U.S. patent application number 16/976287 was filed with the patent office on 2021-01-07 for analysis method of 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, treatment for amyotrophic lateral sclerosis, inhibition of progression of amyotrophic lateral sclerosis, and method of producing drug containing 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug.
This patent application is currently assigned to MITSUBISHI TANABE PHARMA CORPORATION. The applicant listed for this patent is MITSUBISHI TANABE PHARMA CORPORATION. Invention is credited to Aya SANO, Takeshi WAKASUGI.
Application Number | 20210003544 16/976287 |
Document ID | / |
Family ID | |
Filed Date | 2021-01-07 |
United States Patent
Application |
20210003544 |
Kind Code |
A1 |
WAKASUGI; Takeshi ; et
al. |
January 7, 2021 |
ANALYSIS METHOD OF 3-METHYL-1-PHENYL-2-PYRAZOLIN-5-ONE BULK DRUG,
TREATMENT FOR AMYOTROPHIC LATERAL SCLEROSIS, INHIBITION OF
PROGRESSION OF AMYOTROPHIC LATERAL SCLEROSIS, AND METHOD OF
PRODUCING DRUG CONTAINING 3-METHYL-1-PHENYL-2-PYRAZOLIN-5-ONE BULK
DRUG
Abstract
This method of analyzing the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug involves: obtaining a
first measured value by measuring the phenylhydrazine content of a
standard solution that contains phenylhydrazine or a salt thereof,
a first acidic water and a first water-soluble organic solvent, and
that exhibits a phenylhydrazine concentration of 0.01 .mu.g/mL to
10 .mu.g/mL; obtaining a second measured value by measuring the
phenylhydrazine content in a sample solution that contains a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, a second acidic
water and a second water-soluble organic solvent; and detecting the
phenylhydrazine content in the 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug on the basis of the first measured value and the second
measured value. The first acidic water is at least one type
selected from hydrochloric acid, an aqueous acetic acid solution,
and the like, the first water-soluble organic solvent is at least
one type selected from acetonitrile and methanol, the second acidic
water is at least one type selected from hydrochloric acid, an
aqueous acetic acid solution, and the like, and the second
water-soluble organic solvent is at least one type selected from
acetonitrile, and methanol.
Inventors: |
WAKASUGI; Takeshi;
(Osaka-shi, JP) ; SANO; Aya; (Osaka-shi,
JP) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
MITSUBISHI TANABE PHARMA CORPORATION |
Osaka-shi |
|
JP |
|
|
Assignee: |
MITSUBISHI TANABE PHARMA
CORPORATION
Osaka-shi
JP
|
Appl. No.: |
16/976287 |
Filed: |
February 27, 2019 |
PCT Filed: |
February 27, 2019 |
PCT NO: |
PCT/JP2019/007415 |
371 Date: |
August 27, 2020 |
Current U.S.
Class: |
1/1 |
International
Class: |
G01N 33/15 20060101
G01N033/15; A61K 31/4152 20060101 A61K031/4152; G01N 30/06 20060101
G01N030/06 |
Foreign Application Data
Date |
Code |
Application Number |
Feb 28, 2018 |
JP |
PCT/JP2018/007544 |
Claims
1. A method of analyzing the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, which comprises:
obtaining a first measured value by measuring the phenylhydrazine
content in a phenylhydrazine standard solution that contains
phenylhydrazine or a salt thereof, a first acidic water and a first
water-soluble organic solvent, and that exhibits a phenylhydrazine
concentration of 0.01 .mu.g/mL to 10 .mu.g/mL, obtaining a second
measured value by measuring the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution that contains a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, a second acidic
water and a second water-soluble organic solvent, and detecting the
phenylhydrazine content in the 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug on the basis of the first measured value and the second
measured value, wherein the first acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution, an aqueous phosphoric acid solution
and an aqueous perchloric acid solution, and the first
water-soluble organic solvent is at least one type selected from
the group consisting of acetonitrile and methanol, provided that,
when the first water-soluble organic solvent is methanol, the first
acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution, and wherein the
second acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution, an aqueous phosphoric acid solution and an aqueous
perchloric acid solution, and the second water-soluble organic
solvent is at least one type selected from the group consisting of
acetonitrile and methanol, provided that, when the second
water-soluble organic solvent is methanol, the second acidic water
is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution.
2. The method according to claim 1, which further comprises:
preparing the phenylhydrazine standard solution that contains
phenylhydrazine or the salt thereof, the first acidic water and the
first water-soluble organic solvent, and preparing the
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution that contains
the 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, the second
acidic water and the second water-soluble organic solvent.
3. The method according to claim 1, wherein the first measured
value is a value obtained by measuring the phenylhydrazine content
in the phenylhydrazine standard solution using high performance
liquid chromatography, and the second measured value is a value
obtained by measuring the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution using high
performance liquid chromatography.
4. The method according to claim 1, wherein the first acidic water
contains 0.01 mol/L to 5 mol/L hydrochloric acid, and the second
acidic water contains 0.01 mol/L to 5 mol/L hydrochloric acid.
5. The method according to claim 1, wherein the volume ratio of the
first acidic water and the first water-soluble organic solvent
contained in the phenylhydrazine standard solution is in the range
of 3:1 to 4:5, and the volume ratio of the second acidic water and
the second water-soluble organic solvent contained in the
3-methyl-1-phenyl-2-pyrazoline-5-one sample solution is in the
range of 3:1 to 4:5.
6. A method of treating amyotrophic lateral sclerosis, which
comprises: obtaining a first measured value by measuring the
phenylhydrazine content in a phenylhydrazine standard solution that
contains phenylhydrazine or a salt thereof, a first acidic water
and a first water-soluble organic solvent, and that exhibits a
phenylhydrazine concentration of 0.01 .mu.g/mL to 10 .mu.g/mL,
obtaining a second measured value by measuring the phenylhydrazine
content in a 3-methyl-1-phenyl-2-pyrazolin-5-one sample solution
that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, a
second acidic water and a second water-soluble organic solvent,
detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value, to determine
that the phenylhydrazine content is 20 ppm or less, and
administering a drug containing the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug to a patient in need,
wherein the first acidic water is at least one type selected from
the group consisting of hydrochloric acid, an aqueous acetic acid
solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution and an
aqueous perchloric acid solution, and the first water-soluble
organic solvent is at least one type selected from the group
consisting of acetonitrile and methanol, provided that, when the
first water-soluble organic solvent is methanol, the first acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution, and wherein the second acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution, an
aqueous phosphoric acid solution and an aqueous perchloric acid
solution, and the second water-soluble organic solvent is at least
one type selected from the group consisting of acetonitrile and
methanol, provided that, when the second water-soluble organic
solvent is methanol, the second acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution and an aqueous perchloric acid
solution.
7. The method according to claim 6, which further comprises:
preparing the phenylhydrazine standard solution that contains
phenylhydrazine or the salt thereof, the first acidic water and the
first water-soluble organic solvent, and preparing the
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution that contains
the 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, the second
acidic water and the second water-soluble organic solvent.
8. The method according to claim 6, wherein the first measured
value is a value obtained by measuring the phenylhydrazine content
in the phenylhydrazine standard solution using high performance
liquid chromatography, and the second measured value is a value
obtained by measuring the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution using high
performance liquid chromatography.
9. The method according to claim 6, wherein the first acidic water
contains 0.01 mol/L to 5 mol/L hydrochloric acid, and the second
acidic water contains 0.01 mol/L to 5 mol/L hydrochloric acid.
10. The method according to claim 6, wherein the volume ratio of
the first acidic water and the first water-soluble organic solvent
contained in the phenylhydrazine standard solution is in the range
of 3:1 to 4:5, and the volume ratio of the second acidic water and
the second water-soluble organic solvent contained in the
3-methyl-1-phenyl-2-pyrazoline-5-one sample solution is in the
range of 3:1 to 4:5.
11. A method of suppressing progress of amyotrophic lateral
sclerosis, which comprises: obtaining a first measured value by
measuring the phenylhydrazine content in a phenylhydrazine standard
solution that contains phenylhydrazine or a salt thereof, a first
acidic water and a first water-soluble organic solvent, and that
exhibits a phenylhydrazine concentration of 0.01 .mu.g/mL to 10
.mu.g/mL, obtaining a second measured value by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent, detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value, to determine
that the phenylhydrazine content is 20 ppm or less, and
administering a drug containing the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug to a patient in need,
wherein the first acidic water is at least one type selected from
the group consisting of hydrochloric acid, an aqueous acetic acid
solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution and an
aqueous perchloric acid solution, and the first water-soluble
organic solvent is at least one type selected from the group
consisting of acetonitrile and methanol, provided that, when the
first water-soluble organic solvent is methanol, the first acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution, and wherein the second acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution, an
aqueous phosphoric acid solution and an aqueous perchloric acid
solution, and the second water-soluble organic solvent is at least
one type selected from the group consisting of acetonitrile and
methanol, provided that, when the second water-soluble organic
solvent is methanol, the second acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution and an aqueous perchloric acid
solution.
12. The method according to claim 11, which further comprises:
preparing the phenylhydrazine standard solution that contains
phenylhydrazine or the salt thereof, the first acidic water and the
first water-soluble organic solvent, and preparing the
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution that contains
the 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, the second
acidic water and the second water-soluble organic solvent.
13. The method according to claim 11, wherein the first measured
value is a value obtained by measuring the phenylhydrazine content
in the phenylhydrazine standard solution using high performance
liquid chromatography, and the second measured value is a value
obtained by measuring the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution using high
performance liquid chromatography.
