CN115184497A - Method for measuring content of 2, 4-epibrassinolide in dendrobium officinale - Google Patents
Method for measuring content of 2, 4-epibrassinolide in dendrobium officinale Download PDFInfo
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Abstract
The invention provides a method for measuring the content of 2, 4-epibrassinolide in dendrobium officinale, belonging to the technical field of pesticide residue detection. According to the invention, acetonitrile and water are used as extracting agents, ultrasonic extraction and vortex extraction are combined, and the extracted sample is purified by a solid phase extraction column, so that the matrix effect of the sample is reduced, and the operation is simpler than that of conventional derivatization treatment; according to the method, a proper amount of ammonium formate is added into the mobile phase, so that the ionization efficiency of the 2, 4-epibrassinolide can be improved, the obtained solution to be detected is further ensured to be suitable for high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) detection, and the 2, 4-epibrassinolide in the dendrobium officinale can be detected in a quantitative manner by an external standard method, so that the simple, rapid and sensitive detection of the 2, 4-epibrassinolide in the dendrobium officinale can be finally realized, and the method has important significance on the medication safety of the dendrobium officinale.
Description
Technical Field
The invention relates to the technical field of pesticide residue detection, in particular to a method for determining the content of 2, 4-epibrassinolide in dendrobium officinale.
Background
Dendrobium officinale (Dendrobium officinale Kimuraet Migo) is one of the earliest recorded orchidaceous herbaceous plants in China, is a traditional Chinese medicinal material with dual purposes of medicine and food, is called as 'Chinese sprangletop' and 'immortal grass', and has the effects of tonifying stomach, promoting fluid production, nourishing yin, clearing heat, reducing weight, prolonging life and the like. The dendrobium officinale has complex content ingredients and contains active ingredients such as polysaccharide, flavone, polyphenol and the like; the health food has the main effects of reducing blood sugar and blood fat, protecting gastrointestinal tract function, improving immunity, resisting tumor, etc. Wild dendrobium officinale with low yield is listed in national key protection wild plant directory (first batch), and is mainly planted artificially.
Brassinolide has the chemical formula of C 28 H 48 O 6 (see (a) in fig. 1) is a novel green and environment-friendly plant growth regulator, belonging to the sixth class of plant growth regulators, which can significantly increase the growth of plant nutrients and promote fertilization at low concentrations. The brassinolide content in natural compounds is extremely low, the extraction process is complex, and the actual production requirements cannot be met, so that the artificial synthesis of the brassinolide is a trend. At present, 2, 4-epibrassinolide (shown as (b) in figure 1) is most widely applied among the registered synthetic brassinolides in China due to high synthesis efficiency, good biological activity and low cost.
With the wide use of 2, 4-epibrassinolide in the planting of dendrobium officinale, the phenomenon of pesticide abuse occurs, and the safety problem of the dendrobium officinale is concerned more and more. However, 2,4-epibrassinolide does not contain a conjugated structure and a photoelectric response group, and a conventional ultraviolet detector or a fluorescence detector is difficult and inconvenient to detect the content of the epibrassinolide. With continuous development of instrument technology, high performance liquid chromatography detection technology and gas chromatography are widely applied in the field of analysis and detection, but before detection of 2, 4-epibrassinolide, derivatization treatment is required, the operation is complex, and sometimes the column is damaged by derivatization on the column. In addition, the components in the dendrobium officinale are complex, the quality difference is large under the influence of the growth environment, and no report is found at present about the residue analysis of 2, 4-epibrassinolide in the dendrobium officinale. Therefore, how to realize simple and sensitive detection of 2, 4-epibrassinolide in dendrobium officinale is a problem which needs to be solved urgently at present.
Disclosure of Invention
The invention aims to provide a method for measuring the content of 2, 4-epibrassinolide in dendrobium officinale, and the method provided by the invention is simple to operate and high in sensitivity.
In order to achieve the above object, the present invention provides the following technical solutions:
the invention provides a method for measuring the content of 2, 4-epibrassinolide in dendrobium officinale, which comprises the following steps:
mixing herba Dendrobii, acetonitrile and water, performing ultrasonic extraction, and mixing the obtained ultrasonic extraction system with NaCl and MgSO 4 Mixing, performing vortex extraction, layering an obtained vortex extraction system to obtain a water phase and an acetonitrile phase, removing a solvent in the acetonitrile phase to obtain a first remainder, and performing first redissolution on the first remainder by using an acetone-n-hexane solution as a redissolvent to obtain a first redissolution;
using acetone-n-hexane solution as eluent and C 18 Purifying the first redissolved solution by using a solid-phase extraction column, removing a solvent in the obtained purified solution to obtain a second remainder, and carrying out second redissolution on the second remainder by using acetonitrile to obtain a second redissolved solution serving as a solution to be detected;
performing high performance liquid chromatography-tandem mass spectrometry detection on the liquid to be detected to obtain a chromatogram of the liquid to be detected;
obtaining the content of 2, 4-epibrassinolide in the dendrobium officinale according to a blank matrix standard curve of the dendrobium officinale with 2, 4-epibrassinolide and a chromatogram of a liquid to be detected;
the high performance liquid chromatography-tandem mass spectrometry detection high performance liquid chromatography conditions comprise:
the mobile phase system comprises a mobile phase A and a mobile phase B, wherein the mobile phase A is methanol, the mobile phase B is formic acid-ammonium formate-water solution, the volume fraction of formic acid in the mobile phase B is 0.1%, and the concentration of ammonium formate is 2-5 mmol/L; a gradient elution procedure was used, which was: 0-0.5 min, and keeping the volume fraction of the mobile phase A at 10%; 0.5-2.5 min, wherein the volume fraction of the mobile phase A is increased from 10% to 92%; 2.5-4.0 min, wherein the volume fraction of the mobile phase A is kept at 92%; 4.0-4.01 min, and the volume fraction of the mobile phase A is reduced from 92% to 10%; 4.01-5.5 min, the volume fraction of the mobile phase A is kept 10%.
