基于液质联用技术定量分析牛体外受精胚胎中谷胱甘肽含量

doi:10.11843/j.issn.0366-6964.2018.12.011

Abstract

Abstract:

This study aimed to present a method to clarify the effects of exogenous glutathione (GSH) on bovine embryo development. Bovine embryos fertilized in vitro were cultured in media with 3 mmol·L^-1 GSH for 7 days in treatment group and without GSH for 7 days in control group,respectively. The rates of cleavage, morula and blastocyst formation of embryos, and the total cell number of blastocysts were calculated in both groups. The contents of reactive oxygen species (ROS) and GSH in zygotes at different stages(8,18,24,32 hours post-insemination)were detected by fluorescence staining and liquid chromatography mass spectrometry (LC/MS). Additionally, the changes of isotope labelled GSH (GSX) contents of bovine zygotes and culture media were tracked by LC/MS after GSX was added in culture medium. Results showed that:1)Exogenous GSH significantly reduced ROS contents of embryos(18,24,32 hours post-insemination)(P<0.05,P<0.01). 2)The morula rate (58.67% vs. 48.46%), the blastocyst rate (50.83% vs. 34.88%) and the total cell number of blastocysts (102.26 vs. 76.27) in treatment group were significantly higher than that in control group (P<0.05).3) GSH contents of zygotes at 3 stages in treatment group were significantly higher than that in control group (P<0.05). Meanwhile, GSH contents of zygotes in both groups decreased along with the developmental process(18,24,32 hours post-insemination). 4)After addition of GSX, the contents of GSH of zygotes in treatment group increased significantly (P<0.05). In addition, GSX concentrations in culture media decreased steadily with the extension of culture time(18,24,32 hours post-insemination). Conclusively, exogenous GSH improves the development rate of bovine embryo and blastocyst quality with scavenging of ROS. Moreover, LC/MS is a reliable method for quantitative analysis of GSH and isotope labeled GSH. Therefore LC/MS can be used to explore the mechanism of exogenous GSH improving the development of embryo cultured in vitro.

CLC Number:

S823.2

LI Feng, CUI Li-xin, HAO Hai-sheng, LIU Yan, ZHAO Xue-ming, PANG Yun-wei, ZHU Hua-bin, DU Wei-hua. Quantitative Analysis of Glutathione Levels in Bovine in vitro Embryos by LC/MS[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(12): 2622-2631.

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Quantitative Analysis of Glutathione Levels in Bovine in vitro Embryos by LC/MS

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