脱氢氯甲睾酮人体内代谢研究

本文叙述了用GC-MS联用技术研究脱氢氯甲睾酮人体内代谢的方法。尿样中的甾体化合物经大孔树脂吸附提取后,酶解。浓缩并衍生化,进行GC-MS分析。在服药后8～30h的尿样中,检出了脱氢氯甲睾酮原型,并发现了七个代谢产物。分析色谱和质谱数据,得到了这些代谢物的结构及其浓度变化趋势,推测了脱氢氯甲睾酮可能的体内代谢模式,确定了筛检尿中脱氢氯甲睾酮的特定代谢物和特征检测离子,比较了不同的样品处理方法对分析结果的影响。     

人尿中卡鲁睾酮的代谢产物研究

使用GC/MS方法对人尿中卡鲁睾酮(calusterone)的代谢情况进行了研究。尿样经XAD-2树脂柱吸附、酶水解、乙醚萃取及三甲基硅烷衍生化处理后,使用毛细管柱进行GC/MS分析,鉴定了7种代谢物,从而推导出卡鲁睾酮在人体中的代谢模式。     

人尿中卡鲁睾酮的代谢产物研究

使用GC/MS方法对人尿中卡鲁睾酮(calusterone)的代谢情况进行了研究。尿样经XAD-2树脂柱吸附、酶水解、乙醚萃取及三甲基硅烷衍生化处理后,使用毛细管柱进行GC/MS分析,鉴定了7种代谢物,从而推导出卡鲁睾酮在人体中的代谢模式。     

人尿中奥美拉唑代谢物的提取与精制

目的对奥美拉唑在人体内代谢后尿中2种代谢物进行提取与精制后推测其结构并测定其含量。方法24名健康受试者单剂量口服奥美拉唑肠溶胶囊40mg后,收集服药后12h内尿液,用乙醚提取和浓缩,以高效液相色谱法进行分离,将相对纯品进行质谱扫描测定,推测其结构并计算含量。结果经分离得到的2种代谢物推测为吡啶5’—或3’—甲基氧化生成的羟基砜型代谢物、吡啶环上5’—甲基羟化硫醚型代谢物,含量分别为96.54%、97.26%。结论本方法分离得到的奥美拉唑尿中代谢物质纯度较高。     

尿中氯胺酮及其代谢物的定性分析

目的:了解氯胺酮在人体内的代谢情况和代谢产物,为检测滥用氯胺酮者尿液提供定性分析手段.方法:用固相萃取及GC/MS的选择性离子模式(SIM)检测尿中氯胺酮及其2种代谢物.结果:运用GC/MS确认了氯胺酮滥用者尿液中的有效成分氯胺铜及其代谢物去甲基氯胺酮和脱氢去甲基氯胺酮.结论:上述方法可以用于滥用者尿中氯胺酮及其代谢物的测定.     

LC-MS/MS法快速分析大鼠尿液中他喷他多的代谢物

目的建立快速分析大鼠尿液中他喷他多代谢物的方法。方法大鼠灌胃给予他喷他多,收集空白和给药后0～12 h尿液,以液相色谱-串联质谱法,采用多离子反应监测(multiple reactionmonitoring,MRM)及二级全扫描质谱(full scan MS2)方式,分析尿液中他喷他多的代谢物。结果在大鼠尿液中发现了他喷他多原形药物及其15种代谢物,首次发现了他喷他多脱氢化合物、他喷他多脱氢葡萄糖醛酸结合物。结论本方法简便、快速,适用于大鼠尿液中他喷他多代谢物结构及代谢路径分析。     

高效液相-质谱联用法对盐酸关附甲素在大鼠尿中代谢物的研究

目的研究盐酸关附甲素在大鼠尿中的代谢产物。方法大鼠iv盐酸关附甲素后收集尿,用高效液相-质谱联用方法测定。通过与标准化合物的色谱保留时间、分子离子峰、碎片离子峰对照从而鉴定I相代谢物。通过用葡糖醛酸酶和硫酸酯酶酶解鉴定其水解产物(苷元)从而确定II相结合物。结果大鼠尿中发现I相代谢物关附醇胺和关附壬素;尿经过葡糖醛酸酶和硫酸酯酶酶解后,产生关附甲素和关附壬素。结论盐酸关附甲素在大鼠体内可以转化为关附壬素、关附醇胺、关附甲素葡糖醛酸和硫酸结合物、关附壬素葡糖醛酸和硫酸结合物。经过生物转化,代谢产物的极性增加,药效下降。     

