CN110514751A - A method of BHA content in measurement Bertha Luo Ting soft capsule - Google Patents
A method of BHA content in measurement Bertha Luo Ting soft capsule Download PDFInfo
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- CN110514751A CN110514751A CN201810490326.5A CN201810490326A CN110514751A CN 110514751 A CN110514751 A CN 110514751A CN 201810490326 A CN201810490326 A CN 201810490326A CN 110514751 A CN110514751 A CN 110514751A
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- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
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Abstract
The invention discloses a kind of methods of BHA content in measurement Bertha Luo Ting soft capsule, belong to drug quality detection field.This method detects BHA content in Bertha's Luo Ting soft capsule by HPLC method, comprising the following steps: (1) prepares BHA reference substance solution；(2) BHA sample solution to be measured is prepared；(3) by HPLC method, BHA characteristic peak is acquired, determines the content of BHA in Bertha's Luo Ting soft capsule；Wherein, in step (2) BHA sample solution to be measured the preparation method comprises the following steps: methanol is added into volumetric flask in the accurate uniformly mixed content of Bertha's Luo Ting soft capsule that weighs, ultrasound simultaneously shakes frequently, is cooled to room temperature to solution, with methanol constant volume, centrifugation, takes supernatant, filter membrane.By means of the present invention can effectively accurate detection go out BHA content in Bertha's Luo Ting soft capsule, high sensitivity is reproducible, and precision is high, and durability is good.
Description
Technical field
The invention belongs to drug quality detection fields, specifically, the present invention relates to a kind of measurement Bertha Luo Ting soft capsules
The method of middle BHA content.
Background technique
Bexarotene is antineoplastic, by Ligand drugmaker, the U.S. develop, on January 15th, 2000 the U.S. for the first time on
City is clinically mainly used for treating intractable and non-recalcitrant dermal t cell lymphoma (CTCL), needs to follow the doctor's advice and take this
Medicine.Bexarotene is a kind of vitamin A acid analog, and API is unstable, is easily oxidized.Butylated hydroxy anisole (BHA,
Butylated Hydroxyanisole) it is a kind of effective antioxidant, it is added in bexarotene soft capsule as auxiliary material
The oxidative degradation of API can effectively be inhibited.Different from the soft capsule of most of addition BHA at present, bexarotene content is to be suspended
Liquid, during measuring BHA, sample is needed by adequately pretreatment, its content of ability Accurate Determining.
Summary of the invention
The purpose of the present invention is to provide the methods of effective measurement BHA a kind of, and measure to containing for BHA in bexarotene
Calmly method validation is carried out, it was demonstrated that its feasibility, the measurement for BHA content in suspension provide reference frame.
To achieve the above object, described the present invention provides a kind of method of BHA content in measurement Bertha Luo Ting soft capsule
Method is to be detected by HPLC method to BHA content in Bertha's Luo Ting soft capsule, comprising the following steps:
(1) BHA reference substance solution is prepared；
(2) BHA sample solution to be measured is prepared；
(3) by HPLC method, BHA characteristic peak is acquired, determines the content of BHA in Bertha's Luo Ting soft capsule；
Wherein, in step (2) BHA sample solution to be measured the preparation method comprises the following steps: precision weighs the mixing of Bertha's Luo Ting soft capsule
Methanol is added into volumetric flask in uniform content, and ultrasound simultaneously shakes frequently, is cooled to room temperature to solution, with methanol constant volume, from
The heart takes supernatant, filter membrane.
In the present invention, the preparation method of BHA sample solution to be measured can be in the step (2) are as follows: precision weighs 5.6g shellfish
Into 50ml volumetric flask 30ml methanol is added, ultrasound simultaneously shakes frequently, to solution in the uniformly mixed content of Sha Luoting soft capsule
It is cooled to room temperature, with methanol constant volume, centrifugation takes supernatant, crosses 0.45 μm of filter membrane.
In the present invention, the chromatographic condition of the HPLC method further comprises: use specification for 4.6 × 150mm, 3.5 μm
Zorbax Eclipse Plus C18 chromatographic column；Mobile phase A is 0.1% glacial acetic acid: acetonitrile=50:50 (v/v), Mobile phase B are
0.1% glacial acetic acid: acetonitrile=10:90 (v/v)；Detector is UV detector (UV)；Flow rate of carrier gas is 1.0mL/min；Column temperature
It is 35 DEG C, wavelength 276nm, sampling volume is 20 μ l, runing time 29min.
