CN114720605A - Preparation and detection method of azo compound - Google Patents

Preparation and detection method of azo compound Download PDF

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CN114720605A
CN114720605A CN202110005253.8A CN202110005253A CN114720605A CN 114720605 A CN114720605 A CN 114720605A CN 202110005253 A CN202110005253 A CN 202110005253A CN 114720605 A CN114720605 A CN 114720605A
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王仲清
许国彬
朱文民
廖守主
刘子健
罗会
杨小芹
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Sunshine Lake Pharma Co Ltd
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    • G01N30/02—Column chromatography
    • G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
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Abstract

The invention relates to a preparation and detection method of an azo compound, belonging to the field of pharmaceutical chemistry. The preparation method comprises the steps of carrying out catalytic hydrogenation reaction and reoxidation reaction on raw materials to prepare the target compound. The preparation, analysis and detection method provided by the invention can well obtain the target compound and detect and analyze the content of the target compound, thereby controlling the quality of the medicine and improving the safety of the medicine.

Description

Preparation and detection method of azo compound
Technical Field
The invention relates to the field of pharmaceutical chemistry, in particular to a preparation and detection method of an azo compound.
Background
Mirabegron (Mirabegron), chemical name is (R) -2- (2-aminothiazole-4-yl) -4' - [2- [ (2-hydroxy-2-phenylethyl) amino]Ethyl radical]Acetic acid anilides of formula C21H24N4O2S, the structure is as follows:
Figure BDA0002883057870000011
mirabegron is the first beta 3-adrenoceptor agonist drug to treat overactive bladder, and this drug is used to relax the detrusor smooth muscle during the storage phase of the bladder filling-voiding cycle, thus promoting increased bladder capacity, and is mainly used to treat overactive bladder (OAB) with symptoms of urge incontinence, urgency and urinary frequency.
Various impurities can be generated in the production process of the mirabegron, and the impurities with great harm, particularly genotoxic impurities and the like can bring harm to the human body, so that the potential safety hazard exists in the medicine. Therefore, a sufficient study of possible impurities is required.
Disclosure of Invention
The azo compound E and the azoxy compound C shown by the following formulas are impurities which may be generated in the production process of mirabegron. In order to fully study the detection method of impurities, the influence of impurities, and the like, it is necessary to prepare an impurity reference substance.
In a first aspect, the present invention provides a process for preparing mirabegron azo impurity compound E.
A method of preparing compound E comprising:
Figure BDA0002883057870000012
Figure BDA0002883057870000021
1) in the presence of hydrogen and a catalytic reagent, carrying out a reduction reaction on the compound C in a reaction solvent, filtering after the reaction is finished, and evaporating the filtrate to dryness to prepare a compound D;
2) under the protection of inert gas, mixing the compound D, sodium iodide, tert-butyl hypochlorite and acetonitrile, stirring for reaction, and carrying out aftertreatment to obtain a compound E.
In the step 1), the catalytic reagent is selected from one of palladium carbon and raney nickel. In some embodiments, the catalyst is palladium on carbon. In some embodiments, the catalyst is raney nickel.
In step 1), the mass ratio of the catalytic agent to the compound C may be 1:150 to 1: 250. In some embodiments, the mass ratio of the catalytic agent to compound C is 1: 200.
In the step 1), the reaction solvent is at least one selected from methanol, ethanol and ethyl acetate. In some embodiments, the reaction solvent is methanol. In some embodiments, the reaction solvent is ethanol. In some embodiments, the reaction solvent is ethyl acetate.
In the step 1), the mass-to-volume ratio of the compound C to the reaction solvent may be 0.03g/mL-0.08 g/mL. In some embodiments, the mass to volume ratio of compound C to reaction solvent is from 0.03g/mL to 0.05 g/mL. In some embodiments, the mass to volume ratio of compound C to reaction solvent is from 0.05g/mL to 0.08 g/mL. In some embodiments, the mass to volume ratio of compound C to reaction solvent is 0.03 g/mL. In some embodiments, the mass to volume ratio of compound C to reaction solvent is 0.05 g/mL. In some embodiments, the mass to volume ratio of compound C to reaction solvent is 0.08 g/mL.
In step 1), the reaction temperature of the reaction may be 20 ℃ to 40 ℃. In some embodiments, the reaction temperature is 30 ℃.
In the step 1), the reaction time may be 12h to 24 h. In some embodiments, the reaction time is 16 h. In some embodiments, the reaction time is 18 h. In some embodiments, the reaction time is 20 hours.
