New Approach to Reduce the Overhigh Plasma Concentration of

Article pubs.acs.org/crystal

New Approach to Reduce the Overhigh Plasma Concentration of Captopril by the Formation of Zinc Coordination Polymer Jia Yao,† Yong-Hui Mo,‡ Jia-Mei Chen,*,‡ and Tong-Bu Lu*,†,‡ †

MOE Key Laboratory of Bioinorganic and Synthetic Chemistry/School of Chemistry and Chemical Engineering, Sun Yat-Sen University, Guangzhou 510275, China ‡ School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou 510006, China S Supporting Information *

ABSTRACT: Captopril is an antihypertension drug, it has side effects of vertigo, headache, and functional gastrointestinal disorders due to its overhigh plasma concentration. In addition, captopril undergoes oxidation at its thiol to yield captopril disulfide. In order to reduce its release rate and increase its oxidation stability, coordination polymer of captopril with zinc ion was prepared and its structure was determined by single crystal X-ray diffraction. The results of solubility measurements indicate that the apparent solubility and intrinsic dissolution rate of captopril can be dramatically decreased after the formation of coordination polymer. Pharmacokinetic studies in rats reveal that the plasma concentration of captopril can be reduced by the formation of zinc coordination polymer, with maximum plasma concentration reduced from 2.53 μg/mL for captopril to 0.99 μg/mL for the zinc coordination polymer. Moreover, the oxidation stability of captopril is simultaneously improved by the formation of zinc coordination polymer.

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the release rate of captopril have been reported.5−8 We consider that the overhigh maximum plasma concentration of Cap could be reduced simply by the formation of coordination polymers of metal ions with Cap, as Cap contains three potential donor groups (thiol, carboxyl, and amide) which might act as a linker to form coordination polymers with metal ions. In contrast to the formation of salts, which are usually considered as an approach to increase the solubility of a drug, as the drug becomes cation or anion after the formation of a salt, the solubility of the drug can usually be decreased after the formation of coordination polymers, as drug molecules are difficult to escape from the frameworks of coordination polymers. Moreover, the oxidation at the thiol group of Cap may simultaneously be prohibited by the formation of coordination polymers, as the thiol group of Cap can be coordinated to metal ions. To our knowledge, reducing the overhigh maximum plasma concentration of a drug by the formation of coordination polymers has not been reported so far. In the present work, zinc ion was chosen to interact with Cap, as it is an essential element and plays important roles in many biological processes of human body, and zinc salts, such as zinc sulfate, zinc acetate, and zinc gluconate are included in the list of “generally recognized as safe” (GRAS). The reaction of Zn(II) with Cap gave a coordination polymer of captopril-Zn (Cap-Zn) rather than a Cap-Zinc salt. The difference between Cap-Zinc salt

INTRODUCTION Captopril (Cap, Scheme 1), an angiotensin-converting-enzyme inhibitor, is used for treatment of hypertension and heart failure. Scheme 1. Structure of Captopril (Cap) and Captopril Disulfide (Di-Cap)

After an oral administration of 50 mg, the plasma concentration of Cap could reach the maximum of 600 μg/L.1,2 Compared to its therapeutic concentration (50 μg/L), the much higher Cmax of Cap may result in side effects of vertigo, headache, and functional gastrointestinal disorders.3 In addition, Cap undergoes oxidation at its thiol group to yield captopril disulfide (Di-Cap, Scheme 1) in aqueous solution.4 Approaches to utilize cyclodextrin, layered double hydroxides, and polymer-mesoporous silica to slow down © 2014 American Chemical Society

