OA16386A

OA16386A - Hydrate of 1-{(2S)-2-amino-4-[2,4bis(trifluoromethyl)-5,8-di-hydropyrido[3,4d]pyrimidin-7(6H)-yl]-4-oxobutyl}-5,5-difluoropiperidin-2-one tartrate.

Info

Publication number: OA16386A
Application number: OA1201300164
Authority: OA
Inventors: Ki Sook Park; Jung Min Yun; Bong Chan Kim; Kyu Young Kim; Ji Hye Lee
Original assignee: Lg Life Sciences Ltd.
Priority date: 2010-11-01
Filing date: 2011-10-31
Publication date: 2015-10-07

Abstract

The present invention relates to 1.5 hydrate of 1-{(2S)-2-amino-4-[2,4-bis(trifluoromethyl)-5,8dihydropyrido[3,4-d]pyrimidin-7(6H)-yl]-4oxobutyl}-5,5-difluoropiperidin-2-one tartrate, a process for preparing the same, and a pharmaceutical composition for inhibiting DPP-IV, which comprises said compound as the active component.

Description

HYDRATE OF H(2S)-2-AMINO-4-[2,4-BIS(TRIFLUOROMETHYL)5,8-DI- HYDROPYRlDO[3,4-rf]PYRIMIDIN~7(6H)-YL]-4OXOBUTYL}-5,5-DIFLUORO-PIPERIDIN-2-ONETARTRATE

TECHNICAL FIELD

The présent invention relates to 1.5 hydrate of l-{(2S)-2- amino-4-[2,4bis(trifluoromethyl)-5,8-dihydropyrido[3,4-i/]pyrimidin-7(6H)-yl]-4-oxo-butyl}-5,5difluoropiperidin-2-one tartrate represented by the following formula l (hereinafter, referred to as “Compound l”), and a process for preparing the same.

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BACKGROUND ART

Compound l is disclosed in Korean Patent Application No. 10-2006-0029138, which is incorporated herein by reference in its entîrety, and exhibits superior inhibitory activity against dipeptidyl peptidase-IV (DPP IV) and thus is useful as an agent for treating diabètes. Typical examples of diseases caused by DPP IV can include, but are not limited to, diabètes, obesity, etc. Among diabètes, it is particularly useful for the treatment and prévention of type II diabètes. The term “treatment” as used above means that when the compound is used for individuals manifesting symptoms of a disease, it can interrupt or delay the progress of disease; and the term “prévention” as used above means that when the compound is used for individuals who do not manifest symptoms of a disease but have the risk of onset of disease, it can interrupt or delay the sign of disease.

The investigation of physical and chemical properties of a new drug îs necessary for efficient and successful development of the new medicine. Particularly, by studying the presence of polymorphs and pseudopolymorphs of the drug and différences in physical and chemical properties between respective polymorphs the préférable crystal fonn of the drug can be selected in view of the pharmaceutical aspect (Remington’s Pharmaceutics, Chapter 75 Preformulation); (Bym, S.R., Solid State Chemistry of Drugs, Academie Press, New York, 1982). When the polymorphs are présent in the solution, they are chemically identical, but in the solid state they respectîvely have definitely different X-ray diffraction patterns and exhibit différences in various physical and chemical properties. Particularly, respective polymorphs can have différences in bioavailability due to the différences in dissolution rates, and exhibit unexpected properties in the aspect of thermodynamic stabiiity.

When a certain drug is présent in the form of polymorphs, the crystal forms having different structures can be obtained depending on the conditions of recrystallization such as recrystallizing solvent, drug concentration, heating and cooling rates, température, stirring rate, and the like, during the procedures for preparing the drug. Therefore, in order to obtain the same crystal form a spécial attention is required for the management of manufacturing procedures.

Hydrates as one of pseudopolymorphs comprise a water molécule within the crystal of drug, and have a crystal form different from anhydrate. Différence in the crystal structure can be distinguished by X-ray diffraction pattern. Since in hydrates only physical properties such as crystallinity, hygroscopic property, melting point, solubility, dissolution rate, etc. are changed without any change of chemical properties providing pharmacological effects, they have a very important significance in the pharmaceutical aspect, like polymorphs (Morris, K. R. et al., Int. J. Pharm., 108, 1994, 15-206).