In the step 2), the inert gas is selected from one of nitrogen and argon. In some embodiments, the inert gas is nitrogen.
In step 2), the reaction temperature may be 20 ℃ to 50 ℃. In some embodiments, the reaction temperature in step 2) is 25 ℃, 30 ℃, or 35 ℃.
In the step 2), the reaction time can be 3h-8 h. In some embodiments, step 2) has a reaction time of 5 h.
In the step 2), the feeding molar ratio of the sodium iodide to the compound D is 1:1-3: 1. In some embodiments, the molar ratio of sodium iodide to compound D charged is 2: 1.
In the step 2), the charging molar ratio of the tert-butyl hypochlorite to the compound D can be 1:1-3: 1. In some embodiments, the charged molar ratio of tert-butyl hypochlorite to compound D is 2: 1.
In step 2), the post-treatment comprises: filtering the reaction liquid, pulping the filter cake and an alcohol solvent at the temperature of 60-90 ℃ for 0.1-2 h, filtering and drying to obtain the compound E. In some embodiments, the post-processing comprises: filtering the reaction liquid, pulping the filter cake and an alcohol solvent at the temperature of 60-80 ℃ for 0.5h, filtering and drying to obtain the compound E. In some embodiments, the post-processing comprises: filtering the reaction solution, pulping the filter cake with an alcohol solvent at 65 ℃ or 78 ℃ for 0.5h, filtering and drying to obtain the compound E.
The alcohol solvent is selected from one of methanol, ethanol and isopropanol. In some embodiments, the alcoholic solvent is methanol.
In some embodiments, a method of making compound E comprises: 1) in the presence of hydrogen and a catalytic reagent, carrying out a reduction reaction on the compound C in a reaction solvent, filtering after the reaction is finished, and carrying out rotary evaporation on the filtrate to obtain a compound D; wherein the catalytic agent is selected from one of palladium carbon and Raney nickel; the mass ratio of the catalytic reagent to the compound C is 1:150-1: 250; the reaction solvent is selected from one of methanol, ethanol and ethyl acetate; the mass-volume ratio of the compound C to the reaction solvent is 0.03g/mL-0.08 g/mL; the reaction time is within 12h-24 h; the reaction temperature of the reaction is 20-40 ℃; 2) under the protection of inert gas, mixing the compound D, sodium iodide, tert-butyl hypochlorite and acetonitrile, stirring for reaction, and carrying out aftertreatment to obtain a compound E; wherein the inert gas is one of nitrogen and argon; the reaction temperature is 20-50 ℃; the reaction time is 3-8 h; the post-treatment comprises the following steps: filtering the reaction solution, pulping the filter cake at 60-90 ℃ for 0.1-2 h by using an alcohol solvent, filtering and drying, wherein the alcohol solvent is selected from one of methanol, ethanol and isopropanol.
The method provided by the invention can conveniently obtain the compound E, and the purity of the obtained compound E (namely 1, 2-bis (4- (2- (((R) -2-hydroxy-2-phenylethyl) amino) ethyl) phenyl) azo) is more than 95.0 percent or 97.0 percent or 98.0 percent or 99.0 percent.
In the foregoing methods, compound C can be prepared by any suitable method.
In some embodiments, a method of making compound C comprises: under the protection of inert gas, reacting the compound A with a reducing agent in a reaction solvent to generate a compound B, after the reaction is finished, cooling to room temperature, filtering, then contacting the filtrate with air, stirring for reaction, after the reaction is finished, carrying out post-treatment to obtain a compound C; the inert gas is selected from one of nitrogen and argon. In some embodiments, a method of making compound C comprises: reacting the compound A in a reaction solvent in the presence of a catalytic reagent and hydrogen to generate a compound B, cooling to room temperature after the reaction is finished, filtering, contacting the filtrate with air, stirring for reaction, and performing post-treatment after the reaction is finished to obtain a compound C. The reaction formula is shown as the following formula:
Figure BDA0002883057870000031
in the method for preparing the compound C, the inert gas is at least one selected from nitrogen and argon. In some embodiments, the gas is nitrogen.
In the method for preparing the compound C, the reducing agents are zinc powder and ammonium chloride.
