Langmuir 1995,11, 2476-2479
2476
Sodium Dodecyl Sulfate Micellar Aggregation Numbers in the Presence of Cyclodextrins David J. Jobe Atomic Energy of Cariada, Pinawa, Manitoba, Canada ROE 1LO
Vincent C. Reinsborough” and Stacey D. Wetmore Department of Chemistry, Mount Allison University, Sackville, New Brunswick, Canada EOA 3CO Received November 21, 1994. I n Final Form: April 24, 1995@ Micellar aggregation numbers, n, have been obtained for sodium dodecyl sulfate (SDS) micelles in cyclodextrin (CD) solutions through static fluorescence quenching. For a-cyclodextrin, P-cyclodextrin, (hydroxylpropyl)-b-cyclodextrin,maltosyl-@-cyclodextrin,and y-cyclodextrin solutions, n was found to be essentially unchanged from the 59 f5 value found in sodium dodecyl sulfate solutions without cyclodextrin present, even though the critical micelle concentrations as measured conductometrically had increased. The increases in critical micelle concentrations were consonant, with 1:l CDISDS complexation predominating over other complexesin all CD systems except for a-CD solutions,where the 2:l a-CD/SDS complex prevailed. The association constants of the fluorescence quencher, 9-methylanthracene, with SDS and with each of the pure cyclodextrinswere determined to ensure preferred binding of the quencher with SDS micelles rather than with CD. The substituted cyclodextrins, dimethyl-P-CD and trimethylp-CD,were not amenable to the fluorescence quenching method. The implicationsof these findings to drug delivery are examined.
Introduction The complexes formed between surfactants and cyclodextrins (CDs)have recently received much attention,’-1° partly because these systems can be used to model the effect of cyclodextrins on phospholipids, a major constituent of cell membranes. Hemolysis of cell membranes is a major deterrent to the use of cyclodextrins in the sitespecific delivery of drugs.ll Although initial studies deal with the CD inclusion of the surfactant in the absence of any micelles,’-’ more recent studies have focused on the changes occurring within the micelle due to the presence of the 1:l complex formed between unmicellized surfactant and CD’S.~-~O These studies have shown that the cyclodextrin principally acts a s a “vacuum-cleaner”, siphoning away surfactant to form 1:1 inclusion complexes, ultimately leading to a decrease in the concentration of surfactant that can form micelles. Although it is appreciated that cyclodextrins inhibit the formation of micelles in this manner, the mechanism and the exact impact of these complexes on the properties of the micelle are not well-
* Author to whom all correspondence should be addressed. Abstract published in Advance A C S Abstracts, J u n e 15,1995. (1)Satake, i.; Ikenoue, T.; Takeshita, K.; Hayakawa, K.; Maeda, T. @
Bull. Chem. SOC.Jpn. 1985,58, 2746. (2) Satake, I.; Yoshida, K.; Hayakawa, K.; Maeda, T.; Kusumoto, Y. Bull. Chem. SOC.Jpn. 1986,59,3991. (3)Palepu, R.; Richardson,J. E.; Reinsborough,V.C.Langmuir 1989, 5. _ ,218.
(4)Palepu, R.; Reinsborough, V. C. Can. J . Chem. 1988,66,325. (5)Palepu, R.; Reinsborough, V. C. Can. J . Chem. 1989,67,1550. (6)Georges, J.; Desmettre, S. J. J . Colloid Interface Sci. 1987,118, 192. (7) Saint Aman, E.; Serve, D. J . ColloidInterface Sci. 1990,138,365. (8)Jobe, D. J.; Verrall, R. E.; Junquera, E.; Aicart, E. J . Phys. Chem. 1993,97,1243. (9)Jobe, D. J.; Verrall, R. E.; Junquera, E.; Aicart, E. J . Phys. Chem. 1994,97,10814. (10)Junquera, E.; Aicart, E.; Tardajos, G. J . Phys. Chem. 1992,96, 4533. (11)Yamamoto, M.; Aritomi, H.; Irie, T.; Hirayama, F.; Ukkama, K. In Minutes: 5th International Symposium on Cyclodextrins; Duchgne, D.; Ed.; Editions de SantB, Paris, 1990,p 541.
