Pretreatment method and detection method for detecting medicines in plasma

Abstract

The invention belongs to the field of drug detection, and particularly discloses a pretreatment method and a detection method for detecting drugs in plasma. The pretreatment method comprises the following steps: mixing a plasma sample containing metformin, aripiprazole or amisulpride micromolecule drugs after protein removal with carboxylated TiO2 (at) MWCNTs, and carrying out solid-phase dispersion extraction to obtain a supernatant which is used as a subsequent LC-MS / MS injection sample for detection. The carboxylation modified TiO2 (at) MWCNTs are used as an extraction / adsorption material for solid-phase dispersion extraction, so that the interference of matrixes such as phospholipid and the like is greatly reduced, the influence of an ion inhibition phenomenon and background noise on the quantification of small molecule drugs in the subsequent detection and analysis process is reduced, the sensitivity in LC-MS / MS detection is improved, and the linear detection range is expanded; in addition, the method is simple, convenient, rapid and low in cost.

Description

Technical Field

[0001] The present invention belongs to the field of drug detection, and in particular relates to a pretreatment method and a detection method for detecting drugs in plasma. Background Art

[0002] Liquid chromatography-mass spectrometry (LC-MS / MS) is increasingly used in routine clinical laboratories due to its high specificity, high sensitivity, and multi-analyte potential. Sample pretreatment is essential for LC-MS / MS analysis because untreated plasma or urine samples are directly introduced into the ionization source. Interfering substances in these samples (such as hydrogen, sodium, and ammonium) can suppress or enhance the analyte ions. Among these interfering substances, phospholipids, commonly found in clinical samples, have a significant inhibitory effect on ions. Matrix effects can compromise the precision and accuracy of bioanalytical methods, thus affecting the quality of the results.

[0003] Common sample pretreatment methods include protein precipitation (PPT), liquid-liquid extraction (LLE), and solid-phase extraction (SPE). Numerous studies have reported that extracts after PPT contain the highest levels of substances that inhibit analyte ionization compared to SPE and LLE. Currently, using pure acetonitrile to elute drugs from silica-based sorbents is a strategy that reduces phospholipids and, therefore, matrix effects. Another strategy for reducing matrix effects is using a MeOH:H₂O solution containing less than 50% methanol to elute drugs from silica and polymer-based sorbents during SPE, thereby eliminating lysophosphatidylcholine (LPC) from the extract. Selecting organic solvents and optimizing the pH of LLE are also strategies for minimizing phospholipid extraction and matrix effects. Using chloroform or a 80:20% (v / v) mixture of MTBE:chloroform as the extraction solvent can reduce or eliminate matrix effects caused by lysophosphatidylcholine (LPC) and phosphatidylcholine (PC) in LLE. Furthermore, the hybrid SPE-PPT platform with zirconium oxide-coated particles enables efficient extraction of phospholipids; Captiva ND lipid filter plates remove LPCs and PCs from biological fluids without retaining target analytes.

[0004] While using an eluent containing less than 50% methanol to elute the drug in SPE eliminates phosphatidylcholine from the extract, this can significantly impact drug recovery. LLE provides a very clean extraction of the analyte from the matrix and is the most thorough method for removing phospholipids. However, LLE involves selecting an appropriate extraction solvent, extracting the supernatant, and then evaporating and re-dissolving it, often taking several hours. The manual labor associated with these steps is tedious and time-consuming. Summary of the Invention

[0005] The purpose of the present invention is to address the deficiencies of the prior art and provide a rapid, convenient, and efficient pretreatment method and detection method for detecting drugs in plasma. Based on this, the first aspect of the present invention provides a pretreatment method for detecting drugs in plasma, comprising the following steps: mixing a protein-depleted plasma sample with a composite material for solid phase dispersion extraction to obtain a supernatant;

[0006] The plasma sample includes a small molecule drug, wherein the small molecule drug is metformin, aripiprazole or amisulpride;

[0007] The composite material is carboxylated TiO2@MWCNTs; TiO2@MWCNTs are multi-walled carbon nanotubes coated with TiO2.