The feeding molar ratio of the ammonium chloride to the compound A can be 1:1-2: 1. In some embodiments, the molar feed ratio of ammonium chloride to compound a is from 1:1 to 1.5: 1. In some embodiments, the molar charge ratio of ammonium chloride to compound a is from 1.5:1 to 2: 1. In some embodiments, the molar feed ratio of ammonium chloride to compound a is 1: 1. In some embodiments, the molar feed ratio of ammonium chloride to compound a is 1.5: 1. In some embodiments, the molar feed ratio of ammonium chloride to compound a is 2: 1.
The feeding molar ratio of the zinc powder to the compound A can be 2:1-4: 1. In some embodiments, the zinc powder and compound a are fed in a molar ratio of 2:1 to 3: 1. In some embodiments, the zinc powder and compound a are charged in a molar ratio of 3:1 to 4: 1. In some embodiments, the zinc powder to compound a feed molar ratio is 2.5: 1. In some embodiments, the zinc powder to compound a feed molar ratio is 3: 1. In some embodiments, the zinc powder to compound a feed molar ratio is 3.5: 1.
In the method for preparing the compound C, the catalytic reagent is at least one of palladium carbon and Raney nickel. In some embodiments, the catalyst is palladium on carbon. In some embodiments, the catalyst is raney nickel.
In the method for preparing the compound C, the reaction solvent is at least one selected from water, methanol, ethanol and ethyl acetate. In some embodiments, the reaction solvent is water. In some embodiments, the reaction solvent is methanol. In some embodiments, the reaction solvent is ethanol. In some embodiments, the reaction solvent is ethyl acetate.
In the method for preparing the compound C, the mass-to-volume ratio of the compound A to the reaction solvent can be 0.05g/mL-0.15 g/mL. In some embodiments, the mass to volume ratio of compound a to reaction solvent is from 0.05g/mL to 0.10 g/mL. In some embodiments, the mass to volume ratio of compound a to reaction solvent is from 0.10g/mL to 0.15 g/mL. In some embodiments, the mass to volume ratio of compound a to reaction solvent is 0.08 g/mL. In some embodiments, the mass to volume ratio of compound a to reaction solvent is 0.10 g/mL. In some embodiments, the mass to volume ratio of compound a to reaction solvent is 0.12 g/mL.
In the method for preparing the compound C, the reaction temperature of the reaction can be 20-80 ℃ independently. In some embodiments, the reaction temperature is from 20 ℃ to 50 ℃. In some embodiments, the reaction temperature is from 20 ℃ to 65 ℃. In some embodiments, the reaction temperature is from 50 ℃ to 80 ℃. In some embodiments, the reaction temperature is from 50 ℃ to 65 ℃. In some embodiments, the reaction temperature is 65 ℃ to 80 ℃. In some embodiments, the reaction temperature is 25 ℃. In some embodiments, the reaction temperature is 50 ℃. In some embodiments, the reaction temperature is 65 ℃. In some embodiments, the reaction temperature is 75 ℃.
In the method for preparing the compound C, the stirring reaction time is 5-15 h. In some embodiments, the stirring reaction time is from 5h to 10 h. In some embodiments, the stirring reaction time is from 10h to 15 h. In some embodiments, the stirring reaction time is 8 hours. In some embodiments, the stirring reaction time is 11 hours. In some embodiments, the stirring reaction time is 13 hours.
In a process for preparing compound C, the post-treatment comprises: cooling the reaction liquid to room temperature, filtering, pulping a filter cake by using an alcohol solvent, filtering, and drying the obtained solid to obtain a compound C; the alcohol solvent is at least one selected from methanol, ethanol and isopropanol. In some embodiments, the organic solvent is methanol. In some embodiments, the organic solvent is ethanol. In some embodiments, the organic solvent is isopropanol.
In the present invention, the end point of the reaction can be monitored by liquid chromatography (HPLC); when the detection shows that the peak area of the reaction substrate is less than or equal to 1.0 percent, the reaction is considered to be finished.
In a second aspect, the invention provides a method for the analytical detection of compound E and compound C.
An analytical detection method for compound E and compound C comprising: and (3) detecting the sample solution by using HPLC-MS, wherein a chromatographic column with high-strength silica gel particles as a filler is adopted during detection, the mobile phase is divided into an A phase and a B phase, the A phase is an acid aqueous solution, and the B phase is acetonitrile.
The filler of the chromatographic column is high-strength silica gel particles (HSS). In some embodiments, the chromatography column may be Waters
Figure BDA0002883057870000051
HSS T3。
The aqueous acid solution is at least one selected from the group consisting of an aqueous formic acid solution and an aqueous acetic acid solution. In some embodiments, the aqueous acid solution is aqueous formic acid.
