ABSTRACT
Three-dimensional(3D)cell spheroid models combined with mass spectrometry imaging(MSI)enables innovative investigation of in vivo-like biological processes under different physiological and patho-logical conditions.Herein,airflow-assisted desorption electrospray ionization-MSI(AFADESI-MSI)was coupled with 3D HepG2 spheroids to assess the metabolism and hepatotoxicity of amiodarone(AMI).High-coverage imaging of＞1100 endogenous metabolites in hepatocyte spheroids was achieved using AFADESI-MSI.Following AMI treatment at different times,15 metabolites of AMI involved in N-desethylation,hydroxylation,deiodination,and desaturation metabolic reactions were identified,and according to their spatiotemporal dynamics features,the metabolic pathways of AMI were proposed.Subsequently,the temporal and spatial changes in metabolic disturbance within spheroids caused by drug exposure were obtained via metabolomic analysis.The main dysregulated metabolic pathways included arachidonic acid and glycerophospholipid metabolism,providing considerable evidence for the mechanism of AMI hepatotoxicity.In addition,a biomarker group of eight fatty acids was selected that provided improved indication of cell viability and could characterize the hepatotoxicity of AMI.The combination of AFADESI-MSI and HepG2 spheroids can simultaneously obtain spatiotemporal infor-mation for drugs,drug metabolites,and endogenous metabolites after AMI treatment,providing an effective tool for in vitro drug hepatotoxicity evaluation.
ABSTRACT
Deconvolution of potential drug targets of the central nervous system (CNS) is particularly challenging because of the complicated structure and function of the brain. Here, a spatiotemporally resolved metabolomics and isotope tracing strategy was proposed and demonstrated to be powerful for deconvoluting and localizing potential targets of CNS drugs by using ambient mass spectrometry imaging. This strategy can map various substances including exogenous drugs, isotopically labeled metabolites, and various types of endogenous metabolites in the brain tissue sections to illustrate their microregional distribution pattern in the brain and locate drug action-related metabolic nodes and pathways. The strategy revealed that the sedative-hypnotic drug candidate YZG-331 was prominently distributed in the pineal gland and entered the thalamus and hypothalamus in relatively small amounts, and can increase glutamate decarboxylase activity to elevate γ-aminobutyric acid (GABA) levels in the hypothalamus, agonize organic cation transporter 3 to release extracellular histamine into peripheral circulation. These findings emphasize the promising capability of spatiotemporally resolved metabolomics and isotope tracing to help elucidate the multiple targets and the mechanisms of action of CNS drugs.
ABSTRACT
With the rapid development and wide application of traditional Chinese medicine injection (TCMI), a number of adverse events of some TCMIs have incessantly been reported and have drawn broad attention in recent years. Establishing effective and practical analytical methods for safety evaluation and quality control of TCMI can help to improve the safety of TCMIs in clinical applications. In this study, a sensitive and rapid high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method has been developed and validated for the quantitative determination of potentially harmful substance 5,5'-oxydimethylenebis (2-furfural, OMBF) in TCMI samples. Chromatographic separation was performed on a C18 reversed-phase column (150 mm × 2.1 mm, 5 µm) by gradient elution, using methanol-water containing 0.1% formic acid as mobile phase at the flow rate of 0.3 mL/min. MS/MS detection was performed on a triple quadrupole mass spectrometer with positive electrospray ionization in the multiple reaction-monitoring mode. The method was sensitive with a limit of quantification of 0.3 ng/mL and linear over the range of 0.3-30 ng/mL (=0.9998). Intra- and inter-day precision for analyte was <9.52% RSD with recoveries in the range 88.0-109.67% at three concentration levels. The validated method was successfully applied to quantitatively determine the compound OMBF in TCMIs and glucose injections. Our study indicates that this method is simple, sensitive, practicable and reliable, and could be applied for safety evaluation and quality control of TCMIs and glucose injections.
ABSTRACT
To screen the harmful substance 5-hydroxymethyl furfural content in commercially available traditional Chinese medicine injection which are commonly used, and to preliminarily evaluate the quality of these injections, 5-hydroxymethyl furfural was taken as an index. The contents of 5-hydroxymethyl furfural in 56 samples which consist of 23 kinds of traditional Chinese medicine injections and glucose injection were determined using LC-MS/MS, and 5-hydroxymethyl furfural was detected in 52 of these samples. The minimal content was 0.0038 microg x L(-1) and the maximum content was 1420 microg x mL(-1). The contents of 5-hydroxymethyl furfural were significantly different in traditional Chinese medicine injection which came from different kinds, manufacturers or batches. The results showed the quality difference of commercially available traditional Chinese medicine injection is significant taking 5-hydroxymethyl furfural content as assessment index. More attention should be paid to the safety of 5-hydroxymethyl furfural in traditional Chinese medicine injection, and unified limitation standard should be set to improve medication safety of traditional Chinese medicine injection.