NOTES
3012
listed in Table I and calculated from eq 1 may be subject to systematic errors arising from the temperature dependence of &*. Thus, for precise work, our results must be accepted as effective values of &* valid at the quoted temperature difference and mean temperature.
The Methylene Blue-Ferrous Iron Reaction in a Two-Phase System‘
Downloaded by FLORIDA STATE UNIV on September 13, 2015 | http://pubs.acs.org Publication Date: September 1, 1966 | doi: 10.1021/j100881a507
by D. Frapkowiak and E. Rabinowitch Department of Botany and Department of Physiology and Biophysics, University of Illinois, Urbana, Illinois (Received February 26, 1966)
The photosynthetic apparatus of living plants separates from each other the probably highly unstable, intermediate oxidation and reduction products of the primary photochemical process and thus permits their conversion to (relatively) stable final products, carbohydrates and oxygen. This is probably accomplished by conveying these intermediates into different phases in the lamellar structure.2 The failure to imitate in vitro the storage of light energy as chemical energy, achieved by plants in photosynthesis, may be due primarily to not providing such a separation mechanism. Nathai and Rabinowitch3 showed that an in vitro system can be constructed, in which the products of an oxidation-reduction reaction (that between thionine and ferrous ions), which runs in light against the gradient of chemical potential, are separated by distribution between water and ether in an emulsion. The light energy stored in this way can be liberated by permitting the two phases to mix again (e.g., by adding methanol). In continuing this work, Ghosh4 in this laboratory observed that separating is more effective if methylene blue is substituted for thionine; perhaps, more of the neutral species of the leuco dye is present in methylene blue than in thionine a t the pH values used. (This species alone is likely to be involved in the extraction of the leuco dye into ether.) The following experiments deal with the methylene blue-ferrous iron system. They provide some additional information concerning the effectiveness of the separation. The reaction vessel used was a cylindrical Pyrex-glass vessel containing 30 ml of aqueous solution and 15 ml of ether (or another solvent immiscible with water). The approximate initial concentrations of methylene blue and FeS04 were of the order of 10-5 and 5 x low3 The Journal of Physical Chemistry
M , respectively. The mixture was stirred by bubbling through a stream of purified argon. No buffer was used; the pH was 3.5-4. The light used for illumination was either white light from a 1000-w coiled filament incandescent lamp or the same light filtered through an interference filter (Balzer K-6) with maximum transmission of 650 mp (near the peak of the methylene blue absorption band at 664 mp). The experiments were made at room temperature maintained in a water bath. Merck reagent grade methylene blue was used without purification. Chromatography on aluminum oxide and the test of Bergmann and O’Konski5 indicated that the dye was pure enough for our purpose. We tried out several combinations of immiscible solvents, but the most efficient separation of the photoproducts we were able to obtain took place in the system water ethyl ether, already used in Rlathai’s work. The reaction is
+
light
NIB
+ %Fez+
dark
(S + L)
+ nFe3+
Here, MB is methylene blue, S, the semiquinone, and L, the leuco methylene blue; n is between 1 and 2, depending on relative amounts of S and L formed. From the work of Parkera and the earlier work of Rabinowitch,’ it appears that in the photostatior?ary state, partly reduced solutions of methylene blue contain only a small percentage of semiquinone, so that n is close to 2. The predominant ionic species (at the prevailing acid pH’s) are monovalent positive ions of MB and monovalent (or divalent) positive ions of L. The neutral form of L, which we can assume to be the only one soluble in ether, is present only in minute amounts. This probably accounts for the slowness with which the leuco dye is extracted into ether. Further experiments on the effect of pH on the rate of extraction should clarify this point. I n the stationary state, the extent of extraction de(1) This research was supported by research grants from the National Science Foundation (GB 1946 and GB 3305) and from the Atomic Energy Commission [AT(ll-1)-15021. (2) E. Rabinowitch, paper presented at the Biophysical Society Meeting, Washington, D. C., 1962, Abstract FC3. (3) K. G. hlathai and E. Rabinowitch, J . Phys. Chem., 66, 663 (1962). (4) A. K. Ghosh, unpublished. ( 5 ) K. Bergmann and C. T . O’Konski, J. Phys. Chem., 67, 2169 (1963). (6) C. A. Parker in “Photochemistry in the Liquid and Solid States,” L. J. Heidt, et al., Ed., John Wiley and Sons Inc., New York, N. Y., 1960, p 48. (7) E. Rabinowitch, J . Chem. Phys., 8 , 551 (1940); see also L. F. Epstein, F. Karush, and E. Rabinowitch, J . Opt. SOC.A m . , 31, 77 (1941).
NOTES
3013
I .7
I .6
1.5 0 .D. M
Downloaded by FLORIDA STATE UNIV on September 13, 2015 | http://pubs.acs.org Publication Date: September 1, 1966 | doi: 10.1021/j100881a507
1.4
~~
Figure 1. Apparatus used to study the bleaching of methylene blue by Fez+ in light: B1, 1000-w tungsten filament lamp; F1,2 cm thick water filter; L1, Ll, La, lenses; Ft, Balzers broad-band interference filter (Type K-6), transmission peak a t 650 mp; V, reaction vessel; Ph, photomultiplier (RCA 6217); T, water bath; M, Bausch and Lomb monochromator, 600 grooves/mm; Bz, 6-v, 18-amp tungsten ribbon filament lamp.