The volume ratio (V/V) of the acid to the water is 0.5:1000 to 1.5: 1000. In some embodiments, the volume ratio of acid to water (V/V) is 1: 1000.
The volume ratio of the phase A to the phase B in the elution procedure is 70-75: 30-25. In some embodiments, the volume ratio of the a phase to the B phase in the elution procedure is 73: 27.
In some embodiments, the method for detecting compounds E and C can comprise or be implemented by the following steps:
a) taking a proper amount of a sample to be detected, adding methanol for ultrasonic dissolution, fixing the volume to a scale, and shaking up to obtain a sample solution;
b) setting instrument parameters: detector, ion source, scanning mode, flow rate of mobile phase, sample volume, sample tray temperature, run time;
c) and b) injecting the sample solution obtained in the step a) into a high performance liquid chromatography-mass spectrometer to finish the analysis and detection of the compound E and the compound C.
In the step a), each 1mL of reaction solution can contain 0.2mg-1mg of sample to be detected. In some embodiments, step a) comprises 0.3mg, 0.5mg, or 0.8mg of the sample to be tested per 1mL of the reaction solution.
In step b), the detector is a triple quadrupole. The ion source is an AJS-ESI ion source. The scanning mode is MS2 SIM (+).
In step b), the ion polarity adopted by the ion source is positive ion mode, and the detector selects ions as m/z 509.4(+) and m/z 525.3 (+).
In step b), the flow rate of the mobile phase can be 0.8mL/min to 1.2 mL/min. In some embodiments, the flow rate of the mobile phase in step b) is 0.9 mL/min. In some embodiments, the flow rate of the mobile phase in step b) is 1.0 mL/min. In some embodiments, the flow rate of the mobile phase in step b) is 1.1 mL/min.
In the step b), the sample solution is injected into the sample container in an amount of 1 to 5 mu L. In some embodiments, the sample solution is introduced in an amount of 2 μ L. In some embodiments, the sample solution is loaded in an amount of 3 μ L. In some embodiments, the sample solution is loaded in an amount of 4 μ L.
In step b), the temperature of the sample tray is 5-30 ℃. In some embodiments, the sample tray temperature is 5 ℃ to 25 ℃. In some embodiments, the sample tray temperature is 10 ℃. In some embodiments, the sample tray temperature is 15 ℃. In some embodiments, the sample tray temperature is 20 ℃. In some embodiments, the sample tray temperature is 25 ℃.
In step b), the running time is not less than 8 min. In some embodiments, the run time is 8 min.
In some embodiments, the method for separating and detecting compound E and compound C comprises or can be achieved by:
d) taking a proper amount of a sample to be detected, adding methanol for ultrasonic dissolution, fixing the volume to a scale, and shaking up to obtain a sample solution;
b) setting the flow rate of a mobile phase to be 0.8-1.2 mL/min, setting the detector to be a triple quadrupole MS detector, setting the ion source to be an AJS-ESI ion source, setting the scanning mode to be MS2 SIM (+), setting the ion polarity of the ion source to be a positive ion mode, selecting the ions to be m/z 509.4(+) and m/z 525.3(+), setting the temperature of a sample tray to be 5-30 ℃, setting the sample feeding amount to be 1-5 muL, and setting the running time to be not less than 8 min;
c) and b) injecting the sample solution obtained in the step a) into a high performance liquid chromatography-mass spectrometer to complete the separation and determination of the compound E and the compound C.
The high performance liquid chromatography-mass spectrometry combination instrument (HPLC-MS) can be an Agilent 1260 type high performance liquid chromatography-mass spectrometry combination system and a workstation or other suitable and feasible systems.
In some embodiments, the method for separately detecting compound E and compound C comprises: detecting the sample solutions of compound E and compound C by HPLC-MS using a chromatographic column selected from Waters
Figure BDA0002883057870000061
HSS T3。
In some embodiments, the method for separately detecting compound E and compound C comprises: detecting the sample solutions of compound E and compound C by HPLC-MS using a chromatographic column selected from Waters
Figure BDA0002883057870000062
HSS T3; the mobile phase is divided into a phase A and a phase B, wherein the phase A is 0.1% formic acid aqueous solution, and the phase B is acetonitrile.
