OBJECTIVES: To establish a LC-MS/MS method which is accurate and sensitive for determination of koumine, gelsemine, and gelsenicine in biological samples and to verify the method. METHODS: Strychnine was used as internal standard. Analytes in blood, urine and liver with 1% sodium hydroxide solution were extracted by ethyl acetate. Chromatographic separation was achieved on a ZORBAX SB-C₁₈ column （150 mm×2.1 mm, 5 μm）, and gradient elution was performed with the buffer solution of methanol-20 mmol/L ammonium acetate （including 0.1% formic acid and 5% acetonitrile） as mobile phase. Qualitative and quantitative analysis was performed in the multiple reaction monitoring mode coupled with an electrospray ionization source under positive ion mode（ESI⁺）. RESULTS: The linearity of koumine, gelsemine and gelsenicine in blood, urine and liver was good within corresponding linear limitation and the correlation coefficients （r）>0.995 0. The limits of detection were 0.1 ng/mL （0.1 ng/g）, 0.1 ng/mL （0.1 ng/g） and 0.01 ng/mL （0.01 ng/g）, respectively. The extraction recovery and accuracy of the alkaloids ranged from 61.9% to 114.6% and 92.4% to 114.3%, respectively. The relative standard deviations of the intra-day and inter-day precisions were not more than 11.0%. CONCLUSIONS The method is selective, sensitive and suitable for simultaneous determination of koumine, gelsemine and gelsenicine in body fluids and tissues, which offering technical support for clinical diagnosis and treatment and forensic toxicological analysis of Gelsemium elegans poisoning.
Alkaloids/urine* ; Chromatography, High Pressure Liquid ; Chromatography, Liquid ; Forensic Toxicology ; Formates ; Humans ; Indole Alkaloids/urine* ; Liver ; Reproducibility of Results ; Strychnine ; Tandem Mass Spectrometry

AIMTo identify the main metabolites of aconitine in the urine of rabbits.

METHODSAfter oral administration of aconitine (5 mg.kg-1), the urine of male rabbits was collected and extracted by solid phase extraction and analyzed by liquid chromatography-ion trap mass spectrometry.

RESULTSAconitine and 4 metabolites were found in the rabbit urine. Their protonated molecular ions at m/z 632, m/z 604, m/z 590, m/z 500 and multistage fragment ions with neutral loss of 60 u, 32 u, 28 u and 18 u were monitored. Their relative concentration were M1 > Aconitine > M4 > M2 > M3.

CONCLUSIONThe metabolites M1-M4 were deduced as 16-O-demethylaconitine, benzoylaconine, 16-O-demethylbenzoylaconine and aconine, respectively.

Aconitine ; analogs & derivatives ; metabolism ; urine ; Alkaloids ; urine ; Animals ; Chromatography, High Pressure Liquid ; Male ; Rabbits ; Spectrometry, Mass, Electrospray Ionization

This study aimed to profile the chemical constituents of Zi-Shen pill (ZSP) and its metabolites in plasma, urine, and prostate tissue, after administration into rats. Based on the chromatographic retention behavior, fragmentation patterns of chemical components, published literatures, and literature databases, an UPLC-Q-TOF/MS (LC-TOF/MS) method was established to identify the components of ZSP and its metabolites in biological samples. A total of 101 compounds were identified and tentatively characterized from the ZSP, including alkaloids, xanthones, and timosaponins. Except for 33 prototype components, 22 metabolites were detected in the plasma, urine, and prostate, and mainly came from Phellodendri Amurensis Cortex and Anemarrhenae Rhizoma. It was found that glucuronidation and sulfation were the major metabolic processes of xanthones, while oxidation, demethylation, and glucuronidation were the major metabolic pathways of alkaloids. In summary, the present study provided important chemical information on the metabolism of ZSP, indicating that alkaloids might be able to be absorbed into the prostate. The results provided a basis for further studies of the mechanisms of action for ZSP.
Alkaloids ; blood ; urine ; Animals ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; metabolism ; Male ; Plasma ; chemistry ; Rats ; Tandem Mass Spectrometry ; Urine ; chemistry ; Xanthones ; blood ; urine