14. The method according to claim 11, wherein the first acidic
water contains 0.01 mol/L to 5 mol/L hydrochloric acid, and the
second acidic water contains 0.01 mol/L to 5 mol/L hydrochloric
acid.
15. The method according to claim 11, wherein the volume ratio of
the first acidic water and the first water-soluble organic solvent
contained in the phenylhydrazine standard solution is in the range
of 3:1 to 4:5, and the volume ratio of the second acidic water and
the second water-soluble organic solvent contained in the
3-methyl-1-phenyl-2-pyrazoline-5-one sample solution is in the
range of 3:1 to 4:5.
16. A method of producing a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug having a phenylhydrazine content of 20 ppm or less, which
comprises: obtaining a first measured value by measuring the
phenylhydrazine content in a phenylhydrazine standard solution that
contains phenylhydrazine or a salt thereof, a first acidic water
and a first water-soluble organic solvent, and that exhibits a
phenylhydrazine concentration of 0.01 .mu.g/mL to 10 .mu.g/mL,
obtaining a second measured value by measuring the phenylhydrazine
content in a 3-methyl-1-phenyl-2-pyrazolin-5-one sample solution
that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, a
second acidic water and a second water-soluble organic solvent, and
detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value, to determine
that the phenylhydrazine content is 20 ppm or less, wherein the
first acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution, an aqueous phosphoric acid solution and an aqueous
perchloric acid solution, and the first water-soluble organic
solvent is at least one type selected from the group consisting of
acetonitrile and methanol, provided that, when the first
water-soluble organic solvent is methanol, the first acidic water
is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution, and wherein the second acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution, an
aqueous phosphoric acid solution and an aqueous perchloric acid
solution, and the second water-soluble organic solvent is at least
one type selected from the group consisting of acetonitrile and
methanol, provided that, when the second water-soluble organic
solvent is methanol, the second acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution and an aqueous perchloric acid
solution.
17. A method of producing a drug containing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug having a
phenylhydrazine content of 20 ppm or less, which comprises:
obtaining a first measured value by measuring the phenylhydrazine
content in a phenylhydrazine standard solution that contains
phenylhydrazine or a salt thereof, a first acidic water and a first
water-soluble organic solvent, and that exhibits a phenylhydrazine
concentration of 0.01 .mu.g/mL to 10 .mu.g/mL, obtaining a second
measured value by measuring the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution that contains a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, a second acidic
water and a second water-soluble organic solvent, detecting the
phenylhydrazine content in the 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug on the basis of the first measured value and the second
measured value, to determine that the phenylhydrazine content is 20
ppm or less, and mixing the 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug with at least one pharmacologically and pharmaceutically
acceptable additive, wherein the first acidic water is at least one
type selected from the group consisting of hydrochloric acid, an
aqueous acetic acid solution, an aqueous trifluoroacetic acid
solution, an aqueous formic acid solution, an aqueous phosphoric
acid solution and an aqueous perchloric acid solution, and the
first water-soluble organic solvent is at least one type selected
from the group consisting of acetonitrile and methanol, provided
that, when the first water-soluble organic solvent is methanol, the
first acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution, and wherein the
second acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution, an aqueous phosphoric acid solution and an aqueous
perchloric acid solution, and the second water-soluble organic
solvent is at least one type selected from the group consisting of
acetonitrile and methanol, provided that, when the second
water-soluble organic solvent is methanol, the second acidic water
is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution.
18. A drug for treating amyotrophic lateral sclerosis or
suppressing progress of amyotrophic lateral sclerosis, which is
produced by the method according to claim 17.
19. A 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug obtained by the
method according to claim 16.
Description
TECHNICAL FIELD
[0001] The present invention is related to a method of analyzing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, treatment for
amyotrophic lateral sclerosis (hereinafter, may be also referred to
as ALS) and inhibition of progression of ALS, and a method of
producing a drug containing a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug.
BACKGROUND ART
[0002] ALS, which is one of the motor neuron diseases, is an
incurable disease that manifests initial symptoms such as weakness
of the hands, dyskinesia of the fingers and fiber spasm of the
upper limbs, and causes respiratory failure through muscle
atrophy/muscular weakness, bulbar paralysis and muscle fiber spasm.
ALS is classified into upper limb type, bulbar type, lower limb
type and mixed type depending on the onset site, all of which show
that muscle groups of the whole body are affected while the
symptoms progress. As a pharmaceutical product effective in
treating ALS and suppressing progress of ALS, the drugs that
contains a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug have been
approved and clinically used in Japan, the United States, and so on
(for example, generic name "edaravone", brand names "Radicut
(registered trademark)", "Radicava (registered trademark)"
manufactured and sold by Mitsubishi Tanabe Pharma Corporation, and
the like).
[0003] The drugs may contain phenylhydrazine as one of the
impurities. As a method of analyzing phenylhydrazine contained in
3-methyl-1-phenyl-2-pyrazolin-5-one, the method has been reported,
which comprises preparing ammonium dihydrogen phosphate
solution-methanol whose pH is adjusted to 2.0 with phosphoric acid
as a liquid for dissolution, and analyzing phenylhydrazine by HPLC
using the liquid for dissolution of ammonium dihydrogen phosphate
solution-methanol whose pH is adjusted to 3.5 with phosphoric acid
as a mobile phase (Nonpatent Document 1). This disclosure is
incorporated herein by reference.
CITATION LIST
Nonpatent Document
[0004] Nonpatent Document 1: Central South Pharmacy, August 2014,
Vol. 12 No. 8, 814-816.
SUMMARY OF THE INVENTION
Problems to be Solved by Invention
[0005] The purpose of the present invention is to provide an
analysis method for measuring phenylhydrazine contained in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug accurately and
simply.
[0006] The purpose of the present invention is also to provide a
method of treating amyotrophic lateral sclerosis and of suppressing
progress of amyotrophic lateral sclerosis using a drug containing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug in which the
phenylhydrazine content is strictly controlled.
[0007] Further, the purpose of the present invention is to provide
a method of producing a drug containing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug in which the
phenylhydrazine content is strictly controlled.
Means to Solve Problems
[0008] A method of analyzing the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug according to an
embodiment of the present invention comprises:
[0009] obtaining a first measured value by measuring the
phenylhydrazine content in a phenylhydrazine standard solution that
contains phenylhydrazine or a salt thereof, a first acidic water
and a first water-soluble organic solvent, and that exhibits a
phenylhydrazine concentration of 0.01 .mu.g/mL to 10 .mu.g/mL,
[0010] obtaining a second measured value by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent, and
[0011] detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value,
[0012] wherein the first acidic water is at least one type selected
from the group consisting of hydrochloric acid, an aqueous acetic
acid solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution and an
aqueous perchloric acid solution, and the first water-soluble
organic solvent is at least one type selected from the group
consisting of acetonitrile and methanol, provided that, when the
first water-soluble organic solvent is methanol, the first acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution, and
[0013] wherein the second acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution, an aqueous phosphoric acid solution
and an aqueous perchloric acid solution, and the second
water-soluble organic solvent is at least one type selected from
the group consisting of acetonitrile and methanol, provided that,
when the second water-soluble organic solvent is methanol, the
second acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution.
[0014] A method of treating amyotrophic lateral sclerosis according
to another embodiment of the present invention comprises:
[0015] obtaining a first measured value by measuring the
phenylhydrazine content in a phenylhydrazine standard solution that
contains phenylhydrazine or a salt thereof, a first acidic water
and a first water-soluble organic solvent, and that exhibits a
phenylhydrazine concentration of 0.01 .mu.g/mL to 10 .mu.g/mL,
[0016] obtaining a second measured value by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent,
[0017] detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value, to determine
that the phenylhydrazine content is 20 ppm or less, and
[0018] administering a drug containing the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug to a patient in
need,
[0019] wherein the first acidic water is at least one type selected
from the group consisting of hydrochloric acid, an aqueous acetic
acid solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution and an
aqueous perchloric acid solution, and the first water-soluble
organic solvent is at least one type selected from the group
consisting of acetonitrile and methanol, provided that, when the
first water-soluble organic solvent is methanol, the first acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution, and
[0020] wherein the second acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution, an aqueous phosphoric acid solution
and an aqueous perchloric acid solution, and the second
water-soluble organic solvent is at least one type selected from
the group consisting of acetonitrile and methanol, provided that,
when the second water-soluble organic solvent is methanol, the
second acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution.
[0021] A method of suppressing progress of amyotrophic lateral
sclerosis according to yet another embodiment of the present
invention comprises:
[0022] obtaining a first measured value by measuring the
phenylhydrazine content in a phenylhydrazine standard solution that
contains phenylhydrazine or a salt thereof, a first acidic water
and a first water-soluble organic solvent, and that exhibits a
phenylhydrazine concentration of 0.01 .mu.g/mL to 10 .mu.g/mL,
[0023] obtaining a second measured value by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent,
[0024] detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value, to determine
that the phenylhydrazine content is 20 ppm or less, and
[0025] administering a drug containing the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug to a patient in
need,
[0026] wherein the first acidic water is at least one type selected
from the group consisting of hydrochloric acid, an aqueous acetic
acid solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution and an
aqueous perchloric acid solution, and the first water-soluble
organic solvent is at least one type selected from the group
consisting of acetonitrile and methanol, provided that, when the
first water-soluble organic solvent is methanol, the first acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution, and
[0027] wherein the second acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution, an aqueous phosphoric acid solution
and an aqueous perchloric acid solution, and the second
water-soluble organic solvent is at least one type selected from
the group consisting of acetonitrile and methanol, provided that,
when the second water-soluble organic solvent is methanol, the
second acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution.
