4690
J. Phys. Chem. 1982, 86, 4690-4694
Study of the Micelle Formation of Sodium Deoxycholate. Concentration Dependence of Carbon-13 Nuclear Magnetic Resonance Chemical Shift Yoshlo Murata, Gohsuke Suglhara, Kohsuke Fukushlma, Mltsuru Tanaka, Department of Chemlstty, Faculty of Science, Fukuoka University, Nanakuma, Jonan-ku, Fukuoka 8 14-0 1, Japan
and Karuhlro Matsushlta NMR Appllcetion Laboratory, JOEL Ltd., Nakagami 1418, Akishlma, Tokyo 196, Japan (Received: March 15, 1982; I n Final Form: August 4, 1982)
It has generally been recognized that sodium deoxycholate (NaDC) forms polymer-like aggregates or gels only at low pH but does not form them above pH 8. Even at pH 7.8, however, its polymer-like aggregation was observed in this study. The surface tension of aqueous sodium deoxycholate-borate buffer solution (pH 7.8) was measured at 30 "C, and two inflection points were found in the surface tension-concentration curve. The partial molal volume of NaDC in the same borate buffer solution was also determined from precise density measurements, and a remarkable increase in the partial molal volume was observed upon transition from polymer-like aggregates to smaller micelles. In order to know the details of the aggregation of NaDC at the molecular level, we measured 13CNMR chemical shifts at various concentrations. The carbons of the methyl groups attached to the steroid skeleton showed about 0.4-0.5-ppm downfield shift with increasing NaDC concentration. This downfield shift suggests that hydrophobic bonding is involved in the change in the aggregation state. On the other hand, the methyl-group carbon in the side chain was not affected by the concentration change. The chemical shifts of carbinol carbons, C-3 and C-12, moved upfield by 0.6 and 0.3 ppm, respectively, with increasing NaDC concentration. The increase in NaDC concentration resulted in 0.6-ppm upfield shift in the case of the carboxyl-group carbon. From IIB NMR measurements it was revealed that the borate ion does not contributeto the formation of the NaDC aggregate. It was, accordingly,considered that the polymer-like aggregate having somewhat ordered structure is formed by the intermolecular hydrogen bonding between the hydroxyl group at the 3 position and the carboxyl group of the other molecule together with partially hydrophobic interaction (back-to-back).
Introduction Some reports on the association behavior of bile salts in aqueous solutions have been pub1ished.'-l2 Of the various bile salts, sodium deoxycholate (NaDC) has a unique aggregation behavior, such as polymer-like aggregation,9J3-lgthough the polymer-like aggregation was not (1)M. C. Carey and D. M. Small, Arch. Intern. Med., 130,506 (1972). (2)A. Djavanbakht, K. M. Kale, and R. h a , J. Colloid Interface Sci., 59, 139 (1977). (3)D. G.Oakenfull and D. E. Fenwick, A u t . J. Chem., 30,335(1977). (4)Y. Chang and J. R. Cardinal, J. Pharm. Sci., 67, 994 (1978). (5)B. Lindman, N. Kamenka, H. Fabre, J. Ulmius, and T. Wieloch, J. Colloid Interface Sci.. 73. 556 (1980). (6)M. Van&ere, R. Mat&ajan; and S. Lindenbaum, J. Phys. Chem., 84,1900 (1980). (7)G. DArrigo, B. Sesta, and C. La Mesa, J. Chem. Phys., 73, 4562 (1980). (8) C. H. Spink and R. E. Stedwell, J. Phys. Chem., 84,2044(1980). (9)Y. Murata, G.Sugihara, N. Nishikido, and M. Tanaka in "Solution Behavior of Surfactants: Theoretical and Applied Aspects", Vol. 1, K. L. Mittal and E. J. Fendler, Eds., Plenum Press, New York, 1982, pp 611-28. (10)J. Robeson, B. W. Foster, S. N. Rosenthal, E. T. Adams, Jr., and E. J. Fendler, J. Phys. Chem., 85,1254 (1981). (11)D. C. Thomas and S. D. Christian, J . Colloid Interface Sci., 82, 430 (1981). (12)G.Sugihara, K.Yamakawa, Y. Murata, and M. Tanaka, J.Phys. Chem., 86,2784 (1982). (13)A. Rich and D. M. Blow, Nature (London), 423 (1958). (14)D. M. Blow and A. Rich, J. Am. Chem. SOC.,82, 3566 (1960). (15)C. Botr6, P. A. Cicconetti. C. Lionetti. and M. Marchetti. J. Pharm. Sci., 56, 1035 (1967). (16)G.Sugihara, K. Motomura, and R. Matuura, Mem. Fac. Sci., Kyushu Uniu., Ser. C, 7 , 103 (1970). (17)G . Sugihara and M. Tanaka, Bull. Chem. SOC.Jpn., 49, 3457 (1976). ~
~~
0022-3654/82/2086-4690$0 1.25/0
