J. CHEM. SOC. FARADAY TRANS., 1994, 90(13), 1913-1922 Redox Properties of Ubiquinone (UQ,.) adsorbed on a Mercury Electrode Gabriel J. Gordillot and David J. Schiffrin Chemistry Department, The Donnan and Robert Robinson Laboratories, P.O. Box 147, The University of Liverpool, Liverpool, UK L69 3BX The electrochemical behaviour of ubiquinone 10 (UQ,,) adsorbed on mercury in contact with aqueous electro- lytes has been investigated at coverages smaller than a monolayer. Stability regions, acid-base ionisation constants and standard potentials for redox equilibria between conjugate stable species were determined and reaction mechanisms proposed. The standard potential for the ubiquinone/ubihydroquinone couple obtained was 0.276 V vs. SCE (-0.138 at pH 7).The values of pK, obtained for the two acid-base dissociation equilibria for the ubihydroquinone, 12 and 13.6, are higher than those predicted from the Hammett equation and lower than the value obtained in a low-relative- permittivity medium (80% w/w ethanol). The ubisemiquinone ion radical has been found to be stable at pH > 13.6 with a disproportionation constant of 0.4. The corresponding constant for the protonated radical was estimated to be showing the high instability of this form. This fact, together with kinetic considerations, suggests that for pH values lower than 12 the redox chemistry proceeds via dismutation. Two-phase transitions for the reduced and oxidised forms were observed. Ubiquinone (UQ) is known to be an important mitochon- drial redox component of the electron-transport chain.The molecule contains a quinone redox centre and a long iso- prenoid chain, and it has been suggested that its lipidic char- acter is responsible for its location 0 0 ubiquinone 10 in the hydrophobic domain within the phospholipid bilayers in cell membranes.'V2 The biological function of this coenzyme is to act as an electron carrier between membrane- bound redox enzymes. It has been proposed that UQ can be present in the mem- brane in two forms, associated with electron-transport com-plexes and in a pool containing an unbound form.3 Knowledge of the redox properties of UQ is essential for the understanding of its function. In common with other quin- ones, the reduction of ubiquinone follows a two-electron, two-proton reaction.In biological membranes it is now rec- ognised that the intermediate ubisemiquinone radical is sig- nificantly stable, as shown from electron paramagnetic resonance (EPR) spectro~copy.~ However, the values of the redox potentials corresponding to the UQ/UQ'-and UQ'-/UQH2 couples are still under disp~te.~-~ It is appar- ent that the various redox potentials measured are strongly dependent on the environment in which the quinone is present. For this reason, it is important to study electron- transfer reactions under well defined conditions. Classical electrochemical investigations have relied on the use of non- aqueous solvents, such as a~etonitrile.~ It is questionable, however, if the results obtained from aprotic solvent studies can be transposed to the membrane case, where, although the t Present address : Departamento de Quimica Inorganica, Analitica y Quimica Fisica, Universidad de Buenos Aires, Ciudad Universitaria, Pab.11, 1824 Buenos Aires, Argentina. coenzyme is present in a lipidic environment, water is avail- able to participate in intermediate chemical steps. The electrochemistry of UQ in aqueous solutions has been studied by Petrova and co-workers' who used thin films of the insoluble quinone spread on carbon electrodes by evaporation of its toluene solution. Schrebler et al. attached UQ to pyrolytic graphite by absorption from a benzene solution' and Takehara and Ide" studied charge-transfer reactions to UQ attached to glassy carbon by spin coating from an acetone solution.One of the problems with the pre- vious work has been the difficulty of controlling both the amount of electroactive species attached to the surface and the state of the carbon electrode surfaces employed. For these reasons a self-assembly technique based on the transfer of UQ adsorbed at the air/solution interface to mercury was employed in the present work. This method has been suc- cessfully used by Nelson" to attach monolayers of phospho- lipids on mercury. Experimental The adsorption technique consisted of transferring an Hg drop through a layer of UQ adsorbed at the air/solution interface. A hanging mercury drop electrode (HMDE) (Kouteckjr, Poland) was used for preparing the Hg surface prior to coating.The HMDE was mounted on a vernier screw clamp which allowed the Hg drop to be lowered slowly across the surface of the cell solution containing the spread ubiquinone. The electrode entered the cell through a silicone rubber septum that allowed movement of the capillary inside the cell. A three-electrode cell was employed; the counter electrode was a Pt gauze of 1 cm2 area and the reference electrode was a saturated calomel electrode (SCE). All poten- tials are given with respect to this electrode. The