J . Phys. Chem. 19!33,97, 10083-10091
10083
Mechanisms of Transmembrane Electron Transfer: Diffusion of Uncharged Redox Forms of Viologen, 4,4’-Bipyridine, and Nicotinamide with Long Alkyl Chains Leif Hammarstriim’ and Mats Almgren Department of Physical Chemistry, University of Uppsala, Box 532, S-751 21 Uppsala, Sweden Johan Lmd and Gabor Merhyi Department of Nuclear Chemistry, Royal Institute of Technology, S-100 44 Stockholm, Sweden Thomas Norrby and Bjiirn Akermark Department of Organic Chemistry, Royal Institute of Technology, S-100 44 Stockholm, Sweden Received: December 29, 1992; In Final Form: April 6, I993
Transmembrane electron transfer in lecithin (phosphatidylcholine) vesicles was studied by pulse radiolysis. CMV), cetylbipyridine (4Upon reduction, cetylmethylviologen (N-hexadecyl-N’-methyl-4,4’-bipyridinium (N-hexadecylpyridinium-4-yl)pyridine,CB), and cetylnicotinamide (N-hexadecyl-3-(aminocarbonyl)pyridinium, CNA) transferred electrons from the bulk water phase to Fe(CN)a3- in the internal water phase of the vesicles. The transmembrane electron transfer was found in all cases to proceed through diffusion of uncharged forms of the redox mediators (CMVO, CBO, and CNAO, respectively) but the kinetic behavior varied considerably. The mechanisms for CB and C N A were simple, the reaction following first-order kinetics, and the transmembrane diffusion was rate limiting (k = (1.5 f 0.3) X lo3s-l for CB and k = 3.2 f 0.5 s-l for CNA). The mechanism for CMV was more complicated, and the reaction followed second-order kinetics. The rate-determining step was proposed to be the disproportionation of two viologen radicals formed by the radiation pulse (2CMV+ CMVO CMV2+), followed by rapid transmembrane diffusion of CMVO and its subsequent reoxidation by Fe(CN)a3-. In pulse radiolysis, and in phosphorescence quenching experiments with Pt2(P205)sH&, CBO and CB+ were used as models in order to obtain the rates of transmembrane diffusion of CMVO and CMV+, respectively. Our results exclude the possibility of electron tunneling between viologens on opposite sides of the membrane, and they provide strong arguments against transmembrane diffusion of viologen radical (CMV+).
-
+
Introduction The concept of using microheterogeneous media for separation of redox products in systems performing artificial photosynthesis has attracted much attention for a long time. A vesicle can be used as a model of a thylakoid membrane and is therefore suitable for systems mimicking natural photosynthesis as well as nonbiological systems exhibiting photoinduced charge separation.14 Systems with vesicles offer many advantages compared to other microheterogeneous systems: (I) they have an extremely large surface area, compared to, e.g., BLMs, (11) they offer many potential variations since there are different microcompartments, Le., the hydrocarbon core, the interface (whose net charge can be fine-tuned by use of different vesicle-formingsurfactants and cosurfactants), and the bulk and internal water phases, and (111) there is a possibility of separating water-soluble compounds in the inner water phase from those in the bulk phase. The high complexity of vesicular systems is also a problem. Thus, although extensive research has been undertaken in this field, important features are still poorly understood and not under control. The viologens (N,N’-dialkyl-4,4’-bipyridinium, abbreviated C,,,#V) and their derivatives are among the most well-examined electronacceptors and redox mediators in artificial photosynthesis. In spite of