阅读说明：本技术 一种用于同分异构体混合物组成的核磁共振检测方法 (Nuclear magnetic resonance detection method for isomer mixture composition ) 是由 乔岩 赵奇 马慧 王英雄 侯相林 于 2019-10-30 设计创作，主要内容包括：本发明涉及一种用于同分异构体混合物组成的核磁共振检测方法,所述核磁共振检测方法是根据同分异构混合物中各组分信号在一维纯化学位移氢谱中的化学位移值分布不同,从而快速对各组分信号进行分析指认；当所述一维纯化学位移谱对同分异构体混合物的信号指认分析后,通过借助化学选择性滤波激发脉冲提取出与激发质子同属某单一组分的分子信号,对一维纯化学位移谱中的重叠信号做进一步指认分析,从而实现对同分异构体混合物各组分的有效分析。本发明无需对样品进行预处理,能够得到混合物组分的无重叠且去耦氢谱和单一组分的选择性滤波实验激发谱,简化谱图分析；能够定性检测出复杂混合物中的物质组成,并适用于多种复杂混合物的成分的检测。(The invention relates to a nuclear magnetic resonance detection method for composition of an isomer mixture, which is characterized in that the nuclear magnetic resonance detection method is used for rapidly analyzing and identifying each component signal according to different chemical shift value distributions of each component signal in the isomer mixture in a one-dimensional pure chemical shift hydrogen spectrum; after the signal of the one-dimensional pure chemical shift spectrum to the isomer mixture is subjected to identification analysis, a molecular signal which belongs to a single component with an excited proton is extracted by means of chemical selective filtering excitation pulse, and overlapped signals in the one-dimensional pure chemical shift spectrum are subjected to further identification analysis, so that effective analysis of each component of the isomer mixture is realized. According to the invention, the sample is not required to be pretreated, the non-overlapping and decoupling hydrogen spectrum of the mixture components and the selective filtering experiment excitation spectrum of the single component can be obtained, and the spectrogram analysis is simplified; can qualitatively detect the substance composition in the complex mixture and is suitable for detecting the components of various complex mixtures.)

1. A nuclear magnetic resonance detection method for the composition of an isomer mixture is characterized in that the nuclear magnetic resonance detection method is used for rapidly analyzing and identifying each component signal according to different chemical shift value distributions of each component signal in the isomer mixture in a one-dimensional pure chemical shift hydrogen spectrum; after the signal of the one-dimensional pure chemical shift spectrum to the isomer mixture is subjected to identification analysis, a molecular signal which belongs to a single component with an excited proton is extracted by means of chemical selective filtering excitation pulse, and overlapped signals in the one-dimensional pure chemical shift spectrum are subjected to further identification analysis, so that effective analysis of each component of the isomer mixture is realized.

2. The method as claimed in claim 1, wherein the one-dimensional pure chemical shift spectrum of the isomeric mixture is obtained by sampling data using pulse sequence of psyche.

3. The nmr method according to any one of claims 1 or 2, wherein the nmr method comprises the following steps:

firstly, taking a proper amount of each component sample of an isomer, dissolving the component sample in a deuterated solvent, transferring the component sample into a nuclear magnetic resonance sample tube, and further placing the nuclear magnetic resonance sample tube containing the sample in a liquid nuclear magnetic resonance spectrometer;

setting the temperature of a nuclear magnetic resonance spectrometer, adjusting the airflow rate, and testing the sample after the sample is kept constant for a certain number of minutes under the condition;

thirdly, obtaining a conventional one-dimensional hydrogen spectrum of the isomer mixture according to the sample measurement result in the second step;

fourthly, changing a sampling pulse sequence into psyche.mf, setting a gradient field, adjusting sampling parameters, and acquiring pure chemical displacement hydrogen spectrum data of an isomer mixture to obtain pseudo two-dimensional experimental data;

fifthly, processing the pseudo two-dimensional experimental data obtained in the fourth step by using a macro command pshift to obtain a pure chemical displacement one-dimensional hydrogen spectrum of the isomer mixture, and analyzing and identifying attribution according to the chemical displacement value of each component signal;

and sixthly, changing the pulse sequence to selcsfdizs.2 according to the conventional one-dimensional hydrogen spectrum of the isomer mixture obtained in the third step, selecting the chemical shift value of the obvious characteristic peak of each component as an excitation frequency, carrying out a chemical selective filtering experiment, and further performing identification analysis on an overlapped signal in the one-dimensional pure chemical shift spectrum of the isomer mixture, thereby realizing the effective analysis on each component of the isomer mixture.