observed in various studies of NaDC solution above pH In our previous paper the aggregation states of NaDC in aqueous borate buffer solutions (pH 7.8) were studied by means of various physicochemical techniques and an interesting finding was obtained; i.e., the aggregates formed in the lower concentration range are much larger than those formed in the higher concentration range? This phenomenon seems to contradict common knowledge on micelle formation in aqueous solutions of surfactants. The concentration changes in the dissymmetry, ,Z , and in the molar ellipticity, t9220, were obtained through light-scatbring and CD measurements, respectively. An abnormally high value of Z4, was found in the lower concentration range, while in the higher concentration range it was nearly equal to unity. This suggests that the aggregation state in the lower concentration range is quite different from that in the higher range. The same interpretation could be made from the increase in t9zzo with concentration; the aggregate formed in the lower concentration range may be considered to have a fairly ordered structure. Viscosity measurements were made on the same solution systems as above; the reduced viscosity as a function of NaDC concentration also indicates that the aggregation number in the lower concentration range is larger than that in the higher range. The partial molal volume and the surface tension of NaDC solutions were measured (more detailed data obtained recently will be described later). From all 8.192943631'
(18)G.Sugihara, T.Ueda, S. Kaneshina, and M. Tanaka, Bull. Chem. SOC.Jpn., 60, 604 (1977). (19)G . Sugihara, M.Tanaka, and R. Matuura, BULL. Chem. SOC.Jpn., 50,2542 (1977).
0 1982 American Chemical Society
The Journal of Physical Chemistry, Vol. 86,
Micelle Formation of Sodium Deoxycholate Conc, o f NaDC
Conc. o f NoDC
( g/g )
No. 24, 1982 4691
( g/g )
0,0025 0,005 0.01
-
320 I -7,O
-6.0
-5.0
-4,O
In -7.0
-6.0
-5.0 In m
-4.0
-3.0
Flgure 1. Plots of the surface tension of NaDC-buffer solution vs. logarithmic concentration in molality at 30 O C and pH 7.8.
results mentioned above it has been concluded that in aqueous borate buffer solutions of NaDC the aggregate in the lower concentration range has a larger size and a rather ordered structure, and with increasing NaDC concentration the larger aggregate is broken down and changed into smaller micelles formed by hydrophobic bonding. Recent developments in the instrumentation for NMR spectroscopy have enabled us to observe the 13C NMR spectra at natural-abundance levels of various compounds within a short time. Many papers have appeared even in the field of bile salts and related On the other hand, IH or 2H NMR spectroscopy has been used to study the solution state of bile salt^.^^,^^ From the concentration dependence of the relaxation time it was found that hydrophobic (back-to-back) interaction between bile salt molecules occurred in aqueous solution. In the present paper a 13CNMR study has been made in order to confirm our previous work and to obtain molecularstructural information on the aggregates. Experimental Section NaDC samples purchased from E. Merck Co. and Nakarai Co. were recrystallized 7 times from ethanol solution and dried at 110 "C in vacuo for 1-5 days. The purity of NaDC was found to be higher than 99.9% from the elemental analysis. Thin-layer chromatography gave one and the surface tension of the solution gave no minimum (see Figure 1). Other chemicals were of guaranteed reagent grade and used without further purification. The thrice-distilled water was used through all experiments. The surface tension measurements were carried out on the NaDC-buffer solution (pH 7.8, ionic strength 0.35) at 30 "C by means of a Wilhelmy-type surface balance (Kyowa CBVP surface tension meter A3). The density of the solution was measured by the use of Anton Parr densitometer Model 02C and DMA 60. The sample solutions were degassed to obtain more precise values. 13C NMR spectra were obtained on a JOEL FX 200 FT NMR (20)B. Lindman, C.Lindbbm, H. Wenneratrom, and H. Gustausson, 'Micellization, Solubilization, and Microemulsions", Vol. 1,K. L. Mittal, Ed., Plenum Press, New York, 1977,pp 195-227. (21)S. Miyagishi and M. Nishida, J . Colloid Interface Sci.,78,195 (1980). (22)H.Maeda, S. Ozeki, S. Ikeda, H. Okabayashi, and K. Matsushita, J. Colloid Interface Sci..76. 532 (1980). (23)D.M. Small, S. A.P&kett,'and D. Chapman, Biochim. Biophys. Acta, 176,178 (1969). (24)B.M. Fung and M. C. Peden, Biochim. Biophys. Acta,437,273
.--(25)T.Usui, J . Biochem., 54,283 (1963).