Preferably, when the ultrasonic extraction is performed, the dosage ratio of the dendrobium officinale, the acetonitrile and the water is 10g: (10-20) mL: (4-6) mL.
Preferably, the temperature of ultrasonic extraction is 15-40 ℃, the time is 13-30 min, and the power is 220-280 Hz.
Preferably, the NaCl, mgSO 4 The dosage ratio of the acetonitrile used for ultrasonic extraction to the acetonitrile is (1.3-1.7) g: (5-7) g: (10-20) mL.
Preferably, the temperature of the vortex extraction is 15-40 ℃ and the time is 1.5-2.5 min.
Preferably, the volume ratio of acetone to n-hexane in the double solvent and the eluent is independently 6.
Preferably, the dosage ratio of the double solvent, the eluent and the dendrobium officinale is (1.5-2.5) mL: (20 to 25) mL:10g.
Preferably, said C 18 The solid phase extraction column is Cleanert PestiCarb/NH 2 A columella, oasis HLB columella, clearert PEP-2 columella, clearert TPH columella, or clearert NANO columella.
Preferably, the high performance liquid chromatography conditions further comprise:
the flow rate of the mobile phase system is 0.4mL/min, and the chromatographic column is C 18 The temperature of the column and the column incubator is 40 ℃, and the injection volume is 2.0 mu L.
Preferably, the mass spectrometric conditions of the hplc-tandem mass spectrometric detection comprise:
the ionization being ESI + Monitoring by adopting MRM multiple reactions; the capillary voltage is 4.0kV; the temperature of the sheath gas is 350 ℃; the sheath gas flow rate was 11L/min.
The invention provides a method for measuring the content of 2, 4-epibrassinolide in dendrobium officinale, which uses acetonitrile and water as an extracting agent, combines ultrasonic extraction and vortex extraction, and purifies a sample by a solid-phase extraction column after extraction, so that the matrix effect of the sample is reduced, and the method is simpler in operation compared with conventional derivatization treatment; according to the method, a proper amount of ammonium formate is added into the mobile phase, so that the ionization efficiency of the 2, 4-epibrassinolide can be improved, the obtained solution to be detected is further ensured to be suitable for high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) detection, and the 2, 4-epibrassinolide in the dendrobium officinale can be detected in a quantitative manner by an external standard method, so that the simple, rapid and sensitive detection of the 2, 4-epibrassinolide in the dendrobium officinale can be finally realized, and the method has important significance on the medication safety of the dendrobium officinale. The results of the examples show that when the method provided by the invention is used for detecting 2, 4-epibrassinolide in dendrobium officinale, the 2, 4-epibrassinolide has a good linear relation in the concentration range of 0.0005-1 mg/L, the average recovery rate of the 2, 4-epibrassinolide is 81.9-103.2%, the coefficient of variation is 1.1-9.4%, and the detection limit is 0.005mg/kg. The method provided by the invention has the advantages of good purification and separation effects, good repeatability and low detection limit, completely meets the technical requirements of detecting 2, 4-epibrassinolide residue in dendrobium officinale, and is suitable for rapid detection of a large number of samples.
Drawings
FIG. 1 is a structural formula of brassinolide and 2, 4-epibrassinolide;
FIG. 2 is a standard curve of 2, 4-epibrassinolide Dendrobium officinale blank matrix solution;
FIG. 3 is a spectrum of 2, 4-epibrassinolide in a practical sample of Dendrobium officinale Kimura et Migo;
FIG. 4 is a graph of the effect of different conditions on the recovery of 2, 4-brassinolide.
Detailed Description
The invention provides a method for measuring the content of 2, 4-epibrassinolide in dendrobium officinale, which comprises the following steps:
mixing herba Dendrobii, acetonitrile and water, performing ultrasonic extraction, and mixing the obtained ultrasonic extraction system with NaCl and MgSO 4 Mixing, performing vortex extraction, layering an obtained vortex extraction system to obtain a water phase and an acetonitrile phase, removing a solvent in the acetonitrile phase to obtain a first residue, and performing first redissolution on the first residue by taking an acetone-n-hexane solution as a redissolution to obtain a first redissolution;
using acetone-n-hexane solution as eluent and C 18 Purifying the first redissolution by using a solid-phase extraction column, removing a solvent in the obtained purified solution to obtain a second remainder, and performing second redissolution on the second remainder by using acetonitrile to obtain a second redissolution as a solution to be detected;
performing high performance liquid chromatography-tandem mass spectrometry detection on the liquid to be detected to obtain a chromatogram of the liquid to be detected;
obtaining the content of 2, 4-epibrassinolide in the dendrobium officinale according to a blank matrix standard curve of the dendrobium officinale with 2, 4-epibrassinolide and a chromatogram of a liquid to be detected;
the high performance liquid chromatography-tandem mass spectrometry detection high performance liquid chromatography conditions comprise:
the mobile phase system comprises a mobile phase A and a mobile phase B, wherein the mobile phase A is methanol, the mobile phase B is formic acid-ammonium formate-water solution, the volume fraction of formic acid in the mobile phase B is 0.1%, and the concentration of ammonium formate is 2mmol/L; a gradient elution procedure was used, which was: 0-0.5 min, and keeping the volume fraction of the mobile phase A at 10%; 0.5-2.5 min, wherein the volume fraction of the mobile phase A is increased from 10% to 92%; 2.5-4.0 min, wherein the volume fraction of the mobile phase A is kept at 92%; 4.0-4.01 min, and the volume fraction of the mobile phase A is reduced from 92% to 10%; 4.01-5.5 min, and the volume fraction of the mobile phase A is kept 10%.