[0028] A method of producing a drug containing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug having a
phenylhydrazine content of 20 ppm or less according to still
another aspect of the present invention comprises:
[0029] obtaining a first measured value by measuring the
phenylhydrazine content in a phenylhydrazine standard solution that
contains phenylhydrazine or a salt thereof, a first acidic water
and a first water-soluble organic solvent, and that exhibits a
phenylhydrazine concentration of 0.01 .mu.g/mL to 10 .mu.g/mL,
[0030] obtaining a second measured value by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent,
[0031] detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value, to determine
that the phenylhydrazine content is 20 ppm or less, and
[0032] mixing the 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug
with at least one pharmacologically and pharmaceutically acceptable
additive,
[0033] wherein the first acidic water is at least one type selected
from the group consisting of hydrochloric acid, an aqueous acetic
acid solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution and an
aqueous perchloric acid solution, and the first water-soluble
organic solvent is at least one type selected from the group
consisting of acetonitrile and methanol, provided that, when the
first water-soluble organic solvent is methanol, the first acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution, and
[0034] wherein the second acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution, an aqueous phosphoric acid solution
and an aqueous perchloric acid solution, and the second
water-soluble organic solvent is at least one type selected from
the group consisting of acetonitrile and methanol, provided that,
when the second water-soluble organic solvent is methanol, the
second acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution.
Effect of Invention
[0035] According to the present invention, it is possible to
provide an analysis method for measuring phenylhydrazine contained
in a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug accurately and
simply.
[0036] Further, according to the present invention, it is possible
to provide a method of treating amyotrophic lateral sclerosis and
of suppressing progress of amyotrophic lateral sclerosis which
comprises using a drug containing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug in which the
phenylhydrazine content is strictly controlled.
[0037] Furthermore, according to the present invention, it is
possible to provide a method of producing a drug containing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug in which the
phenylhydrazine content is strictly controlled.
BRIEF DESCRIPTION OF THE DRAWINGS
[0038] FIG. 1 is an HPLC chromatogram of a phenylhydrazine solution
prepared with a 0.1 mol/L hydrochloric acid/acetonitrile (6:4)
mixture.
[0039] FIG. 2 is an HPLC chromatogram of a phenylhydrazine solution
prepared with a 0.1 mol/L hydrochloric acid/acetonitrile (4:6)
mixture.
[0040] FIG. 3 is an HPLC chromatogram of a phenylhydrazine solution
prepared with a 0.1 mol/L hydrochloric acid/acetonitrile (2:8)
mixture.
[0041] FIG. 4 is an HPLC chromatogram of a phenylhydrazine solution
prepared with a 1 mol/L hydrochloric acid/acetonitrile (6:4)
mixture.
[0042] FIG. 5 is an HPLC chromatogram of a phenylhydrazine solution
prepared with a 0.1 mol/L hydrochloric acid/acetonitrile (7:3)
mixture.
[0043] FIG. 6 is a graph showing the time-dependent changes in the
ratio (%) of the peak area to the initial value of phenylhydrazine
contained in an addition sample solution prepared with a 0.1 mol/L
hydrochloric acid/acetonitrile (7:3) mixture.
[0044] FIG. 7 is a graph showing the time-dependent changes in the
ratio (%) of the peak area to the initial value of phenylhydrazine
contained in an addition sample solution prepared with a 0.1 mol/L
hydrochloric acid/methanol (5:5) mixture.
MODE FOR CARRYING OUT THE INVENTION
[0045] The embodiments of a method of analyzing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, a method of treating
amyotrophic lateral sclerosis (hereinafter, may be also referred to
as ALS), a method of suppressing progress of amyotrophic lateral
sclerosis, and a method of producing a drug containing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug are described in
detail hereinafter, with reference to the accompanying
drawings.
[0046] A method of analyzing the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug according to the
present invention comprises obtaining a first measured value by
measuring the phenylhydrazine content in a phenylhydrazine standard
solution that contains phenylhydrazine or a salt thereof, a first
acidic water and a first water-soluble organic solvent and that
exhibits a phenylhydrazine concentration of 0.01 .mu.g/mL to 10
.mu.g/mL, obtaining a second measured value by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent, and detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value.
[0047] The method of analyzing the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug according to the
present invention may further comprise other steps, if
necessary.
[0048] 3-Methyl-1-phenyl-2-pyrazolin-5-one may be represented by
the following structural formula. As for
3-methyl-1-phenyl-2-pyrazolin-5-one, the tautomers represented by
the following structural formulas exist.
##STR00001##
[0049] The phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug can be determined by
analyzing a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug through one or more
of the quantitative analytical techniques, and comparing the
resultant test result with the test result obtained in a standard
solution of a substantially pure phenylhydrazine.
[0050] It is preferable that a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug and a standard
solution of phenylhydrazine should be ones in which a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug and phenylhydrazine
or a salt thereof are completely dissolved in a solvent used
therefor. Use of such a solution enables to measure the accurate
content of phenylhydrazine contained in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug.
[0051] It can be visually confirmed whether or not a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug and phenylhydrazine
or a salt thereof are completely dissolved. A dissolved residue in
minute amounts would be checked by evaluating the same filtered
with a filter followed by measuring with HPLC.
[0052] It is desirable that, in a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug and a standard
solution of phenylhydrazine, 3-methyl-1-phenyl-2-pyrazolin-5-one or
a physiologically acceptable salt thereof and/or phenylhydrazine
would not substantially decompose until each solution is prepared
and then analyzed after a certain period of time. Phenylhydrazine
can be exemplified as one of the decomposition products of
3-methyl-1-phenyl-2-pyrazolin-5-one or a physiologically acceptable
salt thereof, and the degree of decomposition of
3-methyl-1-phenyl-2-pyrazolin-5-one or a physiologically acceptable
salt thereof can be checked by determining the change in the
phenylhydrazine content in the solution. The phenylhydrazine
content in a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug and/or a standard
solution of phenylhydrazine can be measured by analysis method
using HPLC as described below.
[0053] Regarding the description that phenylhydrazine would not
substantially decompose, specifically the value measured after a
certain period of time does not change preferably by 20% or more,
more preferably it does not change by 10% or more, and even more
preferably it does not change by 5% or more, compared with the
value of phenylhydrazine measured within 30 minutes after preparing
a phenylhydrazine standard solution.
[0054] Further, regarding the description that
3-methyl-1-phenyl-2-pyrazolin-5-one or a physiologically acceptable
salt thereof would not substantially decompose, specifically the
value of phenylhydrazine measured after a certain period of time
does not change preferably by 4 ppm or more relative to the content
of 3-methyl-1-phenyl-2-pyrazolin-5-one, more preferably, it does
not change by 2 ppm or more, and even more preferably, it does not
change by 1 ppm or more, compared with the value of phenylhydrazine
measured within 30 minutes after preparing a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug.
[0055] A certain period of time is not particularly limited.
Preferably it is 2 hours, 3 hours, 4 hours, 5 hours, 9 hours, 10
hours, 12 hours or 24 hours, more preferably it is 2 hours, 3
hours, 4 hours, 5 hours, 9 hours, 10 hours or 12 hours, and it is
even more preferably 2 hours, 3 hours, 4 hours or 5 hours.
[0056] A sample solution of a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug contains a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug,
an acidic water, and a water-soluble organic solvent.
[0057] The acidic water is at least one type selected from the
group consisting of hydrochloric acid, an aqueous acetic acid
solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution, and an
aqueous perchloric acid solution. These acidic waters may be used
alone, or a mixture prepared by mixing two or more acidic waters at
an appropriate ratio may be used. At least one type selected from
the group consisting of hydrochloric acid, an aqueous acetic acid
solution, an aqueous trifluoroacetic acid solution and an aqueous
formic acid solution is preferable, and hydrochloric acid is more
preferable.
[0058] The acid concentration of the acidic water is preferably
0.01 mol/L to 5 mol/L, more preferably 0.02 mol/L to 1 mol/L, and
even more preferably 0.05 mol/L to 0.2 mol/L., and even more
preferably 0.1 mol/L.
[0059] The water-soluble organic solvent is at least one type
selected from the group consisting of acetonitrile and methanol.
These water-soluble organic solvents may be used alone, or a
mixture prepared by mixing these solvents at an appropriate ratio
may be used. Acetonitrile is preferable.
[0060] When the water-soluble organic solvent is methanol, the
acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution.
[0061] The ratio of the acidic water and the water-soluble organic
solvent can be changed appropriately. The volume ratio of acidic
water:water-soluble organic solvent can be 3:1 to 4:5, preferably
3:1 to 5:5, and more preferably 3:1 to 6:4. For example, the volume
ratio of acidic water:water-soluble organic solvent can be 7:3,
6:4, and 5:5. When the acidic water and the water-soluble organic
solvent are hydrochloric acid and acetonitrile, respectively, a
particularly preferable volume ratio is 7:3. When the acidic water
and the water-soluble organic solvent are hydrochloric acid and
methanol, respectively, a particularly preferable volume ratio is
5:5.
[0062] A method of preparing a sample solution is not particularly
limited as long as the sample solution contains a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, an acidic water and
a water-soluble organic solvent. An acidic water prepared may be
mixed with a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug and a
water-soluble organic solvent to prepare a sample solution, and
further, a water may be mixed with a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug and/or a
water-soluble organic solvent, followed by adding a corresponding
acid to the mixture to prepare a sample solution.
[0063] A standard solution of phenylhydrazine contains
phenylhydrazine or a salt thereof, an acidic water, and a
water-soluble organic solvent.
