CN112903834A - Detection method for morpholine residue in bulk drug and application thereof - Google Patents
Detection method for morpholine residue in bulk drug and application thereof Download PDFInfo
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- 238000001514 detection method Methods 0.000 title claims abstract description 17
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- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical group OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 99
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- KJHOZAZQWVKILO-UHFFFAOYSA-N N-(diaminomethylidene)-4-morpholinecarboximidamide Chemical compound NC(N)=NC(=N)N1CCOCC1 KJHOZAZQWVKILO-UHFFFAOYSA-N 0.000 description 2
- ZKXDGKXYMTYWTB-UHFFFAOYSA-N N-nitrosomorpholine Chemical compound O=NN1CCOCC1 ZKXDGKXYMTYWTB-UHFFFAOYSA-N 0.000 description 2
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- MTJGVAJYTOXFJH-UHFFFAOYSA-N 3-aminonaphthalene-1,5-disulfonic acid Chemical compound C1=CC=C(S(O)(=O)=O)C2=CC(N)=CC(S(O)(=O)=O)=C21 MTJGVAJYTOXFJH-UHFFFAOYSA-N 0.000 description 1
- UIKUBYKUYUSRSM-UHFFFAOYSA-N 3-morpholinopropylamine Chemical compound NCCCN1CCOCC1 UIKUBYKUYUSRSM-UHFFFAOYSA-N 0.000 description 1
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- MKQLBNJQQZRQJU-UHFFFAOYSA-N morpholin-4-amine Chemical compound NN1CCOCC1 MKQLBNJQQZRQJU-UHFFFAOYSA-N 0.000 description 1
- FLCWYEUDIOQXEB-UHFFFAOYSA-N morpholin-4-yl(phenyl)methanone Chemical compound C=1C=CC=CC=1C(=O)N1CCOCC1 FLCWYEUDIOQXEB-UHFFFAOYSA-N 0.000 description 1
- 208000010805 mumps infectious disease Diseases 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- 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
- G01N30/02—Column chromatography
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- 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
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- 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
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
- G01N2030/067—Preparation by reaction, e.g. derivatising the sample
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- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Heterocyclic Carbon Compounds Containing A Hetero Ring Having Nitrogen And Oxygen As The Only Ring Hetero Atoms (AREA)
Abstract
The invention discloses a detection method and application of morpholine residual quantity in a raw material medicine, and relates to the technical field of medicine analysis and detection.
Description
Technical Field
The invention relates to the technical field of drug analysis and detection, in particular to a detection method of morpholine residue in bulk drugs.
Background
Morpholine, also called morphine forest, is a colorless oily liquid at normal temperature, is mainly used for preparing rubber vulcanization accelerator, can be used for synthesizing a surfactant, a textile printing and dyeing auxiliary agent, medicine and pesticide, can be used as a metal corrosion inhibitor and an antirust agent, and is also a solvent for dye, resin, wax, shellac and the like, is usually used for producing moroxydine and moroxydine hydrochloride in the field of medicine, is a commonly used broad-spectrum antiviral drug, and can be used for treating diseases such as influenza, mumps, viral bronchitis, chicken pox, herpes, epidemic pinkeye and the like.
In 27.10.2017, in the initial reference of carcinogen lists published by international cancer research institution of world health organization, morpholine is listed in 3 types of carcinogen lists, the detection method of morpholine content is not accepted in ChP2015, USP42, BP2020 and JP17 national formulary and forum at present, the boiling point of morpholine is 128.3 ℃, the morpholine can be dissolved in water and dissolved in most organic solvents, and the detection can be carried out by gas chromatography, but the morpholine peak tailing is serious, the sensitivity is low, and the limit requirement cannot be met, so that a method for detecting morpholine residue with high sensitivity is necessary for further controlling the safety of products such as moroxydine hydrochloride and the like.
Disclosure of Invention
The invention aims to: the method can overcome the defects of serious tailing and low sensitivity when detecting the morpholine content by a gas chromatography.
The technical scheme adopted by the invention is as follows:
a detection method for morpholine residue in bulk drugs comprises the following steps:
1) dissolving a raw material medicine sample by using a diluent to obtain a diluted solution;
2) adding an acid-binding agent and a derivatization reagent into the diluted solution in the step 1) for derivatization reaction to obtain a morpholine derivative;
3) measuring the content of the morpholine derivative in the step 2) by adopting a high performance liquid chromatography, wherein the chromatographic conditions are as follows:
filling agent: octadecylsilane chemically bonded silica;
mobile phase: mobile phase A and mobile phase B;
a detector: an ultraviolet detector;
and (3) an elution mode: gradient elution;
detection wavelength: between 364nm and 384 nm.
