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Application of 3-hydrazinobenzoic acid derivatized dextran in mass calibration of maldi-tof-ms

A MALDI-TOF-MS, hydrazine benzoic acid technology, applied in the field of mass spectrometry detection, can solve the problems of high separation and purification cost, weakened peak intensity, complicated operation, etc., and achieves simplified operation steps, simple operation and high derivatization efficiency. Effect

Active Publication Date: 2020-06-05
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the substances commonly used as calibrator include protein, peptide mixture, etc., but the cost of separation and purification of such calibrator is high, sensitive to the environment, and has high requirements for storage conditions; metal clusters, such as [Cs n I n-1 ] + and [Cs n-1 I n ] - Clusters can be used for MALDI-MS positive and negative ion mode calibration, but when using this type of calibrator, as the molecular weight increases, the peak intensity is obviously weakened; polymers, such as polyethylene glycol, polyethylene glycol, etc., T.Gruending Use polyalanine to calibrate the mass of MALDI mass spectrometry, and it can be calibrated in the range of m / z 1000-3500, but the calibration of positive and negative ions needs to use DHB and CHCA as matrix respectively, and the operation is cumbersome

Method used

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  • Application of 3-hydrazinobenzoic acid derivatized dextran in mass calibration of maldi-tof-ms
  • Application of 3-hydrazinobenzoic acid derivatized dextran in mass calibration of maldi-tof-ms
  • Application of 3-hydrazinobenzoic acid derivatized dextran in mass calibration of maldi-tof-ms

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0065] Example 1: Optimization of derivatization conditions for reducing sugar target plate based on 3-HBA reactive matrix .

[0066] Prepare 20pmol / μL of maltohexaose (G6) as a model compound, and investigate the temperature, time, acetic acid ratio and DHB concentration ratio of the reaction. The specific steps are as follows:

[0067] (1) Optimization of temperature conditions: a saturated solution of 3-HBA was prepared with a mixed solvent of MeOH:DMSO with a volume ratio of 9:1, and 10% acetic acid was added as a catalyst. Take 1 μL of LG6 and 1 μL of matrix solution and place them on the same well of the target plate in turn, and mix them evenly. After reacting at 30°C, 45°C, 65°C and 85°C for 30 minutes, the derivatization efficiency was calculated by the peak intensity. Such as figure 2 As shown in a, the derivatization efficiency increases with increasing temperature;

[0068] (2) Reaction time optimization: under the conditions of 65°C and 10% acetic acid, take...

Embodiment 2

[0072] Example 2: Optimization of D2000 concentration.

[0073] D2000 aqueous solutions with concentrations of 0.05mg / mL, 0.5mg / mL and 5mg / mL were prepared respectively, and a combination matrix of 0.4M DHB combined with saturated 3-HBA was used. Take 1 μL of D2000 solution and 1 μL of LDHB / 3-HBA mixed matrix solution and spot on the same spotting hole of the target plate in turn, mix well, and react at 65°C for 30 minutes. Such as image 3 As shown, when the D2000 concentration is 0.05mg / mL, the derivatization effect is the best, and the mass distribution range is wide.

Embodiment 3

[0074] Example 3: 3-HBA derivatized D2000 for MALDI-TOFMS calibration

[0075] (1) Prepare the DHB / 3-HBA mixed matrix solution and 0.05 mg / mL D2000 aqueous solution as in Example 2, take 1 μ LD2000 solution and 1 μ LDHB / 3-HBA mixed matrix solution and place them on the same spotting hole of the target plate in turn, and mix Evenly, react at 65°C for 30min. Put the target plate into the mass spectrometer, collect the first-order mass spectrum in positive and negative ion modes and calibrate the mass spectrometer. Figure 4 a is the MALDI mass spectrum of D2000 in positive and negative ion mode, the signal peaks are normally distributed, the mass distribution is uniform, and the accuracy is high. The mass-to-charge ratio difference between adjacent signal peaks is m / z 162, which corresponds to exactly one glucose residue. The primary mass spectrometry calibration data are listed in Table 1.

[0076] Table 1: Primary calibration data for 3-HBA derivatized D2000

[0077]

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Abstract

The invention belongs to the field of mass spectrometry detection, and specifically discloses application of reactive matrix3-hydrazino-benzoic acid derivatized glucan to MALDI-TOF-MS mass calibration. 3-hydrazine-benzoic acid reacts with hemiacetal at the tail end of reducing sugar in a hydrazone mode, reaction is high in derivatized efficiency, a target board is used for online derivation, and operation is simple and convenient; the derivative strategy is used, and derivatized products can be detected in a positive ion mode and a negative ion mode; and carboxylate radical in the 3-hydrazine-benzoic acid significantly improves the ionization efficiency of the reducing sugar in the negative ion mode. According to the application, the 3-hydrazino-benzoic acid derivatized glucan is used as amass calibration object of MALDI mass spectrometry, mass difference between adjacent signal peaks is 162Da, and the accurate calibration is realized simultaneously in the positive ion mode and the negative ion mode corresponding to a glucose residue.

Description

technical field [0001] The invention relates to the technical field of mass spectrometry detection, in particular to the mass calibration of 3-hydrazinobenzoic acid (3-HBA) as a reactive matrix derivatized dextran in MALDI-TOF-MS (matrix-assisted laser desorption ionization time-of-flight mass spectrometry) application. Background technique [0002] Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) is widely used in the mass determination of macromolecules, such as proteins, polysaccharides and polymers. Accurate mass-to-charge ratio calibration is the first prerequisite for mass spectrometry to analyze substances, so it is very important to choose an appropriate mass calibrator. [0003] The compound used as a mass calibrator should have: (1) accurate spectral characteristics and accurate molecular weight; (2) abundant spectral peaks within the calibration range and suitable difference between adjacent mass spectral peaks; (3) easy...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/64G01N33/68G01N1/28
CPCG01N1/28G01N27/64G01N33/6851
Inventor 潘远江赵晓勇黄钰
Owner ZHEJIANG UNIV
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