In this study, UPLC-MS/MS was adopted to determine the contents of five ephedrine alkaloids (Norephedrine, Norpseudoephedrine, Ephedrine, Pseudoephedrine, Methylephedrine) in plasma and urine in rats after the combined administration of Ephedrae Herba-Gypsum Fibrosum and calculate relevant pharmacokinetic parameters, in order to discuss the effect of the combined administration of Ephedrae Herba-Gypsum Fibrosum on plasma pharmacokinetics and urinary excretion characteristics. According to the results, after being combined with Gypsum, the five ephedrine alkaloids showed similar pharmacokinetic changes, such as shortened t(max), accelerated absorption rate, but reduced AUC(0-t) and V(z)/F, which may be related to the increase in urine excretion. Besides, Gypsum was added to enhance C(max) of Pseudoephedrine and prolong MRT(0-t) of Methylephedrine, so as to enhance the anti-asthmatic effect of Ephedrae Herba and resist the toxic effect of Norephedrine and Ephedrine. This study proved the scientific compatibility of Ephedrae Herba-Gypsum Fibrosum and provided a reference for studies on the prescription compatibility regularity and relevant practices.
Alkaloids ; blood ; pharmacokinetics ; urine ; Animals ; Calcium Sulfate ; pharmacokinetics ; Drugs, Chinese Herbal ; pharmacokinetics ; Ephedra ; chemistry ; Male ; Rats ; Rats, Sprague-Dawley ; Urine ; chemistry

AIMTo identify the main metabolites of oxymatrine (OMT) in rats.

METHODSTo optimize the conditions of LC/ESI-ITMS' chromatograms and spectra by oxymatrine and matrine (MT), and summarize their ionization and cleavage rules in ESIMS, then serving as the basis for the metabolite analyses of oxymatrine in rats. To collect the 0-24 h urine samples of the rats after ip 40 mg x kg(-1) oxymatrine, the samples were enriched and purified through C18 solid-phase extraction cartridge. The purified samples were analyzed by LC/ESI-ITMS. The structures of OMT metabolites were identified according to their retention times and ESI-ITMSn rules.

RESULTSSix phase I metabolites and the parent drug OMT were found in the rat urine, and the main metabolite was MT. No phase II metabolites were found.

CONCLUSIONThe developed LC/ESI-ITMSn methods to identify the metabolites of oxymatrine in rats is not only simple and rapid but also sensitive and specific. This technology is one of the most efficient methods for the analysis of drug metabolites.

Alkaloids ; isolation & purification ; pharmacokinetics ; urine ; Animals ; Chromatography, Liquid ; methods ; Plants, Medicinal ; chemistry ; Quinolizines ; isolation & purification ; pharmacokinetics ; urine ; Rats ; Rats, Wistar ; Sophora ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; methods

OBJECTIVE: To establish a new method for determination of strychnine alkaloid in biological fluids based on molecularly imprinted polymers. METHODS: A strychnine molecularly imprinted monolithic column was prepared by in-situ molecularly imprinted technique. The polymer was filled to a 1cm column, and a method was developed to concentrate and determine strychnine alkaloids in biological fluids. RESULTS: the limit of detection of the method was 4.9 ng, and the recoveries were more than 92%. The relative standard deviations were smaller than 6.59%. The linear correlation coefficients of standard curves were 0.999 1 and 0.9966 respectively. This method was applied to concentrate and determine strychnine in plasma and urine of poisoned rabbit. CONCLUSION The new method could concentrate and simultaneously determine strychnine alkaloids in biological fluids, and it was applied to forensic toxicological analysis.
Alkaloids/analysis* ; Animals ; Chromatography, High Pressure Liquid/methods* ; Humans ; Male ; Polymers/chemistry* ; Rabbits ; Sensitivity and Specificity ; Strychnine/urine*

AIMTo study the in vitro and in vivo metabolism of (-)-securinine.