cell was jacketed and the temperature of the circulating water was controlled with a Grant W14 thermostat (kO.01"C) at 25 "C. The solutions were de-aerated with high-purity nitrogen (BOC) and a constant flow of gas was kept over the surface during measurement.The spreading solution was prepared from synthetic ubi- quinone 10 (Sigma). The spreading solvent was pentane (Fluka puriss p.a.) which was distilled before use in order to remove any non-volatile impurities. Before preparing the J. CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 solutions, the pentane was de-aerated; the solutions were 60 kept at -20°C. The purity of the solvent used was checked by measuring the double-layer capacitance of the mercury surface after 40 being lowered through the aqueous solution surface on which only the solvent had been spread. No difference between Hg N 20 drops extruded directly in the bulk of the solution and those previously made above the surface and then transferred across the air/solution interface was observed.The solutions were prepared from triply distilled water, E O O5.‘= -20 once from alkaline permanganate followed by distillation from dilute phosphoric acid and then by a final distillation. -40 Borax (BDH, AnalaR), K,HPO, and KH,PO, (Fluka, BioChemika) were used as received. -60 A Hi-Tek (England) DT 101 potentiostat and PPRl wave- -1.2 -0.8 -0.4 0 form generator were employed. The voltammograms at high sweep rate were recorded with a Phillips PM3302 digital E/v storage oscilloscope and at low sweep rates directly with an X-Y recorder (Phillips PM8277). Capacitance measurements were carried out with a PAR 5210 lock-in amplifier by mea- suring the in-phase and out-of-phase components of the current when a sinusoidal potential of & 5 mV was applied.Results Fig. 1 shows the general voltammetric behaviour of UQ adsorbed on mercury at different values of pH. For pH 26 f2--10 - three groups of peaks are observed; the two groups at the extremes of polarisation (11, 111, IV and V) are pH indepen- dent, whereas peak I shifts with pH. A characteristic sweep- rate dependence of the voltammograms and of the peak potential is shown in Fig. 2 and 3, respectively. At low sweep rates, the behaviour expected for a surface reaction pseudo- -1.2 -0.8 -0.4 0 capacitance is observed, but the lack of linearity at high EP sweep rates shows the onset of irreversible behaviour.This can be clearly seen from the sweep-rate dependence of the peak potential (E,). The values of E, are strongly pH depen- dent as shown in Fig. 4, but the peak separation between the 2030 t ’ ’ ’ anodic and cathodic scans tends to zero at very low sweep rates. The E, values extrapolated to zero sweep rate (E’’) depend on the pH (Fig. 5). Three pH dependences are observed; for 4.1 < pH < 12, dE”/dpH = 59.6 mV decade-’; for 12 < pH 5 13.6, dE”/dpH z 30 mV decade-’ and for pH 2 13.6, dE”/dpH w 0 mV decade- ’. Discussion Adsorption Capacitance and Reorientation Processes It is striking that two different types of voltammetric peak are -30 -I I I IVC 1 observed. At the extremes of polarisation, very sharp peaks can be seen.Since the corresponding peak potentials are independent of pH it can be concluded that reorientation processes are responsible for these. Similar processes have been discussed by Nelson and Beaton for phospholipids adsorbed on Mg.” The model proposed for this behaviour12.13 is based on the decrease in hydrophobicity of the Hg surface for increasing charge densities. This results in an increased interaction between the phospholipid polar head groups and the surface, leading to an inhomogeneous layer. As the potential is increased (both for potentials positive and negative with respect to the pzc), water displaces the lipid Fig. 1 Cyclic voltammetry of ubiquinone 10 adsorbed on mercury at different values of pH for a sweep rate of 0.1 V s-:(a)pH 4.5,O.l mol dm-3, H,KPO,, (.* .) base electrolyte; (---) adsorbed QU,o; (b)pH 6, 0.1 mol dm-3 mixture of HK,PO, and H,PO,) solutions; (c) pH 8.3, 0.1 mol dm-3 mixture of HK,PO, and H,PO, solutions; (6)pH 9.2,O.l mol dm-3 borax -1.2 -0.8 -0.4 EP 0 J. CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 0.8 0.4 Q 30 -0.4 ....... ........ ....... -0.8 . .,,.': ,. .. .. ...... -1.2, -0.5 -0.4 -0.3 4.2 -0.1 0 EP 30 20 ................................ 10Q 30 -1 0 -20 -30 -40 .... ...