characterization of physical forms of donepezil...

254 Acta Chim. Slov. 2007, 54, 254–267

Vrbinc et al.: Characterization of Physical Forms of Donepezil Hydrochloride

1. IntroductionMajority of pharmaceutical solids, both active ingre-

dients and excipients, exhibit a phenomenon of occurancein different unsolvated and solvated crystal forms by thesame solid substance, namely polymorphism and pseudo-polymorphism.1–3 A novel, supramolecular definition ofpolymorphism defines it as »the existance of more thanone type of network superstructure for the same molecularbuilding blocks«, and pseudopolymorphism as the casewhere the solvent is one of molecular components in thenetwork.4 Pseudopolymorphism refers to solvated forms,where solvent in crystal network of majority of pharma-ceuticals is water. Hydrates are further classified accord-ing to the nature of release of water from the crystal latticeas stochiometric (crystal lattice collapses upon dehydra-tion) and non-stochiometric hydrates (crystal lattice is es-sentially retained upon dehydration) and can be used indescription of transformations of crystal forms during tec-hnological processes, e.g. crystallization, milling/grin-ding, granulation, drying, tabletting, and rarely in filmcoating.5

Characterization of solid active ingredient in bulkand solid dosage form is very important in quality assur-ance of final product. The most frequent analytical meth-ods are differential scanning calorimetry (DSC), thermo-

gravimetry (TGA), moisture sorption/desorption isother-mal analysis, infrared (IR) and Raman spectroscopy, X-ray powder diffraction (XRPD), solubility, microscopy,solid-state nuclear magnetic resonance (ss-NMR), andshould be used in a proper combination to determine themost important differences between various forms.6

Donepezil (D), (±)-2,3-dihydro-5,6-dimethoxy-2-((2-(phenylmethyl)-4-piperidinyl)methyl)-1H-inden-1-one, is piperidine-based, long-acting reversible acetyl-cholinesterase (AchE) inhibitor that exhibit a relativelyhigh degree of selectivity for neuronal AchE. It increasesa level of acetylcholine (Ach) in the brain of patientswith mild to moderately severe Alzheimer’s disease(AD).7–9

The most obvious neurochemical change observedin patients is rigorous deficiency of choline acetyltrans-ferase, which is responsible for the synthesis of Ach.

Scientific paper

Characterization of Physical Forms of Donepezil Hydrochloride

Miha Vrbinc,* Berta Jordan-Kotar, Matej Smrkolj, Franc Vre~er

* Corresponding author: Krka, d. d., Novo mesto, Šmarje{ka cesta 6, 8501 Novo mesto, SI-SloveniaTel.: +386 7 33 13 535, Fax: +386 7 33 13 751, E-mail: [email protected]

Received: 20-07-2006

AbstractDifferent crystal forms and amorphous form of donepezil hydrochloride were prepared and characterized, using variousanalytical techniques. Hydrate form I is stable only at lower relative humidity (RH), while at higher RH it sorbes addi-tional water. Forms II, III and V were identified as polymorphs, whereas form III is thermodynamically stable. Form Vwas prepared by heating of hydrate IV at high temperature (about 150 °C). Three different forms of hydrate IV were ob-tained and characterized. Differential scanning calorimetry and X-ray powder diffraction are the most promising analyt-ical methods for differentiation between crystal forms, both polymorphs and hydrates. Methods could be also used indetermination of physical form of a drug, incorporated into solid dosage form. Thermal methods explain the most sig-nificant difference in dehydration of different forms IV: transformation to pure polymorphic forms or mixture thereof.On heating of hydrates I and IV-1, two novel crystal forms were prepared and characterized, namely crystal forms VIIand VIII.

Keywords: donepezil hydrochloride, polymorphs, hydrates, characterization

Figure 1. Chemical structure of donepezil hydrochloride.

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Between several therapeutic approaches, inhibition ofcholinesterase has been most investigated9. Symptomatictreatment of AD may at least maintain function in activi-ties of daily living10.

In solid dosage forms, (D) is in the form ofdonepezil hydrochloride (DH)7–9 (Figure 1). Besides(DH), another halogenide salt, donepezil hydrobromidehas been studied, where six physically distinct forms werecharacterized: four polymorphs, hydrate and amorphousform11. Organic salts of (D) were also obtained by use oftosyllic, mesyllic, benzoic, salicylic, tartaric, citric acidand the like, and are described to be more chemically sta-ble than (D) and (DH) in solution by exposure to light(1000 Lux, 1 month) and temperature (60 °C, 1 month),subsequently12.