[0008] The present invention uses carboxyl-modified TiO2@MWCNTs as the extraction / adsorption material for solid phase dispersion extraction. The titanium atoms bonded to the surface of multi-walled carbon nanotubes act as Lewis acids, and utilize the highly selective Lewis acid-base interaction characteristics to interact with the phospholipid components in the plasma sample, thereby achieving the purpose of removing phospholipids from the plasma sample; at the same time, the large specific surface area of ​​multi-walled carbon nanotubes is utilized to adsorb other matrix interfering substances in the plasma sample, thereby reducing the quantitative impact of ion suppression and background noise on the analyte (small molecule drug) in subsequent detection and analysis. Moreover, since phospholipids are a class of amphiphilic compounds with phosphate groups, they contain both a hydrophobic tail composed of fatty acid chains and a hydrophilic head composed of substituted groups connected to phosphates. Therefore, they generally do not show electrical properties. However, in the carboxyl-modified TiO2@MWCNTs used in the present invention, during the solid-phase separation process, the titanium atoms can bond with the phosphate groups, while the carboxyl groups can combine with the positively charged nitrogen atoms in the trimethylamine groups, thereby further enhancing the adsorption and separation effect of phospholipids, thereby further reducing the impact of phospholipids on analytes (small molecule drugs) during subsequent detection.

[0009] Furthermore, the carboxyl-modified TiO2@MWCNTs employed in the present invention exhibit stronger electronegativity than pure titanium dioxide, are less likely to aggregate in a plasma environment, and exhibit better dispersibility. They also exhibit superior adsorption properties for phospholipids. Compared to the PPT method used in the prior art, the present invention's pretreatment method requires only one additional vortex centrifugation step, making it simple and rapid. Furthermore, the pretreatment method requires only a small amount of carboxyl-modified TiO2@MWCNTs, as low as 2 mg, resulting in a relatively low cost.

[0010] Among them, according to experimental research, the pretreatment method of the present invention can only be applied to the UPLC-MS / MS quantitative detection of three drugs: metformin (MET), amisulpride (ASP), and aripiprazole (APZ); it is not applicable to other small molecule drugs, such as curcumin and omeprazole.

[0011] In addition, possibly because amisulpride contains a sulfonic acid group, the pretreatment method of the present invention has a better effect when applied to amisulpride in terms of matrix effect, accuracy, and precision.

[0012] In some preferred implementations, the preparation method of the composite material comprises the following steps: stirring tetrabutyl titanate and nitric acid in an aqueous solution at 70°C-95°C for 6h-24h (preferably at 70°C for 12h) to obtain a colloidal solution, filtering (using a 0.22 μm PTFE filter membrane), and then redissolving (adding water to redissolve), then adding multi-walled carbon nanotubes, stirring at 70°C-95°C for 12h-24h (preferably at 90°C for 12h), centrifuging to obtain a powder, washing (with water and ethanol in sequence), and drying, and then drying at 5 Annealing at 500°C-600°C for 2-8 hours (preferably annealing at 550°C for 4 hours) to obtain the TiO2@MWCNTs (the mass ratio of TiO2 to multi-walled carbon nanotubes is (0.1-1):1); dispersing the TiO2@MWCNTs in water, adding chloroacetic acid, and stirring at 60°C-95°C for 1-6 hours to perform a carboxylation reaction (preferably stirring at 95°C for 2 hours; relatively high temperatures are more conducive to the carboxylation reaction); and centrifuging to obtain the carboxylated TiO2@MWCNTs. The mixture can be further washed with deionized water, and then an AgNO3 aqueous solution is added to detect the AgCl content. The mixture is then washed with deionized water until no AgCl is detected, and then centrifuged at 10,000 rpm for 15 minutes to obtain a precipitate that is dried at 60°C for 24 hours.

[0013] The volume ratio of tetrabutyl titanate to nitric acid is (5-20) mL:1 mL (preferably 10 mL:1 mL). The concentration of the colloidal solution obtained after filtration and redissolution is 1 mg / mL-10 mg / mL (preferably 2 mg / mL). The ratio of the colloidal solution obtained after filtration and redissolution to multi-walled carbon nanotubes is 100 mL:(100-1000) mg (preferably 100 mL:200 mg). The ratio of the TiO2@MWCNTs to chloroacetic acid is 1 g:(2-30) mL (preferably 1 g:10 mL).