Received: March 1, 2014 Revised: March 25, 2014 Published: April 24, 2014 2599

dx.doi.org/10.1021/cg500302h | Cryst. Growth Des. 2014, 14, 2599−2604

Crystal Growth & Design

Article

(i) A mixture of captopril (217 mg, 1 mmol) and ZnO (41 mg, 0.5 mmol) was added to 3 mL of methanol and allowed to stir overnight at room temperature. The suspension was filtered and the isolated solid was washed with methanol and dried under vacuum. Yield: 128.0 mg, 91.4% based on ZnO. Anal. Calcd for ZnC9H13NO3S: C, 38.52; N, 4.99; H, 4.67%. Found: C, 38.97; N, 5.05; H, 4.83%. (ii) To 2 mL of aqueous ZnCl2 (136 mg, 1 mmol) was added a solution of water (3 mL) containing captopril (87 mg, 0.4 mmol) and NaOH (12 mg, 0.4 mmol). The resulting solution was stirred overnight and filtered. The isolated solid was washed with methanol and dried under vacuum. Yield: 101.2 mg, 90.3% based on captopril. Single crystal of Cap-Zn was obtained by layering a methanol solution of captopril and NaOH with an aqueous solution of ZnCl2. Single Crystal X-ray Diffraction. Diffraction data for Cap-Zn were collected using an Agilent Technologies Gemini A Ultra system. Data reduction and cell refinement were performed with the program of CrysAlis PRO.9 The structure was solved by the direct method using the SHELXS-97 programs10 and refined by the full-matrix least-squares method on F2. All non-hydrogen atoms were refined with anisotropic displacement parameters. All hydrogen atoms were placed in calculated positions with fixed isotropic thermal parameters and included in the structure factor calculations in the final stage of full-matrix least-squares refinement (Table 1). Stability. Stability evaluation was carried out by slurry method. In these experiments, excess solids were stirred in deionized water for 72 h at 40 °C. Then the solvent was removed under reduced pressure, and the solids were dissolved in the mixture of 0.1 M HCl aqueous solution and methanol (3:7) and analyzed by HPLC to detect the residual of captopril. HPLC analysis was performed on a Shimadzu 20A HPLC system, which was equipped with Inertsil ODS-3 (150 mm × 4.6 mm I.D., 5 μm) column. The mobile phase consisted of a mixture of 0.05% H3PO4 aqueous solution and acetonitrile (73:27, v/v) at a flow rate of 1.0 mL/min. The column was kept at room temperature, and the effluents were monitored at 220 nm. Powder Dissolution Experiments. All the solids were milled to powders and sieved using standard mesh sieves to provide samples with the particle size ranges of 75−150 μm. In a typical experiment, a flask containing 300−1500 mg of powder was added to 5 mL of Tris-HCl buffer or 0.1 M HCl aqueous solution, and the resulting suspension was stirred at 37 °C and 200 rpm. At each time interval, an aliquot of the slurry was withdrawn from the flask, followed by adding an equal volume of fresh medium to the suspension and filtered through a 0.22 μm nylon filter. After appropriate dilutions, the solutions were injected into the HPLC system for analysis. Concentrations of Cap and Cap-Zn in Tris-HCl buffer (pH = 6.8) and 0.1 M HCl aqueous solution (pH = 1) were determined by HPLC. HPLC analysis was performed on a Shimadzu 20A HPLC system, which was equipped with a Inertsil ODS-3 (150 mm × 4.6 mm I.D., 5 μm) column. The mobile phase consisted of a mixture of 0.01 M NaH2PO4 aqueous solution (pH 3; adjusted with phosphoric acid) and methanol (65:35, v/v), at a flow rate of 1.0 mL/min. The column was kept at room temperature, and the effluents were monitored at 220 nm. Intrinsic Dissolution Rate. The intrinsic dissolution rate (IDR) experiments of solid materials were carried out on a ZQY-2 Dissolution Tester (Shanghai Huanghai Yaojian instrument distribution Co., Ltd.). One hundred and fifty milligrams of each solid was compressed in a hydraulic press at 1 ton for 5 s in a die of a 5 mm diameter disk. The disk was coated using paraffin wax, leaving only the surface under investigation free for dissolution. Then, the disk was dipped into 150 mL of 0.1 M HCl aqueous solution at 37 °C, with the paddle rotating at 100 rpm. At each time interval, 2 mL of the dissolution medium was withdrawn and replaced by an equal volume of fresh medium to maintain a constant volume. After being filtered through 0.22 μm nylon filters, solutions were injected into the HPLC system for analysis. Pharmacokinetics in Rat Plasma. Male Sprague−Dawley rats weighing 200 ± 20 g for in vivo pharmacokinetics study were purchased from the animal research center of Sun Yat-sen University (Guangzhou, China) and housed in a temperature- and humidity-controlled room with a 12 h light/dark cycle with free access to food and water. All the animal experiments were carried out in accordance with institutional

and Cap-Zn coordination polymer is that only the carboxylic group of Cap is deprotonated in Cap-Zinc salt, while both carboxylic and thiol groups of Cap are deprotonated in Cap-Zn coordination polymer, in which Cap−2 anions alternately connect Zn(II) to form a neutral coordination polymer. Herein, we reported the structure, stability, solubility, and pharmacokinetics of Cap-Zn coordination polymer.