-50 .... I I 1 I I I I -0.7 4.6 -0.5 -0.4 -0.3 -0.2 -0.1 0 E/v Fig. 2 Redox peaks of UQ,, adsorbed on mercury at pH 9.2 (0.1 mol dm-3 borax). A, Sweep rates from (a) to (9): 0.3, 0.2, 0.1, 0.075, 0.05, 0.01 and 0.005 V s-'.B, Sweep rates from (a) to (9): 100,75, 50, 25, 10,7.5,5 and 2.5 V s-I. from the surface to produce discontinuous bilayer structures owing to its higher dipole moment. There are two distinct processes that can be predicted from a statistical thermodynamics model describing the changes of interaction energy of the different parts of the phospholipid molecules with the surface for different states of charge.12 As the interfacial field is increased, there is a competition for the increased polarity of the surface between the polar heads and the hydrocarbon chain. It has been proposed that the first peak at -0.9 V (us. Ag/AgC1/3.5 mol dm3 KCl) corresponds to a reorientation of the film, leading to a change in position of the polar headgroup.The second peak at more negative potentials corresponds to the formation of bilayer structures on the Hg surface, with the polar headgroups pointing both to the metal and to the solution. The experimental justifica-tion for this model is that the values of the interfacial capac-itance at potentials negative of the second peak corresponded to nearly half of those obtained for a lipid-free surface. At all pH values the capacitance at potentials more nega-tive than ca. -1.15 and more positive than ca. 0.06 V us. SCE, i.e. outside the reorientation peaks, is very close to that of a ubiquinone-free surface [see, for example, Fig. l(a)]. The sharp peaks at the extremes of polarisation are related to reorientation processes and no desorption is observed at least up to a potential of ca. -1.3 V, as shown by the repro-ducibility of the voltammograms for multiple scans.In the potential range between the reorientation and the redox peaks, the low value of the capacitance indicates strong adsorption of the molecule with an orientation parallel to the 1915 I I , I c;v (a ) 0-4 1 3 Q 6 3 .-2 0 1 0 0 1 2 3 4 5 vlv s-' I'I'I'I'I'I 30 20 15. .-lo 0 v/v s-' Fig. 3 Dependence of the peak current on sweep rate at pH 9.2 (0.1 mol dm-3 borax): (a) sweep rates under 5 V s-'; (b) over the entire range of sweep rates investigated. surface. For example, at pH 11, a minimum capacitance of 5.5 pF cm-2 is found at -0.6 V for a surface concentration of quinone of 2.6 x lo-" mol crnp2.Ignoring the small diffuse-layer contribution and considering full monolayer 0.00 -0.15 $ -0.30 -0.45 -0.6C -6.0 -2.0 2.0 6.0 In(v/V s-') Fig. 4 Dependence of the peak potential on sweep rate for different value of pH: (a) 5.3,(b) 7.0, (c) 9.2,(d) 13.3.Top curves correspond to the oxidation peak. J. CHEM. SOC. FARADAY TRANS., 1994, VOL. 901916 -0.1 I\ -0.2 % -0.3Lu -0.4 -0.5 -0.6 -11111,111,1 , coverage (see below), the thickness (d)of the adsorbed layer can be calculated from: d=-EEO C where E is the relative permittivity, c0 the permittivity of free space and C is the capacitance. From eqn. (l),and consider- ing E = 2 for a hydrocarbon, d = 0.3 nm.This calculation, of course, is affected by the surface coverage and indicates that the effective relative permittivity is greater than 2. The length of the ubiquinol molecule was found to be 5.6 nm.14 The trans double bond in the isoprenoid chain introduces rigidity in the hydrophobic chain. A molecular-modelling calcu-lation" confirmed this length and indicated an average transversal thickness of ca. 0.7 nm. This would correspond to a total surface reduction charge of the quinone for a 2e- process of 8.2 pC cm-2 for a close-packed monolayer, which should be compared with a measured charge of 5.1 pC cmP2. Considering a thickness of 0.7 nm for a flat orientation at full coverage," E x 4.3. This large value of the relative permit- tivity is probably related to insufficient close packing of the isoprenoid chains leading to areas of the Hg surface exposed to aqueous electrolyte.The higher polarisability of the double bonds in the isoprenoid chain compared with that of a satu- rated hydrocarbon is unlikely to be the reason for the appar- ently high value of E, since non-polar organic compounds have relative permittivities in the range 2-3. In order to distinguish between these possibilities, experi- ments in which the coverage by UQ,, was altered were carried out. Different coverages were obtained by altering the electrode area, i.e. changing the hanging Hg drop volume for a constant amount of adsorbed quinone. Fig. 6 shows the effect of area expansion on the cyclic voltammograms. The peak height and area of the redox process remain constant, but the capacitance baseline current increases linearly with electrode area (Fig.7). The model of the interface considered is that of a surface . which is partially covered by the quinone, with areas where water molecules have free access. This is the classical two- capacitor model used extensively for analysis of the capac- itance of adsorbed organic molecules for which the capacitance is given by:'6 C = OC,, + (1 -e)Co (2) where c is the capacitance per unit area, 6 is the coverage by quinone or quinol depending on the potential range analysed, F -0.2 1#1,1,1,111, -0.7 -0.6 -0.5 -0.4 -0.3 -0.2 E/v Fig. 6 Effect of drop expansion at pH 11 and at a constant amount of adsorbed UQ,, (2.5 x mol) on the cyclic voltammetry of UQ,, at v = 0.1 V s-'. Drop areas (a) 0.022, (b)0.003and (c) 0.039 cm2.and c,, and Coare the ubiquinone monolayer and base elec- trolyte capacitances, respectively. In terms of the area expan- sion experiments, it is covenient to express eqn. (2) as c = A,(&) -C,) + ACo (3) where C is the total measured