The knowledge which is understood up to date from various référencés relating to the technical field to which the présent invention belongs is that there is no general tendency, for example, to prefer the hydrate to the anhydrate or vice versa, for the improvement of pharmaceutical properties including drug stability, hygroscopic property, etc. Ultimately, détermination of the forms having the optimal pharmaceutical properties for respective compounds must be made by a person skilled in the relevant technical field through continuons study case by case.

Particularly, it can never be anticipated among any contemplable forms of a certain drug, i.e. free compound, sait, anhydrate and hydrate, which one can exhibit a stability with the hygroscopic property that is not changed depending on the surrounding humidity. Furthermore, among the hydrates the most stable hydration number cannot be predicted. And, even though the hydration number is the same, it is also unpredictable which crystal form would be the most stable. This is a phenomenon that is inconsi stent l y revealed since it cannot be anticipated and belongs to the experimental area which can be confirmed only through repeated experiments.

SUMMARY OF THE INVENTION

Thus, the présent inventors conducted intensive study to provide a stable polymorph or pseudopolymorph of Compound l. As a resuit, we hâve surprisingly found that 1.5 hydrate of tartrate sait of Compound l exhibits a superior stability against the change of relative humidity as compared to the anhydrate or other hydrates having a similar hydration number, and thus, completed the présent invention. Up to date, the crystal form of Compound l has never been publicly disclosed.

Therefore, the présent invention provides 1.5 hydrate of tartrate sait of Compound

l.

In addition, the présent invention provides a process for preparîng l .5 hydrate of tartrate sait of Compound l.

DISCLOSURE OF THE INVENTION

Since 1.5 hydrate of tartrate sait of Compound l obtained as a crystal form that can be developed according to the présent invention, is superior to the similar crystal foims in terms of a stability, particularly, a storage stability, it can be very advantageously used in preparing the pharmaceutical composition containing Compound l as an active component. That is, 1.5 hydrate according to the présent invention neither loses any water molécule in the crystal nor accepte water molécules any more in the broad range of relative humidity to maintain the water content thereof, and therefore, shows substantially no change of weight depending on the change of humidity. In case of unstable crystal forms, the water content thereof can vary with environments or additives during the storage and formulation process. For instance, in quantifying the standard material and the sample for the purpose of quantification, if the experiment is not conducted in a drying room, some experimental error may be caused to incur the problème of quality control. However, since in the l .5 hydrate according to the présent invention the water content does not sensitively vary with environments in the broad range of relative humidity, the product with a uniform standard can aiways be obtained during the storage and formulation process, and further, an error in the quality control is very small. As above, the 1.5 hydrate according to the présent invention shows a great advantage in terms of handling and quality control.

Further, 1.5 hydrate of tartrate sait of Compound l according to the présent invention does not show any change in the crystal form depending on the change of humidity. On the contrary, the présent inventors hâve identified that 0.5 hydrate and anhydrate of tartrate sait of Compound l absorb much water as the relative humidity is raised, to be converted into a more stable 1.5 hydrate according to the présent invention. It was also identified that even in the experiment for an accelerating stability (40°C/75% RH) the 0.5 hydrate is converted into the 1.5 hydrate after a lapse of some hours.

In addition, it is possible to control the préparation of I.5 hydrate of tartrate sait of Compound l according to various methods as provided in the présent invention.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure l represents the X-ray powder diffraction (XRD) spectrum of the crystal form I as the 1.5 hydrate oftartrate sait of Compound 1.

Figure 2 represents the Infrared spectroscopy (FT-IR) spectrum of the crystal form I as the 1.5 hydrate of tartrate sait of Compound 1.

Figure 3 represents the resuit of Differential scanning calorimetric (DSC) or Thermogravimetric (TG) analysis of the crystal form I as the 1.5 hydrate of tartrate sait of Compound 1.

Figure 4 represents the dynamic vapor adsorption/desorption isotherm of the crystal form I as the 1.5 hydrate of tartrate sait of Compound 1.

Figure 5 represents the X-ray powder diffraction (XRD) spectrum of the crystal form II as the 0.5 hydrate of tartrate sait of Compound 1.

