Physical stability and dissolution behavior of ketoconazole-organic

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Physical stability and dissolution behavior of ketoconazole-organic acid coamorphous systems Michelle H. Fung, K#rlis B#rzi#š, and Raj Suryanarayanan Mol. Pharmaceutics, Just Accepted Manuscript • DOI: 10.1021/acs.molpharmaceut.8b00035 • Publication Date (Web): 12 Mar 2018 Downloaded from http://pubs.acs.org on March 13, 2018

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Molecular Pharmaceutics

Physical stability and dissolution behavior of ketoconazole-organic acid coamorphous systems Michelle Fung1, Kārlis Bērziņš1,#, Raj Suryanarayanan1* 1

Department of Pharmaceutics, University of Minnesota, Minneapolis, Minnesota

55455, USA #

Latvian Institute of Organic Synthesis, Riga, Latvia

*Corresponding Author: Department of Pharmaceutics, University of Minnesota, Minneapolis, Minnesota 55455, United States. Phone: 612-624-9626. Fax: 612-6262125. E-mail: [email protected]

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For Table of Contents Use Only


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ABSTRACT In an earlier investigation, coamorphous systems of ketoconazole (KTZ) prepared with each oxalic (OXA), tartaric (TAR), citric (CIT) and succinic (SUC) acid, revealed drugacid ionic and/or hydrogen bonding interactions in the solid-state (Fung et al, Mol Pharmaceutics, 2018, 15(x), xxxx-yyyy). We showed that the drug-acid interactions in KTZ-TAR were the strongest, followed by KTZ-OXA, KTZ-CIT and KTZ-SUC. In this study, we investigated the crystallization propensity and dissolution behavior of the KTZ-acid coamorphous systems. When in contact with water (either as water vapor or as aqueous phosphate buffer), while KTZ-CIT and KTZ-TAR were physically stable and resisted crystallization, KTZ-SUC and KTZ-OXA crystallized more readily than KTZ alone. The dissolution performance of the coamorphous systems were compared using the area under the curve (AUC) obtained from the concentration-time profiles.

KTZ-

OXA exhibited the highest AUC, while it was about the same for KTZ-TAR and KTZ-CIT and the lowest for KTZ-SUC. The enhancement in dissolution appears to become more pronounced

as

the

strength

of

the

acid

(OXA>TAR>CIT>SUC)

increased.

Coamorphization with acid caused at least a 2-fold increase in AUC when compared with amorphous KTZ. The decrease in pH of the diffusion layer of the dissolving solid, brought about by the acid, is at least partially responsible for the dissolution enhancement.

In addition, the particles of KTZ-OXA, KTZ-TAR and KTZ-CIT were

much smaller than KTZ-SUC. The consequent effect on surface area could be another contributing factor to the initial dissolution behavior. Keywords:

Amorphous, coamorphous, solid dispersions, dissolution, crystallization,

physical stability



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INTRODUCTION Utilizing the amorphous form of a poorly water soluble active pharmaceutical ingredient (API) has the potential to increase its dissolution rate and consequently lead to oral bioavailability enhancement. Preparation of amorphous solid dispersions (ASD), where an API is molecularly mixed with a hydrophilic polymeric carrier, is a popular strategy for stabilizing amorphous APIs1-4. While polymers have been extensively used to fabricate ASDs, formation of stable dispersions required high polymer concentration, thus limiting their use to low dose APIs. Recently, excipients other than polymers were shown to form stable amorphous phases5-7. One promising stabilization strategy is the preparation of coamorphous drugsmall molecule mixtures. For example, two poorly water-soluble APIs, carbamazepine and

indomethacin

were

stabilized

by

amino

acids6-7.