Different crystal forms of (DH) have been preparedand partially characterized. Five crystal forms (I, II, III,IV and V) and amorphous form were characterized bymelting point, XRPD and IR spectroscopy. From the lit-erature data on water content it can be concluded, thatforms I and IV can be described as monohydrate and tri-hydrate, respectively. Crystal forms I–IV are preferentialover amorphous form due to their chemical stability if ex-posed to temperature (–20–80 °C) up to 3 month and sev-eral RH (10.6–100%) at controlled room temperature13.Crystal form VI was prepared from donepezil oxalate14.Further, another four crystal forms (H1, H2, monohy-drate, sesquihydrate) were prepared and characterized byXRPD15.

However, different crystal forms (A, B and C) wereobtained and characterized also in (D)16.

In the present article, preparation and physical char-acterization of different crystal forms and amorphousform of (DH) is highlighted. In order to assure clearnessof nomenclature, »polymorph« and »polymorphic form«refer only to anhydrous crystalline forms, while terms»hydrate« and »hydrate form« refer to hydrated crys-talline forms.

2. Experimental

2.1. Materials(D) and (DH) were obtained by Krka, d.d., Novo

mesto (Slovenia). Polymorphs II, III and hydrates IV-1and IV-2 were prepared by crystallization of (DH) fromdifferent solvents. Hydrates I and IV-3 were prepared byin situ formation of (DH) from (D) by addition of hy-drochloric acid. Polymorph V was prepared by heating ofhydrate forms IV (dehydration). Amorphous form wasprepared by freeze-drying13. Crystal forms VII and VIIIwere prepared by heating of hydrates I and IV-1 on 90 °C.

Solvents used for crystallization were of analyticalgrade and used as received. Hydrochloric acid was ob-tained by Merck (Germany). Solid carbon dioxide was ob-tained by Messer (Slovenia).

2.2. MethodsThermal properties of physical forms were charac-

terized, using Mettler Toledo DSC822e differential scan-ning calorimeter and Perkin Elmer TGA-7 thermogravi-metric analyzer. DSC curves were recorded under nitro-gen atmosphere at heating rate 10 °C/min in temperaturerange 30–250 °C.

Particle size distribution was measured on MalvernMastersizer S/Femtosystems. Samples were dispersed insunflower oil.

Hot stage microscopy was performed on MettlerToledo termosystem FP900 with FP82HT hot-stage cell.

Mid-IR spectroscopy was obtained on Perkin ElmerFT-IR Spectrometer 1720X in range 400–4000 cm–1 (res-olution 4 cm–1) as KBr pellets.

Near-IR spectroscopy was performed on PerkinElmer FT-IR Spectrometer GX in range 10000–4000 cm–1

(resolution 8 cm–1). Raman spectroscopy was performed on Perkin

Elmer FT-IR Spectrometer GX in range 4000–100 cm–1

(resolution 4 cm–1).X-ray powder diffraction (XRPD) analysis was per-

formed on powder diffractometer Philips X’pert Pro undercontrolled conditions (CuKα radiation, λ = 1.54187 Å, 2θrange 2.9280–31.0615°, step 0.0167° 2θ).

Inverse gas chromatography (IGC) was tested inarranged gas chromatograph Agilent Technologies 6890Nby the following adsorptives: methane, n-hexane, n-hep-tane, n-octane, n-nonane, n-decane, tetrahidrofurane(THF), acetone, ethylacetate and chloroform.

True density was determined by Accupyc 1330 heli-um pycnometer.

Dynamic vapour sorption (DVS) was studied on au-tomatic water sorption analyzer DVS-1 (Surface Measure-ment Systems Ltd.) under following conditions: contro-lled room temperature (25 °C), nitrogen flow: 200 mL/min, 2 complete cycles 0–95% RH and backwards in 11steps, minimal time in 1 step (dm/dt < 0.002%): 10 minu-tes, maximal time in 1 step: 360 minutes.

Scanning electron microscopy (SEM) was per-formed on microscope Olympus BX50, connected toCCD camera Sony XC-75CE.

13C-solid state nuclear magnetic resonance (13C ss-NMR) spectra were obtained by Varian Unity Inova 600MHz NMR spectrometer, using RAMP/CW technique at10 kHz.

3. Results and Discussion

DSC curves of polymorphs II, III and V show no dif-ference in basic line until sharp melting, which confirmshigh crystallinity of samples with low degree of admixedamorphous form (Figure 2). Pure amorphous form is soft-ened at about 120 °C, which was confirmed also by hotstage microscopy. Distinction between individual physical

forms is based on melting points for crystal samples andenthalpies (Tables 1 and 2). Amorphous form shows twocrystallization exothermic effects, where the first one ismuch stronger than the second. Broad melting endothermwith a shoulder is observed after both exothermic effects,showing the presence of two crystalline forms. From thecomparison of thermal behaviour of crystalline and amor-phous form, it can be concluded that forms II and III arepresent in the crystallized amorphous form.

