Formation Mechanism of Coamorphous DrugâAmino Acid Mixtures

Article pubs.acs.org/molecularpharmaceutics

Formation Mechanism of Coamorphous Drug−Amino Acid Mixtures Katrine Tarp Jensen,† Flemming Hofmann Larsen,‡ Claus Cornett,† Korbinian Löbmann,† Holger Grohganz,*,† and Thomas Rades† †

Department of Pharmacy and ‡Department of Food Science, University of Copenhagen, Copenhagen, Denmark

ABSTRACT: Two coamorphous drug−amino acid systems, indomethacin−tryptophan (Ind−Trp) and furosemide−tryptophan (Fur−Trp), were analyzed toward their ease of amorphization and mechanism of coamorphization during ball milling. The two mixtures were compared to the corresponding amorphization of the pure drug without amino acid. Powder blends at a 1:1 molar ratio were milled for varying times, and their physicochemical properties were investigated using XRPD, 13C solid state NMR (ssNMR), and DSC. Comilling the drug with the amino acid reduced the milling time required to obtain an amorphous powder from more than 90 min in the case of the pure drugs to 30 min for the coamorphous powders. Amorphization was observed as reductions in XRPD reflections and was additionally quantified based on normalized principal component analysis (PCA) scores of the ssNMR spectra. Furthermore, the evolution in the glass temperature (Tg) of the coamorphous systems over time indicated complete coamorphization after 30 min of milling. Based on the DSC data it was possible to identify the formation mechanism of the two coamorphous systems. The Tg position of the samples suggested that coamorphous Ind−Trp was formed by the amino acid being dissolved in the amorphous drug, whereas coamorphous Fur−Trp was formed by the drug being dissolved in the amorphous amino acid. KEYWORDS: coamorphous, amorphization, furosemide, indomethacin, drug−amino acid (PVP).2 In these systems, a high weight percentage of the polymer is commonly used. Especially with drugs that are given in high doses, this may lead to overall high dosage volumes and lead to problems in the downstream formulation of the glass solution into a tablet or capsule dosage form. Moreover, since many of the polymers are very hygroscopic, this leads to a risk of water uptake, with negative consequences for physical stability, due to the plasticizing nature of water, lowering the glass transition temperature (Tg) of the glass solution. So called coamorphous formulations consisting of two low molecular weight molecules that stabilize each other in the amorphous form are a promising alternative to polymer-based glass solution.3 Coamorphous drug−drug, drug−citric acid, and drug−amino acid systems have previously been shown to improve solubility of poorly water-soluble drugs.3−9 These systems offer an opportunity to stabilize drugs in the

1. INTRODUCTION Amorphous solids lack the long-range orientational and positional order of molecules that characterizes the crystal structure. This leads to changes in most physicochemical properties of a given compound when it is converted from the crystalline state to an amorphous form. For example, amorphous forms of drugs are characterized by improved dissolution rate and solubility compared to their respective crystalline forms. Therefore, the amorphous form represents an obvious advantage in the formulation of poorly water-soluble drugs,1 especially as low solubility of new drug candidates is one of the most pressing problems in formulation and drug development. However, since the amorphous form of a drug is a thermodynamically unstable, high energy solid form, physical instability and recrystallization to a crystalline form upon preparation, storage, and administration is a major obstacle in the formulation of amorphous drugs. A common way of stabilizing an amorphous drug is by formulation of the drug in a solid dispersion, in which the drug is molecularly dispersed in an amorphous polymeric carrier (to form a so-called glass solution) such as polyvinylpyrrolidone © XXXX American Chemical Society

Received: April 16, 2015 Revised: June 8, 2015 Accepted: June 9, 2015

A

DOI: 10.1021/acs.molpharmaceut.5b00295 Mol. Pharmaceutics XXXX, XXX, XXX−XXX

Article

Molecular Pharmaceutics

Figure 1. Chemical structures of the investigated amino acid tryptophan (center) and the drugs indomethacin (left) and furosemide (right).

2. MATERIALS The amino acid L-tryptophan was purchased from SigmaAldrich (St. Louis, MO, USA). The drugs indomethacin and furosemide were acquired from Hawkins Pharmaceutical group (Minnesota, USA) and IPLA Laboratories Limited (Mumbai, India), respectively. The structural formulas are shown in Figure 1. All substances were of reagent grade and used as received.

