A method for determining the content of codeine, morphine, papaverine, noscapine and thebaine in poppy oil

Abstract

The present application provides a method for determining the content of codeine, morphine, papaverine, narcotine and thebaine in poppy oil, and belongs to the technical field of detection methods. The method comprises the following steps: (1) preparation of an isotopic internal standard working solution; (2) adding an adsorbent to the poppy oil sample, stirring and adsorbing, filtering, adding the solid into a solvent, heating and desorbing, filtering, washing, combining the washing liquid and the filtrate, adding the isotopic internal standard working solution, filtering, constant volume, and preparing a test sample solution; (3) preparation of a control stock solution; (4) preparation of a control sample solution; (5) determination of the content of codeine, morphine, papaverine, narcotine and thebaine in poppy oil by LC-MS / MS. The present application uses a simple and effective method to determine the content of codeine, morphine, papaverine, narcotine and thebaine in poppy oil, has a larger linear range, a lower detection limit, a high accuracy, a high sensitivity, and a good linearity, and is a reliable and effective detection method.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of detection methods, and particularly relates to a method for determining the contents of codeine, morphine, papaverine, noscapine and thebaine in poppy oil. BACKGROUND

[0002] Poppy seed oil is a plant oil extracted from the seeds of the poppy plant (Papaver somniferum L.). Although poppy seeds are generally considered to contain no or only trace amounts of alkaloids, the juice of the poppy capsule (containing high concentrations of alkaloids) can easily contaminate the seed surface during actual cultivation, harvesting, processing and storage. In addition, studies have shown that under certain conditions (such as mechanical damage, pests and diseases, and specific varieties), alkaloids can also enter the seed interior through endogenous pathways. Therefore, poppy oil may contain a variety of natural alkaloids derived from contamination or endogenous accumulation, of which the five most important and pharmacologically and toxicologically significant are morphine, codeine, papaverine, noscapine and thebaine. In order to avoid health risks and abuse, it is necessary to detect the contents of the above five substances in poppy oil.

[0003] Poppy oil itself is a high-fat matrix, containing a large amount of lipids such as triglycerides and free fatty acids. These substances can seriously interfere with subsequent chromatographic separation and mass spectrometric detection, leading to column contamination, poor peak shape and decreased sensitivity. Traditional methods for water-soluble samples or plant extracts are usually not applicable. In addition, the chemical structures, polarities, acid-base properties (morphine and codeine have phenolic hydroxyl groups and are weakly acidic; papaverine, noscapine and thebaine are tertiary amines and are weakly basic), and solubilities (dissolve differently in oil matrix and water) of the five alkaloids are significantly different. Developing a method that can effectively extract, purify and separate these five compounds with different properties at the same time is a core difficulty. SUMMARY

[0004] The present application aims to provide a method for determining the contents of codeine, morphine, papaverine, noscapine and thebaine in poppy oil, which can determine the contents of codeine, morphine, papaverine, noscapine and thebaine in poppy oil by a simple and effective method, has a larger linear range, a lower detection limit, a high accuracy, a high sensitivity, and a good linearity with a regression coefficient (r) of 0.990 or more, and is a reliable and effective detection method.

[0005] The technical solution of the present application is implemented as follows:

[0006] The present application provides a method for determining the contents of codeine, morphine, papaverine, noscapine and thebaine in poppy oil, comprising the following steps:

[0007] (1) Preparation of isotopic internal standard working solution: precisely pipette morphine-D3 isotope solution and codeine-D3 isotope solution into a volumetric flask respectively, and then dilute to volume to prepare morphine-D3 and codeine-D3 internal standard working solution;

[0008] (2) Preparation of test sample solution: add adsorbent to the opium oil sample, stir and adsorb, filter, add the solid into solvent, heat and desorb, filter, wash, combine the washing liquid with the filtrate, add isotopic internal standard working solution, filter, and dilute to volume to prepare the test sample solution;

[0009] (3) Preparation of control stock solution: prepare the corresponding stock solution by using codeine, morphine, papaverine, thebaine and noscapine standard;

[0010] (4) Preparation of control sample solution: dilute the control stock solution to prepare the control sample solution;

[0011] (5) Detection: LC-MS / MS is used to detect the contents of codeine, morphine, papaverine, noscapine and thebaine in opium oil.

[0012] The detection method first separates the target objects in the test sample by using liquid chromatography (HPLC); the separated sample is ionized into charged ions in the ion source; then, the first mass spectrometer selects the target parent ion according to the mass-to-charge ratio (m / z) and excludes other interfering ions; the parent ion enters the collision chamber and is fragmented into daughter ions by collision-induced dissociation (CID); finally, the second mass spectrometer detects the daughter ions to generate a fragment spectrum, revealing the chemical structure characteristics of the parent ion.

[0013] As a further improvement of the present application, the adsorbent comprises adsorbent A, adsorbent B and adsorbent C, and the mass ratio is 5-7:4-6:1-3;

[0014] The preparation method of the adsorbent A is as follows:

[0015] Morphine, thebaine, monomer, crosslinking agent, emulsifier and pore-forming agent are mixed and added into ethyl acetate, uniformly mixed, dropped into water, emulsified, and then initiator is added under the protection of inert gas, and the product is filtered, added into a Soxhlet extractor, extracted by mixed solvent until no morphine and thebaine are detected, washed, dried, and the adsorbent A is prepared;

[0016] The preparation method of the adsorbent B is as follows:

[0017] S1. Preparation of chiral amino acid coupled MOF: L-aspartic acid, L-glutamic acid were added to water, N-hydroxysuccinimide and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirring activation, UiO-66-NH2 was added, stirring reaction, filtration, washing, drying, chiral amino acid coupled MOF was prepared;

[0018] S2. Double bond grafting modification: maleic acid was added to water, N-hydroxysuccinimide and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirring activation, chiral amino acid coupled MOF was added, stirring reaction, filtration, washing, drying, modified chiral amino acid coupled MOF was prepared;

[0019] S3. Copolymerization: modified chiral amino acid coupled MOF, methyl methacrylate, styrene, N-isopropyl acrylamide were mixed and added to ethyl acetate, mixed uniformly, added dropwise to water, emulsified, under the protection of inert gas, initiator was added, heated and stirred to react, the product was filtered to prepare adsorbent B;

[0020] The preparation method of the adsorbent C is as follows:

[0021] T1. Preparation of p-aminobenzenesulfonic acid diazonium salt solution: p-aminobenzenesulfonic acid and sodium nitrite were dissolved in lye, heated and mixed uniformly, then cooled to room temperature, hydrochloric acid was added dropwise, stirred to react, and p-aminobenzenesulfonic acid diazonium salt solution was prepared;

[0022] T2. Preparation of sulfonated / double bond co-modified graphene oxide: maleic acid was added to toluene, graphene oxide and p-methylbenzenesulfonic acid were added, heated and refluxed to stir, centrifuged, washed, dried, the product was added to p-aminobenzenesulfonic acid diazonium salt solution, stirred to react, centrifuged, washed, dried, and sulfonated / double bond co-modified graphene oxide was prepared;

[0023] T3. Copolymerization: sulfonated / double bond co-modified graphene oxide, methyl methacrylate, styrene, N-isopropyl acrylamide were mixed and added to ethyl acetate, mixed uniformly, added dropwise to water, emulsified, under the protection of inert gas, initiator was added, heated and stirred to react, the product was filtered to prepare adsorbent C.