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Figure 2. DSC curves of amorphous and polymorphic forms II, IIIand V.

Figure 4. DSC curves of hydrate forms IV-1, IV-2 and IV-3.

Table 1: Melting points and enthalpies of transition of amorphousand polymorphic forms II, III and V.

Parameter Amorphous Form II Form III Form Vform

Tmelting (°C) 221.80* 223.49 229.09 227.62∆Hmelting (J/g) 81.61 105.09 125.78 113.85

* melting of crystallized amorphous form

Parameter Hydrate I Hydrate IV-1 Hydrate IV-2 Hydrate IV-3Tdehydration (°C) 79.33 88.83 90.44 79.25∆Hdehydration (J/g) 150.55 156.64 162.47 119.63

Table 2: Dehydration of hydrate forms I, IV-1, IV-2 and IV-3.

Figure 3. TGA of crystal forms I, II, III, IV-2 and V.

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be defined as monohydrate, although it contains addition-al water (additional hydration), or it can also contain lesswater than theoretical monohydrate, which can be attrib-uted to partial dehydration of monohydrate, or damagedcrystal structure, or to the presence of small amount of hy-groscopic amorphous form.

Form IV-2 contains 11–11.24% of water, dependingon the method of measurement used, which is in good ac-cordance to theoretical trihydrate (11.49% of water). It isassumed that it does not contain significant amount of sur-face bound water. Narrow dehydration peak could be at-tributed to isolated lattice sites of water in crystal form.First loss of weight in TGA (about 1%) is loss of waterfrom sites near the surface of crystals, while the rest ofwater evaporates up to 100 °C. It does not represent a rap-id removal of water from the inner structure which is con-firmed by DSC curve with relatively extended dehydra-tion peak. Endothermic peak after dehydration at about 90°C is connected to transition to polymorph II. Broad en-dothermic peak after transition to form II is most obviousin dehydrated form IV-3.

Dehydration of form IV-3 is easier and starts at sig-nificantly lower temperature (79.25 °C) than at the otherforms IV (88.83 °C for form IV-1 and 90.44 °C for formIV-2). Different forms IV are pressumed also from experi-ments, performed by drying in heating process. Forms IVare finally completely converted to pure form V or mixtureof form V with additional small amount of form II. Thisphenomenon is surprising since polymorphic forms II andV are metastable and spontaneously transit to stable formIII. However, stability studies confirmed good stability of

Table 3: Loss of weight in TGA and water content by Karl-Fischer analysis.

Parameter Hydrate I Form II Form III Hydrate IV-2 Form V Amorphous form∆m1 (%) 1.8 (until 50 °C) 0.3 (until 55 °C) 0.0 (until 200 °C) 1.1 (until 44 °C) 0.2 (until 200 °C) 5.4 (until 100 °C)∆m2 (%) 3.4 (50–115 °C) – – 9.9 (44–120 °C) – –Water content 5.34 0.13 0.05 11.24 0.21 5.67by KFA (%)Theoretical 4.15 0 0 11.49 0 0water content (%)

Table 4: Particle size distribution of amorphous and crystal forms.

Range Particle size distribution (volume %)(µm) Amorphous form Hydrate I Form II Form III Hydrate IV-10.02–2.00 0.0 0.6 0.6 0.9 0.92.00–5.00 0.0 2.9 1.7 3.2 1.35.00–10.00 0.2 6.6 4.7 11.0 1.310.00–20.00 2.1 13.8 5.6 22.1 6.520.00–50.00 7.4 29.0 22.4 18.7 17.850.00–100.00 14.2 21.2 31.0 7.5 25.8100.00–150.00 13.8 9.1 12.0 9.3 22.6150.00–500.00 52.9 15.3 20.9 27.1 23.7500.00–800.00 9.3 1.5 1.2 0.2 0.0

Average size (µm) 241 88 108 95 103

Form III has the highest melting point (229.09 °C)and melting enthalpy (125.78 J/g), which confirms thatthis form is thermodinamically stable. The other two poly-morphic forms are metastable and tend to transform toform III with different kinetics and is influenced by tem-perature and relative humidity. Lower melting point ofthese forms show also lower melting enthalpies: form IIwith melting point 223.49 °C has the lowest melting en-thalpy (105.09 J/g) and form V with melting point 227.62°C and melting enthalpy 113.85 J/g. The absence of waterin forms II, III and V was confirmed by TGA, where lessthan 0.3% loss of weight was observed (Figure 3, Table3). Loss of weight can be attributed to adsorbed, non-crys-talline bound water.