amorphous form without adding large quantities of polymers, thereby keeping the overall volume of the formulation as low as possible. Molar 1:1 ratios have been shown to be preferable, and heterodimer formation has been suggested in most coamorphous systems.3,7,11,12 However, little is known about the mechanism of formation of coamorphous systems, which to date mostly are prepared by comilling.4,11 Amorphous indomethacin (Ind) is a widely used model compound and has been prepared in coamorphous drug−drug binary systems in combination with ranitidine hydrochloride and naproxen and in coamorphous drug−amino acid binary and ternary mixtures together with L-arginine, L-tryptophan (Trp), and L-phenylalanine.4,6,7 The combination of Ind and Trp is of particular interest as the interactions within this coamorphous system consist of π−π interactions including the aromatic regions of the two molecules and hereby represents an alternative to salt formation.27 In addition, the improved dissolution rate of the drug and a high thermal stability with respect to recrystallization, indicated by a Tg of 68.8 ± 2.6 °C for coamorphous Ind−Trp, suggest that Trp could be an interesting amino acid to investigate further.4 Furosemide (Fur) has not previously been prepared as a coamorphous system, but for example as a solid dispersion.10 Therefore, it is of interest to investigate the potential of Fur in a coamorphous formulation in more detail. In the case of drug−polymer mixtures, conversion of the physical mixture into a glass solution can be regarded as a dissolution process of the drug into the polymer which initially is already amorphous. However, the situation is less clear in the case of coamorphous systems, because both starting materials are crystalline. The aim of this work is thus to gain more knowledge on how coamorphous formulations form during a milling process, and second to assess the performance of different analytical methods for quantifying the amorphous fraction in the drug−amino acid powders upon amorphization. Two different coamorphous drug−amino acid systems were investigated consisting of the amino acid Trp in combination with either Ind or Fur. The powder mixtures were prepared by ball milling of the two components at a molar ratio of 1:1. Chemical degradation during mechanical activation is generally considered low, and chemical degradation during milling of coamorphous Ind−Trp has previously been investigated resulting in a recovery of 99.01 ± 0.28%.4 We hypothesize that by varying the milling time it is possible to vary the degree of structural change and hence the degree of amorphization or the composition of the amorphous fraction, and thus to elucidate the amorphization mechanism of the coamorphous forms. All mixtures were investigated by differential scanning calorimetry (DSC), X-ray powder diffraction (XRPD), and 13C solid state nuclear magnetic resonance (ssNMR) spectroscopy.

3. METHODS 3.1. Preparation of Amorphous Materials. Coamorphous mixtures were prepared by ball milling of 1500 mg of powder consisting of a 1:1 molar ratio of crystalline drug and amino acid. All samples were milled in an oscillatory ball mill (Mixer mill MM400, Retsch GmbH & Co., Hann, Germany) at a frequency of 30 Hz in 25 mL jars containing two stainless steel balls with a diameter of 12 mm. The mill was placed in a cold environment (6 °C) to reduce the risk of recrystallization caused by heating during the mechanical process.12 Seven different mixtures were prepared for each of the 2 coamorphous combinations by using different milling times (0, 5, 15, 30, 45, 60, and 90 min, respectively). Before analysis, the “0 min” samples were mixed for 5 min in the mill but without balls in order to obtain homogeneous mixtures. Amorphous Trp, Ind, and Fur were prepared by ball milling for 6 h (amino acid) and 3 h (drugs), respectively. All samples were analyzed at the day of preparation. 3.2. X-ray Powder Diffraction (XRPD). XRPD measurements were performed using an X’Pert PRO X-ray diffractometer (PANalytical, Almelo, The Netherlands) using Cu Kα radiation (λ = 1.54187 Å). An acceleration voltage of 45 kV and current of 40 mA were used. Samples were scanned in reflection mode between 5° and 35° 2θ with a scan speed of 0.067° 2θ/s and a step size of 0.026°. Data was collected and analyzed using the software X’Pert Data Collector 2.2i (PANalytical B.V., Almelo, The Netherlands). 3.3. Solid State NMR (ssNMR). Solid state NMR spectra were recorded on a Bruker Avance 400 spectrometer operating at Larmor frequencies of 400.13 and 100.61 MHz for 1H and 13 C, respectively. The CP/MAS experiments13 were carried out using a double-tuned CP/MAS probe equipped for 4 mm outer diameter rotors, a spin rate of 12.5 kHz, 304 scans, a sample temperature of 40 °C, a recycle delay of 8 s, a contact time of 4.5 ms, and an acquisition time of 40.9 ms during which highpower 1H TPPM decoupling was applied.14 Subsequently the data were apodized by Lorentzian line broadening of 10 Hz prior to Fourier transformation. All spectra were referenced to an external sample of α-glycine at 176.5 ppm. 3.4. Differential Scanning Calorimetry (DSC). DSC measurements were performed on a Discovery DSC (TA Instruments, New Castle, DE, USA). Approximately 5 mg of sample was analyzed in an aluminum Tzero pan sealed with an B


Article


Figure 2. XRPD diffractograms of γ-indomethacin, furosemide (form I), and tryptophan ball milled for 0, 5, 15, 30, 45, 60, and 90 min, respectively.

