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Method for absolutely quantifying mass spectrum of histidine tag-containing recombinant human endostatin

A technology of histidine tag and endostatin, which is applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve the problems that there is no quantitative method of mass spectrometry

Inactive Publication Date: 2012-08-01
CHINA PHARM UNIV
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Problems solved by technology

Existing quantitative methods for recombinant human endostatin mainly include radioisotope labeling (Current Drug Metabolism, 2006, 7, 565-576) and enzyme-linked immunosorbent assay (J.Clin.Oncol.21(2003) 223-231) , there is no mass spectrometric method for quantification

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  • Method for absolutely quantifying mass spectrum of histidine tag-containing recombinant human endostatin
  • Method for absolutely quantifying mass spectrum of histidine tag-containing recombinant human endostatin
  • Method for absolutely quantifying mass spectrum of histidine tag-containing recombinant human endostatin

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Embodiment 1

[0028] Example 1: Determination of pharmacokinetics of recombinant human endostatin containing histidine tag in rats

[0029] The main steps are as follows:

[0030] Eighteen SD rats were randomly divided into three groups (half male and half male, body weight 200±20 g), 6 rats in each group. Tail vein injection of recombinant human endostatin injection 1.5, 4.5, or 13.5mg / kg, after administration, 15 and 30 minutes, 1, 2, 3, 4, 6, 8, 12, 24 hours blood collection about 300μL , placed in an anticoagulant test tube, centrifuged at 3000 rpm at 4°C, and then transferred to another sterile dry tube, and stored in a freezer at -80°C. After the plasma sample was extracted and prepared according to the above biological sample preparation method, the blood drug concentration was determined by LC-MS / MS. Use the pharmacokinetic calculation software WinNonlin to calculate the kinetic behavior of the drug in the blood, and the half-life of the distribution phase t 1 / 2α , Elimination ph...

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Abstract

The invention belongs to the technical field of analysis, and relates to a method for absolutely quantifying mass spectrum of histidine tag-containing recombinant human endostatin. The method is based on LC-MS / MS, and extracts histidine tag-containing recombinant human endostatin from a plasma sample through utilizing Ni<2+> sepharose gel resin, and realizes absolute quantification of histidine tag-containing recombinant human endostatin in the plasma sample through utilizing a quantifying strategy for a specific peptide generated during hydrolysis of recombinant human endostatin with trypsin and taking a synthetic peptide having a sequence similar to that of the specific peptide as an interior label. The invention mainly includes a biological sample preparation method, an LC-MS / MS measuring method, standard curve preparation and precision degree measurement. The method has good linear relation, high accuracy degree and excellent reproducibility, and can be used for measuring the blood concentration of histidine tag-containing recombinant human endostatin and studying the pharmacokinetics.

Description

technical field [0001] The invention relates to a method for absolutely quantifying recombinant human endostatin with a histidine tag using mass spectrometry Background technique [0002] Endostatin has a wide range of anti-tumor angiogenesis effects. Its mechanism of action is to inhibit the formation of tumor angiogenesis by inhibiting the migration of endothelial cells that form blood vessels, and block the nutrient supply of tumors, thereby inhibiting tumor proliferation or metastasis. In 2005, SFDA approved recombinant human endostatin injection (Endostar), as the first new drug for lung cancer with endostatin, which contains a histidine tag. As a marketed drug, the biological sample quantitative method of recombinant human endostatin is of great significance for evaluating the kinetic behavior of the drug in vivo. Existing quantitative methods for recombinant human endostatin mainly include radioisotope labeling (Current Drug Metabolism, 2006, 7, 565-576) and enzyme-l...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/88
Inventor 王广基李盈淳郝海平梁艳汤志远肖亚楠
Owner CHINA PHARM UNIV
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