[0024] As a further improvement of the present application, in the preparation process of the adsorbent A, the monomers include methyl methacrylate, styrene, 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethyl sulfonate) imidazole, N-isopropyl acrylamide, and the mass ratio is 3-5:4-6:1-2:5-7, the crosslinking agent is ethylene glycol dimethacrylate, the emulsifier is a mixture of Tween-80 and Span-80, and the mass ratio is 2-3:3-5, the pore former is cetyltrimethylammonium chloride or cetyltrimethylammonium bromide, the initiator is selected from at least one of azobisisobutyronitrile, sodium persulfate, potassium persulfate, and ammonium persulfate, the temperature of the heating and stirring reaction is 55-65℃, and the time is 4-6h, the mass ratio of morphine, thebaine, the monomer, the crosslinking agent, the emulsifier, the pore former, and the initiator is 0.5-1:0.5-1:8-12:0.2-0.4:0.5-1:0.1-0.2:0.01-0.15, and the mixed solvent is a mixed solvent of acetonitrile and dichloromethane, and the volume ratio is 1:1-3.

[0025] As a further improvement of the present application, in the preparation process of the adsorbent A, the monomers include methyl methacrylate, styrene, 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethyl sulfonate) imidazole, N-isopropyl acrylamide, and the mass ratio is 3-5:4-6:1-2:5-7, the crosslinking agent is ethylene glycol dimethacrylate, the emulsifier is a mixture of Tween-80 and Span-80, and the mass ratio is 2-3:3-5, the pore former is cetyltrimethylammonium chloride or cetyltrimethylammonium bromide, the initiator is selected from at least one of azobisisobutyronitrile, sodium persulfate, potassium persulfate, and ammonium persulfate, the temperature of the heating and stirring reaction is 55-65℃, and the time is 4-6h, the mass ratio of morphine, thebaine, the monomer, the crosslinking agent, the emulsifier, the pore former, and the initiator is 0.5-1:0.5-1:8-12:0.2-0.4:0.5-1:0.1-0.2:0.01-0.15, and the mixed solvent is a mixed solvent of acetonitrile and dichloromethane, and the volume ratio is 1:1-3.

[0026] As a further improvement of the present application, in the preparation process of the adsorbent A, the monomers include methyl methacrylate, styrene, 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethyl sulfonate) imidazole, N-isopropyl acrylamide, and the mass ratio is 3-5:4-6:1-2:5-7, the crosslinking agent is ethylene glycol dimethacrylate, the emulsifier is a mixture of Tween-80 and Span-80, and the mass ratio is 2-3:3-5, the pore former is cetyltrimethylammonium chloride or cetyltrimethylammonium bromide, the initiator is selected from at least one of azobisisobutyronitrile, sodium persulfate, potassium persulfate, and ammonium persulfate, the temperature of the heating and stirring reaction is 55-65℃, and the time is 4-6h, the mass ratio of morphine, thebaine, the monomer, the crosslinking agent, the emulsifier, the pore former, and the initiator is 0.5-1:0.5-1:8-12:0.2-0.4:0.5-1:0.1-0.2:0.01-0.15, and the mixed solvent is a mixed solvent of acetonitrile and dichloromethane, and the volume ratio is 1:1-3.

[0027] As a further improvement of the present application, the detection chromatographic conditions are as follows: chromatographic column: Agilent ZORBAX SB-Aq 3.5 μm 3.0x100mm, column temperature 35-45℃, sample disc temperature 3-7℃, mobile phase: A: 2-7Mmol / L ammonium acetate+0.05-0.15% formic acid (v / v); B: 0.05-0.15% formic acid acetonitrile solution (v / v), flow rate 0.1-0.5mL / min, needle washing solution MeOH:H2O=1:1 (V:V).

[0028] As a further improvement of the present application, the detection mass spectrometry conditions are as follows: ion source ESI, curtain gas 30-40psi, temperature 550-650℃, ionization voltage 5000-6000V, spray gas 45-55psi, auxiliary heating gas 45-55psi, scanning mode positive ion mode.

[0029] As a further improvement of the present application, the linear range of codeine is 62.5ng / mL-187.5ng / mL, the linear range of morphine is 62.5ng / mL-187.5ng / mL, the linear range of papaverine is 12.5ng / mL-37.5ng / mL, the linear range of noscapine is 12.5ng / mL-37.5ng / mL, and the linear range of thebaine is 12.5ng / mL-37.5ng / mL.

[0030] As a further improvement of the present application, the regression coefficient of codeine is 0.995, the regression coefficient of morphine is 0.994, the regression coefficient of papaverine is 0.995, the regression coefficient of noscapine is 0.998, and the regression coefficient of thebaine is 0.999.

[0031] As a further improvement of the present application, the mass ratio of poppy oil, adsorbent and isotopic internal standard working solution is 15-20:1-2:0.000001-0.000002, the solvent is a mixed solution of formic acid and acetonitrile with a volume ratio of 0.5-1:100, the temperature for heating desorption is 40-45℃, and the time is 20-40min.

[0032] The present application has the following beneficial effects:

[0033] The present application uses a simple and effective method to determine the content of codeine, morphine, papaverine, noscapine and thebaine in poppy oil, has a larger linear range, a lower detection limit, a high accuracy and a high sensitivity, and the regression coefficient (r) is greater than or equal to 0.990, so that the linear range is good, and the present application is a reliable and effective detection method.

[0034] The application prepares adsorbent A, adsorbent B and adsorbent C, which have good adsorption on alkaloids in opium oil, and fast desorption rate, and the adsorbents can be repeatedly used, thereby reducing the cost.

[0035] The adsorbent A has good adsorption effect on morphine and thebaine with similar structures, adopts double-template imprinting, effectively separates and adsorbs morphine and thebaine, and the polymer also contains a temperature-sensitive monomer, which has temperature controllability in the recognition of template molecules morphine and thebaine, and the volume swells at low temperature, thereby helping the adsorption of morphine and thebaine, and the volume shrinks at high temperature, and the desorption of morphine and thebaine molecules, so that the affinity of the imprinting cavity combined with the template molecules changes to realize the efficient separation of the molecules, and the polymer also contains an ionic liquid monomer with an imidazole group, which has specific affinity for the indole structure of morphine and thebaine through π-π stacking and hydrogen bonding adsorption, thereby improving the adsorption speed of the adsorbent A on morphine and thebaine.

