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Langmuir 2005, 21, 11034-11039
Preparation of a Highly Stable Niosome and Its Hydrotrope-Solubilization Action to Drugs Tianqing Liu*,† and Rong Guo‡ School of Chemistry and Chemical Engineering, Nanjing University, Jinagsu 210093, P.R. China, and School of Chemistry and Chemical Engineering, Yangzhou University, Yangzhou, Jinagsu 25002, P.R. China Received July 10, 2005. In Final Form: September 6, 2005 Highly stable niosomes are prepared and investigated in Tween80/PEG6000/Span80/H2O system. The mean radius of the niosomes is 0.15-0.2 µm. The contents of PEG6000 and Span80 and the system temperature affect the size and the stability of the niosome. A certain Span80 can remarkably improve the stability. The niosome is provided distinctly with the hydrotrope-solubilization action to the hydrophilic drug and hydrophobic drug, which affects the niosome membrane. The mechanism of the effects of PEG6000 and Span80 on the niosome is discussed in this paper.
Introduction Vesicles are an important form of molecular organizedassembles,1-4 and are unilamellar or multilamellar spheroid structures composed of amphiphilic molecules assembled into bilayers. They can be prepared from the phospholipid, nonionic surfactant and mixed cationic and anionic surfactant systems.5-16 Vesicles are widely used not only as the models for cell membranes, but also as * Corresponding author. E-mail:
[email protected]. Fax: +86514-7975244. Phone: +86-514-7975590-9517. † Nanjing University. ‡ Yangzhou University. Permanent corresponding address. (1) Mao, M.; Huang, J.; Zhu, B.; Ye, J. The Transition from Vesicles to Micelles Induced by Octane in Aqueous Surfactant Two-Phase Systems. J. Phys. Chem. B 2002, 106, 219-225. (2) Bryskhe, K.; Bulut, S.; Olsson, U. Vesicle Formation from Temperature Jumps in a Nonionic Surfactant System. J. Phys. Chem. B 2005, 109, 9265-9274. (3) Svenson, S. Controlling surfactant self-assembly. Curr. Opin. Colloid Interface Sci. 2004, 9, 201-212. (4) Hiemenz, P. C.; Rajagopalan, R. Principle of Colloid and Surface Chemistry, 3rd ed.; Marcel Dekker: New York, 1997; Chapter 8, pp 355-398. (5) Gebicki, J. M.; Hicks, M. Ufasomes are Stable Particles surrounded by Unsaturated Fatty Acid Membranes. Nature 1973, 243, 232-234. (6) Cistola, D. P.; Atkinson, D.; Hamilton, J. A.; Small, D. M. Phase Behavior and Bilayer Properties of Fatty Acids: Hydrated 1: 1 AcidSoaps. Biochmistry 1986, 25, 2804-2812. (7) Kunitake, T.; Okahata, Y.; Shimomura, M.; Yasunami, S.-i.; Takarabe, K. Formation of Stable Bilayer Assemblies in Water from Single-Chain Amphiphiles. Relationship between the Amphiphile Structure and the Aggregate Morphology. J. Am. Chem. Soc. 1981, 103, 5401-5413. (8) Kaler, E. W.; Murthy, A. K.; Rodriguez, B. E.; Zasadzinski, J. A. N. Spontaneous Vesicle Formation in Aqueous Mixtures of Dingle-Tailed Surfactants. Science 1989, 245, 1371-73. (9) Discher, D. E.; Eisenberg, A. Polymer Vesicles. Science 2002, 297, 967-268. (10) Su, Y.-L.; Li, J.-R.; Jiang, L. A study on the interactions of surfactants with phospholipid/polydiacetylene vesicles in aqueous solutions. Colloids Surf. A 2005, 257-258, 25-30. (11) Lee, J.-H.; Gustin, J. P.; Chen, T.; Payne, G. F.; Raghavan, S. R. Vesicle-Biopolymer Gels: Networks of Surfactant Vesicles Connected by Associating Biopolymers. Langmuir 2005, 21, 26-33. (12) Yin, H.; Huang, J.; Gao, Y.; Fu, H. Temperature-Controlled Vesicle Aggregation in the Mixed System of Sodium n-Dodecyl Sulfate/ n-Dodecyltributylammonium Bromide. Langmuir 2005, 21, 2656-2659. (13) Deo, N.; Somasundaran, P. Disintegration of Liposomes by Surfactants: Mechanism of Protein and Cholesterol Effects, Langmuir 2003, 19, 2007-2012. (14) Annable, T.; Buscall, R.; Ettelaie, R.; Shepherd, P.; Whittlestone, D. Influence of Surfactants on the Rheology of Associating. Langmuir 1994, 10, 1060-1070. (15) Yu, W.-Y.; Yang, Y.-M.; Chang, C.-H. Cosolvent Effects on the Spontaneous Formation of Vesicles from 1:1 Anionic and Cationic Surfactant Mixtures. Langmuir, 2005, 21, 6185-6193.