METHODSThe metabolic transformation of (-)-securinine was studied by using phenobarbital-induced rat liver microsomal incubate containing the NADPH-generating system in vitro and the constitution of the system was optimized. A reversed phase HPLC method was established to analyze the parent drug and its metabolites. The major metabolites were isolated and purified by liquid-liquid extraction, preparative TLC and HPLC, and their structures were elucidated as 6-hydroxyl securinine, 6-carbonyl securinine, 5 beta-hydroxyl securinine and 5 alpha-hydroxyl securinine by 1HNMR, 13CNMR and MS spectral analysis. An HPLC method was developed to analyze securinine and its metabolites in biofluids (bile, urine) of rat. The bile, urine and their enzymatic hydrolyzed samples of the rat i.p. administrated with (-)-securinine were determined by using this method.

RESULTSFour main metabolites of (-)-securinine in rat hepatic microsome incubation were obtained and their structures were elucidated. Metabolites from in vitro study were confirmed in biofluids (bile, urine) which were collected from rats given securinine i.p. It was suggested that 6-hydroxyl securinine was excreted in conjugated form as well by analyzing enzymatic hydrolyzed bile.

CONCLUSIONThe main metabolic pathway of (-)-securinine in vitro and in vivo is basically elucidated.

Alkaloids ; isolation & purification ; metabolism ; urine ; Animals ; Azepines ; isolation & purification ; metabolism ; urine ; Bile ; metabolism ; Euphorbiaceae ; chemistry ; Heterocyclic Compounds, 4 or More Rings ; isolation & purification ; metabolism ; urine ; Heterocyclic Compounds, Bridged-Ring ; In Vitro Techniques ; Lactones ; isolation & purification ; metabolism ; urine ; Male ; Microsomes, Liver ; metabolism ; Piperidines ; isolation & purification ; metabolism ; urine ; Plants, Medicinal ; chemistry ; Rats ; Rats, Wistar ; Stereoisomerism

AIMTo establish a high-performance capillary electrophoresis (HPCE) chiral separation method for d-securinine and l-securinine, and use this method to investigate the stereoselective metabolism process of d- and l-securinine in Wistar rats.

METHODSThe electrophoretic condition and parameters were investigated and the optimized conditions were as following: the electrophoretic medium was 40 mmol.L-1 Tris-H3PO4 buffer (pH adjusted to 6.0 with H3PO4) containing 32 mmol.L-1 HP-beta-CD as chiral selector. Determination was carried out with a UV detector at 254 nm. The separations were performed at 16 degrees C with a positive voltage of 15 kV. Samples were injected into the capillary by pressure for 6 s. The biological samples (urine, bile, plasma and feces) of rats were alkalized and extracted with ethyl acetate.

RESULTSThe experimental results showed that the concentration of HP-beta-CD, the concentration of the running buffer and the pH value of the buffer were the main important factors which effected the resolution. d-Securinine and l-securinine were separated at baseline level under the determination conditions. The determination was not interfered by endogenous components and metabolites. After i.p. administration, the rats excreted more d-securinine than l-securinine through bile, urine and feces. The metabolism process in rats was stereoselective.

CONCLUSIONThis method is simple, reliable and suitable for studying the stereoselective metabolism of securinine in rats.

Alkaloids ; chemistry ; isolation & purification ; metabolism ; urine ; Animals ; Azepines ; chemistry ; isolation & purification ; metabolism ; urine ; Bile ; metabolism ; Electrophoresis, Capillary ; methods ; Euphorbiaceae ; chemistry ; Heterocyclic Compounds, 4 or More Rings ; chemistry ; isolation & purification ; metabolism ; urine ; Heterocyclic Compounds, Bridged-Ring ; Lactones ; chemistry ; isolation & purification ; metabolism ; urine ; Male ; Molecular Structure ; Piperidines ; chemistry ; isolation & purification ; metabolism ; urine ; Plants, Medicinal ; chemistry ; Rats ; Rats, Wistar ; Stereoisomerism