[0036] The adsorbent B couples chiral amino acids on the surface of MOF material UiO-66-NH2 through amino carboxyl water coupling, realizes specific adsorption of codeine through chiral cavity induction and electrostatic complementary mechanism. The pore size of MOF matches the size of codeine molecules, codeine is a tertiary amine alkaloid, and can be tightly combined with the side chain of the negatively charged amino acid by forming directional hydrogen bonds, so the adsorption capacity is large, and the prepared chiral amino acid coupled MOF is coupled with maleic acid, so that the surface of the prepared modified chiral amino acid coupled MOF has a double bond, which can be copolymerized with monomers. The temperature-sensitive monomer in the chiral amino acid coupled MOF changes the affinity of the imprinting cavity combined with the codeine molecules to realize the adsorption and desorption separation of the molecules.

[0037] The adsorbent C couples sulfonic acid groups and double bonds on the surface of graphene oxide material, realizes high-efficiency adsorption of papaverine and narcotine through sulfonic acid group ion exchange and π-π stacking, the sulfonic acid group (-SO3H) of GO-SO3H forms strong electrostatic action with the benzyl isoquinoline structure of papaverine and the isoquinoline structure of narcotine, thereby accelerating the separation and providing separation efficiency, in addition, the double bond can be copolymerized with monomers, and the temperature-sensitive monomer in the chiral amino acid coupled MOF changes the affinity of the imprinting cavity combined with the codeine molecules to realize the adsorption and desorption separation of the molecules. DETAILED DESCRIPTION

[0038] The technical solutions in the embodiments of the application will be described below clearly and completely. Obviously, the described embodiments are only part of the embodiments of the application, rather than all the embodiments. Based on the embodiments in the application, all other embodiments obtained by those skilled in the art without creative labor fall within the protection scope of the application.

[0039] Preparation Example 1 Adsorbent A

[0040] The preparation method is as follows:

[0041] 0.5 g of morphine, 0.5 g of thebaine, 8 g of monomers, 0.2 g of ethylene glycol dimethacrylate, 0.5 g of emulsifier and 0.1 g of cetyltrimethylammonium chloride were mixed into 200 mL of ethyl acetate, stirred and mixed for 15 min, added dropwise into 500 mL of water, emulsified at 5000 r / min for 15 min, 0.01 g of potassium persulfate was added under nitrogen protection, heated to 55°C, stirred for 4 h, the product was filtered, added into a Soxhlet extractor, extracted with a mixed solvent until no morphine and thebaine were detected, washed, dried, and adsorbent A was prepared;

[0042] The mixed solvent is a mixed solvent of acetonitrile and dichloromethane, with a volume ratio of 1:1;

[0043] The emulsifier is a mixture of Tween-80 and Span-80, with a mass ratio of 2:3;

[0044] The monomers include methyl methacrylate, styrene, 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethanesulfonyl) imide salt, and N-isopropyl acrylamide, with a mass ratio of 3:4:1:5.

[0045] Preparation Example 2 Adsorbent A

[0046] The preparation method is as follows:

[0047] 1 g of morphine, 1 g of thebaine, 12 g of monomers, 0.4 g of ethylene glycol dimethacrylate, 1 g of emulsifier and 0.2 g of cetyltrimethylammonium chloride were mixed into 200 mL of ethyl acetate, stirred and mixed for 15 min, added dropwise into 500 mL of water, emulsified at 5000 r / min for 15 min, 0.15 g of sodium persulfate was added under nitrogen protection, heated to 65°C, stirred for 6 h, the product was filtered, added into a Soxhlet extractor, extracted with a mixed solvent until no morphine and thebaine were detected, washed, dried, and adsorbent A was prepared;

[0048] The mixed solvent is a mixed solvent of acetonitrile and dichloromethane, with a volume ratio of 1:3;

[0049] The emulsifier is a mixture of Tween-80 and Span-80, with a mass ratio of 3:5;

[0050] The monomers include methyl methacrylate, styrene, 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethanesulfonyl) imide salt, and N-isopropyl acrylamide, with a mass ratio of 5:6:2:7.

[0051] Preparation Example 3 Adsorbent A

[0052] The preparation method is as follows:

[0053] 0.7 g of morphine, 0.7 g of thebaine, 10 g of monomers, 0.3 g of ethylene glycol dimethacrylate, 0.7 g of emulsifier and 0.15 g of cetyltrimethylammonium bromide were mixed into 200 mL of ethyl acetate, stirred for 15 min, added dropwise into 500 mL of water, emulsified at 5000 r / min for 15 min, 0.012 g of azobisisobutyronitrile was added under nitrogen protection, heated to 60°C, stirred for 5 h, the product was filtered, added into a Soxhlet extractor, extracted by mixed solvents until no morphine and thebaine were detected, washed, dried, and the adsorbent A was prepared;

[0054] The mixed solvent is a mixed solvent of acetonitrile and dichloromethane, and the volume ratio is 1:2;

[0055] The emulsifier is a mixture of Tween-80 and Span-80, and the mass ratio is 2.5:4;

[0056] The monomers include methyl methacrylate, styrene, 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethanesulfonyl) imide salt, and N-isopropyl acrylamide, and the mass ratio is 4:5:1.5:6.

[0057] Comparative Preparation Example 1

[0058] Compared with Preparation Example 3, the difference is that 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethanesulfonyl) imide salt is not added. The monomers include methyl methacrylate, styrene, and N-isopropyl acrylamide, and the mass ratio is 4:6.5:6.

[0059] Comparative Preparation Example 2

[0060] Compared with Preparation Example 3, the difference is that N-isopropyl acrylamide is not added. The monomers include methyl methacrylate, styrene, 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethanesulfonyl) imide salt, and N-isopropyl acrylamide, and the mass ratio is 4:11:1.5.

[0061] Preparation Example 4 Adsorbent B

[0062] The preparation method is as follows:

[0063] S1. Preparation of chiral amino acid coupled MOF: 2 g of L-aspartic acid and 1 g of L-glutamic acid were added into 200 mL of water, 2 g of N-hydroxysuccinimide and 2 g of 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirred for 30 min, 10 g of UiO-66-NH2 was added, stirred for 7 h, filtered, washed, and dried to prepare the chiral amino acid coupled MOF;

[0064] S2. Double bond graft modification: 2 g of maleic acid was added to 200 mL of water, 1 g of N-hydroxysuccinimide and 1 g of 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirred and activated for 30 min, 10 g of chiral amino acid coupled MOF was added, stirred and reacted for 5 h, filtered, washed, and dried to obtain the modified chiral amino acid coupled MOF;

[0065] S3. Copolymerization: 5 g of modified chiral amino acid coupled MOF, 4 g of methyl methacrylate, 3 g of styrene, and 6 g of N-isopropyl acrylamide were mixed and added to 200 mL of ethyl acetate, stirred and mixed for 10 min, added dropwise into 300 mL of water, emulsified at 7000 r / min for 15 min, 0.1 g of sodium persulfate was added under nitrogen protection, heated to 60°C, stirred and reacted for 3 h, the product was filtered to obtain adsorbent B.

