RESEARCH ARTICLE

Crystal modification of rifapentine using different solvents

Kun ZHOU, Jun LI (✉), Jianhong LUO, Dongsheng ZHENG

School of Chemical Engineering, Sichuan University, Chengdu 610065, China

© Higher Education Press and Springer-Verlag Berlin Heidelberg 2010

Abstract Rifapentine crystals with different habits wereprepared by recrystallization from selected solvents, suchas methanol, ethanol, chloroform, and acetic acid. Scan-ning electron microscopy, X-ray powder diffractometry,infrared spectrometry, and differential scanning calorime-try were used to investigate the physicochemical char-acteristics of the prepared crystals. The comparativedissolution behaviors of the newly developed crystalsand of rifapentine without being treated were also studied.Results show that the newly developed crystals weredifferent from each other with respect to physical proper-ties but were identical chemically. Needle-shaped crystalswere obtained from methanol, ethanol, and chloroformsolvents, and the block-shaped crystals were obtained fromacetic acid solvent. X-ray diffraction spectra and differ-ential scanning calorimetry investigation on those devel-oped crystals clearly indicate that rifapentine has differentcrystal structure modification. When the crystal wasobtained from acetic acid, the change of crystal habitwas originated from the crystal structure modification. Thedissolution rate of newly developed crystals was found tobe higher than that of rifapentine without being treated.However, the modified crystal obtained from acetic acidshows the lower dissolution rate than crystals obtainedfrom other solvents.

Keywords rifapentine, recrystallization, physicochemicalcharacterization.

1 Introduction

This paper deals with the study of polymorphism ofrifapentine (Fig. 1), a rifamycin derivative with excellentactivity againstMycobacterium tuberculosis in vitro and inanimals. Like most organic compounds, rifapentine canpresent different morphology and physical properties in thesolid state. The different crystal forms of a drug have

different physicochemical characteristics, that is, crystalshape, crystal size, melting point, density, flow properties,solubility pattern, dissolution characteristics, and X-raypowder diffractometry (XRD) pattern, although they arechemically identical. Different physiological and formula-tion factors are responsible for the bioavailability of drugfrom the dosage form. One of the most important physicalfactors, which affect the bioavailability and therapeuticefficacy of drug, is the existence of active ingredients invarious crystal forms having different internal structureand physical properties [1,2]. The drug rifapentine usedherein is practically insoluble in water, but the waterinsolubility and the poor bioavailability are the limitationsof its effective use clinically. Keeping this in view, crystalmodification of rifapentine has been undertaken to improvedissolution and bioavailability. The differences in mor-phology of crystals of the same compound have frequentlybeen reported in literature [3,4]. Possible reasons are achange of the relative growth rates of the different crystalfaces caused by solvent [5,6] and impurity interaction [7]or by a change in supersaturation [8]. An alternativeexplanation is polymorphism, in which the same com-pound shows a different crystal structure. In the presentwork, rifapentine has been recrystallized from selectedsolvents. The newly developed rifapentine crystals werecharacterized by some physicochemical approaches, suchas scanning electron microscopy (SEM), XRD, infrared(IR) spectrometry, and differential scanning calorimetry(DSC).

Received November 4, 2009; accepted November 30, 2009

E-mail: lijun@email.scu.edu.cn Fig. 1 Chemical structure of rifapentine

Front. Chem. Eng. China 2010, 4(1): 65–69DOI 10.1007/s11705-009-0302-6

2 Materials and methods

2.1 Materials

A brick red crystalline rifapentine powder (C47H64N4O12,molecular mass 877.04) was purchased from Leshan SanJiu-Long March Pharmaceuticals Co., Ltd., China. Otherreagents (purity> 99.5%) were methanol, ethanol, chloro-form, and acetic acid from Chengdu Chemical Reagent Co.

2.2 Preparation of rifapentine crystals

The cooling crystallization method used in this study toobserve the effect of solvents on the development of crystalhabits in a changed environment is given below.According to the solubility of rifapentine in different

solvents [9,10], the predetermined amounts of rifapentinewere dissolved in 30 mL of selected solvents (ethanol,methanol, chloroform, and acetic acid) in a jacketed vessel.The solution was stirred at 50°C for 2 h and then cooleddown to 20°C for 4 h. The crystals were then recovered byfiltration under vacuum using a sintered glass funnel andwashed with distilled water. Finally, they were kept inairtight container for further use.

