CN109342609B - Method for identifying chemical components of traditional Chinese medicine fluid extract of compound three-dimensional calcium D-pantothenate syrup - Google Patents
Method for identifying chemical components of traditional Chinese medicine fluid extract of compound three-dimensional calcium D-pantothenate syrup Download PDFInfo
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Abstract
The invention relates to the field of syrup quality detection, in particular to a method for identifying chemical components of a traditional Chinese medicine fluid extract of compound three-dimensional calcium D-pantothenate syrup; the method comprises the steps of establishing a traditional Chinese medicine fluid extract raw material for retrieval, establishing a chemical component database, separating an ion peak through a primary mass spectrum, obtaining an ion fragment through a secondary mass spectrum, analyzing information of the ion peak and the ion fragment and comparing the information with the chemical component database, so as to rapidly and accurately identify each chemical component in the traditional Chinese medicine fluid extract in the compound three-dimensional calcium D-pantothenate syrup.
Description
Technical Field
The invention relates to the field of syrup quality detection, and particularly relates to a method for identifying chemical components of a traditional Chinese medicine fluid extract of compound three-dimensional calcium D-pantothenate syrup.
Background
Compound three-dimensional calcium D-pantothenate syrup, also known as vitamin feier cream and vitamin cream, is composed of 5 western medicines of vitamin B1, vitamin B2, vitamin B6, nicotinamide and calcium pantothenate (also known as vitamin B5), and another 10 traditional Chinese medicines (radix Codonopsis, Atractylodis rhizoma, Poria, Coicis semen, semen euryales, rhizoma Dioscoreae, fructus crataegi, fructus Hordei Germinatus preparata, semen Nelumbinis, and semen lablab album) extract and maltose, and can be used for treating malnutrition and vitamin deficiency, and child anorexia. Because the chemical components of the traditional Chinese medicine fluid extract of the compound three-dimensional calcium D-pantothenate syrup are complex, the prior art does not report the identification of the chemical components of the traditional Chinese medicine fluid extract of the compound three-dimensional calcium D-pantothenate syrup, and the effective quality control of the traditional Chinese medicine fluid extract cannot be carried out.
Disclosure of Invention
The technical problem to be solved by the invention is as follows: provides a quick and accurate identification method for chemical components of the traditional Chinese medicine fluid extract of the compound three-dimensional calcium D-pantothenate syrup.
In order to solve the technical problems, the invention adopts the technical scheme that: a method for identifying chemical components of a traditional Chinese medicine fluid extract of compound three-dimensional calcium D-pantothenate syrup comprises the following steps:
s1: consulting the separated and identified chemical components in the codonopsis pilosula, the poria cocos, the lotus seeds, the coix seeds, the hawthorns, the white hyacinth beans, the bighead atractylodes rhizome, the malts, the gordon euryale seeds and the Chinese yams, and establishing a chemical component database, wherein the chemical component database comprises chemical names, molecular formulas, molecular weights and chemical structure information;
s2: performing primary mass spectrum on the traditional Chinese medicine fluid extract of the compound calcium D-pantothenate syrup, and calculating a molecular formula according to the molecular mass information of an accurate compound obtained by the molecular ion peak of the obtained mass spectrum;
s3: comparing the calculated molecular formula with the chemical name, molecular formula and molecular weight of a chemical component database, and preliminarily determining the chemical components of the molecular ion peak of the mass spectrum;
s4: performing collision induced dissociation on the separated mass spectrum molecular ion peak, and performing secondary mass spectrum cracking to obtain fragment ions corresponding to the compound;
s5: and comparing the obtained fragment ions with chemical structure information of a chemical composition database, and further determining the chemical composition of the molecular ion peak of the mass spectrum.