[0014] By controlling the amount of chloroacetic acid added, TiO2@MWCNTs with different degrees of carboxylation can be prepared (the number of carboxylic acid groups on the surface of carboxylated TiO2@MWCNTs can be determined by acid-base titration), thereby controlling the electronegativity of the final product.

[0015] In some preferred embodiments, the solid phase dispersion extraction includes the following steps: mixing the protein-removed plasma sample with the composite material, vortexing for 0.5 min-3 min, and centrifuging for 5 min-10 min at 4°C-25°C and 12000 rpm-15000 rpm (preferably vortexing for 2 min and centrifuging for 10 min at 4°C and 14000 rpm).

[0016] In some preferred embodiments, the method for preparing the protein-removed plasma sample comprises the following steps: mixing the plasma sample with acetonitrile in a volume ratio of 1:(3-5), vortexing for 0.5-3 min, and centrifuging at 4°C-25°C and 12,000-15,000 rpm for 5-10 min (preferably vortexing for 1 min and centrifuging at 4°C and 14,000 rpm for 10 min). The supernatant obtained is the protein-removed plasma sample.

[0017] A second aspect of the present invention provides a method for detecting a drug in plasma, wherein the drug is metformin, aripiprazole, or amisulpride; the method comprises the following steps: using the supernatant obtained by the pretreatment method as an injection sample, and detecting the injection sample using LC-MS / MS. The LC process utilizes a C18 column for chromatographic separation, with ultrapure water containing 2 mM ammonium formate and 0.0012% formic acid as mobile phase A, and acetonitrile as mobile phase B. The gradient elution conditions are: 0.0 minute, 65% A-35% B; 1.51 minutes, 10% A-90% B; 3 minutes, 0% A-100% B; and 3.01 minutes-5 minutes, 65% A-35% B.

[0018] Based on the above pretreatment method, the sensitivity of the small molecule drugs to be tested (metformin, aripiprazole or amisulpride) in LC-MS / MS detection can be improved and the linear detection range can be expanded.

[0019] The beneficial effects of the present invention are as follows: the present invention uses carboxyl-modified TiO2@MWCNTs as the extraction / adsorption material for solid-phase dispersion extraction, which greatly reduces matrix interference such as phospholipids, reduces the ion suppression phenomenon and background noise in the subsequent detection and analysis process on the quantitative influence of small molecule drugs, improves the sensitivity in LC-MS / MS detection, and expands the linear detection range; in addition, the method of the present invention is simple, fast and low-cost. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Shown is the mass spectrometry spectrum of aripiprazole;

[0021] Figure 2 Shown is the mass spectrometry spectrum of amisulpride;

[0022] Figure 3 Shown is the mass spectrometry spectrum of metformin;

[0023] Figure 4 Shown is the mass spectrometry spectrum of reserpine;

[0024] Figure 5 Shown is the peak diagram of the UPLC-MS / MS process for detecting ion channels of rat plasma samples containing metformin after pretreatment;

[0025] Figure 6 Shown is the peak diagram of the UPLC-MS / MS process for detecting ion channels of rat plasma samples containing amisulpride after pretreatment;

[0026] Figure 7 Shown is the peak diagram of the UPLC-MS / MS process for detecting ion channels of rat plasma samples containing aripiprazole after pretreatment;

[0027] Figure 8 Shown is the peak diagram of the UPLC-MS / MS process for detecting ion channels of rat plasma samples containing reserpine after pretreatment;

[0028] Figure 9 Shown is the peak shape diagram of the UPLC-MS / MS detection channel of the rat plasma sample containing curcumin after protein precipitation only;

[0029] Figure 10 Shown is the peak shape diagram of the UPLC-MS / MS detection channel of the rat plasma sample containing curcumin after pretreatment;

[0030] Figure 11 The figure shows the linear concentration curve of omeprazole-containing rat plasma samples detected by UPLC-MS / MS after pretreatment. DETAILED DESCRIPTION

[0031] The following will provide a clear and complete description of the concept and technical effects of the present invention in conjunction with the embodiments and drawings to fully understand the purpose, scheme and effects of the present invention.