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EXPERIMENTAL SECTION

Materials and General Methods. Captopril was purchased from Suizhou Hongqi Chemical Co., Ltd. All of the other chemicals and

Table 1. Crystallographic Data and Refinements for Cap-Zn Cap-Zn ZnC9H13NO3S 280.63 tetragonal P41 9.5790(12) 9.5790(12) 12.110(2) 90 90 90 1111.2(3) 4 1.677 576 0.08 × 0.05 × 0.02 −8 ≤ h ≤ 11 −10 ≤ k ≤ 9 −14 ≤ l ≤ 12 0.0742 1.018 0.0613 0.1589 0.04(7)

formula formula weight crystal system space group a (Å) b (Å) c (Å) α (deg) β (deg) γ (deg) V (Å3) Z Dc (g·cm−3) F(000) crystal size (mm) range of indices

Rint GOF R1[ I > 2σ(I)]α wR2 [all data]α Flack parameter

α R1 = Σ∥Fo| − |Fc∥/Σ|Fo|. wR2 = [Σ[w(Fo2 − Fc2)2]/Σw(Fo2)2]1/2, and w = 1/[σ2(Fo)2 + (aP)2 + bP], where P = [(Fo2) + 2Fc2]/3.

Table 2. Mean Pharmacokinetic Parameters for Cap and CapZn in Male Rat (n = 5)a

a

parameter

Cap

Cap-Zn

tmax (min) Cmax (μg/mL) AUC0‑∞ (μg·min/mL) MRT0‑∞ (min)

10 2.53 ± 0.41 58.74 ± 5.54 31.56 ± 6.10

23 ± 2.74b 0.99 ± 0.15b 42.03 ± 11.33b 43.64 ± 5.73b

Each value represents the mean ± SD (n = 5). bP < 0.05.

solvents were commercially available and used as received. Elemental analyses (EA) were carried out by Elementar Vario EL elemental analyzer. The infrared spectra (IR) were recorded in the 4000 to 400 cm−1 region using KBr pellets and a Bruker EQUINOX 55 spectrometer. Thermogravimetric analyses (TGA) were recorded on a Netzsch TG-209 instrument and platinum crucible in nitrogen atmosphere, with a heating rate of 10 °C/min. Differential scanning calorimetry (DSC) was recorded on a Netzsch STA 409PC instrument and aluminum sample pans in nitrogen atmosphere, with a heating rate of 10 °C/min. X-ray powder diffraction (XRPD) patterns were obtained on a Bruker D2 PHASER with Cu Kα radiation (30 kV, 10 mA) Cap-Zn Coordination Polymer. This compound was prepared via the following two methods: 2600



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Figure 1. (a) Coordination environments of Zn and coordination mode of Cap. (b) Side view of 1D right-handed helical chain along the c axis. (c) Coordination bonds between the adjacent helical chains. (d) Three-dimensional structure of Cap-Zn (view from the c axis). guidelines in compliance with regulations formulated by Sun Yat-sen University. The experimental protocol was approved by the Institutional Animal Care and Use Committee of Sun Yat-sen University. Pharmacokinetics was evaluated in five male Sprague−Dawley rats (200 ± 20 g) per formulation, which were fasted for 12 h before drug administration. The solid of Cap or Cap-Zn was added to 0.5% CMCNa aqueous solution (pH = 6.9) and then was orally administered at 20 mg/kg (the equivalent amount of free captopril) to rats, respectively. Blood samples were extracted from retro-orbital sinus using a Pasteur pipet, at the following time intervals: 0, 5, 10, 15, 20, 25, 30, 40, 50, 60, 90, 120, and 180 min after oral dosing, respectively. The plasma samples were separated by centrifugation (5 min, 6000 rpm). The total amount of captopril in plasma was determined by treating the plasma with pbromophenacyl bromide (p-BPB) to reduce the oxidized form of captopril (Di-Cap) and used p-BPB as a chromophore to detect captopril.11−13 To 200 μL of plasma was added 20 μL of 1 mg/mL p-BPB acetonitrile solution. The mixture was vortex-mixed for 5 min and allowed to stand in dark for 1 h. Then, to the mixture was added 80 μL of 1 M HCl aqueous solution, and it extracted with 800 μL of benzene-ethyl acetate (1:1, v/v). The organic layer was evaporated to dryness under vacuum and reconstituted in 50 μL of acetonitrile. A portion of 20 μL was injected into the HPLC.