The method comprises the step of mixing dendrobium officinale, acetonitrile and water for ultrasonic extraction to obtain an ultrasonic extraction system. The invention preferably takes the leaves and stems of dendrobium officinale as samples for detection; the leaves and stems of the dendrobium officinale are preferably subjected to dry ice crushing and sample preparation before use, and the obtained sample is preferably stored at the temperature of-20 ℃. In the invention, the preferable dosage ratio of the dendrobium officinale, the acetonitrile and the water is 10g: (10-20) mL: (4-6) mL, more preferably 10g:15mL of: 5mL. Preferably, water is added into the dendrobium officinale, then acetonitrile is added, and the obtained mixed system is subjected to ultrasonic extraction; according to the invention, water is firstly added into the dendrobium officinale, so that the dendrobium officinale can be fully infiltrated, and then acetonitrile is added for ultrasonic extraction, so that the extraction efficiency can be effectively improved. In the present invention, the temperature of the ultrasonic extraction is preferably 15 to 40 ℃, more preferably 20 to 30 ℃, and specifically, the ultrasonic extraction may be performed at room temperature (25 ℃); the time for ultrasonic extraction is preferably 13-30 min, and more preferably 15min; the power of the ultrasonic extraction is preferably 220 to 280Hz, and more preferably 250Hz.
After an ultrasonic extraction system is obtained, the ultrasonic extraction system is mixed with NaCl and MgSO 4 Mixing, and performing vortex extraction to obtain a vortex extraction system. In the present invention, the NaCl and MgSO 4 The dosage ratio of the acetonitrile used in ultrasonic extraction is preferably (1.3-1.7) g: (5-7) g: (10-20) mL, more preferably 1.5g:6g:15mL. In the present invention, the temperature of the vortex extraction is preferably 15 to 40 ℃, more preferably 20 to 30 ℃, and particularly the vortex extraction can be performed under the condition of room temperature (25 ℃); the time of the vortex extraction is preferably 1.5 to 2.5min, more preferably 2min. In the present invention, the NaCl and MgSO 4 Has salting-out effect, and can be used for obtaining water phase and acetonitrile phase for subsequent layering.
After a vortex extraction system is obtained, the vortex extraction system is layered to obtain a water phase and an acetonitrile phase, and a solvent in the acetonitrile phase is removed to obtain a first remainder. The present invention preferably centrifuges the vortex extraction system to facilitate stratification; the rotating speed of the centrifugation is preferably 7000-9000 r/min, more preferably 8000r/min, and the time of the centrifugation is preferably 8-12 min, more preferably 10min; the centrifugation is preferably carried out at 2 to 6 ℃, more preferably at 4 ℃. The solvent in the acetonitrile phase is preferably removed by rotary evaporation, and the acetonitrile phase is particularly subjected to rotary evaporation and near drying under the condition of water bath at 40 ℃.
After the first residue is obtained, the first residue is subjected to first redissolution by using an acetone-n-hexane solution as a redissolution to obtain a first redissolution. In the present invention, the volume ratio of acetone to n-hexane in the double solvent is preferably 6 to 8, more preferably 6; the preferable dosage ratio of the double solvent to the dendrobium officinale is (1.5-2.5) mL:10g, more preferably 2mL:10g.
After the first complex solution is obtained, the invention takes acetone-n-hexane solution as eluent and adopts C 18 And purifying the first redissolution by using a solid-phase extraction column to obtain a purified solution. In the present invention, the composition of the eluent is preferably the same as that of the double solvent, and is not described herein again; the preferable dosage ratio of the eluent to the dendrobium officinale is (20-25) mL:10g, more preferably 25mL:10g. In the present invention, said C 18 The solid phase extraction column is preferably Cleanert Pesticocarb/NH 2 A columella, oasis HLB columella, cleanert PEP-2 columella, cleanert TPH columella or Cleanert NANO columella, more preferably Cleanert Pesticocarb/NH 2 A pillar; in an embodiment of the present invention, the Cleanert Pesticocarb/NH 2 The size of the column is preferably 500mg/6mL, the size of the Oasis HLB column is preferably 500mg/6mL, the size of the Cleanert PEP-2 column is preferably 500mg/6mL, the size of the Cleanert TPH column is 2g/12mL, and the size of the Cleanert NANO column is 1mL. In the present invention, said C 18 The solid phase extraction cartridge is preferably activated before use, the reagent used for activation is preferably the same as the double solvent and is not described herein, and in the present invention, the volume of the reagent used for activation is preferably 8 to 12mL, and more preferably 10mL. The invention preferably loads the first re-solution on activated C 18 Solid phase extracting the small column, then eluting the C by eluent 18 And (5) performing solid phase extraction on the small column to obtain a purified solution.