(1976). - -,-
-3,O
-2.0
m
Flgure 2. Concentration change in the partial molal volume change of NaDC in the borate buffer solution (pH 7.8) at 30 "C.
spectrometer at 50.1 MHz. All spectra were recorded under proton noise decoupling, deuterium internal lock mode, a pulse recycling time of 1 s, a flip angle of 40-50" and 1000-20 000 transients. 13Cchemical shift measurements were accurate to 0.02 ppm. To prevent the sample heating due to proton decoupling, spectra were observed under a gated decoupling mode, and the ambient probe temperature was 25 f 1 "C. The reproducibility of 13C chemical shifts was within f0.02 ppm. On the other hand, llB NMR spectra were measured under a spectral width of 50 KHz, a pulse recycling time of 1 s, 16K data points, a pulse width of 10 ks, and 20 transients. Borate buffer solution without NaDC was used as an external reference. Results and Discussion In order to examine the purity of NaDC and obtain more detailed information on the aggregation states, we carried out surface tension measurements. The results are shown in Figure 1, which shows that there exist two inflections in the curve of the concentration dependence of the surface tension. The concentration which gives the inflection point at the higher-concentration side coincides with those which have been observed in the experiments of the light scattering, CD, and so on. This inflection, accordingly, seems to reflect the transition from the polymer-like aggregate to the micelle-like one as mentioned b e f ~ r e .The ~ other inflection point at the lower concentration in the curve may be considered to be the concentration at which the polymer-like aggregates begin to form. From density data the partial molal volume of NaDC was calculated as a function of NaDC concentration as is shown in Figure 2. Here it should be noted that the apparent partial molal volume data used for Figure 2, though they are not given here, show a decreasing tendency with increasing NaDC concentration up to ca 6.0 X m (from 370 to 335 cm3/mol) and then become nearly constant (its general feature is shown in a previous paperg). On the other hand, Lindenbaum et al.'s result obtained at higher pH (pH 9) showed at the lower concentration ranges an increasing tendency with increasing concentration, and volumes were lower than those of this paper.6 This difference supports the conclusion that the aggregation state at pH 7.8 is quite different from that at pH 9 and that the polymer-like aggregation takes place at pH 7.8. In Figure 2 a remarkable discontinuity was found in the curve of the concentration vs. the partial molal volume of NaDC in the solution. The concentration where the discontinuity appears coincides with that found in the other properties. It is well-known that hydrophobic bonding accompanies an increase in the partial molal
4692
The Journal of Physical Chemistry, Vol. 86, No. 24, 1982
Murata et al.
m0,03
21
CH 3
0,O
0,01
0,02
0,011
0.05
I " " ' " " '
OH 18 13,
I
C0ONa
24.0
LA/J
HO
Flgure 3. Molecular structure of NaDC. Numerals indicate the numbers assigned to respective carbons. 13.
( B ) C-18
Figure 4. I3C NMR chart of NaDC In the aqueous solution.