Figure 3. XRPD diffractograms of the furosemide (Fur)−tryptophan (Trp) and indomethacin (Ind)−Trp mixtures ball milled for 0, 5, 15, 30, 45, 60, and 90 min, respectively.

which is commonly referred to as the appearance of a “halo” in the diffractogram. Figure 2 presents the XRPD patterns for γ-Ind,17 Fur form 18 I, and Trp19 analyzed after various milling times. Neither of the compounds were completely amorphous within 90 min of milling as crystalline reflections are still visible, in contrast to the fully amorphous halo obtained after individually milling the drugs for 3 h and the amino acid for 6 h. The XRPD reflections in both drug−amino acid mixtures decreased during milling (Figure 3). As expected, the reflections in crystalline Ind−Trp and Fur−Trp mixtures (ball milled for “0 min”) are comparable to a combination of the reflections of crystalline drug and Trp. In stark contrast to the individual drugs and the amino acid, reduction in reflection intensities is already pronounced after milling for 5 min and no Trp reflections are visible in the Fur−Trp sample at this milling time, indicating that in this mixture Trp is amorphous after milling for only 5 min. However, an amorphous halo without any crystalline reflections requires milling of the coamorphous mixtures for 30 min. These data suggest that milling of binary drug−amino acid powders induces and facilitates amorphization compared to the individual drugs as the halo in the XRPD diffractograms of Fur−Trp and Ind−Trp appear after shorter milling times compared to pure Fur, Ind, and Trp, respectively (Figure 2, Figure 3). In comparable amorphous systems which are also prepared by milling, 30 min was required in order to

aluminum Tzero lid. The surface of the sample was smoothed out to ensure an equal sample layer and proper contact between sample and pan. Measurements were performed in modulated temperature mode in the temperature range from −20 to 180 °C with an applied heating rate of 2 K/min, modulation amplitude of 0.2120 °C, and a period of 40 s. A constant nitrogen flow rate of 50 mL/min was applied. Routine calibration of temperature and enthalpy was performed on an indium standard. Cell constant and Tzero calibrations were performed with sapphire. Analysis was performed using Trios software (TA Instruments-Waters LLC, New Castle, DE, USA). The Tg was calculated as the midpoint of onset and end temperature of three independent samples. 3.5. Multivariate Analysis. Multivariate analysis of the ssNMR data was performed by principal components analysis (PCA) on mean-centered spectral data (SIMCA-P, Version 13.0.3, Umetrics AB, Umeå, Sweden). Prior to PCA the area under the curve in each spectrum was normalized to one.15

4. RESULTS AND DISCUSSION 4.1. X-ray Powder Diffraction (XRPD). XRPD is considered one of the most definitive methods for detecting molecular order in a solid system.16 In contrast, disorder, for example if the sample is present in a (partly) amorphous form, is implied indirectly by the lack of order, observed as diffuse scattering and the absence of Bragg reflections in the pattern, C


Article


Figure 4. 13C CP/MAS NMR spectra of indomethacin (Ind), furosemide (Fur), and tryptophan (Trp) ball milled for 0, 5, 15, 30, 45, 60, and 90 min and 3 or 6 h. In addition the PC 1 loadings from a PCA on the Ind (explained variance 88.4%), Fur (explained variance 84.6%), and Trp (explained variance 92.4%) samples are shown.

Figure 5. 13C CP/MAS NMR spectra of indomethacin (Ind)−tryptophan (Trp) and furosemide (Fur)−Trp ball milled for 0, 5, 15, 30, 45, 60, and 90 min. In addition the PC 1 loadings from a PCA on the Ind−Trp (explained variance 92.8%) and Fur−Trp (explained variance 92.3%) samples are presented.

prepare amorphous Ind comilled together with silica.21,22 180 and 60 min of ball milling were required in order to obtain amorphous Ind−ranitidine hydrochloride and amorphous Ind− PVP in a molar ratio of 1:1, respectively.7,21 4.2. Solid State NMR. Solid state NMR is a sensitive technique to assess crystalline and amorphous material.23 Previously the transition from crystalline to amorphous forms of a compound has been assessed by detecting line width and shape changes in ssNMR spectra.24 Asymmetrical broadening of the overall spectrum occurs in the amorphous state due to inhomogeneous distribution of isotropic chemical shifts resulting in a static distribution of bond or torsion angles.25 Hence the NMR spectrum in principle contains information on the crystalline order, amorphous disorder, and molecular interactions. In agreement with this, a continuous line broadening is observed during milling of pure Fur, Ind, and Trp (Figure 4). Generally, subtle differences and correlations between spectral features are often hard to describe by visual evaluation. Mathematically, in a set of spectra, each measured response at a certain input parameter can be seen as a variable in a multivariate matrix. One possible method to reduce the