After the purified solution is obtained, the solvent in the purified solution is removed to obtain a second residue, acetonitrile is adopted to carry out second redissolution on the second residue, and the obtained second redissolution is used as a solution to be detected. The solvent in the purifying liquid is preferably removed by rotary evaporation, and the purifying liquid is particularly subjected to rotary evaporation to be nearly dried under the condition of water bath at 40 ℃. In the invention, acetonitrile and the second remainder are preferably mixed and ultrasonically dissolved for 1min, and the obtained mixed solution is filtered through a 0.22 mu m filter membrane to obtain a solution to be detected.
After the liquid to be detected is detected, the high performance liquid chromatography-tandem mass spectrometry detection is carried out on the liquid to be detected, and the chromatogram of the liquid to be detected is obtained. In the invention, the high performance liquid chromatography-tandem mass spectrometry detection high performance liquid chromatography conditions comprise:
the mobile phase system comprises a mobile phase A and a mobile phase B, wherein the mobile phase A is methanol, the mobile phase B is formic acid-ammonium formate-water solution, the volume fraction of formic acid in the mobile phase B is 0.1%, and the concentration of ammonium formate is 2mmol/L; a gradient elution procedure was used, which was: 0-0.5 min, wherein the volume fraction of the mobile phase A is kept at 10%; 0.5-2.5 min, wherein the volume fraction of the mobile phase A is increased from 10% to 92%; 2.5-4.0 min, wherein the volume fraction of the mobile phase A is kept at 92%; 4.0-4.01 min, and the volume fraction of the mobile phase A is reduced from 92% to 10%; 4.01-5.5 min, and the volume fraction of the mobile phase A is kept 10%.
In the present invention, the high performance liquid chromatography conditions preferably further comprise: the flow rate of the mobile phase system is 0.4mL/min, and the chromatographic column is C 18 The temperature of the column and the column incubator is 40 ℃, and the injection volume is 2.0 mu L.
In the present invention, the mass spectrometric conditions for the hplc-tandem mass spectrometric detection preferably comprise: the ionization being ESI + Monitoring by adopting MRM multiple reactions; the capillary voltage is 4.0kV; the temperature of the sheath gas is 350 ℃; the sheath gas flow rate was 11L/min.
After obtaining the chromatogram of the test solution, the content of the 2, 4-epibrassinolide in the dendrobium officinale can be obtained according to the blank matrix standard curve of the dendrobium officinale with the 2, 4-epibrassinolide and the chromatogram of the test solution. In the invention, the blank matrix standard curve of the dendrobium officinale with 2, 4-epibrassinolide is preferably a linear regression equation of the mass concentration of the 2, 4-epibrassinolide in the blank matrix solution of the dendrobium officinale with the label and the chromatographic peak area of the 2, 4-epibrassinolide in the chromatogram of the blank matrix solution of the dendrobium officinale with the label, and the concentrations of the 2, 4-epibrassinolide in the blank matrix solution of the dendrobium officinale with the label are preferably 0.0005mg/L, 0.001mg/L, 0.0025mg/L, 0.025mg/L, 0.5mg/L, 0.25mg/L, 0.5mg/L and 1mg/L respectively. In the invention, the labeled dendrobium officinale blank matrix solution is specifically obtained by diluting 2, 4-epibrassinolide standard mother liquor with the dendrobium officinale blank matrix solution; the preparation method of the dendrobium officinale blank matrix solution is preferably the same as that of the solution to be detected, and is not repeated herein; the concentration of the 2, 4-epibrassinolide standard mother liquor is preferably 100mg/L, and the solvent is preferably acetonitrile. In the invention, the chromatogram of the blank matrix solution of the dendrobium officinale added with the standard is preferably obtained by carrying out high performance liquid chromatography-tandem mass spectrometry detection on the blank matrix solution of the dendrobium officinale added with the standard; the conditions for performing high performance liquid chromatography-tandem mass spectrometry detection on the labeled dendrobium officinale blank matrix solution are preferably consistent with the conditions for performing high performance liquid chromatography-tandem mass spectrometry detection on the solution to be detected, and are not repeated herein. According to the blank matrix standard curve of the dendrobium officinale with 2, 4-epibrassinolide and the chromatogram of the liquid to be detected, the content of the 2, 4-epibrassinolide in the liquid to be detected can be obtained, and further the content of the 2, 4-epibrassinolide in the dendrobium officinale can be obtained.
The technical solution of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. It is to be understood that the described embodiments are merely exemplary of the invention, and not restrictive of the full scope of the invention. All other embodiments, which can be obtained by a person skilled in the art without making any creative effort based on the embodiments in the present invention, belong to the protection scope of the present invention.
Example 1
1. Materials and methods
1.1 Main instruments and chemical reagents
The main apparatus comprises: high performance liquid chromatography-triple quadrupoleMass spectrometer (G6470, agilent, usa); the Agilent Master Qualitative Analysis 8.0 software is used for data acquisition and Analysis; the chromatographic column is Waters C 18 Columns (100 mm. Times.2.1mm, 1.7 μm); an ultrasonic cleaner (SK-7200H, shanghai Ke dao ultrasonic Instrument Co., ltd.); vortex oscillators (NMSG-12, mino medical science, tex.); high-speed refrigerated centrifuges (6430R, pendorf Co.).