[0066] Preparation Example 5 Adsorbent B

[0067] The preparation method is as follows:

[0068] S1. Preparation of chiral amino acid coupled MOF: 3 g of L-aspartic acid and 2 g of L-glutamic acid were added to 200 mL of water, 4 g of N-hydroxysuccinimide and 4 g of 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirred and activated for 30 min, 10 g of UiO-66-NH2 was added, stirred and reacted for 10 h, filtered, washed, and dried to obtain the chiral amino acid coupled MOF;

[0069] S2. Double bond graft modification: 4 g of maleic acid was added to 200 mL of water, 2 g of N-hydroxysuccinimide and 2 g of 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirred and activated for 30 min, 10 g of chiral amino acid coupled MOF was added, stirred and reacted for 7 h, filtered, washed, and dried to obtain the modified chiral amino acid coupled MOF;

[0070] S3. Copolymerization: 7 g of modified chiral amino acid coupled MOF, 6 g of methyl methacrylate, 5 g of styrene, and 8 g of N-isopropyl acrylamide were mixed and added to 200 mL of ethyl acetate, stirred and mixed for 10 min, added dropwise into 300 mL of water, emulsified at 7000 r / min for 15 min, 0.2 g of potassium persulfate was added under nitrogen protection, heated to 70°C, stirred and reacted for 5 h, the product was filtered to obtain adsorbent B.

[0071] Preparation Example 6 Adsorbent B

[0072] The preparation method is as follows:

[0073] S1. Preparation of chiral amino acid coupled MOF: 2.5 g L-aspartic acid, 1.5 g L-glutamic acid were added into 200 mL water, 3 g N-hydroxysuccinimide and 3 g 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirring activation for 30 min, 10 g UiO-66-NH2 was added, stirring reaction for 8 h, filtration, washing, drying, to obtain chiral amino acid coupled MOF;

[0074] S2. Double bond grafting modification: 3 g maleic acid was added into 200 mL water, 1.5 g N-hydroxysuccinimide and 1.5 g 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirring activation for 30 min, 10 g chiral amino acid coupled MOF was added, stirring reaction for 6 h, filtration, washing, drying, to obtain modified chiral amino acid coupled MOF;

[0075] S3. Copolymerization: 6 g modified chiral amino acid coupled MOF, 5 g methyl methacrylate, 4 g styrene, 7 g N-isopropyl acrylamide were mixed and added into 200 mL ethyl acetate, stirring mixing for 10 min, dropping into 300 mL water, emulsifying at 7000 r / min for 15 min, under nitrogen protection, 0.15 g ammonium persulfate was added, heating to 65℃, stirring reaction for 4 h, the product was filtered, to obtain adsorbent B.

[0076] Comparative Preparation Example 3

[0077] Compared with Preparation Example 3, the difference lies in that no L-aspartic acid is added in step S1.

[0078] Specifically as follows:

[0079] S1. Preparation of chiral amino acid coupled MOF: 2.5 g L-aspartic acid, 1.5 g L-glutamic acid were added into 200 mL water, 3 g N-hydroxysuccinimide and 3 g 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirring activation for 30 min, 10 g UiO-66-NH2 was added, stirring reaction for 8 h, filtration, washing, drying, to obtain chiral amino acid coupled MOF.

[0080] Comparative Preparation Example 4

[0081] Compared with Preparation Example 3, the difference lies in that no L-glutamic acid is added in step S1.

[0082] Specifically as follows:

[0083] S1. Preparation of chiral amino acid coupled MOF: 4 g L-aspartic acid was added to 200 mL water, 3 g N-hydroxysuccinimide and 3 g 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirred for activation for 30 min, 10 g UiO-66-NH2 was added, stirred for reaction for 8 h, filtered, washed, dried, and the chiral amino acid coupled MOF was prepared.

[0084] Comparative Preparation Example 5

[0085] Compared with Preparation Example 3, the difference is that step S1 is not performed.

[0086] The specific process is as follows:

[0087] S1. Double bond grafting modification: 3 g maleic acid was added to 200 mL water, 1.5 g N-hydroxysuccinimide and 1.5 g 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride were added, stirred for activation for 30 min, 10 g UiO-66-NH2 was added, stirred for reaction for 6 h, filtered, washed, dried, and the modified MOF was prepared;

[0088] S2. Copolymerization: 6 g modified MOF, 5 g methyl methacrylate, 4 g styrene, and 7 g N-isopropyl acrylamide were mixed and added to 200 mL ethyl acetate, stirred for mixing for 10 min, added dropwise to 300 mL water, emulsified at 7000 r / min for 15 min, 0.15 g ammonium persulfate was added under nitrogen protection, heated to 65°C, stirred for reaction for 4 h, and the product was filtered to prepare adsorbent B.

[0089] Preparation Example 7 Adsorbent C

[0090] The preparation method is as follows:

[0091] T1. Preparation of p-aminobenzenesulfonic acid diazonium salt solution: 0.04 g p-aminobenzenesulfonic acid, 1 g sodium nitrite were dissolved in 20 mL 2 wt% NaOH solution, heated to 50°C, stirred and mixed for 15 min, cooled to room temperature, 6 g 36 wt% hydrochloric acid was added dropwise, stirred for reaction for 3 h, and the p-aminobenzenesulfonic acid diazonium salt solution was prepared;

[0092] T2. Preparation of sulfonated / double bond co-modified graphene oxide: 2 g maleic acid was added to 200 mL toluene, 8 g graphene oxide and 0.5 g p-toluenesulfonic acid were added, heated to reflux and stirred for reaction for 4 h, centrifuged, washed, dried, and the product was added to 15 g p-aminobenzenesulfonic acid diazonium salt solution, stirred for reaction for 10 h, centrifuged, washed, dried, and the sulfonated / double bond co-modified graphene oxide was prepared;

[0093] T3. Copolymerization: 4 g of the sulfonated / double bond co-modified graphene oxide, 3 g of methyl methacrylate, 3 g of styrene, and 5 g of N-isopropyl acrylamide were mixed into 200 mL of ethyl acetate, stirred and mixed for 20 min, added dropwise into 300 mL of water, emulsified at 8000 r / min for 15 min, 0.1 g of sodium persulfate was added under nitrogen protection, heated to 55°C, and stirred for 4 h. The product was filtered to obtain adsorbent C.

