J . Phys. Chem. 1989, 93, 2510-2575
2570
Asymmetric Photogeneration of the Magnesium Octaethylporphyrin Cation in Vesicular Bilayers John F. Smalley,* Stephen W. Feldberg, Department of Applied Science, Brookhaven National Laboratory, Upton, New York 11973
and 'Steven H. Wool Plum Island Animal Disease Center, Greenport. New York 11944 (Received: June 22, 1988; In Final Form: October 13, 1988)
Three examples of inside/outside asymmetric behavior result from the flash photolysis of aqueous suspensions of small (-250 A in diameter) egg yolk phosphatidylcholine (PC) vesicles containing magnesium octaethylporphyrin in the presence of water-soluble electron acceptors (either methylviologen or ferricyanide): ( 1) Many more porphyrin cations are produced by reaction with the electron acceptor on the inside of the vesicle than by reaction with the electron acceptor on the outside. ( 2 ) The lifetime of the porphyrin triplet is much shorter when the electron acceptor is on the inside of the vesicle than when it is present only on the outside of the vesicle. (3) The lifetime of a porphyrin cation produced by reaction with an electron acceptor on the inside of a vesicle is much shorter than that of a porphyrin cation produced by reaction with this same electron acceptor on the outside. All of these observations are shown to be consistent with a model which presumes that there are two populations of the porphyrin within the vesicle bilayer on or near each surface and that the concentration of porphyrin associated with the inside vesicle surface is much higher than that associated with the outside vesicle surface. This model agrees with the one proposed by Tollin and co-workers for the distribution of chlorophyll a molecules in small PC vesicles.
Introduction There is currently a great interest in the study of heterogeneous electron-transfer processes at membrane interfaces. There are many reasons for this interest. Among these are that an understanding of these electron-transfer processes may be relevant to biological transmembrane electron transfer] as well as the possible (if not probable) application to the design and construction of solar energy conversion devicesS2 Several reports have noted that the photoinitiated processes occurring at the inner and outer surfaces of spherical vesicular membranes are significantly different. In general, examples involve porphyrin moieties (magnesium octaethylp~rphyrin~ or chlorophyll a4g5)present at presumably identical inner or outer surfaces of vesicular bilayers. For chlorophyll a (Chl), this asymmetric behavior manifests itself as significantly more Chl+ produced on the inner surface of an egg yolk phosphatidylcholine (PC) vesicle than on the outer surface upon flash photolysis of the Chl when a water-soluble electron acceptor (methylviologen, MV2+; ferri9,lOcyanide, Fe( CN)63-; 9,10-anthraquinone-2,6-disulfonate; anthraquinone-2-sulfonate; Fe(CN)5N02-;and 5-(2-methyl- 1,4naphthoquinonyl-3)glutathione) is present in comparable concentrations at either surface. With magnesium octaethylporphyrin (P) in small PC vesicles, striking asymmetric behavior is also noted in similar flash photolysis experiments. The production of P+ and the rate of back-reaction of P+ with the reduced acceptor (MV', Fe(CN)64-) are significantly greater on the inner surface of the vesicle. Furthermore, the rate of quenching (by either MV2+or ferricyanide) of the magnesium octaethylporphyrin triplet (T) is apparently much faster on the inner surface. This paper augments our preliminary r e p ~ r t .We ~ suggest that the asymmetry in the photochemistry occurring at the inner and outer vesicle surfaces arises because the inside vesicle monolayer contains most of the porphyrin. This explanation is also consistent with Tollin's observations of the behavior of chlorophyll a in similar vesicle^.^,^ ( I ) For example, see: Light-Induced Charge Separation in Biology and Chemistry; Gerischer, H., Katz, J. J., Eds.; Verlag Chemie: New York, 1979. (2) For example, see: Calvin, M. Acc. Chem. Res. 1978, 11, 369, and
references therein. (3) Smalley, J. F.; Feldberg, S. W.; Wool, S . H. Abstracts of Papers, 183rd ACS National Meeting; American Chemical Society: Washington, DC, 1982; PHYS 108. (4) Ford, W. E.; Tollin, G . Photochem. Photobiol. 1982, 36, 641. (5) Ford, W. E.; Tollin, G . Photochem. Photobiol. 1984, 40, 249.