HPLC analysis was performed on a Shimadzu 20A HPLC system, which was equipped with a Inertsil ODS-3 (150 mm × 4.6 mm I.D., 5 μm) column. The mobile phase consisted of a mixture of 0.5% acetic aqueous solution and acetonitrile (55:45, v/v), at a flow rate of 1.0 mL/ min. The column was kept at room temperature, and the effluents were monitored at 260 nm. The plasma concentration−time data were fitted using WinNonlin 5.0.1 software, and the pharmacokinetics parameters such as tmax, Cmax, AUC, and MRT were obtained (Table 2). All the values were presented as mean ± SD for five rats. Statistically significant differences between two groups were evaluated by Student’s t test. A probability (P) of less than 0.05 is considered statistically significant.

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RESULTS AND DISCUSSION Crystal Structure of Cap-Zn. The asymmetric unit of CapZn contains one zinc cation and one captopril anion, in which both carboxyl and thiol groups of captopril are deprotonated (Figure 1a). The Zn(II) ion displays a tetrahedral geometry by coordinating with two sulfur atoms and two carboxylate oxygen atoms from four individual captopril anions. In Cap-Zn, the Zn(II) ions are alternately connected by captopril anions to form a right-handed helical chain along the c axis (Figure 1b). Each helix contains four Zn(II) and four captopril anions, with a pitch of 12.11 Å. Each right-handed helical chain connects with four 2601



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Figure 4. XRPD patterns for Cap-Zn after powder dissolution experiment in Tris-HCl buffer of pH 6.8 (blue) and 0.1 M HCl aqueous solution (red) (black, simulated XRPD pattern from the singlecrystal diffraction of Cap-Zn).

Figure 2. DSC and TG curves for Cap-Zn (red) and Cap (blue).

Figure 5. IDR profiles for Cap and Cap-Zn in 0.1 M HCl aqueous solution.

Figure 3. Powder dissolution profiles for Cap and Cap-Zn in Tris-HCl buffer of pH 6.8 (a) and 0.1 M HCl aqueous solution (b).

adjacent right-handed helical chains through Zn−S and Zn−O1 coordination interactions, resulting in a 3D homochiral coordination polymer of Cap-Zn (Figure 1c,d). Stability. The thermal behaviors of Cap-Zn were investigated by DSC and TGA, and the results are presented in Figure 2. From the TG curve it can be found that no weight loss belonging to residual solvent was detected. Cap-Zn can be stable up to 250 °C and then begins to decompose upon further heating.

Figure 6. Plasma concentration−time profiles for Cap and Cap-Zn after oral administration. Each point represents the mean ± SD (n = 5).

Compared to the decomposition temperature of Cap at 170 °C, Cap-Zn shows the enhanced thermal stability (250 °C). 2602



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molecule, as Cap-Zn can be transformed to free Cap in 0.1 M HCl aqueous solution, as evidenced by the ESI-MS measurement (Figure S1, Supporting Information). It can be expected that the lower Cmax of Cap-Zn may reduce the side effects of captopril. Although Cap-Zn has a relatively lower AUC, its MRT is longer than that of Cap, which means the therapeutic effect of Cap-Zn may sustain for a longer period of time.