In some embodiments, the method for separately detecting compound E and compound C comprises: detecting the sample solutions of compound E and compound C by HPLC-MS using a chromatographic column selected from Waters
Figure BDA0002883057870000063
HSS T3; the mobile phase is divided into a phase A and a phase B, wherein the phase A is 0.1% formic acid aqueous solution, and the phase B is acetonitrile; the volume ratio (V/V) of formic acid to water in the aqueous formic acid solution in the phase A is 0.5:1000 to 1.5: 1000; the volume ratio of the A phase to the B phase was 73: 27.
By adopting the separation method, the time for separating and detecting the compound E and the compound C can be completed within 10 minutes.
Description of terms:
in the description herein, references to the description of the term "one embodiment," "some embodiments," "an example," "a specific example," or "some examples," etc., mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above are not necessarily intended to refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, various embodiments or examples and features of different embodiments or examples described in this specification can be combined and combined by one skilled in the art without contradiction.
In the present invention, the expression "compound a" and "compound represented by formula a" means the same compound.
The term "room temperature" means a temperature of about 20 ℃ to 35 ℃ or about 23 ℃ to 28 ℃ or about 25 ℃.
In the foregoing or following text, all numbers disclosed herein are approximate, regardless of whether the word "about" or "approximately" is used. The numerical value of each number may vary by 1%, 2%, 5%, 7%, 8%, or 10%.
According to the method, the purity of the reactant can be detected by adopting liquid chromatography (HPLC).
According to the method provided by the invention, by controlling appropriate reaction conditions, the purity of the obtained compound E (namely 1, 2-bis (4- (2- (((R) -2-hydroxy-2-phenylethyl) amino) ethyl) phenyl) azo) is more than 95.0 percent, or 97.0 percent, or 98.0 percent, or 99.0 percent.
Drawings
FIG. 1 shows the mass spectrum of Compound E in example 1;
FIG. 2 shows an HPLC-MS spectrum of a white solution in example 2, with the abscissa representing Acquisition Time (Acquisition Time) in units of minutes (min) and the ordinate representing response intensity (Counts);
FIG. 3 shows an HPLC-MS spectrum of a control solution in example 2, with Acquisition Time (Acquisition Time) on the abscissa and response intensity (Counts) in units of minutes (min) on the ordinate;
FIG. 4 shows an HPLC-MS spectrum of a detection limit solution in example 2, with Acquisition Time (Acquisition Time) on the abscissa and response intensity (Counts) in units of minutes (min) on the ordinate;
FIG. 5 shows an HPLC-MS spectrum of the test solution in example 2, with Acquisition Time (Acquisition Time) on the abscissa and response intensity (Counts) on the ordinate in units of minutes (min);
FIG. 6 shows an HPLC-MS spectrum of the sample-spiked solution of example 2, with Acquisition Time (Acquisition Time) on the abscissa and response intensity (Counts) on the ordinate.
Detailed Description
In order to make the technical solutions of the present invention better understood by those skilled in the art, some non-limiting examples are further disclosed below, and the present invention is further described in detail.
The reagents used in the present invention are either commercially available or can be prepared by the methods described herein.
In the embodiment of the present invention, the ratio (R) of the peak area of the standard-added impurity in the standard-added solution of the test sample to the average peak area of the corresponding impurity in the reference solution, which is related to the accuracy, is calculated as follows:
Figure BDA0002883057870000071
in the formula: a. theT+SAdding the peak area of the corresponding impurity in the standard solution to the sample;
ATthe peak area of the corresponding impurity in the test solution is shown;
ASthe average value of the peak areas of the corresponding impurities in 3-needle reference substance solutions;
WTweighing the sample amount of the sample in the sample solution, namely mg;
WT+Sthe sample amount is the sample weight of the sample in the standard solution.
In the invention, HPLC-MS represents the combination of high performance liquid chromatography and mass spectrometry; MS denotes a mass spectrometer detector; mL for mL, h for h, g for g, mg for mg, mg/mL for mg per mL, c for degrees celsius, HPLC for liquid chromatography.
For a further understanding of the present invention, reference will now be made in detail to the following examples.