Experimental Section Vesicles were prepared according to the procedures described by Barenholz et aL6 with some modifications to introduce magnesium octaethylporphyrin (P) into the bilayer. Five milliliters of chloroform containing 25 (or 50) mg of phosphatidylcholine (PC) was added to a test tube containing 0.4 (or 0.8) mL of methanol with 30-80 p g of P dissolved in it. The solution was then slowly evaporated to dryness under vacuum, and 5 mL of an aqueous buffer solution containing 0.010 M MOPS, 34Nmorpho1ino)propanesulfonic acid, at a pH of 7.2, was added to the test tube. If an electron acceptor, methylviologen (MV2+, 0.01-0.067 M) or potassium ferricyanide (16.7 mM), was to be present on both the inside and outside of the vesicles, it was added to this buffer solution, and the ionic strength of this solution was made equal to 0.1 1 M (or 0.17 or 0.21 M for [MV2+] > 0.033 M) with KCI. The test tube was then placed in an ice bath, and the suspension was subjected to six 5-min sonications using the maximum power of a Heat System Ultrasonics Model W-220F sonicator equipped with a Model 420 microtip, allowing at least 1 min for cooling between each sonication. The vesicle suspension was then centrifuged for 0.5 h at 1 X lo5 g and 3.5 h at 1.5 X IO5 g in a Beckman Model L2-65B ultracentrifu e in order to separate a monodisperse suspension of small (- 250 in diameter) vesicles from the polydisperse suspension resulting from sonication. When the MV2+ was to be present only on the outside of the vesicles, the ionic strength of the buffer solution prior to sonication was made equal to 0.1 1 or 0.21 M with KCI. An aqueous solution of the acceptor was added to the suspension after it had been ultracentrifuged. The ionic strength of this electron acceptor solution (either 0.11 M for [MV2+] = 0.033 M or 0.21 M for [MV2+] = 0.067 M) was the same as that of the salt (KCI) solution used to make the suspension, and it was also buffered at pH = 7.2 with 0.010 M MOPS. Vesicle suspensions prepared according to these procedures have an optical density of -0.005 at X = 633 nm. Since none of the species present are expected to absorb at 633 nm, we attribute this small optical density to the light scattering of the vesicles. If we assume that the vesicles are spherical with a radius less than 0.05X (Le,, 1.0 X M is (8.6 f 1.0) X lo4 s-l with the uncertainty being two standard deviations. At [MV2+]= 1.00 X 10-* M, this decay is a bit slower (Le., k , = (6.2 f 0.7) X lo4 s-I) because the reaction T
co
(4)
can no longer be considered to be instantaneous on the time scale of the experiment. The entire course of the decay of the long-lived radical ions produced on the outer surfaces of the vesicles could not be studied because of the small absorption changes and the slight instabilities in the intensity of xenon arc lamp producing the interrogating s) of the slow light.ls The initial portion (up to t = 1.6 X decay of these signals is quite complex, being neither pure first nor pure second order. Additionally, the spectrum of the long-lived transient is invariant u p to t = 1.6 X s, which indicates that the decay of the radical ions produced on the outside of vesicles is simply due to a reaction between P+ and MV+ at least up to (20) When only one ion pair can exist in a vesicle, then the ratio between PI (the probability that a vesicle has one ion pair) and n (the average number of ion pairs per vesicle) will be unity. If it is assumed that the distribution
of these ion pairs among the vesicles is described by Poisson statistics, then the probability that a vesicle has a certain number (i) of these ion pairs is given by Pi = de-"/i!. When i = 1, then P , / n = e'", which will only approach unity when n
