Snake Venom Thrombin Peptide Species Identification via LC-MS
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Solution Overview
Problem
Current methods for identifying the species of snake venom thrombin drugs are inadequate, as they cannot determine the source of the biochemical drugs, leading to differences in mechanism of action and pharmacological effects, necessitating a more specific and reliable identification method for quality control and safety assurance.
Innovation Solution
A snake venom thrombin-like enzyme from Agkistrodon Halys Pallas, with a specific amino acid sequence (LDSPVSNSAHIAPLSLPSSAPSVGSVCR), is used to identify Hemocoagulase for Injection through a method involving alkylation reduction, enzymolysis, and liquid chromatography-mass spectrometry analysis, providing a rapid, specific, and convenient means to determine the species origin.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If biochemical reaction, chemical chromogenic reaction, liquid chromatography-peptide graph control method or gel electrophoresis method is used for identification, then the identification process is established, but the source of species of the biochemical drugs cannot be identified
Solution Approach 1:
The patent segments the snake venom thrombin protein into specific peptide fragments through enzymatic digestion. By focusing on characteristic peptide sequences (such as the KQKQK motif) that are unique to specific snake species, the method divides the complex identification problem into manageable peptide-level analyses, enabling species-specific identification that whole-protein methods cannot achieve
Solution Approach 2:
The patent introduces mass spectrometry as an intermediary technique between traditional biochemical methods and species identification. The mass spectrometry system acts as a mediator that translates peptide mass patterns into species-specific information, bridging the gap between general protein detection and precise species determination by analyzing the unique mass-to-charge ratios of species-characteristic peptides
2Reliability
If traditional identification methods are used, then the quality control process is maintained, but the specificity for species identification is insufficient
Solution Approach 1:
The patent changes the analytical parameters from general protein detection (used in traditional methods) to specific peptide mass parameters. By measuring the mass-to-charge ratios of digested peptides and comparing them against species-specific reference patterns, the method transforms the identification process into a precise parameter-based measurement that can reliably distinguish between different snake species while maintaining quality control standards
3Manufacturing precision
If a new species-specific identification method is established, then the quality control level is improved, but the method complexity increases
Solution Approach 1:
The patent applies preliminary enzymatic digestion to break down the snake venom thrombin protein into characteristic peptide fragments before analysis. This preliminary action simplifies the subsequent mass spectrometry analysis by reducing the molecular complexity and generating species-specific peptide patterns that are easier to detect and identify, thereby improving quality control precision without requiring overly complex analytical equipment
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 significantly enhances the quality control of snake venom thrombin drugs, ensuring safety and effectiveness by specifically identifying the species of Hemocoagulase for Injection, filling the gap in existing quality standards and providing a reliable means to differentiate between snake venom thrombin sources.
Implementation Method 1
conducting alkylation reduction treatment on the test solution and the reference solution with dithiothreitol and iodoacetamide
Implementation Method 2
adding enzyme liquid for hydrolysis
Implementation Method 3
injecting a supernatant into a liquid chromatography-mass spectrometer for analysis
Data Source
AI summary
The present invention provides a snake venom thrombin marker peptide of Agkistrodon Halys Pallas and an application of the snake venom thrombin-like enzyme in identifying species of Hemocoagulase for Injection. The application includes the following steps of: dissolving a to-be-detected sample and a reference substance of the marker peptide respectively to prepare a test solution and a reference solution, and conducting alkylation reduction on the test solution and the reference solution with dithiothreitol and iodoacetamide; after diluting products with an ammonium bicarbonate solution, adding enzyme for hydrolysis; and after enzymolysis is finished, conducting centrifugation at a high speed, and injecting a supernatan into a liquid chromatography-mass spectrometer for analysis. This method is simple, convenient and rapid, is strong in specificity, fills the gap in identifying the source of species of the snake venom thrombin-like enzyme of Agkistrodon Halys Pallas and improves the quality control level.