capacitance, A is the electrode area and A,, is the surface area covered by the quinone when the amount of quinone present on the Hg surface is constant and determined by the transfer of the HMDE through the monolayer spread at the air/solution interface. From the results shown in Fig. 7 and eqn. (3), Co = 24.2 pF cm-2. The value of CuQwas calculated from capacitance measurements as a function of total surface coverage by quinone.These experiments were carried out by successive immersions of the mercury electrode through the quinone layer adsorbed at the air/solution interface and typical results are shown in Fig. 8. The capacitance reaches a nearly con- stant value of 5.1 pF cm-2 for a surface charge which is con- sidered to be due to a monolayer of quinone (QM)of ca. 5.2 pC cm-2; for increasing amounts of adsorbed quinone, the capacitance shows a small decrease with increasing coverage, probably due to the formation of surface aggregates. From eqn. (3) and the results in Fig. 7, A,, = 8.7 x lop3cm2. Con- sidering that the charge corresponding to the adsorbed 0.7 0.6 0.4 0.3 0.02 0.03 0.04 area/cm2 Fig.7 Dependence of the capacitance on area at pH 11 and E = -0.28 V on drop expansion with 2.5 x mol of UQ,, adsorbed on it J. CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 ~ ~~~~ 0 5 10 15 QudpCcm-2 Fig. 8 Dependence of the specific capacity on surface concentra- tions calculated from the charge involved in the redox process [reaction (I)] quinone is 0.05 pC for the experiment shown in Fig. 7, a monolayer charge of ca. 5.7 pC cm-, is obtained. This is in reasonable agreement with the value calculated from the results in Fig. 8. Furthermore, from eqn. (2), the slope dc/dQ,, for surface coverages of less than a monolayer, should be given by: (4) From the results in Fig.8, and taking QM= 5.2 pC cm-,, a value of c,, -coof -18.6 pF cm-, is calculated in good agreement with that calculated from the data in Fig. 7, of -19.1 pF ern-,. The similarity in the values obtained gives confidence in the two-parallel-capacitor model employed since different experimental approaches lead to similar calcu- lated quantities. The similarity of the capacitance values observed with those of the base electrolyte is due to the low surface area covered by the quinol molecule after reorientation. If the cathodic peak is due to a change in adsorption geometry from parallel to perpendicular to the metal surface, the area covered by the organic compound will be only small and related to the cross-sectional area of the quinol molecule. For a reorientation process for a packed monolayer from a paral- lel to a perpendicular orientation, the coverage will be given by the ratio of projected areas in the two orientations.For the quinol molecule this is approximately given by 0.7,/ 0.7 x 5.6 = 0.12. For Co x 20 pF cm-2, the measured capac- itance will be only 6% lower than that of the base electrolyte, in agreement with experimental results. Redox Peaks At low sweep rates, the UQ/UQH, reaction is reversible, as shown by the characteristic symmetry of the voltammetric waves (Fig. 1 and 2), a ratio of unity for the charge of the anodic and cathodic peaks, the linear dependence of the peak current on sweep rate (Fig. 3) and the constant value of the peak potential for different sweep rates.The peak half-width can be used for calculating the number of electrons transferred. From the theory of cyclic voltammetry for adsorbed compounds under ideal behavi~ur'~the peak current density (i,) is given by: . nFvQ 1, = -4RT 1917 Table 1 Dependence of AE,,,/mV on sweep rates for different values of the pH PH v/V s-' 6.0 7.0 8.3 9.2 10 11 12.4 13.5 0.005 50 49 51 31 51 51 --0.05 51 49 51 31 51 52 62 80 0.1 52 51 51 52 52 52 62 80 -0.5 58 60 54 56 55 54 -where n is the number of electrons, v is the sweep rate, Q is the total charge per unit area associated with the voltam- metric wave and the other symbols have their usual signifi- cance. The peak half-width (AEI,,) is given by: 90.6AE,,, = -mV; at 25°C n The redox processes will be analysed for the three pH regions shown in Fig.5. pH < 12 The limiting value of AE,,, at low sweep rates, measured to sweep rates down to 10 mV s-l or below, was 51 f2 mV between pH 11 and 6. Results for different sweep rates are given in Table 1. There are two ways to analyse these results; first, from eqn. (6) it can be considered that n = 2 and sec- ondly, that there is a non-ideal term representing lateral interactions between the adsorbed molecules to account for the slight difference with a theoretical prediction of AE,,, = 45.3 mV. The value of AE,,, was found to be independent of the degree of coverage by the quinone, from 8 = 0.2 to ca. 1. At first sight, this is an unusual result; if lateral interactions are effective in broadening the redox wave, it would be expected that AE1,, should decrease at low coverages.The voltammetric waves can be analysed following the treatment of Laviron18 and of Brown and Anson." The activity coefficients of the oxidised