2.3 Scanning electron microscopy

Electron micrograph of crystals was obtained using a SEM(HITACHI S-450) operating at 20 kV. The specimens weremounted on a metal stub (with double-sided adhesive tape)and coated under vacuum with gold in an argon atmo-sphere prior to observation.

2.4 X-ray powder diffractometry

The cavity of the metal sample holder of X-raydiffractometer (XRD) was filled with ground samplepowder and then smoothed out with a spatula. XRDpattern of rifapentine crystals was obtained with the XRD(X′ Pert Pro MPD) operated at 40 kV and 40 mA. CuKα

radiation was utilized in the measurements. The diffractionangles 2θ ranged from 5° to 50°. All XRD measurementswere performed at ambient temperature.

2.5 IR spectroscopy

The spectra were recorded on an IR spectrophotometer(FT-IR, 670 NEXUS, USA) after respective samples weremixed with dried KBr powder and compressed to a 12 mmdisc by a hydraulic press at 10 tonnes compression for 30 s.

2.6 Thermal analysis

The DSC of the samples, 3.5 mg, was carried out using athermal analysis system (NETZSCH DSC 204 F1).

Calibration with standard was undertaken prior to subject-ing the samples, which were heated at 10°C/min in analuminum pan under a nitrogen atmosphere, and a similarempty pan was used as the reference. The instrumentautomatically calculated onsets of melting points andenthalpy of fusion.

2.7 Dissolution studies

Rifapentine and its crystals with specific size range (90–250 μm), 25 mg in each case, were accurately weighed andthe dissolution profile of the drug was determined in a USPno. 1 dissolution test apparatus at 37°C, with basket(100 mesh) with a stirring speed of 50 rpm. The dissolutionmedium was 900 mL of water containing 0.5% sodiumlauryl sulfate. Samples were withdrawn from the dissolu-tion vessels at selected time intervals and analyzed forrifapentine content at 474 nm on a UV spectrophotometrysystem (Shimadzu, Japan).

3 Results and discussion

3.1 Morphology of crystals

Figure 2 shows the SEM of untreated and recrystallizedrifapentine from different solvents under cooling method.It is clear from the figure that the untreated rifapentine areslightly irregular needle-shaped crystals (Fig. 2(a)),whereas the crystals obtained from methanol is needleshaped (Fig. 2(b)). Recrystallization of rifapentine fromethanol and chloroform solution produced regular needle-shaped crystals (Figs. 2(c) and 2(d)), whereas the crystalsobtained from acetic acid is block shaped (Fig. 2(e)).Comparing SEM of crystals obtained from acetic acid andother solutions shows that the crystal shape of rifapentinewas significantly changed to the block shape, which isquite different from the previous needle shape. Thevariations in face dimensions or the appearance ordisappearance of some faces could be the cause of thechange in morphology of rifapentine crystals, obtainedfrom different solvents. Results also showed that the size ofcrystals produced from recrystallization is somewhatdifferent from the size of untreated rifapentine , which issmaller.

3.2 X-ray diffraction

To obtain information on the physicochemical character-istics of the prepared crystals, XRD measurements wereconducted. XRD spectra for all crystals are presented inFig. 3. In the powder diffractogram, sharp peaks atdiffraction angles (2θ) 5.57°, 6.65°, 7.9°, 10.29°, 14.83°,15.66°, 21.28°, and 22.74° were obtained in the case of thedrug rifapentine and the modified crystals obtained fromuntreated, ethanol, methanol, chloroform, and acetic acid,

66 Front. Chem. Eng. China 2010, 4(1): 65–69

respectively. The presence of these sharp peaks is clearlyevident in the comparative diffractogram presented inFig. 3 and the data recorded therein. From the recordeddata, it is evident that there is a significant difference in theentire diffraction pattern or d-spacing values betweentreated and untreated rifapentine samples. The intensity of

the peak in untreated rifapentine is the lowest than that ofall other modified crystals reported herein. This is probablydue to higher crystal perfection in this condition ofcrystallization. At the same time, in Fig. 3, two diffractionpeaks are present in the 2θ range from 15° to 17°.However, for profiles in Fig. 3, a slight but obvious

Fig. 2 SEM of untreated rifapentine (a) and rifapentine recrystallized from methanol (b), ethanol (c), chloroform (d), and acetic acid (e)

Kun ZHOU et al. Crystal modification of rifapentine using different solvents 67

difference from those mentioned above is observed: onlyone diffraction peak is present in the 2θ range from 15° to17°. The change of XRD profile of rifapentine crystalsindicates that crystal structure is changed when acetic acidis present in solution [11].