Figure 6 represents the Infrared spectroscopy (FT-IR) spectrum of the crystal form II as the 0.5 hydrate of tartrate sait of Compound 1.

Figure 7 represents the resuit of Differential scanning calorimetric (DSC) or Thermogravimetric (TG) analysis of the crystal form II as the 0.5 hydrate of tartrate sait of Compound 1.

Figure 8 represents the dynamic vapor adsorption/desorption isotherm of the crystal form II as the 0.5 hydrate of tartrate sait of Compound 1.

Figure 9 represents the X-ray powder diffraction (XRD) spectrum of the crystal form II as the 0.5 hydrate of tartrate sait of Compound 1, during the course of moisture adsorptîon study.

Figure 10 represents the X-ray powder diffraction (XRD) spectrum of the crystal form III as the anhydrate of tartrate sait of Compound 1.

Figure 11 represents the Infrared spectroscopy (FT-IR) spectrum of the crystal form III as the anhydrate of tartrate sait of Compound 1.

Figure 12 represents the resuit of Differentiai scanning calorimetric (DSC) or Thermogravimetric (TG) analysis of the crystal form III as the anhydrate of tartrate sait of Compound l.

Figure 13 represents the dynamic vapor adsorption/desorption isotherm of the crystal form III as the anhydrate of tartrate sait of Compound 1.

Figure 14 represents the moisture adsorption curve ofthe crystal form III as the anhydrate of tartrate sait of Compound 1, with time lapse at normal température and normal humidity.

Figure 15 represents the X-ray powder diffraction (XRD) spectrum of the crystal form III as the anhydrate of tartrate sait of Compound 1, during the moisture adsorption study.

Figure 16 represents the X-ray powder diffraction (XRD) spectrum obtained after conducting the experiment of Test Example 8, indicating XRD of the crystal form II, XRD of the sample of the crystal form II after 8-week storage at 60°C/5% RH, XRD of the sample of the crystal form II after 8-week storage at 40°C/75% RH, and XRD of the crystal form I, in the order from the bottom.

BEST MODE FOR CARRYING OUT THE INVENTION

As identified to respectively hâve the characteristic crystal forms according to the présent invention, in the présent spécification the 1,5 hydrate of tartrate sait of Compound 1 is designated as “the crystal form I”; the 0.5 hydrate is designated as “the crystal form II”; and the anhydrate is designated as “the crystal form III”. Herein, the water content of

1.5 hydrate is 3.0-5.5%, that of 0.5 hydrate is 1.0~2.5%, and that of anhydrate is 0-1.0%.

The crystallinities of the crystal forms I, II and III are different from each other as can be identified from X-ray diffraction diagrams shown in Figures 1, 5 and 10.

As a resuit from the analysis by Differentiai Scanning Calorimetry for the crystal ' form I, the endothermie peak occurring with releasing water contained in the crystal lattice at 90~l30°C has a broad section in which the melting point is included, and the endothermie peak occurs one more time in the section of13O~16O°C with undergoing the chemical déhydration of tartrate sait of Compound l. In addition, the thermogravimetric 5 analysis shows the weight loss of about 4.0% that is équivalent to the water contained in the lattice in the first endothermie section of differential scanning calorimetry and the weight loss of about 2.5% that is équivalent to the water removed by déhydration from the structure of tartrate sait of Compound l in the second endothermie section (Figure 3). The analysis for the crystal forms II and III by Differential Scanning Calorimetry shows 10 three endothermie points. Specifically, the first endothermie point occurs with releasing water contained in the lattice, the second endothermie point occurs with the melting point, and the third endothermie point occurs via chemical déhydration reaction. Further, according to the thermogravimetric analysis the first endothermie section shows the weight loss that is équivalent to the water contained in the lattice (about l.3% in the 15 crystal form II, and about 0.6% in the crystal form III), and the third endothermie section shows the weight loss of about 2.4% that is équivalent to the water removed by déhydration (Figures 7 and 12). Since the water content in the lattice of tartrate sait of Compound l as above was consistent with the water content quantified by Karl-Fischer method (Mettler Toledo DL37 KF Coulometer), it was proved that the endothermie peaks 20 were caused by évaporation of water molécules.