and reduced components of the adsorbed redox couple are given by:'9 70 = exp -l-0 + rOR rd (7) and YR = ~XP (rRR rR + PRO ro) (8)-where roo and TRR are the interaction parameters between molecules of the same type, whereas yOR and yRO are those between different species; Ti is the surface concentration of species i. The peak current is given by : (9) where rT is the total surface concentration of electroactive species (Q = t+rT), ro = roo -roR and rR = TRR -rRO.From the expression for the total current" and the surface Nernst equation : RT royoE=Eo+-In -nF (rR7a) the values of AE,,, as a function of the interaction param- eters can be derived. The expression obtained is: 2RT [In( e)AE,,, = --arT r]nF 1-a where a = -J( +) '1 1918 From eqn. (ll),the theoretical dependence of AE,,, on the product rTr can be calculated. Surprisingly, this relationship is almost linear and was found to be given by: AE,,, = 45.28 -27.426rTr (13) with a correlation coefficient of 0.9999. Following Brown and An~on,'~ the interaction parameters for quinone and quinol can be considered to be equal. The argument used for this assumption was the ifidependence of peak potential on surface concentration for a series of quin- ones.A similar situation was observed in the present work (see Fig. 6). Furthermore, the value of the interaction param- eters will be determined, to a large extent, by the long hydro- carbon unsaturated chain of the molecule, and therefore, differences in the interaction energies between the oxidised and reduced forms will be much smaller compared with unsubstituted quinones. Thus, the peak potential, given by:" E, = E'O -RTT-,-(r, -rR) 2nF should be independent of surface coverage, as is indeed observed. Also, note that the value of r = ro = rR calculated from eqn. (11) is -8 x lo9 mol-I cm2, which is five times larger than that calculated for 1,4-naphthoquinone adsorbed on graphite, reflecting the increased intermolecular inter- actions due to the isoprenoid chain present in the ubiquinone molecule.An alternative interpretation for the observed peak broadening is the existence of two well defined fast one-electron transfer reactions with close values of the standard potential and ideal surface behaviour. The analysis for this type of reaction has been carried out by Laviron." From their results it can be concluded that AE,,, should be strongly dependent on the equilibrium constant of dispro- portional, K, . This is given by: where, in the present case, E: and E: correspond to the stan- dard potentials for the formation of the ubisemiquinone and the quinol in the adsorbed state, respectively. If EY G E:, K, = 00 and the value of AE,,, is 45.3 mV.Following the analysis by Plichon and Laviron,,' the dependence of the peak half-width on K,, and hence on standard potential dif- ferences, is shown in Fig. 9. For the values of AE,,, found, of i 8o t > E..70 -%i 60 -50 - 1 I , I , I , I , I 1 -0.120 -0.080 -0.040 (f? -e)/V 0.0 0.040 Dependence of the half-width peak potential on the standard Fig.potential difference of two consecutive one-electron transfer reactions calculated from ref. 20 J. CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 I I 0.18 1' '4 0.12 - 0.06 - 3 0.00 - -0.06 - 4.12 - -0.18 b -I -0.75 4.65 4.55 -0.45 4.35 -0.25 EP Fig. 10 Effect of drop expansion at pH 12.4 and constant amount of adsorbed UQ (4.8 x mol).Area/cm-l (a) 0.015, (b)0.024 and (c)0.031; v = 0.1 V s-'. around 51 mV (Table l), the difference between the two potentials should be E! -EY = 61 mV, with Kd = 11. pH > 12 For pH values higher than 12 the shape of the voltammetric curves depends on the surface concentration, rT.Fig. 10 shows this effect, obtained by the expansion of a mercury drop with a constant amount of adsorbed UQ (r+.As can be seen, when the area is increased, the base capacitative cur- rents increase while i, decreases. Consequently, higher values of AE,,, are found at lower coverages. In the range 12-13.5, the limiting value of AE,,, obtained by lowering rT depends on the pH. This behaviour can be explained by taking into account that at pH > pK, the products of the reduction of UQ are charged species.Repulsive interactions between the adsorbed anions should be dominant and an activity surface coefficient greater than unity should be expected. If only this interaction is considered, two effects should be found: (a) a narrowing of the peaks when r is increased, and (b) AE,!, lower than the theoretical value of 45.3 mV. The first effect is in agreement with experimental results. However, a limiting value of ca. 80 mV has been found for AE,,, at pH > 13.6. This shows that AE,,, must be related to a two-step process with very close values of the formal potentials. pH-Potential Stability Diagram In order to analyse the E,-pH diagram (Fig. 9,the reactions that need to be considered are: UQ + 2Hf + 2e-eUQH2 (1) where Ki is the corresponding acid dissociation constant.As can be seen in Fig. 5, for pH > 13.5 no dependence of E, on pH is apparent. In this pH range the