Thermal Analysis of Albendazole investigated by HSM
Transcription
Thermal Analysis of Albendazole investigated by HSM
Microscopy: advances in scientific research and education (A. Méndez-Vilas, Ed.) __________________________________________________________________ Thermal Analysis of Albendazole investigated by HSM, DSC and FTIR J.R. Moyano1, J. Liró1, J.I. Pérez1, M.J. Arias1 and P.J. Sánchez-Soto2 1 Departamento de Farmacia y Tecnología Farmacéutica, Facultad de Farmacia, Universidad de Sevilla, c/Profesor García González 2, 41012 Sevilla, Spain, email: [email protected] 2 Instituto de Ciencia de Materiales de Sevilla (ICMS), CSIC-Universidad de Sevilla, Centro de Investigaciones Científicas Isla de La Cartuja de Sevilla, c/Américo Vespucio 49, 41092 Sevilla, Spain, e-mail: [email protected] The thermal behavior of albendazole, an antihelmintic drug, crystallized in different solvents, was analyzed using Differential Scanning Calorimetry and complemented by Hot Stage Microscopy Studies. Fourier Transform Infrared Spectroscopy was applied to monitoring the integrity of albendazole during thermal treatment. Thermal analyses revealed the existence of a low melting form of albendazole. A hypothetic new crystal form generated by crystallization of the melt was isolated and studied by FTIR spectroscopy, but in this case corresponds to an oxidized form of the original drug. Keywords: Albendazole; Recrystallization; HSM; DSC; Thermal stability 1. Introduction Polymorphism is defined as the existence of at least two different solid phases for a molecule. Solid phases can be crystalline, amorphous or solvates, e.g. crystalline phases that include solvent molecules in their structures (hydrates when water is the solvent). The research of the nature of different crystalline forms of compounds constitutes a very important step in pharmaceutical development, not only for active pharmaceutical ingredients (APIs) but also for excipients. The physicochemical properties of interesting pharmaceutical compounds are strongly related with the structure at the solid state, such as apparent solubility, hygroscopicity, dissolution rate or rheological behaviour. Consequently, the study of polymorphism becomes one of the most-important tasks in a serious preformulation process. Screening for detection of crystal habits, solid phases, determination of their relative stabilities, differentiation between stable and metastable forms, thermal decomposition and the identification of transition factors are mandatory during the development of new APIs or dosage forms [1]. For these objectives, the isolation of solid phases by systematic recrystallization processes is combined by different characterization techniques. The most usual techniques are Differential Scanning Calorimetry (DSC) and Hot Stage Microscopy (HSM) that constitutes the first step in polymorphism studies. In particular, HSM enables direct observation of a sample during heating, cooling or under isothermal conditions. Such investigations provide reliable firsthand evidence about the processes occurring [2,3]. In a second step, a depth characterization is developed by the use of vibrational spectroscopy, as Fourier Transform Infrared Spectroscopy (FTIR) or RAMAN, X-ray diffraction (powder, single crystal) and solid-state nuclear magnetic resonance (NMR) [4,5]. Albendazole (ABZ) is an anthelmintic active in neurocysticercosis by larvae of Taenia solium. It is active against other species such as Ascaris lumbricoides, Wuchereria bancrofti, Trichuris trichiura, Enterobius vermicularis, Ancylostoma duodenale, Strongyloides stercoralis, Echinococcus granulosus, Necator American, Hymenolepis nana, Taenia sp. and Giardia lambia [6-8]. ABZ is closely related with mebendazole, another antihelminitc drug, which have been described up to three polymorphic forms [9,10]. Our investigations are oriented through the thermal history of albendazole (ABZ). Thermal behaviour of samples was investigated by using DSC. Polarized light optical microscopy and Hot Stage Microscopy (HSM) were applied to study the crystal patterns of ABZ in stock form and crystallized from different solvents. Different ABZ crystals were prepared through recrystallization form oversaturated solutions in different common organic solvents. FTIR was used as complementary technique to confirm the nature of compounds produced during thermal treatment. 