Quantitative analysis of veralipride in plasma and urine by gas chromatography-mass spectrometry and gas chromatography with flame-ionization detection

A highly sensitive and selective quantitative assay for unchanged veralipride has been developed. The compound is extracted from alkalized samples (plasma or urine) with dichloromethane and converted to its trimethylated derivative by reaction with trimethylanilinium hydroxide. The reaction mixture is then chromatographed on a 3% OV-1 column. Trimethylated derivatives of plasma samples were assayed by selected-ion monitoring in the chemical-ionization mode and quantified by comparing the intensity of the quasi-molecular ion m/z 426 (M + H) with the intensity of the corresponding ion from trideuterated internal standard, m/z 429 (M + H). Flame-ionization detection was used for the assay of urine samples. The peak height ratio of trimethylated veralipride over trimethylated sulpiride, the internal standard, was used for quantitation of urine samples. A relative standard deviation of less than 10% was found when quantifying 10 ng/ml veralipride in plasma or 1 microgram/ml in urine.     

基于中空纤维液相微萃取的大鼠体内大黄素及其代谢物分析

建立了中空纤维液相微萃取(HFLPME)耦合高效液相色谱法(HPLC)用于测定血浆和尿液中大黄素及其代谢物的浓度,比较了中药有效成分大黄素在不同性别大鼠体内的吸收和代谢能力,阐述了大黄素在体内的代谢和转化过程。本实验以聚偏氟乙烯纤维为溶剂载体,正辛醇为萃取溶剂,对血浆和尿液样品进行HFLPME处理,萃取后挥干有机溶剂,用50 μL甲醇溶解,进行HPLC测定。在优化的微萃取条件下,血浆和尿液样品中大黄素及其代谢物标准曲线线性良好(相关系数(r)大于0.9960);检出限为0.1～3.0 μg/L;富集倍数为12.2～26.3;日内、日间精密度(以相对标准偏差(RSD)计)小于11.0%;血浆和尿液中代谢物的平均回收率为97.9%～103%。HFLPME操作简单,富集倍数高,能有效去除生物样品中复杂基体的干扰,适用于复杂样品中微量、痕量成分分析物的分析测定。     

High-performance liquid chromatographic analysis of amphotericin B in plasma, blood, urine and tissues for pharmacokinetic and tissue distribution studies.

A sensitive and reproducible high-performance liquid chromatographic method was developed to assay ampherotericin B in plasma, blood, urine and various tissue samples. Amphotericin B was isolated from each sample matrix by solid-phase extraction (Bond-Elut). The eluate from Bond-Elut containing amphotericin B was injected onto a reversed-phase C18 column (Waters, mu Bondpak, 10 microns, 300 mm x 3.9 mm I.D.) with a mobile phase of 45% acetonitrile in 2.5 mM Na2EDTA at 1 ml/min. Detection of amphotericin B was by ultraviolet absorption at 382 nm. Blood and tissues were homogenized and extracted with methanol prior to Bond-Elut extraction. The extraction efficiencies of amphotericin B from plasma, blood and tissues were approximately 90, 70 and 75%, respectively. The sensitivity of the assay was less than or equal to 5 ng/ml for plasma, less than or equal to 25 ng/ml for blood, 2.5 ng/ml for urine and 50 ng/g for tissues. The linearity of the assay method was up to 2.5 micrograms/ml for plasma, 5 micrograms/ml for blood, 500 ng/ml for urine and 500 micrograms/g for tissues. The assay was reproducible with an intra-day coefficient of variation (C.V., n = 3) of less than 5% in general for plasma, blood and tissues. The inter-day C.V. of the assay was less than 5% for plasma (n = 5), less than 10% for blood (n = 4) and less than 5% for tissues (n = 3). The overall variability in the urine assay was generally less than 10%. This method has demonstrated significant improvement in the sensitivity and reproducibility in assaying amphotericin B in plasma and especially in blood, urine and tissues. We have employed this assay to compare the pharmacokinetic and tissue distribution profiles of amphotericin B in rats and dogs following administration of Fungizone and ABCD (amphotericin B-cholesteryl sulfate colloidal dispersion), a lipid-based dosage form. In addition, the assay method for plasma and urine samples can also be applied to pharmacokinetics studies of amphotericin B in man.     