[0094] Preparation Example 8: Adsorbent C

[0095] The preparation method is as follows:

[0096] T1. Preparation of p-aminobenzenesulfonic acid diazonium salt solution: 0.06 g of p-aminobenzenesulfonic acid and 1 g of sodium nitrite were dissolved in 20 mL of 2 wt% NaOH solution, heated to 50°C, stirred and mixed for 15 min, cooled to room temperature, 8 g of 36 wt% hydrochloric acid was added dropwise, and stirred for 5 h to obtain the p-aminobenzenesulfonic acid diazonium salt solution;

[0097] T2. Preparation of sulfonated / double bond co-modified graphene oxide: 3 g of maleic acid was added to 200 mL of toluene, 10 g of graphene oxide and 1 g of p-toluenesulfonic acid were added, heated to reflux and stirred for 7 h, centrifuged, washed and dried. The product was added to 20 g of p-aminobenzenesulfonic acid diazonium salt solution, stirred for 12 h, centrifuged, washed and dried to obtain the sulfonated / double bond co-modified graphene oxide;

[0098] T3. Copolymerization: 6 g of the sulfonated / double bond co-modified graphene oxide, 5 g of methyl methacrylate, 4 g of styrene, and 7 g of N-isopropyl acrylamide were mixed into 200 mL of ethyl acetate, stirred and mixed for 20 min, added dropwise into 300 mL of water, emulsified at 8000 r / min for 15 min, 0.2 g of ammonium persulfate was added under nitrogen protection, heated to 65°C, and stirred for 6 h. The product was filtered to obtain adsorbent C.

[0099] Preparation Example 9: Adsorbent C

[0100] The preparation method is as follows:

[0101] T1. Preparation of p-aminobenzenesulfonic acid diazonium salt solution: 0.05 g of p-aminobenzenesulfonic acid and 1 g of sodium nitrite were dissolved in 20 mL of 2 wt% NaOH solution, heated to 50°C, stirred and mixed for 15 min, cooled to room temperature, 7 g of 36 wt% hydrochloric acid was added dropwise, and stirred for 4 h to obtain the p-aminobenzenesulfonic acid diazonium salt solution;

[0102] T2. Preparation of sulfonated / double bond co-modified graphene oxide: 2.5 g of maleic acid was added to 200 mL of toluene, 9 g of graphene oxide and 0.7 g of p-toluenesulfonic acid were added, heated to reflux and stirred for 5.5 h, centrifuged, washed, dried, and the product was added to 17 g of p-aminobenzenesulfonic acid diazonium salt solution, stirred for 11 h, centrifuged, washed, dried, and sulfonated / double bond co-modified graphene oxide was prepared;

[0103] T3. Copolymerization: 5 g of sulfonated / double bond co-modified graphene oxide, 4 g of methyl methacrylate, 3.5 g of styrene, and 6 g of N-isopropyl acrylamide were mixed and added to 200 mL of ethyl acetate, stirred for 20 min, added dropwise to 300 mL of water, emulsified at 8000 r / min for 15 min, 0.15 g of potassium persulfate was added under nitrogen protection, heated to 60°C, and stirred for 5 h. The product was filtered to obtain adsorbent C.

[0104] Comparative Preparation Example 6

[0105] Compared with Preparation Example 9, the difference is that no sulfonation is performed in step T2.

[0106] The specific process is as follows:

[0107] T1. Preparation of double bond modified graphene oxide: 2.5 g of maleic acid was added to 200 mL of toluene, 9 g of graphene oxide and 0.7 g of p-toluenesulfonic acid were added, heated to reflux and stirred for 5.5 h, centrifuged, washed, dried, and double bond modified graphene oxide was prepared.

[0108] T2. Copolymerization: 5 g of double bond modified graphene oxide, 4 g of methyl methacrylate, 3.5 g of styrene, and 6 g of N-isopropyl acrylamide were mixed and added to 200 mL of ethyl acetate, stirred for 20 min, added dropwise to 300 mL of water, emulsified at 8000 r / min for 15 min, 0.15 g of potassium persulfate was added under nitrogen protection, heated to 60°C, and stirred for 5 h. The product was filtered to obtain adsorbent C.

[0109] Comparative Preparation Example 7

[0110] Compared with Preparation Example 9, the difference is that no N-isopropyl acrylamide is added in step T3.

[0111] The specific process is as follows:

[0112] T3. Copolymerization: 5 g of sulfonated / double bond co-modified graphene oxide, 4 g of methyl methacrylate, and 9.5 g of styrene were mixed into 200 mL of ethyl acetate, stirred and mixed for 20 min, added dropwise into 300 mL of water, emulsified at 8000 r / min for 15 min, added 0.15 g of potassium persulfate under nitrogen protection, heated to 60°C, stirred for 5 h, and the product was filtered to obtain adsorbent C. Example

[0113] The present embodiment provides a method for determining the content of codeine, morphine, papaverine, narcotine and thebaine in poppy oil, comprising the following steps:

[0114] (1) Preparation of isotope internal standard working solution: accurately pipette morphine-D3 isotope solution and codeine-D3 isotope solution into a volumetric flask, and dilute to the mark with water to prepare an internal standard working solution with a concentration of 5.0 μg / mL for morphine-D3 and codeine-D3;

[0115] (2) Preparation of test sample solution: add 1 g of adsorbent to 20 g of poppy oil sample, stir for 20 min, filter, add solvent to the solid, heat to 40°C, stir for 20 min, filter, wash, combine the washings and the filtrate, add 2 μg of isotope internal standard working solution, filter through a 0.22 μm filter, and dilute to the mark to prepare the test sample solution;

[0116] The solvent is a mixed solution of formic acid and acetonitrile with a volume ratio of 0.5:100;

[0117] The adsorbent includes adsorbent A prepared in Preparation Example 1, adsorbent B prepared in Preparation Example 4, and adsorbent C prepared in Preparation Example 7, with a mass ratio of 5:4:1;

[0118] (3) Preparation of control stock solution: the concentrations of codeine, morphine, papaverine, thebaine and narcotine stock solutions are 50.0 mg / L, 50.0 mg / L, 10.0 mg / L, 10.0 mg / L and 10.0 mg / L, respectively;

[0119] Prepare stock solution 1 (codeine 5000 ng / mL, morphine 5000 ng / mL, papaverine 1000 ng / mL, narcotine 1000 ng / mL, thebaine 1000 ng / mL): accurately pipette 0.5 mL of stock solution into a 5 mL volumetric flask, dilute to the mark with solvent and mix well, and label as stock solution 1.

