Construction method of stable isotope labeling internal standard, quantitative analysis method of stable isotope labeling internal standard and working solution

By screening and validating the mixture of deuterium aberrants in the deuterium-hydrogen exchange reaction products, a stable isotope-labeled internal standard was constructed, solving the problems of complex synthesis and inaccurate quantification in existing technologies, and realizing efficient quantitative analysis in complex biological matrices.

CN122449039APending Publication Date: 2026-07-24HUNAN NORMAL UNIVERSITY
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Patent Information

Application Number
CN202610615362.4
Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Filing Date
2026-05-07
Publication Date
2026-07-24

AI Technical Summary

Technical Problem

In existing technologies, the synthesis routes of stable isotope-labeled internal standards are complex and costly. Furthermore, when performing quantitative analysis in complex biological matrices, interference from natural isotope peaks and chromatographic isotope effects can affect the accuracy and reproducibility of quantification.

Method used

By preparing the deuterium-hydrogen exchange reaction products of the target compound, a mixture of deuterium isomers satisfying Δm≥3 was screened as a candidate component for internal standard. The retention time difference Δt < 0.01 min between the internal standard and the target compound was verified by mass spectrometry and chromatography to ensure co-eluting of the internal standard and the analyte and the absence of chromatographic isotope effects, thus constructing a stable isotope-labeled internal standard.

Benefits of technology

It effectively avoids interference from natural isotope peaks and chromatographic isotope effects, improves the accuracy and robustness of quantitative correction, and is suitable for quantitative analysis of complex biological samples that lack commercial isotope internal standards or are cost-sensitive.

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Abstract

The invention discloses a stable isotope labeling internal standard construction method, a quantitative analysis method and a working solution, and the method comprises the following steps: obtaining a deuterium-hydrogen exchange reaction product of a target compound, then carrying out mass spectrometric detection, and screening deuterium isomers of which the mass difference between the deuterium isomers and the target compound meets the condition that delta m is greater than or equal to 3 from the deuterium-hydrogen exchange reaction product as internal standard candidate components; detecting the target compound and the internal standard candidate component through chromatography, and if the retention time difference between the target compound and the internal standard candidate component is delta tlt; if the time is 0.01 min, determining that the internal standard candidate component can be used as a stable isotope labeling internal standard; if the retention time difference delta t of the two compounds is larger than or equal to 0.01 min, the steps are executed again after the hydrogen-deuterium exchange reaction conditions are adjusted until the retention time difference delta tlt with the target compound is obtained; the stable isotope labeling interior label is obtained after 0.01 min. The stable isotope labeling internal standard constructed by the invention can avoid natural isotope peak interference, and the internal standard and an analyte chromatogram meet delta tlt; the co-outflow requirement of 0.01 min is met, and the effectiveness of quantitative correction can be improved.
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