[0064] The phenylhydrazine standard solution may further contain a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug.
[0065] Examples of the salt of phenylhydrazine include salts with
mineral acids such as hydrochloric acid, sulfuric acid, hydrobromic
acid and phosphoric acid. As the phenylhydrazine or a salt thereof,
a salt of phenylhydrazine is preferable. As the salt of
phenylhydrazine, phenylhydrazine hydrochloride is preferable.
[0066] According to the analysis method of the present invention,
phenylhydrazine hydrochloride is more preferable than
phenylhydrazine as the former is less likely to be decomposed
during storage, and therefore is most preferable as phenylhydrazine
or a salt thereof used in the standard solution of the present
invention.
[0067] It is desirable that phenylhydrazine or a salt thereof would
be substantially pure. Specifically, the purity is preferably at
least 90% or more, more preferably at least 95% or more, at least
98% or more, and even more preferably at least 99% or more.
Phenylhydrazine hydrochloride having a purity of 98% or more and a
purity of 99% or more is commercially available (for example,
phenylhydrazine hydrochloride, 99%; Alfa Aesar; code: A14645), and
such a commercially available product may be used.
[0068] The acidic water is at least one type selected from the
group consisting of hydrochloric acid, an aqueous acetic acid
solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution, and an
aqueous perchloric acid solution. These acidic waters may be used
alone, or a mixture prepared by mixing two or more acidic waters at
an appropriate ratio may be used. At least one type selected from
the group consisting of hydrochloric acid, an aqueous acetic acid
solution, an aqueous trifluoroacetic acid solution and an aqueous
formic acid solution is preferable, and hydrochloric acid is more
preferable.
[0069] The acid concentration of the acidic water is preferably
0.01 mol/L to 5 mol/L, more preferably 0.02 mol/L to 1 mol/L, and
even more preferably 0.05 mol/L to 0.2 mol/L., And even more
preferably 0.1 mol/L.
[0070] The water-soluble organic solvent is at least one type
selected from the group consisting of acetonitrile and methanol.
These water-soluble organic solvents may be used alone, or a
mixture prepared by mixing these solvents at an appropriate ratio
may be used. Acetonitrile is preferable.
[0071] When the water-soluble organic solvent is methanol, the
acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution. The concentration
of phenylhydrazine in a phenylhydrazine standard solution can be
appropriately adjusted according to the concentration of
phenylhydrazine in a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug
to be analyzed.
[0072] For example, the concentration of phenylhydrazine in a
phenylhydrazine standard solution (the concentration of
phenylhydrazine in the case of a phenylhydrazine free form, and the
concentration equivalent to phenylhydrazine in the case of a salt
of phenylhydrazine) may be adjusted from 0.01 .mu.g/mL to 10
.mu.g/mL, preferably 0.05 .mu.g/mL to 2 .mu.g/mL, and more
preferably 0.1 .mu.g/mL to 1 .mu.g/mL.
[0073] Alternatively, the concentration of phenylhydrazine in a
phenylhydrazine standard solution can be adjusted to a
concentration equivalent to 1 ppm to 200 ppm corresponding to the
dilution ratio of a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug. The concentration
equivalent to ppm means a concentration corresponding to the
dilution ratio of a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug. For example, when
0.10 g of 3-methyl-1-phenyl-2-pyrazolin-5-one is mixed with an
acidic water and a water-soluble organic solvent to make 10 mL, the
concentration equivalent to 1 ppm is 0.01 .mu.g/mL. The
concentration equivalent to 5 ppm to 100 ppm is preferable, and the
concentration equivalent to 10 ppm to 50 ppm is more
preferable.
[0074] An embodiment of the present invention comprises obtaining a
first measured value by measuring the phenylhydrazine content of a
phenylhydrazine standard solution that exhibits a phenylhydrazine
concentration of 0.01 .mu.g/mL to 10 .mu.g/mL. The number of the
first measured value is not particularly limited as long as it can
be compared with a second measured value obtained by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution, which number can be one or more.
[0075] The ratio of the acidic water and the water-soluble organic
solvent can be changed appropriately. The volume ratio of acidic
water:water-soluble organic solvent can be 3:1 to 4:5, preferably
3:1 to 5:5, and more preferably 3:1 to 6:4. For example, the volume
ratio of acidic water water-soluble organic solvent can be 7:3,
6:4, and 5:5. Particularly preferred is 7:3. When the acidic water
and the water-soluble organic solvent are hydrochloric acid and
acetonitrile, respectively, a particularly preferable volume ratio
is 7:3. When the acidic water and the water-soluble organic solvent
are hydrochloric acid and methanol, respectively, a particularly
preferable volume ratio is 5:5.
[0076] As long as a phenylhydrazine standard solution contains
phenylhydrazine or a salt thereof, an acidic water, and a
water-soluble organic solvent, the method of preparing the
phenylhydrazine standard solution is not particularly limited. The
acidic water prepared may be mixed with phenylhydrazine or a salt
thereof and a water-soluble organic solvent to prepare a
phenylhydrazine standard solution, and further, water may be mixed
with phenylhydrazine or a salt thereof and/or a water-soluble
organic solvent, followed by adding a corresponding acid to the
mixture to prepare a phenylhydrazine standard solution.
[0077] An acidic water in a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug is preferably the
same as an acidic water in a phenylhydrazine standard solution. A
water-soluble organic solvent in a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug is preferably the
same as a water-soluble organic solvent in a phenylhydrazine
standard solution. A volume ratio of acidic water to water-soluble
organic solvent in a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug is preferably the
same as a volume ratio of acidic water to water-soluble organic
solvent in a phenylhydrazine standard solution.
[0078] More preferably, an acidic water, a water-soluble organic
solvent and a volume ratio of the acidic water to the water-soluble
organic solvent in a sample solution of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug are the same,
respectively, as an acidic water, a water-soluble organic solvent
and a volume ration of the acidic water to the water-soluble
organic solvent in a phenylhydrazine standard solution.
[0079] Quantitative analysis techniques include high performance
liquid chromatography (HPLC). High performance liquid
chromatography includes high performance liquid chromatography
(HPLC) and ultra high performance liquid chromatography (UHPLC).
When high performance liquid chromatography is used, the
theoretical plate number of the phenylhydrazine peak is preferably
as large as possible, and for example, the theoretical plate number
N determined by the half-width method can be 1000 plates or more,
and more preferably 3000 plates or more.
Theoretical plate number: N=5.54.times.(tr/W.sub.0.5).sup.2 Formula
1
wherein
[0080] tr: retention time, and
[0081] W.sub.0.5: Width of half peak height
[0082] Symmetry coefficient S is an index representing the degree
of symmetry of the peak on the chromatogram, and the closer the
symmetry coefficient S is to 1, the more symmetrical the peak is
(normal distribution). The closer the symmetry coefficient S is to
1, the higher the performances as analysis method such as accuracy,
precision and limit of quantification, becomes, which is
preferable. For example, symmetry coefficient S may be 2.0 or less,
and more preferably 1.5 or less.
Symmetry factor: S=W.sub.0.05h/2f Formula 2
wherein
[0083] W.sub.0.05h: Peak width at a height of 1/20 of the peak
height from the peak baseline
[0084] 2f: Distance on the rising side of the peak width of
W.sub.0.05h obtained by dividing the peak width by a perpendicular
line drawn from the peak top to the horizontal axis.
[0085] Examples of physiologically acceptable salts of
3-methyl-1-phenyl-2-pyrazolin-5-one or a physiologically acceptable
salt thereof include salts with mineral acids such as hydrochloric
acid, sulfuric acid, hydrobromic acid, phosphoric acid and the
like; salts with organic acids such as methanesulfonic acid,
p-toluenesulfonic acid, acetic acid, oxalic acid, citric acid,
malic acid, fumaric acid and the like; salts with alkali metals
such as sodium, potassium and the like; salts with alkaline earth
metals such as magnesium, and the like; and salts with amines such
as ammonium, ethanolamine, 2-amino-2-methyl-1-propanol and the
like. Physiologically acceptable salts also include hydrates and
solvates.
[0086] 3-Methyl-1-phenyl-2-pyrazolin-5-one bulk drug means
3-methyl-1-phenyl-2-pyrazolin-5-one or its physiologically
acceptable salt as the active ingredient. A
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug may contain
phenylhydrazine as one of the impurities. According to the
preparation method described in European Patent Publication No.
208874 (or Japanese Patent Publication (Kokoku) No. H5-31523),
phenylhydrazine is used as a precursor for synthesizing
3-methyl-1-phenyl-2-pyrazolin-5-one.
[0087] A solution preparation containing a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug is prone to increase
in phenylhydrazine during the storage. Phenylhydrazine is a
mutagenic compound, and its exposure limit for industrial health is
set in each country. The United States has legislated the strictest
exposure limit for phenylhydrazine, which is 2,860 .mu.g/kg/year.
Phenylhydrazine is easily decomposed by light and oxygen.
[0088] It is preferable that the amount of impurity phenylhydrazine
contained in a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug is as
small as possible. For example, it is preferably 20 ppm or less,
and more preferably 10 ppm or less.
[0089] When 3-methyl-1-phenyl-2-pyrazolin-5-one is used for the
purpose of treating ALS or suppressing progress of ALS, it is
preferable to continue the pharmacotherapy with this drug for a
lifetime starting administration of the drug. Therefore, it is
preferable that the phenylhydrazine content should be strictly
controlled.