Indomethacin-arginine

coamorphous system showed an approximately 200-fold increase in intrinsic dissolution rate relative to amorphous indomethacin. In an earlier study, we utilized several organic acids (oxalic, tartaric, citric and succinic acid) to stabilize a weakly basic API, ketoconazole (KTZ), in the amorphous state8. KTZ exhibited ionic and/or hydrogen bonding interactions with each acid. As the strength of KTZ-acid interaction increased, there was an increase in α-relaxation time, a measure of molecular mobility. Moreover, in the two systems with structurally similar dicarboxylic acids, KTZ-succinic and KTZ-oxalic, the slower mobility of the latter was likely responsible for its increased stability.

In addition, the KTZ-tartaric and KTZ-citric

systems were exceptionally stable in the solid-state.

These amorphous systems were

prepared with the ultimate goal of achieving high apparent aqueous solubility. 4 ACS Paragon Plus Environment

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However, for the solubility improvement to be useful (i.e. lead to enhanced bioavailability), the coamorphous systems should resist crystallization in timescales of practical importance. Several methods were used to evaluate the ability of each coamorphous system to resist crystallization when placed in contact with water. Acting as a plasticizer, water lowers the glass transition temperature (Tg) of amorphous materials9, increases molecular mobility and consequently accelerates crystallization. First, the coamorphous systems were allowed to sorb water by storage under different RH conditions, following which the samples were evaluated (Tg, crystallization propensity) by differential scanning calorimetry. As a second step, each coamorphous system was wetted with dissolution medium and crystallization was monitored, in real time, by X-ray diffractometry (XRD) using synchrotron radiation. Thirdly, each coamorphous system was slurried in dissolution medium. Again, by synchrotron XRD, the crystallizing phase was identified and the crystallinity of the solid in contact with the dissolution medium was quantified. The aqueous solubility of KTZ, a weak base, is highly pH dependent10. When the KTZacid undergoes dissolution, the acid in the coamorphous system will influence the diffusion layer pH around each dissolving particle. We hypothesize that as the strength of the acid increases, the dissolution enhancement will be more pronounced - an effect attributed to the magnitude of decrease in diffusion layer pH. The dissolution behavior of the coamorphous systems were compared from the areas under the curve (AUC) obtained from the concentration-time profiles. Finally, since it is not possible to directly determine the diffusion layer pH, the slurry pH was measured. 5 ACS Paragon Plus Environment


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EXPERIMENTAL Materials.

The model drug, ketoconazole (KTZ) was a gift from Laborate

Pharmaceuticals (Haryana, India). Oxalic acid, citric acid anhydrous, D, L-tartaric acid and succinic acid were purchased from Sigma-Aldrich (Missouri, USA). Preparation of amorphous systems. Ketoconazole and each of the following organic acids: oxalic, tartaric, citric and succinic acid in 1:1 molar ratio were spray dried using Buchi mini spray dryer B-290 (Delaware, USA). A methanolic solution (5% w/v total solid content) composed of KTZ and each acid in 1:1 molar ratio was spray dried at a rate of 3.5 ml/min. The inlet and outlet temperatures were maintained at ~55 ºC and ~35 ºC respectively. The spray dried samples were dried under vacuum for 20 hours (RT) to remove residual solvent. Crystalline ketoconazole-acid samples. KTZ-succinic acid cocrystal and KTZ-oxalate salt were prepared by slow evaporation. Methanolic solutions, at 1:1 KTZ to acid molar ratio, were prepared (1% w/v total solid content) from which single crystals were obtained. The product formation was confirmed using X-ray diffractometry (Supporting Information, Figures S3 and S4) Physical mixtures. Samples were prepared by geometric mixing of crystalline KTZ with each acid in 1:1 molar ratio using a motor and pestle. Differential Scanning Calorimetry (DSC).