this, the mechanism for viologen-mediated transmembrane electron transfer is still a matter of controversy: diffusion of the reduced viologen radical (Cn,n!V+),5-10electron tunneling between Cn,,+V+and Cn,~V2+on opposite sides of the vesicle bilayer,10-13 or a combination of those two mechanismsl0.1616 has been proposed. Electron tunneling has been suggested to occur between interface-bound viologens through the bilayer over a distance of at least 30 A, which is too far according to simple tunneling calculations, or between viologens 0022-3654/93/2097- 10083$04.00/0
that for some reason are permanently or transiently located deeper in the vesicle membrane, thus reducing the tunneling distance. We have recently’’ presented evidence, based mainly on stopped-flowexperiments,that excludes the possibility of electron tunneling between cetylmethylviologens (CMV) bound to the interfaces of egg lecithin (phosphatidylcholine)vesicles. Instead we proposed a mechanism controlled by a disproportionation (eq l), followed by transmembrane diffusion of the doubly reduced 2CMV’
-
CMVO
+ CMV2+
(1)
CMVO and subsequent reaction with an internal electron acceptor, Fe(CN)63-. The most important arguments were that the rate of transmembrane electron transfer from external CMV+ to internal Fe(CN)& was the same for vesicles with and without CMV on the inside and that it was second order in external CMV+, both facts excluding the possibility of electron tunneling. We now present results, based on experiments with pulse radiolysis, that confirm the earlier interpretations. We have used three different amphiphilic redox mediators (Figure 1): cetylmethylviologen (N-hexadecyl-Nt-methyl-4,4’-bipyridinium, abbreviated CMV), cetylbipyridine (4-(N-hexadecylpyridinium4-yl)pyridine, abbreviated CB), and cetylnicotinamide (Nhexadecyl-3-(aminocarbonyl)pyridinium,abbreviated CNA). All redox mediators were bound to the outer interface of the vesicles with the redox-active group exposed to the water phase and the alkyl chain probably intercalated in the hydrocarbon core (Figure 2). Fe(CN)63- was dissolved in the internal water phase and acted as electron acceptor. Since pulse radiolysis offers the possibility to vary the extent of reduction of the redox mediator in a convenient way, it was possible to investigate the dependence Q 1993 American Chemical Society
Hammarstram et al.
10084 The Journal of Physical Chemistry, Vol. 97, No. 39, 1993 -+”+-
-+” d
N
H
2
- N + Figure 1. The redox mediators used in this study (from top): CMV, CB, and CNA.
bipyridiniumdich1oride(CMVC12).l74-(N-methylbipyridinium4-y1)pyridine was obtained as an intermediate in the synthesisof the latter. Pt2(PzOs)4H&was synthesizedaccordingto reference 34. Ethanol, 99.5%, was used as supplied by Kemetyl, Sweden. Diethyl ether and dichloromethane were supplied by Kebo AB, Sweden, grade purum, and were distilled prior to use. All other solvents were of Aldrich HPLC grade unless stated otherwise. All other chemicals were of the highest commercially available purity and were used as received. Instrumentation. Absorption spectra were recorded on a Varian CARY 2400 spectrophotometer. Emission spectra were recorded on a SPEX Fluorolog 2 Series spectrofluorimeter. A Nz laser (LSI Laser Science, Inc., Model VSL 337ND) was used for the determination of excited-state lifetimes of Ptz(P205)4H&. Proton NMR spectra were run on Bruker AM 400 and ACF250 spectrometers. Chemical shifts are reported in parts per million downfield from TMS (d), in coupling patterns ABX and AA’XX’ as supported by