4. The nuclear magnetic resonance detection method for the composition of the isomer mixture according to claim 3, wherein the nuclear magnetic resonance spectrometer used in the nuclear magnetic resonance detection method is a liquid nuclear magnetic resonance spectrometer with a gradient field of 300-950 MHz.

5. The method of claim 3, wherein the deuterated reagent used in the method is deuterium oxide, deuterated methanol, deuterated chloroform, deuterated acetone, deuterated dimethyl sulfoxide or deuterated-N, N-dimethylformamide; the dosage of the deuterated reagent is 400-800 mu l.

6. The method according to claim 3, wherein the temperature of the NMR spectrometer in the second step is 273K-323K, the airflow rate is 100-900 lph, and the sample holding time is 15-50 minutes.

7. The method of claim 3, wherein the third step comprises measuring the hydrogen spectrum using a pulse sequence selected from the group consisting of zg30 and zgpr.

8. The method of claim 3, wherein the gradient field used in the fourth step is a CHIRP-shaped pulse.

9. The NMR detection method of claim 3, wherein the sampling parameters adjusted in the fourth step are respectively as follows: TD (F2) is 8-32 k, TD (F1) is 20, SWH (F2) is 10000Hz, SWH (F1) is 50-100 Hz, CNST20 is 10-25 degrees, and the sampling times NS are positive integer multiples of 2.

10. The NMR detection method of an isomer mixture composition according to claim 3, wherein the chemoselective filter experiment in the sixth step sets the value of the sampling point TD to 8k to 64k, the sampling frequency NS to be a positive integer multiple of 2, the mixing time to be 0.02 to 0.09s, and the suppressed adjacent signal range CNST20 to be 2 to 10 Hz.

Technical Field

The invention relates to the technical field of nuclear magnetic resonance detection, in particular to a nuclear magnetic resonance detection method for isomer mixture composition.

Background

With the rapid development of the current research fields of chemical industry, biological medicine, agriculture and the like, more and more natural products, including the extraction and separation of natural active molecules with high added values and the separation of effective components in the synthesis process of fine chemicals, particularly in the synthesis process of drug intermediates, need to establish a rapid and efficient modern separation technical method, and the nuclear magnetic resonance method provides a new way for strengthening a drug development support system in the fields. Because isomers have similarity in physical and chemical properties, and have the same relative molecular weight, small polarity difference, similar boiling points and the same characteristic ions of mass spectrum, gas chromatography and liquid chromatography are generally difficult to effectively separate; for a conventional nuclear magnetic resonance method such as a one-dimensional hydrogen spectrum, spectral peak signal overlapping can not be identified, and for a two-dimensional nuclear magnetic resonance method such as a diffusion sequencing spectrum, the test setting is complicated, the time is long, and rapid and effective detection can not be carried out on isomers with the same molecular weight and functional groups, so that the research of a separation method for rapidly and accurately identifying the isomers is very important.

Disclosure of Invention

To solve the above technical problems. The invention aims to provide a nuclear magnetic resonance detection method for the composition of an isomer mixture, so that the composition of the isomer mixture can be rapidly, efficiently and sensitively detected.

