 2932−2939. (4) Wu, Z.-Y.; Li, C.-Y.; Guo, X.-L.; Li, B.; Zhang, D.-W.; Xu, Y.; Fang, F. Nanofracture on fused silica microchannel for Donnan exclusion based electrokinetic stacking of biomolecules. Lab Chip 2012, 12 (18), 3408−3412. (5) Grygolowicz-Pawlak, E.; Crespo, G. A.; Ghahraman Afshar, M.; Mistlberger, G. n.; Bakker, E. Potentiometric Sensors with IonExchange Donnan Exclusion Membranes. Anal. Chem. 2013, 85 (13), 6208−6212. (6) Jagerszki, G.; Takacs, A.; Bitter, I.; Gyurcsanyi, R. E. Solid-State Ion Channels for Potentiometric Sensing. Angew. Chem., Int. Ed. 2011, 50 (7), 1656−1659. (7) Wipf, M.; Stoop, R. L.; Tarasov, A.; Bedner, K.; Fu, W.; Wright, I. A.; Martin, C. J.; Constable, E. C.; Calame, M.; Schönenberger, C. Selective sodium sensing with gold-coated silicon nanowire field-effect transistors in a differential setup. ACS Nano 2013, 7 (7), 5978−5983. 10132

dx.doi.org/10.1021/la502320m | Langmuir 2014, 30, 10127−10132