the literature,lg and by simulations.20 Infrared spectroscopy was performed on a Perkin Elmer FT-IR 1725X. Melting point measurementswere made on a Gallenkamp melting point apparatus (uncorrected). TLC was performed on Merck aluminum oxide 60 F254 neutral (type E). Ion exchange chromatography was performed on Dowex 1-X8 (20-50 mesh, 1.4 mequiv/mL) anion exchange resin, freshly charged with chloride ions by treatment with 5 vol % aqueous HCl and then with deionized water until neutral washings were obtained. Column gel chromatography was performed on Aldrich alumina (neutral, 150 mesh, Brockmann activity type I). Synthesis. 44N-Hexadecylpyridinium-4-yl)pyridineBromide. A 2.7-g (9 mmol) sample of hexadecyl bromide (Merckp.a.) was added to 0.94 g (6 mmol) of 4,4’-bipyridine (Aldrich, 98%) in 100 mL of acetonitrile (99%Aldrich), and the mixture was heated at reflux for 24 h. The product, which precipitated upon cooling (2.26 g), was filtered off, washed with a small amount of cold acetonitrile in order to remove any unreacted starting material, air dried by suction, and placed in a Soxhlet extractor. Extraction with acetonitrile (250 mL, 3 h) transferred 4-(N-hexadecylpyridinium-4-y1)pyridine bromide into the acetonitrile solution, and any dialkylated product (N,N’-dihexadecyl-4,4’-bipyridinium dibromide) was retained in the Soxhlet filter. The acetonitrile was evaporated, and the product was dried at the pump. Yield: 1.57 g (3.4 mmol, 56%;purity 96%). The reaction and all workup steps were followed by TLC, eluent: ethyl acetate (15 parts by volume), methanol ( 5 parts), glacial acetic acid (1 part), water (1 part). 4,4’-bipyridine, Rf= 0.79,4-(N-hexadecylpyridinium4-y1)pyridine bromide Rf= 0.59, N,N’-dihexadecyl-4,4’-bipyridinium dibromide Rf 0.29. 1-H NMR (solvent mixture, 1:l by volume of CDCl3 and d6DMSO, with DMSO as internal standard at 2.49 ppm versus TMS): d 9.27 (2 H, H2 and H6, d, J = 6.7 Hz), 8.79 (2 H, H2’ and H6’, d, J = 6.0 Hz), 8.46 (2 H, H3 and HS, d, J = 6.7 Hz), 7.81 (2 H, H3’ and H5’, d, overlaps CDCI3 signal), 4.66 (2 H, NCHzCH2, t, J = 7.4 Hz), 1.94 (2 H, NCH,CH2-, m), 1.17 (-13 H, -(CH2)13-, m), 0.79 (3 H, 4 H 3 , t, J = 6.4 Hz). 4-(N-Hexadecylpyridinium-4-yl)pyridine Chloride. A 2.1-g (-4.5 mmol) sample from the previous step consisting mainly of slightly yellow 4-(N-hexadecylpyridinium-4-y1)pyridine bromide (96%, the remainder being N,N’-dihexadecyl-4,4’-bipyridinium dibromide) was dissolved in 50 mL of a 1:1 (by volume) mixture of deionized water and ethanoland passed through a 30-mL Dowex 1-X8 (20-50 mesh, =40 mequiv) anion exchange resin, which was further eluted with the same solvent mixture. Three fractions of about 40 mL each were collected, and the solvent was removed at the rotary evaporator. The resulting white solid was dissolved in absolute ethanol, filtered, and transferred to a smaller vessel, the solvent was evaporated, and the product was dried at the pump yielding a white precipitate, 1.7 g (4.1 mmol, 90%). Separation of the mono- and dialkylated products using a Soxhlet
-
Figure 2. Schematic picture of the vesicle systems used in pulse radiolysis, before any experiment is performed. For simplicity, the vesicle bilayer is depicted without indicatingthe individual lipid molecules or specifying the head-group region. The redox mediators are depicted with the hydrophilic head-group as circles and the alkyl chain penetrating the bilayer. The exact position of the head-group of each redox mediator relative to the head-groups of the phospholipids in the bilayer is not
known.