understood. This complicated ternary system poses a number of interesting questions; the first and most important asks if the complex is incorporated into the micelle and the second, if the size and shape of the micelles change in the presence of the complex. Ultrasonic absorption spectroscopicinvestigations have shown that the kinetics of the surfactant-micelle solution process for decyltrimethylammonium bromide8 and sodium perfluoroo~tanoate~ are unaffected by the presence of the complex. Furthermore, when the Aniansson model12 is applied to the ultrasonic data, the analysis is satisfactory, which argues for a n unaltered size and shape for the micelles. However, no direct measurements of these properties, e.g., micellar aggregation number ( n ) ,hydrodynamic radius, have been made. Micellar aggregation numbers can be obtained by the quenching of fluorescent probes.13-17 Under certain conditions, a particularly convenient and simple variant is the static fluorescence quenching method due to Turro and Yekta.13 In this method, both the fluorophore and quencher must be preferentially absorbed into the hydrophobic environment of the micelle, yet both must a t the same time be sufficiently hydrophillic to be soluble in water. An extra constraint in the present situation is that neither molecule, although hydrophobic, can be strongly complexed by cyclodextrins. Since it is chiefly through hydrophobic interactions that cyclodextrins include great molecules into their cavities, this restriction may greatly limit the choice of a suitable fluorophore/ quencher pair. (12)Aniansson, E. A. G.; Wall, S. N. In Chemical and Biological Applications ofRelamtion Spectrometry;Wyn-Jones, E.; Ed.; D. Riegel Publishing: Dordrecht, Holland, 1975;p 223. (13)Turro, N.; Yekta, A. J . Am. Chem. SOC.1987,100, 5951. (14)Yekta, A.; Aikawa, M.; Turro, N. J. Chem. Phys. Lett. 1978,63, 543. (15)Lianos, P.; Zana, R. J. J . Colloid Interface Sci. 1981,84, 100. (16)Almgren, M.; Lofroth, J.-E. J . Colloid Interface Sci. 1981,81, 486. (17)Jobe, D. J.; Skalski, B. D.; Verrall, R. E. Langmuir 1993,9, 2814.
0743-7463/95/2411-2476$09.00/00 1995 American Chemical Society
Langmuir, Vol. 11, No. 7, 1995 2477
SDS Micelles in Cyclodextrin Solutions A possible candidate for the fluorophore is the trisbipyridyl complex of Ru2+ used in the original paper13 since ruthenium bipyridyl complexes bind only weakly with cyclodextrins.l8 However, introducing a fluorescence probe and a quencher into a n already complex system gives rise to a number ofother possible mechanisms, most of which involve cyclodextrin complexation. Therefore, it is imperative to choose a quencher/fluorophore pair that will minimize the effect of these alternate routes in order to obtain meaningful data in these systems. In this study, we have obtained dodecyl sulfate micellar aggregation numbers ( n ) in the presence of various cyclodextrins using the Turro and Yekta method with the fluorophore-quencher pair of Ru(bipy)S2+and 9-methylanthracene (9-MeA). Both molecular species proved to be preferentially absorbed into the micellar medium rather than into cyclodextrin cavities. The effect of CD concentration, 1 : l complex formation, and CD cavity size on n was also investigated.
601
d 0 -0.01
Results and Discussion (a) Binding of Donor and Quencher by SDS and CD’s. In order to have effective quenching of the donor by the quencher in micellar SDS solutions containing CDs, it is important that both the donor and quencher be preferentially absorbed into the micellar phase rather than be encapsulated by CD’s. Thus, it is first necessary to determine the relative strengths of these interactions. As explained above, the fluorophore Ru(bipy)S2+can reasonably be assumed to have been assimilated almost entirely into the SDS micelles. By way of confirmation, when CD’s were introduced into Ru(bipy)32+solutions, no perceptible change in the visible spectrum was noted. Spectral displacement is often used to obtain CD binding constants.l8 The anthracene moiety of the quencher 9-MeA should show affinity for the cyclodextrin cavity especially for that of y-CD, which has roughly the correct diameter (800 pm) for a snug fit. A rough indicator of the competitiveness of CD inclusion over micelle solubilization is to compare the fluorescence quenching with fluorophore and quencher present in micellar solution with and without CD present. If the fluorescence readings were the same, then any incorporation of 9-MeA by CD was probably of little consequence as compared with its incorporation into the SDS micelles. This was, in fact, the situation with all of the CD’s except DIMEB and TRIMEB which both caused the fluorescence to increase. This would be the direction expected if CD/9-MeAcomplexformation was significantly (18)Johnson, M.D.;Reinsborough, V. C.; Ward, S. Inorg. Chem. 1992,31, 1085.