proton is not involved in the overall reaction. In all this analysis it is considered that at low sweep rates the oxidised and fully reduced form of the quinone follow a Nernstian equilibrium. Therefore, the reac- J. CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 Table 2 Formal potential of ubiquinone 10 in aqueous solutions at a!, + = 1 ;potentials us. SCE Table 3 Hammett constants used in the calculation of pK, of sub-stituted phenols for different positions of the substituent electrode condition Ea/V reference no.mercury 80% ethanol 0.272 7 pyrolitic carbon coated 0.267 9 glassy carbon coated 0.248 10 pyrolytic graphite coated 0.264 8 mercury coated 0.276 present work tion in this pH range is a combination of eqn. (15) and (16): UQ +2e-=UQ2-(VII) with a formal potential of E" = -0.495 V us. SCE. At pH values between 13.5 and 12, a slope dE,/dpH of 30 mV decade-' indicates that an overall 2e-, 1H' reaction occurs, which can be considered to be : UQ +H+ +2e-=UQH-(VIII) The formal potential of this reaction (for uH+ = 1) is -0.09 V us. SCE. For pH < 12, a 2e-, 2H' process is present and reaction (I) must be considered. The value of Eo (at pH 0) obtained from the results in Fig. 5 is 0.276 V us. SCE. The above results are in reasonable agreement with those obtained by other techniques, as shown in Table 2.In order to analyse the breaks in the E,pH dependence (Fig. 5), the ionisation constants of reaction (1V)-(VI) must be known. Since the changes in slope are a consequence of the change in reaction stoichiometry, the break points corre-spond to the pK of the ionisation of the reduction product, ubiquinol, from which pK,, = 12 and pK,, = 13.6. The differ- ence between the formal potentials of reaction (+) and (VII) is related to the dissociation constants through: From the E'' values (Table 2), pK,, +pK,, = 25.7, which compares favourably with the value obtained from the stabil- ity boundaries indicated above, of 25.6. Of course, no new information can be obtained from the E'' values calculated and the above discussion simply shows that the results quoted are self-consistent.It is difficult to compare the pK, values obtained with those reported in the literature. The pKs refer, in the present case, to adsorbed molecules. Morrison et uL7 obtained pK,, = 13.3 for ubiquinone dissolved in 80% ethanol-water mixtures, which was very large compared with that for other quinones. Petrova and co-workers' investigated the pH dependence of the electrochemistry of ubiquinone adsorbed on graphite and obtained pK,, =9.9. gi 0- m- P- ref. CH3 -0.13 -0.06 -0.17 21, 28 C2H5 -0.09 -0.07 -0.15 21, 28 OCH, 0 0.1 1 28 OH 0.04 0.13 -0.03 28 0- -1.1 -0.47 -0.66 28 QBr 0.68 0.70 0.37 0.39 0.23 0.27 21 21 NO* 1.24 0.7 1 0.78 21 The experimental pK, values can be compared with those predicted from linear Gibbs energy relationships, since there is no simple way of estimating independently the acid disso- ciation constants.The Hammett equation for the pK of sub-stituted phenols is:,' pK =9.92 -2.23Z0, (17) where CT~is the Hammett constant for substituent i. The values of ui used in the calculations are shown in Table 3. o for the isoprene chain was taken as 0.10 for the o-position, slightly higher than that of the ethyl group, but lower than that of -CH,. The reason for this is that the inductive effect of the first secondary -CH, group in the chain is bound to be small. The possible error due to this choice of -B is minor._~ Table 4 shows a comparison of calculations of ionisation constants of several quinols and of ubiquinol with measured quantities. The pKas of the various quinols are presented for comparison purposes only, in order to ascertain the reli- ability that can be ascribed to the Hammett equation. Most of the experimental results agree with predictions within f4%. There is a significant difference in the values of pK,, measured both in the present work and by Morrison et al. in aqueous ethan01,~ compared with those calculated from eqn. (17). In the latter case, the difference is most likely to be related to the low value of the relative permittivity of the 80% EtOH-H,O mixture used, of 32.8.22 The results obtained in the present work, although referring to an aqueous solution, are approximately two pK units greater than calculated.Since the electroactive species is adsorbed on the Hg surface, this result is not unexpected and probably reflects the reduced value of the relative permittivity in the interfacial region. The analysis of the pK is made more difi- cult by the expected n interactions with the metal surface, which would act, in this case, as equivalent to a substituent with a positive value of 6.The pKa values measured by Petrova and co-workers' are close to those predicted from eqn. (25). However, there is an uncertainty regarding the way parent compound p-benzoquinone 2-meth yl-p-benzoquinone duroquinone 2-bromo-p-benzoquinone 2-chloro-p-benzoquinone 2,6-dichloro-p-benzoquinone 2-nitro-p-benzoquinone UQ6 UQ9 UQio UQio UQio Table 4 pKa values for