High-performance liquid chromatographic determination of proquazone and its m-hydroxy metabolite in spiked human plasma and urine

A simple and rapid high-performance liquid chromatographic method for the determination of proquazone (PQZ) and its major metabolite, m-hydroxyproquazone, in spiked human plasma and urine was developed. Plasma samples were purified using acetonitrile as a protein precipitant, while urine samples were diluted only with the mobile phase and filtered prior to injection. Samples containing the parent compounds and glafenine (internal standard) were eluted from a reversed-phase C8 column using acetonitrile-0.025 M sodium acetate (60 + 40) adjusted to pH 5 as the mobile phase and detected at 234 nm. Peak area ratios of the analytes versus internal standard were used for calibration. The mean recoveries from plasma and urine samples spiked with PQZ and its m-hydroxy metabolite ranged from 97.87 to 103.88%. The relative standard deviation for the within- and between-day analyses were < 4%. The proposed method was applied for the assay of PQZ in laboratory-made tablets.     

Determination of famotidine in low-volume human plasma by normal-phase liquid chromatography/tandem mass spectrometry

A rapid, sensitive and robust assay procedure using liquid chromatography coupled with tandem mass spectrometry (LC/MS/MS) for the determination of famotidine in human plasma and urine is described. Famotidine and the internal standard were isolated from plasma samples by cation-exchange solid-phase extraction with benzenesulfonic acid (SCX) cartridges. The urine assay used direct injection of a diluted urine sample. The chromatographic separation was accomplished by using a BDS Hypersil silica column with a mobile phase of acetonitrile-water containing trifluoroacetic acid. The MS/MS detection of the analytes was set in the positive ionization mode using electrospray ionization for sample introduction. The analyte and internal standard precursor-product ion combinations were monitored in the multiple-reaction monitoring mode. Assay calibration curves were linear in the concentration range 0.5--500 ng ml(-1) and 0.05--50 microg ml(-1) in plasma and urine, respectively. For the plasma assay, a 100 microl sample aliquot was subjected to extraction. To perform the urine assay, a 50 microl sample aliquot was used. The intra-day relative standard deviations at all concentration levels were <10%. The inter-day consistency was assessed by running quality control samples during each daily run. The limit of quantification was 0.5 ng ml(-1) in plasma and 0.05 microg ml(-1) in urine. The methods were utilized to support clinical pharmacokinetic studies in infants aged 0-12 months.     

Quantitative determination of the novel anticancer drug E7070 (indisulam) and its metabolite (1,4-benzenedisulphonamide) in human plasma, urine and faeces by high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry

E7070 (indisulam) is a novel anticancer drug currently undergoing clinical investigation. We present a sensitive and specific method for the quantitative determination of E7070 and its metabolite M1 (1,4-benzenedisulphonamide) in human plasma, urine and faeces. The analytes and their tetra-deuterated analogues, which were used as internal standards, were isolated from the biological matrix by solid-phase extraction with OASIS cartridges (0.5 mL plasma or 1 mL urine) and by liquid-liquid extraction with ethyl acetate at pH 5 (1 mL faecal homogenate). The analytes were separated on a C8 reversed-phase chromatographic column and analyzed using electrospray ionization and tandem mass spectrometric detection in the negative ion mode. The validated concentration ranges in plasma were 0.1-20 microg/mL for E7070 and 0.01-2 microg/mL for M1. In urine and faecal homogenate, a concentration range from 0.05-10 microg/mL or microg/g, respectively, was validated for both analytes. Validation of the plasma assay was performed according to the most recent FDA guidelines. The assay fulfilled all generally accepted requirements for linearity (r > 0.99, residuals between -8 and 10%), accuracy (-13.5 to 1.4%) and precision (all less than 11%) in the tested matrices. We investigated recovery, stability (working solutions at -20 degrees C and at room temperature, biological matrices at -20 degrees C, room temperature and after 3 freeze/thaw cycles; final extracts at room temperature) and robustness. All these parameters were found acceptable. This method is suited for mass balance studies or therapeutic drug monitoring, as demonstrated by a case example showing plasma concentrations and cumulative excretion of E7070 and M1 in urine and faeces. Furthermore, we show the presence of E7070 metabolites in patient urine.     

High-performance liquid chromatographic determination of spironolactone and its metabolites in human biological fluids after solid-phase extraction.

A simple and sensitive high-performance liquid chromatographic procedure to determine spironolactone and its three major metabolites in biological specimens is described. The assay involves sequential extraction on C18 and CN solid phases, and subsequent separation on a reversed-phase column. In plasma samples, spironolactone and its metabolites were completely separated within 8 min using an isocratic mobile phase, while in urine samples a methanol gradient was necessary to achieve a good separation within 14 min. Recoveries for all analytes were greater than 80% in plasma and 72% in urine. Linear responses were observed for all compounds in the range 6.25-400 ng/ml for plasma and 31.25-2000 ng/ml for urine. The plasma and urine methods were precise (coefficient of variation from 0.8 to 12.5%) and accurate (-12.1% to 7.4% of the nominal values) for all compounds. The assay proved to be suitable for the pharmacokinetic study of spironolactone in healthy human subjects.     