drug carriers to delivery to the targets of tumors and viruses.17-22 The size and the stability of vesicles are very important on the pharmacokinetics of vesicle-encapsulated drugs. The vesicles formed from liposome have been reported to have such functions as increasing drug stability, enhancing therapeutic effects, prolonging circulation time, and promoting uptake of the entrapped drugs into target site while drug toxicity is diminished.23 The vesicle formed from nonionic surfactant has been known as niosomes or nonionic surfactant vesicles. Noisomes are usually similar to liposomes in structure and properties. Liposomes are unilamellar or multilamellar spheroid structures composed of lipid molecules, often phospholipid, assembled into bilayers. Liposomes can carry hydrophilic drugs by encapsulating or hydrophobic drugs by partitioning these drugs into hydrophobic domains. However, the relatively low cost of the materials used to prepare niosomes make the vesicles more attractive and more useful than liposomes for industrial productions both in pharmaceutical and daily chemical applications. In addition, liposomes have some problems regarding degradation by hydrolysis of phospholipid molecules in aqueous system.17,23 The advantage of the simple method for the routine and large-scale production of niosomes is that it avoids the use of unacceptable solvents. (16) Ding, Y.; Liu, T.; Guo, R. Preparation of Vesicles from Lamellar Liquid Crystal in Triton X-100/n-C10H21OH/H2O System. Wuli Huaxue Xuebao 2000, 16, 481-486. (17) Alsarra, I. A.; Bosela, A. A.; Ahmed, S. M.; Mahrous, G. M. Proniosomes as a drug carrier for transdermal delivery of ketorolac. Eur. J. Pharm.Biopharm. 2005, 59, 485-490. (18) Nagayasu, A.; Uchiyama, K.; Kiwada, H. The size of liposomes: a factor which affects their targeting efficiency to tumors and therapeutic activity of liposomal antitumor drugs. Adv. Drug Delivery Rev. 1999, 40, 75-87. (19) Harashima, H.; Kiwada, H. Liposomal targeting and drug delivery: kinetic consideration. Adv. Drug Delivery Rev. 1996, 19, 425444. (20) Nagayasu, A.; Shimooka, T.; Kinouchi, Y.; Uchiyama, K.; Takeichi, Y.; Kiwada, H. Effects of fluidity and vesicle size on antitumor activity and myelosuppressive activity of liposomes loaded with daunorubicin. Biol. Pharm. Bull. 1994, 17, 935-939. (21) Schreier, H.; Bouwstra, J. Liposomes and niosomes as topical drug carriers: dermal and transdermal drug delivery. J. Controlled Release 1994, 30, 1-15. (22) Menger, F. M.; Angelova, Ml. Giant vesicles: imitating the cytological processes of cell membranes. Acc. Chem. Res. 1998, 31, 789797. (23) Manosroi, A.; Wongtrakul, P.; Manosroi, J.; Sakai, H.; Sugawara, F.; Yuasa, M.; Abe, M. Characterization of vesicles prepared with various nonionic surfactants mixed with cholesterol. Colloids Surf. B 2003, 30, 129-138.