[0120] Prepare L-1000% solution (codeine 1250 ng / mL, morphine 1250 ng / mL, papaverine 250 ng / mL, noscapine 250 ng / mL, thebaine 250 ng / mL) as follows: accurately transfer 5.0 mL of stock solution 1 to a 20 mL volumetric flask, dilute to the mark with solvent and mix well, and label as L-1000% solution;

[0121] (4) Preparation of control solution: prepare control solution (codeine 125 ng / mL, morphine 125 ng / mL, papaverine 25 ng / mL, noscapine 25 ng / mL, thebaine 25 ng / mL) as follows: accurately transfer 1.0 mL of L-1000% solution to a 10 mL volumetric flask, dilute to the mark with solvent and mix well, and label as control solution;

[0122] (5) Sensitivity preparation: prepare sensitivity solution (codeine 25 ng / mL, morphine 25 ng / mL, papaverine 5 ng / mL, noscapine 5 ng / mL, thebaine 5 ng / mL) as follows: accurately transfer 0.2 mL of L-1000% solution to a 10 mL volumetric flask, dilute to the mark with solvent and mix well, and label as sensitivity solution;

[0123] (6) Detection: use LC-MS / MS to determine the content of codeine, morphine, papaverine, noscapine and thebaine in opium;

[0124] The chromatographic conditions of the detection are as follows: chromatographic column: Agilent ZORBAX SB-Aq 3.5 μm 3.0×100 mm, column temperature 40℃, injection volume 5 μL, injection disc temperature 5℃, mobile phase: A: 5Mmol / L ammonium acetate + 0.1% formic acid (v / v); B: 0.1% formic acid acetonitrile solution (v / v), flow rate 0.3 mL / min, needle washing solution MeOH:H2O=1:1 (V:V); mass spectrometry conditions: ion source ESI, curtain gas 35 psi, temperature 600℃, ionization voltage 5500V, spray gas 50 psi, auxiliary heating gas 50 psi, scanning mode positive ion mode. Example

[0125] The present embodiment provides a method for determining the content of codeine, morphine, papaverine, noscapine and thebaine in opium, comprising the following steps:

[0126] (1) Preparation of isotopic internal standard working solution: accurately pipette morphine-D3 isotopic solution and codeine-D3 isotopic solution into a volumetric flask, dilute to the mark with water, and prepare an internal standard working solution with a concentration of 5.0 μg / mL for morphine-D3 and codeine-D3;

[0127] (2) Preparation of test solution: 1 g of adsorbent was added to 20 g of opium oil sample, stirred for 20 min, filtered, and the solid was added to solvent and heated to 45℃, stirred for 40 min, filtered, washed, and the washing liquid was combined with the filtrate, 2 μg of isotope internal standard working solution was added, filtered through a 0.22 μm filter, and made up to volume to prepare the test solution;

[0128] The solvent is a mixed solution of formic acid and acetonitrile in a volume ratio of 1:100;

[0129] The adsorbent includes adsorbent A prepared in Preparation Example 2, adsorbent B prepared in Preparation Example 5, and adsorbent C prepared in Preparation Example 8, in a mass ratio of 7:6:3;

[0130] (3) Preparation of control stock solution: the concentrations of the stock solutions of codeine, morphine, papaverine, thebaine and noscapine were 50.0 mg / L, 50.0 mg / L, 10.0 mg / L, 10.0 mg / L and 10.0 mg / L, respectively;

[0131] Prepare stock solution 1: (codeine 5000 ng / mL, morphine 5000 ng / mL, papaverine 1000 ng / mL, noscapine 1000 ng / mL, thebaine 1000 ng / mL), as follows: accurately transfer 0.5 mL of stock solution to a 5 mL volumetric flask, make up to the mark with solvent and mix well, and label as stock solution 1.

[0132] Prepare L-1000% solution (codeine 1250 ng / mL, morphine 1250 ng / mL, papaverine 250 ng / mL, noscapine 250 ng / mL, thebaine 250 ng / mL), as follows: accurately transfer 5.0 mL of stock solution 1 to a 20 mL volumetric flask, make up to the mark with solvent and mix well, and label as L-1000% solution;

[0133] (4) Preparation of control solution: prepare control solution (codeine 125 ng / mL, morphine 125 ng / mL, papaverine 25 ng / mL, noscapine 25 ng / mL, thebaine 25 ng / mL), as follows: accurately transfer 1.0 mL of L-1000% solution to a 10 mL volumetric flask, make up to the mark with solvent and mix well, and label as control solution;

[0134] (5) Sensitivity preparation: prepare sensitivity solution (codeine 25 ng / mL, morphine 25 ng / mL, papaverine 5 ng / mL, noscapine 5 ng / mL, thebaine 5 ng / mL), as follows: accurately transfer 0.2 mL of L-1000% solution to a 10 mL volumetric flask, make up to the mark with solvent and mix well, and label as sensitivity solution;

[0135] (6) Detection: LC-MS / MS was used to detect the contents of codeine, morphine, papaverine, noscapine and thebaine in poppy oil;

[0136] The chromatographic conditions of the detection are as follows: chromatographic column: Agilent ZORBAX SB-Aq 3.5 μm 3.0x100 mm, column temperature 40℃, injection volume 5 μL, injection disc temperature 5℃, mobile phase: A: 5Mmol / L ammonium acetate + 0.1% formic acid (v / v); B: 0.1% formic acid acetonitrile solution (v / v), flow rate 0.3 mL / min, needle washing liquid MeOH:H2O=1:1 (V:V); mass spectrometry conditions: ion source ESI, curtain gas 35 psi, temperature 600℃, ionization voltage 5500V, spray gas 50 psi, auxiliary heating gas 50 psi, scanning mode positive ion mode. Example

[0137] The present embodiment provides a method for determining the contents of codeine, morphine, papaverine, noscapine and thebaine in poppy oil, comprising the following steps:

[0138] (1) Preparation of isotopic internal standard working solution: accurately pipette morphine-D3 isotopic solution and codeine-D3 isotopic solution into a volumetric flask, and dilute to the mark with water to prepare an internal standard working solution with a concentration of 5.0 μg / mL for morphine-D3 and codeine-D3;

[0139] (2) Preparation of test sample solution: add 1 g of adsorbent to 20 g of poppy oil sample, stir for 20 min, filter, add solvent to the solid, heat to 42℃, stir for 30 min, filter, wash, combine the washing liquid and the filtrate, add 2 μg of isotopic internal standard working solution, filter through a 0.22 μm filter membrane, and dilute to prepare a test sample solution;

[0140] The solvent is a mixed solution of formic acid and acetonitrile with a volume ratio of 0.7:100;

[0141] The adsorbent includes adsorbent A prepared in Preparation Example 3, adsorbent B prepared in Preparation Example 6 and adsorbent C prepared in Preparation Example 9, and the mass ratio is 6:5:2;

[0142] (3) Preparation of control stock solution: the concentrations of codeine, morphine, papaverine, thebaine and noscapine stock solutions are 50.0 mg / L, 50.0 mg / L, 10.0 mg / L, 10.0 mg / L and 10.0 mg / L, respectively;

[0143] Preparation of stock solution 1: (codeine 5000 ng / mL, morphine 5000 ng / mL, papaverine 1000 ng / mL, noscapine 1000 ng / mL, thebaine 1000 ng / mL), as follows: precisely transfer 0.5 mL stock solution into a 5 mL volumetric flask, dilute to the mark with solvent and mix well, and mark as stock solution 1.