Further, the HPLC method carries out gradient elution by the following conditions:
| Time/min | Mobile phase A/% | Mobile phase B/% |
| 0.0 | 100 | 0 |
| 4.0 | 100 | 0 |
| 16.0 | 0 | 100 |
| 25.0 | 0 | 100 |
| 25.1 | 100 | 0 |
| 29.0 | 100 | 0 |
In the present invention, in the step (1) BHA reference substance solution the preparation method comprises the following steps: precision weigh 20mg BHA control
Product dissolve in 200ml volumetric flask, with methanol and are diluted to scale, mix, and shift 10ml into 100ml volumetric flask, use methanol
Dilution is settled to scale.
In the present invention, in the step (2) ultrasonic time can be 30min, centrifugation can be at 3500rpm/min from
Heart 10min.
Method of the invention effectively accurate detection can go out BHA content in Bertha's Luo Ting soft capsule, high sensitivity, repeatability
Good, precision is high, and durability is good, and in addition to this, advantages of the present invention part will become apparent from the description below, or passes through
Practice of the invention obtains.
Detailed description of the invention
Fig. 1 is the liquid chromatogram of diluent methanol of the present invention；
Fig. 2 is the canonical plotting at the corresponding peak BHA of the present invention；
Fig. 3 is the canonical plotting at another the corresponding peak BHA of the present invention；
Fig. 4 is the liquid chromatogram of blank solution of the present invention；
Fig. 5 is the liquid chromatogram of reference substance solution of the present invention；
Fig. 6 is the liquid chromatogram of sample solution of the present invention.
Specific embodiment
In order to make the objectives, technical solutions, and advantages of the present invention clearer, with reference to the accompanying drawings and embodiments, right
The present invention is further elaborated.The embodiments described below for explaining only the invention, and should not be understood as to this hair
Bright limitation.Particular technique or condition are not specified in embodiment, according to the literature in the art described technology or conditions
Or it is carried out according to product description.Reagents or instruments used without specified manufacturer, being can be by commercially available acquisition
Conventional products.
In one aspect of the invention, the invention proposes a kind of methods of BHA content in measurement Bertha Luo Ting soft capsule.
The method that BHA content in measurement Bertha Luo Ting soft capsule is described in detail below according to the embodiment of the present invention.Reality according to the present invention
Example is applied, this method comprises:
Step (1): preparation reference substance solution
In this step, according to the following steps prepare BHA reference substance solution: precision weigh 20mg BHA reference substance in
In 200ml volumetric flask, scale is dissolved and be diluted to methanol, is mixed, and is shifted 10ml into 100ml volumetric flask, is used methanol dilution
It is settled to scale.
The concentration of BHA reference substance of the present invention is not particularly restricted, those skilled in the art can according to actual needs into
The concentration of row selection, a specific embodiment according to the present invention, BHA reference substance can be 10 μ g/ml, and inventor passes through experiment
It is measured in Bertha's Luo Ting soft capsule it was unexpectedly observed that selecting the BHA solution of the concentration that can significantly improve in following detection step
The accuracy of BHA content.
Step (2): testing sample solution is prepared
In this step, testing sample solution is prepared according to the following steps: taking 15~20 capsules, careful to cut, transfer
Content.Precision weighs the uniformly mixed content of 5.6g into 50ml volumetric flask, and 30ml methanol is added, and ultrasound simultaneously shakes frequently
30min.It is cooled to room temperature to solution, uses methanol constant volume.Sample is centrifuged, supernatant is taken, crosses 0.45 μm of filter membrane.
Step (3): acquisition BHA characteristic peak
In this step, the characteristic peak of BHA reference substance solution and BHA sample solution to be measured is acquired by liquid chromatography.