2. Materials ABZ (lot number 035K1421) was purchased from Sigma-Aldrich Co. (St. Louis MI, USA). All of the solvents employed (Chloroform, Methanol, Acetone, Acetonitrile, Ethyl acetate, Cyclohexane, Diethyl ether, Dichloromethane and Methyl amine) were of HPLC purity grade. © FORMATEX 2014 1043 Microscopy: advances in scientific research and education (A. Méndez-Vilas, Ed.) __________________________________________________________________ 3. Methods 3.1 Recrystallization process Recrystallization from stock ABZ was carried out by its dissolution in different solvents aided by agitation and heating. The obtained solutions were filtered under vacuum in order to get a particle-free limpid oversaturated solution. The resultant clean solutions were transferred into a crystallizer, covered and stocked in a quiet place to cool slowly and to allow crystallization. Alternatively, filtered solutions were crash cooled (by using a blend of ice/salt or liquid nitrogen) to induce fast precipitation. Solid samples were collected by filtration, dried under vacuum and stored at ambient temperature prior to analysis. A summary of solvents employed and treatments to induce precipitation/crystallization is reported in Table 1. Table 1 Experimental conditions for ABZ recrystallization; a: slow cooling to ambient T; b: crash-cooling in liquid nitrogen and filtration under vacuum of resultant precipitate; c: crash cooling in ice/salt bath and filtration under vacuum; d: slow solvent evaporation at ambient temperature and filtration. *: Crystals obtained after fusion of ABZ. Solvent Treatment type Chloroform a Chloroform a Methanol a Acetonitrile a Methanol a Ethyl acetate a Acetonitrile a Chloroform b Methanol b Methanol c Chloroform+acetonitrile (1:1) a Chloroform+acetonitrile (1:1) c Ethyl acetate+chloroform (1:1) a Cyclohexane a Chloroform+cyclohexane+acetone d Diethylether+acetone d Methylamine d Notes Nucleation induced by seed plain crystals* Nucleation induced by seed plain crystals* Nucleation induced by seed plain crystals* 3.2 DSC DSC analyses were carried out through a Mettler FP 80 series equipment or a Mettler DSC 821e. Samples of about 10 mg were accurately weighed in open aluminum crucibles. Heating runs were carried out under static air or inert (N2) atmosphere at 5 or 10 ºC·min-1 rates. 3.3 HSM Hot stage microscopy studies were performed by using a Mettler FP82HT hot plate linked to a FP85 control unit. The optical equipment was an Olympus BH2 microscopy and an Olympus Camedia 5600 as recording unit. Polarized light filter was employed to distinguish between amorphous and crystalline phases. The applied heating rates were the same as DSC experiments. Isothermal conditions were used in some interesting transitions. 3.4 FTIR FTIR analyses of samples, before and after thermal analysis, were performed through a Jasco FT/IR 410 apparatus controlled by Spectra Manager for Windows™, by using KBr press tablet method. Wavenumbers ranged between 600 and 4000 cm-1, with 20 scans for sample and a resolution of 4 cm-1. 1044 © FORMATEX 2014 Microscopy: advances in scientific research and education (A. Méndez-Vilas, Ed.) __________________________________________________________________ 4. Results and discussion DSC runs of selected samples are collected in Figs. 1 and 2. Stock ABZ is characterized by the presence of two peaks: first, an acute endothermic effect (onset T 186 ºC; peak T: 202 ºC), which overlaps with another second sharp peak (peak T: 205 ºC). These data differs from the declared melting point of the substance (215 ºC). DSC thermogram of ABZ (30-225 ºC, N2 atmosphere, heating rate 10 ºC·min-1). Fig. 1 Figure 2 reports the DSC analyses of recrystallized samples compared with original ABZ. One of the most remarkable differences in the DSC runs of crystals obtained by recrystallization in methanol (see Figs. 2b-c) appears as a clear endo-exo effect at 143 ºC, followed with the endothermic effects described for the stock ABZ. The peaks above 200 ºC appear overlapped thus difficulting their interpretation. An analogous behaviour was observed for other recrystallized samples. This endo-exo effect corresponds to a