[0144] Preparation of L-1000% solution (codeine 1250 ng / mL, morphine 1250 ng / mL, papaverine 250 ng / mL, noscapine 250 ng / mL, thebaine 250 ng / mL), as follows: precisely transfer 5.0 mL stock solution 1 into a 20 mL volumetric flask, dilute to the mark with solvent and mix well, and mark as L-1000% solution.

[0145] (4) Preparation of control solution: prepare control solution (codeine 125 ng / mL, morphine 125 ng / mL, papaverine 25 ng / mL, noscapine 25 ng / mL, thebaine 25 ng / mL), as follows: precisely transfer 1.0 mL L-1000% solution into a 10 mL volumetric flask, dilute to the mark with solvent and mix well, and mark as control solution.

[0146] (5) Sensitivity preparation: prepare sensitivity solution (codeine 25 ng / mL, morphine 25 ng / mL, papaverine 5 ng / mL, noscapine 5 ng / mL, thebaine 5 ng / mL), as follows: precisely transfer 0.2 mL L-1000% solution into a 10 mL volumetric flask, dilute to the mark with solvent and mix well, and mark as sensitivity solution; limit of detection (LOD) is also called sensitivity; limit of detection refers to the minimum amount of the measured substance in the sample that can be detected. Here, the method for configuring the limit of quantification (LOQ) is described; limit of quantification refers to the minimum amount of the measured substance in the sample that can be quantitatively determined.

[0147] (6) Detection: LC-MS / MS is used to determine the content of codeine, morphine, papaverine, noscapine and thebaine in opium;

[0148] The chromatographic conditions for the detection are as follows: chromatographic column: Agilent ZORBAX SB-Aq 3.5 μm 3.0 x 100 mm, column temperature 40 °C, injection volume 5 μL, injection disc temperature 5 °C, mobile phase: A: 5 Mmol / L ammonium acetate + 0.1% formic acid (v / v); B: 0.1% formic acid acetonitrile solution (v / v), flow rate 0.3 mL / min, needle washing solution MeOH:H2O=1:1 (V:V); mass spectrometry conditions: ion source ESI, curtain gas 35 psi, temperature 600 °C, ionization voltage 5500 V, spray gas 50 psi, auxiliary heating gas 50 psi, scanning mode positive ion mode.

[0149] Comparative Example 1-2

[0150] The difference from Example 3 is that the adsorbent A is prepared by Comparative Preparation Example 1-2.

[0151] Comparative Example 3-5

[0152] The difference from Example 3 is that the adsorbent B is prepared by Comparative Preparation Example 3-5.

[0153] Comparative Example 6-7

[0154] The difference from Example 3 is that the adsorbent C is prepared by Comparative Preparation Example 6-7. Example

[0155] Limit of detection (LOD):

[0156] The results are shown in Table 1.

[0157] Table 1

[0158]

[0159] From the above table, the signal-to-noise ratio meets the requirements. The signal-to-noise ratio is used for an analytical method that can show baseline noise, that is, the signal measured by a known low concentration sample is compared with the signal measured by a blank sample, and the lowest concentration or amount of the measured substance that can be reliably detected is calculated. Generally, the detection limit is determined when the signal-to-noise ratio is 3:1.

[0160] Limit of quantitation (LOQ):

[0161] The results are shown in Table 2. Generally, the limit of quantitation is determined when the signal-to-noise ratio is 10:1.

[0162] Table 2

[0163]

[0164] From the above table, the signal-to-noise ratio of the limit of quantitation meets the requirements.

[0165] Linear range: The linear range is within the LOQ to 200% limit concentration range, and the regression coefficient (r) is ≥0.990, which is good in linear.

[0166] The results are shown in Table 3.

[0167] Table 3

[0168]

[0169] (4) Accuracy

[0170] The average recovery rate of the sample spiked solution at the three limit levels of accuracy of 50%, 100% and 150% was between 70.0% and 130.0%, and the precision of the recovery rate of the 9 spiked sample solutions was not more than 15.0%. The results are shown in Tables 4 to 8.

[0171] Table 4 Codeine

[0172]

[0173] Table 5 Morphine

[0174]

[0175] Table 6 Papaverine

[0176]

[0177] Table 7 Narcotine

[0178]

[0179] Table 8 Thebaine

[0180]

[0181] Test Example 1

[0182] The average recovery rate and precision of the spiked detection results of the results of the determination of the contents of codeine, morphine, papaverine, narcotine and thebaine in the same batch of poppy oil sample (determined by the methods in Example 3 and Comparative Examples 1-7, respectively) are shown in Tables 9-13.

[0183] Table 9 Codeine

[0184]

[0185] Table 10 Morphine

[0186]

[0187] Table 11 Papaverine

[0188]

[0189] Table 12 Narcotine

[0190]

[0191] Table 13 Thebaine

[0192]

[0193] From the above table, the average recovery of codeine, morphine, papaverine, noscapine and thebaine determined by the methods in Examples 1-3 of the present application was 93.60-98.96%, and the precision was 1.2-3.3%.