Specifically, use specification for 4.6 × 150mm, 3.5 μm of Zorbax Eclipse Plus C18 chromatographic column；Mobile phase A is
0.1% glacial acetic acid: acetonitrile=50:50 (v/v), Mobile phase B are 0.1% glacial acetic acid: acetonitrile=10:90 (v/v)；Detector is purple
External detector (UV)；Flow rate of carrier gas is 1.0mL/min；Column temperature is 35 DEG C, wavelength 276nm, and sampling volume is 20 μ l, when operation
Between be 29min；Gradient elution is carried out by the following conditions:
| Time/min | Mobile phase A/% | Mobile phase B/% |
| 0.0 | 100 | 0 |
| 4.0 | 100 | 0 |
| 16.0 | 0 | 100 |
| 25.0 | 0 | 100 |
| 25.1 | 100 | 0 |
| 29.0 | 100 | 0 |
Inventor is by experiment it was unexpectedly observed that the chromatographic condition in detecting step can significantly affect the shellfish that detection obtains
The accuracy of BHA content in the soft capsule of Bimbisara spit of fland.In view of this, inventor has therefrom determined above-mentioned chromatography by many experiments
Condition significantly improves the accuracy for measuring BHA content in bexarotene soft capsule as a result,.
Below with reference to specific embodiment, present invention is described, it should be noted that these embodiments are only to describe
Property, without limiting the invention in any way.
Embodiment 1: the investigation of pretreatment condition
Investigate the influence of ultrasound and centrifugation to BHA content in sample in the preparation process of sample solution.
(1) condition: ultrasonic 20min is centrifuged 5min at 3500rpm/min.
| Sample | Sample weighting amount/g | Peak area | BHA/% |
| It is not ultrasonic not to be centrifuged | 5.61600 | 88283 | 88.4 |
| Not ultrasonic centrifugation | 5.70420 | 95451 | 94.1 |
| Ultrasound is not centrifuged | 5.59086 | 92064 | 92.6 |
| Ultrasound centrifugation | 5.04770 | 85543 | 95.3 |
(2) condition: ultrasound 5min, 10min, 20min, 30min, 40min respectively；5min is centrifuged at 3500rpm/min.
| Ultrasonic time/min | Sample weighting amount/g | Peak area | BHA/% |
| 0 | 5.61600 | 88283 | 88.4 |
| 5 | 5.68145 | 90827 | 89.9 |
| 10 | 5.45522 | 89683 | 92.5 |
| 20 | 5.40930 | 90998 | 94.6 |
| 30 | 5.62520 | 97930 | 97.9 |
| 40 | 5.53801 | 96217 | 97.7 |
(3) condition: ultrasonic 30min, centrifugation rate are respectively 2000rpm/min, 3000rpm/min, 3500rpm/min,
4000rpm/min, 5000rpm/min, centrifugation time 5min.
(4) condition: ultrasonic 30min, centrifugation rate 3500rpm/min, centrifugation time are respectively 5min, 10min,
15min, 20min.
Pretreatment condition is determined as a result, are as follows: is centrifuged 10min under ultrasonic 30min, 3500rpm/min.
Embodiment 2:HPLC methodology validation
1. linear
1.1 linear stock solutions: precision weighs 10mg BHA in 100ml volumetric flask, is dissolved and is diluted with methanol and is settled to
Scale, the concentration of stock solution are 100 μ g/ml.
1.2 accordings to the form below prepare linear solvent, with methanol dilution constant volume
| Level of linearity | Stock solution volume (mL) | Constant volume (mL) | Final concentration (μ g/ml) |
| 20% | 2.0 | 100 | 2 |
| 50% | 5.0 | 100 | 5 |
| 100% | 5.0 | 50 | 10 |
| 150% | 15.0 | 100 | 15 |
| 200% | 10.0 | 50 | 20 |
2. precision and Intermediate precision
Two analysts prepare 6 parts of parallel samples respectively, measure the content of BHA, as a result well.
3. recycling
3.1 recycling stock solutions: precision weighs 10mg BHA in 100ml volumetric flask, is dissolved and is diluted with methanol and is settled to
Scale, the concentration of stock solution are 100 μ g/ml.
3.2 using auxiliary material as blank, according to the form below preparation recycling solution, with methanol dilution constant volume.
3.3 experimental result
4. durability
4.1 durability conditions
(1) flow velocity ± 0.1ml/min；
(2) column temperature ± 5 DEG C；
(3) Detection wavelength ± 2nm；
(4) mobile phase ratio ± 10%
4.2 result
The above results explanation, method of the invention can effectively accurate detection go out BHA content in bexarotene soft capsule, spirit
Sensitivity is high, reproducible, and precision is high, and durability is good.
As it will be easily appreciated by one skilled in the art that embodiment described above is only exemplary, not to limit this
Invention, any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in this hair
Within bright protection scope.