Further, the diluent is methanol or ethanol.
Further, the acid-binding agent is triethylamine or sodium hydroxide.
Further, the derivatization reagent is 2, 4-dinitrofluorobenzene, and the concentration of the derivatization reagent is 0.16 mg/ml-1.9 mg/ml.
Further, the reaction temperature of the derivatization reaction in the step 2) is 55-75 ℃, and the reaction time is 30-120 minutes.
Further, in 3), the mobile phase a is an aqueous phosphoric acid solution, and the mobile phase B is acetonitrile.
Further, in 3), the mobile phase a is water, and the mobile phase B is acetonitrile.
Further, the proportion of the mobile phase A is 45-55%, and the proportion of the mobile phase B is 45-55%.
Further, the raw material medicines are N- (2-guanidino-imino methyl) -morpholine and moroxydine hydrochloride.
In summary, due to the adoption of the technical scheme, the invention has the beneficial effects that:
(1) the method adopts the pre-column derivatization high performance liquid chromatography to detect the morpholine content in the bulk drug, indirectly measures the morpholine content by a method for detecting the morpholine derivative content, solves the problems of serious morpholine peak tailing, low sensitivity and incapability of reaching the detection limit of a gas chromatography, has strong specificity and high sensitivity, is mild in derivatization reaction condition, can be carried out at room temperature to 75 ℃, and is rapid and complete in reaction; the generated derivatization product can be detected by using an ultraviolet detector, the maximum absorption exists in the range of 364nm-384nm, the maximum absorption wavelength is in a higher waveband, the interference of a solvent or related impurities is not easy to occur, the sensitivity is obviously improved, and the detection limit can reach 0.2 ng.
(2) The derivatization reagent adopted by the method is 2, 4-dinitrofluorobenzene, has low toxicity, is a common chemical intermediate and is easy to purchase.
Drawings
FIG. 1 shows the reaction scheme of morpholine derivatization.
FIG. 2 shows the UV absorption spectrum of morpholine derivative.
FIG. 3 is a liquid chromatogram of morpholine derivatives using different types of diluents.
FIG. 4 is a liquid chromatogram of morpholine derivatives using different types of acid-binding agents.
FIG. 5 is a liquid chromatogram of a morpholine derivative with different amounts of derivatizing agent.
FIG. 6 is an X-Y scattergram of the amount of derivatizing agent and the peak area of morpholine derivative.
FIG. 7 is a liquid chromatogram of a morpholine derivative at different derivatization times.
FIG. 8 is an X-Y scattergram of derivatization reaction time versus morpholine derivative peak area.
FIG. 9 is a liquid chromatogram of a morpholine derivative at different derivatization temperatures.
FIG. 10 is an X-Y scattergram of derivatization reaction temperature versus morpholine derivative peak area.
FIG. 11 is a liquid chromatogram of a blank solvent, a reference solution, and a sample solution under the same derivatization conditions.
FIG. 12 is a liquid chromatogram of morpholine derivatives from solutions of different concentrations of morpholine.
FIG. 13 is an X-Y scattergram of morpholine solution concentration and morpholine derivative peak area.
FIG. 14 is a liquid chromatogram of a morpholine derivative from a control solution after being left for various periods of time.
FIG. 15 is a liquid chromatogram of a morpholine derivative in a test solution taken for different periods of time.
FIG. 16 is a liquid chromatogram of morpholine derivatives from two different batches of crude drugs in a blank solvent and a reference solution.
Detailed Description
For the purpose of making the objects, technical solutions and advantages of the present invention more apparent, the present invention will be further described in detail with reference to the accompanying drawings and embodiments, it being understood that the specific embodiments described herein are only for the purpose of explaining the present invention and are not intended to limit the present invention.
Example 1
Determination of ultraviolet wavelength of morpholine derivative:
instruments and conditions: the Shimadzu LC-20A liquid chromatograph (DAD detector) was used, and ZORBAX SB-C18, 250mm × 4.6mm, 5 μm, column temperature 30 deg.C, flow rate of 1.0 ml/min, sample amount of 10 μ l, mobile phase A water, and mobile phase B acetonitrile were used as chromatographic column.