The invention has the beneficial effects that: according to the chemical component identification method of the traditional Chinese medicine fluid extract of the compound three-dimensional calcium D-pantothenate syrup, provided by the invention, the raw materials of the traditional Chinese medicine fluid extract are searched, a chemical component database is established, ion peaks are separated through a primary mass spectrum, ion fragments are obtained through a secondary mass spectrum, and the chemical components in the traditional Chinese medicine fluid extract of the compound three-dimensional calcium D-pantothenate syrup are quickly and accurately identified through the analysis of the information of the ion peaks and the ion fragments and the comparison with the chemical component database.
Drawings
FIG. 1 is a positive ion flow diagram of UHPLC-Q active orbital hydrazine high-resolution mass spectrum of a compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract sample;
FIG. 2 is a negative ion flow diagram of UHPLC-Q active orbital hydrazine high-resolution mass spectrum of a compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract sample.
Detailed Description
In order to explain technical contents, achieved objects, and effects of the present invention in detail, the following description is made with reference to the accompanying drawings in combination with the embodiments.
The most key concept of the invention is as follows: the method comprises the steps of establishing a traditional Chinese medicine fluid extract as a raw material for retrieval, establishing a chemical component database, separating an ion peak through a primary mass spectrum, obtaining an ion fragment through a secondary mass spectrum, and rapidly and accurately identifying each chemical component in the traditional Chinese medicine fluid extract in the compound three-dimensional calcium D-pantothenate syrup through the analysis of the information of the ion peak and the ion fragment and the comparison with the chemical component database.
The invention relates to a method for identifying chemical components of a traditional Chinese medicine fluid extract of compound three-dimensional D-calcium pantothenate syrup, which comprises the following steps:
s1: consulting the separated and identified chemical components in the codonopsis pilosula, the poria cocos, the lotus seeds, the coix seeds, the hawthorns, the white hyacinth beans, the bighead atractylodes rhizome, the malts, the gordon euryale seeds and the Chinese yams, and establishing a chemical component database, wherein the chemical component database comprises chemical names, molecular formulas, molecular weights and chemical structure information;
s2: performing primary mass spectrum on the traditional Chinese medicine fluid extract of the compound calcium D-pantothenate syrup, and calculating a molecular formula according to the molecular mass information of an accurate compound obtained by the molecular ion peak of the obtained mass spectrum;
s3: comparing the calculated molecular formula with the chemical name, molecular formula and molecular weight of a chemical component database, and preliminarily determining the chemical components of the molecular ion peak of the mass spectrum;
s4: performing collision induced dissociation on the separated mass spectrum molecular ion peak, and performing secondary mass spectrum cracking to obtain fragment ions corresponding to the compound;
s5: and comparing the obtained fragment ions with chemical structure information of a chemical composition database, and further determining the chemical composition of the molecular ion peak of the mass spectrum.
According to the chemical component identification method of the traditional Chinese medicine fluid extract in the compound three-dimensional calcium D-pantothenate syrup, the raw materials of the traditional Chinese medicine fluid extract are searched, a chemical component database is established, ion peaks are separated through a primary mass spectrum, ion fragments are obtained through a secondary mass spectrum, and the chemical components in the traditional Chinese medicine fluid extract in the compound three-dimensional calcium D-pantothenate syrup are quickly and accurately identified through analysis of information of the ion peaks and the ion fragments and comparison with the chemical component database.
Further, in the method for identifying chemical components of the compound three-dimensional D-calcium pantothenate syrup traditional Chinese medicine fluid extract, the mass spectrum conditions of the primary mass spectrum and the secondary mass spectrum are as follows:
the Q-active four-stage rod-electrostatic field orbit trap high-resolution mass spectrum adopts two modes of positive ions and negative ions of an electrospray ion source: the spray voltage in the positive ion mode was 3.8kV, the spray voltage in the negative ion mode was 3.1kV, the capillary temperature was 320 ℃, the sheath gas flow rate was 45psi, the auxiliary gas flow rate was 15psi, the mass-to-charge ratio window width was set to 3, and the collision energy gradient was the NCE mode: 20,40,60eV, and the scanning rate is 7 Hz; collecting mass spectrum measurement data in a full scanning mode, wherein the data collection range is m/z 70-1000;
the resolution of the primary mass spectrum is 70000; the resolution of the secondary mass spectrum was 17500.