[0032] In the following description, vortexing was performed using a "Vortex Mixer XW-80A, Germany" and centrifugation was performed using an "Eppendorf 5418R, Germany".

[0033] The preparation method of carboxylated TiO2@MWCNTs includes the following steps:

[0034] (1) 10 mL of tetrabutyl titanate, 1 mL of nitric acid and 100 mL of deionized water were stirred at 70°C for 12 hours to obtain a uniform milky white colloidal solution; the solution was filtered using a 0.22 μm PTFE filter membrane, and then deionized water was added to adjust the concentration of the colloidal solution to 2 mg / mL; 100 mL of the colloidal solution prepared above was taken, 200 mg of multi-walled carbon nanotubes were added, and the mixture was stirred at 90°C for 12 hours, and then centrifuged at 10,000 rpm for 10 minutes to obtain a white powder, which was washed with deionized water and ethanol in turn, and then dried in a forced air drying oven at 120°C for 2 hours; finally, the mixture was annealed at 550°C in a tube furnace for 4 hours to obtain TiO2@MWCNTs powder, i.e., TiO2-coated multi-walled carbon nanotube powder;

[0035] (2) 1 g of the TiO2@MWCNTs synthesized in step (1) was dispersed into 50 mL of deionized water, ultrasonically dispersed for 10 min, and then 10 mL of chloroacetic acid was added and mixed. The mixture was heated at 95°C and stirred for 2 hours to perform a carboxylation reaction. The mixture was then centrifuged at 10,000 rpm for 15 min to obtain carboxylated TiO2@MWCNTs. After washing twice with deionized water, an AgNO3 aqueous solution was added. No AgCl was detected. The mixture was then rinsed twice with deionized water and centrifuged at 10,000 rpm for 1 minute.

[0036] The centrifugal precipitate was dried in a vacuum oven at 60 °C for 24 h.

[0037] A method for detecting drugs in plasma includes a pretreatment process and a detection process; wherein the pretreatment process includes the following steps:

[0038] Step 1: Add 300 μL of acetonitrile to 100 μL of a rat plasma sample (Nanjing Senbei Coffee) containing a small molecule drug to obtain a mixture. Vortex the mixture for 1 minute and then centrifuge it at 14,000 rpm for 10 minutes at 4°C. The supernatant is obtained, i.e., the protein-depleted plasma sample.

[0039] Step 2: Add 2 mg of carboxylated TiO2@MWCNTs and protein-depleted plasma sample to a 1.5 mL EP tube, vortex for 2 minutes, and then centrifuge at 14,000 rpm for 10 minutes at 4°C to obtain the supernatant after solid phase dispersion extraction.

[0040] The detection process includes the following steps: taking 100 μL of the supernatant prepared in the above step 2 as an injection sample and adding it into an injection bottle for LC-MS / MS detection and analysis.

[0041] The above detection method was used to detect the small molecule drugs metformin MET (LogP = -2.31), aripiprazole APZ (LogP = 5.59), and amisulpride ASP (LogP = 1.60). An ultra-high performance liquid chromatography (UPLC) system (Waters) was used at 40 ° C using a C18 column (3 μm, 100 mm × 2.1 mm, Chromatographic separation was performed using ultrapure water containing 2 mM ammonium formate and 0.0012% formic acid as mobile phase A, and acetonitrile as mobile phase B. The gradient elution flow rate was 0.3 mL / min. The gradient elution conditions were: 65% A to 35% B (0.0 min); 10% A to 90% B (1.51 min); 0% A to 100% B (3 min); and 65% A to 35% B (3.01 min to 5 min). The sample vial was placed in an autosampler at 15°C, and 100 μL of the rat plasma matrix solution containing the small molecule drug was injected into the LC system for analysis.