Preparation of compound C:
Figure BDA0002883057870000081
the method comprises the following steps: compound A (12.47g, 1.0eq), and methanol (125mL, 10V) were added to a 250mL two-necked flask at room temperature, followed by addition of NH4Preparing a solution from Cl (3.1g, 1.5eq) and water (60mL, 5V) and zinc powder (7.58g, 3.0eq), performing nitrogen vacuum replacement for three times, heating to 65 ℃, reacting for 5 hours, monitoring by HPLC that the content of the compound A is less than 5.0%, cooling to room temperature after the reaction is finished, filtering, and transferring the filtrate to a 250mL single-port bottle; heating to 50 ℃, stirring for 10h, sampling, monitoring by HPLC (high performance liquid chromatography) that the content of the compound B is less than 1.0%, cooling to room temperature after the reaction is finished, filtering, pulping a filter cake by using methanol (50mL) for 1h, performing suction filtration, and drying a wet product at 60 ℃ in vacuum (-0.09Mpa) for 16h to obtain a compound C: 6.27g, purity 96.65% and yield 54.3%; and (3) detection: ESI-MS (M/z):525.3(M + H +);1H NMR(400MHz,DMSO-d6)δ9.18(brs,4H),8.22(d,J＝8.0Hz,2H),8.10(d,J＝8.0Hz,2H),7.53(d,J＝8.0Hz,2H),7.42(m,8H),7.33(m,2H),6.28(d,2H),5.15(d,2H),2.96-3.26(m,12H)。
the method 2 comprises the following steps: adding the compound A (10.0g, 1.0eq), 5% palladium carbon (0.3g) and water (100mL, 10V) into a 250mL high-pressure hydrogenation kettle at room temperature, performing nitrogen vacuum replacement for three times, then performing hydrogen replacement, pressurizing hydrogen to 2.5Mpa, heating to 50 ℃ for reaction for 1.5h, monitoring the compound A by HPLC (high performance liquid chromatography) to be less than 5.0%, cooling to room temperature, filtering, and transferring the filtrate into a 250mL single-port bottle; heating to 50 ℃, stirring for 10h, sampling, monitoring by HPLC (high performance liquid chromatography) that the content of the compound B is less than 1.0%, cooling to room temperature after conversion is finished, filtering, pulping a filter cake by using methanol (50mL) for 1h, performing suction filtration, and drying a wet product at 60 ℃ in vacuum (-0.09Mpa) for 16h to obtain a compound C: 4.76g, purity 95.92%, yield 51.4%.
Example 1
Figure BDA0002883057870000082
Adding a compound C (3g), 10% Pd/C (0.15g) and methanol (60mL) into a 100mL double-mouth bottle, performing vacuum replacement with hydrogen three times, increasing the hydrogen supply of a balloon to 30 ℃, supplementing hydrogen during the period, sampling for 24h, inspecting, detecting the content of the compound C by HPLC (high performance liquid chromatography) to be lower than 1.0%, after the reaction is finished, performing suction filtration, and performing rotary evaporation on the filtrate to obtain a solid, thus obtaining a compound D: 2.7g, purity 96.32%, yield 92.2%; ESI-MS (M/z):511.3(M + H +).
Figure BDA0002883057870000083
Adding a compound D (1.0g,1.0eq), sodium iodide (0.59g,2.0eq) and acetonitrile (30mL) into a 50mL double-mouth bottle, performing nitrogen vacuum replacement three times, adding t-BuOCl (0.43g,2.0eq) by using a syringe, stirring and reacting at room temperature (25 ℃) for 6 hours, sampling and inspecting, detecting that the reaction compound D is lower than 1.0% by HPLC (high performance liquid chromatography), performing suction filtration after the reaction is finished, leaching and draining acetonitrile (3mL), adding a wet product (1.05g) into methanol (10mL), thermally pulping (internal temperature 65 ℃) for 0.5 hour, filtering, adding a wet product (0.85g) into ethanol (8mL), thermally pulping for 0.5 hour (internal temperature 78 ℃), filtering, and drying the wet product at 60 ℃ under vacuum (-0.09MPa) for 14 hours to obtain a compound E: 0.53g, the purity is 97.78 percent, and the yield is 53.2 percent; and (3) detection: ESI-MS (M/z):509.3(M + H +);1H NMR(400MHz,DMSO-d6)δ9.15(brs,4H),7.87(brd,J＝8.0Hz,4H),7.50(brd,J＝8.0Hz,4H),7.42(m,8H),7.33(m,2H),6.23(d,2H),5.02(d,2H),3.03–3.268(m,12H)。
example 2
Apparatus and conditions
Figure BDA0002883057870000091
Experimental procedure
Mobile phase a phase: adding 1mL of formic acid into 1000mL of ultrapure water, uniformly mixing, and performing ultrasonic treatment for 10min to obtain a 0.1% formic acid aqueous solution;
mobile phase B phase: acetonitrile;
diluent/blank solution: methanol;