Form I is defined as a hydrate crystal form by sto-chiometric ratio of molecules of (DH) to water 1:1 andtheoretical water content 4.15%. Further, hydrate form IVis defined by stochiometric ratio of molecules of (DH) towater 1:3 and water content 11.49%13. DSC curves of hy-drate forms IV-1, IV-2 and IV-3 (Figure 4) show dehydra-tion until 120 °C, which is confirmed by TGA (Figure 3).Very complex thermal behaviour was obtained by all threeforms of hydrate form IV.

Water content at room temperature in form I is 1.3moles of water on 1 mole of (DH). Water can be sorbed indifferent ways, while TGA confirms loss of weight in twosteps: in lower temperature surface adsorbed water (until50 °C), and crystal bound water of hydrate mainly at high-er temperature (50–115 °C). However, form I can be alsodefined by total amount of water, determined by Karl-Fischer titrimetric method. It is proposed that form I can

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all crystal forms, even if exposed directly to extreme con-ditions (e.g. 40 °C/75% RH) for period of 10 days; stresswith increased both temperature and RH show transfor-mation of metastable forms to thermodynamically stable

polymorph III. Double melting peak of dehydrated formIV-3 could be attributed either to non-homogeneous parti-cle size distribution and/or high proportion of bigger par-ticles (100.00–500.00 µm) (Table 4), or to the presence of

Figure 5. Hot stage microscopy of amorphous form.

T = 200 °C – crystal form T = 211–220 °C – melting of crystal form

T = 30 °C – amorphous form T = 120 °C – beginning of softening

T = 125 °C – glass transition T = 155–180 °C – recrystallization

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mixture of polymorphic forms II (223.49 °C) and V(227.62 °C).

Form V was prepared by heating of hydrate formsIV-1 and IV-2 at about 150 °C in a drying chamber. Theprocess is, however, not repeatable. By using the same,known parameters, prevailing form V was prepared withadmixed form II. It was observed that form II that also ap-peared by heating of hydrate form IV-3. However, poly-morph II obtained by dehydration of hydrate IV-3, re-mains stable even at heating to 170 °C. It is obvious thattransformation of hydrate IV upon heating is throughform II to form V or directly to form V, depending on pa-rameters of samples preparation and difference betweenforms IV-1, IV-2 and IV-3. Nevertheless, form V cannotbe prepared directly from form II, prepared by crystalliza-tion, upon heating, but only from hydrate forms IV. Theprocess is influenced by factors which are not explained

yet, with possible presence of nuclei that relieve appear-ance of prevailing form V, because samples were preparedin the same laboratory. If all crucial parameters are set-tled, every process is repeatable, as described in the litera-ture17–20.

In amorphous form, glass transition temperature couldonly be determined by hot stage microscopy (Figure 5).

Glass transition of amorphous form begins at 120 °C.Wide exothermic peak at about 150 °C represents crystal-lization. Melting of crystal form is at about 220 °C.

Intensive absorption in mid-IR spectra at 1690 cm–1

is connected to vibrations of C=O group in the structure of(DH). Frequency of absorption of C=O group in hydrateform IV-1 (1683 cm–1) is less intensive than in polymor-phic forms II, III and V (1690, 1697 and 1693 cm–1, re-spectively), probably because of forming of hydrogenbonds to water molecules (Figures 6 and 7).

Crystal forms are distinguished in vibration area ofterc-amino group in range 2200–2700 cm–1. Stongest ab-sorption of sp3 hybridized carbon atoms of C–H bonds isat 2922 cm–1 in all polymorphic forms, with exception ofform II (2928 cm–1). Double absorption band at 1590 and1600 cm–1 is of vibrations of C=C aromatic bonds.

The most significant differences between hydratesand polymorphs are in the functional groups area(1200–4000 cm–1) and are effected by interactions of wa-ter molecules through hydrogen bonding21. Additionalabsorption band at 1650 cm–1 that is present only in hy-drate forms, is contributed by water molecules that areannoted to carbonyl group through hydrogen bonding. Inhydrates IV, absorption bands at 3590 cm–1 are indicated,but are overlayed because of presence of multiple OHgroups of water molecules in crystal structure of trihy-drate. They are forming hydrogen bonds, because ofwhich they can be seen as wide absorption bands in re-gion 3100–3700 cm–1 with most intensive absortionbands at 3320, 3350 and 3420 cm–1. Absorption bands inthis region indicates possible presence of free OH group(i.e. tautomerism of (DH)).

Partial dehydration of hydrate form IV-1, obtainedby heating, results in formation of form II in a mixturewith hydrate form. Presence of crystal bound water was

Figure 6. Mid-IR spectra of hydrate forms IV-1, IV-2 and IV-3.