A differential scanning calorimeter (TA

Instruments Q2000, DE, USA) equipped with a refrigerated cooling unit was used. The instrument was calibrated with indium. The samples were heated in hermetically sealed (Tzero) aluminum pans (TA Instruments, DE, USA) at 10 ºC/min, from 0 to 175-205 ºC


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(depending on sample) under nitrogen purge (50 ml/min). Glass transition temperature (Tg) was the midpoint of the transition and the crystallization onset temperature (Tc) was the onset temperature of the crystallization peak. Water sorption studies. The coamorphous KTZ-acids were placed in open DSC pans and stored at room temperature in chambers containing saturated salt solutions of lithium chloride (~11% RH), potassium carbonate (~43% RH) and sodium chloride (~75% RH). The sample weights were recorded before and after storage for 24 hours. The DSC pans containing the samples were hermetically sealed and characterized using a differential scanning calorimeter (TA Instruments Q2000, DE, USA). Synchrotron X-ray Diffractometry (SXRD).

Preparation of freeze dried samples.

Approximately 50 mg of amorphous KTZ and KTZ-acid coamorphous systems were exposed to 250 µL of dissolution media for a pre-determined time period (15 min, 30 min, 1 h, 1.5 h, 2 h, 3 h, 6 h, 12 h and 24 h). At each time point, samples were frozen in liquid nitrogen and then placed in a bench top freeze dryer (VirTis AdVantage, Gardiner, NY). The shelf of the freeze dryer was pre-cooled to -10 ºC and the samples were dried at -10 ºC, under reduced pressure (65 ± 25 mTorr) for 24 hours. The dried samples were transferred to DSC pans and hermetically sealed in a glove box at controlled RH (RH < 5%) and stored at -20 ºC until analyzed. The DSC pans were mounted on a custom-made holder and exposed to synchrotron radiation. In situ experiments. In the second study, approximately 20 mg of ASDs were weighed in DSC Tzero pans (TA instruments, DE) and 25 µL of phosphate buffer (pH 6.8) was added to uniformly wet the samples. The wetted samples were hermetically sealed and, as before, mounted on the custom-made holder. The samples were analyzed, at 7 ACS Paragon Plus Environment


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approximately 20-minute intervals, for slightly over 3 hours. The specific sampling times are given in the figures. Both sets of experiments were performed in the transmission mode using synchrotron radiation in the 17-BM-B beam line at Argonne National Laboratory (Lemont, IL). A monochromatic X-ray beam [wavelength 0.72768 Å, beam size 250 µm (horizontal) & 160 µm (vertical)] and a two-dimensional area detector (XRD-1621, PerkinElmer) were used. The sample to detector distance was set at 900 mm and the pan was oscillated (±1 mm from the center) along the horizontal axis using a stepper motor. A triplebounce channel-cut Si single crystal monochromator with [111] faces polished was used, which limited the line broadening to its theoretical limit, i.e., the Darwin width. The flux of the incident X-ray was 8 x 1011 photons/sec at 17 keV. The calibration was performed using Al2O3 (NIST, SRM-647a) standard. The two-dimensional (2D) data were integrated to yield one-dimensional (1D) 2θ (º) scans using the GSAS-II software. Comparison with laboratory X-ray data was enabled by converting the scans for the wavelength of Cu Kα radiation (1.5406 Å) using JADE 2010 (Material data, Inc.). Powder dissolution. A USP type II apparatus (Varian 705 DS, Agilent Technologies, Santa Clara, CA) was used. The powder sample was passed through US no. 35 mesh and 50 mg (KTZ equivalent) was dispersed in 500 mL of dissolution medium (pH 6.8; 37 °C) with a paddle speed of 100 rpm. Aliquots were taken at pre-determined time points and filtered with PTFE (0.45 µm) syringe filter, diluted with 0.1 N HCl and analyzed by HPLC with UV detection (Shimadzu, Kyoto, Japan). The dissolution volume was kept constant throughout the experiments. Chromatographic separation was performed at RT using Luna C18 column (250 x4.6mm, 100Å) under isocratic conditions (mobile 8 ACS Paragon Plus Environment

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phase 70:30 0.01 N HCl to acetonitrile) at a flow rate of 1 ml/min. Standard curves were linear over the concentration range of 1 to 100 µg/ml of KTZ (r2 = 0.999; RSD < 6.0%). After the dissolution run, the pH of the medium was determined. The electrode was calibrated at 25 °C using standard buffer solutions (pH 2.00, 4.01 and 10.00). The dissolution profiles were analyzed using Phoenix WinNonlin version 6.4 (Certara USA, Inc., Princeton, NJ). The area under the curve (AUC) was determined by noncompartmental analysis (NCA) using trapezoidal rule integration to the last time point (AUC(0→t)).

