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Method for detecting residual quantity of sodium cyanoborohydride in protein sample or product

A technology of sodium cyanoborohydride and a detection method, which is applied in the direction of color/spectral characteristic measurement, etc., can solve problems such as only qualitative analysis, and achieve the effects of low cost, low toxicity, and simple method

Active Publication Date: 2016-06-22
深圳未名新鹏生物医药有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the existing methods for the analysis of cyanide residues can only achieve qualitative analysis, or require highly toxic substances, such as hydrogen bromide, benzidine, etc., such as the 060 cyanide inspection method in the Pharmacopoeia

Method used

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  • Method for detecting residual quantity of sodium cyanoborohydride in protein sample or product
  • Method for detecting residual quantity of sodium cyanoborohydride in protein sample or product
  • Method for detecting residual quantity of sodium cyanoborohydride in protein sample or product

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] With reference to the above-mentioned main analysis steps, Example 1 is briefly described below.

[0055] Carry out this test (check its stability) by two days:

[0056] (1) Test on the first day:

[0057] Preparation of sodium cyanoborohydride standard gradient solution: Take the standard mother liquor of sodium cyanoborohydride and dilute it with purified water to 0.25mmol / L, 0.20mmol / L, 0.15mmol / L, 0.10mmol / L, 0.05mmol / L, 0.025 mmol / L solution.

[0058] Then, put the standard solutions of different concentrations in a boiling water bath for 15 minutes, and centrifuge to get the supernatant;

[0059] Finally, the staining solution was added respectively, kept in a water bath at 37° C. for 30 min in the dark, and measured at 595 nm.

[0060] (2) Test on the second day:

[0061] Group A: Same as the first day.

[0062] Group B: The method of preparing the solution is the same as the first day, but without boiling water bath treatment; then, add the staining solutio...

Embodiment 2

[0065] With reference to the above-mentioned main analysis steps, Example 2 is briefly described below.

[0066] 1. Prepare standard gradient solution of sodium cyanoborohydride: take the standard mother liquor of sodium cyanoborohydride and dilute it with purified water to 0.25mmol / L, 0.20mmol / L, 0.15mmol / L, 0.10mmol / L, 0.05mmol / L ,0.025mmol / L solution.

[0067] 2. Take a batch of PEG-rhG-CSF stock solution (pure stock solution, without sodium cyanoborohydride), perform pretreatment (to remove PEG-rhG-CSF protein), and take the supernatant after centrifugation;

[0068] Take three supernatants and add 1mmol / L standard sodium cyanoborohydride solution respectively to make the final concentrations respectively 0.05mmol / L, 0.10mmol / L, and 0.15mmol / L.

[0069] 3. Add the staining solution respectively, keep in a water bath at 37°C for 30 minutes in the dark, and measure at 595nm. The measurement results are shown in Table 1 below.

[0070] Table 1 embodiment 1 sample addition ...

Embodiment 3

[0073] With reference to the above-mentioned main analysis steps, Example 2 is briefly described below.

[0074] 1. Prepare standard gradient solution of sodium cyanoborohydride: take the standard mother liquor of sodium cyanoborohydride and dilute it with purified water to 0.25mmol / L, 0.20mmol / L, 0.15mmol / L, 0.10mmol / L, 0.05mmol / L ,0.025mmol / L solution.

[0075] 2. Take the rhG-CSF stock solution (pure stock solution, without sodium cyanoborohydride), add 20mmol / L sodium cyanoborohydride to obtain rhG-CSF sample;

[0076] Sample pretreatment (removal of rhG-CSF protein), supernatant after centrifugation;

[0077] Then the obtained sample is diluted 400 times (purified water dilution), that is, the sample contains 0.05mmol / L sodium cyanoborohydride;

[0078] Divide the obtained sample into three parts A, B, and C according to the volume ratio of 1:1:1. Add an equal volume of 0.05mmol / L standard sodium cyanoborohydride solution to A sample, and add 0.10mmol / L standard cyanobo...

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Abstract

The invention discloses a method for detecting a residual quantity of sodium cyanoborohydride in a protein sample or product. The method comprises the steps of subjecting the protein sample or product to pretreatment including protein removal, then, adding the pretreated protein sample or product into a mixed solution of a Coomassie brilliant blue G-250 solution and a polysorbate 20 solution so as to form a mixture system, determining the light absorbancy of the mixture system by adopting an ultraviolet-visible spectrophotometer, and finally, determining the content of sodium cyanoborohydride in the protein sample or product by using an external standard method, wherein an external standard substance of the external standard method is sodium cyanoborohydride. The method has the advantages of simple operating steps, quickness, low cost and low toxicity.

Description

technical field [0001] The invention relates to the field of chemical substance detection, in particular to a method for detecting residual sodium cyanoborohydride. Background technique [0002] Sodium cyanoborohydride is a commonly used complex type hydride, which has excellent reducibility, but also has certain toxicity. In the pharmaceutical industry, the safety problems caused by sodium cyanoborohydride residues have long attracted people's attention, and three methods for the detection of cyanide residues have appeared in the Pharmacopoeia. However, the existing methods for the analysis of cyanide residues can only achieve qualitative analysis, or require highly toxic substances, such as hydrogen bromide, benzidine, etc., such as the 060 cyanide inspection method in the Pharmacopoeia. Contents of the invention [0003] In order to solve the above-mentioned technical problems, the object of the present invention is to propose a method for detecting the residual amount...

Claims

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Application Information

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IPC IPC(8): G01N21/31
CPCG01N21/31
Inventor 张晖张静贺淳晓
Owner 深圳未名新鹏生物医药有限公司
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