Hereinafter, we intend to explain that the crystal form I according to the présent invention has unexpected superior characteristics from any aspect in comparison with other crystal form II or III by more specifically comparing the characteristics of respective crystal forms.

Crystal Form I

The présent invention relates to the crystal form of 1.5 hydrate of tartrate sait of

Compound 1. As a resuit of the analysis of the properties thereof the crystal form I exhibîts the following characteristics:

(a) The water content of the crystal form I is in the range of 3.0-5.5%.

(b) The characteristic peak values (20) of XRD spectrum measured at CuKa, 40kV, 30mA are 15, 18, 20, 21 and 23°.

(c) Infrared (IR) spectrum shows a characteristic absorbance at about 3591, 3401, 3128, 1712,1655, 1636, 1229, 1205, 1129 and 1058 cm’¹.

(d) Differential scanning calorimetry spectrum shows the endo-thermal peaks in two broad température ranges of about 90-130°C and 130~160°C.

(e) The water content measured by Karl-Fisher method îs about 4.0%.

(f) When the température is raised from 25°C to 250°C, the weight loss of about 4.0% and about 2.5% occur in the range of 70-110°C and 140-170°C, respectively (This is the resuit obtained from TG of Figure 3. Since energy change occurs slightly later than the weight change, there is generally a différence between the results of DSC and TG.).

(g) Tartrate sait of Compound 1 can be crystallized from water, acetonitrile /water, ethanol/water, ethanol/hexane or ethyl acetate/hexane solvent, and preferably crystallized from water.

(h) Crystal form I can be prepared by subjecting the crystal form II or III to moisture absorption.

(i) The weight change is absent or, if any, 0.8% or less depending on the change of extemal humidity in the range of 5-95% RH, and the crystal form is not changed with the change of humidity.

Crystal Form II

In order to find out other crystal forms than the crystal form I the présent inventors prepared 0.5 hydrate of tartrate sait of Compound 1 (crystal form II). Although the 0.5 hydrate is the same tartrate sait of the same compound and has a similar hydration number, it shows unstable storage stability as follows, in comparison with the I.5 hydrate (crystal form I).

(a) The water content ofthe crystal form II is in the range of 1.0-2.5%.

(b) The characterîstîc peak values (20) of XRD spectrum measured at CuKa, 40kV, 30mA are 14, 15, 17, 18, 19, 21 and 23°.

(c) Infrared (IR) spectrum shows a characteristic absorbance at about 3455, 2891, 1721, 1655, 1571, 1228, 1209, 1131, 1086 and 1059 cm'¹.

(d) Differential scanning caiorimetry spectrum shows the endo-thermal peaks in three broad température ranges of about 80~l 15°C, 115~135°C and I35-173°C, and the melting point at about 117°C.

(e) The water content measured by Karl-Fisher method is about 2.0%.

(f) When the température is raised from 25°C to 250°C, the weight loss of about 1.3% and about 2.4% occur in the range of 70-104°C and 137-168°C, respectively.

(g) The weight change of 4.0% or more occurs over the change of external humidity in the range of 5-95% RH and moisture is rapidly absorbed from 45% RH, giving a weight increase of 3.7% at 75% RH. If a total of 4.0% or more moisture is contained, the crystal form II is converted into the crystal form I. It has been shown that the crystal form II is converted into the crystal form 1 from about 60% RH (see Figures 8 and 9). That is, the crystal form varies with the change of humidity to reach the more stable crystal form I. In addition, it is converted into the crystal form I within 8 weeks in the accelerating (40°C/75 %RH) stability test.

Crystal Form III

In order to fïnd out other crystal forms than the crystal form I the présent inventors prepared anhydrate of tartrate sait of Compound 1 (crystal form III). The ' anhydrate shows unstable storage stability as follows, in comparison with the 1.5 hydrate (crystal form I).

(a) The water content of the crystal form Π1 is in the range of 0-1.0%.

(b) The characteristic peak values (20) of XRD spectrum measured at CuKa, 40kV, 30mA are 6, 17, 21, 23, 24, 26 and 30°.