血、尿中安眠酮及其代谢物的测定

通过一例安眠酮中毒病人血、尿中安眠酮及其代谢物的测定,描述了用紫外光谱(uv)、气相色谱(GC)和气相色谱质谱(GC/MS)法测定安眠酮及其代谢物的系统分析方法。样品的提取净化采用液一液萃取和固相萃取两种方法,都得到了很好的结果。紫外光谱用于测定血、尿中安眠酮和其代谢物的总量;气相色谱用于测定血、尿中安眠酮原药的含量;气相色谱质谱则用于鉴定血、尿中的安眠酮及其代谢物。除安眠酮外,血、尿中共检出10种安眠酮代谢物,其中包括两种乙酰化代谢物。此法还为临床救治提供指导。     

左旋紫草素体内代谢研究

本文对紫草素在大鼠体内的生物转化进行了研究,对经苯巴比妥预处理（i.p.60mg.kg^-1.day,3天）的Wistar种健康雄性大鼠腹腔注射紫草素（20mg.kg^-1),收集到的胆汁和血样分别经β－葡萄糖醛酸酶水解后,用乙酸乙酯提取,建立PP－HPLC／DAD方法对紫草素及其代谢物大大鼠胆汁和尿样中的分布进行了初步考察,根据色谱峰的保留时间和紫外光谱分析,大鼠的胆汁和尿样中分别有10个代谢物。     

刺激剂类药物及代谢物的分离鉴定

报道了41种刺激剂的气相色谱和质谱数据以及人尿中原药和它们的游离型和结合型代谢物的分离和鉴定方法。用分辨率高的毛细管气相色谱分离,以灵敏度高专属性好的氮磷检测器检测志愿者24 h尿样中游离型母体药及代谢物。选择部分时间收集尿样进行直接气质联用分析以及采用三氟醋酰化、三甲基硅烷化和两者并用的衍生化方法鉴定母体药及游离型代谢物。尿样酸水解后,再进行以上选择性衍生化,可测定它们的结合型代谢物。     

In vivo biotransformation of metoprolol in the horse and on-column esterification of the aminocarboxylic acid metabolite by alcohols during solid phase extraction using mixed mode columns

The in vivo biotransformation of metoprolol tartrate in the thoroughbred racehorse was studied after administration of a single oral dose. Metoprolol and its basic and bifunctional phase I metabolites were isolated from urine and plasma using mixed mode solid phase extraction (SPE) cartridges. The isolates were derivatised as trimethylsilyl ethers and analysed by capillary column gas chromatography--positive ion electron ionisation and ammonia chemical ionisation mass spectrometry. Metabolism was primarily confined to the oxidative transformations of the p-(2-methoxy)ethyl substituent. Metoprolol and five phase I metabolites were detected in horse urine. In common with man, rat and dog, the zwitterionic compound (+/-)-4-(2-hydroxy-3-isopropylaminopropoxy)-phenylacetic acid (H117/04), was the principle metabolite in the horse. This compound was readily isolated from both plasma and urine samples by SPE and, in addition, an unusual on-column esterification of the carboxylic acid moiety by alcohols was observed. Metoprolol and the major aliphatic acid metabolite were detected for about 10 and 40 h, respectively in unhydrolysed urine. After enzymatic hydrolysis, the detection period increased to 15 and 60 h, respectively indicating some phase II metabolism of metoprolol and its metabolites in the horse.     

Comparative metabolism of codeine in man, rat, dog, guinea-pig and rabbit: identification of four new metabolites.

The metabolism and excretion of codeine and its metabolites in untreated urine of man, rat, dog, guinea-pig and rabbit have been examined. Metabolites were identified by gas chromatography mass spectrometry operated in the chemical ionization mode (methane). Concentrations of codeine and metabolites were measured by selected ion monitoring. Both codeine and norcodeine were detected in the urine of all species but a new metabolite, hydrocodone, was found only in the urine from man, guinea-pig and dog. Additional metabolites (presumably resulting from the metabolism of hydrocodone) were also detected in man and guinea-pig. Overall recoveries of drug and metabolites from untreated urine were low for all species.     

Comparative metabolism of codeine in man,rat, dog,guinea-pig and rabbit: identification of four new metabolites

The metabolism and excretion of codeine and its metabolites in untreated urine of man, rat, dog, guinea-pig and rabbit have been examined. Metabolites were identified by gas chromatography mass spectrometry operated in the chemical ionization mode (methane). Concentrations of codeine and metabolites were measured by selected ion monitoring. Both codeine and norcodeine were detected in the urine of all species but a new metabolite, hydrocodone, was found only in the urine from man, guinea-pig and dog. Additional metabolites (presumably resulting from the metabolism of hydrocodone) were also detected in man and guinea-pig. Overall recoveries of drug and metabolites from untreated urine were low for all species.     