number of dimensions and give us an understandable overview of the correlations in a system is the application of a projection method such as principal component analysis (PCA).28 PCA was performed on the 13C NMR spectra of the samples milled for varying milling times for Ind, Fur, Trp, Ind−Trp, and Fur− Trp, respectively. Hereby it was possible to detect variations between the NMR spectra upon amorphization. These variations were to a high degree (always above 84%) explained by the first principal component (PC 1). The second principal component usually explained 3−9% of the spectral variation and was omitted from further analysis as the loadings could not be connected to clear spectral features. The PC 1 loadings are presented together with the NMR spectra, and generally the spectral loadings with positive intensity and narrow line widths originate from the crystalline components in each mixture, whereas the spectral loadings with negative intensity and broader lines originate from the amorphous parts of the mixture (Figure 4, Figure 5). In crystalline Fur form I a single carbonyl resonance at 172.4 ppm is observed,20,26 which is converted into two resonances of equal intensities at 172.3 and 169.2 ppm in amorphous Fur, ball D


Article


Figure 6. 13C NMR spectra of amorphous drug, tryptophan (Trp), coamorphous drug−Trp, amorphous drug and Trp combined, and amorphous drug−Trp subtracted from coamorphous drug−Trp. Drug: indomethacine (Ind; left), furosemide (fur; right).

dominated by resonances with both positive and negative intensities as well as different line widths in the aromatic region, a narrow carbonyl resonance, and resonances of very low intensity in the aliphatic region. This difference between the NMR spectra may be explained by π−π interactions between the two extended π-systems in both Ind and Trp that would also position the aliphatic carbons close to the other molecule. In contrast the slightly extended π-system in Fur consisting of the aromatic ring and the carboxylic acid can interact via π−π interactions without bringing the aliphatic part close to other parts of Trp. With regard to other polymorphs of the APIs, the NMR spectrum of coamorphous Ind−Trp contains single resonance signals similar to crystalline γ-Ind. Therefore, the interactions within coamorphous Ind−Trp are expected to be more similar to the interactions within γ-Ind than α-Ind, as the latter NMR spectrum is characterized by signals with splitting.17 Crystalline Fur form II is characterized by splitting of the carbonyl carbon resonance in comparison to form I.20 However, in coamorphous Fur−Trp this peak is overlapping with Trp resonances. Additional line broadening upon milling makes it difficult to observe if the interactions within coamorphous Fur−Trp are similar to the interactions in the unit cell of Fur form I or II. In order to assess the amorphous content, the evolution in scores from the five PCAs on the ssNMR spectra was evaluated. Usually the amorphous content in a sample is calculated from a standard curve based on samples with known crystallinity.16 However, physical mixing of amorphous and crystalline powder for a standard curve possibly influences the crystallinity of the samples, thereby potentially introducing an error.29 Instead, the crystallinity of the sample was calculated on the basis of a fully amorphous and fully crystalline sample. The crystalline drug− crystalline amino acid samples should be exposed to minimal mechanical energy, however the mixture should be homogeneous as the sample size for analysis is small and should be representative for the overall mixture.30 Mixing for 5 min in the ball mill but without balls was chosen as the preferred preparation method in order to ensure sample homogeneity. The amorphous reference for the individual drugs and the amino acid were prepared by milling for 3 and 6 h, respectively, whereas the coamorphous powders where considered completely amorphous after 90 min of milling. A graph representing the normalized PC 1 scores from a PCA of the ssNMR data as a function of milling time is shown

milled for 3 h (Figure 4). A lower frequency chemical shift change of the carbonyl group in Fur has previously been assigned to hydrogen-bond interactions involving this group.26 The integrals of the two resonances indicate that around 50% of the molecules in amorphous Fur are involved in hydrogen bonds including the carbonyl group. Pairwise hydrogen bonding of drug molecules upon amorphization has previously been reported.6 In the case of Fur it is likely that the hydrogen bond occurs between the carbonyl groups and the sulfonamide moiety of the molecules. The Ind carbonyl group is characterized by a single resonance at 179.1 ppm in amorphous as well as crystalline γ-Ind.17 Apart from line broadening no significant chemical shift changes were observed upon amorphization. Previous analysis of Ind in the stable γ-form, the metastable α-form, and the amorphous form have led to the conclusion that amorphous Ind consists of cyclic homodimers interacting via carboxylic acid hydrogen bonds, similar to the interactions in the γ-form.2 As a result, no additional carbonyl resonance is expected in the NMR spectrum of amorphous Ind. Amorphization of the drug−Trp systems (Figure 5) is observed as line broadening in the ssNMR spectra, similar to the single component powders. No interactions could be detected between the drug and the amino acid in coamorphous Ind−Trp when analyzed by Fourier-transform infrared spectroscopy (FTIR) in a previous study.27 In order to test this hypothesis, the NMR spectra resulting from the subtraction of the NMR spectrum of pure amorphous drug and amorphous Trp from coamorphous drug−Trp were carefully analyzed (Figure 6). If the coamorphous spectrum was identical to the combination of the spectra of the individual amorphous components, the subtraction of the spectra would result in a baseline, whereas resonances in the spectrum after subtraction indicate additional interactions between the two components in the coamorphous mixture. When evaluating the difference spectra, the resonances with positive intensity originate from the carbon sites which are more abundant in the coamorphous mixture than in the corresponding binary mixture. The opposite is true for the resonances with negative intensity. Comparing the difference spectra associated with the two drugs, it is noticed that the spectrum relating to Ind−Trp is mainly characterized by broad resonances of positive intensity that include both aliphatic and aromatic as well as carbonyl resonances, whereas the spectrum associated with Fur−Trp is E