substituted p-benzoquinols experimental theoretical PKa, PKa 2 PKa, ~Ka2 ref.9.85 11.4 9.98 11.4 29 10.15 11.75 10.12 11.68 29 11.25 12.8 11.83 12.24 29 8.67 10.68 8.43 10.52 30 8.81 10.78 8.47 10.57 30 7.30 9.99 6.95 9.74 30 7.42 10.1 1 7.22 9.8 1 30 10.15 - 10.10 11.6 8 9.98 - 10.10 11.6 8 9.94 - 10.10 11.6 8 13.3 - 10.10 11.6 7 12.0 13.6 10.10 11.6 this work 1920 5.5 5.0 (LLa 4.5 4.0 -0.40 -0.32 -0.24 -0.16 -0.08 0.00 C%+m Fig. 11 Relationship between the pK, of substituted p-benzosemiquinones in aqueous solutions and the sum of the Ham- mett substituent constants, Coo+m.The linear correlations shown on the graph are pK, = 9.92 -2.33 (2.58 + Zoo+,,,)using a value of op of 2.58 for 0-.Parent compounds: (a)p-benzoq~inone;~~”~(b)2-methyl-p-benz~quinone;~~ 2,3-dirnethyl-p-benzoq~inone;~~(c) (6) 2,5-dimethyl-p-ben~oquinone;~~,~~*~’(e) 2,6-dimethyl-p-benzoquin-J.CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 and UQH’ + eeUQH-; Eiy = -0.014 (XI) Disproportionation Constants The disproportionation reaction of the radical anion is: 24‘-+ Q + Q2~ (XII) The corresponding constant, Kd2 , calculated from reaction (XII) and the difference between the formal potentials for reactions (11)and (111) is 0.4. This value shows that the radical anion is stable at very high pH.The disproportionation con- stant for the protonated semiquinone: 2QH’=Q + QH,; Kd, (XIII) can be estimated from : Kd, = Kd2 G - (18) Ka, Ka2 The value obtained, Kd, % 2 x loi4, indicates that at low pH the radical UQH’ is very unstable and that the overall process should occur via reactions (IX) and (XIII). one;23(f) 2,3,5-trimethyl-p-ben~oquinone;~~(9)dur~quinone.~~.~~ in which these results were obtained, since multilayers depos- ited on carbon were used. Therefore, it is likely that these results’ are not comparable with those obtained in the present work. The potentials for the various steps can be calculated from the estimated pK,, values and from those corresponding to the ubisemiquinone.From AE,,, obtained for pH > 13.5 and using the data in Fig. 9, E&--E&-= 24 mV. Consider- ing that i(EGQ.--E&-) = -0.495 V the following formal potentials have been estimated: E&.-= -0.483 V and EgWlP2--0.507 V us. SCE. In order to obtain a complete reaction scheme, the acid dissociation constant of the ubise- miquinone radical, KR, must be known. The pK, of semi- quinones has been found to be highly dependent on the Rate Constant and the Transfer Coefficient Deviations from the reversible behaviour become evident when the sweep rate is increased, and the cathodic and anodic peak potentials shift symmetrically from the reversible potential. The degree of reversibility depends on the pH and the reaction becomes less reversible at low pH.At higher pH, higher sweep rates are required to obtain the same peak separation. These effects are shown in Fig. 4. The theoretical analysis for linear sweep voltammetry of adsorbed layers has been developed by La~iron.~~ For the general reaction : Oads+ ne-Rads (JW and when adsorption obeys the Langmuir isotherm, the surface formal potential is defined by: polarity of the solvent. For example, Patel and Will~on~~ have determined pK, between 4.9 and 5.1 for durosemiquin- one in aqueous solution of isopropyl alcohol (1 mol dmP3) and acetone (1 mol drnp3), while in aqueous isopropyl alcohol (7 mol dm-3) and acetone (1 mol drnp3) a value of pK, = 6 was found. For UQH’, Land and Swallow24 have obtained pK, = 6.5 in methanol solutions while Patel and Will~on,~found a value of 5.9 in aqueous solutions of isopro- pyl alcohol (7 mol dm-3) and acetone (1 mol dm-3).Owing to the aromatic character of the semiquinones, it is possible to use the Hammett equation to estimate the value of pK,. Fig. 11 shows a very good correlation between predictions and measurement if a value of bp of 2.57 is assigned to the substituent 0’for the pKR determined in aqueous isopropyl alcohol (1 mol drnp3) and acetone (1 rnol dmP3). From this equation a pKR of 4.4 is obtained for UQ,, . This is very low, but shows the limiting value expected in a solution with a relative perimittivity close to that of water. As has been discussed above, the relative permittivity at the interface may be quite different when compared with the corresponding value for the bulk solution.Similar to the analyses of pK,, the pK, for the system under study should be between 6.5 and 4.4. Taking pK, as 5.5, the formal poten- tials (aH+= 1, us. SCE) for the following reactions were esti- mated : UQ + H+ + e-eUQH’; EL: = -0.157 (IX) UQH’ + H+ + e-eUQH,; EL: = 0.710 (X) where b, and b, are the adsorption coefficients of 0 and R. There is no adsorption equilibrium in the present system since ubiquinone is practically insoluble in aqueous solutions, but a similar mathematical formalism can be used to analyse the system under study. The current is given by: i = -nFAX,/at = nFAk’* x {r,exp[-af(E -EO)] -rRexpC(1 -ct)f(~ -EO)]) (20) where k“ is the rate constant, A is the area, ct the transfer coefficient,f= F/RT, To and rR are the