Gradient elution was used, the elution procedure is shown in table 1:
table 1 example 1 gradient elution procedure
| Time/min | Mobile phase A/%) | Mobile phase B/%) |
| 0-15 | 55 | 45 |
| 15-22 | 45 | 55 |
| 22-30 | 55 | 45 |
The specific implementation steps are as follows:
preparing a derivatization reagent: newly preparing, weighing 158mg of 2, 4-dinitrofluorobenzene, placing in a 10ml measuring flask, ultrasonically dissolving by using methanol, diluting to a scale, and shaking up to obtain the product;
② preparing 2mg/ml sodium hydroxide/methanol solution: weighing 0.1g of sodium hydroxide, placing the sodium hydroxide into a 50ml volumetric flask, adding methanol for ultrasonic dissolution, diluting the sodium hydroxide to a scale with the methanol, and shaking up to obtain the sodium hydroxide solution;
preparing a blank solvent: sequentially weighing 1ml of methanol, 0.1ml of 2mg/ml sodium hydroxide/methanol solution and 1ml of derivatization reagent, putting the materials into a same 10ml measuring flask, shaking up, heating the materials in a 65 ℃ water bath for 60min, taking out the materials, cooling the materials, diluting the materials to a scale with methanol, and shaking up to obtain the reagent;
preparing morpholine stock solution: taking about 1.0g of morpholine, precisely weighing, placing in a 100ml measuring flask, diluting to scale with methanol, and shaking up; precisely measuring 1ml, placing in a 50ml measuring flask, diluting with methanol to scale, and shaking; precisely measuring 5ml, placing in a 100ml measuring flask, diluting with methanol to scale, and shaking up (about 10 μ g/ml) to obtain;
preparing a reference solution: precisely measuring 1ml of morpholine stock solution, placing the morpholine stock solution into a 10ml measuring flask, sequentially adding 0.1ml of 2mg/ml sodium hydroxide/methanol solution and 1ml of derivatization reagent, shaking up, heating the mixture in a 65 ℃ water bath for 60min, taking out the mixture, cooling the mixture, diluting the mixture to a scale with methanol, and shaking up to obtain the morpholine stock solution;
sixthly, chromatographic analysis: precisely measuring blank solvent and reference solution, respectively 10 μ l, injecting into chromatograph, and recording chromatogram, the result is shown in figure 2.
FIG. 2 shows the UV absorption spectrum of the derivative, from which it can be seen that the derivative has UV absorption in the range of 364nm-384nm and the maximum absorption wavelength is 374 nm.
Example 2
Investigation of different diluents:
instruments and conditions: using an Agilent type 1260II liquid chromatograph, column: agilent EC-C18, 150mm × 4.6mm, 4 μm, detection wavelength of 374nm, column temperature of 30 deg.C, flow rate of 1.0ml per minute, sample amount of 10 μ l, mobile phase A of 0.1% phosphoric acid solution, and mobile phase B of acetonitrile.
The gradient elution procedure is shown in table 2.
Table 2 example 2 gradient elution procedure
| Time/min | Mobile phase A/%) | Mobile phase B/%) |
| 0-10 | 60 | 40 |
| 10-11 | 45 | 55 |
| 11-15 | 25 | 75 |
| 15-25 | 60 | 40 |
The derivatization conditions are shown in Table 3.
Table 3 derivatization conditions of example 2
| Diluent | Amount of derivatizing agent/ml | Reaction temperature/. degree.C | Reaction time/min | Acid-binding agent |
| Methanol and | 1 | 60 | 30 | Sodium hydroxide |
Referring to example 1, the results are shown in FIG. 3, and the derivatization reaction was completed in the above diluent, and the peak area difference was small.
Example 3
Investigation of different acid-binding agents:
instruments and conditions: the same as in example 2.
The derivatization conditions are shown in Table 4.
Table 4 derivatization conditions of example 3
| Acid-binding agent | Amount of derivatizing agent/ml | Reaction temperature/. degree.C | Reaction time/min | Diluent | |
| Sodium hydroxide, | 1ml | 60℃ | 30 minutes | Methanol |
The results are shown in figure 4, and the two different acid-binding agents of sodium hydroxide and triethylamine can be used for complete reaction.
Example 4
Examination of the amount of derivatizing agent:
instruments and conditions: the same as in example 1.