Article

Molecular Pharmaceutics in Figure 7. The normalization was performed by dividing all PCA scores by the score of the 100% amorphous sample after

Figure 8. DSC heating curves (reversing and total heat flow) for coamorphous indomethacin (Ind)−tryptophan (Trp) and amorphous Ind. The glass transition temperature is indicated by the arrows. Figure 7. PC 1 scores (explained variation >84.6%) of ssNMR spectra of indomethacin (Ind), furosemide (Fur), tryptophan (Trp), Ind−Trp, and Fur−Trp. After PCA, the scores were normalized according to 100% and 0% amorphous standards and plotted against milling time. PCA was performed on ssNMR data normalized according to AUC equal to one and preprocessed by mean centering. The data point of Ind−Trp is overlapping with that of Fur−Trp at 90 min.

subtracting the score of the crystalline sample. From the data it is again observed that the rate of amorphization of Trp is faster than for the two drugs when ball milled individually (Figure 7), whereas the amorphization process was even faster for the two coamorphous systems. 4.3. Thermal Analysis (DSC). DSC analysis is commonly used to directly analyze the amorphous content in solids. The glass transition temperature (Tg) reflects the amorphous fraction in an amorphous crystalline mixture (via the magnitude of the heat capacity change at the Tg) in contrast to indirect estimation of the amorphous fraction based on reduced crystallinity (peak intensities) detected by the XRPD. In the modulated DSC scans, the Tg is observed in the reversing heat flow and is differentiated from possible water evaporation, enthalpic relaxation, and recrystallization, which are all described by the nonreversing heat flow. The heating curves of the pure amorphous drugs are compared to the thermograms of the corresponding coamorphous system in Figure 8 and Figure 9. As expected, the Tg is clearly observed in the reversing heat flow signals for coamorphous Ind−Trp and Fur−Trp and amorphous Ind and Fur, as indicated by the arrows in Figures 8 and 9. The single Tg in the coamorphous systems indicates that the two components are forming a single phase coamorphous blend, although it has to be considered that a certain domain size of approximately 30 nm is required for detection.31,32 In addition the deviation in solubility parameters (Δδ) is less than 7.5 for each drug−amino acid system, calculated as group contributions for Ind, Fur, and Trp,33,34 thereby supporting that the drugs and the amino acid are completely miscible, thus resulting in one amorphous phase.35 Trp acts as an antiplasticizer in both mixtures as the Tgs of the coamorphous systems are higher than the Tg of Ind or Fur alone. In the total heat flow curve of coamorphous Ind−Trp two overlapping exothermic events representing the recrystallization

Figure 9. DSC heating curves (reversing and total heat flow) for coamorphous furosemide (Fur)−tryptophan (Trp) and amorphous Fur. The glass transition temperature is indicated by the arrows.

of Ind and Trp are followed by a large endothermic melting peak (Figure 8). The melting peak only represents the drug as the amino acid does not melt in this region. Furthermore, the recrystallization peaks observed in coamorphous Ind−Trp are at higher temperatures than the recrystallization peak of pure amorphous Ind, indicating that the coamorphous Ind−Trp mixtures are thermally more stable with respect to recrystallization. In Figure 9, no melting peak is present within the investigated range. Similar to the Ind−Trp mixture, the recrystallization peak is shifted to higher temperatures in coamorphous Fur−Trp mixtures compared to amorphous Fur alone, indicating a thermally more stable system. Determination of the heat capacity change (ΔCp) at the Tg has previously been shown to be a suitable method for calculating the amorphous content in a drug powder.36 Figure 10 shows amorphization of Ind, Fur, Ind−Trp, and Fur−Trp based on ΔCp as a function of milling time. These data suggest that neither of the two drugs is completely amorphous upon F


