5-BROMO-2-(alpha-HYDROXYPENTYL)BENZOIC ACID SODIUM SALTS IN DIFFERENT CRYSTAL FORMS, AND PREPARATION METHOD THEREOF
20180127349 ยท 2018-05-10
Assignee
Inventors
Cpc classification
C07C65/03
CHEMISTRY; METALLURGY
C07C51/60
CHEMISTRY; METALLURGY
C07C65/03
CHEMISTRY; METALLURGY
A61P9/10
HUMAN NECESSITIES
International classification
C07C65/03
CHEMISTRY; METALLURGY
Abstract
The present invention discloses 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt in different crystal forms and the preparation methods thereof, and belongs to the field of pharmaceutical chemistry. Said different crystal forms of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt include: amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt, crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt, and crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt. The different crystal forms of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt obtained according to the present invention have better stability and water-solubility than the mixed forms of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt, thus is advantageous for pharmaceutical use. Moreover, the different crystal forms of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt possess much better therapeutic effect than 5-bromo-2-(-hydroxypentyl)benzoic acid potassium salt.
Claims
1. Crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt, defined by a XRPD pattern with the diffraction peaks at 20 values of 5.60, 16.46, 16.78, 18.77, 21.45, 32.28, 33.30, 33.49, 34.12, 36.14, 36.61, 37.87, 40.24, 41.32, 43.76, 44.92, 45.18, 47.23, 55.12, 56.73, 57.12, 62.78, wherein the error range of 20 value is 0.2.
2. Crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt, defined by a XRPD pattern with the diffraction peaks at 20 values of 5.72, 6.69, 9.93, 11.09, 11.84, 14.55, 16.42, 17.27, 17.80, 18.28, 19.86, 20.98, 22.21, 23.43, 23.88, wherein the error range of 2 value is 0.2.
3. A method for preparing amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt, the method comprising: placing 6-bromo-3-butyl-1(3H)-isobenzofuranone and sodium hydroxide in a crystallizer to form a mixture, adding tetrahydrofuran and water solvents to the mixture; and reacting the mixture containing the solvents under a water bath at 60 C. to form a reaction mixture; wherein after the reaction is complete, the reaction mixture is distilled under reduced pressure to remove the tetrahydrofuran and water solvents therefrom and to yield amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt.
4. A method for preparing the crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 1, the method comprising: adding a solution of NaOH in methanol to amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid to form a mixture; dissolving the amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid within the mixture under ultrasonic conditions; and volatilizing the methanol solvent slowly to precipitate solids from the mixture; wherein after the solids are precipitated, the solids are dried under a vacuum.
5. A method for preparing the crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 2, the method comprising: dissolving amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid in a tetrahydrofuran solvent, stirring the resulting solution at room temperature, and adding an aqueous solution of sodium hydroxide dropwise to the solution; wherein after evaporation of the tetrahydrofuran and water solvents, the resulting solids are dried under a vacuum.
6. A method for preparing the crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 2, the method comprising: weighing and placing 6-bromo-3-butyl-1(3H)-isobenzofuranone and sodium hydroxide in a flask, adding methanol to the flask, heating the resulting mixture under reflux to react the contents, evaporating and removing the methanol, adding ethyl acetate, shaking the resulting mixture and filtering and removing the undissolved solids, evaporating and removing the ethyl acetate to yield a solid compound, adding anhydrous diethyl ether to dissolve the solid compound, and filtering the resulting solution after standing overnight.
7. A method of using the amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt prepared in accordance with claim 3 to reduce brain tissue injury induced by obstruction of cerebral arteries.
8. A method of using the amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt prepared in accordance with claim 3 to reduce brain infarction volume.
9. A method of using the amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt prepared in accordance with claim 3 to reduce brain edema volume.
10. A method of using the crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 1 to reduce brain tissue injury induced by obstruction of cerebral arteries.
11. A method of using the crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 1 to reduce brain infarction volume.
12. A method of using the crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 1 to reduce brain edema volume.
13. A method of using the crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 2 to reduce brain tissue injury induced by obstruction of cerebral arteries.