Further, in the method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract, the step S2 is specifically as follows:
s2: performing primary mass spectrum on the traditional Chinese medicine fluid extract of the compound calcium D-pantothenate syrup, and calculating the molecular formula in a mass deviation range of 5ppm through Xcaliibur 3.0 software according to the accurate compound molecular mass information obtained by the molecular ion peak of the obtained mass spectrum.
Further, in the method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract, the step S4 is specifically as follows:
s4: and (3) performing collision induced dissociation on the separated mass spectrum molecular ion peak, obtaining fragment ions corresponding to the compound through the cracking of a secondary mass spectrum, and analyzing the chemical structure information of the fragment ions by adopting MS DataAnlysis software.
Further, in the method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract, after the step S5, the method further comprises the following steps:
s6: preparing a reference substance solution of the identified chemical components, preparing a test solution of the compound three-dimensional calcium D-pantothenate syrup, and analyzing whether the retention time and MRM chromatograms of the identified chemical components in the test sample and the reference sample are consistent under the same chromatographic condition and mass spectrum condition.
As can be seen from the above description, the compound identified in S5 is purchased as a control for the corresponding compound, and a control solution is prepared, and the retention time of the identified chemical component in the test sample and the control and the MRM chromatogram are analyzed to be consistent under the same chromatographic conditions and mass spectrometry conditions as those of the test sample solution, thereby further accurately determining the identified chemical component.
Further, in the method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract, the chromatographic conditions in S6 are as follows:
using a Waters CORTECS UPLC C18 chromatographic column; taking acetonitrile solution as a mobile phase A and aqueous solution containing 0.1% formic acid as a mobile phase B, and performing gradient elution for 0-0.5 min by using 2% of A; 0.5-2.5 min, 2% A → 7% A; 2.5-4.5 min, 7% A → 13% A; 4.5-6.5 min, 13% A → 20% A; 6.5-9.5 min, 20% A → 55% A; 9.5-12.5 min, 55% A → 90% A; 12.5-13.5 min, 90% A; 13.5-13.6 min, 90% A → 2% A; 13.6-15 min, 2% A, flow rate of 0.25ml/min, column temperature of 45 ℃, and sample injection amount of 2 mul.
Further, in the method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract, the mass spectrum conditions in S6 are as follows:
monitoring multiple reactions by adopting an electrospray ion source positive ion mode; the capillary voltage is 3.5 KV; the flow rate of desolventizing nitrogen is 800L/h; the desolventizing temperature is 500 ℃; taper hole N2The airflow is 50L/h; the ion source temperature is 150 ℃; the secondary taper hole extraction voltage is 3.00V; the retention time is 5 ms; argon is used as collision gas; the carrier gas is nitrogen.
Further, in the method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract, the preparation method of the test solution in the S6 specifically comprises the following steps:
precisely measuring 1.0mL of compound three-dimensional calcium pantothenate syrup traditional Chinese medicine fluid extract, transferring into a 5mL measuring flask, diluting with 50% methanol water to scale, shaking, filtering with 0.22 μm microporous membrane, and collecting filtrate to obtain sample solution.