The maximum concentration (Cmax) and time to reach the maximum

concentration (Tmax) were also obtained. The extent of dissolution enhancement was quantified from the AUC ratio ([AUC(0→t), sample]/[AUC(0→t),crystalline KTZ]). pH measurements. Amorphous samples containing 5.0 mg equivalent of KTZ were added to HPLC vials containing 1 ml of deionized water. The slurry pH was measured with a micro pH electrode after 24 hours at RT. Scanning Electron Microscopy (SEM). SEM was used to characterize the particle size.

Spray dried powders were sprinkled on aluminum stubs with a double-sided

carbon tape, coated with platinum (20 Å), and imaged in a scanning electron microscope (Jeol 6500 F microscope, Hitachi, Japan). Isothermal X-ray powder diffraction (PXRD). ASDs were evaluated using an X-ray diffractometer (D8 ADVANCE, Bruker AXS, WI, USA) equipped with a variable temperature stage (TTK 450; Anton Paar, Graz-Straβgang, Austria) and a Si strip onedimensional detector (LynxEye). Cu Kα radiation (1.54 Å; 40 kV x 40mA) was used, and



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data were collected in the range of 5-35° 2θ with a step size of 0.02° and a dwell time of 0.5 s.


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RESULTS Thermal analysis after water sorption The KTZ-acid coamorphous systems, in the dry state, were characterized in detail earlier8. When the coamorphous systems were stored under different relative humidity (RH) conditions (~11, 43, and 75% RH; RT), the amount of water sorbed increased as a function of RH (results are presented in Supporting Information, Table S1). The stored samples were assessed by DSC (Figure 1a; Supporting Information Figures S1 and S2) and the results are summarized in Table 1. Following storage at 11% RH, there was only a small decrease in Tg, reflecting minimal plasticization under these RH conditions. However, the plasticization was pronounced when stored at 43 and 75% RH, particularly for the coamorphous systems (Figure 1b). When heated after storage at 43% RH, in the KTZ, KTZ-OXA and KTZ-SUC systems, the glass transition was followed by an exotherm attributable to crystallization and finally a melting endotherm. The melting temperatures (Tm, in Table 1) were in excellent agreement with the reported melting temperatures of KTZ-OXA salt and KTZ-SUC cocrystal11. We therefore concluded that KTZ-OXA and KTZ-SUC had crystallized as a salt and a cocrystal respectively.

The KTZ-CIT and KTZ-TAR systems resisted

crystallization (Figure 1a). Following storage at 75% RH, there was substantial crystallization of the KTZ-SUC system.

This conclusion was based on the absence of both Tg and crystallization

exotherm in the DSC heating curve and the presence of melting endotherm (Supporting Information; Figure S2). In KTZ and KTZ-OXA systems, crystallization was observed



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when the samples were heated in the DSC (Supporting Information; Figure S2). The KTZ-CIT and KTZ-TAR remained amorphous even after heating to 165 ºC.

Figure 1. (a) Representative DSC heating curves of amorphous KTZ and KTZ-acid coamorphous systems after storage at 43% RH for 24 hours at room temperature. The heating curves following storage at 11 and 75% RH are presented in the Supporting Information (Figures S1 and S2). (b) The change in glass transition temperatures, with respect to their dry Tg values [∆ Tg = Tg (after storage) - Tg (dry)], following storage at 11, 43 and 75% RH (mean ± SD; n=3). Table 1. Thermal behavior of the coamorphous systems following storage at 11, 43 and 75% RH (RT) The Tg values at

Physical stability and dissolution behavior of ketoconazole-organic

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