14. A method of using the crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 2 to reduce brain infarction volume.
15. A method of using the crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 2 to reduce brain edema volume.
16. A method of using the amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt prepared in accordance with claim 3 to prevent and treat heart and brain ischemic diseases, to improve heart and brain circulatory disorders, or to produce antithrombotic effect.
17. A method of using the crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 1 to prevent and treat heart and brain ischemic diseases, to improve heart and brain circulatory disorders, or to produce antithrombotic effect.
18. A method of using the crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt of claim 2 to prevent and treat heart and brain ischemic diseases, to improve heart and brain circulatory disorders, or to produce antithrombotic effect.
Description
DESCRIPTION OF FIGURES
[0017]
[0018]
[0019]
[0020]
[0021]
[0022]
EXAMPLES
[0023] The following examples are used to help a skilled person in the art understand the present invention more deeply, but are not intended to limit the present invention in any ways.
Example 1: Preparation of Amorphous 5-bromo-2-(-hydroxypentyl)benzoic Acid Sodium Salt
[0024] 3.5 g of 6-bromo-3-butyl-1(3H)-isobenzofuranone and 520 mg of sodium hydroxide were weighed and placed in a crystallizer, to which 40 ml tetrahydrofuran and 8 ml water were added, then the mixture was dissolved and reacted under a water bath at 60 C. After 3 hours, the reaction mixture was distilled under reduced pressure at 60 C. to remove all the solvents, to yield amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt.
Example 2: Preparation of Crystal Form a of 5-bromo-2-(-hydroxypentyl)benzoic Acid Sodium Salt
[0025] 350 mg of amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid was weighed, to which 2 ml solution of NaOH in methanol was added, dissolved under ultrasound, and then the solvent was volatilized slowly for 14 days. Solids were precipitated. The solids were dried under vacuum to yield crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt.
Example 3: Preparation of Crystal Form B of 5-bromo-2-(-hydroxypentyl)benzoic Acid Sodium Salt (Method I)
[0026] 1.15 g of amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid was weighed and placed in a 20-ml sample bottle with magnetically stirring at room temperature, to which 1 ml tetrahydrofuran was added. 120 mg of sodium hydroxide was weighed and dissolved in 8 ml water, and then the NaOH solution was slowly added into the above-mentioned solution dropwise. The resulting solution was subjected to rotary evaporation at 50 C. Solids were obtained, and dried under vacuum at room temperature to yield crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt.
Example 4: Preparation of Crystal Form B of 5-bromo-2-(-hydroxypentyl)benzoic Acid Sodium Salt (Method II)
[0027] 20 g of 6-bromo-3-butyl-1(3H)-isobenzofuranone and 4.46 g of sodium hydroxide were weighed and placed in a 500-ml round bottom flask. 300 ml methanol was added to the flask, and the resulting mixture was heated under reflux to react. Then methanol was evaporated off. 200 ml ethyl acetate was added. After the resulting mixture was shook adequately, the undissolved solids were filtrated off. Ethyl acetate was evaporated off to yield viscous white solid compound, and 300 ml anhydrous diethyl ether was added to dissolve. The resulting solution was maintained standing overnight and a large amount of white solid precipitated. The mixture was filtrated to yield crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt.
Example 5: Effect of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt on Brain Infarction Volume and Brain Edema Volume in Rats with Local Brain Ischemia
(1) Experimental Materials and Methods
[0028] Wistar rats (50% of rats is female and another 50% of rats is male); 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP, in the form of amorphous form, crystal form A and crystal form B, prepared according to the present invention), prepared in double distilled water; TTC is in the form of white powder, manufactured by Beijing Chemical Works.
[0029] Administration method: 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP) was intravenously injected at one hour before MCAO, with the dosage of 12 mg/kg; 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP) was used to intragastric administration at one hour before MCAO, with the dosage of 20 mg/kg. 2-(-hydroxypentyl)benzoic acid potassium salt (NBPK) was used as positive control, and its dosage (calculated in moles) is the same as that of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt, that is, the dosage of intravenous injection is 9.6 mg/kg, and the dosage of intragastric administration is 15.9 mg/kg.