Example 1
A method for identifying chemical components of a traditional Chinese medicine fluid extract of compound three-dimensional calcium D-pantothenate syrup comprises the following steps:
s1: consulting the separated and identified chemical components in the codonopsis pilosula, the poria cocos, the lotus seeds, the coix seeds, the hawthorns, the white hyacinth beans, the bighead atractylodes rhizome, the malts, the gordon euryale seeds and the Chinese yams, and establishing a chemical component database, wherein the chemical component database comprises chemical names, molecular formulas, molecular weights and chemical structure information;
s2: performing primary mass spectrum on the traditional Chinese medicine fluid extract of the compound calcium D-pantothenate syrup, and calculating the molecular formula in a mass deviation range of 5ppm through Xcaliibur 3.0 software according to the accurate compound molecular mass information obtained by the molecular ion peak of the obtained mass spectrum;
s3: comparing the calculated molecular formula with the chemical name, molecular formula and molecular weight of a chemical component database, and preliminarily determining the chemical components of the molecular ion peak of the mass spectrum;
s4: performing collision induced dissociation on the separated mass spectrum molecular ion peak, performing secondary mass spectrum cracking to obtain fragment ions corresponding to the compound, and analyzing the chemical structure information of the fragment ions by adopting MS DataAnlysis software;
s5: comparing the obtained fragment ions with chemical structure information of a chemical composition database to determine chemical compositions of the molecular ion peaks of the mass spectrum;
s6: preparing a reference substance solution of the identified chemical components, preparing a test solution of the compound three-dimensional calcium D-pantothenate syrup, and analyzing whether the retention time and MRM chromatogram of the identified chemical components in the test substance and the reference substance are consistent under the same chromatographic condition and mass spectrum condition;
the preparation method of the test solution in the S6 specifically comprises the following steps:
precisely measuring 1.0mL of compound three-dimensional calcium pantothenate syrup traditional Chinese medicine fluid extract, transferring into a 5mL measuring flask, diluting with 50% methanol water to scale, shaking, filtering with 0.22 μm microporous membrane, and collecting filtrate to obtain sample solution.
The chromatographic conditions in S6 were as follows:
using a Waters CORTECS UPLC C18 chromatographic column; taking acetonitrile solution as a mobile phase A and aqueous solution containing 0.1% formic acid as a mobile phase B, and performing gradient elution for 0-0.5 min by using 2% of A; 0.5-2.5 min, 2% A → 7% A; 2.5-4.5 min, 7% A → 13% A; 4.5-6.5 min, 13% A → 20% A; 6.5-9.5 min, 20% A → 55% A; 9.5-12.5 min, 55% A → 90% A; 12.5-13.5 min, 90% A; 13.5-13.6 min, 90% A → 2% A; 13.6-15 min, 2% A, the flow rate is 0.25ml/min, the column temperature is 45 ℃, and the sample injection amount is 2 mul;
the mass spectrometry conditions in S6 are as follows:
monitoring multiple reactions by adopting an electrospray ion source positive ion mode; the capillary voltage is 3.5 KV; the flow rate of desolventizing nitrogen is 800L/h; the desolventizing temperature is 500 ℃; the airflow of the taper hole N2 is 50L/h; the ion source temperature is 150 ℃; the secondary taper hole extraction voltage is 3.00V; the retention time is 5 ms; argon is used as collision gas; the carrier gas is nitrogen;
example 2
Taking chlorogenic acid as an example: firstly, according to a molecular ion peak 353.0877 given by a first-order mass spectrum, a mass spectrum software calculates the accurate molecular formula to be C16H18O9, and then the molecular mass of chlorogenic acid and an isomer thereof is consistent with one of the molecular mass of the chlorogenic acid and the isomer thereof by comparing with a chemical component database of the traditional Chinese medicine fluid extract of the compound three-dimensional calcium D-pantothenate syrup, so that the chlorogenic acid or the isomer thereof is preliminarily presumed to be the chlorogenic acid or the isomer thereof. And according to the fact that TOF-MS secondary fragment ions have M/z191, 179, 173, 161 and 135, the compound is consistent with a secondary mass spectrum ion of chlorogenic acid reported in the literature, and the M/z191 peak on the secondary mass spectrum is estimated to be [ M-H-cafefoyl ] -, the M/z179 peak is [ cafefeic acid-H ] -, the M/z173 peak is [ M-H-cafefeoyl-H2O ] -, the M/z161 peak is [ cafefeic acid-H2O-H ] -, the M/z135 peak is [ M-H-cafefeoyl-CO 2] -, and the chromatographic retention time, the molecular ion peak and the secondary mass spectrum fragment are completely consistent with a reference product, so that the compound is identified as chlorogenic acid. The other compounds were identified by similar methods, and finally 179 compounds were identified, 34 of which were determined by comparison with a control.