solid-solid transition polymorphic transformation of ABZ obtained by crystallization to stock ABZ [11]. These two polymorphic forms were successfully isolated by other authors, being characterized to have an enantiotropic relationship [12]. a b c Fig. 2 DSC thermograms of: a) stock ABZ; b) recrystallized from methanol (slow cooling); c) recrystallized from methanol (crash cooling with liquid N2) (30-225 ºC, N2 atmosphere, heating rate 10 ºC·min-1). While the nature of endo-exo effect was described in detail in literature, the endothermic events over 200 °C are not sufficiently explained. In order to elucidate the nature of these phenomena, direct observation by HSM was performed. Photomicrographs of stock ABZ together with recrystallized samples are reported in Fig. 3. ABZ (Fig. 3a) can be described as a micronized microcrystalline powder, confirmed by the observation of birefringence under polarized light. © FORMATEX 2014 1045 Microscopy: advances in scientific research and education (A. Méndez-Vilas, Ed.) __________________________________________________________________ Fig. 3 Microphotographs (100x magnification) at ambient T of: a) Stock ABZ; recrystallized ABZ from b) methanol (slow cooled); c) methanol (crash cooled); d) methyl amine; e) acetonitrile:chloroform; f) ethyl acetate:chloroform; g) methanol (seed with plain crystal); h) cyclohexane. Crystal growth from different experimental conditions become to the formation of a wide variety of crystal shapes. Slow cooling from methanol (Fig. 3b) generates the biggest crystals. Crystals from methylamine (Fig. 3d) are irregular, being similar in shape with those obtained from acetonitrile:chloroform (Fig. 3e). Figure 3g show the ABZ crystal from methanol seeded with plain crystals (seeds were obtained with thermal treatment of ABZ over 200 ºC) originating acicular-shaped crystals, very different compared with other methanol samples. Finally, only very small crystals were obtained from cyclohexane (Fig 3h). Figure 4 summarizes the thermal behaviour of stock ABZ sample observed under microscopy. ABZ first melting was registered at 186-190 ºC, followed by a fast recrystallization of the melt. The formed microcrystals melt again at 216 ºC, 1046 © FORMATEX 2014 Microscopy: advances in scientific research and education (A. Méndez-Vilas, Ed.) __________________________________________________________________ which is in agreement with DSC data. Cooling of this melt to 180 ºC and subsequent heating to 190 ºC did not result in fusion, indicating the presence of a new solid phase. Fig. 4 Microphotographs (100x magnification) of stock ABZ at different T: a) 186 ºC (melting onset); b) 190 ºC; c) 190 ºC (isotherm, recrystallization); d) 190 ºC (isotherm, recrystallized); e) 216 ºC (new melting); f) 190 ºC (isotherm after cooling from 220 to 180 ºC). Between all the ABZ solid phases elaborated, only HSM analysis of recrystallized ABZ sample from methanol was selected to be included in this work, because is representative of other recrystallized samples with a similar recorded behaviour. Photographs at 140 ºC (Fig. 5c), which correspond to the endo-exo effect, did not show any evident change in crystal shape. Therefore, transformation is present because sample behaviour is similar as compared with stock ABZ, indicating that at 140 ºC ABZ reverts to its original form. Accordingly, melting-crystallization and final melting (Figs. 5d-h) are similar to stock ABZ. On the other hand, the change in appearance after first melting could be due either to a polymorphic transformation or a decomposition process. In fact, following the first hypothesis, some crystals generated by heating over 200 ºC were employed as seeds, trying to reproduce a similar solid phase by nucleation from some saturated solution of ABZ. In view of the results obtained by thermal analysis, it was not possible to determine the nature of the material after melting. Therefore, we used an alternative technique such as FTIR in order to confirm the hypothesis of thermal decomposition, as literature data suggest with respect to the related compound mebendazole [13] (see Fig. 6). © FORMATEX 2014 1047 Microscopy: advances in scientific research and education (A. Méndez-Vilas, Ed.) __________________________________________________________________ Fig. 5 Microphotographs of ABZ recrystallized from methanol: a) at ambient T; b) ambient T (polarized light); c) 140 ºC; d) 186 ºC (melting); e) 189 ºC (melting); f) 192.5 ºC (melting); g) 201 ºC; 7) 217 ºC (second melting); h) 220 ºC. FTIR spectra of selected samples are reported in Fig. 7. The FTIR spectra of the original product and recrystallized from methanol the same profile (data not shown). However, samples heated up to 220 °C show a new absorption band at 1029.8 cm-1, which is related to the formation of sulfoxide groups, thereby being demonstrated that melting is followed by an oxidative process. Consequently, the presence of plain crystal after recrystallization does not match with a new ABZ form, but a new compound that differs of the starting substance (see Fig. 8). As described in the literature [7-8], ABZ sulfoxide is the active form of ABZ. In fact, ABZ actually constitutes a prodrug that only becomes active after biotransformation in plasma in its oxidized form. 1048 © FORMATEX 2014 Microscopy: advances in scientific research and education (A. Méndez-Vilas, Ed.) __________________________________________________________________ Fig. 6 Fig. 7 Fig. 8 Thermal degradation of mebendazole. FTIR spectra of: (a) stock ABZ and (b) sample heated to 220 °C. Original ABZ structure (a) and proposed (b) degradation product (ABZ sulfoxide). References [1] [2] [3] [4] [5] [6] WHO Expert Committee on Specifications for Pharmaceutical Preparations. Forty-sixth report. Annex 4: Guidelines on submission of documentation for a multisource (generic) finished pharmaceutical product for the WHO Prequalification of Medicines Programme: Quality part. p. 141-2 [Internet]. Geneva, Switzerland; 2012 [cited 2014 May 5]. Available from: http://www.who.int/medicines/areas/quality_safety/quality_assurance/expert_committee/TRS-970-pdf1.pdf Marti E. Applied chemical thermodynamics and kinetics on pharmaceutical compounds. J Thermal Anal. 1988; 33(1): 37-45. Szelagiewicz M, Marcolli, C, Cianferani S, Hard, A, Vit A, Burkhard A, von Raumer M, Hofmeier U, Zilian A, Francotte E, Schenker R. In Situ Characterization of Polymorphic Forms: The Potential of Raman Techniques. J Thermal Anal Cal. 1999; 57(1): 23-43. Brittain HG. Spectral methods for the characterization of polymorphs and solvates. J Pharm Sci. 1997; 86(4):405-12. Hilfiker R. (Ed.) Polymorphism: In the Pharmaceutical Industry. Weinheim, Germany: John Wiley & Sons; 2006. Navarrete-Vázquez G, Yépez L, Hernández-Campos A, Tapia A, Hernández-Luis F, Cedillo R, González J, MartínezFernández A, Martínez-Grueiro M, Castillo R. Synthesis and antiparasitic activity of albendazole and mebendazole analogues. Bioorg Med Chem. 2003; 11(21):4615-22. © FORMATEX 2014 1049 Microscopy: advances in scientific research and education (A. Méndez-Vilas, Ed.) __________________________________________________________________ [7] Ingold K, Bigler P, Thormann W, Cavaliero T, Gottstein B, Hemphill A. Efficacies of albendazole sulfoxide and albendazole sulfone against In vitro-cultivated Echinococcus multilocularis metacestodes. Antimicrob Agents Chemother. 1999; 43:105261. [8] Vuitton DA. Benzimidazoles for the treatment of cystic and alveolar echinococcosis: what is the consensus?. Expert Rev Anti Infect Ther. 2009; 7(2):145-9. [9] Himmelreich M, Rawson BJ, Watson TR. Polymorphic forms of mebendazole, Aust J Pharm Sci. 1977; 6:123-5. [10] Swanepoel E, Liebenberg W, de Villiers MM. Quality evaluation of generic drugs by dissolution test: changing the USP dissolution medium to distinguish between active and non-active mebendazole polymorphs. Eur J Pharm Biopharm. 2003; 55(3):345-9. [11] Liró J. Estudio del polimorfismo en albendazol mediante análisis térmico y espectroscopía de infrarrojos. Trabajo fin de máster. Máster en Ciencia, Tecnología y Uso Racional del Medicamento. Departamento de Farmacia y Tecnología Farmacéutica. Universidad de Sevilla; 2008. [12] Pranzo MB, Cruickshank D, Coruzzi M, Caira MR, Bettini R. Enantiotropically related albendazole polymorphs. J Pharm Sci. 2010; 99(9):3731-42. [13] Brusau EV, Cami GE, Narda GE, Cuffini S, Ayala AP, Ellena J. Synthesis and characterization of a new mebendazole salt: mebendazole hydrochloride. J Pharm Sci. 2008; 97(1):542-52. 1050 © FORMATEX 2014