10.1021/la051868b CCC: $30.25 © 2005 American Chemical Society Published on Web 10/12/2005
Preparation of a Highly Stable Niosome
Tween80 and Span80 are pharmaceutically acceptable, innocuous, nonionic biological surfactants.24,25 Innocuous PEG-based surfactants show high selectivity in disrupting vesicular membrane.26-28 For this work, Tween80 and Span80 with poly(ethylene glycol) (PEG) were selected to prepare highly stable niosomes (the period of the stability is more than a year). Then, in this paper, the size, the stability, and the dilutability of the niosomes are discussed, and finally, a possible mechanism of the stability is put forward. The niosomes prepared were provided with hydrotrope-solubilization action to ribavirin (hydrophilic drug)29-31 and ibuprofen (hydrophobic drug)32-34 in the solution. Materials and Methods Materials. Tween80 was bought from Shanghai Biological Engineering Ltd (99%), Span 80 from Shanghai Commonage Pharmacy Co. (99%), and ribavirin and ibuprofen from Hubei Qianjiang Pharmaceutical Co. Ltd (99%, material drug). PEG800-12000 was from Shanghai Pudong Goulian Chemical Plant. Doubly distilled and deionized water was used for the preparation of the solutions. The molecular structures of the materials are represented as follows:
Niosome Preparation. Tween80, PEG, and water were vortex mixed at a certain molar fraction. The niosome was prepared by sonicating the mixed nonionic surfactant solution for 6-25 min (CQX25-06 sonicator, Shanghai Bilieng Sonicator Co. Ltd). Span80 was added to the Tween80/PEG/water niosome. Then, the sample was sonicated for 5-25min. The stability of (24) Arnesen, S.; Eriksen, S. H.; Olsen, J.; Jensen, B. Increased production of a-amylase from Thermomyces lanuginosus by the addition of Tween 80. Enzyme Microb. Technol. 1998, 23, 248-252. (25) Gianasi, E.; Cociancich, F.; Uchegbu, I. F.; Florence, A. T.; Duncan, R. Pharmaceutical and biological characterization of a doxorubicin-polymer conjugate (PK1) entrapped in sorbitan monostearate Span 60 niosomes. Int. J. Pharm. 1997, 148, 139-148. (26) Park, M.-J.; Chung, Y.-C.; Chun, B. C. PEG-based surfactants that show high selectivity in disrupting vesicular membrane with or without cholesterol. Colloids Surf. B 2003, 32, 11-18. (27) Needham, D.; Kim, D. H. PEG-covered lipid surfaces: bilayers and monolayers. Colloids Surf. B 2000, 18, 183-195. (28) Hui, S. W.; Kuhl, T. L.; Guo, Y. Q.; Israelachvili, J. Use of poly(ethylene glycol) to control cell aggregation and fusion. Colloids Surf. B 1999, 14, 213-222. (29) Enserink, M. SARS treatment: Interferon shows promise in monkeys. Science 2004, 303 (5662), 1273-1274. (30) Crabb, C. Hard-won advances spark excitement about hepatitis C. Science 2001, 294 (5542), 506-507. (31) Vo, N. V.; Young, K.-C.; Lai, M. M. C. Mutagenic and Inhibitory Effects of Ribavirin on Hepatitis C Virus RNA Polymerase. Biochemistry 2003, 42, 10462-10471. (32) Pathak, P.; Meziani, M. J.; Desai, T.; Sun, Y.-P. Nanosizing Drug Particles in Supercritical Fluid Processing. J. Am. Chem. Soc. 2004, 126, 10842-10843.
Langmuir, Vol. 21, No. 24, 2005 11035 niosome was studied by measuring the particle size and the mean size distribution as a function of time following vesicle preparation using a computerized micrographs image analyzing meter (model SK-882, Sanko Co., Japan) (imaging microgram). The particle size distribution of the niosome varied very little after 48 h of the preparation. Freeze Fracture Replication-Electron Microscopy (FFTEM) of Niosome. The samples for transmission electron microscopy were prepared by freeze-fracture replication according to standard techniques. Fracturing and replication were carried out in a high vacuum freeze-etching system (Balzers BAF-400D). Replicas were examined in a transmission electron microscope (model TECNAIR, Philip