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Ninhydrin detection method as improvement to free amino quantitative method in solid-phase polypeptide synthesis

A technology for solid-phase peptide synthesis and free amino group, which is applied in the field of quantitative detection of amino residues

Inactive Publication Date: 2013-02-13
TECON BIOLOGY CO LTD
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Problems solved by technology

[0002] In the process of peptide synthesis by solid-phase method, the number of free amino groups is a direct reflection of the degree of synthesis completion, but there is no accurate method to obtain the specific number of residual amino groups to verify the reliability and accuracy of the results ; Therefore, it is of great practical value to make standard samples and standard curves for quantitative analysis of free amino groups in unknown resin peptides

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  • Ninhydrin detection method as improvement to free amino quantitative method in solid-phase polypeptide synthesis
  • Ninhydrin detection method as improvement to free amino quantitative method in solid-phase polypeptide synthesis
  • Ninhydrin detection method as improvement to free amino quantitative method in solid-phase polypeptide synthesis

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Embodiment Construction

[0032] The present invention is further clarified in conjunction with embodiment.

[0033] The implementation steps of this method:

[0034] (1) Preparation of reaction reagents:

[0035] Buffer A: Mix 5ml pyridine + 1.185g phenol + 5ml ethanol;

[0036] Buffer B: 500mg ninhydrin + 10ml ethanol;

[0037] 25% piperidine solution: mix 25ml piperidine solution + 75ml nitrogen methyl pyrrolidone;

[0038] (2) Sample handling:

[0039] 1) Solid-phase synthesis process: Weigh 2g of Rink_Amide_MBHA resin, and calculate the required resin mole number A according to the formula for calculating the number of resin moles, select amino acids with Fmoc protecting groups, take 2A moles of each amino acid, and use 1A moles of Rink_Amide_MBHA Resin and 2A moles of amino acids with Fmoc protecting groups were reacted under the action of 8% carbodiimide nitrogen methyl pyrrolidone solution for 2 hours to obtain resin peptides, which were then treated with 25% piperidine solution for 30 minu...

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Abstract

The invention provides a ninhydrin detection method as improvement to a free amino quantitative method in solid-phase polypeptide synthesis. Synthesized Rink_Amide_MBHA (4-Methyl-benzhydrylamine) resin is used as a standard sample and is subjected to continuous weight increment weighing to obtain a group of standard samples; and light absorption values are read under OD570 through a ninhydrin reagent reaction to acquire a standard curve; and free amino quantitative detection analysis can be performed on unknown peptide resin according to the standard curve.

Description

technical field [0001] The invention relates to the quantitative detection of amino residues, especially suitable for the synthesis of new solid-phase polypeptides. The absorbance value is measured by ninhydrin reagent, and a standard curve is prepared to determine the residual amino content in unknown resin peptides. Background technique [0002] In the process of peptide synthesis by solid-phase method, the number of free amino groups is a direct reflection of the degree of synthesis completion, but there is no accurate method to obtain the specific number of residual amino groups to verify the reliability and accuracy of the results ; Therefore, it is of great practical value to make standard samples and standard curves for quantitative analysis of free amino groups in unknown resin peptides. [0003] The conventional ninhydrin detection method is used for the quantitative detection of -NH2 residues in solid-phase peptide synthesis. Generally, the absorbance value of the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/31G01N21/78G01N1/28
Inventor 赵毅郑先强谢仁军肖璐恒李延涛黄炯唐磊
Owner TECON BIOLOGY CO LTD
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