Referring to fig. 1 and fig. 2, fig. 1 is a positive ion flow diagram of UHPLC-Q active orbital hydrazine high-resolution mass spectrum of a compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract sample; FIG. 2 is a negative ion flow diagram of UHPLC-Q active orbital hydrazine high-resolution mass spectrum of a compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract sample.
Referring to table 1, table 1 shows some chemical components (179 compounds in total, which are not listed herein) identified in the compound three-dimensional calcium pantothenate syrup Chinese medicinal fluid extract;
TABLE 1 identification of chemical components of compound three-dimensional calcium pantothenate syrup Chinese medicinal fluid extract
MS DataAnlysis is adopted to analyze the secondary mass spectrum cracking fragments, the cracking characteristics of chemical components of the fragments are presumed through high-resolution data, the chemical cracking rules of various identified compounds are summarized, and the mass spectrum cracking process is as follows (taking liensinine as an example):
example 3
In a method for identifying chemical components of a fluid extract of a compound three-dimensional calcium D-pantothenate syrup according to example 1, the reference substance includes: ferulic acid, butin, catechin, ellagic acid, vitexin, orientin, vitexin rhamnoside, protocatechuic aldehyde, gallic acid, caffeic acid, ethyl gallate, apigenin, oroxylin, kaempferol, luteolin, quercetin, orcinol glucoside, 1-caffeoylquinic acid, neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, vitexin, phloxine, phlorizin, luteolin, quercitrin, isoquercitrin, isochlorogenic acid B, isochlorogenic acid A, isochlorogenic acid C, schaftoside, isoschaftoside, kaempferol-3-O-rutinoside, rutin and narcissin, and adding methanol respectively to prepare a reference solution of 10 μ g/ml.
The retention time of the chemical components identified in S6 in the test sample and the control sample, and the alignment of MRM chromatogram are as follows (34 compounds in total, the alignment of the disclosed partial compounds is as follows):
TABLE 2 comparison of test and control samples for some of the compounds
In summary, in the method for identifying chemical components of the traditional Chinese medicine fluid extract in the compound three-dimensional calcium pantothenate syrup provided by the invention, the raw materials of the traditional Chinese medicine fluid extract are searched, a chemical component database is established, the ion peak is separated by the primary mass spectrum, the ion fragment is obtained by the secondary mass spectrum, and the chemical components in the traditional Chinese medicine fluid extract in the compound three-dimensional calcium pantothenate syrup can be quickly and accurately identified by analyzing the information of the ion peak and the ion fragment and comparing the information with the chemical component database.
The above description is only an embodiment of the present invention, and not intended to limit the scope of the present invention, and all equivalent changes made by using the contents of the present specification and the drawings, or applied directly or indirectly to the related technical fields, are included in the scope of the present invention.