volume at least at atmospheric pressure.26 The abrupt increase in the partial molal volume of NaDC also indicates that the hydrophobic bonding is accompanied or enhanced by the transition from polymer-like aggregates to micelle-like ones. In order to understand the mechanism of the aggregation of the molecular level, we carried out a 13C NMR study. The molecular structure of NaDC is given in Figure 3, in which C-24 denotes the carbon of the carboxyl group; C-3 and C-12 denote carbons bonded to the hydroxyl groups; and C-18, C-19, and C-21 denote the carbons of the methyl groups. (2-24 is likely to play an important role in forming an ordered (polymer-like) aggregate between NaDC molecules, because it may be possible for the C-24 carboxyl group to form a hydrogen bond with the C-3 or C-12 hydroxyl groups of other NaDC molecules. Since C-18, C-19, and C-21 belong to the hydrophobic part of the molecule, they will probably give useful information on the hydrophobic bonding. Quaternary carbons C-10 and C-13 are so distinguishable from the other carbon atoms that one should pay attention to these two carbon atoms to know the behavior of the steroid skeleton on the aggregation of NaDC. The I3C NMR spectrum of NaDC solution (0.004 g/g = 0.0096 m) obtained is shown in Figure 4. The assignment of the spectral peaks has been determined by Leibfritz et al.27 The peaks of the three methyl groups are found in the most upfield region. The carbons of the methylene groups of the steroid skeleton, the quaternary carbons, and the C-3 and (2-12 bonded to the hydroxyl groups appear in the downfield region. In the most downfield region there exists the spectrum of C-24 of the carboxyl group. Respective states of carbon atoms in the NaDC molecule have been observed by measuring the NMR spectra as a function of the NaDC concentration. Here, it should be noted that the 13C chemical shift, in general, is related to the electron density of the carbon atom along with the steric configuration around it. In other words, the nucleus of a carbon atom which has lower electron density is less screened by electrons, so that it gives a downfield shift. Therefore, the downfield shift of the carbon of the carboxyl group is explained in terms of the decreased electron density. It has also been revealed that steric crowding of other groups around the carbon (26) M. Tanaka, S. Kaneshina, K. Shin-no, T. Okajima, and T. Tomida, J . Colloid Interface Sci., 46, 132 (1974). (27) D. Leibfritz and J. D. Roberta, J. Am. Chem. Soc., 95, 4996, (1973).
12,5
0.0
0,Ol
0,005
0.02
0.015
Conc. o f NaDC
( g/g
)
Figure 5. Concentration dependencies of the chemical shifts of methylgroup carbons attached to the steroid ring: C-19 (A) and C-18 (B).
, 0.p1
m
, o,p3
, 0.p2
, O'p4
, 0*:5,
?aL
17.0
'
0.0
I
0.005
0,Ol Conc. o f NoDC
I
0.015 ( g/g
0,02 )
Flgure 6. Concentration dependency of the chemical shift of the methyl group in the side chain, C-21.
atom generally causes an upfield shift.28 However, a downfield shift due to a steric crowding effect between methyl groups was also observed for some substance^.^^^^^ Taking into account these factors, we will interpret the concentration dependence of the NMR spectra as follows. Curves A and B in Figure 5 show the concentration dependences of the chemical shift of (3-18 and C-19. C-18 and C-19 give downfield shifts up to 0.5 and 0.4 ppm with increasing NaDC concentration, respectively. This downfield shift is an interesting contrast with the upfield shift shown by the terminal methyl group of aliphatic surfactants on micelle formation.22 This fact suggests that the aggregation states of NaDC may be quite different from the micellar state of the aliphatic surfactants. These downfield shifts in the case of NaDC occur for the most part up to the concentration, ca. 0.006 g/g = 0.0145 m, which correspondsto the discontinuity in the partial molal volume change, etc. The downfield shifts may be attributed to the steric crowding effect accompanied by the (28) G. J. Martin, M. L. Martin, and S. Odiot, Org. Magn. Reson., 7, 2 (1975). (29) J. I. Kroschwitz, M. Winokur, H. J. Reich, and J. D. Roberts, J. Am. Chem. Soc., 91, 5927 (1969). (30) S. H. Grover and J. B. Stothers, Can. J. Chem., 52, 870 (1974).
The Journal of Physical Chemistry, Vol. 86, No. 24, 1982
Micelle Formation of Sodium Deoxycholate
,
0.p1
, o,p2 ,
4693
311 0’;,
, ;0.
,
0.95
185,5
184.5
1 I
0.0 0.005
0.0
0.01
0,015
Conc. o f NaDC
0.02
( g/g )
,
Flgure 7. Concentration dependencies of the chemical shift of quaternary carbons: C-13 (A) and (3-10 (B).
or
0,o
m , 0,;’
,
0 005 I
0,,02
,
0,Ol
Conc. of NaDC
0,:’
, o,p4
0,015
, Of5
0.02
( g/g )
Flgwe 8. Conceniratbn dependencies of the chemical shm of carbons wRh hydroxyl groups: C-3 (A) and C-12 (B).
