A method for simultaneous detection of five anti-tuberculosis drugs in plasma by uplc-ms/ms method

An anti-tuberculosis and plasma technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems of not fully meeting the needs of clinical determination, large mechanism effects, long analysis time, etc., and achieve short processing time, small matrix effect, and analysis short time effect

Active Publication Date: 2022-04-19
MENGCHAO HEPATOBILIARY HOSPITAL OF FUJIAN MEDICAL UNIV
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0004] Currently, ultra-high performance liquid chromatography-tandem mass spectrometry is the main method for the detection of anti-tuberculosis drugs. Meet clinical assay needs

Method used

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  • A method for simultaneous detection of five anti-tuberculosis drugs in plasma by uplc-ms/ms method
  • A method for simultaneous detection of five anti-tuberculosis drugs in plasma by uplc-ms/ms method
  • A method for simultaneous detection of five anti-tuberculosis drugs in plasma by uplc-ms/ms method

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Different chromatographic columns, mobile phase ratios and gradient elution methods were used to analyze the detected standard substances.

[0050] (1) The chromatographic column Shim-pack XR-ODSⅢ (2.0mm i.d×50mm, 1.6μm); the pre-column Shim-pack GIST-HP(G) (2μm C18, 2.1×10mm) was used to separate and analyze the standard. The mobile phase AB adopts different proportions and gradient elution methods. The peaks of isoniazid and ethambutol in the analyte both elute at about 0.5 min, and the above two analytes are not retained on this chromatographic column. Wherein the chromatographic mobile phase is set as follows: A (water phase): 0.01% (w / w) formic acid aqueous solution, B (organic phase): acetonitrile solution with formic acid concentration of 0.01% (w / w).

[0051]

[0052] The chromatographic peaks of the five standard substances measured are as follows: figure 1 ,Depend on figure 1 It can be seen that ethambutol and isoniazid are almost not retained, and the pe...

Embodiment 2

[0071] Three plasma samples from patients infected with Mycobacterium tuberculosis were collected respectively, named sample 1, sample 2, and sample 3.

[0072] (1) a. Standard product handling:

[0073] Take 90 μL of blank plasma, add 10 μL of mixed standard solutions of different concentrations, vortex and mix, add 10 μL of mixed isotope internal standard working solution, then add 300 μL of acetonitrile, vortex for 1 min, and centrifuge at 12000 r / min for 5 min; absorb 300 μL of supernatant in another Add 300 μL of ultrapure water to a clean 1.5 mL centrifuge tube, vortex for 30 seconds, and filter through a 0.45 μm filter to obtain the test solution; take 2 μL of the supernatant for analysis, and record the chromatogram;

[0074] The quantitative serial concentrations of rifampicin are: 0.039, 0.078, 0.156, 0.313, 0.625, 1.25, 2.5, 5μg / mL,

[0075] The quantitative serial concentrations of rifabutin are: 0.016, 0.031, 0.063, 0.125, 0.25, 0.5, 1, 2μg / mL,

[0076] The quan...

Embodiment 3

[0086] (1) Matrix effect detection

[0087] Take 90 μL of blank plasma, add 10 μL of mixed standard solutions of different concentrations, vortex and mix, add 10 μL of mixed isotope internal standard working solution, then add 300 μL of acetonitrile, vortex for 1 min, and centrifuge at 12000 r / min for 5 min; absorb 300 μL of supernatant in another Add 300 μL of ultrapure water to a clean 1.5 mL centrifuge tube, vortex for 30 seconds, and filter through a 0.45 μm filter to obtain the test solution; prepare 6 parallel samples, and take 2 μL of the supernatant for analysis.

[0088] Use 40% acetonitrile aqueous solution to prepare low and high working solutions of two concentrations (i.e. the concentrations corresponding to the compounds in the quality control sample in the summary of the invention), and prepare 6 copies in parallel, and take 2 μL for analysis. The results are shown in Table 3.

[0089] Table 3: Matrix Effects of Protein Precipitation Experimental Methods

[009...

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Abstract

The invention discloses a method for detecting five anti-tuberculosis drugs (including rifampicin, rifabutin, pyrazinamide, ethambutol and isoniazid) in blood plasma by UPLC-MS / MS. First, accurately measure blank plasma, add a series of mixed standard standard working solutions, and add one-to-one corresponding isotope internal standard standard working solutions. After pretreatment by protein precipitation, use UPLC-MS / MS for analysis to obtain each sample. For the chromatogram, take the peak area ratio of the analyte and its corresponding internal standard as the abscissa, and use the concentration of the analyte as the ordinate to establish a standard curve; then accurately measure the plasma to be tested, and then add a one-to-one corresponding isotope internal standard standard The sample working solution was pretreated by the protein precipitation method, and analyzed by UPLC-MS / MS to obtain the chromatograms of each sample, and the concentration of the plasma sample was calculated using the standard curve. The method is simple and quick to operate, has high sensitivity, accuracy and precision, and has little matrix effect, and can meet the needs of drug concentration monitoring of five anti-tuberculosis drugs in clinical application.

Description

(1) Technical field [0001] The invention relates to a method for detecting five anti-tuberculosis drugs (rifampicin, rifabutin, pyrazinamide, ethambutol, and isoniazid) in blood plasma by ultra-high performance liquid chromatography-mass spectrometry, The invention can be used for monitoring the therapeutic drug concentration of these five drugs, and belongs to the technical field of pharmacokinetic analysis. (2) Background technology [0002] Rifampicin (rifampin RFP) and rifabutin (LM427, rifabutinRFB) both belong to the rifamycin fungicides, and inhibit the RNA polymerase of Mycobacterium tuberculosis so that the bacteria cannot complete transcription process and death; pyrazinamide (pyrazinamind PZA) is a derivative of nicotine amine, has antibacterial or bactericidal effect, and is a half-effect fungicide; The synthesis of sugar leads to the formation of Mycobacterium tuberculosis cell wall and inhibits the growth of bacteria; isoniazid (INH) causes the defect of Mycob...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N30/02G01N30/06
CPCG01N30/02G01N30/06
Inventor 吴灵洁刘景丰刘小龙郑玲叶珍洁储楠楠刘辉
Owner MENGCHAO HEPATOBILIARY HOSPITAL OF FUJIAN MEDICAL UNIV
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