3.3 IR spectroscopy

For Fig. 4, the spectra of all modified crystals wereidentical and the main absorption bands of rifapentineappeared in all of the spectra. For all crystals in Fig. 4,there was a single peak by –OH in the range of 3250–3800 cm–1, –CH on the benzene ring in the range of 2800–3000 cm–1, and –CN at 1250–1. This indicates that therewas no difference between the internal structure and theconformations of these samples because these were notassociated with changes at the molecular level.

3.4 Thermal analysis

DSC data for the drug rifapentine (untreated) and themodified crystals are shown in Table 1. It should be notedthat the DSC thermograms of modified crystals obtainedfrom untreated, ethanol, methanol, and chloroform showedonly slight variation. However, the modified crystalobtained from acetic acid shows significant changes dueto the crystal structure modification.The DSC curve of crystals from untreated, ethanol,

methanol, and chloroform shows a single endothermic

peak at about 173°C corresponding to the melting of thedrug. There is a very slight but insignificant variation intransition temperature and a little difference (not signifi-cant) in enthalpy of fusion. This may be due to oxidation orphase transformation. Crystals obtained from acetic acidshow that there is significant variation in transitiontemperature and significant difference in enthalpy offusion. The melting endotherm occurred in 164.1°C.Results from IR spectroscopy, XRD analysis, and DSC

taken together led to the conclusion that the crystal habitmodifications were observed during recrystallization ofrifapentine from various solvents.

3.5 Dissolution studies

The dissolution profile of untreated rifapentine and itsmodified crystals from different solvents are shown inFig. 5. Recrystallization of the parent drug from varioussolvents (ethanol, methanol, and chloroform) resulted inthe increase of the dissolution rate of different modifiedcrystals than untreated rifapentine. Especially, crystalsobtained from ethanol and chloroform show higherdissolution rate than untreated rifapentine because of thebetter crystallinity of the modified crystals in these cases.The dissolution rate was closely related to the crystalparticle and the crystal shape. Because the crystal shape ofthe acetic acid is block and that of other solvents is needle,the voidage of acetic acid is lower than others. As a result,crystals obtained using only acetic acid show lowerdissolution rate than other crystals obtained. As indicated

Fig. 3 XRD pattern of untreated rifapentine and rifapentinerecrystallized from methanol, ethanol, chloroform, and acetic acid,respectively

Fig. 4 IR spectroscopy of untreated rifapentine and rifapentinerecrystallized from methanol, ethanol, chloroform, and acetic acid,respectively

Table 1 DSC data of rifapentine crystals

type of crystals untreated sampletreated sample using

methanol ethanol chloroform acetic acid

fusion temperature/°C 173.2 174.1 174.3 171.7 164.1

onset temperature/°C 170.3 171.5 169.8 167.6 156.7

enthalpy of fusion/(J$g–1) 12.18 12.05 9.13 8.84 28.42

68 Front. Chem. Eng. China 2010, 4(1): 65–69

by the SEM photographs, crystals obtained from aceticacid exhibit the most crystalline state and therefore themost stable and hence the least soluble form [12].

4 Conclusions

In conclusion, it can be said that the crystallization mediumhas major effect on rifapentine crystal habit. The crystalsunder different solvents showed significant changes in theshape, size, melting points, dissolution rate, XRD patterns,and DSC curves. This suggests that the newly developedcrystals of rifapentine under ambient conditions havedifferent crystalline modification facilitating significantlyimproved dissolution rate compared to untreated rifapen-tine. Therefore, it can be safely concluded that theimprovement obtained in the present study in the modifiedcrystals will give better bioavailability and better ther-apeutic activity clinically [13]. When the crystal wasobtained from acetic acid, the habit of produced rifapentinecrystals was greatly modified from needle to block. Thechange of crystal habit was originated from the crystalstructure modification.

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Fig. 5 Dissolution profile of untreated rifapentine and modifiedcrystals obtained using various solvents

Kun ZHOU et al. Crystal modification of rifapentine using different solvents 69