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Method for detecting release of anaphylactic medium in Chinese medicament injection

An injection and allergy technology, which is applied in the field of rapid detection of the release of allergic mediators caused by traditional Chinese medicine injections, and the detection of endogenous trace substances, can solve the problems of low specificity of histamine, cumbersome operation steps, and difficulty in accurate quantification.

Inactive Publication Date: 2010-06-02
ZHEJIANG UNIV
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Problems solved by technology

The disadvantage of ELISA for measuring histamine is that it has low specificity and is not easy to quantify accurately. It is not suitable for the determination of histamine in biological samples with low histamine content.
However, the disadvantage of fluorescence spectrophotometry is that the operation steps are cumbersome, and the half-life of histamine is extremely short. [6] , anaphylaxis is not detectable for a few minutes after onset

Method used

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  • Method for detecting release of anaphylactic medium in Chinese medicament injection
  • Method for detecting release of anaphylactic medium in Chinese medicament injection
  • Method for detecting release of anaphylactic medium in Chinese medicament injection

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] 1. Preparation of reaction solution

[0031] Prepared by mixing 1110 μL of absolute ethanol, 240 μL of triethylamine, 50 μL of 2 μg / mL 9-methylanthracene, and 3.6 mL of acetone;

[0032] 2. Preparation of derivatization solution

[0033] It is composed of derivatization reagent Ethylchloroformate (ethyl chloroformate) mixed with chloroform in a certain proportion, specifically ethyl chloroformate and chloroform in a volume ratio of 1:9;

[0034] 3. Derivatization preparation of the test solution

[0035] Add 1350 μL of reaction solution to 100 μL of plasma or cell culture supernatant, vortex on a vortex for 3 seconds, add 550 μL of derivatization solution, and continuously vortex for 10 seconds; add 3.5 mL of water and vortex for 5 seconds; centrifuge at 1500 rpm for 5 minutes , take the organic phase and put it in a clean bottle, add 10 mg of anhydrous sodium sulfate for dehydration, and place it in a GC automatic sampling bottle with a liner for analysis.

[0036] ...

Embodiment 2

[0044] 1. Preparation of reaction solution

[0045] It is prepared by mixing 1000 μL of absolute ethanol, 200 μL of triethylamine, 50 μL of 2 μg / mL 9-methylanthracene, and 2.5 mL of acetone; that is, absolute ethanol: triethylamine: 9-methylanthracene solution: acetone = 30:6: 1:80~20:4:1:50,

[0046] 2. Preparation of derivatization solution

[0047] It is composed of derivatization reagent Ethylchloroformate (ethyl chloroformate) which is miscible with chloroform in a certain proportion, specifically ethyl chloroformate and chloroform in a volume ratio of 2:8;

[0048] 3. Derivatization preparation of the test solution

[0049] Add 1350 μL of reaction solution to 100 μL of plasma or cell culture supernatant, vortex on a vortex for 3 seconds, add 550 μL of derivatization solution, and continuously vortex for 10 seconds; add 3.5 mL of water and vortex for 5 seconds; centrifuge at 1500 rpm for 5 minutes , take the organic phase and put it in a clean bottle, add 10 mg of anhy...

Embodiment 3

[0051] 1. Linear relationship inspection

[0052] The linear relationship is to investigate the degree to which the change of the concentration of the tested sample is directly proportional to the result of the sample, and it is often determined by drawing a standard curve. Accurately draw 0.01, 0.1, 0.25, 0.5, 1, 2.5ml of histamine reference substance stock solution (2μg / mL) respectively and put them in a 10ml measuring bottle. , the method under the three items is measured, take reference substance concentration as abscissa, reference substance and internal standard substance peak area ratio as ordinate, draw standard curve, carry out regression analysis, get regression equation Y=1.0726X+0.031, r=0.9993 , indicating that histamine has a good linear relationship in the range of 0.002-2 μg / ml. see Figure 4 .

[0053] 2. Repetitive experiments

[0054] Take the same solution of the test product, repeat the sample injection 5 times according to the method under item 2 and ...

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Abstract

The invention provides a method for detecting the release of an anaphylactic medium in a Chinese medicament injection, which is a method for measuring the trace secretion of the anaphylactic medium namely histamine, capable of causing human body anaphylactic reactions, in the Chinese medicament injection. The method comprises the following steps of: adopting an ethyl chloroformate derivatization method; performing qualitative and quantitative analysis on the histamine by using a GC-MS detection method; and performing determination by analyzing a plurality of important parameters during measuring. Test results show that a chromatogram map obtained by the method has a smooth baseline and a good peak form, and the method is simple, convenient, quick and sensitive, has good repeatability, and can effectively detect the endogenous released histamine caused by the Chinese medicament injection.

Description

technical field [0001] The invention belongs to the field of drug analysis and relates to the detection of endogenous trace substances, in particular to an analysis method for rapidly detecting the release of allergic mediators caused by traditional Chinese medicine injections. Background technique [0002] With the continuous improvement and research of modern Chinese medicine chemistry and Chinese medicine preparation technology, Chinese medicine injections have been widely used clinically in my country. Traditional Chinese medicine injections have quick onset and strong effects, especially in the treatment of critical diseases, infectious diseases, cardiovascular and cerebrovascular diseases, and malignant tumors. However, with the widespread clinical application of traditional Chinese medicine injections and most of them are not single chemical substances, it will inevitably bring some safety problems of traditional Chinese medicine preparations [1] . From the most com...

Claims

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Application Information

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IPC IPC(8): G01N30/02G01N30/72G01N1/28
Inventor 陈艳李连达周长新张永勇
Owner ZHEJIANG UNIV
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