(c) Infrared (IR) spectrum shows a characteristic absorbance at about 3470, 3187, 2940, 1640, 1570, 1229, 1206, 1130 and 1056 cin¹.

(d) Differential scanning calorîmetry spectrum shows the endo-thermal peaks in three broad température ranges of about 65-100°C, 100-130°C and 132~170°C, and the melting point at about 104°C.

(e) The water content measured by Karl-Fisher method is about 0.1%.

(f) When the température is raised from 25°C to 250°C, the weight loss of about 0.6% and about 2.4% occur in the range of 62-110°C and 120-173°C, respectively.

(g) The weight change of 3.5% or more occurs over the change of extemal humidity in the range of 5-95% RH and moisture is rapidly absorbed from 5% RH, so that the crystal form III is converted into the ciystal form I from about 15% RH (see Figures 13, 14 and 15). That is, the crystal form varies with the change of humidity to arrive at the more stable crystal form I.

The above results suggest that in the range of relative humidity under which the formulation is conventionally practiced the crystal forms II and III are unstable and thus automatîcally converted into the crystal form I. Such tendency is shown more remarkably in case of the crystal form III,

1.5 Hydrate of tartrate sait of Compound 1 according to the présent invention exhibits a strong DPP-IV inhibitory activity in the same way as the corresponding free base disclosed in Korean Patent Application No. 10-2006- 0029138. In addition, the 1.5 hydrate of the présent invention shows improved physical and chemical properties as compared to the crystal forms having other hydration state. Therefore, the 1.5 hydrate according to the présent invention is considerably easy to handle, quality control and formulate as compared to the crystal forms having other hydration state.

The 1.5 hydrate according to the présent invention has the DPP-IV inhibitory activity as mentioned above, and therefore, can be formulated for convenient administration in the pharmaceutical and veterinary field. Formulation can be conducted according to the techniques and methods known in the art in relation to other formulations having DPP-IV inhibitory activity, particularly with reference to the disclosure of Korean Patent Application No. 10-2006-0029138 which is incorporated herein by reference in its entirety.

Therefore, a pharmaceutical composition for inhibiting DPP-IV, comprising the

1.5 hydrate according to the présent invention as the active component together with a pharmaceutically acceptable carrier, is covered by the scope of the présent invention. The composition according to the présent invention is characterized in that it is used particularly for the treatment and prévention of diabètes or obesity.

The présent invention is illustrated more in detail by means of the following Examples and Test Examples. However, the following Examples and Test Examples are provided only to assist the understanding of the présent invention but it is not intended that the scope of the présent invention Îs limited in any manner by these Examples and 20 Test Examples.

Example 1

Préparation of 1.5 hydrate of tartrate sait of Compound 1 (crystal form I)

Compound 2 Tartrate sait of Compound l

1.87 Kg of the compound 2 was dissolved in about 9 L of éthanol. 0.94 Kg of SOCl₂ was added at O-lOü and then stirred while maintaining low température. After concentrating under reduced pressure, the concentrate was dissolved in 11.2 L of MTBE (methyl t-butyl ether), and the resulting mixture was adjusted with 10 N NaOH solution to pH 7-8. After separating the layers, the aqueous layer was extracted with about 3.7 L of MTBE and twice with 3.7 L of MTBE, and then concentrated under reduced pressure. The resulting brown turbid solution was dissolved in 12 L of éthanol, 0.47 kg of Ltartaric acid dissolved in about l.5 Lof water was added thereto, and then stirred for l hour. The resulting crystalline slurry was filtered, washed with water and éthanol ( l :8), and then dried to obtain 1.13 kg (yield 97.5%) of the title compound.

'H NMR (500 MHz, CD₃OD) δ 2.38 (m, 2H), 2.59 (m, 2H), 2.82 - 2.99 (m, 2H), 3.H (bt, lH),3.2l (bt, IH), 3.50 - 3.55 (m, IH), 3.72 - 3.9i (m, 5H), 3.98 (t, J=5.2 Hz, IH), 4.38 (s, 2H), 4.97 - 5.00 (m, 2H).