[0090] An embodiment of the present invention is a method of
producing a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug having a
phenylhydrazine content of 20 ppm or less, which comprises:
[0091] obtaining a first measured value by measuring the
phenylhydrazine content in a phenylhydrazine standard solution that
contains phenylhydrazine or a salt thereof, a first acidic water
and a first water-soluble organic solvent, and that exhibits a
phenylhydrazine concentration of 0.01 .mu.g/mL to 10 .mu.g/mL,
[0092] obtaining a second measured value by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent, and
[0093] detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value, to determine
that the phenylhydrazine content is 20 ppm or less,
[0094] wherein the first acidic water is at least one type selected
from the group consisting of hydrochloric acid, an aqueous acetic
acid solution, an aqueous trifluoroacetic acid solution, an aqueous
formic acid solution, an aqueous phosphoric acid solution and an
aqueous perchloric acid solution, and the first water-soluble
organic solvent is at least one type selected from the group
consisting of acetonitrile and methanol, provided that, when the
first water-soluble organic solvent is methanol, the first acidic
water is at least one type selected from the group consisting of
hydrochloric acid, an aqueous acetic acid solution, an aqueous
trifluoroacetic acid solution, an aqueous formic acid solution and
an aqueous perchloric acid solution, and
[0095] wherein the second acidic water is at least one type
selected from the group consisting of hydrochloric acid, an aqueous
acetic acid solution, an aqueous trifluoroacetic acid solution, an
aqueous formic acid solution, an aqueous phosphoric acid solution
and an aqueous perchloric acid solution, and the second
water-soluble organic solvent is at least one type selected from
the group consisting of acetonitrile and methanol, provided that,
when the second water-soluble organic solvent is methanol, the
second acidic water is at least one type selected from the group
consisting of hydrochloric acid, an aqueous acetic acid solution,
an aqueous trifluoroacetic acid solution, an aqueous formic acid
solution and an aqueous perchloric acid solution.
[0096] A drug according to an embodiment of the present invention
is the drug for treating amyotrophic lateral sclerosis or
suppressing progress of amyotrophic lateral sclerosis, which
contains a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug wherein
the phenylhydrazine content in the bulk drug is 20 ppm or less, and
which is produced by the method of the production according to the
present invention.
[0097] A drug according to a further embodiment of the present
invention contains a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug
for treating amyotrophic lateral sclerosis or suppressing disease
progression, which bulk drug is analyzed for the phenylhydrazine
content therein to determine that the phenylhydrazine content is 20
ppm or less.
[0098] In addition, the drug may contain a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, and a
pharmacologically and pharmaceutically acceptable additive, and
further may contain one or more bulk drugs other than the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug.
[0099] A drug containing a pharmacologically and pharmaceutically
acceptable additive according to an embodiment of the present
invention may be produced by a method of the production which
comprises mixing a 3-methyl-1-phenyl-2-pyrazoline-5-one bulk drug
having a phenylhydrazine content of 20 ppm or less with a
pharmacologically and pharmaceutically acceptable additive.
[0100] Although an active ingredient according to an embodiment of
the present invention, 3-methyl-1-phenyl-2-pyrazolin-5-one or a
physiologically acceptable salt thereof, may be administered to a
patient solely, it is preferable that the active ingredient would
be administered in a form of a drug as described above, which is
prepared by adding a pharmacologically and pharmaceutically
acceptable additive.
[0101] As a pharmacologically and pharmaceutically acceptable
additive, the followings may be used: for example, an excipient, a
disintegrating agent or disintegrating accelerator, a binder, a
lubricant, a coating agent, a dye, a diluent, a base, a
solubilizing agent or solubilizing accelerator, an isotonizing
agent, a pH adjusting agent, a stabilizer, a propellant, an
adhesive agent and the like. Examples of dosage forms suitable for
oral administration include, for example, a tablet, a capsule, a
powder, a fine granule, a granule, a liquid, a syrup and the like,
and examples of dosage forms suitable for parenteral administration
include an injection, a drip infusion, a patch, a suppository and
the like.
[0102] As an additive in the drug suitable for oral administration,
the followings may be used: for example, excipients such as
glucose, lactose, D-mannitol, starch or crystalline cellulose;
disintegrating agents or disintegrating accelerators such as
carboxymethylcellulose, starch, or carboxymethylcellulose calcium;
binders such as hydroxypropylcellulose,
hydroxypropylmethylcellulose, polyvinylpyrrolidone or gelatin;
lubricants such as magnesium stearate or talc; coating agents such
as hydroxypropylmethylcellulose, sucrose, polyethylene glycol or
titanium oxide; and bases such as petroleum jelly, liquid paraffin,
polyethylene glycol, gelatin, kaolin, glycerin, purified water, or
hard fat.
[0103] As a pharmaceutical additive in the drug suitable for
injection or drip infusion, the followings may be used:
solubilizing agents or solubilizing accelerators that can
constitute an aqueous or in-use dissolution type injection such as
distilled water for injection, physiological saline, and propylene
glycol; isotonizing agents such as glucose, sodium chloride,
D-mannitol, and glycerin; and pH adjusting agents such as an
inorganic acid, an organic acid, an inorganic base or an organic
base.
[0104] Generic name "edaravone", brand name "Radicat (registered
trademark)", or "Radicava (registered trademark)" manufactured and
sold by Mitsubishi Tanabe Pharma Corporation, and so on, which has
been already clinically used as a cerebral protective agent and a
treating agent for ALS and a suppressing agent for progress of ALS
containing 3-methyl-1-phenyl-2-pyrazolin-5-one as an active
ingredient, may be utilized as 3-methyl-1-phenyl-2-pyrazolin-5-one
or a physiologically acceptable salt thereof used in the drug and
the method according to the present invention.
[0105] A method of treating amyotrophic lateral sclerosis or
suppressing the disease progression according to an embodiment of
the present invention, comprises administering a drug containing an
effective amount of a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug
to a patient in need of the treatment or the suppression of the
disease progression, wherein the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug in the drug has a
phenylhydrazine content of 20 ppm or less as a result of the
analysis of the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug.
[0106] This embodiment may further comprises obtaining a first
measured value by measuring the phenylhydrazine content in a
phenylhydrazine standard solution that contains phenylhydrazine or
a salt thereof, a first acidic water and a first water-soluble
organic solvent, and that exhibits a phenylhydrazine concentration
of 0.01 .mu.g/mL to 10 .mu.g/mL, obtaining a second measured value
by measuring the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution that contains a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, a second acidic
water and a second water-soluble organic solvent, and detecting the
phenylhydrazine content in the 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug on the basis of the first measured value and the second
measured value to determine that the phenylhydrazine content is 20
ppm or less.
[0107] A method of treating amyotrophic lateral sclerosis or
suppressing progress of amyotrophic lateral sclerosis according to
an embodiment of the present invention may comprises providing a
rest period during a pharmacotherapy period, setting a dosing
period and a rest period as one (1) unit, thereby repeating the
unit. For example, a rest period of 14 days is provided twice or
more during a pharmacotherapy period, a dosing period and a rest
period is set as one unit, and the unit is repeated twice or more.
For example, two repeats of a dosing period and a rest period mean
"dosing period, rest period, dosing period, and rest period", but
the last rest period may be omitted to imply "dosing period, rest
period, and dosing period".
[0108] A rest period means a period in which no drug is
administered continuously for example 7 days or more.
[0109] A dosing period may be for example 14 days, or 10 days out
of 14 days. Ten days out of 14 days means any 10 days out of 14
consecutive days, and the 10 days to be administered may be 10
consecutive days or 10 inconsecutive days intervened by 1 to 4 days
of a rest period. It may be 10 consecutive days that are separated
by. A dosing period may be determined to be a preferred period
while observing the patient's condition. A rest period in an
embodiment of the present invention is preferably 14 days.
[0110] The number of repeat in the case of repeating 14-days dosing
period and 14-days rest period is not particularly limited as long
as it is 2 or more.
[0111] When a 14-days initial dosing period is followed by a
14-days initial rest period, and then a 10-days dosing period out
of 14 days and a 14-days rest period are repeated, the number of
repeat of a 10-day dosing period out of 14 days and a 14-days rest
period is not particularly limited as long as it is once or more.
In other embodiments, daily or near daily dosing may be repeated
without any rest period.
[0112] The daily dose of the active ingredient can be appropriately
selected depending on conditions such as the age and status of a
patient. In general, the dose of
3-methyl-1-phenyl-2-pyrazolin-5-one (when the active ingredient is
3-methyl-1-phenyl-2-pyrazolin-5-one, the dose is the dose of
3-methyl-1-phenyl-2-pyrazolin-5-one, or when the active ingredient
is a physiologically acceptable salt of
3-methyl-1-phenyl-2-pyrazolin-5-one, the dose is the dose
equivalent to 3-methyl-1-Phenyl-2-pyrazolin-5-one, this is the same
as below) for adults, is preferably about 15 mg to about 240 mg,
more preferably about 30 mg to about 180 mg, and even more
preferably about 60 mg to about 120 mg, particularly even more
preferably being about 60 mg.
[0113] When the administration is repeated daily or almost daily
without any rest period, in general, the daily dose of
3-methyl-1-phenyl-2-pyrazolin-5-one for adults is preferably about
60 mg, about 120 mg or about 180 mg, and more preferably about 60
mg or about 120 mg.
[0114] The number of administration per day during the dosing
period is not limited, and may be selected while observing the
patient's status. However, considering the burden on the patient
and the like, the number is preferably three times, twice or once,
and more preferably once.
[0115] The administration route of the active ingredient is not
particularly limited, and the active ingredient may be administered
orally or parenterally. Further, the active ingredient may be
administered by bolus dosing and continuous dosing, and continuous
dosing is preferable. Examples of continuous dosing include
intravenous dosing by drip infusion, transdermal dosing, oral
dosing using sublingual tablets, and oral and rectal dosing using
sustained-release formulation, and intravenous dosing by drip
infusion is preferable. In the case of bolus dosing by injection or
intravenous dosing by drip infusion, for example, the injections
described in JPA63-132833 and JPA2011-62529 may be used. These
disclosures are incorporated herein by reference.