of the rate of reaction on the surface concentration of reduced redox mediator. Another advantage was the absence of external reductant during the transmembrane reaction, a circumstance that simplified the interpretation of the results, especially compared to systems where the reductant is an excited state.18 It was also possible to instantaneously reduce CB and CNA to great extents, in spite of the large negative value of their redox potentials. The structural similarity of CMV and CB made it possible toestimate the rates of transmembranediffusionofCMV+ and CMVO by measuring the rates for CB+ and CBO. The rates of transmembrane electron transfer were found to vary by several orders of magnitude for the different redox mediators, and we propose that the mechanism in all cases involved diffusion of the uncharged forms (CMVO, CBO, and CNAO). The large differences in the kinetic behavior show both the difficulties and the possibilities when designing systems of this type. Both a more detailed and a more general understanding of the factors controllingtransmembrane electron transfer is necessary in order to be able to judiciously design systems including more sophisticated molecular organization within the vesicles. In the text the redox state of the redox mediators are indicated in the abbreviations only when it is necessary to distinguish between them.
Experimental Section M8terials. Egg lecithin (first grade, Lipid Products, Nutfield, England) was used as received. 4-(N-hexadecylpyridinium-4y1)pyridine chloride (CBCL) was synthesized by a modification of the procedure described for N-methyl-N‘-hexadecyl-4,4’-
-
Mechanisms of Transmembrane Electron Transfer method” was not successful, and in order to obtain pure monoalkylated product, column gel chromatography was performed on Aldrich alumina (neutral, 150 mesh, Brockmann activity type I) eluted with a gradient consisting of acetonitrilemethanol (30-mL portions, containing 0, 2.5, 5, 10, 20 vol % methanol). A 250-mg sample yielded 195 mg of product (mp = 90-91 “C) which was subsequentlyused in thevesicleexperiments. TLC Rf= 0.45 in acetonitrile-methanol 80:20 (vol %). 1-H NMR (d4-methanol as solvent and internal standard at 3.31 ppmversusTMS): a9.15 ( 2 H , H 2 a n d H 6 , d , J = 6.9Hz), 8.84 (2 H, H2’ and H6’, AA’ portion of AA‘XX’ system, J u = J A X ~= +4.6 Hz, J A ~ X = Jmf= -1.7 Hz, or J u = JA~x’= -4.6 Hz, J A ~ X= JUT = +1.7 Hz), 8.52 (2 H, H3 and 5H, 3, J = 6.8 Hz), 7.99 (2 H, H3’ and HS’, XX’ portion of AA’XX’ system, J u = JAtx’ +4.6 Hz, J A ~ X= J u t = -1.7 Hz, or Jm = JA’X’ -4.6 Hz, JAX = J u t = +1.7 Hz), 4.67 (2 H, NCHzCH2, t, J = 7.6 Hz), 2.07 (2 H, NCH2CH2-, m), 1.28 (a13 H, -(CHz) 13-, m), 0.90 (3 H, -CH3, t, J = 6.6 Hz). N-Hexadecyl-3-(aminocarbonyl)pyridiniumIodide. A 2.0-g (1 6.4 mmol) sample of nicotinamide (3-pyridinecarboxamide, Merck, p.a.) and 6.3 g (5.6 mL, 18 mmol) of hexadecyl iodide (Aldrich 95%) were heated at reflux in a solvent mixture (60:40 by volume) of absoluteethanol and toluene for 72 h. The progress of the reaction was followed by TLC; the eluant (95:s by volume) 0.15, was dichloromethane-methanol. (Nicotinamide Rf hexadecyl iodide R p 0.84, product immobile). The crude product that precipitated from the reaction mixture upon cooling to ambient temperature was filtered off in a Biichner funnel and washed with diethyl ether. A slight yellowing of the product was observed upon drying. Since the crude product NMR revealed a small amount of unreacted nicotinamide, it was repeatedly recrystallized from ethanol until TLC confirmed absence of any starting material. Yield: 2.25 g (4.7 mmol, 29%). 1-H NMR (solvent mixture CDC13:d~methanol2:1by volume, d4-methanol as internal standard at 3.31 ppm versus TMS): a 9.66 (H2, s), 9.1 1 (H6, B portion of ABX system, JBX = 6.1 Hz, JBA=