[0042] A triple quadrupole mass spectrometer (AB Sciex, Triple Quad 4500) coupled to a LC system was used for mass spectrometry analysis. Electrospray ionization was used in positive ion mode. The MS / MS product ion scan function was used for tandem mass spectrometry analysis. The product ion scan time was 0.1 s, and the scan masses for MET, ASP, and APZ were m / z 130.2→60.1, m / z 370.2→242.0, and m / z 448.3→285.0, respectively. The optimized collision energies for MET, ASP, and APZ were 17 V, 45 V, and 37 V, respectively. The optimized declustering voltages for MET, ASP, and APZ were 20 V, 135 V, and 138 V, respectively. Reserpine (RES) was scanned at m / z 609.6 → 195.2, optimized collision energy 45 V, optimized declustering voltage 65 V; argon was used as the collision gas for MS / MS analysis; other general mass spectrometry parameters are as follows: spray voltage 5.5 kV; curtain gas 40 psi; nebulizer temperature 500°C; collision gas 9 psi; nebulizer gas 50 psi; auxiliary gas 50 psi. Nebulizer gas and auxiliary gas are GS1 and GS2 in the AB4500 mass spectrometer parameters, standing for nebulizer gas (GS1) and heater gas (GS2). GS1: Nebulizer gas, applied to the needle to vaporize liquid entering the needle; GS2: Auxiliary heater gas, applied to the two heater heads to evaporate liquid exiting the needle.

[0043] Figure 1 Shown is the mass spectrometry spectrum of aripiprazole; Figure 2 Shown is the mass spectrometry spectrum of amisulpride; Figure 3 Shown is the mass spectrometry spectrum of metformin; Figure 4Shown is the mass spectrum of reserpine. Figures 1-4 It is the ion peak diagram produced by the drug standard entering the mass spectrometer in the mass spectrometer, without entering the liquid phase. It is used to determine the mass-to-charge ratio corresponding to the ion channel of each drug in the subsequent UPLC-MS / MS detection for detection.

[0044] Figure 5 The figure shows the peak diagram of the UPLC-MS / MS process for detecting ion channels of rat plasma samples containing metformin after being treated with the aforementioned pretreatment method; Figure 6 Shown is the peak diagram of the UPLC-MS / MS process for detecting ion channels of a rat plasma sample containing amisulpride after being treated with the aforementioned pretreatment method; Figure 7 Shown is the peak diagram of the UPLC-MS / MS process for detecting ion channels of rat plasma samples containing aripiprazole after being treated with the aforementioned pretreatment method; Figure 8 Shown is the peak diagram of the UPLC-MS / MS process for detecting ion channels in rat plasma samples containing reserpine after being treated with the aforementioned pretreatment method.

[0045] All LC-MS / MS data were collected using software (Analyst), and statistical analysis was performed using Microsoft Office Excel (2016, USA).

[0046] To validate a method for quantifying three small molecule drugs, ASP, APZ, and MET, and to determine the feasibility of a pretreatment method involving solid-phase dispersion extraction (SPE) using carboxylated TiO2-coated multi-walled carbon nanotubes, linearity, accuracy, limit of quantification, inter- and intra-assay precision, method recovery, and matrix effects were evaluated. The drug assay used an internal standard (IS: reserpine, 10 μg / mL).

[0047] (1) Standard curve

[0048] A weighted method was used to calculate the standard curve. A minimum of six concentrations were used to construct the standard curve. Acceptable ranges were defined as ±20% deviation for the lowest concentration and ±15% for the remaining concentrations. A correlation coefficient of r > 0.99 was considered good.

[0049] (2) Accuracy, inter-batch and intra-batch precision

[0050] QC samples were used for testing. Four concentrations of QC samples, high, medium, low, and the lower limit of quantification, were selected for simultaneous testing of the method's precision and accuracy. The concentration of the low-concentration QC sample was no more than three times the lower limit of quantification, the concentration of the high-concentration QC sample was approximately 75% of the upper limit of quantification, and the concentration of the medium-concentration QC sample was selected near the mean geometric concentration. The same procedures were used for the accompanying standard curve, with each sample measured once. When determining intra-batch precision, at least five samples were prepared and measured at each concentration. To achieve inter-batch precision, analytical results for no fewer than 45 samples were obtained in at least three analytical batches, performed on at least two days. Accuracy should be within 85%-115% (with a relative standard deviation (RE) not exceeding ±15%) and within 80%-120% (with a relative standard deviation (RE) not exceeding ±20%) near the lower limit of quantification (LOQ). Precision generally requires an RSD of no more than 15%, and at the lower limit of quantification (LOQ) no more than 20%.