[0116] In the case of intravenous dosing by drip infusion, the
dosing rate is preferably about 0.5 mg/min to about 1 mg/min for
the amount of 3-methyl-1-phenyl-2-pyrazolin-5-one, which is about
15 minutes to about 480 minutes in terms of time, being preferably
about 30 minutes to about 120 minutes, more preferably about 30
minutes to about 60 minutes, and even more preferably about 60
minutes.
[0117] Other dosage forms include a dosage form that is
substantially equivalent to the intravenous dosing by drip infusion
at a dose of about 0.5 mg to about 1 mg of
3-methyl-1-phenyl-2-pyrazolin-5-one per minute. The dosage form
that is substantially equivalent to the intravenous dosing by drip
infusion at a dose of about 0.5 mg to about 1 mg of
3-methyl-1-phenyl-2-pyrazolin-5-one per minute may be substantially
equivalent to the intravenous dosing in view of pharmacokinetics. A
specific example of the dosage form is a dosage form in which the
time-dependent changes in the concentration of the unchanged
3-methyl-1-phenyl-2-pyrazolin-5 in plasma after the administration
of 3-methyl-1-phenyl-2-pyrazolin-5-one or a physiologically
acceptable salt thereof is confirmed to be substantially equivalent
to the intravenous dosing. Examples of such dosing forms include
transdermal dosing, oral dosing using sublingual tablets, and oral
and rectal dosing using sustained-release formulations, and the
like.
[0118] A method of analyzing the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug according to an
embodiment of the present invention may comprises preparing a
phenylhydrazine standard solution that contains phenylhydrazine or
a salt thereof, a first acidic water and a water-soluble organic
solvent; and/or preparing a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent.
[0119] In accordance with a method of analyzing the phenylhydrazine
content in a 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug
according to another embodiment of the present invention, the first
measured value of phenylhydrazine is a value obtained by measuring
a phenylhydrazine standard solution using high performance liquid
chromatography, and/or the second measured value of phenylhydrazine
is a value obtained by measuring a
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution using high
performance liquid chromatography.
[0120] A method of treating amyotrophic lateral sclerosis or
suppressing progress of amyotrophic lateral sclerosis according to
another embodiment of the present invention comprises administering
a drug containing an effective amount of a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug to a patient in need
thereof, wherein the 3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug
was a 3-methyl-1-phenyl-2-pyrazolin-5-one that is analyzed and
determined that the phenylhydrazine content is 20 ppm or less. The
analysis of the phenylhydrazine content comprises obtaining a first
measured value by measuring the phenylhydrazine content in a
phenylhydrazine standard solution that contains phenylhydrazine or
a salt thereof, a first acidic water and a first water-soluble
organic solvent, and that exhibits a phenylhydrazine concentration
of 0.01 .mu.g/mL to 10 .mu.g/mL, to obtaining a second measured
value by measuring the phenylhydrazine content in a
3-methyl-1-phenyl-2-pyrazolin-5-one sample solution that contains a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug, a second acidic
water and a second water-soluble organic solvent, and detecting the
phenylhydrazine content in the 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug on the basis of the first measured value and the second
measured value, to determine that the phenylhydrazine content is 20
ppm or less.
[0121] A method of producing a drug for treating amyotrophic
lateral sclerosis or suppressing progress of amyotrophic lateral
sclerosis according to an embodiment of the present invention
comprises mixing a 3-methyl-1-phenyl-2-pyrazoline-5-one bulk drug
determined by analyzing a phenylhydrazine content therein to
determine that the content of a phenylhydrazine content is 20 ppm
or less, together with at least one pharmacologically and
pharmaceutically acceptable additive. The analysis of the
phenylhydrazine content comprises obtaining a first measured value
by measuring the phenylhydrazine content in a phenylhydrazine
standard solution that contains phenylhydrazine or a salt thereof,
a first acidic water and a first water-soluble organic solvent, and
that exhibits a phenylhydrazine concentration of 0.01 .mu.g/mL to
10 .mu.g/mL, obtaining a second measured value by measuring the
phenylhydrazine content in a 3-methyl-1-phenyl-2-pyrazolin-5-one
sample solution that contains a 3-methyl-1-phenyl-2-pyrazolin-5-one
bulk drug, a second acidic water and a second water-soluble organic
solvent, and detecting the phenylhydrazine content in the
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug on the basis of the
first measured value and the second measured value, to determine
that the phenylhydrazine content is 20 ppm or less.
[0122] A drug according to yet another embodiment of the present
invention is a drug for intravenous dosing which contains a
3-methyl-1-phenyl-2-pyrazolin-5-one bulk drug having a
phenylhydrazine content of 20 ppm or less.
[0123] A method of producing a drug for treating amyotrophic
lateral sclerosis or suppressing progress of amyotrophic lateral
sclerosis according to an embodiment of the present invention
involves at least one pharmacologically and pharmaceutically
acceptable additive which is distilled water for injection, or
physiological saline.
EXAMPLES
[0124] Hereinafter, the embodiments of the present invention is
illustrated by means of the following examples, but the present
invention is not limited to them in any aspect. In those examples,
3-methyl-1-phenyl-2-pyrazolin-5-one, which is a bulk drug, is
referred to as an edaravone bulk drug (may be also referred to as
EDVP).
Test Example 1
Investigation of Liquid for Dissolution
[0125] The concentration of an acidic water, a ratio of an acidic
water to a water-soluble organic solvent and so on are changed to
investigate the conditions under which an edaravone bulk drug and
phenylhydrazine or a salt thereof can be dissolved. Further, the
peak shape of phenylhydrazine is also checked by the HPLC
measurement to investigate an optimum concentration of an acidic
water and an optimal ratio of an acidic water to a water-soluble
organic solvent.
[0126] The following liquids for dissolution (a) to (f) were used
to prepare sample solutions, and the dissolution states of the
edaravone bulk drug in those solutions were investigated. Then, a
liquid for dissolution that enabled to dissolve the edaravone bulk
drug was used to prepare a solution of phenylhydrazine, and the
solution was measured using HPLC to check the peak shape of
phenylhydrazine.
Liquids for Dissolution
[0127] (a) 0.1 mol/L hydrochloric acid test liquid (b) 0.1 mol/L
hydrochloric acid/acetonitrile (8:2) mixture (c) 0.1 mol/L
hydrochloric acid/acetonitrile (6:4) mixture (d) 0.1 mol/L
hydrochloric acid/acetonitrile (4:6) mixture (e) 0.1 mol/L
hydrochloric acid/acetonitrile (2:8) mixture (f) 1 mol/L
hydrochloric acid/acetonitrile (6:4) mixture
Preparation of Sample Solutions
[0128] The edaravone bulk drug weighed to about 0.10 g was
dissolved in liquids for dissolution (a) to (f), and the solutions
were adjusted exactly to 10 mLs to provide sample solutions.
Preparation of Phenylhydrazine Solutions
[0129] To 5 .mu.L of phenylhydrazine collected, 0.1 mol/L
hydrochloric acid test liquid was added, followed by adjusting
exactly to 100 mL. To the solution measured precisely to 0.1 ml,
liquids for dissolution (a) to (f) were added, and the solutions
were adjusted exactly to 10 mLs to provide phenylhydrazine
solutions (0.5 .mu.g/mL).
Test Conditions
[0130] The measurement by HPLC was performed under the conditions
of the feeding liquid for mobile phase as follows.
Mobile phase A: 0.05 mol/L aqueous sodium perchlorate solution at
pH 2.5/acetonitrile (90:10) mixture Mobile phase B: 0.05 mol/L
aqueous sodium perchlorate solution at pH 2.5/acetonitrile (10:90)
mixture Feeding liquid for mobile phase: The concentration gradient
was controlled by changing the mixing ratio of mobile phase A and
mobile phase B as follows:
TABLE-US-00001 TABLE 1 Time after Mobile phase A Mobile phase B
injection (minutes) (vol %) (vol %) 0-7 100 0 7 - 7.1 7.1 _ - 20
##EQU00001## 100 -> 0 0 0 -> 100 100
[0131] The result of each liquid for dissolution is shown in Table
2, and the chromatograms of the phenylhydrazine solutions are shown
in FIGS. 1 to 4. Judging from the solubility of EDVP and the peak
shape of phenylhydrazine, it was found that (c) 0.1 mol/L
hydrochloric acid/acetonitrile (6:4) mixture was the most suitable,
and (f) 1 mol/L hydrochloric acid/acetonitrile (6:4) mixture could
also be used.
TABLE-US-00002 TABLE 2 Result of each liquid for dissolution sample
solutions phenylhydrazine solutions solubility peak liquids for
dissolution of EDVP pH*.sup.5 shape pH*.sup.5 (a) insoluble -- --
-- 0.1 mol/L hydrochloric acid test liquid (b) insoluble -- -- --
0.1 mol/L hydrochloric acid/acetonitrile (8:2) mixture (c) soluble
2 excellent 1 0.1 mol/L hydrochloric acid/acetonitrile (6:4)
mixture (d) soluble 2 bad 1 0.1 mol/L hydrochloric
acid/acetonitrile (4:6) mixture (e) soluble 3 bad 1 0.1 mol/L
hydrochloric acid/acetonitrile (2:8) mixture (f) soluble 1 good 1 1
mol/L hydrochloric acid/acetonitrile (6:4) mixture *.sup.5Measured
with pH test paper
[0132] In the test example, it was revealed that phenylhydrazine
hydrochloride was less likely to decompose during storage and was
easier to handle than phenylhydrazine, showing that the
hydrochloride is more suitable as a standard substance.