(19) Connolly, T.J.;Reinsborough, V. C.; Xiang, X. Aust. J. Chem. 1992,45, 769.
0
0.01 0.02 0.03 [SOSl,,/ M
Figure 1. Solubility of 9-methylanthracene in micellar SDS solutions ([SDSl,ic = [SDSI - cmc). 301
Experimental Section Fluorescence measurements were made in sodium dodecyl sulfate solutions (SDS) a t excitation and emission wavelengths of 450 and 630 nm, respectively, with a Turner Model 430 spectrofluorometer with Ru(bipy)?+ as the luminescent donor and 9-MeA as the luminescence quencher. The concentration of the latter in solution was determined spectrophotometrically while the former was maintained at approximately 5 x M Ru(bipy)32+. Solution conductivity4 and solubility datalg were obtained as described elsewhere. The cyclodextrins(CD)used were a-CD (Sigma),j3-CD (Sigma), y-CD (Wacker and American Maize), 2,6-0-dimethyI-P-CD (DIMEB) (Cyclolab), 2,3,6-0-trimethyl-P-CD (TRIMEB) (Cyclolab), hydroxypropyl-P-CD (HPBCD) (American Maize), and maltosyl-P-CD or G2-P-CD (a gift from Ensuiko Sugar Refining consisting of 99.8% total branched p-CD; 44.2% of the monomaltosyl form, 41.4% of the dimaltosyl form).
P P
50
20mi \
O Y * , 0
I
5
I
I
10
I
1
15
IGCDI / m M Figure 2. Solubility of 9-methylanthracene in y-CD solutions.
competing with assimilation of 9-MeA into the SDS micelles. As a result of this observation, DIMEB and TRIMEB were not investigated further. However, a better indicator of the prevailing degree of competitiveness would be to measure the binding constants for 9-MeA with SDS micelles and independently with each of the CDs. When the solute is colored in solution, as 9-MeAis, spectrophotometrically determined solubility measurements are a convenient route to these binding constants. If S and So are the solubilities of a solute in micellar SDS and in water, respectively, and if the concentration of the micellar SDS or [SDS],i, can be approximated by [ S D S l b ~ cmc (cmcis the critical micelle concentration) (see section c), then the association constant, KA, for a solute A may be defined as (S - So)/ S&[SDSIktal- cmc).lg The corresponding equilibrium process would be SDS,i, -t Amic. Values of KAemerge as the slopes of the linear plots of ( S - So)/Soagainst [SDSltotal- cmc (Figure 1). A similar expression holds for the 1 : l association of the solute with CD; i.e., KA= ( S - So)/So[CDl,where S is now the solubility of the solute in the CD solution (Figure 2). The determined values of& were 2300 f300 M-’ for the binding of 9-MeA with micellar SDS, 120 f20 M-l for the formation constant of the 9-MeNa-CD complex, 280 f 40 M-l for the formation constant of the 9-MeNp-CD complex, and 1700 f 300 M-’ for the formation constant of the 9-MeNy-CD complex. As anticipated from steric considerations, y-CD formed the most stable complex among the CD’s. More importantly for the project a t hand, it was noted that 9-MeA was preferentially solubilized by micellar SDS rather than encapsulated by a-CD, p-CD, and even y-CD. It could be argued that a more realistic and appropriate micellar SDS concentration term would be the concentration of the micelles rather than that of the SDS monomers within the micelle. In this case, the value of KAfor the binding of 9-MeA with SDS micelles would be 1.2 x IO5 M-l (assuming a value of n = 60) which makes the disparity in binding strengths even more
+
Jobe et al.
