UPLC-MS/MS Method for Detection of Etomidate and Its Metabolite Etomidate Acid Quantity in Blood

doi:10.12116/j.issn.1004-5619.2023.330901

Abstract

Abstract:

Objective To establish a method for the simultaneous quantitative analysis of etomidate and its metabolite etomidate acid in blood, and to discuss its application value in actual cases. Methods Acetonitrile precipitate protein method was used, and C₁₈ column was selected. Gradient elution was performed with acetonitrile and 5 mmol/L ammonium acetate within 6 min. Electrospray ionization source in positive ion mode was used. The internal standard etomidate acid-d₅ was obtained by etomidate-d₅ alkaline hydrolysis reaction. Ultra-high performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS） was used for quantitative analysis. The methodological verification was conducted. Results Etomidate and etomidate acid in blood showed good linear relationship in the quantitative linear range （r>0.999）, with the lower limit of quantification was 2.5 ng/mL and 7.5 ng/mL, respectively. The accuracy, precision, recovery rate, and matrix effect of the method met the professional verification standards. The practical application results showed that etomidate and etomidate acid could be detected in the blood of the abusers, and their mass concentrations ranged from 17.24 to 379.93 ng/mL. Conclusion The method established in this study can simultaneously quantify etomidate and etomidate acid in blood, which is simple and convenient to operate with accuracy. It can meet the detection needs of actual cases and provide technical support for law enforcement to crack down on etomidate abuse.

Key words: forensic medicine, toxicological analysis, etomidate, etomidate acid, ultra-high performance liquid chromatography-tandem mass spectrometry, blood

CLC Number:

Xing HAN, Xin LIU, Ming-luo DU, Ruo-lun XU, Jia-rong LI, Chao LIU, Wei-guo LIU. UPLC-MS/MS Method for Detection of Etomidate and Its Metabolite Etomidate Acid Quantity in Blood[J]. Journal of Forensic Medicine, 2023, 39(6): 564-570.

Figures/Tables 10

References 16

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目标物	保留时间/min	定量离子对	定性离子对	Q1偏差电压/V	碰撞能量/eV	Q3偏差电压/V
依托咪酯	3.30	245.2/141.1	245.2/141.1	-10	-10	-13
			245.2/105.1	-10	-26	-17
			245.2/95.1	-13	-24	-16
依托咪酯-d₅	3.30	250.2/141.1	250.2/141.1	-10	-10	-13
			250.2/110.1	-10	-26	-17
			250.2/95.1	-13	-24	-16
依托咪酯酸	2.43	217.2/113.1	217.2/113.1	-17	-10	-20
			217.2/105.1	-12	-22	-18
			217.2/95.1	-25	-22	-15
依托咪酯酸-d₅	2.43	222.2/113.1	222.2/113.1	-17	-10	-20
			222.2/110.1	-12	-22	-18
			222.2/95.1	-25	-22	-15

目标物	保留时间/min	定量离子对	定性离子对	Q1偏差电压/V	碰撞能量/eV	Q3偏差电压/V
依托咪酯	3.30	245.2/141.1	245.2/141.1	-10	-10	-13
			245.2/105.1	-10	-26	-17
			245.2/95.1	-13	-24	-16
依托咪酯-d₅	3.30	250.2/141.1	250.2/141.1	-10	-10	-13
			250.2/110.1	-10	-26	-17
			250.2/95.1	-13	-24	-16
依托咪酯酸	2.43	217.2/113.1	217.2/113.1	-17	-10	-20
			217.2/105.1	-12	-22	-18
			217.2/95.1	-25	-22	-15
依托咪酯酸-d₅	2.43	222.2/113.1	222.2/113.1	-17	-10	-20
			222.2/110.1	-12	-22	-18
			222.2/95.1	-25	-22	-15

温度	时间/h	依托咪酯-d₅		依托咪酯酸-d₅
温度	时间/h	保留时间/min	峰面积	保留时间/min	峰面积
室温	0	3.260	2.25×10⁷	/	/
	2	3.260	1.91×10⁶	2.382	1.42×10⁷
	4	3.273	3.34×10⁵	2.419	1.56×10⁷
	6	3.257	6.14×10⁴	2.372	1.63×10⁷
	18	3.253	2.79×10³	2.374	1.77×10⁷
40 ℃	0	3.260	2.25×10⁷	/	/
	2	3.269	6.45×10³	2.384	1.57×10⁷
	4	3.259	6.76×10³	2.384	1.60×10⁷
	6	3.267	3.71×10²	2.389	1.61×10⁷
	18	3.267	1.85×10²	2.372	1.59×10⁷

温度	时间/h	依托咪酯-d₅		依托咪酯酸-d₅
温度	时间/h	保留时间/min	峰面积	保留时间/min	峰面积
室温	0	3.260	2.25×10⁷	/	/
	2	3.260	1.91×10⁶	2.382	1.42×10⁷
	4	3.273	3.34×10⁵	2.419	1.56×10⁷
	6	3.257	6.14×10⁴	2.372	1.63×10⁷
	18	3.253	2.79×10³	2.374	1.77×10⁷
40 ℃	0	3.260	2.25×10⁷	/	/
	2	3.269	6.45×10³	2.384	1.57×10⁷
	4	3.259	6.76×10³	2.384	1.60×10⁷
	6	3.267	3.71×10²	2.389	1.61×10⁷
	18	3.267	1.85×10²	2.372	1.59×10⁷

化合物	线性范围/（ng·mL^-1）	线性方程	相关系数（r）	LOD/（ng·mL^-1）	LLOQ/（ng·mL^-1）
依托咪酯	2.5~500	y=1.256 0 x+0.001 1	0.999 6	0.25	2.5
依托咪酯酸	7.5~1 500	y=0.033 0 x+0.023 4	0.999 3	0.30	7.5