CEdG Quantification via Stable Isotope Dilution LC-MS/MS

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Solution Overview

Problem

Current methods for quantifying advanced glycation end-products (AGEs), particularly N2-(1-carboxyethyl)-2′-deoxyguanosine (CEdG), are unreliable due to lack of suitable isotopically enriched standards and methodological inconsistencies, hindering their use as biomarkers for diabetic complications and other metabolic disorders.

Innovation Solution

The use of liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) with stable isotope dilution and internal standards, such as 15N5-CEdG, for precise quantification of CEdG in biological samples, allowing for accurate diagnosis, monitoring, and treatment evaluation of metabolic disorders.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Measurement precision

If conventional quantification methods are used for CEdG, then the analysis can be performed, but the measurement precision and reliability are insufficient due to lack of isotopically enriched standards and methodological inconsistencies

Engineering Contradiction:

ImproveCEdG quantification accuracyVSAvoidbiomarker reliability

Core Design Contradiction:

Measurement precisionVSReliability

Solution Approach 1:

The patent applies parameter changes by introducing stable isotope labeling (13C and 15N) to create isotopically enriched internal standards. These labeled standards have different mass-to-charge ratios that allow them to be distinguished from endogenous CEdG by mass spectrometry, enabling accurate quantification despite the absence of natural isotopic enrichment in biological samples.

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The patent uses isotopically enriched synthetic CEdG analogs as intermediary substances. These analogs serve as internal standards that mimic the chemical behavior of endogenous CEdG while being distinguishable by mass spectrometry, thereby mediating the quantification process and improving both measurement precision and reliability.

Inventive Principle:

Principle #24Intermediary (Mediator)

2Adaptability or versatility

If multiple quantitative and semi-quantitative methods are used for AGE measurement, then more options are available, but the difficulty of detecting and measuring increases due to lack of standardization

Engineering Contradiction:

Improvemethod optionsVSAvoidmeasurement standardization

Core Design Contradiction:

Adaptability or versatilityVSDifficulty of detecting and measuring

Solution Approach 1:

The patent establishes a universal quantification approach based on stable isotope dilution mass spectrometry that can be applied to measure multiple AGEs in various biological matrices. The method provides standardized protocols for sample preparation, internal standard addition, and mass spectrometry analysis, enabling consistent measurement across different studies and laboratories.

Inventive Principle:

Principle #6Universality (Multi-functionality)

3Measurement precision

If HbA1c is used as a biomarker for diabetes, then diagnosis and treatment monitoring are possible, but the measurement precision is limited compared to direct CEdG quantification

Engineering Contradiction:

Improvebiomarker sensitivityVSAvoidassay complexity

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The patent replaces indirect protein-based biomarkers (HbA1c) with direct nucleic acid-based biomarkers (CEdG) measured by mass spectrometry. This substitution provides more precise measurement of glycation status at the DNA level, offering superior sensitivity for detecting diabetic complications while maintaining manageable assay complexity through standardized LC-MS/MS protocols.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

This approach provides a reliable and sensitive method for quantifying CEdG, enhancing diagnostic precision and treatment monitoring, enabling early identification of diabetic complications and predicting disease progression.

Implementation Method 1

liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS)

Methodology Applied

Scientific EffectElectrospray ionization: Ionisation

Implementation Method 2

liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) with stable isotope dilution and internal standards

Methodology Applied

Scientific EffectMass spectrometry:

Implementation Method 3

quantification is achieved by a stable isotope dilution method using an internal standard. When the AGE is CEdG, the internal standard is 15N5-CEdG

Methodology Applied

Scientific EffectStable isotope dilution:

Data Source

PatentUS11179361B2Methods of quantifying N<sup>2</sup>-(1-carboxyethyl)-2′-deoxy-guanosine (CEdG) and synthesis of oligonucleotides containing CEdG

Publication Date: 2021.11.23 CITY OF HOPE

AI summary

Methods of quantifying a N2-(1-carboxyethyl)-2′-deoxyguanosine (CEdG) levels in biological samples and comparing those levels to known normal levels can diagnose a number of metabolic disorders or complications associated therewith, including diabetes, its associated complications, and cancer. Methods can also determine whether therapies for disorders are effective by measuring CEdG levels before and after treatment. Measurement of CEdG levels is achieved by using liquid chromatography electrospray ionization tandem mass spectrometry.