2478 Langmuir, Vol. 11, No. 7, 1995 concentration:
1
In I = constant - n[Ql/[SDSl,i, +
+ t
0
10
20 lG2BCDl / mM
30
Figure 3. Effect of GPBCD addition on molar conductivities ofmicellar SDS: 0 , 9 . 5 mM SDS;W, 12.0mM SDS; A, 15.0 mM SDS; +, 20.0 mM SDS.
0 0
5
10 [GZBCD] / mM
15
Figure 4. Dependence of cmc of SDS solutions on G2BCD concentration.
striking. It is clear that, in the presence in solution of these three CD’s,9-MeA will be bound preferentially into the SDS micelles. (b) Dependence of cmc upon CD Concentration. Cyclodextrins in surfactant solutions elevate cmc values because CD encapsulation of surfactant monomers competes with the micellization p r o c e ~ s .The ~ expression for the determination of the micellar aggregation number through static fluorescence quenching13requires accurate values of the free monomer concentration. cmc values in the presence of CD are given by the maxima in the solution conductivity curves when micellar SDS solutions are titrated with CD.4 Figure 3 shows the conductivity curves generated when G2BCD was added to various micellar SDS solutions, and Figure 4 depicts the linear relationship obtained when the maxima of these curves or cmc’s were plotted against [GBBCD]. All the CD systems showed similar behavior. The cmc elevation for each CD system was expressed as the increase in cmc per CD concentration: 0.55 for a-CD, 1.04 for yCD, 0.82 for GBBCD, and 0.80 for HPBCD. A value of 0.70 for p-CD had been determined p r e v i ~ u s l y . ~ SDS forms principally a 2 : l complex with a-CD along with some 1 : l complex; the 1:l complexpredominates withp-CD; only a 1:1complex forms with Y - C D . ~The cmc elevation constants correspond remarkably well with these stoichiometries, Le., the elevation in cmc occasioned by the addition of CD for the formation of a 1:1complex should be approximately unity and, for the formation of a 2:l CDIsurfactant complex, about one-half. This is further confirmation that the norm in CD/SDS complexation is 1:l with a-CD being the exception a t 2:l. ( c ) SDS Micellar Aggregation Numbers. In the Turro and Yekta13fluorescence quenching technique, the micellar aggregation number n is determined from the slope of the plot of In1against [SDSImic-l at fixed quencher
where I is the intensity of fluorescence. In agreement with the literature, n for pure SDS was found to be 59 f 5 at 25.0 ‘C. In these calculations, the concentration of free SDS monomer in micellar solutions was obtained from ref 20. To approximate the free SDS concentration by the cmc value would lead only to a 10%error in n because [SDSlmic > [SDSlfiee. When cyclodextrins were added to the micellar SDS solutions and the correct cmc values as determined conductimetrically (see section b) were used in Turro and Yekta’s equation, the calculated values for n for the SDS micelles were found to be 61 f4 for a-CD, 63 f 6 for p-CD, 64 f3 for y-CD, 58 f8 for GBBCD, and 52 f 8 for HPBCD. Each n value is the average over four to six runs taken a t different CD concentrations in the 2- 10 mM CD range. An average value of n taken over all the cyclodextrins is 60 f 6, which agrees within experimental error with n determined without any CD present. In effect, the cyclodextrins in solution appear to have minimal effect on the SDS aggregation number. The work of Dharmawardana et a1.21suggests that a further refinement in estimating the free SDS concentration in the presence of CD might be to take account of the opposing effects on the activity of the surfactant monomer of increased ionic strength and extra liberated counterions. Once again, this correction would have minimal effect on the SDS concentration in the aggregated state. The linearity of the In I plots (Figure 5) and the consistency in the calculated values give some assurance that [SDSlfi,, has been reasonably approximated from the work of Almgren and Swarup.20 The apparent indifference of the micelle aggregation number to the presence ofp-CD is in agreement with the ultrasonic relaxation results for the p-CD decyltrimethylammonium bromide8 and BCD sodium perfluorooctanoategsystems. Both studies revealed that the rate constants and other kinetic parameters associated with the monomer-micelle exchange process were minimally influenced by the presence of ,!?-CD in solution. The kinetics of this process is markedly dependent upon the micelle aggregation number and its polydispersity or size distribution. The competing equilibria operative in ,!?-CD micellar surfactant systems would be