Main reagents and consumables: 2, 4-epibrassinolide technical (93% purity, CAS: 78821-43-9) was purchased from Zhejiang Shijia scientific Co., ltd; 2, 4-epibrassinolide standard substance (purity is more than or equal to 98 percent, CAS: 78821-43-9) is purchased from Heifengbo-Mei Biotech Limited liability company; methanol, acetonitrile (HPLC grade, merck); formic acid (HPLC grade, ACS corporation, usa); ammonium formate (HPLC grade, honeywell, germany), acetone, n-hexane (AR grade) were purchased from the pharmaceutical group chemicals ltd. Extract salt package (containing 1.5g NaCl and 6g MgSO) 4 ) Purchased from Hangzhou Yimeichu technologies, inc.; cleanert Pesticocarb/NH 2 (CARB/NH 2 ) Solid Phase Extraction (SPE) cartridges (500mg, 6mL) were purchased from Shanghai Anpu laboratory science and technology, inc.; ceramics homogeneous protons are purchased from Hefeibomei biotechnology, limited liability company; drochen distilled water (purchased from drochen group ltd).
1.2 methods
1.2.1 apparatus conditions
High performance liquid chromatography conditions:
a chromatographic column: waters C 18 Columns (100 mm. Times.2.1 mm,1.7 μm);
mobile phase: the method comprises a mobile phase A and a mobile phase B, wherein the mobile phase A is methanol, the mobile phase B is formic acid-ammonium formate-water solution, the volume fraction of formic acid in the mobile phase B is 0.1%, and the concentration of ammonium formate is 2mmol/L; gradient elution was used, as shown in table 1;
flow rate: 0.4mL/min;
column oven: at 40 ℃;
sample introduction volume: 20 mu L of the solution;
operating time: 5.5min.
TABLE 1 mobile phase gradient elution conditions
Mass spectrum conditions:
an ionization mode: ESI + Monitoring by adopting MRM multiple reactions;
capillary voltage: 4.0kV;
temperature of sheath gas: 350 ℃;
sheath gas flow rate: 11L/min;
the detection parameters are shown in Table 2.
TABLE 2 Mass spectrometric parameters
* To quantify ion pairs.
1.2.2 sample pretreatment
Extraction: respectively weighing 10.00g of leaf and stem sample of Dendrobium officinale Kimura et Migo in 50mL centrifuge tube, sequentially adding 5.0mL water and 15.0mL acetonitrile, ultrasonic extracting for 15min, then adding 1.5g NaCl and 6.0g MgSO 4 Placing a ceramic homogeneous proton in each centrifuge tube, performing vortex oscillation for 2min, centrifuging for 10min at 8000r/min at 4 ℃, transferring all upper acetonitrile phases to a round-bottom flask, performing vortex evaporation for near drying at 40 ℃ in a water bath, and re-dissolving with 2mL of acetone-n-hexane solution (the volume ratio of acetone to n-hexane is 6);
purification: the SPE cartridge (CARB/NH) was purified using 10mL of acetone-n-hexane solution (volume ratio of acetone to n-hexane is 6) 2 ) After activation, 2mL of the reconstituted solution was loaded, and then 25mL of an acetone-n-hexane solution (volume ratio of acetone to n-hexane was 6: 4) And (3) leaching the small column, collecting all purified solution, carrying out rotary evaporation in water bath at 40 ℃ to obtain a near dry solution, re-dissolving the near dry solution with acetonitrile, carrying out ultrasonic treatment for 1min, and filtering the solution with a 0.22 mu m filter membrane to be detected.
2. As a result, the
2.1 determination of the Linear relationship of the analytical methods
Accurately weighing 2, 4-epi-brassin0.0111g of ester standard substance is put in a volumetric flask with 100mL, acetonitrile is adopted to fix the volume to a scale mark, and a standard mother solution with 100mg/L is prepared and stored at 4 ℃; diluting a blank matrix solution (prepared by a method of '1.2.2 sample pretreatment') of dendrobium officinale to prepare standard solutions with concentrations of 0.0005mg/L, 0.001mg/L, 0.0025mg/L, 0.025mg/L, 0.5mg/L, 0.25mg/L, 0.5mg/L and 1mg/L, measuring according to a method of '1.2.1 instrument conditions', repeating for 3 times, drawing a standard curve by taking the concentration as a horizontal coordinate and taking the peak area as a vertical coordinate, and calculating a regression equation and a correlation coefficient. The results are shown in Table 3, the regression equation of the substrate standard curve is y =2E +06x +7520.9 (shown in FIG. 2) for 2,4-epibrassinolide in the concentration range of 0.0005-1 mg/L, the correlation coefficient: r is 2 ＝0.9988。
TABLE 3, 4 Standard Curve for epibrassinolide
2.2 recovery and detection limits of the Process
Adding 2, 4-epibrassinolide standard solution into the dendrobium officinale hollow white matrix solution, and performing addition recovery tests with three concentrations of 0.005mg/kg, 0.05mg/kg and 0.5mg/kg, wherein 6 concentrations are arranged in parallel; wherein, the extraction and purification are carried out according to the method of '1.2.2 sample pretreatment', the detection is carried out according to the method of '1.2.1 instrument condition', the addition recovery rate test is completed, and the addition recovery rate and the Relative Standard Deviation (RSDs) of the 2, 4-epibrassinolide in the dendrobium officinale sample are obtained, and the specific data are shown in the table 4.
