Communication pubs.acs.org/crystal
Bile Salts as Crystallization Inhibitors of Supersaturated Solutions of Poorly Water-Soluble Compounds Jie Chen,† Laura I. Mosquera-Giraldo,† James D. Ormes,‡ John D. Higgins,§ and Lynne S. Taylor*,† †
Department of Industrial and Physical Pharmacy, College of Pharmacy, Purdue University, West Lafayette, Indiana 47907, United States ‡ Discovery Pharmaceutical Sciences, Merck Research Laboratories, Merck & Co., Inc., Rahway, New Jersey 07065, United States § Discovery Pharmaceutical Sciences, Merck Research Laboratories, Merck & Co., Inc., West Point, Pennsylvania 19486, United States S Supporting Information *
ABSTRACT: Poor aqueous solubility of new therapeutic agents is one of the most pressing current challenges in drug delivery. Supersaturated solutions, generated in vivo either by virtue of the intrinsic compound properties (i.e., ionization behavior) or by dissolution of formulations containing a high energy form of the drug, are considered one of the most promising approaches to improve drug absorption, when it is limited by solubility. The challenge is to maintain supersaturation, typically achieved by adding polymeric crystallization inhibitors, for biologically relevant time periods. In this study, we present exciting preliminary observations about the potential role of bile salts as crystallization inhibitors. Focusing on sodium taurocholate, an endogenous bile salt also widely used in simulated intestinal media, extended experimental nucleation induction times were observed in the presence of this surfactant for a group of 11 diverse compounds, relative to induction times in buffer. Limited studies with two additional bile salts suggested other members of the bile salt family may also have crystallization inhibition properties. Given the endogenous nature of bile salts, as well as their use in simulated intestinal media, these observations are clearly of potential importance when assessing the supersaturation behavior of poorly water-soluble compounds.
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family, was found to have an inhibitory effect on the nucleation of celecoxib from supersaturated solutions.16 In contrast, in another study, STC was found to promote the crystallization of carbamazepine dihydrate during dissolution of the anhydrous crystal.17 STC and other bile salts are synthesized in the liver and are cholesterol derivatives. Micelle-forming bile salts can increase the solubility of poorly water-soluble compounds. Together with phospholipids and phospholipid hydrolysis products present in vivo, mixed micelles can be formed, and these are also good solubilizers of drugs.18 Because of their ability to lower the interfacial tension, bile salts contribute to the emulsification of fats in the intestine and their subsequent absorption from the gastrointestinal (GI) tract. Bile salts have an anionic hydrophilic headgroup and a rigid steroid ring system that exhibits facial polarity (hydrophilic and hydrophobic edges) as shown in Figure 1. The goal of the current study was to evaluate the ability of STC to delay nucleation from supersaturated solutions of a group of structurally diverse poorly soluble compounds. The nucleation induction time of each compound from a medium containing a bile salt was compared to the nucleation induction time from the same medium without the bile salt. For comparative purposes, similar experiments were performed in
p to 80% of candidates in the contemporary drug discovery and development pipelines suffer from poor water solubility which directly limits both bioavailability and potential for further development.1 Some of these drug candidates can be rescued from failure by formulation strategies, e.g., solubilization in surfactant micelles and lipids,2 cosolvents,3 amorphous solid dispersions,4−6 and nanocrystals.7−9 Solid dispersions contain the drug as an amorphous form, the theoretical solubility of which is usually several times higher than that of the crystalline form.10−13 Amorphous drugs can quickly dissolve in aqueous media under nonsink conditions and reach a concentration much higher than the equilibrium solubility of the crystalline form; however, this high concentration is thermodynamically unstable, and the system has a tendency to nucleate to the more stable crystalline form. Once nucleation of the crystalline form commences, the solution concentration of the drug will begin to drop back toward the equilibrium solubility of the crystalline form. Therefore, it is of great interest to find effective ways to slow/ inhibit nucleation from supersaturated solutions so that the solubility enhancement can persist long enough to allow drug molecules to be absorbed following oral administration. Polymeric additives, e.g., poly vinylpyrrolidone (PVP) based polymers, and cellulose derivatives, are the primary type of additives currently being used to stabilize solid dispersions and to inhibit nucleation to achieve solubility enhancement.14,15 Recently, sodium taurocholate (STC), a member of the bile salt © 2015 American Chemical Society
Received: March 20, 2015 Revised: April 26, 2015 Published: April 27, 2015 2593
DOI: 10.1021/acs.cgd.5b00392 Cryst. Growth Des. 2015, 15, 2593−2597
Crystal Growth & Design
Communication
Figure 1. Structures of sodium taurocholate (STC), sodium glycocholate (SGC), and sodium glycodeoxycholate (SGDC).
