Application of HPLC-MS/MS in the analysis of six tricarboxylic acid cycle metabolites in a mouse model of prenatal DEHP exposure
10.19405/j.cnki.issn1000-1492.2025.05.017
- Author:
Wenkang Tao
1
;
Lun Zhang
2
;
Qianqian Huang
2
;
Yun Yu
2
;
Yue Jiang
1
;
Mengzhen Hou
1
;
Xutao Ling
1
;
Fang Xie
1
;
Jianqing Wang
1
Author Information
1. School of Pharmaceutical Sciences , Anhui Medical University , Hefei 230032 ; Dept of Pharmacy , The First Afiliated Hospital of Anhui Medical University North District , Hefei 230012 ; Anhui Public Health Clinical Center , Hefei 230012
2. Dept of Pharmacy , The First Afiliated Hospital of Anhui Medical University North District , Hefei 230012 ; Anhui Public Health Clinical Center , Hefei 230012
- Publication Type:Journal Article
- Keywords:
tricarboxylic acid cycle;
tricarboxylic acid cycle metabolites;
high performance liquid chromatography_mass spectrometry system;
pregnant mice;
methodology;
DEHP
- From:
Acta Universitatis Medicinalis Anhui
2025;60(5):897-905
- CountryChina
- Language:Chinese
-
Abstract:
Objective :To establish a method for measuring major organic acids in the tricarboxylic acid cycle using a high-performance liquid chromatography-tandem mass spectrometry(HPLC-MS/MS) system, and to investigate the changes in six tricarboxylic acid cycle organic acids(fumaric acid, malic acid, succinic acid, α-ketoglutaric acid, cis-aconitic acid, and citric acid) in the serum, liver, and placenta of mice exposed to di(2-ethylhexyl) phthalate(DEHP) during pregnancy.
Methods :The serum, liver and placental samples from pregnant mice were processed and eluted through a Waters ACQUITY UPLC BEH Amide Column(130 Å, 1.7 μm, 2.1 mm × 150 mm) using a gradient elution program. Mobile phase A comprised an aqueous solution of 10 mmol/L ammonium acetate and 5 μmol/L methanephosphonic acid, while mobile phase B consisted of a 90% acetonitrile aqueous solution containing 10 mmol/L ammonium acetate and 5 μmol/L methanephosphonic acid, with a flow rate maintained at 0.35 ml/min. The mass spectrometry detection system utilized an electrospray ionization technique with negative ion mode for multiple reaction monitoring.
Results : The correlation coefficients of the standard curves for the six tricarboxylic acid cycle organic acid metabolites were all above 0.996 within the quantitative range. The method's accuracy ranged from 97.14% to 108.26%, with inter-day and intra-day precision relative standard deviation between 1.35% and 6.73%. The matrix effect was between 93.29% and 107.47%, and the extraction recovery rate ranged from 94.82% to 112.57%. Analysis of six tricarboxylic acid cycle organic acids in the liver, serum, and placenta of DEHP-exposed mice during pregnancy showed significant reductions in fumaric acid, malic acid, α-ketoglutaric acid, cis-aconitic acid, and citric acid compared to the control group(P<0.05).
Conclusion : The HPLC-MS/MS method established in this study for detecting six tricarboxylic acid cycle organic acids in the serum, liver, and placenta of DEHP-exposed pregnant mice is stable, highly sensitive and selective. Prenatal DEHP exposure induced alterations in the levels of tricarboxylic acid(TCA) cycle organic acid metabolites in the liver, serum, and placenta of mice, suggesting that DEHP exposure during pregnancy may interfere with mitochondrial TCA cycle processes. These findings indicate potential value in the diagnosis and treatment of diseases associated with prenatal DEHP exposure.
- Full text:202602051533506839HPLC-MS_MS检测6种三羧酸循环代谢物在孕期DEHP暴露小鼠模型中的应用_陶文康.pdf