TABLE 4, 4-results of experiments on the recovery of epibrassinolide from Dendrobium officinale
As can be seen from Table 4, when the 2, 4-epibrassinolide is added into the dendrobium officinale at concentrations of 0.005mg/kg, 0.05mg/kg and 0.5mg/kg, the average recovery rates are 94.1%, 95.7.0% and 81.9%, respectively, and the relative standard deviations are 3.3%, 3.9% and 2.6%, respectively; the average recovery rate of 2, 4-epibrassinolide is between 81.9 and 103.2 percent, and the coefficient of variation is between 1.1 and 9.4 percent.
According to the response condition of the 2, 4-epibrassinolide on an ultra performance liquid chromatography tandem mass spectrometer and the response value of a sample with the lowest added concentration of the 2, 4-epibrassinolide of the dendrobium officinale sample on the instrument being 0.005mg/kg is larger than the signal-to-noise ratio of the instrument being 3 times, the lowest detection concentration of the 2, 4-epibrassinolide in the dendrobium officinale sample is 0.005mg/kg. Therefore, the method provided by the invention can completely meet the requirements on accuracy, precision and sensitivity in pesticide residue analysis.
3. Actual sample detection
5 parts of commercially available samples of dendrobium officinale are detected according to a method of 1.2, and the results show that 2, 4-epibrassinolide is not detected (figure 3).
Test example 1
The test example examined the effect of the extraction method, extraction time, type of extractant, volume of extractant, type of SPE column, type of eluent, and volume of eluent on the recovery (%) of 2, 4-epibrassinolide (the result is shown in FIG. 4), and the specific operation steps were as follows (conditions not mentioned in the following steps were performed according to "1.2 method"):
(1) The extraction method adopts the combination of ultrasound, vortex, suction filtration, ultrasound and vortex, the extraction time is 17min, when the combination of ultrasound and vortex is adopted, the ultrasound time is 15min, and the vortex time is 2min; the extractant is acetonitrile-water solution (acetonitrile 15mL, water 5 mL), and clean Pesticocarb/NH 2 Column (500mg, 6ml) purified, eluent was acetone-n-hexane solution (acetone to n-hexane volume ratio 6.
(2) The fixed extraction method is ultrasonic and vortex combined, the extractant is acetonitrile-water solution (acetonitrile is 15mL, water is 5 mL), and clean Pesticocarb/NH is used 2 Purifying by a small column (500mg, 6mL), wherein an eluent is an acetone-n-hexane solution (the volume ratio of acetone to n-hexane is 6; wherein the extraction time is 10min, 15min, 20min,30min (b in FIG. 4).
(3) The fixed extraction method is ultrasonic and vortex combined, the volume of the extractant is 15mL, the extraction time is 15min, and clean Pesticocarb/NH is used 2 Purifying by a small column (500mg, 6mL), wherein an eluent is an acetone-n-hexane solution (the volume ratio of acetone to n-hexane is 6; wherein the extracting agents are acetonitrile, acetone, methanol, acetonitrile-water solution (the volume ratio of acetonitrile to water is 3.
(4) The fixed extraction method is ultrasonic and vortex combined, the extraction agent is acetonitrile-water solution, the extraction time is 15min, and clean PesticCarb/NH is used 2 Purifying by a small column (500mg, 6mL), wherein an eluent is an acetone-n-hexane solution (the volume ratio of acetone to n-hexane is 6; wherein the volume of the extracting agent is determined by acetonitrile: water shows 5mL:5mL, 10mL:5mL, 15mL:5mL, 20mL:5mL (d in FIG. 4).
(5) The fixed extraction mode is ultrasonic and vortex combined, the extraction agent is acetonitrile-water solution (acetonitrile is 15mL, water is 5 mL), the extraction time is 15min, the eluent is acetone-n-hexane solution (the volume ratio of acetone to n-hexane is 6), and the volume of the eluent is 25mL; wherein, the SPE small columns are Cleanert NH respectively 2 Small column (500mg, 6mL), clean PestiCarb/NH 2 Column (500mg, 6mL), oasis HLB column (500mg, 6mL), cleanert PEP-2 column (500mg, 6mL), cleanert TPH column (2g, 12mL), cleanert NANO column (1 mL) (e in FIG. 4).
(6) The fixed extraction method is combination of ultrasound and vortex, the extraction agent is acetonitrile-water solution (acetonitrile is 15mL, water is 5 mL), the extraction time is 15min, and clean Pesticocarb/NH is used 2 Purifying (500mg, 6mL), wherein the volume of the eluent is 25mL; the eluents are an acetonitrile-toluene solution (acetonitrile to toluene volume ratio of 3, 1), an acetone-n-hexane solution (acetone to n-hexane volume ratio of 8, 2), an acetone-n-hexane solution (acetone to n-hexane volume ratio of 6, 4), an acetone-n-hexane solution (acetone to n-hexane volume ratio of 2.
(7) The fixed extraction method isThe acoustic vortex extraction is carried out by mixing acetonitrile-water solution (acetonitrile 15mL, water 5 mL) with clean Pesticocarb/NH for 15min 2 Purifying (500mg, 6mL), wherein an eluent is an acetone-n-hexane solution (the volume ratio of acetone to n-hexane is 6; the eluent volumes were 10mL, 15mL, 20mL, and 25mL (g in FIG. 4), respectively.
As can be seen from a in fig. 4, the combined effect of the ultrasonic extraction and the vortex extraction is the best.
As can be seen from b in FIG. 4, the extraction efficiency was the best at 15 min. Meanwhile, it can be seen that the extraction efficiency is rather decreased when the extraction time is further prolonged, and it is presumed that the matrix effect is enhanced to affect the recovery rate due to leaching of other impurities caused by long-time extraction.