[0194] The above description is merely preferred embodiments of the present application, but not to limit the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A method for determining the content of codeine, morphine, papaverine, noscapine and thebaine in poppy oil, characterized in that, The method comprises the following steps: (1) Preparation of an isotopic internal standard working solution: precisely pipette morphine-D3 and codeine-D3 isotopic solutions into a volumetric flask, respectively, and dilute to volume to prepare morphine-D3 and codeine-D3 internal standard working solutions; (2) Preparation of a test sample solution: add an adsorbent to the opium oil sample, stir and adsorb, filter, add the solid to a solvent, heat and desorb, filter, wash, combine the washing liquid and the filtrate, add the isotopic internal standard working solution, filter, and dilute to volume to prepare a test sample solution; (3) Preparation of a control stock solution: prepare a corresponding stock solution by using codeine, morphine, papaverine, thebaine and noscapine standard samples, respectively; (4) Preparation of a control sample solution: dilute the control stock solution to prepare a control sample solution; (5) Detection: LC-MS / MS is used to detect the contents of codeine, morphine, papaverine, noscapine and thebaine in the opium oil; The adsorbent comprises adsorbent A, adsorbent B and adsorbent C, and the mass ratio is 5-7:4-6:1-3; The preparation method of the adsorbent A is as follows: The morphine, thebaine, monomer, crosslinking agent, emulsifier and pore-forming agent are mixed and added into ethyl acetate, uniformly mixed, added dropwise into water, emulsified, an initiator is added under the protection of inert gas, heated and stirred to react, the product is filtered, added into a Soxhlet extractor, extracted by mixed solvents until no morphine and thebaine are detected, washed, dried, and the adsorbent A is prepared; The preparation method of the adsorbent B is as follows: S1. Preparation of a chiral amino acid coupled MOF: L-aspartic acid and L-glutamic acid are added into water, N-hydroxysuccinimide and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride are added, stirred and activated, UiO-66-NH2 is added, stirred to react, filtered, washed, and dried to prepare a chiral amino acid coupled MOF; S2. Double bond grafting modification: maleic acid is added into water, N-hydroxysuccinimide and 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride are added, stirred and activated, the chiral amino acid coupled MOF is added, stirred to react, filtered, washed, and dried to prepare a modified chiral amino acid coupled MOF; S3. Copolymerization: the modified chiral amino acid coupled MOF, methyl methacrylate, styrene and N-isopropyl acrylamide are mixed and added into ethyl acetate, uniformly mixed, added dropwise into water, emulsified, an initiator is added under the protection of inert gas, heated and stirred to react, the product is filtered, and the adsorbent B is prepared; The preparation method of the adsorbent C is as follows: T1. Preparation of a p-aminobenzenesulfonic acid diazonium salt solution: p-aminobenzenesulfonic acid and sodium nitrite are dissolved in an alkali solution, uniformly mixed after heating, cooled to room temperature, hydrochloric acid is added dropwise, stirred to react, and the p-aminobenzenesulfonic acid diazonium salt solution is prepared; T2. Preparation of sulfonated / double bond co-modified graphene oxide: maleic acid is added into toluene, graphene oxide and p-methylbenzenesulfonic acid are added, heated to reflux and stirred to react, centrifuged, washed, dried, the product is added into the p-aminobenzenesulfonic acid diazonium salt solution, stirred to react, centrifuged, washed, and dried to prepare sulfonated / double bond co-modified graphene oxide. T3. Copolymerization: sulfonated / double bond co-modified graphene oxide, methyl methacrylate, styrene, N-isopropyl acrylamide were mixed into ethyl acetate, mixed uniformly, dropped into water, emulsified, under the protection of inert gas, initiator was added, heated and stirred to react, the product was filtered to obtain adsorbent C.

2. The assay method according to claim 1, characterized by In the preparation process of adsorbent A, the mass ratio of the monomers including methyl methacrylate, styrene, 1-vinyl-3-(aminopropyl) imidazole bis(trifluoromethanesulfonyl) imidazolium salt and N-isopropyl acrylamide is 3-5:4-6:1-2:5-7, the crosslinking agent is ethylene glycol dimethacrylate, the emulsifier is a mixture of Tween-80 and Span-80 with a mass ratio of 2-3:3-5, the pore former is cetyltrimethylammonium chloride or cetyltrimethylammonium bromide, the initiator is at least one selected from azobisisobutyronitrile, sodium persulfate, potassium persulfate and ammonium persulfate, the temperature of the heating and stirring reaction is 55-65℃, the time is 4-6h, the mass ratio of morphine, thebaine, monomers, crosslinking agent, emulsifier, pore former and initiator is 0.5-1:0.5-1:8-12:0.2-0.4:0.5-1:0.1-0.2:0.01-0.15, and the mixed solvent is a mixture of acetonitrile and dichloromethane with a volume ratio of 1:1-3.

3. The assay method according to claim 1, characterized by, The mass ratio of L-aspartic acid, L-glutamic acid, N-hydroxysuccinimide, 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride and UiO-66-NH2 in step S1 is 2-3:1-2:2-4:2-4:10; the mass ratio of maleic acid, N-hydroxysuccinimide, 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride and chiral amino acid coupled MOF in step S2 is 2-4:1-2:1-2:10; the mass ratio of modified chiral amino acid coupled MOF, methyl methacrylate, styrene, N-isopropyl acrylamide and initiator in step S3 is 5-7:4-6:3-5:6-8:0.1-0.2, the temperature of the heating and stirring reaction is 60-70℃, the time is 3-5h, and the initiator is at least one selected from sodium persulfate, potassium persulfate and ammonium persulfate.

4. The assay method according to claim 1, characterized by, The mass ratio of p-aminobenzenesulfonic acid, sodium nitrite and hydrochloric acid in step T1 is 0.04-0.06:1:6-8, and the stirring reaction time is 3-5h; the mass ratio of maleic acid, graphene oxide, p-toluenesulfonic acid and p-aminobenzenesulfonic acid diazonium salt solution in step T2 is 2-3:8-10:0.5-1:15-20, the heating and reflux stirring reaction time is 4-7h, and the stirring reaction time is 10-12h; the mass ratio of sulfonated / double bond co-modified graphene oxide, methyl methacrylate, styrene, N-isopropyl acrylamide and initiator in step T3 is 4-6:3-5:3-4:5-7:0.1-0.2, and the heating and stirring reaction time is 55-65℃, the time is 4-6h.

5. The assay method according to claim 1, characterized by, The detection chromatographic conditions are as follows: chromatographic column: Agilent ZORBAX SB-Aq 3.5μm 3.0×100mm, column temperature 35-45℃, sample disc temperature 3-7℃, mobile phase: A: 2-7Mmol / L ammonium acetate + 0.05-0.15% formic acid (v / v); B: 0.05-0.15% formic acid acetonitrile solution (v / v), flow rate 0.1-0.5mL / min, needle washing solution MeOH:H2O=1:1 (V:V).

6. The assay method according to claim 1, characterized by, The mass spectrometry conditions of the detection are as follows: ion source ESI, curtain gas 30-40psi, temperature 550-650℃, ionization voltage 5000-6000V, spray gas 45-55psi, auxiliary heating gas 45-55psi, scanning mode positive ion mode.

7. The assay method according to claim 1, characterized by Wherein, The linear range of codeine is 62.5ng / mL-187.5ng / mL, the linear range of morphine is 62.5ng / mL-187.5ng / mL, the linear range of papaverine is 12.5ng / mL-37.5ng / mL, the linear range of thebaine is 12.5ng / mL-37.5ng / mL, and the linear range of tybaine is 12.5ng / mL-37.5ng / mL.

8. The assay method according to claim 1, characterized by Wherein, The regression coefficient of codeine is 0.995, the regression coefficient of morphine is 0.994, the regression coefficient of papaverine is 0.995, the regression coefficient of thebaine is 0.998, and the regression coefficient of tybaine is 0.

999.

9. The assay method of claim 1, wherein The mass ratio of the poppy oil, adsorbent, and isotopic internal standard working solution is 15-20:1-2:0.000001-0.000002, the solvent is a mixed solution of formic acid and acetonitrile with a volume ratio of 0.5-1:100, the temperature of the heating desorption is 40-45℃, and the time is 20-40min.

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