change in the aggregation state which brings the methyl groups into a sterically hindered state as a result of hydrophobic bonding between NaDC molecules. On the other hand, as is shown in Figure 6 the chemical shift of C-21 constituting the methyl group in the side chain seems not to depend on the NaDC concentration, which means that C-21 is hardly affected by the change in the aggregation state. This behavior contrasts with those of methyl groups C-18 and C-19 attached to the steroid ring. Figure 7 shows that the chemical shift for the quaternary carbons (2-10 and C-13, chosen as the representatives of the steroid ring, show little concentration dependency, irrespective of the change in aggregation state of NaDC. This seems to be due to the stiffness of the steroid skeleton. Next, let us focus on the carbons with hydroxyl groups. The curves in Figure 8 show the relations between NaDC concentration and the chemical shifts of C-12 and C-3. (3-12, which is located nearly at the center of the NaDC molecule, shows an upfield shift of ca. 0.3 ppm in contrast with the carbons of the methyl groups. The upfield shift may be ascribed to the increase in the electron density of C-12 due to the breakdown of hydrogen bonding upon the
I
0,005
0,Ol
Conc, o f NaDC
I
0.015 ( g/g
0,02 )
Flgure 9. Concentration dependency of the chemical shift of the carboxyl-group carbon, C-24.
change in aggregation state. Another carbon atom with a hydroxyl group, C-3, is located at an end of the steroid skeleton. As is shown in Figure 8, the shift changes toward the upfield by 0.6 ppm with increasing NaDC concentration. The larger shift of C-3, twice that of C-12, suggests that the OH group of the 3 position plays a more important role than the OH of the 12 position upon the change in the aggregation state. Finally, the chemical shift of C-24 changes by ca. 0.6 ppm toward the upfield with increasing NaDC concentration, as is shown in Figure 9. The upfield shifts of C-3, C-12, and C-24 are considered to correspond to the decrease in the steric crowding and/or the increase in the electron density due to the breakdown of hydrogen bonding. This agrees with the idea, mentioned in previous papers>ls that the polymer-like aggregates are formed by means of intermolecular hydrogen bonding in the lower concentration range in the aqueous NaDC solution. Summarizingthe results from 13CNMR chemical shifts, the carbons of the methyl groups (2-18 and C-19, give downfield shifts, and, on the other hand, carbons with hydroxyl grmps or in the carboxyl group give upfield shifts. It is clear that the methyl groups which constitute the hydrophobic part of the NaDC molecule behave differently from the two OH groups and the one COOH group when the aggregation state is changed. It has been reported that the aqueous borate ion-poly(vinyl alcohol) (PVA) system shows a thickening and gelling phenomenon, which has been considered to result from the complex formation of monodiol or didiol types between the OH groups of PVA and borate ion, similar to those in the simple low-molecular-weight polyol-borate systems.31 This seems to suggest that there is a possibility that in the NaDC-borate buffer system also borate ions might contribute to the formation of polymer-like aggregates. However, the polymer-like aggregate can be formed by addition of only HC1 or NaC1.17-20So the borate ion itself is not essential to form the aggregate. Further llB NMR measurements were carried out in order to examine the possibility. The chemical shift of llB was independent of the NaDC concentration. This result suggests that the possibility of complex formation between the borate ion and the deoxycholate molecule is negligible. In an aqueous solution hydrogen-bonding interactions of H 2 0with the COO- group and with the C-3 OH groups ought to be as strong as or stronger than the intermolecular (31) H. Ochiai, S. Shimizu, Y. Tadokoro, and I. Murakami, Polymer, 22, 1456 (1981).
4894
J. Phys. Chem. 1982, 86. 4694-4700
hydrogen bonds between C-3 OH and the carboxyl. It seems difficult to invoke an intermolecular hydrogen bond without having some sort of added hydrophobic contribution in aqueous solution. This kind of effect has been observed in aliphatic carboxylic acids-larger aliphatic chains leading to stronger dimerization effects in aqueous solution.32 To describe the 13C NMR shifts as due to intermolecular hydrogen bonding seems too simple for aqueous solutions of NaDC. Perhaps some partial hydrophobic interactions, which may be back-to-back hydrophobic interaction between bile salt molecules, occur in the initial aggregation, and this then changes when the “normal” micelles begin to form. D e ~ n o y e r suggested s~~ that the model of the partial hydrophobic interaction in the polymer-like aggregate may be Fujiyama’s model; i.e., water molecules lie between hydrophobic molecules.” The decrease of the partial molal volume with the concentration (32) P. Mukerjee, K. J. Mysels, and C. I. Dulin, J. Phys. Chem., 62, 1390 (1958). (33) J. E. Desnoyers, private communication at the 6th International
Symposium on Solute-Solute-Solvent Interaction, Minoo Osaka, Japan, 1982. (34) N. Ito, T. Kato, and T. Fujiyama, Bull. Chem. SOC.Jpn., 54,2573 (1981).