As can be seen from c in fig. 4, different extractants have a great influence on the recovery rate, and although acetone has the best recovery rate effect, the different extractants have strong solubility on substances such as lipids and pigments, and a large amount of impurities are not easy to purify in the extraction process, so that the matrix effect is enhanced, so that the acetonitrile-water solution is selected as the extractant.
As can be seen from d in fig. 4, under the other conditions, the recovery efficiency was the best when the amounts of acetonitrile and water were 15mL and 5mL, and the recovery rate was decreased when the amounts of acetonitrile and water were 20mL and 5mL, and the substrate effect may be affected by the matrix effect due to the increased acetonitrile ratio, which resulted in the dissolution of other impurities from dendrobium officinale.
From e in FIG. 4, it can be seen that the NH is Cleanert NH 2 Besides the small column (500mg, 6 mL), the recovery rate of other SPE small columns reaches the standard (more than or equal to 70 percent), and because the matrix is complex and contains a large amount of chlorophyll, the selection of clean Pesticocarb/NH is comprehensively considered 2 A small column.
As can be seen from f in fig. 4, the recovery rate was best when the eluent was acetone-n-hexane solution and the volume ratio of acetone to n-hexane was 6.
As can be seen from g in FIG. 4, in the case of the combination of sonication and vortexing, the mixture was extracted with an acetonitrile-water solution (15 mL of acetonitrile and 5mL of water) for 15min and then cleaned Pesticocarb/NH 2 Purifying with a small column (500mg, 6mL), eluting with 25mL acetone-n-hexane solution (the volume ratio of acetone to n-hexane is 6The yield thereof was found to be 103.2%.
Test example 2
The test example investigates the influence of the mobile phase and the chromatographic column on the detection result, and concretely comprises the following steps:
(1) The detection was performed under the gradient elution conditions shown in table 5, and other conditions were performed under the "1.2.1 apparatus conditions" in example 1, and the results showed that the peak of the target substance had a tailing, the peak shape was poor, and the retention time was 2.4min. The target substance is detected under the gradient elution conditions shown in the table 1, the chromatographic peak shape of the target substance is better, and the retention time is about 3.4 min. Thus, the gradient elution conditions shown in Table 1 were finally selected.
TABLE 5 comparative mobile phase gradient elution conditions
(2) By T 3 The retention time of the target substance is about 1.6min, and the operation procedure is generally different between 7min and 15min (the detection method is HPLC or HPLC-MS/MS).
The invention selects C 18 And (3) detecting the chromatographic column according to the method of '1.2.1 instrument condition' in the embodiment 1, wherein the retention time of the target is about 3.4min, the running time is 5.5min, the detection time is shortened, the peak-appearance time of the target is moderate, and the problem that the detection result is influenced by impurities in the sample due to too short peak-appearance time is solved.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.
Claims (10)
1. A method for measuring the content of 2, 4-epibrassinolide in dendrobium officinale comprises the following steps:
mixing herba Dendrobii, acetonitrile and water, performing ultrasonic extraction, and mixing the obtained ultrasonic extraction system with NaCl and MgSO 4 Mixing, performing vortex extraction, and mixingLayering the obtained vortex extraction system to obtain a water phase and an acetonitrile phase, removing the solvent in the acetonitrile phase to obtain a first remainder, and carrying out first redissolution on the first remainder by using an acetone-n-hexane solution as a redissolution to obtain a first redissolution;
using acetone-n-hexane solution as eluent and C 18 Purifying the first redissolved solution by using a solid-phase extraction column, removing a solvent in the obtained purified solution to obtain a second remainder, and carrying out second redissolution on the second remainder by using acetonitrile to obtain a second redissolved solution serving as a solution to be detected;
performing high performance liquid chromatography-tandem mass spectrometry detection on the liquid to be detected to obtain a chromatogram of the liquid to be detected;
obtaining the content of 2, 4-epibrassinolide in the dendrobium officinale according to a blank matrix standard curve of the dendrobium officinale with 2, 4-epibrassinolide and a chromatogram of a liquid to be detected;
the high performance liquid chromatography-tandem mass spectrometry detection high performance liquid chromatography conditions comprise:
the mobile phase system comprises a mobile phase A and a mobile phase B, wherein the mobile phase A is methanol, the mobile phase B is formic acid-ammonium formate-water solution, the volume fraction of formic acid in the mobile phase B is 0.1%, and the concentration of ammonium formate is 2-5 mmol/L; a gradient elution procedure was used, which was: 0-0.5 min, wherein the volume fraction of the mobile phase A is kept at 10%; 0.5-2.5 min, wherein the volume fraction of the mobile phase A is increased from 10% to 92%; 2.5-4.0 min, wherein the volume fraction of the mobile phase A is kept at 92%; 4.0-4.01 min, and the volume fraction of the mobile phase A is reduced from 92% to 10%; 4.01-5.5 min, the volume fraction of the mobile phase A is kept 10%.
2. The method according to claim 1, wherein the ultrasonic extraction is performed with a dosage ratio of dendrobium officinale, acetonitrile and water of 10g: (10-20) mL: (4-6) mL.
3. The method according to claim 1 or 2, wherein the temperature of the ultrasonic extraction is 15-40 ℃, the time is 13-30 min, and the power is 220-280 Hz.
4. The method of claim 1, wherein the NaCl, mgSO 4 The dosage ratio of the acetonitrile used for ultrasonic extraction to the acetonitrile is (1.3-1.7) g: (5-7) g: (10-20) mL.