[0051] (3) Lower limit of quantification

[0052] At least five independent quality control samples were prepared, the accuracy of which should be within the range of 80%-120% of the labeled concentration, and the relative standard deviation (RSD) should be less than 20%.

[0053] (4) Method recovery rate

[0054] Take high, medium, and low concentration drug samples, with at least five replicates for each. Substitute the measured values ​​into the accompanying standard curve, compare with the spiked amount, and calculate the recovery. Generally, the average value measured for each concentration should deviate from the actual spiked amount by less than 15%, and the limit of quantification should be less than 20%.

[0055] (5) Matrix effect

[0056] For each batch of matrix, the matrix factor for each analyte and internal standard should be calculated by calculating the ratio of the peak area in the presence of matrix (measured by extracting blank matrix followed by the addition of analyte and internal standard) to the corresponding peak area in the absence of matrix (pure solutions of analyte and internal standard). Furthermore, the matrix factor normalized to the internal standard should be calculated by dividing the matrix factor of the analyte by the matrix factor of the internal standard. The coefficient of variation of the matrix factor should not exceed 15%. The determination should be performed at both low and high concentrations.

[0057] Here are the results:

[0058] (1) Standard curve

[0059] Table 1 Quantitative calibration curves of ASP, APZ and MET in plasma

[0060]

[0061] (2) Accuracy, inter-batch and intra-batch precision

[0062] Table 2 Validation results of the accuracy, inter-batch and intra-batch precision of the quantitative method for MET detection in plasma

[0063]

[0064] Table 3 Validation results of the accuracy, inter-batch and intra-batch precision of the quantitative method for ASP detection in plasma

[0065]

[0066] Table 4 Validation results of the accuracy, inter-batch and intra-batch precision of the quantitative method for APZ detection in plasma

[0067]

[0068]

[0069] (3) Lower limit of quantification

[0070] Table 4 Methodological validation results of the lower limit of quantification method for the detection of MET, ASP and APZ in plasma

[0071]

[0072] (4) Method recovery rate

[0073] Table 5 Methodological validation results for quantitative recovery of MET, ASP and APZ in plasma

[0074]

[0075] (5) Matrix effect

[0076] Table 6 Validation results of quantitative matrix effect method for detecting MET, ASP and APZ in plasma

[0077]

[0078] Based on the results of the above methodological validation, it can be proved that the pretreatment method of solid phase dispersion extraction using carboxylated TiO2-wrapped multi-walled carbon nanotube materials is suitable for the UPLC-MS / MS quantitative detection of three small molecule drugs: metformin (MET), amisulpride (ASP), and aripiprazole (APZ).

[0079] However, comparing matrix effects, it is clear that among the three small molecule drugs treated with carboxylated TiO2-coated multi-walled carbon nanotubes, MET still has a relatively significant matrix effect, with a matrix factor of 27.32% ± 1.45% at the low quality control concentration and 37.69% ± 1.14% at the high quality control concentration. The matrix factors of the other two drugs (ASP and APZ) were both within 100% ± 15%, achieving a level of no significant matrix effect after treatment. Comparing accuracy, ASP maintained good accuracy at the lower limit of quantification (LOQ), with an RE of only -3.68%. While APZ's accuracy at the LOQ also met the sample testing requirements, the RE was relatively high at -13.76%.