The derivatization conditions are shown in Table 5.
Table 5 derivatization conditions of example 4
| Amount of derivatizing agent/ml | Diluent | Reaction temperature/. degree.C | Reaction time/min | Acid-binding agent |
| 0.1-1.2 | | 60℃ | 30 minutes | Sodium hydroxide |
The specific implementation steps are as described in example 1, and the results are shown in Table 6 and FIG. 5.
TABLE 6 relationship between amount of derivatizing agent and morpholine derivative peak area
| Amount of derivatizing agent/ml | Peak area of morpholine derivative |
| 0.1 | 2.03 |
| 0.2 | 4.30 |
| 0.3 | 6.55 |
| 0.5 | 9.72 |
| 0.8 | 10.93 |
| 1.0 | 11.73 |
| 1.2 | 12.08 |
As shown in fig. 5, the morpholine derivative peak area increases with increasing amounts of derivatizing agent.
Taking the dosage of the derivatization reagent as a horizontal coordinate and the peak area of the morpholine derivative as a vertical coordinate, making an X-Y scatter diagram and fitting a smooth curve, as shown in figure 6, when the dosage of the derivatization reagent is more than 1ml, the peak area of the derivative tends to be stable.
Example 5
Investigation of derivatization time:
instruments and conditions: the same as in example 1.
The derivatization conditions are shown in Table 7.
TABLE 7 derivatization conditions of example 5
| Reaction time/min | Amount of derivatizing agent/ml | Diluent | Reaction temperature/. degree.C | Acid-binding agent |
| 30-120 | 1 | | 60 | Sodium hydroxide |
The specific procedure was as described in example 1 and the results are shown in Table 8 and FIG. 7:
TABLE 8 derivatization time vs. morpholine derivative peak area
| Derivatization time/min | Peak area of |
| 30 | 11.54 |
| 60 | 15.29 |
| 90 | 16.81 |
| 120 | 17.33 |
As shown in FIG. 7, the peak area of morpholine derivative increased with the time of derivatization.
Taking the derivatization time as a horizontal coordinate and the morpholine derivative peak area as a vertical coordinate, making an X-Y scatter diagram and fitting a smooth curve, as shown in figure 8, when the derivatization reaction time is more than 60 minutes, the derivative peak area changes gently.
Example 6
Examination of derivatization temperature:
instruments and conditions: the same as in example 1.
The derivatization conditions are shown in Table 9.
TABLE 9 derivatization conditions of example 5
| Reaction temperature/. degree.C | Amount of derivatizing agent/ml | Diluent | Reaction time/min | Acid-binding agent | |
| 55-75 | | Methanol | 60 | Sodium hydroxide |
The specific procedure was as described in example 1, and the results are shown in Table 10 and FIG. 9:
TABLE 10 correlation of derivatization temperature with morpholine derivative peak area
| Derivatization temperature/. degree.C | Peak area of |
| 55 | 14.15 |
| 60 | 15.29 |
| 65 | 17.59 |
| 70 | 17.21 |
| 75 | 17.10 |
As shown in fig. 9, the morpholine derivative peak area increased with increasing derivatization temperature.
Taking the derivatization temperature as a horizontal coordinate and the morpholine derivative peak area as a vertical coordinate, making an X-Y scatter diagram and fitting a smooth curve, as shown in figure 10, when the derivatization reaction temperature is higher than 65 ℃, the morpholine derivative peak area has no obvious change.
Example 7
And (3) special investigation:
instruments and conditions: the same as in example 1.
The derivatization conditions are shown in Table 11.
TABLE 11 derivatization conditions of example 7
| Reaction temperature/. degree.C | Amount of derivatizing agent/ml | Diluent | Reaction time/min | Acid-binding |
| 65 | 1 | | 60 | Sodium hydroxide |
The specific implementation steps are as follows:
preparing a derivatization reagent, a 2mg/ml sodium hydroxide/methanol solution, a blank solvent, a morpholine stock solution and a reference solution: reference example 1;
preparing a test solution: precisely weighing 10mg of a raw material medicine sample by taking N- (2-guanidino-iminomethyl) -morpholine as a raw material medicine, placing the raw material medicine sample into a 10ml measuring flask, adding 1ml of methanol into the measuring flask for ultrasonic treatment for 5 minutes, sequentially adding 0.1ml of sodium hydroxide/methanol solution and 1ml of derivatization reagent, shaking up, heating the raw material medicine sample in a water bath at 65 ℃ for 60 minutes, taking out the raw material medicine sample, cooling the raw material medicine sample, diluting the raw material medicine sample to a scale by using methanol, and shaking up to obtain the finished product;
③ chromatographic analysis: precisely measuring blank solvent, reference solution and sample solution 10 μ l each, injecting into chromatograph, and recording chromatogram, the results are shown in Table 12 and figure 11.