In order to achieve the purpose, the invention adopts the technical scheme that:

a nuclear magnetic resonance detection method for the composition of an isomer mixture is characterized in that the nuclear magnetic resonance detection method is used for rapidly analyzing and identifying each component signal according to the distribution of each component signal in the isomer mixture in a one-dimensional pure chemical displacement hydrogen spectrum and the difference of the resonance frequency of hydrogen nuclei; after the signal of the one-dimensional pure chemical shift spectrum to the isomer mixture is subjected to identification analysis, a molecular signal which belongs to a single component with an excited proton is extracted by means of chemical selective filtering excitation pulse, and overlapped signals in the one-dimensional pure chemical shift spectrum are subjected to further identification analysis, so that effective analysis of each component of the isomer mixture is realized. The chemical shift values of all protons of the mixture are similar due to the fact that all components of the isomeric mixture have the same molecular weight and functional groups, and the further coupling and splitting among all protons can cause that all component signals are seriously overlapped on a conventional one-dimensional hydrogen spectrum and can not be subjected to attribution identification. The chemical selectivity filtering experiment (CSSF-TOCSY) can obtain the proton information directly or indirectly coupled with the excited proton by exciting the proton with specific resonance frequency of each component one by one, so the technology can extract the hydrogen spectrum corresponding to the single component from the hydrogen spectrum of the complex mixture and further identify the attribution of the signal. The invention can realize the fast and high-efficiency analysis and identification of the composition of the isomer mixture by combining the one-dimensional pure chemical shift spectrum and the chemical selectivity filtering experiment.

Furthermore, the one-dimensional pure chemical shift spectrum of the isomer mixture is obtained by sampling data by using a pulse sequence psyche. Mf pulse sequence is used for data sampling to obtain pseudo two-dimensional experimental data, a macro command pshift is needed to be used for converting the data to obtain a one-dimensional pure chemical displacement spectrum without coupling splitting, and different proton signals have different chemical displacement values due to different proton resonance frequencies of all components, so that spectrogram signals are simplified, and attribution identification is facilitated.

Still further, the specific test steps of the nuclear magnetic resonance detection method are as follows:

firstly, taking a proper amount of each component sample of an isomer, dissolving the component sample in a deuterated solvent, transferring the component sample into a nuclear magnetic resonance sample tube, and further placing the nuclear magnetic resonance sample tube containing the sample in a liquid nuclear magnetic resonance spectrometer;

setting the temperature of a nuclear magnetic resonance spectrometer, adjusting the airflow rate, and testing the sample after the sample is kept constant for a certain number of minutes under the condition;

thirdly, obtaining a conventional one-dimensional hydrogen spectrum of the isomer mixture according to the sample measurement result in the second step;

fourthly, changing a sampling pulse sequence into psyche.mf, setting a gradient field, adjusting sampling parameters, and acquiring pure chemical displacement hydrogen spectrum data of an isomer mixture to obtain pseudo two-dimensional experimental data;

fifthly, processing the pseudo two-dimensional experimental data obtained in the fourth step by using a macro command pshift to obtain a pure chemical displacement one-dimensional hydrogen spectrum of the isomer mixture, and analyzing and identifying attribution according to the chemical displacement value of each component signal;

and sixthly, changing the pulse sequence to selcsfdizs.2 according to the conventional one-dimensional hydrogen spectrum of the isomer mixture obtained in the third step, selecting the chemical shift value of the obvious characteristic peak of each component as an excitation frequency, carrying out a chemical selective filtering experiment, and further performing identification analysis on an overlapped signal in the one-dimensional pure chemical shift spectrum of the isomer mixture, thereby realizing the effective analysis on each component of the isomer mixture. The constant temperature and airflow during the testing process can improve the stability and high repeatability of the experiment.

Furthermore, the deuterated reagent used in the nuclear magnetic resonance detection method is deuterium oxide, deuterated methanol, deuterated chloroform, deuterated acetone, deuterated dimethyl sulfoxide or deuterated-N, N-dimethylformamide; the dosage of the deuterated reagent is 400-800 mu l. And a proper deuterated reagent is selected as a solvent, so that the isomer can be fully dispersed in the solvent, and the spectrometer test is facilitated.

Furthermore, the nuclear magnetic resonance spectrometer selected in the nuclear magnetic resonance detection method is a liquid nuclear magnetic resonance spectrometer with a gradient field of 300-950 MHz. Spectrometers of different field strengths will have different finesse differences, and hydrogen spectra produced by spectrometers of high field strengths will have higher resolution and sensitivity.