Apparatus Co.). The polydispersity was less than 0.03, which indicates that the distribution is homogeneous. Viscosity Determination. Viscosity measurement of the solution was performed on a low-shear rheoanalyzer (Haake Rheostress 600, Thermo Electron Corporation, German) with Haake Rheopwin data manager. All samples were equilibrated for 48 h before the measurement. Each result was the average of six runs. Influence of Drug on Niosome Membrane. The influence of drug on the niosome membrane was reflected by the steadystate fluorimetry with pyrene (1.4 × 10-6 mol‚L-1) as probe by using a RF-5310PC fluorescence spectrophotometer (Shimadzu Co., Japan). Pyrene has five emission peaks when it is excited at 338 nm. The intensity ratio (I1/I3) of the first emission peak (373 nm) to the third emission peak (383 nm) indicates the microenvironment where the probe exists, from which the influence of drug added to the niosome membrane can be understood.35-37 All the solutions were deoxygenated by bubbling pure nitrogen for 15 min before measurements. Content of the Component Determination by HPLC. Each sample was centrifuged at 13000 rpm for 10 min. The supernatant and the precipitate were separated out. A 20% ethanol aqueous solution was added to the supernatant and the precipitate, mixed round. Then, the temperature of each sample was increased in order to disrupt the niosome entirely. Later, the sample was centrifuged at 13000 rpm for 10 min again. Finally, the contents of the components for the each sample were measured by a CLASS-VP HPLC system (Shimadzu Co., Japan) with model SPD-M10AVP UV-vis detectors and an autosampler. The chromatographic data were analyzed using a CLASS-VP chromatographic manager (Shimadzu). A Shim-pack CLC-ODS column (150 mm × 6.0 mm, i.d. 10 µm) was used for the HPLC separations. The flow rate was set to 1.0 mL/min and the effluent was monitored with UV-vis double wavelength detection. The mobile phases were methanol and water. Determination of the Distribution Coefficient of Drug in Niosome. The UV-vis difference spectra of the niosome in water were taken on an RF-5301PC UV-vis Spectrum spectrometer (Shimadzu Co. Japan). The range of the scanning wavelength was 700-190 nm. The size of the quartz utensil was 1 cm × 1 cm. From the absorbance spectrum of the drug in the niosome system, the distribution coefficient KD vales of the drug between the niosome and continuous phase were calculated by least-squares method. All measurements had been carried out at 25 °C. (33) Andersson, J.; Rosenholm, J.; Areva, S.; Linde’n, M. Influences of Material Characteristics on Ibuprofen Drug Loading and Release Profiles from Ordered Micro- and Mesoporous Silica Matrices. Chem. Mater. 2004, 16, 4160-4167. (34) Melillo, M.; Gun’ko, V. M.; Tennison, S. R.; Mikhalovska, L. I.; Phillips; G. J.; Davies, J. G.; Lloyd, A. W.; Kozynchenko, O. P.; Malik, D. J.; Streat, M.; Mikhalovsky, S. V. Structural Characteristics of Activated Carbons and Ibuprofen Adsorption Affected by Bovine Serum Albumin. Langmuir 2004, 20, 2837-2851. (35) Thomas, J. K. Radiation-Induced Reactions in Organized Assemblies. Chem. Rev. 1980, 80, 283-299. (36) Kalyanasundaram, K.; Thomas, J. K. Solvent-Dependent Fluorescence of Pyrene-3-carboxaldehyde and Its Applications in the Estimation of Polarity at Micelle-Water Interfaces. J. Phys. Chem. 1977, 81, 2176-2180. (37) Kalynasundaram, K.; Thomas, J. K. Environmental Effects on Vibronic Band Intensities in Pyrene Monomer Fluorescence and Their Application in Studies of Micellar Systems. J. Am. Chem. Soc. 1977, 99, 2039-2044.
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Figure 1. Micrographs of the niosome in Tween 80/PEG6000/H2O system. (a) Imaging micrograph (1:5000). (b) FF-TEM micrograph. The molar ratio of Tween 80/PEG6000/H2O ) 92.1/1.00/667.