centrifugation at 35 000 rpm for 20 min at 25 °C (37 °C for telaprevir), using an Optima L-100 XP ultracentrifuge equipped with a swinging-bucket rotor SW 41 Ti (Beckman Coulter, Inc., Brea, CA). The concentration of the supernatant was determined either using an Agilent 1260/1290 Infinity Series HPLC system (Agilent Technologies, Santa Clara, CA) or a SI Photonics UV/vis spectrometer (Tuscon, Arizona), fiber optically coupled with a dip probe, dependent on the drug. Details of the solubility determination method for each drug can be found in the Supporting Information (Tables S1−S3). 2. Nucleation Induction Time Measurements. It is not possible to directly measure nucleation time, because the nuclei formed can only be detected after they have grown to an experimentally detectable size. Therefore, the experimental nucleation induction time, tind, can be defined as the sum of the time for critical nucleus formation (true nucleation time, tn) and growth to detectable size, tg.21
a medium containing a commonly used anionic surfactant, sodium dodecyl sulfate (SDS). Through these explorations, we hope to shed light on the potential role of bile salts as crystallization inhibitors for supersaturating dosage forms. Materials and Methods. (a) Materials. Sodium taurocholate (hydrate, ≥ 97%, Sigma, MO), sodium glycocholate (hydrate, ≥ 97%, Sigma, MO), sodium glycodeoxycholate (≥97%, Sigma, MO), and SDS (≥99.0%, Sigma, St. Louis, MO) were used at received. Eleven drug compounds with different physicochemical properties were studied, as listed in Table 1. Compound 1 Table 1. List of Compounds Used, Their Ionization Properties, Supplier, and Media Used for the Induction Time Experiments name
property
celecoxib
neutral
griseofulvin
neutral
felodipine
neutral
tadalafil
neutral
telaprevir
neutral
warfarin
acid, pKa of 5.0 acid, pKa of 4.15 acid, pKa of 4.5 base, pKa of 6.22 base, pKa of 2.8 base, pKa 1.9 and 3.4
naproxen indomethacin bifonazole nevirapine compound 1
source Attix Pharmaceuticals Sigma-Aldrich Attix Pharmaceuticals Attix Pharmaceuticals Attix Pharmaceuticals Sigma-Aldrich Sigma-Aldrich Hawkins, Inc. Spectrum Chemical Chempacific Merck
media HCl solution, pH 3 phosphate buffer pH 2, 50 mM phosphate buffer pH 6.5, 50 mM phosphate buffer pH 6.5, 50 mM phosphate buffer pH 6.5, 50 mM phosphate buffer pH 2, 50 mM citric buffer pH 3.5, 50 mM citric buffer pH 3.5, 50 mM carbonate-bicarbonate buffer, pH 9.6, 50 mM phosphate buffer pH 6.5, 50 mM citric buffer pH 4, 50 mM
t ind = tn + tg
(1)
Supersaturation was generated by titrating an appropriate amount of a concentrated drug stock solution (drug dissolved in an organic solvent) to 50 mL of the test solution (buffer or buffer containing 0.1% surfactant) while stirring using a crossshaped magnetic stirrer at 300 rpm. The induction time was determined from the increase in intensity of light scattered (extinction) from the drug solutions upon evolution of particles. Light scattering was detected by monitoring the extinction at a nonabsorbing wavelength (350−450 nm) using a SI Photonics UV/vis spectrometer (Tuscon, Arizona), fiber optically coupled with a dip probe (path-length was typically 10 mm, a shorter path length was used for higher concentration solutions). A typical example of changes in the extinction signal as a function of time is shown in Figure S1, Supporting Information. The plot also shows changes in the absorption signal at an absorption maximum whereby the signal decreases in concert with the increase in extinction. Each experiment was repeated three times to calculate the average induction time. SDS solutions were not tested for bifonazole and compound 1. For compound 1 this was because the targeted initial drug concentration could not be achieved without adding an excess amount of stock solution and hence organic solvent. For bifonazole, experimental issues were encountered with running the systems at the high pH used for this system. Results and Discussion. (a) Solubility of Drugs in the Presence of STC and SDS. The equilibrium solubility values of drugs in buffer, buffer with 0.1% STC, or buffer with 0.1% SDS are listed in Table 2. It can be seen that all the compounds exhibit low aqueous solubility, with values ranging from 0.2 to 80 μg/mL. STC and SDS solutions both enhance the drug solubility compared to buffer; however, the level of enhancement varies dramatically from drug to drug. In general, SDS
was synthesized at Merck and Company, Rahway, NJ.19,20 The medium used for the induction time measurements for each compound is also listed in Table 1 and was chosen to ensure that the compound was un-ionized. Buffer salts were obtained from either Macron Fine Chemicals (Center Valley, PA) or Mallinckrodt Chemical Inc. (St. Louis, MO), methanol was obtained from Macron Fine Chemicals, and N,N-dimethylformamide was obtained from Fisher Chemicals (Pittsburgh, PA). (b) Methods. 1. Determination of Drug Solubility. The solubility of crystalline drugs in various media with or without 0.1% surfactant was determined by adding an excess amount of the drug to 15 mL of solution. The mixture was stirred and allowed to equilibrate for 48 h in a water bath at 25 °C for all drugs except telaprevir, which was equilibrated at 37 °C. The solution was separated from the excess solid by ultra2594
DOI: 10.1021/acs.cgd.5b00392 Cryst. Growth Des. 2015, 15, 2593−2597
Crystal Growth & Design
Communication
Figure 2. Model compound structures.
Table 2. Equilibrium Solubility of Drugs and the C/Ceq Ratios Used in the Nucleation Induction Time Measurementsa solubility in medium blank (μg/mL) celecoxib griseofulvin felodipine tadalafil telaprevir at 37 °C warfarin naproxen indomethacin bifonazole nevirapine compound 1 a
solubility in 0.1% STC (μg/mL)
solubility in 0.1% SDS (μg/mL)
C/Ceq
1.1 15.0 0.6 1.9 4.6
± ± ± ± ±
0.1 0.4 0.1 0.1 0.1
1.8 17.4 1.1 3.8 11.3
± ± ± ± ±
0.1 0.9 0.1 0.3 0.5
1.6 57.8 21.7 10.3 19.0
± ± ± ± ±
0.1 1.6 2.4 0.1 1.8
5 1.5 8 15 13
3.6 20.3 1.7 0.2 81.3 0.2
± ± ± ± ± ±
0.1 0.1 0.1 0.1 1.3 0.1
4.4 21.8 1.8 0.7 98.2 0.3
± ± ± ± ± ±
0.3 0.5 0.1 0.1 1.8 0.1
24.5 36.1 10.1 7.4 98.9 53.1
± ± ± ± ± ±
0.23 1.1 0.7 0.1 1.0 0.3
5 1.5 10 10 4 10
other compounds showed solubility enhancements varying between a factor of ∼1.5 and 37. The very insoluble compounds (solubility