Article


milling for 90 min, which correlates well with the observations from the corresponding XRPD diffractograms and the normalized NMR PCA scores (Figure 2, Figure 7). Calculation of crystallinity based on heat capacity change further appears to be suitable for a very rough estimate of the amorphous content during the formation of coamorphous Ind−Trp. However, large variation between the individual data points did occur for the Fur−Trp samples, resulting in several values exceeding 100% amorphization, for example after 10 min of ball milling. These data points appear as potential outliers and indicate that an alternative method to quantify the degree of amorphization is preferable (i.e., lead to more stable values) for this system. The Tg position for the single component samples remains constant for all time points in the experiment, as outlined above. However, the Tg position of the coamorphous systems changes until a milling time of 30 min, after which it remains constant (Figure 11). The data suggests that coamorphous Ind−Trp is formed by the amino acid being dissolved in the amorphous drug as the Tg increases upon milling from approximately the Tg of the pure Ind. This suggests that Trp

is antiplasticizing the amorphous drug as the milling time increases until the powder mixture is completely coamorphous (i.e., milling time exceeds 30 min). The formation of coamorphous Fur−Trp also seems to be completed after milling for 30 min. However, the Tg position decreases upon milling, which indicates that Trp becomes amorphous first and the drug is dissolved in the amino acid upon milling. Hence during the formation of the coamorphous drug−amino acid systems the amino acid is dissolved in the amorphous drug (Ind−Trp) in one case and the drug is dissolved in the amorphous amino acid (Fur−Trp) in the other. It can be seen in Figure 2 that the amorphization kinetics for the single compounds follow the order IND > TRP > FUR. This indicates that the faster amorphizing single component might serve as solvent for the other component in both systems. As the current study was based on the most stable polymorphs of the APIs, it cannot finally be concluded if this mechanism also would apply when less stable polymorphs were used as starting material. It can, however, be hypothesized that less stable polymorphs would turn amorphous faster. In the case of Ind, this would have no influence on the mechanism, while the mechanism for Fur might be inversed. It is possible to calculate the coamorphous fraction in the coamorphous samples based on the Tg as, from the evolution of the Tg position, it is known what component is dissolved in the other. The Tg of the component which initially becomes amorphous is used as the 0% coamorphous standard and the Tg of the completely coamorphous mixture as 100% coamorphous standard, i.e., the Tg of amorphous Ind is used as 0% amorphous standard for the Ind−Trp samples, and the Tg of amorphous Trp is used as the 0% amorphous standard for the Fur−Trp samples. The coamorphous fractions in the drug−amino acid samples based on normalized Tgs are presented in Figure 12. According to these data the formation of coamorphous Fur−Trp is slightly faster than the formation of coamorphous Ind−Trp. Interestingly, no outliers are observed among the Fur−Trp samples even though these data are based on the same heating curves as the data presented in Figure 10.

Figure 11. Evolution of the glass transition temperature (Tg) of indomethacin−tryptophan and furosemide−tryptophan ball milled for 3, 5, 7, 10, 15, 30, 45, 60, and 90 min, respectively. The Tgs of fully amorphous references for furosemide, indomethacin and tryptophan are shown on the y-axis.

Figure 12. Coamorphization of furosemide (Fur)−tryptophan (Trp) and indomethacin (Ind)−Trp samples ball milled for 3, 5, 7, 10, 15, 30, 45, 60, and 90 min, as well as additional samples milled for 3, 7, and 10 min. The glass transition temperature (Tg) of the comilled mixtures is normalized in relation to the Tg of the coamorphous mixture as well as the Tg for amorphous Trp and Ind, respectively. Nonlinear curve fit (one phase decay) is applied to each data set.

Figure 10. Amorphization upon milling of furosemide (Fur) and indomethacin (Ind) with and without tryptophan (Trp), calculated from the heat capacity change at the glass transition temperature and normalized in relation to the values of the purely amorphous powders equal to 100%.