Results and Discussion Effects of PEG on the Radius and the Stability of Niosome. Tween80, PEG, and H2O are mixed and surged at the mass ratio of Tween80/PEG/H2O ) 0.500/0.0100.060/0.150-0.500 for 2 min, sonicated for 6-25min. Niosome has not been found in Tween80/PEG800/H2O system. But niosome appears and can stabilize in the Tween80/PEG6000/H2O system (Figure 1). The size of the niosome is almost uniform which can be seen from the FF-TEM in Figure 1b (the size is apparently different in Figure 1a because the image appears larger if the niosomes are close up to the ocular, while the image looks smaller if the niosomes are far off the ocular). The size in the imaging micrograph is almost consistent with that in the FF-TEM micrograph. The niosome is also not found at the discretional mass ratio of Tween80/PEG12000/H2O. If the molecular mass of PEG is at a low level, its chain is short and the viscosity of the system is low.27 Its strong hydrophilic ability makes it difficult to form the niosome structure with organized-assembles. Even if niosome is formed, the thickness and intensity of the niosome film is not great. Therefore, the niosome is easily disrupted.38,39 However, if the molecular mass of PEG is very large, its chain length can be very long. The viscidity of the system is high and the surface tension is low. The accessional pressure and the inner stress are so high that the formation of noisomes are handicapped. From the imaging microgram and FF-TEM of the niosome, the stability and the radius of the niosome depend on the composition of the system and the preparation technique (Figure 2). Single PEG6000 in aqueous solution cannot assemble to form vesicles. But the niosome can be stable in the Tween80/PEG6000/H2O system. Its mean radius is 0.15-0.2 µm. The viscosity of the system (1.081.15 × 10-3 kg‚m-1‚s-1) is greater than the viscosity of water (1.00 × 10-3 kg‚m-1‚s-1), which indicates that the niosome membrane may be a multilamellar vesicle but not a unilamellar vesicle. With the molar fraction of PEG6000 increasing, the stability enhances and the radius increases. Within 2.42-3.01 × 10-4 (molar fraction) PEG6000, the stability of the niosome is more than 360 days. Besides, the radius and the number of the niosome do not change obviously in a period of one year (year period). But, when the molar fraction of PEG6000 is more (38) Nicholas, A. R.; Scott, M. J.; Kennedy, N. I.; Jones, M. N. Effect of grafted polyethylene glycol (PEG) on the size, encapsulation efficiency and permeability of vesicles. Biochim. Biophys. Acta 2000, 1463, 167178. (39) Nikolova, A. N.; Jones, M. N. Effect of grafted PEG-2000 on the size and permeability of vesicles. Biochim. Biophys. Acta 1996, 1304, 120-128.
Figure 2. Stability time and radius of the niosome related to the molar fraction of PEG600. Parts a and b point to the stability time and the radius, respectively. The filled squares and circles correspond to Tween80/PEG6000/H2O system. The hollow squares and circles correspond to Tween80/PEG6000/Span80/ H2O system. The hollow and filled squares correspond to 25 °C. The hollow and filled circles correspond to 37 °C.
than 3.01 × 10-4, the formation rate of the niosome, the stability, and the radius all decrease. More hydrophilic PEG6000 leads to the increasing hydrophilicity of the niosome and the decreasing thickness and the intensity of the film until the structure with organized assembly is disrupted. During the experiments, the radius and the number of the niosome decrease when the system temperature goes up from 25 to 37 °C (Figure 2). On one hand, the increase
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Figure 3. Effect of Span80 content on the niosome at the molar ratio of Tween80/PEG6000/H2O ) 92.1/2.00/667 (imaging microgram) (1:3000). Parts a, b, c, and d correspond to the molar fraction of Span80 from 8.70 × 10-4, 2.61 × 10-3, 4.19 × 10-3, and 5.76 × 10-3, respectively.
Figure 4. Stability time of the niosome varies with the content of water added in the niosome. The molar ratio of Tween80/ PEG6000/Span80/H2O ) 70.2/1.00/94.3/1333. The filled squares and circles correspond to 25 and 37 °C, respectively.