Figure 8. Mid-IR spectra of polymorphic form III and amorphous

form.Figure 7. Mid-IR spectra of polymorphic forms II, III and V.

confirmed by a band at 1650 cm–1, which is not present inform II. Karl-Fischer titration method confirms presenceof 0.8% of water in partial dehydrated form IV-1. Partialdehydration of form IV-1 to form II and hydrate form,again confirms transformation of forms IV through formII to form V. Kinetics of transformation differs in hydrateforms IV-1, IV-2 and IV-3, which was observed also inthermal analysis. Differences in dehydration result in ap-pearance of different forms and mixture thereof: dehydra-tion of form IV-1 results in form V or mixture of forms IIand V, while hydrate IV-3 dehydrates to pure form II,which remain in this form even at heating to 170 °C.

Amorphous form presents weaker and widened ab-sorption bands, characteristic for non-crystalline formsand are significantly different from crystalline forms(Figure 8 shows comparison to thermodinamically stableform III).

Absorption of hydrate forms and amorphous form innear-IR spectra at 5000 and 7000 cm–1 are significantlydifferent from polymorphic forms II and III (Figure 9).The bands are typical for pure water (5150 and 6900cm–1)21. Connecting of water through hydrogen bondingon other molecules shifts values towards higher frequen-cies. Near-IR spectra of form I with splitted signal of OHgroup of water at 5000 cm–1 is atributed to its bonding inthe crystal lattice.

Polar groups (CO, OH) of mid-IR absorption spec-tra are less visible in Raman emission spectra. However,non-polar simetric and almost simetric bonds like C–Cand C=C are more intensive in the latter and influence of

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Figure 9. Near-IR spectra of crystal forms I, II, III, IV-1 and amor-phous form.

Figure 10. Raman spectra of crystal forms I, II, III, IV-1 and amor-

phous form.

Figure 11. X-ray powder diffactograms of crystal forms II, III, IV-1, V and amorphous form.

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water is minimized. Absorption peaks due to hydrate bound water are not signifi-cant and difference between polymorphs and hydrates is not obvious (Figure 10).Due to faster depreciation of Raman influence, neighbouring molecules have onlya minor influence on vibrating molecules, which result in sharper peaks and lessoverlaying.

XRPD analysis is shown to be suitable tool for detection of mixture of crys-tal forms and presence of them in amorphous form, e.g. during shelf-life in bothbulk drug (Figure 11) and solid dosage form, containing (DH).

XRPD after exposing to different temperatures were obtained for hydrates Iand IV-1 (Figures 12 and 13).

Heating of hydrate I results in appearing of a novel crystal form at 80 °C,that is named »crystal form VII«. At 90 °C–120 °C, it is present in pure form.

Table 5: XRPD data of crystal form VII.

No. Pos. d-spacing[[°2Th.]] [[Å]]

1 5.1 17.42 5.2 16.93 5.6 15.84 10.5 8.55 11.5 7.76 12.5 7.17 12.9 6.98 14.4 6.29 15.2 5.8

10 16.1 5.511 17.1 5.212 18.0 4.913 18.4 4.814 19.3 4.615 20.0 4.4416 20.4 4.3517 21.0 4.2218 22.0 4.0419 23.1 3.8520 23.8 3.7421 24.7 3.6122 25.0 3.5623 25.9 3.4424 27.0 3.3025 27.7 3.2226 28.1 3.18

No. Pos. d-spacing [[°2Th.]] [[Å]]

1 6.8 13.02 8.8 10.03 11.2 7.94 13.1 6.75 13.7 6.46 15.3 5.87 15.6 5.78 16.4 5.49 17.8 5.0

10 19.1 4.611 21.6 4.1112 22.5 3.9613 23.2 3.8414 24.1 3.6915 24.9 3.5816 25.5 3.4917 26.4 3.3818 27.7 3.2219 28.2 3.1720 29.0 3.0821 29.5 3.03

Figure 12. X-ray powder diffactograms of heated hydrate form I.

Figure 13. X-ray powder diffactograms of heated hydrate form IV-1.

Table 6: XRPD data of crystal form VIII.

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polymorphic form V with admixed polymorphic form II;the latter disappears at 190 °C, where only polymorph Vis presented. Crystal form VIII is characterized by XRPDdata (Table 6).

Amorphous form was exposed to both increasedtemperature (40 °C) and controlled RH (60%). Crysta-llization and formation of crystal forms was observatedover 67 hours. After 1 hour, crystallization to form V is

Further heating up to 200 °C results in appearing of purepolymorphic form III. Crystal form VII is characterizedby XRPD data (Table 5).

Heating of hydrate IV-1 results in appearing of anovel crystal form at 70 °C, namely »crystal form VIII«that is present in pure form at 90 °C. At 130 °C, the sam-ple shows low crystallinity with appearing of polymorphV. At 140 °C, better crystallinity results in appearing of

Table 7: Parameters of surface characteristics of crystal forms and amorphous form.