[0133] Accordingly, in the test example, a standard solution
prepared with phenylhydrazine hydrochloride was used to perform the
measurement. The peak of phenylhydrazine provides a theoretical
plate number and a symmetry coefficient.
[0134] In addition, according to the test example, the viewpoint of
the solubility of EDVP and the peak shape of phenylhydrazine
revealed that it was not possible to prepare any liquid for
dissolution optimal for use in the analysis of an edaravone bulk
drug by merely adjusting the pH to 2.
Test Example 2
Further Investigation of Liquids for Dissolution
[0135] Since the results of Test Example 1 revealed that liquid for
dissolution (c), which is 0.1 mol/L hydrochloric acid/acetonitrile
(6:4) mixture, was the most suitable liquid for dissolution, liquid
for dissolution (g), which is 0.1 mol/L hydrochloric
acid/acetonitrile (7:3) mixture, was used to prepare a standard
solution, and the comparison between the liquids for dissolution in
order to obtain a more preferable peak shape of a phenylhydrazine
solution.
[0136] Liquid for dissolution (c) and liquid for dissolution (g) as
described below were used to prepare standard solutions, and the
peak shapes of phenylhydrazine were further investigated.
Liquids for Dissolution
[0137] (c) 0.1 mol/L hydrochloric acid/acetonitrile (6:4) mixture
(g) 0.1 mol/L hydrochloric acid/acetonitrile (7:3) mixture
Preparation of Standard Solutions
[0138] Phenylhydrazine hydrochloride weighed precisely to about
13.4 mg (about 10 mg as a free form) was dissolved in liquid for
dissolution (c) and liquid for dissolution (g) respectively, and
the solutions were adjusted exactly to 100 mLs. To the solutions
measured precisely to 2 mLs, liquid for dissolution (c) and liquid
for dissolution (g) were added, respectively, and the solutions
were adjusted exactly to 20 mLs. To the solutions measured
precisely to 2 mLs, liquids for dissolution were added, and the
solutions were adjusted exactly to 20 mLs to provide standard
solutions (phenylhydrazine: 1 .mu.g/mL).
Test Conditions
[0139] The measurement was performed under the conditions of the
feeding liquid for mobile phase conditions as follows.
Mobile phase A: 0.05 mol/L sodium perchlorate solution at pH
2.5/acetonitrile (90:10) mixture Mobile phase B: 0.05 mol/L sodium
perchlorate solution at pH 2.5/acetonitrile (40:60) mixture Feeding
liquid for mobile phase: The concentration gradient was controlled
by changing the mixing ratio of mobile phase A and mobile phase B
as follows.
TABLE-US-00003 TABLE 3 Time after Mobile phase A Mobile phase B
injection (minutes) (vol %) (vol %) 0-7 100 0 7 - 7.1 7.1 _ - 20
##EQU00002## 100 -> 0 0 0 -> 100 100
[0140] When liquid for dissolution (g), which is 0.1 mol/L
hydrochloric acid/acetonitrile (7:3) mixture, was tried to be used
to prepare a sample solution, it was found that the edaravone bulk
drug was soluble.
[0141] The measurement results of the standard solutions prepared
with liquid for dissolution (c) and liquid for dissolution (g) as
described above are shown in Table 4 and FIGS. 1 and 5.
[0142] The test revealed that use of liquid for dissolution (g)
enabled to provide any better peak shape of the phenylhydrazine
solution.
[0143] Theoretical Plate Number and a Symmetry Coefficient
TABLE-US-00004 TABLE 4 Result of each standard solution sample
standard solutions solutions theoretical liquids for solubility
plate symmetry dissolution of EDVP number coefficient (c) 0.1 mol/L
soluble*.sup.7 3271 0.9 hydrochloric acid/acetonitrile (6:4)
mixture (g) 0.1 mol/L soluble 5563 1.0 hydrochloric
acid/acetonitrile (7:3) mixture
Example 1
[0144] The edaravone bulk drug (0.1020 g) was dissolved in a 0.1
mol/L hydrochloric acid/acetonitrile mixture (volume ratio 7:3;
hereinafter referred to as Liquid for Dissolution), and the
solution was adjusted exactly to 10 mL to prepare a first sample
solution.
[0145] Separately, 13.693 mg of phenylhydrazine hydrochloride
(about 10 mg as phenylhydrazine) was dissolved in Liquid for
Dissolution, and the solution was adjusted exactly to 100 mL. To
the solution measured precisely to 2 mL, Liquid for Dissolution was
added, and the solution was adjusted exactly to 20 mL. To the
solutions measured precisely to 2 mL, Liquid for Dissolution was
added, and the solution was adjusted exactly to 20 mL to prepare a
first standard solution (phenylhydrazine: 1 .mu.g/mL).
[0146] Separately, 5 .mu.L of phenylhydrazine was dissolved in
Liquid for Dissolution, and the solution was adjusted exactly to
100 mL. To the solutions measured precisely to 1 mL, Liquid for
Dissolution was added, and the solution was adjusted exactly to 10
mL. The resulting solution measured precisely to 1 mL was mixed
with 0.1010 g of the edaravone bulk drug, Liquid for Dissolution
was added to the mixture, and the solution was adjusted exactly to
10 mL to prepare a first addition sample solution.
[0147] After a defined period of time, each 10 .mu.L of the first
standard solution, the first sample solution, and the first
addition sample solution was precisely measured and collected, and
was subjected to the test under the following conditions by high
performance liquid chromatography, followed by measuring the peak
area of phenylhydrazine (measured value of phenylhydrazine) in each
solution according to the automatic integration process.
[0148] The phenylhydrazine content in the edaravone bulk drug can
be calculated by Equation 3 as shown below.
Test Conditions
[0149] Degassing of liquid for dissolution/nitrogen bubbling:
None
[0150] Storage temperature of a first sample solution, a first
standard solution and a first addition sample solution after
preparation: 5.degree. C.
[0151] Vessel (preparation and storage of a first sample solution,
a first standard solution and a first addition sample solution):
Brown glass bottle
[0152] Detector: UV absorption photometer (measurement wavelength:
225 nm)
[0153] Column: Octadecyl silylated silica gel packed column
(YMC-Pack Pro C18)
[0154] Column temperature: Constant temperature around 40.degree.
C.
[0155] Mobile phase: Concentration gradient controlled by starting
from 100% mobile phase A up to 100% mobile phase B [0156] Mobile
phase A: 0.05 mol/L aqueous sodium perchlorate solution at pH
2.5/acetonitrile (volume ratio 9:1) mixture [0157] Mobile phase B:
0.05 mol/L aqueous sodium perchlorate solution at pH
2.5/acetonitrile (volume ratio 2:3) mixture [0158] 0.05 mol/L
aqueous sodium perchlorate solution at pH 2.5:7.0 g of sodium
perchlorate was dissolved in 1000 mL of water, perchloric acid was
added to the solution, and the pH of the resultant solution was
adjusted to 2.5.
[0159] The results of the stability study of phenylhydrazine and/or
3-methyl-1-phenyl-2-pyrazolin-5-one in the first standard solution,
the first sample solution and the first addition sample solution
are shown in Table 5 and FIG. 6.
TABLE-US-00005 TABLE 5 Stability of phenylhydrazine and/or
3-methyl-1-phenyl-2-pyrazolin- 5-one in the solutions of a
hydrochloric acid/acetonitrile mixture Peak area of Elapsed
phenylhydrazine Ratio to initial Solution time (h) (.mu.V second)
value (%) First standard 0.0 32018 100 solution 9.3 32224 100.6
28.2 30139 94.1 First sample 0.0 0 solution 1.0 0 2.1 0 3.1 0 5.1 0
24.1 534 Content of phenylhydrazine: 1.7 ppm First addition 0.0
17313 100 sample solution 1.0 17227 99.5 2.1 17042 98.4 3.1 16815
97.1 5.1 16639 96.1 24.0 14315 82.7
[0160] As to elapsed time (h), the time point at which the first
standard solution, the first sample solution or the first addition
sample solution was injected at the beginning into the HPLC within
30 minutes after preparing the solution was set as zero "0" hour of
the elapsed time, and the subsequent elapsed times were calculated
from the zero hour.
Content of phenylhydrazine in an edaravone bulk drug
(ppm)=M.sub.S/M.sub.T.times.A.sub.T/A.sub.S.times.1.times.0.748
Equation 3:
wherein M.sub.S: Weighed amount of phenylhydrazine hydrochloride in
a first standard solution (mg) M.sub.T: Weighed amount of an
edaravone bulk drug in a first sample solution (g) A.sub.T: Peak
area of phenylhydrazine in a first sample solution (.mu.V second)
A.sub.S: Peak area of phenylhydrazine in a first standard solution
at 0 hour of the elapsed time (.mu.V second) 0.748: Conversion
factor from phenylhydrazine hydrochloride into phenylhydrazine
[0161] The ratio of the peak area to the initial value of
phenylhydrazine in the first standard solution was 90% or more even
after 28 hours elapsed, showing that phenylhydrazine in the
solution was stable.
[0162] Further, the ratio of the peak area to the initial value of
phenylhydrazine in the first addition sample solution was 90% or
more even after 5 hours elapsed, showing that phenylhydrazine,
3-methyl-1-phenyl-2-pyrazoline-5 in the solution was stable.
Furthermore, in the first sample solution, the peak area of
phenylhydrazine was not changed even after 5 hours elapsed, showing
that 3-methyl-1-phenyl-2-pyrazolin-5-one in the solution was
stable.