in the lower concentration range (see Figure 2) can probably be interpreted by the model. Further investigation is needed to decide whether the partial hydrophobic interaction in the polymer-like aggregate is of Fujiyama’s model or not. It may be concluded that, as has been suggested from various experiments, the intermolecular hydrogen bonding between the OH group at the 3 position and the COOH group and partial hydrophobic interaction (back-to-back) contribute to the formation of polymer-like (ordered) aggregates in the lower concentration range and that, with increasing NaDC concentration, the polymer-like aggregates change into smaller aggregates accompanying the breakdown of hydrogen bonding and the enhancement of hydrophobic bonding, as is indicated by the upfield shift of methyl-group carbons and by the abrupt increase in the partial molal volume. Acknowledgment. We thank Professor J. P. Kratohvil, Professor J. E. Desnoyers, and the reviewers of this paper for a great deal of advice. We are indebted to our coworkers for experimental assistance. The present work has been performed within a research program sponsored by Central Research Institute of Fukuoka University.
Viscosity Dependence of the Rotational Reorientation of Rhodamine B in Mono- and Polyalcohols. Picosecond Transient Grating Experiments R. S. Moog, M. D. Ediger, S. 0. Boxer, and M. D. Fayer’ Department of Chemlsby, Stanford University, Stanford. California 94305 (Recelvecl: M a y 24, 1982; I n Flnal Form: August 17, 1982)
Rotational reorientation times (TROT)were obtained for rhodamine B in a series of n-alcohols and polyalcohols of varying viscosities by a transient grating technique. The general trend of the Debye-Stokes-Einstein (DSE) theory was shown to extend to much higher viscosities than previously reported. Detailed analysis of the results suggesta that the n-alcohols and polyalcohols provide two distinct hydrodynamic boundary conditions for the rotating species. In the n-alcohols, stick boundary conditions are observed, while the polyalcohols provide approximately slip boundary conditions. The question of a difference in the rotation times for the ground and excited electronic states is addressed and an upper bound is placed on this difference for rhodamine B.
Introduction The study of rotational reorientation of molecules in liquid solvents has been used as a probe of solute confomations,i solvent-solu~hteradions,z-6 and local solvent structwe~ undersb&,g of molec* and of rotation in liquids is also important in the reaction dynmics.s over10 years ago, the first
direct observation of rotational reorientation of molecules in a liquid solvent using picosecond laser pulses was rePorted.g Since then several other workers have applied picosecond spectroscopic techniques to the study of rotational m ~ t i o n . ~ J + Various l~ dye molecules in a series of solvents of different viscosities have been examined, and the results have been compared to the predictions of De-
(1) Fleming, G. R.; Knight, A. E. W.; Morris, J. M.; Robbins, R. J.; Robinson, G. W. Chem. Phys. Lett. 1977, 49, 1. (2) Scholz, M.; Teuchner, K.; Nather, M.; Becker, W.; Dahne, S. Acta Phys. Pol. A 1978,54, 823. (3) Klein, U. K. A.; Haar, H. P. Chem. Phys. Lett. 1978, 58, 531. (4) Millar, D. P.; Shah, R.; Zewail, A. H. Chem. Phys. Lett. 1979,66,
(8) Steiger, U. R.; Keizer, J. J. Chem. Phys. 1982, 77, 777. (9) Eisenthal, K. B.; Drexhage, K. H. J. Chem. Phys. 1969,51,5720. (10) Phillion, Donald W.; Kuizenga, Dirk J.; Siegman, A. E. Appl. Phys. Lett. 1975, 27, 85. (11) Fleming, G. R.; Morris, J. M.; Robinson, G. W. Chem. Phys. 1976, 17, 91. (12) Eichler, H. J.; Klein, U.; Langhans, D. Chem. Phys. Lett. 1979, 67, 21. (13) Beddard, Godfrey, S.; Doust, Tom; Porter, George Chem. Phys. 1981, 61, 17. (14) van Resandt, R. W. W.; de Maeyer, L. Chem. Phys. Lett. 1981, 78, 219. Spears, K. G.; Cramer, L. E. Chem. Phys. 1978, 30, 1.
435. (5) von Jena, A.; Lessing, H. E. Chem. Phys. 1979, 40, 245; Chem. Phys. Lett. 1981, 78, 187. (6) Shapiro, S. L.; Winn, K. R. Chem. Phys. Lett. 1980, 71, 440. (7) Beddard, G. S.; Doust, T.;Hudales, J. Nature (London) 1981,294, 145.
0022-365418212086-4694$01.2510
0 1982 American Chemical Society