surface concentra- tions of 0 and R, respectively, and the other symbols have their usual significance.Considering that To + rR is constant, eqn. (20) can be numerically integrated and different param- eters correlated. For reversible behaviour AEp = E,, -E,, is zero (Ep, and E,, are the cathodic and anodic peak potentials, respectively). When this difference is larger than 0.2 V the theory predicts totally irreversible behaviour. In the present reaction and for all values of the pH, a transition between these two cases is observed when the sweep rate is increased. The value of x can be determined from the potential dependence of the ratio R = (Epa-Ep)/(Epc-Ep).25R was found to be independent J.CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 0.20 0.15 < L' 0.102 0.05 0.00 0 10 20 30 40 50 vp s-' Fig. 12 Observed dependence of nAE, (n = 2) on sweep rate for dif- ferent values of pH: (*) 7.0, (A)8.3, (0)9.2, (e)10.5, (0)13.3 of potential and equal to one, which corresponds to a = 0.5. When nAE, is greater than 0.2 V, the reaction can be con- sidered totally irreversible. In this case, the transfer coefficient can be obtained from:25 d AE, RT -d log v 2.3n,aF where n, is the number of electrons involved up to the rate- determining step. From Fig. 4, a value of an, x 0.5 is obtained at all pH values, indicating that only one electron is transferred up to the rate-determining step.The rate constant can be determined from the dependence of BE, on the reduced sweep rate given by l/m = nFv/RTk", when nAE, c 200 mV.25 The theory of quasi-reversible surface reactions2' predicts a generalised relationship between nAE, and l/m for each value of a. This is a simple and powerful approach for the calculation of rate constants. Fig. 12 shows the dependence of nAE, as a function of the sweep rate for different values of pH. In order to put the results on a common basis, a value of a = 0.5, as previously found, was considered and a best fit was found for the results at any value of the pH by fitting to the theoretical curve.,' The result of this generalised plot is shown in Fig.13, where it can be seen that the data at different pH values are consis- tent with theory. In order to compare the results at different 0.20 0.15 a2 0.10 0.05 P 0 5 10 15 m-' Fig. 13 Experimental results (Fig. 12) fitted to the theoretical curve (solid line) for the dependence of nAE, on the reduced sweep rate, m-' = ~FV/RT~''.'~ 9.2,(0) 10.5, pH: (+) 5.3, (*) 7.0, (A)8.3, (0)(0)13.3. 3.0 F-2.0 IIn1 0 s 0, 2 1.0 0.0 5.0 7.0 9.0 11.0 13.0 PH Fig. 14 Dependence of the rate constant k" on pH. Results from the data shown in Fig. 12 and 13. pH and calculate the rate constants, values of the sweep rate for the points plotted on Fig. 12 were multiplied by a factor x at each pH, such as to obtain the best fit to the theoretical curve of nAE,,, us.m-l. Since l/m = nfv/RTk", the corre- sponding k" was obtained from x = nF/RTk". The corre- sponding pH dependence of the rate constants thus calculated is given in Fig. 14. At low pH values, a linear dependence is observed, whereas in highly alkaline solutions, the rate constant appears to reach a constant value. The pH dependence indicates the involvement of the proton prior to the rate-determining step. Furthermore, from the slope in acid solution, k" a [H']o.5. This corresponds to a le-, 1H' reaction leading to reduction to the quinol. The proposed sequence corresponds to reaction (11)followed by : UQ'-+ H+(H,O)= UQH'( +OH-) (XV) The disproportionation of the radical anion is expected to be slow owing to electrostatic repulsions.The reaction order with respect to the proton suggests that the rate-determining step is the deproportionation reaction : UQH' + UQ'-+ UQH-+ UQ (XVI) since the value of the pK, of QH' is ca. 5.5, the surface cover- age by QH' is always smaller than that of Q*-, and therefore the disproportionation reaction UQH' + UQH' + UQ + UQH, (XVII) will be much slower than reaction (XVI). In alkaline solutions, where the surface concentration of QH' becomes vanishingly small, the reaction cannot proceed according to a disproportionation pathway and the rate becomes determined by electron transfer according to reac- tion (11) followed by (111). This reaction pathway leads to a pH-independent rate constant in the alkaline range, which is indeed observed.G.J.G. gratefully acknowledges the support of CONICET (Consejo Nacional de Investigaciones Cientificas y Tecnicas de la Republica Argentina). References 1 M. Ondarroa and P. J. Quinn, Biochern. J., 1986,240, 325. 2 M. Ondarroa and P. J. Quinn, Eur. J. Biochem., 1986,155, 353. 3 C. I. Regan and C. Heron, Biochern. J., 1978,178,783. 1922 J. CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 4 5 6 7 8 9 10 11 12 T. Ohnishi, J. C. Salerno, H.Blum, J. S. Leigh and W. I. Ingle- dew, in Bioenergetics of Membranes, ed. L. Parker, G. C. Papa- georgiou and A. Trebst, Elsevier, Amsterdam, 1977, p. 209. P. F. Urban and M. Klingenberg, Eur. J. 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