Determination of Norcantharidin in Mouse Tissues by Liquid Chromatography Coupled to Tandem Mass Spectrometry and its Tissue Distribution Study

Abstract

The purpose of this study is to determine the concentrations of norcantharidin (CAS NO: 5442-12-6) in mouse tissues and investigate its tissue distribution after intragastric administration of disodium norcantharidate solution. A highly sensitive and specific liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated, using ribavirin (CAS NO: 36791-04-5) as the internal standard (IS). Norcantharidin and IS were extracted from 0.3 mL tissue homogenates using protein precipitation with acetone under acid condition. The analyte was separated on a C18 reverse phase column and analyzed by MS/MS in the multiple reaction monitoring (MRM) mode using ESI with positive ionization, m/z 169→123 for norcantharidin and m/z 267→135 for IS. The developed method was validated over a linear range of concentrations 0.01~5 μg·mL^− 1 in liver, lung, kidney, stomach, small intestine, uterus and testis, 0.005~0.5 μg·mL^− 1 in heart, spleen and brain, the correlation coefficients (r²) were between 0.9918 and 0.9976. The tissue distribution study result was as follows: The AUC_0-t of norcantharidin in tissues was in the order as follows: small intestine, stomach, uterus, kidney, testis, liver, lung, spleen, heart, brain.

Key words

norcantharidin - tissue distribution - LC-MS/MS

• References

• 1 Zheng YL, Shi CL, He YX. Clinical studies on liver cancer of norcantharidin. Hebei Yi Xue 2005; 11: 820-821
• 2 Dong LH, Qu YQ, Liu F et al. Clinical observation on late stage of esophageal carcinoma treated with norcantharidin coupled with radiotherapeutics. Jilin Yi Xue 2002; 5: 301
• 3 An WW, Gong XF, Wang MW et al. Norcantharidin induces apoptosis in HeLa cells through caspase, MAPK, and mitochondrial pathways. Acta Pharmacol Sin 2004; 25: 1502-1508
• 4 An WW, Xue L, Wang MW et al. Apoptosis of L929 cells induced by norcantharidin. China Oncology 2005; 15: 22-25
• 5 Fan YZ, Fu JY, Zhao ZM et al. Effect of norcantharidin on proliferation and invasion of human gallbladder carcinoma GBC-SD cells. World J Gastroenterol 2005; 16: 2431-2437
• 6 Huang XQ, Ling CQ, Song TB et al. Study on Apoptosis of Human Acute T Lymphocyte Leukemia Cell Line 6t-CEM induced by Norcantharidin. Zhejiang Zhong Yi Xue Yuan Xue Bao 2001; 25: 49-52
• 7 Liang FY, Rong Y, Chen L et al. The killing effect and possible mechanism of norcantharidin on mammalian cells. Xi Bei Da Xue Xue Bao (Natural Science Edition) 1999; 29: 262-264
• 8 Massicot F, Dutertre-Catella H, Pham-Huy C et al. In vitro assessment of Renal Toxicity and Inflammatory Events of Two Protein Phosphatase Inhibitors Cantharidin and Norcantharidin. Basic Clin Pharmacol Toxicol 2005; 96: 26-32
• 9 Wei CM, Wang BJ, Ma Y et al. Pharmacokinetics and biodistribution of ³H-norcantharidin in mice. Yao Xue Xue Bao 2007; 42: 516-519
• 10 Wei CM, Zhang R, Wang BJ et al. Determination and pharmacokinetic study of norcantharidin in human serum by HPLC-MS/MS method. Biomed Chromatogr 2008; 22: 44-49