Oxidation stability was evaluated by slurry method. After slurrying the solids in deionized water at 40 °C for 3 days, the residuals of Cap are 90 and 98% for Cap and Cap-Zn, respectively, demonstrating the oxidation stability of Cap is enhanced after the formation of Cap-Zn coordination polymer. From the crystal structure of Cap-Zn it can be found that thiol is coordinated with zinc ion, which results in the improved stability of Cap-Zn. Powder Dissolution. Powder dissolution profiles for Cap and Cap-Zn are shown in Figure 3. From Figure 3 it can be found that after formation of Cap-Zn coordination polymer, the apparent solubility of Cap in the Tris-HCl buffer solution (pH = 6.8) was dramatically decreased from 140 mg/mL for the free Cap to 0.3 mg/mL for the Cap-Zn coordination polymer (Figure 3a) . In 0.1 M HCl aqueous solution, Cap-Zn displays different dissolution behavior; the apparent solubility value of Cap-Zn is one-seventh as much as that of Cap (Figure 3b). After the dissolution experiments, the undissolved solids were filtered and dried under vacuum. The result of XRPD analyses indicates that Cap-Zn solid does not change during the experiment (Figure 4), while the pH value of the solution changed from 1 to 3 after the experiment, which is attributed to the protonation of dissolved captopril anion, and this was confirmed by the ESI-MS measurement (Figure S1, Supporting Information). Intrinsic Dissolution Rate. IDR is a key physicochemical parameter commonly used to assess the dissolution-rate controlled absorption of a new compound. Owing to its kinetic nature, IDR assumes a better correlation with in vivo drug dissolution dynamics than solubility.14 To quantitatively evaluate the impact of the solid-state modification on the dissolution behavior, as well as to estimate the clinical potential of Cap-Zn, studies of intrinsic dissolution rate were carried out in 0.1 M HCl aqueous solution and Tris-HCl buffer (pH = 6.8). As the concentration of Cap-Zn in Tris-HCl buffer is too low to be detected by HPLC during the IDR experiment, the discussion is focused on the results in 0.1 M HCl aqueous solution. The intrinsic dissolution profiles within the first hour of the studied systems are shown in Figure 5. The calculated R2 for Cap shows excellent linearity over the entire time interval, while R2 is relatively low (0.98) for Cap-Zn, which can be ascribed to the disproportionation of Cap-Zn in 0.1 M HCl aqueous solution. The IDR curve for Cap-Zn is a result of two processes, namely, the dissolution accompanied by disproportionation, so it does not show a perfect linearity. From the result of the IDR experiments it can be found that Cap-Zn shows a slower dissolution rate than Cap, which coincides with the result of the powder dissolution experiments. Cap-Zn displays the reduced solubility because Cap and zinc ion are connected by coordination bonds, which are stronger than hydrogen bonds. Since solubility and bioavailability are often related, it is supposed that Cap-Zn may reduce the overhigh plasma concentration peak. Pharmacokinetics in Rat Plasma. The plasma concentration−time profiles of Cap and Cap-Zn are shown in Figure 6. The curves of two formulations were both fitted in a noncompartmental model. The mean pharmacokinetic parameters are summarized in Table 2. From Table 2, it can be found that the pharmacokinetic parameters of Cap-Zn are statistically different from those of Cap. The low apparent solubility and slower dissolution rate of Cap-Zn result in lower Cmax and longer tmax, which change from 2.53 μg/mL and 10 min for Cap to 0.99 μg/mL and 23 min for Cap-Zn. We believe that Cap-Zn was absorbed through a dissociated form rather than a complex

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CONCLUSIONS A coordination polymer of captopril with zinc ion was synthesized, and its structure was determined by single crystal X-ray diffraction in which both thiol and carboxyl groups of captopril are deprotonated and coordinated with zinc ion to form a 1D coordination polymer of Cap-Zn. The oxidation stability of captopril is improved after the formation of coordination polymer. The results of solubility and IDR experiments indicate that Cap-Zn shows a reduced apparent solubility and dissolution rate compared to Cap. In 0.1 M HCl aqueous solution, Cap-Zn was disproportionate to captopril molecule during dissolution experiments. Pharmacokinetic research indicates that the maximum plasma concentration reduced from 2.53 μg/mL for captopril to 0.99 μg/mL for Cap-Zn. The results presented herein provide a simple, efficient, and new approach to reduce the release rate and overhigh plasma concentration of a given drug.

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ASSOCIATED CONTENT

S Supporting Information *

ESI-MS of Cap-Zn in 0.1 M HCl aqueous solution; XRPD patterns, IR spectrum for Cap-Zn, and HPLC for Cap and CapZn. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Authors

*(J.-M.C.) Fax: +86-20-84112921. E-mail: chenjm37@mail. sysu.edu.cn. *(T.-B.L.) E-mail: [email protected]. Funding

This work was financially supported by NSFC (grant no. 21101173, 91127002, and 21331007), NSF of Guangdong Province (S2012030006240), and Guangzhou Pearl River New Star Fund Science and Technology Planning Project to J.-M.C. (2013J2200054). Notes

The authors declare no competing financial interest.

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New Approach to Reduce the Overhigh Plasma Concentration of

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