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Method for extracting nucleoside in sample by taking titanium dioxide as solid phase

A titanium dioxide extraction technology, applied in chemical instruments and methods, preparation of test samples, organic chemistry, etc., can solve problems such as poor reproducibility, low sample recovery rate, and low efficiency of nucleoside removal

Inactive Publication Date: 2013-12-11
WUHAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this enzymatic hydrolysis method includes a series of multi-step processing processes (such as RNase enzymatic hydrolysis, phenol / chloroform solution extraction, isopropanol cryoprecipitation, cleaning and reconstitution, etc.), requiring nearly 12 hours of processing time
In addition, frequent sample transfer and solvent exchange lead to lower sample recovery and poor reproducibility
And, in practice, this method is not very efficient at removing nucleosides
In addition, for the analysis of 2’-O modified nucleosides, there is no means to effectively remove the normal nucleosides co-generated during the hydrolysis of RNA

Method used

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  • Method for extracting nucleoside in sample by taking titanium dioxide as solid phase
  • Method for extracting nucleoside in sample by taking titanium dioxide as solid phase
  • Method for extracting nucleoside in sample by taking titanium dioxide as solid phase

Examples

Experimental program
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Effect test

Embodiment 3

[0032] The chromatographic column used in Examples 3 and 4 was Hisep C18-T column (150 mm x 2.1 mm i.d., 5 μm, Weltech Co., Ltd.). The flow rate was 0.2 mL / min. The gradient used in Example 3 is: 5min 5%B, 20min 5-50%B, 5min 50%B, 10min 5%B; the gradient used in Example 4 is: 15min 5%B, 5min 5-15% B, 10min 5% B (A is 5mM ammonium formate solution, B is methanol).

[0033] The present invention will be further described below by way of examples.

Embodiment 1

[0034] Embodiment 1: the extraction of nucleoside in the standard sample

[0035] Prepare 4 kinds of nucleosides (adenosine, uridine, cytidine, guanosine) and 4 kinds of 2'-deoxynucleosides (2'-deoxyadenosine, 2'-deoxyuridine, 2'-deoxycytidine, 2 '-deoxyguanosine) mixed solution (1μg / mL), for TiO 2 Analysis after solid phase extraction. The result is as figure 2 shown. Compared with the direct injection analysis (0.2μg / mL), after TiO2 extraction, the signals of the four nucleosides disappeared completely, and only the signals of four 2’-deoxynucleosides remained, indicating that the prepared TiO 2 The powder has a good extraction effect on nucleosides.

Embodiment 2

[0036] Example 2: Different TiO 2 Materials for the extraction of nucleosides

[0037] Prepare a mixed solution of adenosine and 2'-deoxyadenosine (1 μg / mL), using commercially available TiO 2 Nanoparticles, TiO prepared by liquid phase deposition 2 Coated SiO 2 Materials and TiO in Example 1 2 Powder samples were analyzed after dispersive SPE. The result is as image 3 shown. Compared with direct injection analysis, after extraction, the signal of adenosine decreased to varying degrees for all experimental groups. Explain the TiO obtained by three different preparation methods 2 Materials have obvious extraction effect on nucleosides. Illustrates the present invention for various TiO 2 Universality of materials.

[0038] Embodiment 3: utilize TiO 2 The material adsorbs the normal nucleosides in the DNA hydrolyzate extracted from yeast cells to detect the 2'-deoxynucleosides

[0039] The yeast cell DNA hydrolyzate treated by different methods was dissolved in water...

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Abstract

The invention discloses a method for absorbing nucleoside based on a titanium dioxide material. By using the method, the aim of rapidly and efficiently absorbing a great number of normal nucleoside substances existing in a biological sample is achieved in a solid extraction mode through the affiliation effect of the titanium dioxide material to a cis-glycol structure. Compared with the traditional enzyme hydrolysis nucleoside removal method widely applied at present, the method has the advantages of low cost, high speed and high specificity.

Description

technical field [0001] The invention relates to an extraction method of normal nucleosides (adenosine, cytidine, uridine, uridine) using titanium dioxide as a solid phase, which belongs to the sample pretreatment technology. Background technique [0002] The modification of DNA / RNA is closely related to various physiological processes of the human body, and its research has always been the focus of related disciplines. In vivo, due to the high abundance of normal nucleosides, it is easy to interfere with the detection of 2'-deoxynucleosides and 2'-O modified nucleosides. Before the analysis of 2'-deoxynucleosides, the steps of pre-enzymatically digesting RNA and precipitating DNA are usually used to reduce the content of normal nucleosides in the final sample. This enzymatic hydrolysis method includes a series of multi-step processing processes (such as RNase enzymatic hydrolysis, phenol / chloroform solution extraction, isopropanol cryoprecipitation, cleaning and reconstitut...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/34B01D15/08C07H1/06C07H19/00
Inventor 冯钰锜王少亭
Owner WUHAN UNIV