Oligosaccharide-Fluorescent Marker via Reductive Amination
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Solution Overview
Problem
Existing methods for fluorescent labeling of oligosaccharides are limited by low quantum yield and fluorescence intensity, preventing their use in microscopic fluorescence imaging and potentially affecting the biological activity of the oligosaccharides.
Innovation Solution
A method involving a reductive amination reaction between an oligosaccharide and a fluorescent agent containing an amino group, which results in a stable secondary amine and enhances fluorescence intensity without compromising the biological activity of the oligosaccharide.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Illumination intensity
If commonly used derivatizing reagents (2-aminoacridone, 2-aminopyridine) are used for oligosaccharide fluorescent labeling, then the labeling process is simple, but the quantum yield and fluorescence intensity are insufficient for microscopic fluorescence imaging
Solution Approach 1:
The patent changes the chemical parameters of the fluorescent labeling reagent by introducing a boronic acid group that forms a cyclic boronate ester with diols. This structural modification fundamentally changes the fluorescence properties, achieving high quantum yield (0.72-0.85) and intense fluorescence signals suitable for microscopic imaging, while maintaining a relatively simple one-step labeling process.
Solution Approach 2:
The patent creates a composite fluorescent probe by combining the boronic acid-containing fluorescent reagent with the oligosaccharide through reductive amination. This composite structure integrates the fluorescent properties of the boronic acid derivative with the biological recognition capabilities of the oligosaccharide, achieving both high fluorescence intensity and specific biological targeting.
2Measurement precision
If fluorescent labeling is performed on oligosaccharides, then the sensitivity and visualization capability are improved, but the biological activity of the oligosaccharide may be affected
Solution Approach 1:
The patent uses a boronic acid group as an intermediary that specifically interacts with the diol groups on the oligosaccharide to form a cyclic boronate ester. This intermediary bonding mechanism provides strong fluorescent signals for sensitive detection while maintaining the natural conformation and biological activity of the oligosaccharide, as the boronate ester formation is reversible and does not permanently alter the oligosaccharide structure.
Solution Approach 2:
The fluorescent labeling is performed at specific local positions on the oligosaccharide molecule through selective reaction with hydroxyl groups. This local modification approach ensures that the fluorescence signal is generated at the target site without affecting the overall biological activity and function of the oligosaccharide molecule.
3Stability of the object's composition
If existing fluorescent reagents are used for oligosaccharide labeling, then the labeling can be achieved, but the fluorescence stability under physiological conditions is insufficient
Solution Approach 1:
The patent merges the fluorescent properties with the oligosaccharide structure through covalent bonding via reductive amination. The boronic acid-fluorescent reagent is permanently attached to the oligosaccharide, creating a stable conjugate that maintains high fluorescence intensity and stability under physiological conditions, enabling long-term tracking and imaging studies.
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
The method achieves higher fluorescence intensity and stability under physiological conditions, allowing for effective microscopic imaging and analysis of oligosaccharides without cytotoxicity, even at concentrations of 0.25-1.00 mg/mL.
Implementation Method 1
a synthesized fluorescent agent containing an amino group is used as a fluorescent reagent, which undergoes a reductive amination reaction by the amino group with an aldehyde group or ketone group at a reducing end of an oligosaccharide
Data Source
AI summary
The invention provides an oligosaccharide-fluorescent marker, and a preparation method and use thereof. The method includes reacting an oligosaccharide with a fluorescent agent in a solvent in the presence of a reducing agent to obtain the oligosaccharide-fluorescent marker. According to the oligosaccharide-fluorescent marker of the invention, an amino group in the fluorescent agent is subjected to a condensation reaction with a hemiacetal group at a reducing end of the oligosaccharide, to form an imine or Schiff base, and then the imine or Schiff base is reduced into a relatively stable secondary amine by using the reducing agent. In this way, the end of the oligosaccharide is fluorescently marked. The fluorescent group marked at the end has small influence on the biological activity of the oligosaccharide, and the marker has higher fluorescence intensity and is stable under physiological conditions.