of the temperature quickens the molecular moving rate. The organized assembly is partially disordered and disrupted; on the other hand, the molecular interaction is weakened when the temperature increases, and the surface tension descends. The intensity of the niosome film falls. So, the stability of the niosome decreases. The average radius is about 0.162 µm with 1.80-3.31 × 10-4 (molar fraction) PEG6000 at 37 °C. Effects of Span80 on the Stability of Niosome. The stability of the niosome is about 20 days in the Tween80/ PEG6000/H2O system. However, vesicles are not easily formed in the Span80/PEG6000/H2O system. If a certain amount of Tween80 is added to the Span80/PEG6000/ H2O system, niosome can be formed much more easily. With the increasing molar fraction of Span80 in the Tween80/PEG6000/H2O system, the stability of the niosome is markedly enhanced. Its radius also increases (Figures 2 and 3). Here, the PEG6000 molar fraction that forms the stable niosome is enlarged to 9.02 × 10-4. At the molar ratio Tween80/PEG6000/H2O ) 92.1/2.00/667, Span80 can obviously prolong the stable period of the niosome to about a year (the niosome is already placed for one year, its size and number do not evidently change). But when the molar fraction of Span80 is more than 3.61 × 10-4, Span80 with more hydrophobic ability decreases the radius and the stability. Niosomes can all be formed in the molar ratio of Tween80/PEG6000/Span80/H2O ) 45.8-124/1.00-25.0/210/600-2200. If water is added to the Tween80/PEG6000/Span80/ H2O niosome, the niosome is still stable for some days. Figure 4 shows the relation of the stability time of the niosome to water content. The niosome diluted with 4 times water can be stable for more than 50 h. However,
vesicles cannot experimentally be formed by any methods at the molar ratio of Tween80/PEG6000/Span80/H2O ) 70.2/1.00/94.3/5332 in the 10-40 °C range. This result indicates that the film of the niosome is provided with stronger intensity and stability. Once the niosome is formed, it is difficult to be disrupted. This also implies that the niosome possesses good diluted capability. The diluted niosome is quite sensitive to the temperature (Figure 4). With increasing temperature, the molecular thermodynamic moving rate goes up. The molecular interaction and the interface tension are weakened. The intensity and stability of the niosome film all decrease. However, this phenomenon can provide more important information for the niosome as a drug carrier in human practice applications. Microstructure and Stable Mechanism of Niosome. The contents of PEG6000 and Span80 all affect the size and the stability of the niosome, which can be seen from the imaging microgram and FF-TEM (Figures 1-3). The influence of Span80 on the niosome size is quite obvious (Figures 3). If the niosome is dropped on a quartz piece, the niosome is dried gradually with the volatilization of water. The surface of the niosome begins to crinkle (Figure 5). This phenomenon indicates that the niosome film is provided with definite intensity and tenacity. This may be the important reason of the niosome stability. The desiccated niosomes disappear at last (Figure 5d). The mechanism of the effect of PEG6000 and Span80 on the niosome stability may be as follows: The rigidity of the niosome formed from Tween80 with more hydrophilic ability is weak. The long hydrophilic chains in PEG6000 are mostly coiled and adsorbed on the niosome and act as a stabilizing agent; some insert partially in the film phase of the niosome (Figure 6). The rigidity and stability of the niosome film thus are enhanced. Alternately, PEG can cause the niosome fusion or disruption of the bilayer structure. The addition of PEG can give rise to faceted vesicles. Finally, the addition of an associating polymer can result in a “vesicle gel”, where adjacent vesicles are bridged by polymer chains.40 Adding Span80 with more hydrophobic ability and moderate hydrophilic ability to the niosome can improve the hydrophobic capability and the intensity of the niosome film. Span80 exists in the interphase of the film phase, decreases the interaction between polar heads of the amphiphilic molecules, and thus stabilizes the niosome. The hydrophobic interaction results in the insertion of the alkyl chain of the surfactants into the hydrophobic domain of the niosome.23 When the molar fraction of Span80 is more (40) Antunes, F. E.; Marques, E. F.; Gomes, R.; Thuresson, K.; Lindman, B.; Miguel, M. G. Network Formation of Catanionic Vesicles and Oppositely Charged Polyelectrolytes. Effect of Polymer Charge Density and Hydrophobic Modification. Langmuir 2004, 20, 46474656.
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Figure 5. Imaging microgram of the microstructure of the niosome in dehydrating and drying process (1:2000): (a) niosome in the solution, (b) niosome after dehydrating 5 min, (c) niosome after dehydrating 10 min, (d) niosome after dehydrating 15 min. Table 1. Effect of Ribavirin Content on the Micropolarity xribavirin × 103
0
1.03
3.09
5.15
7.21
9.27
11.3
I1/I3
1.23
1.20
1.17
1.13
1.11
1.10
1.09
Table 2. Effect of Ibulefen Content on the Micropolarity
Figure 6. Mechanism of the effects of PEG6000 and Span80 on the niosome stability.