[0030] Preparation of MCAO model: Establish a focal brain ischemia-reperfusion injury model by cerebral artery obstruction in rats with suture-occluded method. Rats were intraperitoneally injected with 10% chloral hydrate to anesthetize. Rats were fixed on back and the skin of neck center was incised. The left common carotid artery, external carotid artery, and internal carotid artery were subjected to blunt dissection. The pterygopalatine artery was ligated, only remaining cranial trunk of internal carotid artery; a prepared nylon rope (burning diameter 0.3 mm) was inserted via external carotid artery incision into the bifurcation of the jugular vein, and was pushed along the cranial direction of the internal carotid artery. Using the place of the bifurcation as starting point, after the rope was pushed forward about 17 mm, hand can feel force of resistance, indicating that the head section of nylon rope has been through the beginning of the middle cerebral artery. This indicates one side of the middle cerebral artery occlusion model was completed. Sutured the incision, remained the end of nylon line out of the skin. For reperfusion, the nylon line was gently pulled out. If feel force of resistance, this indicates the end of nylon line has returned to the trunk of external carotid artery, thereby accomplishing reperfusion.
[0031] Determination of infarction volume: After being subjected to ischemia for 2 hours and reperfusion for 24 hours, the heads of rats were cut off. Forebrain was removed and cut into six pieces, each piece being 2 mm. The pieces were stained with TTC, and incubated at 37 C. for 30 minutes. Normal issues were stained to be red, but the infarcted issues were stained to be white. Took photos with digital camera, and calculated the percentage of the infarcted area relative to the spherical area of brain by Photoshop.
(2) Results
[0032] Experimental results of model group: brain infarction volume was 53.424.65%, and brain edema volume was 13.663.46%.
[0033] The effect of intravenous injection and intragastric administration of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP) on brain infarction volume in rat MCAO model and the effect of intravenous injection and intragastric administration of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP) on brain edema volume in rat MCAO model were as follows:
[0034] Effect of intravenous injection of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP) on brain infarction volume in rat MCAO model
TABLE-US-00001 Inhibition of Infarction infarction Dosage volume volume Drug (mg/kg) n (%) (%) Pseudo-surgery 0 0 group Model group 10 53.42 4.65.sup. BZP (amorphous 12 10 3.46 2.94 aa 93.52 form) BZP (crystal form 12 10 4.54 1.73 aa 91.50 A) BZP (crystal form 12 10 4.76 1.45 aa 91.09 B) NBP-K 9.6 10 24.35 4.32 aa 54.42 aa: P < 0.01, as compared with the model group; ** P < 0.01, as compared with the NBP-K group.
[0035] Effect of intravenous injection of 5-bromo-2-((-hydroxypentyl)benzoic acid sodium salt (BZP) on brain edema in rat MCAO model
TABLE-US-00002 Inhibition Dosage Edema of edema Drug (mg/kg) n (%) (%) Pseudo-surgery 0 0 group Model group 10 13.66 3.46 BZP (amorphous 12 10 .sup.2.56 1.63 aa 88.58 form) BZP (crystal form 12 10 .sup.3.24 4.11 aa * 81.28 A) BZP (crystal form 12 10 4.20 2.03a 78.71 B) NBP-K 12 10 12.38 1.29 9.37 aa: P < 0.01, as compared with the model group; * P < 0.05, as compared with the NBP-K group; ** P < 0.01, as compared with the NBP-K group.
[0036] Effect of intragastric administration of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP) on brain infarction volume in rat MCAO model
TABLE-US-00003 Inhibition of Infarction infarction Dosage volume volume Drug (mg/kg) n (%) (%) Pseudo-surgery 0 0 group Model group 10 53.42 4.65.sup. 0 BZP (amorphous 20 10 5.59 2.35.sup.aa 89.05 form) BZP (crystal form 20 10 7.13 2.98.sup.aa 86.65 A) BZP (crystal form 20 10 7.13 2.98.sup.aa** 86.03 B) NBP-K 15.9 10 22.28 2.65 .sup.aa 58.3 .sup.aaP < 0.01, as compared with the model group; **P < 0.01, as compared with the NBP-K group.