Claims (6)
1. A method for identifying chemical components of a traditional Chinese medicine fluid extract of compound three-dimensional calcium D-pantothenate syrup is characterized by comprising the following steps:
s1: consulting the separated and identified chemical components in the codonopsis pilosula, the poria cocos, the lotus seeds, the coix seeds, the hawthorns, the white hyacinth beans, the bighead atractylodes rhizome, the malts, the gordon euryale seeds and the Chinese yams, and establishing a chemical component database, wherein the chemical component database comprises chemical names, molecular formulas, molecular weights and chemical structure information;
s2: performing primary mass spectrum on the traditional Chinese medicine fluid extract of the compound calcium D-pantothenate syrup, and calculating a molecular formula according to the molecular mass information of an accurate compound obtained by the molecular ion peak of the obtained mass spectrum;
s3: comparing the calculated molecular formula with the chemical name, molecular formula and molecular weight of a chemical component database, and preliminarily determining the chemical components of the molecular ion peak of the mass spectrum;
s4: performing collision induced dissociation on the separated mass spectrum molecular ion peak, and performing secondary mass spectrum cracking to obtain fragment ions corresponding to the compound;
s5: comparing the obtained fragment ions with chemical structure information of a chemical composition database to determine chemical compositions of the molecular ion peaks of the mass spectrum;
s6: preparing a reference substance solution of the identified chemical components, preparing a test solution of the compound three-dimensional calcium D-pantothenate syrup, and analyzing whether the retention time and MRM chromatogram of the identified chemical components in the test substance and the reference substance are consistent under the same chromatographic condition and mass spectrum condition; the chromatographic conditions were as follows:
using a Waters CORTECS UPLC C18 chromatographic column; taking acetonitrile solution as a mobile phase A and aqueous solution containing 0.1% formic acid as a mobile phase B, and performing gradient elution for 0-0.5 min by using 2% of A; 0.5-2.5 min, 2% A → 7% A; 2.5-4.5 min, 7% A → 13% A; 4.5-6.5 min, 13% A → 20% A; 6.5-9.5 min, 20% A → 55% A; 9.5-12.5 min, 55% A → 90% A; 12.5-13.5 min, 90% A; 13.5-13.6 min, 90% A → 2% A; 13.6-15 min, 2% A, flow rate of 0.25ml/min, column temperature of 45 ℃, and sample injection amount of 2 mul.
2. The method for identifying chemical components in the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract according to claim 1, wherein the mass spectrum conditions of the primary mass spectrum and the secondary mass spectrum are as follows:
the Q-active four-stage rod-electrostatic field orbit trap high-resolution mass spectrum adopts two modes of positive ions and negative ions of an electrospray ion source: the spray voltage in the positive ion mode was 3.8kV, the spray voltage in the negative ion mode was 3.1kV, the capillary temperature was 320 ℃, the sheath gas flow rate was 45psi, the auxiliary gas flow rate was 15psi, the mass-to-charge ratio window width was set to 3, and the collision energy gradient was the NCE mode: 20,40,60eV, and the scanning rate is 7 Hz; collecting mass spectrum measurement data in a full scanning mode, wherein the data collection range is m/z 70-1000;
the resolution of the primary mass spectrum is 70000; the resolution of the secondary mass spectrum was 17500.
3. The method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract according to claim 1, wherein S2 specifically comprises:
s2: performing primary mass spectrum on the traditional Chinese medicine fluid extract of the compound calcium D-pantothenate syrup, and calculating the molecular formula in a mass deviation range of 5ppm through Xcaliibur 3.0 software according to the accurate compound molecular mass information obtained by the molecular ion peak of the obtained mass spectrum.
4. The method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract according to claim 1, wherein S4 specifically comprises:
s4: and (3) performing collision induced dissociation on the separated mass spectrum molecular ion peak, performing secondary mass spectrum cracking to obtain fragment ions corresponding to the compound, and analyzing the chemical structure information of the fragment ions by adopting MSDAANLYSIS software.
5. The method for identifying chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract according to claim 1, wherein the mass spectrometry conditions in S6 are as follows:
monitoring multiple reactions by adopting an electrospray ion source positive ion mode; the capillary voltage is 3.5 KV; the flow rate of desolventizing nitrogen is 800L/h; the desolventizing temperature is 500 ℃; the airflow of the taper hole N2 is 50L/h; the ion source temperature is 150 ℃; the secondary taper hole extraction voltage is 3.00V; the retention time is 5 ms; argon is used as collision gas; the carrier gas is nitrogen.
6. The method for identifying the chemical components of the compound three-dimensional calcium D-pantothenate syrup traditional Chinese medicine fluid extract according to claim 1, wherein the preparation method of the test solution in S6 specifically comprises the following steps:
precisely measuring 1.0mL of compound three-dimensional calcium pantothenate syrup traditional Chinese medicine fluid extract, transferring into a 5mL measuring flask, diluting with 50% methanol water to scale, shaking, filtering with 0.22 μm microporous membrane, and collecting filtrate to obtain sample solution.
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