than 4.19 × 10-3, more hydrophobic action leads to the relaxation or the reversal of the niosome film and its stability decreases and finally reaches disruption. Hydrotrope-Solubilization Action of Niosome to Drug. Ibuprofen dissolved slightly in water and ribavirin with the solubility about 8.5% can be successfully encapsulated in the niosome at the molar ratio of Tween80/ PEG6000/Span80/H2O ) 70.2/1.00/94.3/1333. The maximal solubilities of ibuprofen and ribavirin are 0.45% and 11.23% (their molar fractions are 5.05 × 10-4 and 1.16 × 10-2) in the niosome, respectively. Because of the hydrophilicity of the film surface and the hydrophobic ability of the film cavity, the hydrotrope-solubilization action of the niosome is obvious to hydrophilic drug (ribavirin) and hydrophobic drug (ibuprofen) in the solution. The encapsulation of drugs in niosomes can decrease drug toxicity, increase drug absorption, and retard the loss of drug from the circulation due to slow drug release. When the molar fractions of the drug are less than 2.06 × 10-3, the drug content does not affect the size and the stability of the niosome distinctly. If the molar fractions are more than 2.06 × 10-3, the increase of the molar fractions causes the size and the stability to decrease slowly.
xbuleden × 104
0
0.561
1.12
1.68
2.24
3.37
4.49
I1/I3
1.23
1.21
1.19
1.16
1.14
1.11
1.07
The effect of drug on the niosome membrane can be confirmed by the micropolarity (I1/I3) change of pyrene as probe. Tables 1 and 2 show the effects of ribavirin and ibulefen on the micropolarity of the niosome. With the molar fraction of drug increasing, the micropolarity decreases. This result indicates that the drug is solubilized in the film bilayer of the niosome. The solubilized drug makes the pyrene probe gradually move inside the film phase. A little content of the small molecular drug and the small change of the probe position are not sufficient to change the membrane structure and stability of the niosome. However, the greater content of the small molecular drug with interspace structure and short chain in the film phase loosens the film structure and reduces the film stability slightly. The behaviors of ribavirin are disabled in water after 48 h. However, the ribavirin solubilized in the niosome can be stable for 30 days according to the UV-vis difference spectra of ribavirin and the HPLC, while the niosome is destroyed with the increase of the system temperature and the addition of ethanol. The ibulefen solubilized is stabilized for 45 days. The probable reason is that the drug is solubilized in the niosome in which hidebound water and oxygen are difficult to penetrate into the niosome. Distribution of Drug in Niosome. The distribution coefficient KD of ribavirin between the niosome and water continuous phase can be determined by hypervelocity sonicating centrifugal separation and the distribution coefficient method of UV-vis difference spectra.41
1 1 1 1 × + ) Eψ - Ew KD(Em - Ew) cS Em - Ew
(1)
Here, cS is the surfactant concentration. EΨ is the apparent mole absorption coefficient of drug at a given wavelength. Ew and Em are the apparent mole absorption coefficients of the drug in water continuous phase and in niosome, respectively. EΨ and Ew can be determined experimentally. So, plotting 1/(EΨ - Ew) against 1/cS (Figure 7), the distribution coefficient KD can be calculated from the slope (41) Wenbin, Q.; Lizhong, Z. Study of the effect of mixed ionic -nonionic surfactants on color reactions and its application. III. Distribution constants of color reactions between micellar phase and water. Huaxue Xuebao 1987, 45, 707.
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The molar Gibbs free energy change, ∆Gm, of ribavirin distributed between the niosome and water continuous phase is given as
∆Gm ) -RT ln KD
(2)
∆Gm for ribavirin is -32.56 and -23.25 kJ‚mol-1 at 25 and 37 °C, respectively. The hydrotrope-solubilization of the niosome to drug is spontaneous. The increase of temperature results in the increasing molecular moving rate and the stability and the decreasing encapsulation efficiency of the niosome to drug. Conclusion
Figure 7. Plotting of 1/(EΨ - EW) against 1/cD. The filled squares and circles correspond to 25 and 37 °C, respectively.
and the intercept of the linearity. KD for ribavirin is 5.092 × 105 and 8.735 × 103 at 25 and 37 °C, respectively. Because ibulefen is not dissolved in water, it may be entirely solubilized in the niosome. Therefore, the encapsulation efficiency of the niosome to hydrophilic drug and hydrophobic drug is quite large.
Biologically stable niosome can be prepared in the existence of PEG with moderate chain length in the Tween80/PEG/Span80/H2O system. The niosome proves to have good diluted capability, and the niosome film has stronger intensity and stability. PEG and Span80 all affect the size and the stability of the niosome. The hydrotropesolubilization action of the niosome is remarkable for the hydrophilic drug and hydrophobic drug in the solution. Acknowledgment. This work was supported by the National Natural Science Foundation of China (20233010, 20573091). LA051868B