[0037] Effect of intragastric administration of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP) on brain edema in rat MCAO model
TABLE-US-00004 Inhibition Dosage Edema of edema Drug (mg/kg) n (%) (%) Pseudo-surgery 0 0 group Model group 10 13.66 3.46.sup. BZP (amorphous 20 10 2.24 1.66.sup.aa** 88.36 form) BZP (crystal form 20 10 2.7 1.30.sup.aa** 87.55 A) BZP (crystal form 20 10 2.87 1.56 .sup.aa** 85.09 B) NBP-K 15.9 10 11.04 1.48 .sup.aa** 42.65 .sup.aa P < 0.01, as compared with the model group; * P < 0.05, as compared with the NBP-K group; **P < 0.01, as compared with the NBP-K group.
(3) Conclusion
[0038] 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt can obviously reduce brain tissue injury induced by obstruction of cerebral arteries, reduce brain infarction volume and reduce brain edema volume in rats. At the same dosage, the activity of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt is obviously better than 2-(-hydroxypentyl)benzoic acid potassium salt, and the activity of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt is also better than butylphthalide and halogenated 2-benzo[c]furanone known in the art.
Example 6: Results of Primary Toxicity Study of 5-bromo-2-(-hydroxypentyl)benzoic Acid Sodium Salt
[0039] Data of acute toxicity and sub-acute toxicity of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt BZP (in amorphous form)
TABLE-US-00005 BZP NBP-K Items ig iv ig iv LD50 Mouse 927 323 880 303 (mg/Kg) Rat 726 246 623 230 Sub-acute Rat >20 >12 >20 >12 (mg/Kg) Note: ig 20 mg/kg; iv 12 mg/kg.
Example 7: Sub-Acute Toxic Effect of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (in amorphous form) on Liver and Kidney Functions in Rats
[0040]
TABLE-US-00006 Groups Control ig iv Items Sex group (20 mg/kg) (12 mg/kg) ALT F 43.38 6.69 35.10 8.61 38.00 9.82 AST F 219.40 55.38 168.00 17.19* 202.40 68.12 M 5.30 0.87 4.92 0.75 5.34 1.33 AST/ALT F 246.20 129.61 135.29 48.13* 184.80 68.59 M 4.35 0.05 2.74 1.62* 3.96 1.20 Total Proteins F 48.52 2.46 46.63 1.07 49.00 0.79 (TP) M 41.70 2.04 40.26 15.78 53.40 8.69* Albumin F 33.52 1.98 32.43 1.14 33.96 1.71 (ALB) M 27.74 1.38 28.98 7.34 36.90 6.72* Total Bilirubin F 2.58 0.63 2.10 0.40 2.72 0.67 (TBIL) M 5.20 3.88 2.24 1.50** 3.10 1.23* Blood urea nitrogen F 3.69 0.46 3.80 0.59 4.00 0.43 (BUN) M 4.54 0.72 2.76 1.09** 3.46 0.72* Creatinine F 59.80 5.02 57.50 3.11 55.40 2.30 (CRE) M 60.00 7.25 38.80 25.62* 66.60 15.09 Creatinine kinase F 924.20 445.66 776.00 437.68 636.80 536.05 (CK) M 634.50 37.48 938.60 553.43* 762.00 351.72 Note: *P < 0.05, **P < 0.01, as compared with the same gender of control group.