Example 2

Recrystallization of 1.5 hydrate of tartrate sait of Compound l (crystal form I) from water g of tartrate sait of Compound l obtained from Example l was added to

250-500 ml of water, and dissolved in water while adjusting the solution with ΙΟ N

NaOH to pH 6~7. IL7 g of L-tartaric acid dissolved in 23.5 ml of water was added, and

I2 crystals were obtained with varying the température, stirring rate and stirring time as shown in the following Table l. Then, the crystals were filtered and dried to obtaîn the crystal form I. The stirring rate was varied in the range of 50-400 rpm, and the température was varied in the range of 5-32□. The volume of water used for 5 recrystallization, the stirring rate, température and stirring time are represented in the following Table 1.

[Table 1 ]

Entry	Volume (mL)	RPM	Température	Time (h)	Yield (%)
1	250	50	25.0	0.50	96.0 (48.0g)
2	250	100	25.0	1.00	92.8 (46.4g)
3	250	400	25.0	5,00	93.6 (46.8g)
4	375	400	25.0	1.00	92.4 (46.2g)
5	500	50	25.0	5.00	46.6 (23.3g)
6	500	400	25.0	3.00	88.2 (44.1g)
7	375	225	31.8	2.75	94.0 (47.0g)
8	375	225	15.0	2.75	91.0 (45.5g)
9	250	50	5.0	0.50	81.4 (40.7g)
10	250	400	5.0	5.00	101.8 (50.9g)

Example 3

Recrystallization of 1.5 hydrate of tartrate sait of Compound 1 (crystal form I) from the mixed solvent g of tartrate sait of Compound 1 was dissolved in 25-60 ml of solvent mixtures comprised of water and acetonitrile in different ratios. Crystals were precipitated with varying the température in the presence or absence of stirring, filtered and dried to recrystallize the tartrate sait of Compound l. The conditions used for recrystallization are listed in the following Table 2.

[Table 2]

Entry	Acetonitrile/water	Stirring	Température (□)
1	4/1	N	25.0
3	6/1	N	25.0
4	6/1	N	0.0
5	8/1	N	25.0
6	8/1	Y	25.0
8	1/1	N	25.0
9	1/1	Y	20.0

Test Example 1

Powder X-ray Diffractometry

About 20 mg of the sample was charged in the sample holder and mounted on

Powder X-ray diffractometer to obtain the diffraction pattern in the range of 3~40°/20.

The diffraction patterns as obtained are attached to the présent spécification as Figures 1, and 10, respectively. Spécifie conditions for analysis are as follows.

Instrument: Bruker 4D Endeavor

Time per step: 0.3 s

Stepsize: 0.03 ⁰

Scan Mode: step

Voltage/ Current : 40 kV /30 mA

Cu-target (Ni-filter)

Divergence slit: 0.3

Detector: PSD: LynxEye

Instrument: Philips X-ray Generator (PW1710)

Time per step: 0.5 s

Stepsize: 0.03°

Scan Mode: step

Voltage/ Current : 40 kV /30 mA

Cu-target (Ni-filter)

Source Slit: 1.0 mm

Detector Siits: 0.15 mm, 1.0 mm

Test Example 2

Infrared Spectroscopy

The infrared spectra for respective crystal forms according to the présent invention were obtained using ZASCO FT-IR 4200 provided with DTGS detector. The resolution of respective spectra was 4cm*¹, and the number of scan was 16. in the présent Test Example 1~2 mg of the sample was placed on the accessory of ATR (Attenuated Total Réflectance) and the equipment was operated to obtain the spectrum. Background data was obtained by operating the equipment without any material in ATR. The spectra 15 thus obtaîned are attached to the présent spécification as Figures 2, 6 and 11, respectively.

Test Example 3

Differential Scanning Calorimetry (PSC)

Differential Scanning Calorimetry (DSC) was conducted using Mettler Toledo’s DSC82l^c. About 2-3 mg of the sample was charged into the aluminum pan, and the weight thereof was accurately recorded. The pan was covered with a lid through which a hole was punched. The pan was mounted on the equipment and heated from 25 to 250D in the rate of lOÛ/mîn under nitrogen purge. Indium métal was used as the standard for calibration. The spectra thus obtaîned are attached to the présent spécification as Figures 3, 7 and 12, respectively.

