Preparation and Characterization of the Micelle-Forming Polymeric Drug Indomethacin-Incorporated Poly(ethylene oxide)-Poly(-benzyl L-aspartate) Block Copolymer Micelles SUNG BUM LA † ,TERUO OKANO †‡ , AND KAZUNORI KATAOKA †§X Received May 18, 1995, from the † International Center for Biomaterials Science, Research Institute for Biosciences, Science University of Tokyo, Yamazaki 2669, Noda-shi, Chiba 278, Japan, ‡ Institute of Biomedical Engineering, Tokyo Women's Medical College, Kawada-cho, Tokyo, Japan, and § Department of Materials Science and Technology and Research Institute for Bioscience, Science University of Tokyo, Yamazaki 2641, Noda-shi, Chiba 278, Japan. Accepted for publication September 12, 1995 X . Abstract 0 To estimate the feasibility of novel containers for drugs, poly- (ethylene oxide)-poly(-benzyl L-aspartate) (PEO-PBLA) micelles were prepared by dialysis against water using different solvents. The solvent selected is very important because it drastically affects the stability of polymeric micelles. The critical micelle concentration (cmc) of the prepared micelles in distilled water was determined by a fluorescence probe technique using pyrene. Indomethacin (IMC) as a model drug was incorporated into the micelles by dialysis and an oil/water emulsion method. Characteristics of PEO-PBLA micelles without and with the physically trapped IMC in the inner core of the micelles (IMC/PEO-PBLA) were studied by dynamic light scattering and gel permeation chromatography/ HPLC as well as an in vitro release test of IMC from the micelles. For the PEO-PBLA block copolymers, N,N-dimethylacetamide (DMAc) was found to be the best of the solvents tested to form stable polymeric micelles with a narrow size distribution and avoid its aggregation, and the cmc of PEO-PBLA micelles thus prepared was determined to be ca. 18 mg/L in water. The diameters of PEO-PBLA micelles and IMC/PEO- PBLA micelles in number averaged scale were observed to be ca. 19 and 25-29 nm, respectively. The release study of IMC from IMC/PEO- PBLA micelles in various buffer solutions at the pH range from 1.2 to 7.4 at 37 °C revealed that the release rate of IMC from the micelles was increased by increasing the pH of the medium and indicated that the release rate of IMC from the micelles are controlled by the partition coefficient of IMC based on the pH of the medium and interaction between IMC and the hydrophobic portion of the micelles. An amphiphilic AB type block copolymer is made of two segments of different chemical structure with hydrophilic and hydrophobic components. This system has been the focus of numerous investigations, because of the intrinsic interest in self-assembling systems. 1-5 This type of diblock copolymer can form a spherical micelle structure in an aqueous milieu. The hydrophobic segment forms the hydrophobic core of the micelle, while the hydrophilic segment surrounds this core as a hydrated outer shell. This core-shell structure provides polymer micelles with the potential for use as vehicles for drug delivery, since the hydrophobic core may serve as a micro- container for drugs which is segregated from the outer environment by a palisade-like hydrophilic segment. Kataoka and co-workers have been investigating micelle-forming block copolymer-drug conjugates 6-8 and block copolymer micelles with physically entrapped drugs in the field of drug deliv- ery. 9,10 Since most drugs have a hydrophobic character, these drugs can be easily incorporated into the micelle by chemical conjugation to block copolymers or by simple physical entrap- ment into the micelle core through dialysis or an oil/water (O/W) emulsion method. Poly(ethylene oxide) (PEO) is known to be a nontoxic and nonimmunogenic water soluble polymer. In recent years, derivatives of PEO have been proven to be valuable in a variety of diverse chemical and biological endeavors. Such applications include peptide synthesis, phase transfer cataly- sis, pharmaceutical modification, protein and cell purifica- tions, polymer-bound reagents, and binding assays. 11 A final property of PEO which has been of much importance in its commercial application is its lack of toxicity. 12 It has been used in protein modification to decrease the antigenicity 13,14 of the intact protein and to prolong its half-life in the blood stream. 15,16 Furthermore, PEO has been used for protein modification 17 and surface modification of microspheres 18 to decrease hepatic uptake. Considering these properties, PEO is suitable for the hydrophilic segment of the block copolymer to form the outer shell of the micelle. IMC, 1-(4-chlorobenzoyl)-5-methoxy-2-methyl-1H-indole-3- acetic acid, as shown in Figure 1A, is one of the most potent nonsteroidal anti-inflammatory drugs (NSAIDs) for the treat- ment of conditions such as rheumatoid arthritis, ankylosing spondylitis, osteoarthritis, and acute gouty arthritis. Also, it has analgesic properties distinct from its anti-inflammatory effects, and there is evidence for both a central and a peripheral action; it is also an antipyretic. 19 The mechanism of action of IMC is to suppress prostaglandin synthesis in the tissues via inhibition of cyclooxygenase activity. 20 IMC, however, has side effects such as irritation of gastrointestinal mucosa from direct exposure to drug and toxicity to central nervous system as a necessary consequence of high plasma levels following administration of the conventional nonfor- mulated product. 21 In order to overcome the side effects of IMC, several experimental models, such as encapsulation, 22 film coating, 23 and matrix form, 24 were studied. Lin and Kawashima also studied the solubility behavior and stability of IMC by using ABA triblock copolymers, poly(ethylene oxide)-poly(oxypropylene)-poly(ethylene oxide) (Pluron- ics). 25,26 In this paper, IMC-incorporated micelles based on AB block copolymers of PEO and poly(-benzyl L-aspartate) (PBLA) were prepared as an alternative approach to decrease the side effects of IMC. In particular, PBLA was chosen as hydropho- bic poly(amino acid) segments because of its biodegradable nature with lower toxicity. One of the principal advantages of poly(ethylene oxide)-poly(-benzyl L-aspartate) (PEO- PBLA, Figure 1B) block copolymers is based on its simple structure, which leads to an easier determination of its physicochemical properties, including micelle formation. The objective of this research is to evaluate the possibility of using PEO-PBLA polymeric micelles as novel carriers for IMC delivery. Experimental Section MaterialssR-Methoxy-ω-amino-poly(ethylene oxide) [CH3O-poly- (ethylene oxide)-NH2, MW ) 12 000, Mw/Mn ) 1.10, purity ) 95.3%] and -benzyl L-aspartate were supplied by Nippon Oil and Fat, Co., X Abstract published in Advance ACS Abstracts, October 15, 1995. © 1996, American Chemical Society and 0022-3549/96/3185-0085$12.00/0 Journal of Pharmaceutical Sciences / 85 American Pharmaceutical Association Vol. 85, No. 1, January 1996