G


Article


(9) Hoppu, P.; Jouppila, K.; Rantanen, J.; Schantz, S.; Juppo, A. M. Characterisation of blends of paracetamol and citric acid. J. Pharm. Pharmacol. 2007, 59, 373−381. (10) Nielsen, L.; Gordon, S.; Holm, R.; Selen, A.; Rades, T.; Müllertz, A. Preparation of an amorphous sodium furosemide salt improves solubility and dissolution rate and leads to a faster Tmax after oral dosing to rats. Eur. J. Pharm. Biopharm. 2013, 85 (3), 942−951. (11) Löbmann, K.; Laitinen, R.; Grohganz, H.; Gordon, K. C.; Strachan, C.; Rades, T. Coamorphous drug systems: enhanced physical stability and dissolution rate of indomethacin and naproxen. Mol. Pharmaceutics 2011, 8 (5), 1919−1928. (12) Allesø, M.; Chieng, N.; Rehder, S.; Rantanen, J.; Rades, T.; Aaltonen, J. Enhanced dissolution rate and synchronized release of drugs in binary systems through formulation: Amorphous naproxen− cimetidine mixtures prepared by mechanical activation. J. Controlled Release 2009, 136 (1), 45−53. (13) Peersen, O. B.; Wu, X.; Kustanovich, I.; Smith, S. O. Variableamplitude cross-polarization MAS NMR. J. Magn. Reson., Ser. A 1993, 104 (3), 334−339. (14) Bennett, A. E.; Rienstra, C. M.; Auger, M.; Lakshmi, K.; Griffin, R. G. Heteronuclear decoupling in rotating solids. J. Chem. Phys. 1995, 103 (16), 6951−6958. (15) Winning, H.; Larsen, F. H.; Bro, R.; Engelsen, S. B. Quantitative analysis of NMR spectra with chemometrics. J. Magn. Reson. 2008, 190 (1), 26−32. (16) Shah, B.; Kakumanu, V. K.; Bansal, A. K. Analytical techniques for quantification of amorphous/crystalline phases in pharmaceutical solids. J. Pharm. Sci. 2006, 95 (8), 1641−1665. (17) Masuda, K.; Tabata, S.; Kono, H.; Sakata, Y.; Hayase, T.; Yonemochi, E.; Terada, K. Solid-state 13 C NMR study of indomethacin polymorphism. Int. J. Pharm. 2006, 318, 146−153. (18) Matsuda, Y.; Tatsumi, E. Physicochemical characterization of furosemide modifications. Int. J. Pharm. 1990, 60, 11−26. (19) Görbitz, C. H.; Törnroos, K. W.; Graeme, M. D. Single-crystal investigation of L-tryptophan with Z′=16. Acta Crystallogr., B 2012, 68, 549−557. (20) Doherty, C.; York, P. Furosemide crystal forms; solid state and physicochemical analyses. Int. J. Pharm. 1988, 47, 141−155. (21) Watanabe, T.; Hasegawa, S.; Wakiyama, N.; Kusai, A.; Senna, M. Comparison between polyvinylpyrrolidone and silica nanoparticles as carriers for indomethacin in a solid state dispersion. Int. J. Pharm. 2003, 250 (1), 283−286. (22) Watanabe, T.; Wakiyama, N.; Usui, F.; Ikeda, M.; Isobe, T.; Senna, M. Stability of amorphous indomethacin compounded with silica. Int. J. Pharm. 2001, 226 (1), 81−91. (23) Gustafsson, C.; Lennholm, H.; Iversen, T.; Nyström, C. Comparison of solid-state NMR and isothermal microcalorimetry in the assessment of the amorphous component of lactose. Int. J. Pharm. 1998, 174 (1), 243−252. (24) Lefort, R.; De Gusseme, A.; Willart, J. F.; Danede, F.; Descamps, M. Solid state NMR and DSC methods for quantifying the amorphous content in solid dosage forms: an application to ball-milling of trehalose. Int. J. Pharm. 2004, 280 (1), 209−219. (25) Berendt, R. T.; Sperger, D. M.; Munson, E. J.; Isbester, P. K. Solid-state NMR spectroscopy in pharmaceutical research and analysis. Trends Anal. Chem. 2006, 25 (10), 977−984. (26) Goud, N. R.; Gangavaram, S.; Suresh, K.; Pal, S.; Manjunatha, S. G.; Nambiar, S.; et al. Novel furosemide cocrystals and selection of high solubility drug forms. J. Pharm. Sci. 2012, 101 (2), 664−680. (27) Löbmann, K.; Laitinen, R.; Strachan, C.; Rades, T.; Grohganz, H. Amino acids as co-amorphous stabilizers for poorly water-soluble drugs - Part 2: Molecular interactions. Eur. J. Pharm. Biopharm 2013, 85, 882−888. (28) Davis, T.; Fearn, T. Back to basics: The principle of principle component analysis. Spectrosc. Eur. 2004, 13, 20−23. (29) Phillips, E. M. An approach to estimate the amorphous content of pharmaceutical powders using calorimetry with no calibration standards. Int. J. Pharm. 1997, 149 (2), 267−271.

5. CONCLUSION Comilling the drugs Ind and Fur together with the amino acid Trp reduced the milling time required to obtain a fully amorphous powder compared to the pure drugs, as detected by XRPD, DSC, and 13C ssNMR. Analysis of 13C ssNMR spectra suggested hydrogen bonds and π-interactions in coamorphous Ind−Trp and Fur−Trp involving the carbonyl of the drug and the aromatic parts of Ind, Fur, and Trp, respectively. Quantification of the amorphous content in the coamorphous mixtures based on the Tg indicated that amorphization was complete after 30 min, which was confirmed by a halo observed in the XRPD diffractogram and by amorphization based on normalized NMR PCA scores. These results suggest that normalization of the Tgs and NMR PCA scores is a suitable method for the quantification of amorphization of coamorphous powders in these systems. The heat capacity change can be used as a method to quantify amorphization of amorphous and coamorphous powders as previously suggested, although this method appeared much less robust. Based on the DSC data it was further possible to identify the formation mechanism of the two coamorphous systems. The Tg position of the samples suggested that coamorphous Ind−Trp was formed by the amino acid being dissolved in the amorphous drug, whereas coamorphous Fur−Trp was formed by the drug being dissolved in the amorphous amino acid.