Example 8: Sub-Acute Toxic Effect of 5-bromo-2-(-hydroxypentyl)benzoic Acid Sodium Salt (in amorphous form) on Blood Routine Tests
[0041]
TABLE-US-00007 Groups Control ig iv Items Sex group (20 mg/kg) (12 mg/kg) RBC F 6.97 1.80 4.51 3.30 6.95 1.06 M 7.67 0.53 7.61 0.28 7.86 0.40 HGB F 126.60 30.22 117.00 19.70 125.60 16.68 M 140.33 5.69 134.00 19.13 141.40 7.20 PLT F 1129.75 109.48 1146.00 74.22 1075.00 41.70 M 1118.67 53.26 1258.20 281.33 1087.20 56.34 WBC F 5.50 2.49 4.44 2.24 5.20 1.53 M 4.67 2.22 5.18 0.71 5.66 2.23 Neutrophil F 22.20 7.96 28.44 10.20 21.44 2.74 (NEU) M 17.23 5.66 23.88 5.48 22.16 4.14 Lymphocyte F 71.26 7.86 66.45 11.13 73.62 2.16 (LYM) M 73.53 9.11 70.84 3.99 72.50 4.66 Monocyte F 4.56 2.12 2.05 1.74** 3.62 0.63 (MON) M 7.20 323 4.93 1.91* 3.80 0.70* Eosinophil F 0.70 0.50 0.23 0.12** 0.76 0.50 (EOS) M 0.63 0.45 0.56 0.50 0.60 0.29 Basophil F 1.28 0.85 1.43 1.17 0.56 0.15** (BAS) M 1.40 0.20 0.62 0.25* 0.40 0.12** Note: *P < 0.05, **P < 0.01, as compared with the same gender of control group.
Example 9: Sub-Acute Toxic Effect of 5-bromo-2-(-hydroxypentyl)benzoic Acid Sodium Salt (in Amorphous Form) on Blood Coagulation Functions in Rats
[0042]
TABLE-US-00008 Groups Control ig iv Items Sex group (20 mg/kg) (12 mg/kg) APTT F 25.6 2.3 22.8 5.9 22.3 8.5 (sec) M 25.5 3.2 26.5 3.2 23.7 2.1 PT F 18.0 2.1 17.5 2.2 18.8 3.3 (sec) M 18.4 1.8 16.4 3.4 17.5 2.0 FIB F 7.31 0.51 7.22 0.97 6.63 0.95 (g/l) M 6.66 1.08 7.03 0.29 6.28 0.55 TT F 26.3 2.5 27.3 2.4 27.5 2.8 (sec) M 28.5 1.7 26.9 5.9 27.1 3.8
Example 10: Effect of 5-bromo-2-(-hydroxypentyl)benzoic Acid Sodium Salt (in Amorphous Form) on Body Weight of Rats
[0043]
TABLE-US-00009 Day 1 Day 6 Day 11 Day 16 Day 21 Day 24 Groups Sex BW BW W (%) BW W (%) BW W (%) BW W (%) BW W (%) Control M 178 6 207 8 16 5 253 4 22 3 295 10 66 6 330 20 85 9 341 24 92 10 F 172 6 188 6 9 2 216 10 15 3 260 6 37 5 258 11 50 6 259 10 52 7 ig group M 183 8 210 8 15 3 245 12 17 5 280 18 53 14 320 18 75 14 337 18 84 14 F 179 12 192 16 7 4 205 14 7 2* 224 11 25 4* 230 15 29 6** 235 12 31 5** iv group M 179 13 204 7 14 6 238 6 14 2* 270 6 51 10 296 8 66 15* 304 10 71 16* F 182 13 195 12 7 2 200 15 3 2** 212 14 21 7** 223 20 23 6** 221 19 21 6** Note: W (%) means the percentage of increase of body weight of rats at Day X as compared with the body weight at Day 1. *P < 0.05, as compared with the percentage of increase of body weight of the same gender in control group, **P < 0.01, as compared with the percentage of increase of body weight of the same gender in control group. Note: ig: 20 mg/kg; iv: 12 mg/kg.
Example 11
[0044] Comparison of stability and solubility of amorphous 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP, in the form of amorphous form), crystal form A of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP crystal form A) and crystal form B of 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (BZP crystal form B) with 5-bromo-2-(-hydroxypentyl)benzoic acid sodium salt (a mixture, CN ZL200810230890.X) in water and methanol solution
TABLE-US-00010 Solubility Solubility in water Stability in methanol Stability Drag (g/mL) (hours) (g/mL) (hours) BZP (amorphous) 2.98 >24 1.89 >24 BZP (Form A) 2.36 >20 1.80 >24 BZP (Form B) 2.43 >24 1.63 >24 BZP (mixture) 2.13 >12 1.58 >12