control stock solution 1: accurately weighing 37.5mg to 100mL of each of the compound C and the compound E in volumetric flasks, adding a proper amount of methanol, ultrasonically dissolving, and fixing the volume to a scale mark to obtain the compound E;
control stock solution 2: precisely transferring 1mL of reference stock solution into a volumetric flask of 1-50 mL, performing constant volume with methanol, and shaking up;
control stock solution 3: precisely transferring 1mL of reference substance stock solution into a 2-10 mL volumetric flask, performing constant volume with methanol, and shaking up;
control solution: precisely transferring 1mL of reference stock solution into a volumetric flask of 3-50 mL, performing constant volume with methanol, and shaking up;
detection limiting solution: precisely transferring 3mL of reference substance stock solution into a 10mL volumetric flask, performing constant volume by using methanol, and shaking up;
test solution: taking about 25mg of a test sample, precisely weighing, putting into a 50mL volumetric flask, dissolving methanol, fixing the volume, and shaking up;
adding a standard solution into a test sample: taking about 25mg of a test sample, precisely weighing the test sample into a 50mL volumetric flask, adding 1.0mL of reference substance stock solution 3, adding methanol for dissolving, and fixing the volume to obtain 100% test sample standard solution, and preparing 3 parts in parallel.
Taking blank solution, detection limit solution, reference solution, sample solution and sample adding solution, performing LC-MS analysis according to the following sequence table (Table A) and the above conditions, recording chromatogram, and showing the results in tables 1-6 as shown in figures 1-6.
TABLE A
Name of solution Number of sample introduction needles
Blank solution 1-2 needles
Detection limiting solution 1 needle
Control solution
3 needles
Test solution
1 needle
Sample adding solution 1 needle
Detection limiting solution 3 needles
Control solution for 1.56h 1 needle
Control solution for 2.56h 1 needle
Control solution for 3.84h 1 needle
The test solution is 1.15h 1 needle
The test solution is 2.15h 1 needle
The test solution is 3.15h 1 needle
Sample solution (batch number 1) 1 needle
Sample solution (batch 2) 1 needle
Sample solution (batch 3) 1 needle
Sample solution (batch number 4) 1 needle
Sample solution (batch number 5) 1 needle
Sample solution (batch number 6) 1 needle
Sample solution (batch number 7) 1 needle
Sample solution (batch number 8) 1 needle
Sample solution (batch number 9) 1 needle
Sample solution (batch number 10) 1 needle
Control solution
1 needle
And (4) analyzing results:
TABLE 1 System suitability results
Figure BDA0002883057870000101
Figure BDA0002883057870000111
TABLE 2 specificity results
Figure BDA0002883057870000112
TABLE 3 detectability results
Figure BDA0002883057870000113
Figure BDA0002883057870000121
TABLE 4 detection Limit results
Figure BDA0002883057870000122
TABLE 5 solution stability results
Figure BDA0002883057870000123
Figure BDA0002883057870000131
TABLE 6 results of sample testing
Batch number Compound E Compound C
Sample solution (batch number 1) Not detected out Not detected out
Sample solution (batch 2) Not detected out Not detected out
Sample solution (batch 3) Undetected Not detected out
Sample solution (batch 4) Not detected out Not detected out
Sample solution (batch number 5) Not detected out Not detected out
Sample solution (batch number 6) Not detected out Not detected out
Sample solution (batch number 7) Not detected out Not detected out
Sample solution (batch number 8) Not detected out Not detected out
Sample solution (batch number 9) Not detected out Not detected out
Sample solution (batch number 10) Not detected out Undetected
Detection Limit (LOD) 8.8ppm 8.8ppm
While the methods of the present invention have been described in terms of preferred embodiments, it will be apparent to those of ordinary skill in the art that variations and modifications of the methods and applications described herein, as well as other suitable variations and combinations, may be made to implement and use the techniques of the present invention within the context, spirit and scope of the invention. Those skilled in the art can modify the process parameters appropriately to achieve the desired results with reference to the disclosure herein. It is specifically noted that all such substitutions and modifications will be apparent to those skilled in the art and are intended to be included within the present invention.