TABLE 12 comparison of the retention times of control solution and test solution
| Name (R) | Retention time/min |
| Blank solvent | / |
| Control solution | 9.140 |
| Test solution | 9.144 |
Example 8
Linear and range investigation:
instruments and conditions: the same as in example 1.
The specific implementation steps are as follows:
preparing a derivatization reagent, a 2mg/ml sodium hydroxide/methanol solution, a blank solvent and a 10 mu g/ml morpholine stock solution: reference example 1;
chromatographic analysis: precisely measuring 0.1ml, 0.2ml, 0.5ml, 1.0ml and 1.5ml of morpholine stock solution with the concentration of 10 mu g/ml, respectively placing the morpholine stock solution into a 10ml measuring flask, sequentially adding 0.1ml of sodium hydroxide/methanol solution with the concentration of 2mg/ml and 1ml of derivatization reagent, shaking up, heating in a water bath with the temperature of 65 ℃ for 60min, taking out, cooling, diluting with methanol to scale, shaking up to obtain linear solutions (a) and (b), precisely measuring blank solvents and linear solutions (a) and (b) and (c) respectively injecting 10 mu l into a chromatograph, and recording a chromatogram, wherein the results are shown in a table 13 and an attached figure 12.
TABLE 13 correlation of morpholine solution concentration with morpholine derivative peak area
Taking the concentration of the morpholine solution as an abscissa and the peak area of the morpholine derivative as an ordinate, making an X-Y scattergram, and fitting a trend line, as shown in figure 13, it can be known that the concentration of the morpholine solution and the peak area are linearly related within the concentration range of 0.020-0.300 mug/ml, the correlation coefficient r is 0.9970, and the function formula of the trend line is that Y is 67.77X + 0.2744.
Example 8
And (3) solution stability investigation:
instruments and conditions: the same as in example 1.
The specific implementation steps are as follows:
preparing a derivatization reagent, a 2mg/ml sodium hydroxide/methanol solution, a blank solvent, a 10 mu g/ml morpholine stock solution and a reference solution: reference example 1;
preparing a test solution: refer to example 7.
③ chromatographic analysis: the reference solution and the sample solution are respectively placed for 1-12 hours, 10 mul of each of the sample solution and the reference solution are precisely measured and injected into a chromatograph, and chromatograms are recorded, wherein the results are shown in table 14 and figures 14-15.
TABLE 14 correlation of morpholine derivative peak area to solution standing time
| Standing time/hour | Peak area of control solution | Peak area of |
| 0 | 18.03 | 34.16 |
| 1 | 18.50 | 35.39 |
| 2 | 18.54 | 36.56 |
| 4 | 19.00 | 38.79 |
| 6 | 19.24 | 42.05 |
| 8 | 20.12 | 45.59 |
| 12 | 20.62 | 55.14 |
The peak areas of the reference solution and the test solution are gradually increased in the standing process, and the solutions are unstable, so that the solutions need to be newly prepared.
Example 9
Measuring the residual quantity of morpholine in a raw material medicine sample:
instruments and conditions: using an Agilent type 1260II liquid chromatograph, column: agilent EC-C18, 150mm 4.6mm, 4 μm, detection wavelength of 374nm, column temperature of 30 deg.C, flow rate of 1.0ml per minute, sample amount of 10 μ l, mobile phase A of water, and mobile phase B of acetonitrile.
The gradient elution procedure is shown in Table 15.
Table 15 example 9 gradient elution procedure
| Time/min | Mobile phase A/%) | Mobile phase B/%) |
| 0-10 | 60 | 40 |
| 10-11 | 45 | 55 |
| 11-15 | 25 | 75 |
| 15-25 | 60 | 40 |
The derivatization conditions are shown in Table 16.