Test Example 4

Thermogravimetry (TG)

Thermogravimetry (TG) was conducted using Mettler Toledo TGA850. About 4—5 mg of the sample was charged into the aluminum pan. The pan was mounted on the equipment and then heated from 25 to 250ü in the rate of lOD/min under nitrogen purge. Nickel and Aluminum™ were used as the standard for calibration. The results of TG analysis as obtaîned are attached to the présent spécification as Figures 3, 7 and 12, respectively, together with the results of DSC analysis according to said Test Example 3.

Test Example 5

Dynamic Vapor adsorption/desorption analysis

The dynamic vapor adsorption/desorption data were collected on VTI-SA Vapor

Sorption Analyzer. While maintaining 25□, the vapor adsorption and desorption were repeated three times at intervals of 5% RH in the range of relative humidity (RH) 5-95%. The samples were not dried prior to the analysis. The equilibrium standard used for analysis was that the weight change within 2 minutes is less than 0.01%.

As the resuit of analysis, Figure 4 shows a resuit of the vapor adsorption/desorption isotherm of the crystal form I, from which it can be seen that the crystal form I shows the weight change of 0.8% or less according to the change of extemal humidity in the range of 5-95% RH. That is, the crystal form I according to the présent invention is very stable against the change of relative humidity. Figure 8 shows a resuit of the vapor adsorption/desorption isotherm of the crystal form II, from which it can be seen that in the crystal form II the weight increases by 4.3% when the relative humidity is raised up to 95% in the initiai moisture adsorption test. After moisture desorption, the crystal form II gives the same graph of moisture behavior as the crystal form I. From such resuit, it could be confirmed that the crystal form II was converted into the crystal form I during the course of the initial moisture adsoiption. Figure 13 shows a resuit of the vapor adsorption/desorption isotherm of the crystal form III, from which it can be seen that in the crystal form III the weight increases by 3.6% when the relative humidity is raised up to 95%. Furthermore, it can be seen that decreasing the humidity causes the moisture desorption of 0.6% or less so that the crystal form III becomes to contain the same moisture content as the crystal form I.

Test Example 6

Powder X-ray diffraction test of the crystal form II during moisture adsorption

About 50 mg of the crystal form II was placed in a glass vial, put under the relative humidity of 11%, 32%, 53%, 64%, 78% and 97%, respectively, for two or more days to induce the moisture adsorption, and then subjected to the powder X-ray diffraction test according to the conditions as presented in said Test Example 1 to identify any change of the crystal form at the time of moisture adsorption (see Figure 9). w

Respective relative humidity was provided by making the saturated aqueous solution of sait as shown in the following Table 3 and placing the solution in a desiccator, which was then sealed.

[Table 3]

Relative humidity 11%	Saturated aqueous solution of LiCl
Relative humidity 32%	Saturated aqueous solution of MgCl₂
Relative humidity 53%	Saturated aqueous solution of Mg(NO3)₂-6H₂O
Relative humidity 64%	Saturated aqueous solution of NaNO₂
Relative humidity 78%	Saturated aqueous solution of NaCI
Relative humidity 97%	Saturated aqueous solution of K.NO3

Test Example 7

Powder X-ray diffraction test of the crystal form III during moisture adsorption

The crystal form III was placed on a XRD holder and subjected to XRD test over the time (after 30 minutes, l hour and 5 hours) while allowing the moisture adsorption at room température under atmosphère (see Figure 15). In addition, the weight change at room température under atmosphère was recorded by times to obtain a graph (see Figure 14). It could be known that the crystal form III rapidly absorbs water so that it is converted into the crystal form I within about one hour.