+
+
+
M + p-CD 5 WP-CD and
M
+ Mn.-l 5 M,
where A4 and Mn-l are the surfactant monomer and micelle, respectively. The results of our findings and the ultrasonic work show that during the short time period of the ultrasonic relaxation, ca. loT8 s, the only effect W C D complex has on the monomer/micelle exchange kinetics is to “siphon”away monomeric surfactant, leaving the exchange rates and the micelle relatively unaffected. This implies that Kc >> Kmic, where Kmic can be ap(20) Almgren, M.; Swamp, S. J.Colloid Interface Sci. 1983,91,256. (21)Dharmawardana, U.R.; Christina, S. D.; Tucker, E. E.; Taylor, R. W.; Scamehorn, J. F. Langmuir 1993,9, 2258. (22) Okabu, T.; Maeda, Y.; Kitano, H. J.Phys. Chem. 1989,93,3721.
Langmuir, Vol. 11, No. 7, 1995 2479
SDS Micelles in Cyclodextrin Solutions
-
I -1.8
--2.0 t
20
i0
60
ISOSl;
80 / mM-‘
160
Figure 5. Fluorescence intensities of Ru(bipy)32+in SDS solutions with 8.06 mM p-CD present. proximated by the inverse ofthe cmc. For SDS, this means that K, >> 125 M-l or that the Gibbs free energy for a surfactant in the 1:l complex is lower than for the monomer in the micelle. Also it implies that the formation kinetics of the 1 : l complex are slower than those of the monomer-micelle exchange process. This is in agreement with stopped-flow studies21where the rate of formation of the a-CD/SDS complex is 110M-l whereas monomer exit times from micelles are on the order of lo6s--l.12Ifthe opposite was true, the amount of “free” or uncomplexed surfactant would decrease and could not be easily approximated by the cmc in micellar solution. Thus, even the presence of the 1 : l WP-CD complex appears to have little effect on the size and shape of the surrounding micelles in which they are likely not solubilized.
Conclusions For micellar surfactant systems, the effect of cyclodextrins on the micelles is probably minimal, even if the Gibbs free energy indicates that the surfactant is more stable in the complex than the micelle. Furthermore, for those
systems also containing 9-MeA, the Gibbs energy of transfer from the complex to the micelle is greater for the surfactant than for 9-MeA, except for the substituted cyclodextrins DIMEB and TRIMEB, where the surfactant was found to compete successfully with 9-MeA for the cyclodextrin cavity. Our results support the view that CDs or their 1:l surfactant complexes have little effect on either the monomer-micelle exchange kinetics, the average micelle aggregation number, or the polydispersity of the micelles sizes. Also, the kinetics of 1 : l complex formation is slower than the exit rate of the monomer from the micelle. For site specific delivery of drugs, the ramifications of these results are 2-fold: (1)CD does not alter the fast exchange kinetics between monomer and micelle, and thus dissolution of the membrane depends primarily on the Gibbs energies of transfer for the membrane material and the drug, and (2)if a drug molecule binds more strongly to the membrane than the complex, it can be delivered, as evidenced by the removal of 9-MeA, to the SDS micelle. However, this process could enhance the remove of cellular material from the membrane by favoring the formation of phospholipidCD complexes following the dissolution ofthe drug/CD complex. The CD complexis not solubilized or incorporated into SDS micelles, which suggests that it will have even less affinity for the more hydrophobic phospholipid membrane. The ideal situation would have both drug and the membrane surfactant show a stronger affinity for the membrane than for the complex.
Acknowledgment. We wish to thank the Natural Sciences and Engineering Council of Canada for financial support. LA940922D