γγbs ± SD (mJ/m2) ∆∆Gsp ± SD (kJ/mol)

THF acetone ethylacetate ChloroformHydrate I 46.20 ± 0.51 4.07 ± 0.31 6.27 ± 0.32 5.58 ± 0.33 5.40 ± 0.17Form II 45.08 ± 0.41 2.80 ± 0.35 5.79 ± 0.24 4.78 ± 0.13 4.30 ± 0.03Form III 41.07 ± 1.20 2.67 ± 0.22 6.13 ± 0.18 5.15 ± 0.20 4.37 ± 0.12Hydrate IV-1 43.80 ± 2.85 2.20 ± 0.64 6.02 ± 0.84 4.75 ± 0.36 4.96 ± 0.09Amorphous form 27.23 ± 1.66 3.81 ± 0.33 4.93 ± 0.64 3.99 ± 0.31 1.99 ± 0.01

Figure 14. X-ray powder diffactograms of amorphous form, exposed to 40 °C/60%RH, over the time.

Table 8: Relation of specific components of free enthalpies of adsorptives for crystal forms and amorphous form.

Ratio Hydrate I Form II Form III Hydrate IV-1 Amorphous form∆Gsp

chloroform/∆GspTHF 1.33 1.54 1.64 2.25 0.52

∆Gspchloroform/∆Gsp

ethylacetate 0.97 0.90 0.85 1.04 0.50∆Gsp

chloroform/∆Gspacetone 0.86 0.74 0.71 0.82 0.40

∆Gspethylacetate/∆Gsp

acetone 0.89 0.83 0.84 0.79 0.81∆Gsp

THF/∆Gspethylacetate 0.73 0.59 0.52 0.46 0.96

∆GspTHF/∆Gsp

acetone 0.65 0.48 0.44 0.37 0.77

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completed with additional new crystal form with the fol-lowing 2θ values: 11.3, 13.8, 14.1, 15.9, 22.6, 23.2, 28.6and 29.8°. After 67 hours, the new crystal form is com-pletely converted to crystal form V (Figure 14).

IGC could be used for differentiation on the basis ofspecific surface characteristics of crystal and amorphousform. Both acetone and ethylacetate are capable of inter-actions of Lewis acid and base. Due to stronger electronacceptor action of acetone, ∆Gsp values of it are higherthan of ethylacetate (Table 7, results are average of 3measurements). Reproducibility of measurements doesnot offer a strong confidence, which is confirmed by rela-tively high standard deviation (SD).

Relation ∆Gspchloroform/∆Gsp

THF is appropriate for differ-entiation between individual forms with exception ofpolymorphic forms II and III (Table 8). Chemical groupson surface of crystal forms are capable of formation ofstronger interactions to Lewis acids than bases. HydrateIV-1 have the weakest interactions with basic adsorptives,and strongest interactions to Lewis acids. Hydrate I has

relatively strongest interactions to Lewis bases, whichcould be important in bounding of water at lower RH val-ues (i.e. OH groups of water act as Lewis acids)22.∆Gsp

chloroform/∆GspTHF of amorphous form is significantly low-

er than in the case of crystal forms. Weaker electron donorsurface characteristics of amorphous form are confirmedalso by ∆Gsp

chloroform/∆Gspethylacetate and ∆Gsp

chloroform/∆Gspacetone.

However, interactions of amorphous surface to electrondonor functional groups of adsorptives are stronger whencomparing to crystal forms. Weaker electron donor orstronger electron acceptor characteristics of amorphousform surface in comparison to crystal forms are confirmedby ∆Gsp

THF/∆Gspacetone and ∆Gsp

ethylacetate/∆Gspacetone, too.

True density (ρT) is not the most significant parame-ter to distinct between individual forms (Table 9). Thelowest value of true density was obtained for hydrate formI. True density of polymorphic form III has the highestvalue (1.442 ± 0.004 g/cm3) which is attributed to smallproportion of free spaces between molecules in thermodi-namically stable form. High value of true density of amor-phous for is explainable by presence of bigger particles(Table 4).

No significant difference in solubility and dissolu-tion rate was observed between different forms, which isattributed to good solubility of (DH) according to BCS,23

which is also in good agreement with difference betweenboth polymorphic forms and polymorphic and hydrateforms; the ratio of solubility between them is typicallyless than 2.24

DVS distinguishes forms I and III and is suitablemethod form determination of presence of them in mix-tures (Figures 15 a and 15 b).

Thermodynamically stable form III adsorbes only aminor amount of water, i.e. only 0.13% up to 95% RH. Infirst cycle, replacement of residual methanol, which issolvent of crystallization is observed. Open hysteresisloop explains presence of pores, that retain water afterdrying.

