Identification of biomarkers for bongkrekic acid poisoning by liquid chromatography-tandem mass spectrometry
XU Xiaomin1,2,3, AN Yuxin1,2,3, XU Meijia3, XU Jiaojiao1,2,3, CAI Zengxuan1,2,3, HAN Jianlong1,2,3
1. NHC Specialty Laboratory of Food Safety Risk Assessment and Standard Development, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, Zhejiang 310051, China; 2. Zhejiang Key Laboratory of Public Health Detection and Pathogenesis Research, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, Zhejiang 310051, China; 3. Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, Zhejiang 310051, China
Abstract：Objective To explore the distribution and metabolic characteristics of bongkrekic acid in plasma, serum and urine, and to identify potential biomarkers for bongkrekic acid poisoning, so as to provide evidence for etiological identification and therapeutic effect evaluation. Methods Blood samples were centrifuged to separate plasma or serum and then stored frozen, while urine samples were directly frozen for preservation. After thawing, the samples were treated with internal standard working solution and 0.5% formic acid acetonitrile solution, and detected after centrifugation and filtration. A C18 chromatographic column with gradient elution was adopted, and electrospray ionization in negative multiple reaction monitoring mode was used with optimized ion pair parameters for the determination of bongkrekic acid. The method was evaluated in terms of matrix effect, linear range, coefficient of determination (R2), limit of detection, limit of quantification, average spiked recovery rate, and relative standard deviation (RSD). Determination of the distribution and mass concentration of bongkrekic acid in three types of samples by liquid chromatography-tandem mass spectrometry, and analysis of its metabolic characteristics. Results The matrix effects of bongkrekic acid in plasma, serum and urine were 86.8%, 88.1% and 93.3%, respectively. The results of methodological validation demonstrated that bongkrekic acid exhibited good linearity within the concentration range of 0.2-100 ng/mL, with R2 of 0.999. The limit of detection and limit of quantification were determined to be 1.0 ng/mL and 2.0 ng/mL, respectively. No interfering peaks were observed in the chromatograms. The average spiked recovery rates ranged from 92.1% to 104.0%, with RSD ranging from 2.9% to 6.8%. Among the 12 patients with bongkrekic acid poisoning, the mass concentrations of bongkrekic acid in plasma and serum from 8 patients within 1-14 days of poisoning ranged from 168.0 to 967.0 ng/mL and from 163.0 to 1 017.0 ng/mL, respectively, with average mass concentration proportions of 50.3% and 49.7%, respectively. Among them, bongkrekic acid mass concentration decreased overall in 2 patients after plasma exchange treatment. Conclusion Bongkrekic acid in plasma or serum can serve as an effective biomarker for poisoning diagnosis and therapeutic effect evaluation of bongkrekic acid poisoning.
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