Determination of growth hormone releasing peptides metabolites in human urine after nasal administration of GHRP‐1, GHRP‐2, GHRP‐6, Hexarelin,and Ipamorelin

Growth hormone releasing peptides (GHRPs) stimulate secretion of endogenous growth hormone and are listed on the World Anti‐Doping Agency (WADA) Prohibited List. To develop an effective method for GHRPs anti‐doping control we have investigated metabolites of GHRP‐1, GHRP‐2, GHRP‐6, Hexarelin, and Ipamorelin in urine after nasal administration. Each compound was administrated to one volunteer. Samples were collected for 2 days after administration, processed by solid‐phase extraction on weak cation exchange cartridges and analyzed by means of nano‐liquid chromatography ‐ high resolution mass spectrometry. Six metabolites of GHRP‐1 were identified. GHRP‐1 in the parent form was not detected. GHRP‐1 (2‐4) free acid was detected in urine up to 27 h. GHRP‐2, GHRP‐2 free acid and GHRP‐2 (1‐3) free acid were detected in urine up to 47 h after administration. GHRP‐6 was mostly excreted unchanged and detected in urine 23 h after administration, its metabolites were detectable for 12 h only. Hexarelin and Ipamorelin metabolized intensively and were excreted as a set of parent compounds with metabolites. Hexarelin (1‐3) free acid and Ipamorelin (1‐4) free acid were detected in urine samples after complete withdrawal of parent substances. GHRPs and their most prominent metabolites were included into routine ultra‐pressure liquid chromatography‐tandem mass spectrometry procedure. The method was fully validated, calibration curves of targeted analytes were obtained and excretion curves of GHRPs and their metabolites were plotted. Our results confirm that the detection window after GHRPs administration depends on individual metabolism, drug preparation form and the way of administration. Copyright © 2015 John Wiley & Sons, Ltd.     

Sulfate metabolites as alternative markers for the detection of 4‐chlorometandienone misuse in doping control

Sulfate metabolites have been described as long‐term metabolites for some anabolic androgenic steroids (AAS). 4‐chlorometandienone (4Cl‐MTD) is one of the most frequently detected AAS in sports drug testing and it is commonly detected by monitoring metabolites excreted free or conjugated with glucuronic acid. Sulfation reactions of 4Cl‐MTD have not been studied. The aim of this work was to evaluate the sulfate fraction of 4Cl‐MTD metabolism by liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) to establish potential long‐term metabolites valuable for doping control purposes. 4Cl‐MTD was administered to two healthy male volunteers and urine samples were collected up to 8 days after administration. A theoretical selected reaction monitoring (SRM) method working in negative mode was developed. Ion transitions were based on ionization and fragmentation behaviour of sulfate metabolites as well as specific neutral losses (NL of 15 Da and NL of 36 Da) of compounds with related chemical structure. Six sulfate metabolites were detected after the analysis of excretion study samples. Three of the identified metabolites were characterized by liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) and gas chromatography‐tandem mass spectrometry (GC‐MS/MS). Results showed that five out of the six identified sulfate metabolites were detected in urine up to the last collected samples from both excretion studies. Copyright © 2016 John Wiley & Sons, Ltd.     

Metabolism of chlormezanone in man

The metabolism of chlormezanone (Muskel Trancopal) in man was studied by the aid of gas chromatography/mass spectrometry and high performance liquid chromatography after an oral dose of 400 mg. Six metabolites and/or degradation products were identified in the urine. Some of the metabolites are formed at least partially by nonenzymatic hydrolysis in the stomach. In contrast to previous publications, no unchanged drug was detected in plasma and urine. The main metabolite in plasma is generated by cleavage of the amide bond in the six-membered heterocyclic ring. This derivative is easily formed by in vitro hydrolysis at pH 1, too. It structurally resembles baclofene. About 40% of the dose is excreted with the urine. The major metabolite in urine is 4-chlorohippuric acid. Additionally, 4-chloro-benzoyl-N-methylamide, 4-chlorobenzoic acid, N-methylimino-4-chlorobenzaldehyde, 4-chlorobenzaldehyde, and "hydrolized" chlormezanone were identified.     