Further, the third step is to test the hydrogen spectrum with the selected pulse sequence zg30 or zgpr. The data from small angle excitation and pressurized water pulse testing will yield a spectrum with better results.

Further, the gradient field used in the fourth step is a CHIRP shaped pulse. Selecting this shape of the pulse will have a better excitation effect.

Further, the sampling parameters adjusted in the fourth step are respectively: TD (F2) is 8-32 k, TD (F1) is 20, SWH (F2) is 10000Hz, SWH (F1) is 50-100 Hz, CNST20 is 10-25 degrees, and the sampling times NS are positive integer multiples of 2. Setting parameters TD (F2) to be 8-32 k and TD (F1) to be 20, so that the test time can be reduced under the condition of ensuring the common quality; meanwhile, parameters SWH (F2) are set to be 10000Hz, SWH (F1) is set to be 50-100 Hz, and the sampling times NS are set to be 2N (N is a positive integer), so that sampling can be smoothly carried out to obtain a clean spectrogram.

Furthermore, in the chemoselective filtering experiment in the sixth step, the value of the sampling point number TD is 8k to 64k, the sampling frequency NS is an integral multiple of 2, the mixing time is 0.02 to 0.09s, and the range of the pressed adjacent signal CNST20 is 2 to 10 Hz. And the proper mixing time and the accurate pressing range are adjusted, so that all signals can be excited to obtain a selective filter spectrogram.

Compared with the prior art, the invention has the following beneficial effects:

the invention relates to a nuclear magnetic resonance method for analyzing and detecting the composition of an isomer mixture, which analyzes and identifies each component of the isomer mixture by combining a one-dimensional pure chemical shift spectrum and a chemoselectivity filtering experiment. Compared with the chromatographic analysis technology, the nuclear magnetic resonance method does not need to preprocess the sample, does not damage the sample composition, and needs a small amount of sample, and can obtain the non-overlapping and decoupling hydrogen spectrum of the mixture components and the selective filtering experiment excitation spectrum of the single component, thereby greatly simplifying the spectrogram analysis.

The invention removes proton coupling information based on the characteristics of a one-dimensional pure chemical shift simplified spectrogram, only provides single chemical shift information, effectively reduces the spectrogram crowding degree and improves the spectrogram resolution. The invention also discloses a method for accurately positioning a target object based on the CSSF-TOCSY which can extract pure compound signals from a complex mixture one by one. The method has the advantages of visual and clear result, high detection resolution, simple operation, accurate result and high accuracy, can qualitatively detect the substance composition in the complex mixture, and is suitable for detecting the components of various complex mixtures.

Drawings

FIG. 1 is a pure chemical shift PSYCHE spectrum and a conventional hydrogen spectrum of example 1 of the present invention;

FIG. 2 is a spectrum of a chemical selective filter experiment (CSSF-TOCSY) in example 1 of the present invention;

FIG. 3 is a comparison of pure chemical shift PSYCHE spectra of example 1 of the present invention with conventional hydrogen spectra and with chemical selectivity filter experiment (CSSF-TOCSY) spectra;

FIG. 4 is a pure chemical shift PSYCHE spectrum and a conventional hydrogen spectrum of example 2 of the present invention;

FIG. 5 is a spectrum of a chemical selective filter experiment (CSSF-TOCSY) in example 2 of the present invention;

FIG. 6 is a pure chemical shift PSYCHE spectrum and a conventional hydrogen spectrum of example 3 of the present invention;

FIG. 7 is a spectrum of a chemical selective filter experiment (CSSF-TOCSY) in example 3 of the present invention;

FIG. 8 is a pure chemical shift PSYCHE spectrum and a conventional hydrogen spectrum of example 4 of the present invention;

FIG. 9 is a spectrum of a chemical selective filter experiment (CSSF-TOCSY) in example 4 of the present invention.

Detailed Description

The technical solution of the present invention is further described below with reference to the accompanying drawings and specific embodiments. It should be understood by those skilled in the art that the specific embodiments are only for the understanding of the present invention and should not be construed as the specific limitations of the present invention.