Test Exemple 8

Thermal stability of the crystal form I and the crystal form II

About 50 mg of each of the crystal form I and the crystal form II was placed in

Duma bottle, and then kept at 40±2D, 75±5% RH or 60±2D, 5±5% RH. After 2 weeks, 4 weeks and 8 weeks, each sample was removed from Duma bottie and analyzed by XRD for identifying any change of the crystal form and HPLC for Îdentifying the stability. For HPLC analysis the sample was dissolved in a mixture of acetonitrile/water/trifluoroacetîc acid = 30/70/0.1 (v/v/v) and then subjected to the analysis. The conditions for HPLC analysis are as follows:

Conditions for HPLC analysis

Column: Atlantis dCl8 (4.6 mm I.D x 250 mm L, Particle Size 5/zni, Waters)

Column Température: 10D

Mobile phase:

Mobile phase A: MeCN/TFA= 100/0.1 (v/v)

Mobile phase B: H₂O/TFA = 100/0.1 (v/v)

Gradient condition:

Time (min.)	A(%)	B (%)
Initial	38	62
25	38	62
35	80	20
40	38	62
55	38	62

Flow rate: 0.7 ml/min.

Détection: 256 nm, UV

Injection volume: 10/zC

Total analysis time: 55 min.

The results of the stability for the crystal form I and the crystal form II are shown in the following Table 4.

[Table 4]

Results of the thermal stability for the crystal form I and the crystal form II

Time (week)	40ü/75% RH	60D/5% RH
	Crystal form I	Crystal form II	Crystal form I	Crystal form II
0	99.4	98.4	99.4	98.4
2	99.3	98.4	98.8	98.1
4	99.2	98.3	98.6	97.8
8	99.2	98.2	98.4	97.5

As shown in Table 4, it could be confirmed that upon keeping the crystal form I and the crystal form II at 40±2EJ, 75±5% RH or 60±2D, 5±5% RH they exhibit a superior 10 stability up to 8 weeks. However, according to the resuit of XRD analysis the crystal form I did not show any change up to 8 weeks, but the crystal form II was converted into the crystal form 1 at 8 week under the condition of 40°C/75% RH (see Figure 16).

Test Example 9

Détermination of density of the crystal form I and the crystal form II

About 20-30 ml of each of the crystal form I and the crystal form II was introduced into a 50 ml measuring cylinder, Here, the samples were slowly introduced so that they are not closely contacted with each other. After reading a scale on the measuring cylinder, the weight thereof was measured to calculate the bulk-density. After measuring the bulk-density, the measuring cylinder containing the sample was put on ER.WEKA densitometer and then subjected to impulses 250 times to measure the tapdensity (Table 5). The resuit of measurement shows that both the bulk-density and the tap-density of the crystal form l were higher than those of the crystal form II. The Carr index was calculated from the bulk-density and the tap-density. As a resuit, it was lower in the crystal form I than in the crystal form IL Since it is generally understood that the lower the Carr index, the better the flowability, it could be identified by comparing the Carr indexes that the crystal form I can be easily handled in processing procedures including tabletting as compared to the crystal form II.

[Table 5]

Détermination of density and particle size of the crystal form l and the crystal form II

Form	Density
Bulk-density Tap-density	Carr index
I	0.45	0.54	16.22
II	0.34	0.44	23.47

Claims

1. 1.5 Hydrate of l-{(2S)-2-amino-4-[2,4-bis(trifluoromethyl)-5,8-dihydropyrido[3,4-f/]pyrimidin-7(6H)-yl]-4-oxobutyl}-5,5-difluoropiperidin-2-one tartrate sait having a water content in the range of 3.5-5.5%.

2. The 1.5 hydrate according to Claim l, wherein characteristic peak values (2Θ) in XRD diffractogram are 15, 18, 20, 21 and 23°.

3. A process for preparing the 1.5 hydrate according to Claim 1, characterized in that l-{(2S)-2-amino-4-[2,4-bis(trifiuoromethyl)-5,8-dihydropyrido[3,4-i/]-pyrimidin- 7(6H)yl]-4-oxobutyl)-5,5-difluoropiperïdin-2-one tartrate sait is recrystallized from water, acetonitrile/water, ethanol/water, ethanol/hexane or ethyl acetate/hexane solvent.

4. The process according to Claim 3, wherein the recrystallizing solvent is water.

5. A pharmaceutical composition for inhibiting DPP-IV, which comprises the 1.5 hydrate according to Claim 1 as the active component together with a pharmaceutically acceptable carrier.

6. The composition according to Claim 5, wherein it is used for the treatment or prévention of diabètes or obesity, sx/ Λ

LG Llfe Sciences Ltd.