■

AUTHOR INFORMATION

Corresponding Author

*Department of Pharmacy, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark. Tel: +45 35336473. E-mail: [email protected]. Notes

The authors declare no competing financial interest.

■

REFERENCES

(1) Hancock, B. C.; Zografi, G. Characteristics and significance of the amorphous state in pharmaceutical systems. J. Pharm. Sci. 1997, 86 (1), 1−12. (2) Taylor, L. S.; Zografi, G. Spectroscopic characterization of interactions between PVP and indomethacin in amorphous molecular dispersions. Pharm. Res. 1997, 14 (12), 1691−1698. (3) Löbmann, K.; Strachan, C.; Grohganz, H.; Rades, T.; Korhonen, O.; Laitinen, R. Co-amorphous simvastatin and glipizide combinations show improved physical stability without evidence of intermolecular interactions. Eur. J. Pharm. Biopharm. 2012, 81 (1), 159−169. (4) Löbmann, K.; Grohganz, H.; Laitinen, R.; Strachan, C.; Rades, T. Amino acids as co-amorphous stabilizers for poorly water soluble drugs − Part 1: Preparation, stability and dissolution enhancement. Eur. J. Pharm. Biopharm. 2013, 85, 87−881. (5) Jensen, K. T.; Löbmann, K.; Rades, T.; Grohganz, H. Improving co-amorphous drug formulations by the addition of the highly water soluble amino acid, proline. Pharmaceutics 2014, 6 (3), 416−435. (6) Löbmann, K.; Laitinen, R.; Grohganz, H.; Strachan, C.; Rades, T.; Gordon, K. C. A theoretical and spectroscopic study of co-amorphous naproxen and indomethacin. Int. J. Pharm. 2013, 453, 80−87. (7) Chieng, N.; Aaltonen, J.; Saville, D.; Rades, T. Physical characterization and stability of amorphous indomethacin and ranitidine hydrochloride binary systems prepared by mechanical activation. Eur. J. Pharm. Biopharm. 2009, 71 (1), 47−54. (8) Patel, J.; Carlton, R.; Yuniatine, F.; Needham, T.; Wu, L.; Vogt, F. Preparation and Structural Characterization of Amorphous SprayDried Dispersions of Tenoxicam with Enhanced Dissolution. J. Pharm. Sci. 2012, 101, 641−663. H


Article

Molecular Pharmaceutics (30) Heinz, A.; Savolainen, M.; Rades, T.; Strachan, C. J. Quantifying ternary mixtures of different solid-state forms of indomethacin by Raman and near-infrared spectroscopy. Eur. J. Pharm. Sci. 2007, 32 (3), 182−192. (31) Qian, F.; Huang, J.; Zhu, Q.; Haddadin, R.; Gawel, J.; Garmise, R.; Hussain, M. Is a distinctive single Tg a reliable indicator for the homogeneity of amorphous solid dispersion? Int. J. Pharm. 2010, 395 (1−2), 232−235. (32) Paudel, A.; Geppi, M.; Van den Mooter, G. Structural and dynamic properties of amorphous solid dispersions: the role of solid state nuclear magnetic resonance spectroscopy and relaxometry. J. Pharm. Sci. 2014, 103, 2635−2662. (33) Breitkreutz, J. Prediction of Intestinal drug absorption properties by three-dimensional solubility parameters. Pharm. Res. 1998, 15 (9), 1370−1375. (34) Jouybam-Gharamaleki, A.; Dastmalchi, S.; Chan, H.-K.; Barzegar-Jalali, M. Solubility prediction for furosemide in watercosolvent mixtures using the minimum number of experiments. Drug Dev. Ind. Pharm. 2001, 27 (6), 577−583. (35) Greenhalgh, D. J.; Williams, A. C.; Timmins, P.; York, P. Solubility parameters as predictors of miscibility in solid dispersions. J. Pharm. Sci. 1999, 88 (11), 1182−1190. (36) Guinot, S.; Leveiller, F. The use of MTDSC to assess the amorphous phase content of a micronised drug substance. Int. J. Pharm. 1999, 192 (1), 63−75.

I


Formation Mechanism of Coamorphous DrugâAmino Acid Mixtures

Recommend Documents

Formation Mechanism of Coamorphous DrugâAmino Acid Mixtures