Comparing different hydrate forms IV, a significantdifference is observed in sorption of water at different RHvalues (Figures 16 a, 16 b and 16 c). Form IV-1 representsthe most accurate course of sorption and desorptionisotherms, that at already 20% RH sorbes amount of waterthat corresponds to 3 moles per 1 mole of (DH), which isin good accordance with theoretical trihydrate. Further,only small amount of water is adsorbed to the surface.First sorption cycle of form IV-2, brings adsorption of 2moles of water per 1 mole of (DH) up to 90% RH. Up to95% RH, additional 1 mole of water is adsorbed for for-mation of trihydrate, while amount of water over theoreti-cal trihydrate is bound to the surface. In the second cycle,

Table 9: True densities of crystal forms and amorphous form.

Form Hydrate I Form II Form III Hydrate IV-1 Amorphous formρT ± SD (g/cm3) 1.263 ± 0.000 1.337 ± 0.014 1.442 ± 0.004 1.366 ± 0.019 1.585 ± 0.000

Figure 15 a. Sorption-desorption isotherms of polymorphic formIII.

Figure 15 b. Comparison of sorption-desorption isotherms of hy-drate I and mixture of hydrate I and polymorphic form III.

adsorption course of form IV-2 is similar as for form IV-1.It can be concluded that a physical change happened dur-ing the first cycle to the form similar to hydrate IV-1.Form IV-3 adsorbes 1.5 mole of water per 1 mole of (DH)up to 20% RH, while it further lineary adsorbes 1.5 molesof water per 1 mole of (DH). Sorption over 90% RH isconnected to deliquescence. For all forms IV, it is charac-teristic that in desorption cycle below 10% RH, 3 moles ofwater per 1 mole of (DH) are released. Difference in sorp-tion of water between hydrate forms IV is ascribable bydifferent porosity of samples, i.e. different surface charac-

teristics. Similarity of forms IV-1 and IV-2 are ascribed tothe same method of preparation. Difference betweenforms IV has been noticed also in thermal behaviourwhere dehydration occurred on heating of different sam-ples (Figure 4).

Polymorph II uptakes 4.5 moles of water per 1 moleof (DH) up to 90% RH. In the first cycle, the water is re-tained until 50% RH, while further dehydration dropdown to the value, which is characteristic for trihydrate(Figure 17). Transformation to “meso” form is seemed be-cause of a large uptake of water, which can induce phasechange. In that case, desorption is denoted by minor dropof water content, which depends on stability of formatedhydrate, until final release of water at low RH.

Polymorphic form V uptakes <0.5 moles water per 1mole of (DH) up to 60% RH in first sorption cycle. Above60% RH, it is transformed through monohydrate to trihy-drate with additional surface bound water. Transition inthe second cycle is not finished yet which is confirmed bynot fulfilled desorption in first cycle (Figure 18).

Surface characteristics of crystal and amorphousforms were observated also by SEM (Figure 19). Diffe-rences between crystal forms can be described as differentcrystallinity; the term is ascribed to crystal habit. Zero-crystallinity of amorphous form is determined only byXRPD and not by specific SEM micrograph.

264 Acta Chim. Slov. 2007, 54, 254–267


Figure 16 a. Sorption-desorption isotherm of hydrate form IV-1.

Figure 16 b. Sorption-desorption isotherm of hydrate form IV-2.

Figure 16 c. Sorption-desorption isotherm of hydrate form IV-3.

Figure 17. Sorption-desorption isotherm of polymorphic form II.

Figure 18. Sorption-desorption isotherm of polymorphic form V.

265Acta Chim. Slov. 2007, 54, 254–267


13C-solid state NMR is the only analytical methodwhere it can be possible to determine a structure of indi-vidual physical forms on the basis of chemical shifts forall C atoms in the structure (Table 10, Figure 20). Themethod is very useful also in determination of mixtureof different crystal forms, where specific chemicalshifts are connected to individual crystal forms.Amorphous form with broad peaks is different from allanalyzed crystal forms with sharp peaks for certain Catoms.

4. ConclusionDifferent crystal forms and amorphous form of

(DH) were prepared and characterized by various analyti-cal techniques. 3 polymorphs were identified (II, III andV) and thermodynamically stable form III was determined(the highest: melting point, enthalpy and true density).Among hydrates, two generally different forms were iden-tified: monohydrate I and trihydrate IV. The latter brings anew aspect with existence of different crystal forms of tri-hydrates, namely IV-1, IV-2 and IV-3; term »hydratemor-phism« of (DH) trihydrates is proposed. Difference be-tween trihydrates is attributed to different methods ofpreparation and results mainly by changes in surface char-acteristics, e.g. orientation of functional groups of (DH) incrystal lattice. They were determined by thermoanalyticalmethods, mid-IR spectroscopy and DVS, but less appliedwith IGC that is otherwise appropriate analytical tool fordetermination of difference in surface characteristics.Obvious difference among them was observed also inboth partial and total dehydration with appearance offorms II and V and mixtures thereof. Appearance of poly-morphs II and V depends on heating rate: by heating of

Figure 19. SEM micrographs of crystal forms I (a), II (b), III (c),IV-1 (d) and amorphous form (e).

