Optimization of enzymatic digestion conditions of shrimp allergens in mass spectrometry by response surface methodology
10.19485/j.cnki.issn2096-5087.2021.08.003
- Author:
PAN Xiaodong
;
HUANG Baifen
;
CAI Zengxuan
;
WANG Mengli
;
HAN Jianlong
;
CHEN Qing
- Publication Type:Journal Article
- Keywords:
allergen protein response surface methodology enzymatic digestion mass spectrometry
- From:
Journal of Preventive Medicine
2021;33(8):768-771
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To optimize the enzymatic digestion conditions of trypsin, so as to improve the testing capacity of mass spectrometry for shrimp allergens.
Methods:The enzymatic digestion test was carried out by response surface methodology for optimizing pH, temperature and time. After enzymatic hydrolysis, the peptides were separated by chromatography and determined by high-resolution mass spectrometry with Q-orbitrap. The allergen protein was identified and quantified by UniProt database and MaxQuant software.
Results:Two allergen proteins, tropomyosin and arginine kinase, were isolated from shrimp, and their intensities ranged from 100.2×106 to 436.5×106. Response surface analysis showed that when the digestion time was 4.29 hours, the temperature was 44.15 ℃ and pH value was 6.55, the maximal intensity of the allergen proteins was 457.48×106. The experiment was validated with the digestion time of 4.2 h, pH value of 6.5, and temperature of 44 ℃, then resulted in the average intensity of 448.1×106. The deviation from the predicted value was 2.1%.
Conclusions:The conditions of enzymatic digestion can be optimized by response surface methodology. The enzyme may have the best performance with the pH value of 6.5, temperature of 44 ℃ and digestion time of 4.2 hours.
- Full text:虾过敏原蛋白质谱分析方法的酶解条件优化研究.pdf