[0080] In the process of experimental verification of the removal of interfering substances in the matrix by carboxylated TiO2-coated multi-walled carbon nanotube materials, drugs that are not suitable for this material were also found, such as curcumin. According to the established UPLC-MS / MS instrument detection method, the peak shape of this drug after protein precipitation treatment is thin and high, which can clearly capture the retention time of the drug to be tested, curcumin (such as Figure 9 However, after being treated with carboxylated TiO2-wrapped multi-walled carbon nanotubes, the peak shape of curcumin became significantly unstable, with the peak width becoming very wide and the peak height significantly reduced (as shown in Figure 2). Figure 10 The retention time of curcumin cannot be accurately determined for quantitative analysis. Solid phase dispersion extraction of carboxylated TiO2-coated multi-walled carbon nanotubes is also not suitable for UPLC-MS / MS detection of omeprazole. In the experiment, different concentrations of omeprazole (0.01, 0.05, 0.25, 1.25, 6.25, 31.25 ng / mL) were not linear (as shown in Figure 2). Figure 11 Sample testing is not possible.

[0081] The above description is merely a preferred embodiment of the present invention. The present invention is not limited to the above-described embodiments. As long as the technical effects of the present invention are achieved by the same means, they shall fall within the scope of protection of the present invention. Within the scope of protection of the present invention, various modifications and variations of the technical solutions and / or implementation methods may be made.

Claims

1. A pretreatment method for detecting drugs in plasma, characterized in that: The following steps are involved: The plasma sample after protein removal is mixed with the composite material for solid phase dispersion extraction to obtain a supernatant; The plasma sample includes a small molecule drug, wherein the small molecule drug is metformin, aripiprazole or amisulpride; The composite material is carboxylated TiO2@MWCNTs; TiO2@MWCNTs are multi-walled carbon nanotubes coated with TiO2.

2. The pre-treatment method according to claim 1, characterized in that: The small molecule drug is amisulpride.

3. The pre-treatment method according to claim 1, characterized in that: The preparation method of the composite material comprises the following steps: stirring tetrabutyl titanate and nitric acid in an aqueous solution system at 70°C-95°C for 6h-24h to obtain a colloidal solution, filtering and redissolving, then adding multi-walled carbon nanotubes, stirring at 70°C-95°C for 12h-24h, centrifuging to obtain powder, washing and drying, and then annealing at 500°C-600°C for 2h-8h to obtain the TiO2@MWCNTs; dispersing the TiO2@MWCNTs in water, adding chloroacetic acid to mix, stirring at 60°C-95°C for 1h-6h to carry out a carboxylation reaction, centrifuging to obtain the carboxylated TiO2@MWCNTs.

4. The pre-treatment method according to claim 3, characterized in that: The volume ratio of tetrabutyl titanate to nitric acid is (5-20) mL:1 mL.

5. The pre-treatment method according to claim 3, characterized in that: The concentration of the colloidal solution obtained by re-dissolving after filtration is 1 mg / mL-10 mg / mL, and the ratio of the colloidal solution obtained by re-dissolving after filtration to the multi-walled carbon nanotubes is 100 mL: (100-1000) mg.

6. The pre-treatment method according to claim 3, characterized in that: The ratio of the TiO2@MWCNTs to chloroacetic acid is 1 g: (2-30) mL.

7. The pre-treatment method according to claim 1, characterized in that: The solid phase dispersion extraction comprises the following steps: mixing the plasma sample after protein removal with the composite material, vortexing for 0.5 min-3 min, and centrifuging for 5 min-10 min at 4° C.-25° C. and 12000 rpm-15000 rpm.

8. The pre-treatment method according to claim 1, characterized in that: The method for preparing the protein-removed plasma sample comprises the following steps: mixing the plasma sample with acetonitrile in a volume ratio of 1:(3-5), vortexing for 0.5 min-3 min, and centrifuging for 5 min-10 min at 4°C-25°C and 12000 rpm-15000 rpm, and the obtained supernatant is the protein-removed plasma sample.

9. A method for detecting drugs in plasma, characterized in that: The method comprises the following steps: using the supernatant obtained by the pretreatment method according to any one of claims 1 to 8 as an injection sample, and detecting the injection sample by LC-MS / MS; the drug is metformin, aripiprazole or amisulpride.

10. The detection method according to claim 9, characterized in that: Chromatographic separation was performed using a C18 chromatographic column, ultrapure water containing 2 mM ammonium formate and 0.0012% formic acid was used as mobile phase A, and acetonitrile was used as mobile phase B.

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