5. The method according to claim 1 or 4, wherein the temperature of the vortex extraction is 15-40 ℃ for 1.5-2.5 min.
6. The method according to claim 1, wherein the volume ratio of acetone to n-hexane in the double solvent and eluent is independently 6 to 8.
7. The method according to claim 6, wherein the dosage ratio of the double solvent, the eluent and the dendrobium officinale is (1.5-2.5) mL: (20 to 25) mL:10g.
8. The method of claim 1, wherein C is 18 The solid phase extraction column is Cleanert PestiCarb/NH 2 A columella, oasis HLB columella, clearertpep-2 columella, clearert TPH columella, or clearertnano columella.
9. The method of claim 1, wherein the high performance liquid chromatography conditions further comprise:
the flow rate of the mobile phase system is 0.4mL/min, and the chromatographic column is C 18 The temperature of the column and the column incubator is 40 ℃, and the injection volume is 2.0 mu L.
10. The method of claim 1, wherein the mass spectrometric conditions for hplc-tandem mass spectrometric detection comprise:
the ionization being ESI + Monitoring by adopting MRM multiple reactions; the capillary voltage is 4.0kV; the temperature of the sheath gas is 350 ℃; the flow rate of the sheath gas was 11L/min.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116106467A (en) * | 2023-04-13 | 2023-05-12 | 江西省药品检验检测研究院 | Method for identifying raw pinellia tuber in Huoxiang Zhengqi water |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103936494A (en) * | 2013-01-22 | 2014-07-23 | 新宁县湘斛种植科技开发有限公司 | Drug agent used in dendrobium officinale plantation and application method thereof |
| WO2019196233A1 (en) * | 2018-04-09 | 2019-10-17 | 深圳市宇驰检测技术股份有限公司 | Method for measuring contents of various endocrine disruptors in estuary sediment |
| CN110887926A (en) * | 2019-12-19 | 2020-03-17 | 浙江省农业科学院 | Sample pretreatment method for determining residual quantity of various pesticides in dendrobium officinale and detection method thereof |
| CN111373914A (en) * | 2020-03-27 | 2020-07-07 | 江西瑞博特生物科技有限公司 | Microbial nutrient for planting dendrobium officinale, and preparation method and application thereof |
| US20200326352A1 (en) * | 2019-04-09 | 2020-10-15 | Peking University | Method and a kit for simultaneous analyses of thyroid hormones and related metabolites in serum |
| AU2021105035A4 (en) * | 2021-08-06 | 2021-09-30 | Zhejiang Fangyuan Test Group Co., Ltd. | High performance liquid chromatography tandem mass spectrometry method for determining quaternary ammonium salt in food packaging paper on the basis of dispersive solid phase extraction |
-
2022
- 2022-07-18 CN CN202210838830.6A patent/CN115184497B/en active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103936494A (en) * | 2013-01-22 | 2014-07-23 | 新宁县湘斛种植科技开发有限公司 | Drug agent used in dendrobium officinale plantation and application method thereof |
| WO2019196233A1 (en) * | 2018-04-09 | 2019-10-17 | 深圳市宇驰检测技术股份有限公司 | Method for measuring contents of various endocrine disruptors in estuary sediment |
| US20200326352A1 (en) * | 2019-04-09 | 2020-10-15 | Peking University | Method and a kit for simultaneous analyses of thyroid hormones and related metabolites in serum |
| CN110887926A (en) * | 2019-12-19 | 2020-03-17 | 浙江省农业科学院 | Sample pretreatment method for determining residual quantity of various pesticides in dendrobium officinale and detection method thereof |
| CN111373914A (en) * | 2020-03-27 | 2020-07-07 | 江西瑞博特生物科技有限公司 | Microbial nutrient for planting dendrobium officinale, and preparation method and application thereof |
| AU2021105035A4 (en) * | 2021-08-06 | 2021-09-30 | Zhejiang Fangyuan Test Group Co., Ltd. | High performance liquid chromatography tandem mass spectrometry method for determining quaternary ammonium salt in food packaging paper on the basis of dispersive solid phase extraction |
Non-Patent Citations (4)
| Title |
|---|
| HONGHONG FAN等: "Molecular cloning and expression of 1-deoxy-d-xylulose-5-phosphate synthase and 1-deoxy-d-xylulose-5-phosphate reductoisomerase in Dendrobium officinale", 《 PLANT CELL, TISSUE AND ORGAN CULTURE (PCTOC)》, vol. 125, no. 2, pages 381 - 385 * |
| WANG GUANGJU等: "A study on the synthesis and application of 24-epibrassinolide", vol. 40, no. 6, pages 12 - 15 * |
| 曹立冬等: "高效液相色谱-蒸发光散射检测器检测 24-表芸苔素内酯原药含量", vol. 16, no. 1, pages 72 - 77 * |
| 虞冰等: "QuEChEＲS-超高效液相色谱-串联质谱法快速测定 草莓中2，4-表芸苔素内酯残留量", vol. 58, no. 9, pages 1538 - 1540 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116106467A (en) * | 2023-04-13 | 2023-05-12 | 江西省药品检验检测研究院 | Method for identifying raw pinellia tuber in Huoxiang Zhengqi water |
| CN116106467B (en) * | 2023-04-13 | 2023-07-28 | 江西省药品检验检测研究院 | Method for identifying raw pinellia tuber in Huoxiang Zhengqi water |
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