Claims (10)

1. A method of making compound E comprising:
Figure FDA0002883057860000011
1) in the presence of hydrogen and a catalytic reagent, the compound C is subjected to a reduction reaction in a reaction solvent, after the reaction is finished, the filtering is carried out, and the filtrate is evaporated to dryness to prepare a compound D;
2) under the protection of inert gas, mixing the compound D, sodium iodide, tert-butyl hypochlorite and acetonitrile, stirring for reaction, and carrying out aftertreatment to obtain a compound E; the inert gas is at least one of nitrogen and argon.
2. The method of claim 1, the catalytic agent is selected from at least one of palladium on carbon, raney nickel; wherein the mass ratio of the catalytic agent to the compound C is 1:150-1: 250.
3. The process of claim 1, wherein the reaction solvent is selected from at least one of methanol, ethanol, ethyl acetate; the mass-to-volume ratio of the compound C to the reaction solvent is 0.03g/mL-0.08 g/mL.
4. The method according to claim 1, wherein the reaction temperature in the step 1) is 20-40 ℃, and the reaction time is 12-24 h; the reaction temperature in the step 2) is 20-50 ℃, and the reaction time is 3-8 h.
5. The process of claim 1, wherein the molar ratio of sodium iodide to compound D charged is from 1:1 to 3: 1; the feeding molar ratio of the tert-butyl hypochlorite to the compound D is 1:1-3: 1.
6. The method of claim 1, step 2), the post-processing comprising: filtering the reaction solution, pulping the filter cake and an alcohol solvent at the temperature of 60-90 ℃ for 0.1-2 h, filtering and drying; wherein the alcohol solvent is selected from one of methanol, ethanol and isopropanol.
7. The method of claim 1, further comprising: under the protection of inert gas, reacting the compound A with a reducing agent in a reaction solvent to generate a compound B, after the reaction is finished, cooling to room temperature, filtering, then contacting the filtrate with air, stirring for reaction, after the reaction is finished, carrying out post-treatment to obtain a compound C; the inert gas is selected from one of nitrogen and argon; the reducing agent is zinc powder and ammonium chloride; the feeding molar ratio of the ammonium chloride to the compound A is 1:1-2:1, and the feeding molar ratio of the zinc powder to the compound A is 2:1-4: 1;
or reacting the compound A in a reaction solvent in the presence of a catalytic reagent and hydrogen to generate a compound B, cooling to room temperature after the reaction is finished, filtering, contacting the filtrate with air, stirring for reaction, and performing post-treatment after the reaction is finished to obtain a compound C; the catalytic agent is at least one of palladium carbon and Raney nickel;
Figure FDA0002883057860000021
wherein the reaction solvent is selected from one of water, methanol, ethanol and ethyl acetate, the mass volume ratio of the compound A to the reaction solvent is 0.05g/mL-0.15g/mL, and the reaction temperature is 20-80 ℃ respectively and independently.
8. A method of analytically detecting compound E and compound C as described in claim 1, comprising: detecting a sample solution by HPLC-MS, wherein a chromatographic column with high-strength silica gel particles as a filler is adopted during detection, a mobile phase is divided into an A phase and a B phase, the A phase is an acid aqueous solution, and the B phase is acetonitrile; the acid aqueous solution is at least one of formic acid aqueous solution or acetic acid aqueous solution, and the volume ratio (V/V) of the acid to the water is 0.5:1000 to 1.5: 1000.
9. The method of claim 8, wherein the packing material of the chromatographic column is high-strength silica gel particles, and the volume ratio of the phase A to the phase B is 70-75: 30-25.
10. The method according to claim 8 or 9, comprising the steps of:
a) taking a proper amount of a sample to be detected, adding methanol for ultrasonic dissolution, fixing the volume to a scale, and shaking up to obtain a sample solution;
b) setting the flow rate of a mobile phase to be 0.8-1.2 mL/min, setting the detector to be a triple quadrupole MS detector, setting the ion source to be an AJS-ESI ion source, setting the scanning mode to be MS2 SIM (+), setting the ion polarity of the ion source to be a positive ion mode, selecting the ions to be m/z 509.4(+) and m/z 525.3(+), setting the temperature of a sample tray to be 5-30 ℃, setting the sample feeding amount to be 1-5 muL, and setting the running time to be not less than 8 min;
c) and b) injecting the sample solution obtained in the step a) into a high performance liquid chromatography-mass spectrometer to complete the separation and determination of the compound E and the compound C.
CN202110005253.8A 2021-01-05 2021-01-05 Preparation and detection method of azo compound Pending CN114720605A (en)

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