Table 16 example 9 derivatization conditions
| Reaction temperature/. degree.C | Amount of derivatizing agent/ml | Diluent | Reaction time/min | Acid-binding |
| 65 | 1 | | 60 | Sodium hydroxide |
The specific implementation steps are as follows:
preparing a derivatization reagent, a 2mg/ml sodium hydroxide/methanol solution, a blank solvent, a morpholine stock solution and a reference solution: reference example 1;
preparing a test solution: taking two different batches of finished moroxydine hydrochloride products as raw material samples, precisely weighing 10mg of the raw material samples, putting the raw material samples into a 10ml measuring flask, adding 1ml of methanol for ultrasonic treatment for 5 minutes, sequentially adding 0.1ml of sodium hydroxide/methanol solution and 1ml of derivatization reagent, shaking up, heating the mixture in a water bath at 65 ℃ for 60 minutes, taking out the mixture, cooling the mixture, diluting the mixture to a scale with methanol, and shaking up to obtain the moroxydine hydrochloride;
③ chromatographic analysis: precisely measuring blank solvent, reference solution, and two sample solutions 10 μ l each, injecting into chromatograph, and recording chromatogram, the result is shown in figure 16;
fourthly, calculating the morpholine content in the raw material medicine sample:
measuring the peak areas of morpholine derivatives in a control solution and two test solutions, using C1Represents the concentration of morpholine in the control solution, S1Represents the peak area of morpholine derivative in the control solution, C2Represents the concentration of morpholine in the test solution, S2Represents the peak area of the morpholine derivative in the test solution, C2=C1×（S2/S1) The measurement and calculation results are shown in table 17.
TABLE 17 test results of control solutions and two crude drug samples
The raw material for synthesizing moroxydine is also the main raw material for synthesizing moroxydine hydrochloride, and the method for detecting the morpholine content in moroxydine hydrochloride is also suitable for detecting the morpholine content in moroxydine hydrochloride, and the specific implementation mode refers to example 9, and the detailed description is omitted.
In addition, the downstream products of morpholine such as 4-acetyl morpholine, tri (4-morpholino) phosphine, N- (3-aminopropyl) morpholine, N-nitrosomorpholine, N-formyl morpholine, 4-benzoyl morpholine, N-amino morpholine, etc. may also produce morpholine residue when morpholine is used as main raw material for producing downstream products.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents and improvements made within the spirit and principle of the present invention are intended to be included within the scope of the present invention.
Claims (9)
1. A detection method for morpholine residue in bulk drugs is characterized by comprising the following steps:
1) dissolving a raw material medicine sample by using a diluent to obtain a diluted solution;
2) adding an acid-binding agent and a derivatization reagent into the diluted solution in the step 1) for derivatization reaction to obtain a morpholine derivative;
3) measuring the content of the morpholine derivative in the step 2) by adopting a high performance liquid chromatography, wherein the chromatographic conditions are as follows:
filling agent: octadecylsilane chemically bonded silica;
mobile phase: mobile phase A and mobile phase B;
a detector: an ultraviolet detector;
and (3) an elution mode: gradient elution;
detection wavelength: between 364nm and 384 nm.
2. The method of claim 1, wherein the diluent is methanol or ethanol.
3. The method for detecting the residual quantity of morpholine in bulk drug according to claim 1, characterized in that the acid-binding agent is triethylamine or sodium hydroxide.
4. The method for detecting the residual quantity of morpholine in bulk drugs according to claim 1, wherein the derivatization reagent is 2, 4-dinitrofluorobenzene, and the concentration of the derivatization reagent is 0.16mg/ml to 1.9 mg/ml.
5. The method for detecting the residual quantity of morpholine in bulk drug according to claim 1, wherein the reaction temperature of the derivatization reaction in 2) is 55 ℃ to 75 ℃, and the reaction time is 30 minutes to 120 minutes.
6. The method for detecting the residual quantity of morpholine in bulk drug according to claim 1, characterized in that, in 3), the mobile phase A is phosphoric acid aqueous solution, and the mobile phase B is acetonitrile.
7. The method for detecting the residual quantity of morpholine in bulk drug according to claim 1, characterized in that, in 3), the mobile phase A is water, and the mobile phase B is acetonitrile.
8. The method for detecting morpholine residues in a bulk drug according to claims 6-8, wherein the proportion of mobile phase A is 45% -55%, and the proportion of mobile phase B is 45% -55%.
9. The use of the detection method according to claim 1, wherein the drug substance is N- (2-guanidino-iminomethyl) -morpholine or moroxydine hydrochloride.
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