Claims (6)
1. a kind of method of BHA content in measurement Bertha Luo Ting soft capsule, which is characterized in that this method is by HPLC method to Bertha
BHA content is detected in Luo Ting soft capsule, comprising the following steps:
(1) BHA reference substance solution is prepared；
(2) BHA sample solution to be measured is prepared；
(3) by HPLC method, BHA characteristic peak is acquired, determines the content of BHA in Bertha's Luo Ting soft capsule；
Wherein, in step (2) BHA sample solution to be measured the preparation method comprises the following steps: precision weighs Bertha's Luo Ting soft capsule is uniformly mixed
Content into volumetric flask, methanol is added, ultrasound simultaneously shakes frequently, is cooled to room temperature to solution, with methanol constant volume, be centrifuged,
Take supernatant, filter membrane.
2. the method for BHA content in measurement Bertha Luo Ting soft capsule according to claim 1, which is characterized in that the step
Suddenly in (2) BHA sample solution to be measured the preparation method comprises the following steps: accurate weigh the uniformly mixed content of 5.6g Bertha's Luo Ting soft capsule
Into 50ml volumetric flask, 30ml methanol is added, ultrasound simultaneously shakes frequently, is cooled to room temperature to solution, with methanol constant volume, it is centrifuged,
Supernatant is taken, 0.45 μm of filter membrane is crossed.
3. the method for BHA content in measurement Bertha Luo Ting soft capsule according to claim 1, which is characterized in that described
The chromatographic condition of HPLC method further comprises: using specification for 4.6 × 150mm, 3.5 μm of Zorbax Eclipse Plus
C18 chromatographic column；Mobile phase A is 0.1% glacial acetic acid: acetonitrile=50:50 (v/v), and Mobile phase B is 0.1% glacial acetic acid: acetonitrile=
10:90 (v/v)；Detector is UV detector (UV)；Flow rate of carrier gas is 1.0mL/min；Column temperature is 35 DEG C, wavelength 276nm,
Sampling volume is 20 μ l, runing time 29min.
4. the method for BHA content in measurement Bertha Luo Ting soft capsule according to claim 3, which is characterized in that described
HPLC method carries out gradient elution by the following conditions:
。
5. the method for BHA content in measurement Bertha Luo Ting soft capsule according to claim 1, which is characterized in that the step
Suddenly in (1) BHA reference substance solution the preparation method comprises the following steps: precision weighs 20mgBHA reference substance in 200ml volumetric flask, use methanol
It dissolves and dilutes and release to scale, mix, shift 10ml into 100ml volumetric flask, be settled to scale with methanol dilution.
6. the method for BHA content in measurement Bertha Luo Ting soft capsule according to claim 1 or 2, which is characterized in that described
Ultrasonic time is 30min in step (2), and centrifugation is to be centrifuged 10min at 3500rpm/min.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2004233312A (en) * | 2003-02-03 | 2004-08-19 | Sumitomo Chem Co Ltd | B vitamin a, inducer of the same and analysis method of carotenoid |
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2018
- 2018-05-21 CN CN201810490326.5A patent/CN110514751A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2004233312A (en) * | 2003-02-03 | 2004-08-19 | Sumitomo Chem Co Ltd | B vitamin a, inducer of the same and analysis method of carotenoid |
Non-Patent Citations (6)
| Title |
|---|
| QIANG-SHENG XIE等: "Determination of synthetic antioxidants in fish oil soft", 《JOURNAL OF FOOD SAFETY AND QUALITY》 * |
| 佟晓波等: "丁基羟基茴香醚(BHA)原料含量及有关物质的检测方法的建立", 《山东化工》 * |
| 夏英姿等: "丁基羟基茴香醚的合成", 《精细石油化工进展》 * |
| 孙玉侠: "CCT软胶囊制剂研发及相关质量控制", 《中国优秀博硕士学位论文全文数据库(硕士)医药卫生科技辑》 * |
| 张红等: "超高效液相色谱法测定辛伐他汀胶囊中叔丁基-4-羟基茴香醚与2,6-二叔丁基对甲酚的含量", 《西北药学杂志》 * |
| 李杨杰等: "HPLC法测定丁基羟基茴香醚含量及其有关物质", 《广州化工》 * |
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Application publication date: 20191129 |