Bioformation of boldenone and related precursors/metabolites in equine feces and urine,with relevance to doping control

Boldenone (1‐dehydrotestosterone) is an exogenous anabolic‐androgenic steroid (AAS) but is also known to be endogenous in the entire male horse and potentially formed by microbes in voided urine, the gastrointestinal tract, or feed resulting in its detection in urine samples. In this study, equine fecal and urine samples were incubated in the presence of selected stable isotope labeled AAS precursors to investigate whether microbial activity could result in 1‐dehydrogenation, in particular the formation of boldenone. Fecal matter was initially selected for investigation because of its high microbial activity, which could help to identify potential 1‐dehydrogenated biomarkers that might also be present in low quantities in urine. Fecal incubations displayed Δ1‐dehydrogenase activity, as evidenced by the use of isotope labeled precursors to show the formation of boldenone and boldione from testosterone and androstenedione, as well as the formation of Δ1‐progesterone and boldione from progesterone. Unlabeled forms were also produced in unspiked fecal samples with Δ1‐progesterone being identified for the first time. Subsequent incubation of urine samples with the labeled AAS precursors demonstrated that Δ1‐dehydrogenase activity can also occur in this matrix. In all urine samples where labeled boldenone or boldione were detected, labeled Δ1‐progesterone was also detected. Δ1‐progesterone was not detected any non‐incubated urine samples or following an administration of boldenone undecylenate to one mare/filly. Δ1‐progesterone appears to be a candidate for further investigation as a suitable biomarker to help evaluate whether boldenone present in a urine sample may have arisen due to microbial activity rather than by its exogenous administration.     

Etodolac in equine urine and serum: determination by high-performance liquid chromatography with ultraviolet detection, confirmation, and metabolite identification by atmospheric pressure ionization mass spectrometry.

A high-performance liquid chromatographic method was used for the detection of etodolac in equine serum and urine. The method consisted of a one-step liquid-liquid extraction, separation on a reversed-phase (RP-18) column and detection using an ultraviolet detector. Additional confirmation methods included a HPLC coupled with an atmospheric pressure chemical ionization mass spectrometer (APCI-MS). Free (unbound) etodolac and its conjugates were present in the samples. Concentrations of the drug in the serum and urine samples collected from four standardbred mares after a single oral administration of Ultradol were determined. Maximum etodolac concentrations of 712, 716, 568, and 767 microg/mL in urine and 4.1, 3.6, 3.1, and 2.2 microg/mL in serum were observed. The peak concentrations of the drug were detected 2-10 h (urine) and 40 min-6 h (serum) after administration to four horses. The maximum detection time was 79 h in urine and 48 h in serum after the drug administration. The drug-elimination profiles for both urine and serum are presented and discussed. Method ruggedness and precision and stability studies of etodolac in serum and urine are presented. Three major metabolites were detected in the urine by liquid chromatography-APCI-MS. All three metabolites were identified as monohydroxylated etodolac.     

Liquid chromatography electrospray ionization tandem mass spectrometry for the detection of mesocarb abuse in horse doping

A method is described for the determination of mesocarb abuse in equestrian sport by combining gradient liquid chromatography and electrospray ionization tandem mass spectrometry. Mesocarb was administrated orally to two horses at a dose of 50 µg/kg. Urine samples were collected up to 120 h post administration. Hydrolyzed and conjugated urine fractions were handled using liquid‐liquid extraction (LLE). The identity of the parent drug and metabolites was confirmed using liquid chromatography combined with tandem mass spectrometry (MS/MS). Mesocarb and seven metabolites were detected in horse urine. Mono‐ and two di‐hydroxylated metabolites were the main metabolites observed in horse urine samples. Based on the differences in MS/MS spectra it was supposed that these metabolites were been formed by the hydroxylation of the phenylisopropyl moiety of mesocarb whilst the main process of hydroxylation of mesocarb in human occurred in the phenylcarbamoyl moiety. The main metabolites were almost completely glucuroconjugated. Minor metabolites such as p‐hydroxymesocarb and three di‐hydroxylated metabolites together with parent mesocarb were also presented in the free urine fraction. This study has shown that two mono‐ and two di‐hydroxylated metabolites are useful for controlling the abuse of mesocarb in horses. Copyright © 2011 John Wiley & Sons, Ltd.     

Study of the metabolism of the new anti-ulcer drug quiditene [(3-quinuclidyl)-di(2-thienyl)carbinol]

Thin-layer chromatography and mass spectrometry were used to study the metabolism of the new anti-ulcer drug Quiditene. After oral administration in a dose of 100 mg/kg, urine samples taken 2 h later showed no metabolites. Later samples (taken 4, 6, 8, and 10 h after administration) revealed three spots belonging to its metabolites in the chromatograms, in addition to the drug itself. Still later samples (taken 24 and 30 h after administration) revealed negligible amounts of the drug and no metabolites. The greatest amounts of metabolites were found in the urine in 6 and 8 h after administration of the drug. The structure of the metabolites was (3-quinuclidyl)-2-thienyl)ketone and its N-oxide. They were synthsized by alternate route to establish their identity and tested for anti-ulcer activity.