pends on the partition coefficient of L between the two solvents. For small relative concentrations of L (i.e., when only a small part of the dye is reduced), the amount extracted into ether is proportional to the total concentration of La8 For a larger change of concentration (or for dimerized solutions), the amount of leuco dye extracted into ether ceases to be proportional to its concentration in water. I n a one-phase system, the stationary ratio [JIB]:[L] is established, in intense light, within a few seconds after the beginning of illumination.’ If the bleaching is measured (with the instrument shown in Figure 1) in the two-phase system, while the illumination is in progress (by reducing momentarily the stirring before each measurement), the bleaching of the aqueous phase is found to occur as rapidly as in the absence of ether. However, this fast process is superimposed by a much slower shift, caused by progressive extraction of the leuco dye into ether. This slow extraction is demonstrated by the following three experiments. (1) Change of Absorption in the Reoxidized Water Phase. After a period of illumination, the gas flow is stopped and the two phases are allowed to separate. An aliquot of the water phase is transferred by a syringe from the photolytic vessel into a Bausch and Lomb spectrophotometer cuvette. During this operation, the dye in the aqueous phase is reoxidized by reaction of the leuco dye with Fe3+,but because part of the leuco dye had been extracted into ether, the reoxidized
1.3
1.2
620
660
700
mtL Figure 2. Absorption spectrum of the water phase after prolonged illumination: 1, t = 0; 2, t = 50 min; 3, t = 90 min.
aqueous solution contains less dye than it did before the illumination. After this measurement of the absorption spectrum, the sample can be returned to the reaction vessel, the illumination resumed, and measurements repeated several times. The decrease of the absorption band area, shown in Figures 2 and 3, is proportional to the total time of illumination. This, of course, means that the illumination times used in these experiments were short compared to the time needed to reach a stationary distribution of the leuco dye between the two phases. (2) The Potential Diflerence between the Water and the Ether Phase. For this measurement, a small volume (0.5 ml) of the ether phase was removed, after a certain illumination time, and placed in one half-cell of a galvanic cell, where it was mixed with 0.5 ml of methanol. A standard half-cell containing a (AIB FeS04) solu-
+
(8) J. H. Hildebrand and R. L. Scott in “Solubility of Non-electrolytes,” 3rd ed, Reinhold Publishing Corp., New York, N. Y., 1950, p 205.
Volume 70,Number 9 September 1966
NOTES
3014
01
0 0
20
40
60
Downloaded by FLORIDA STATE UNIV on September 13, 2015 | http://pubs.acs.org Publication Date: September 1, 1966 | doi: 10.1021/j100881a507
TIME,
80
100
120
140
Figure 3. Changes of potential difference ( A e ) between water and ether phases (crosses) and concomitant increase it1 abqorption, &OD (integrated over the band) in water phase (circles); squares show the concomitant increase i n ab3orption in reoxidized ether phase.
tion of known concentration was connected by an agar bridge saturated with KCl. Beckman platinum electrodes were used, and the difference of potential (A€) between the half-cells was measured by a Leeds and Northrup universal pH indicator assembly. The results are shown in Figure 3 (crosses, scale at right). The curve indicates growing accumulation of the reduced dye in the ether phase. (3) Reoxiclation of the Extracted Leuco Dye. The separated ether phase was exposed to air after dilution by methanol (to prevent precipitation of MB). The increasing amount of 3IB found in this phase after prolonged illumination is shown in Figure 4. (Recoloratioii could be observed upon exposure to air also in methanol-free ether-although solid MB does not dissolve niarliedly in ether, and no hIB is extracted into puic ether from an aqueous solution.) These three sets of experiments indicate slowly growing extraction of the leuco form of JIB into ether during prolonged illumination. The maximum amount extracted i n sonic of our experiments, about 35% of the total dye present, seemed to be close to saturation. The whole process, bleaching and extraction, is fully reversible-at least, after a not excessively long illumination period. If both phases are mixed together again and the ether is permitted to evaporate, the absorption spectrum of the remaining aqueous solution is found to be the same as before the experiment. With larger concentrations of MB (25 X M), the shape of the absorption curve changed during illumination. This must be attributed to the dimerization of the dye,9 Since the total concentration of MB in water was reduced by the extraction of a part of it (as leuro dye) into ether, the dimerization equilibrium must have been shifted. These results confirm that a storage of photochemical energy can be achieved by dividing the photoproducts The Journal of Physical Chemistry
540
min.
500
620
660
700
mP Figure 4. Absorption by reoxidized leuco dye in the ether phase after different times of illumination of the two-phase system: 1, 10 min; 2, 35 min; 3, 95 min; 4, 120 min.
between two phases in an inhomogeneous system. This could be considered as a model of the storage of photochemical energy in photosynthesis.2 Experiments should be made on the change in effectiveness of the separation of leuco methylene blue from illuminated solutions as function of concentration of the components, the pH, and the interface area between the two phases. (Competition between homogeneous recombination in the aqueous phase and the removal of the reduction product into another phase, preventing this recombination, must depend on the diffusion path between the locus of the primary reaction and the water-ether interface.) I n order to reproduce better the situation in the living cell, one could prepare very thin layers-only a few molecules thick-alternately hydrophilic and hydrophobic, and study the separation process in such systems. Finally, one should try to find and use dyes in which a higher proportion of the leuco form is in the neutral state under the pH conditions used. (9) E. Rabinowitch and L. F. Epstein, J . Am. Chem. SOC.,63, 69 (1941).
The First Ionization Potentials of Samarium, Europium, Gadolinium, Dysprosium, Holmium, Erbium, Thulium, and Ytterbium by the Electron-Impact Method
by K. F. Zmbov and J. L. Margrave Department of Chemistry, Rice University, Houston, Texas (Receiaed April 1 , 1966)
During a mass spectrometric study of gaseous equilibria involving rare earth subfluorides, it was possible