266 Acta Chim. Slov. 2007, 54, 254–267


Table 10: 13C solid state NMR assignments for amorphous and crystal forms.

Amorphous form Hydrate I Form II Form III Hydrate IV-1 Form V Assignment21–68 26.5–60.0 26.0–59.0 22.0–57.0 27–56 22–58 C10 C10' C9 C3 C8 C2

C11 C11' C12 (OMe)2

100–112 101.1 101.3 102.7 102.0 102.0–109.8 C4 C7102.5 102.7 109.0 107.0

107.3108.3

124–135 128–130 128–135 128–132 128–132 127.6–136.5 C4' C12' C13 C13' C14 C14' C15

100–112 101.1 101.3 102.7 102.0 102.0–109.8 C4 C7102.5 102.7 109.0 107.0

107.3108.3

124–135 128–130 128–135 128–132 128–132 127.6–136.5 C4' C12' C13 C13' C14 C14' C15

148 149.6 149.0 149.7 149.2 148.7 C5 C6 C7'

155 155.6 154.0 156.2 156.2 157154.6

207 208.6 205.5 205.8 209.1 206.8 C1

Figure 20. 13C solid state NMR spectra of crystal and amorphous forms of (DH).

hydrates by DSC at heating rate 10 °C/min, preferentialform II is formed and further transformation to form V isnot detected, while at heating to 150 °C, form V is prefer-ently formed with possible admixture of minor quantitiesof form II; the latter depends on starting form IV. Heating

of forms IV-1 and IV-2 results in formation of form Vthrough form II, while hydrate IV-3 dehydrates to form IIwhich remain in this form even at heating on 170 °C. Ithas to be focused out that hydrate form IV-1 is most simi-lar to the known trihydrate form IV13.

267Acta Chim. Slov. 2007, 54, 254–267


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929–939.

Two novel crystal forms were prepared by heatingof hydrates I and IV-1 and tracked by XRPD. Accordingto the first settled nomenclature for crystal forms of(DH)13, the novel crystal forms were named »crystal formVII« and »crystal form VIII«. They are stable at certaintemperature range. Upon heating, crystal forms VII andVIII transform to polymorphic forms III and V, subse-quently.

Form I contains 1 mole of water per 1 mole of (DH)already at 10% RH, while later adsorption of water ismainly to the surface of the crystal lattice.

Near-IR resolves hydrates from polymorphs, whilethermal analysis brings difference between crystal formsalthough minor difference is observed in melting pointsand enthalpies of both polymorphs and hydrates. A signif-icant difference is observed by XRPD among all crystalforms, while amorphous form brings expected diffrac-togram with broad peak.

The confirmation of the structure difference be-tween polymorphs cannot be determined by universal an-alytical technique – different methods should be used tocomplete characterization of (DH), although 13C ss-NMRbrings the most obvious differences between individualphysical forms on the basis of chemical shifts for all Catoms of the structure.

5. Acknowledgement

The authors kindly acknowledge to Assoc Prof.Odon Planin{ek from University of Ljubljana, Faculty ofPharmacy, for IGC and SEM analysis.

6. References

1. J. Haleblian, W. McCrone, J. Pharm. Sci. 1969, 58, 911–929.2. J. K. Haleblian, J. Pharm. Sci. 1975, 64, 1269–1288.3. T. L. Threlfall, Analyst 1995, 120, 2435–2460.

PovzetekPripravili in karakterizirali smo razli~ne kristalne oblike in amorfno obliko donepezil hidroklorida. Hidrat I je stabilenle pri nizki relativni vla`nosti in pri vi{ji relativni vla`nosti absorbira vodo. Oblike II, III in V so polimorfi, termodinam-sko je stabilna oblika III. Oblika V nastane s segrevanjem hidrata IV pri temperaturi okrog 150 °C. Pripravili in karak-terizirali smo tri oblike hidrata IV. Kot najprimernej{i metodi za dolo~itev kristalnih oblik, tako polimorfov kot hidratov,sta se izkazali diferen~na dinami~na kalorimetrija (DSC) in rentgenska pra{kovna analiza. Metodi sta uporabni tudi zadolo~itev oblike donepezil hidroklorida v trdni farmacevtski obliki. S termi~no analizo smo ugotovili razlike v dehidra-taciji razli~nih oblik hidrata IV, to je prehod v ~isto polimorfno obliko ali v zmes polimorfov. Segrevanje hidratov I inIV-1 daje novi kristalni obliki VII in VIII .

characterization of physical forms of donepezil...

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