[0163] In addition, the use of the liquid for dissolution of 0.1
mol/L hydrochloric acid/acetonitrile mixture (volume ratio 7:3)
enabled to dissolve completely the edaravone bulk drug and
phenylhydrazine, as well as to measure exactly the phenylhydrazine
content in the solution.
[0164] Furthermore, the adoption of mobile phase A and mobile phase
B as described above when performing HPLC enabled to reliably
separate the edaravone bulk drug contained in the solution.
Further, the composition ratio of mobile phase A and mobile phase B
as described above enabled to perform quantitative measurement
without other peaks in the vicinity of the retention time of
phenylhydrazine, and to elute the edaravone bulk drug in the sample
solution in a short time, leading to any efficient HPLC
irrespective of involving in a two-step mobile phase.
[0165] From the above, it has been found that the method enables to
accurately, quantitatively and easily measure the phenylhydrazine
content in the edaravone bulk drug without performing the
complicated procedures such as degassing/nitrogen bubbling of
liquid for dissolution nor measurement immediately after preparing
the solution.
Example 2
[0166] Phenylhydrazine hydrochloride (13.517 mg, about 10 mg as
phenylhydrazine) was dissolved in a 0.1 mol/L hydrochloric
acid/acetonitrile mixture (volume ratio 7:3; hereinafter referred
to as Liquid for Dissolution), and the solution was adjusted
exactly to 100 mL. To the solution measured precisely to 2 mL,
Liquid for Dissolution was added, and the solution was adjusted
exactly to 20 mL. To the solution measured precisely to 2.5 mL,
Liquid for Dissolution was added, and the solution was adjusted
exactly to 100 mL to prepare a second standard solution
(phenylhydrazine: 0.25 .mu.g/mL).
[0167] Ten .mu.L of the second standard solution was precisely
measured and collected, and was subjected to the test by high
performance liquid chromatography under the following conditions,
followed by measuring the peak area of phenylhydrazine (measured
value of phenylhydrazine) in the solution according to the
automatic integration process.
Test Conditions
[0168] Degassing of liquid for dissolution/nitrogen bubbling:
None
[0169] Storage temperature of a second sample solution and a second
standard solution after preparation: 5.degree. C.
[0170] Vessel (preparation and storage of a second sample solution
and a second standard solution): Brown glass bottle
[0171] Detector: UV absorption photometer (measurement wavelength:
225 nm)
[0172] Column: Octadecyl silylated silica gel packed column
(YMC-Pack Pro C18 or an equivalent column)
[0173] Column temperature: Constant temperature around 40.degree.
C.
[0174] Mobile phase: Concentration gradient controlled by starting
from 100% mobile phase A up to 100% mobile phase B [0175] Mobile
phase A: 0.05 mol/L aqueous sodium perchlorate solution at pH
2.5/acetonitrile (volume ratio 9:1) mixture [0176] Mobile phase B:
0.05 mol/L aqueous sodium perchlorate solution at pH
2.5/acetonitrile (volume ratio 2:3) mixture [0177] 0.05 mol/L
aqueous sodium perchlorate solution at pH 2.5:7.0 g of sodium
perchlorate was dissolved in 1000 mL of water, perchloric acid was
added to the solution, and the pH of the resultant solution was
adjusted to 2.5.
[0178] The result of the stability study of phenylhydrazine in the
second standard solution is shown in Table 6.
TABLE-US-00006 TABLE 6 Stability of phenylhydrazine in the
solutions of a hydrochloric acid/acetonitrile mixture Peak area of
Elapsed phenylhydrazine Ratio to initial Solution time (h) (.mu.V
second) value (%) Second 0.00 10016 100 standard 1.02 9872 98.6
solution 2.04 9820 98.0 3.06 9693 96.8 5.09 9113 91.0 12.21 9328
93.1
[0179] As for the elapsed time (h), the time point at which the
second standard solution was at the beginning injected into the
HPLC within 30 minutes after preparing the solution was set as zero
"0" hour of the elapsed time, and the subsequent elapsed times were
calculated from the zero hour.
[0180] Further, when the second standard solution is used, the
phenylhydrazine content contained in the edaravone bulk drug in the
first sample solution can be calculated by Equation 4.
Content of phenylhydrazine in an edaravone bulk drug
(ppm)=M.sub.S/M.sub.T.times.A.sub.T/A.sub.S.times.0.25.times.0.748
Equation 4:
wherein M.sub.S: Weighed amount of phenylhydrazine hydrochloride in
a second standard solution (mg) M.sub.T: Weighed amount of an
edaravone bulk drug in a first sample solution (g) A.sub.T: Peak
area of phenylhydrazine in a first sample solution (.mu.V second)
A.sub.S: Peak area of phenylhydrazine in a second standard solution
at 0 hour of the elapsed time (.mu.V second) 0.748: Conversion
factor from phenylhydrazine hydrochloride into phenylhydrazine
[0181] The ratio of the peak area to the initial value of
phenylhydrazine in the second standard solution was 90% or more
even after 12 hours elapsed, showing that phenylhydrazine in the
solution was stable. In addition, it was revealed that the second
standard solution as prepared in this example was enabled to use in
the analysis method of the present invention, similarly to the
first standard solution as prepared in Example 1.
[0182] From this, it has been found that the method enables to
accurately, quantitatively and easily measure the phenylhydrazine
content in the edaravone bulk drug without performing complicated
procedures such as degassing/nitrogen bubbling of liquid for
dissolution nor measurement immediately after preparing the
solution.
Example 3
[0183] The stability of phenylhydrazine and/or
3-methyl-1-phenyl-2-pyrazolin-5-one in a solution was investigated
provided that an organic solvent in a liquid for dissolution was
changed from acetonitrile to methanol.
[0184] Mixtures of 0.1 mol/L hydrochloric acid test liquid and
methanol were prepared, and a liquid for dissolution was selected
so that the ratio of organic solvent became lower in the range
where EDVP was possibly dissolved (since the previous studies
showed that a peak shape was improved when the ratio of organic
solvent be close to that of the mobile phase). In accordance with
the conditions of Example 1 other than the liquids for dissolution,
the stability of phenylhydrazine and/or
3-methyl-1-phenyl-2-pyrazolin-5-one in the solutions was
investigated.
[0185] Sample solutions were prepared with the following three
liquids for dissolution.
[1] 0.1 mol/L hydrochloric acid/methanol (7:3) mixture [2] 0.1
mol/L hydrochloric acid/methanol (6:4) mixture [3] 0.1 mol/L
hydrochloric acid/methanol (5:5) mixture
[0186] The results showed that EDVP was soluble in liquid for
dissolution [3]. Therefore, 0.1 mol/L hydrochloric acid/methanol
(5:5) mixture was used to prepare the addition sample solution
(edaravone bulk drug: 0.10111 g, phenylhydrazine: 5 .mu.L), the
standard solution (phenylhydrazine hydrochloride: 13.154 mg), and
the sample solution (edaravone bulk drug: 0.10275 g), and the
stability of the solutions was investigated.
[0187] The phenylhydrazine content in the edaravone bulk drug can
be determined by the method according to the calculation method
based on Equation 3.
[0188] The results of the stability study of phenylhydrazine in the
solution of 0.1 mol/L hydrochloric acid/methanol (5:5) mixture are
shown in Table 7 and FIG. 7. The phenylhydrazine in the solution
tended to be faster decomposed than the results of Example 1 as
prepared using 0.1 mol/L hydrochloric acid/acetonitrile (7:3)
mixture.
TABLE-US-00007 TABLE 7 Stability in the solutions using
hydrochloric acid/methanol mixtures Elapsed Peak area of
time*.sup.15 phenylhydrazine Ratio to initial Solution (h) (.mu.V
second) value (%) Addition 0.0 17853 100.0 sample 2.1 17343 97.1
solution 4.1 17057 95.5 Standard 0.0 34144 100.0 solution 2.1 33831
99.1 4.1 33406 97.8 10.3 31594 92.5 Sample 0.0 0 solution (Rot 2.1
0 No. 240010) 4.1 0
[0189] 15) As for the elapsed time (h), the time point at which the
addition sample solution, the standard solution and the sample
solution were at the beginning injected into the HPLC within 30
minutes after preparing the solutions was set as zero "0" hour of
the elapsed time, and the subsequent elapsed times were calculated
from the zero hour.
[0190] Phenylhydrazine is easily decomposed by light and oxygen.
Therefore, in order to prevent the decomposition of phenylhydrazine
in the solution, complicated procedures normally should be
conducted such as degassing/nitrogen bubbling of a liquid for
dissolution before the measurement, the measurement immediately
after preparing the solution and the like. However, according to
the embodiment of the present invention, it has been found that it
is not necessary to perform degassing and nitrogen bubbling of a
liquid for dissolution before the measurement, and that the
decomposition of phenylhydrazine in the solution is enabled to be
prevented for a long period of time.
[0191] The results of Examples 1 to 3 have demonstrated that the
analysis method based on the embodiments of the present invention
using a hydrochloric acid/acetonitrile mixture (liquid for
dissolution) and a hydrochloric acid/methanol mixture (liquid for
dissolution) enables to quantitatively and easily measure a
phenylhydrazine content in an edaravone bulk drug, without
performing degassing and nitrogen bubbling of a liquid for
dissolution, while preventing the decomposition of phenylhydrazine
and the decomposition of 3-methyl-1-phenyl-2-pyrazolin-5-one in a
solution for a long period of time.
[0192] Further modifications and variations of the present
invention can be made on the basis of the above teachings.
Therefore, it is to be understood that, within the scope of the
claims attached hereto, the invention can be practiced according to
any aspect other than the descriptions as specifically described
herein.
* * * * *