Simultaneous quantitation of d7-nefazodone, nefazodone, d7-hydroxynefazodone, hydroxynefazodone, m-chlorophenylpiperazine and triazole-dione in human plasma by liquid chromatographic-mass spectrometry

A rapid and sensitive LC-MS assay was developed and validated for the simultaneous determination of d7-nefazodone (d7-NEF), nefazodone (NEF), d7-hydroxynefazodone (d7-OH-NEF), hydroxynefazodone (OH-NEF), m-chlorophenylpiperazine (mCPP), and triazole-dione (Dione) in human plasma using trazodone (TRZ) as the internal standard (IS). A 0.1 mL aliquot of the plasma sample was precipitated with 0.1 mL of acetonitrile and vortexed for 2 min. After centrifugation, 50 microL of supernatant was mixed with 100 microL of 10 mM ammonium formate (pH = 4.0), and a 50 microL aliquot was injected onto a BDS Hypersil C18 column at a flow rate of 0.3 mL/min. The mobile phase consisting of 10 mM ammonium formate (pH = 4) and acetonitrile, 55:45 v/v, was used in an isocratic system. The mass spectrometer was programmed to admit the protonated molecules at m/z 477.2 (d7-NEF), 493.3 (d7-OH-NEF), 197.0 (mCPP), 372.0 (IS), 470.4 (NEF), 458.0 (Dione) and 486.2 (OH-NEF). Standard curves were linear (r(2) >/= 0.994) over the concentration range of 4-1000 ng/mL for Dione and 2-500 ng/mL for all other analytes. The lowest standard concentrations were the lower limits of quantitation for each analyte. The mean predicted quality control concentrations for all analytes deviated by less than 14.3% from the corresponding nominal values; the intra-assay and inter-assay precisions of the assay for all analytes were within 10.5% relative standard deviation. All analytes including the internal standard were stable in the injection solvent at room temperature for at least 24 h. The extraction recovery of the various analytes ranged from 79.2 to 109.1%. The validated assay was applied to the analysis of clinical samples obtained from a human subject who simultaneously received d7-NEF and NEF orally.     

Liquid chromatography tandem mass spectrometry method for the quantification of clonidine with LLOQ of 10 pg/mL in human plasma

A sensitive high-performance liquid chromatography-positive ion electrospray tandem mass spectrometry method was developed and validated for the quantification of clonidine in human plasma. Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reverse-phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M + H](+) ions, m/z 230 to 44 for clonidine and m/z 254 to 44 for the internal standard. The assay exhibited a linear dynamic range of 10-2000 pg/mL for clonidine in human plasma. The lower limit of quantification was 10 pg/mL with a relative standard deviation of less than 6.8%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 2.5 min for each sample made it possible to analyze more than 250 human plasma samples per day. The validated method was successfully used to analyze human plasma samples for application in pharmacokinetic studies. Copyright (c) 2008 John Wiley & Sons, Ltd.     

Trace determination of anthracyclines in urine: a new high-performance liquid chromatography/tandem mass spectrometry method for assessing exposure of hospital personnel

Health-care workers handling antineoplastic agents may be exposed to extremely low doses of these drugs. Very sensitive and specific analytical methods are therefore needed for biological monitoring. The aim of this study was to develop and validate a method for trace level determination of doxorubicin, epirubicin, daunorubicin and idarubicin in human urine, using epi-daunorubicin as an internal standard. Solid-phase extraction (SPE) was used for sample preparation. Urine samples were loaded onto Bond Elut C18 cartridges. The analytes were eluted in methylene chloride/2-propanol (1:1, v/v) and then evaporated to dryness. The residue was reconstituted with the mobile phase prior to high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) analysis. Quantitation of each analyte was performed using the multiple reaction monitoring (MRM) method. The urine assay was linear over the range 0.1-2.0 microg/L, with a lower limit of quantification (LLOQ) of 0.10 microg/L for doxorubicin and epirubicin, and 0.03 microg/L for daunorubicin and idarubicin. The respective limits of detection (LODs) were 0.04 and 0.01 microg/L. The precision and accuracy of the assay were determined on three different days. The within-series precision was found to be always less than 13.9% for all the analytes. The overall precision expressed as relative standard deviation (RSD) was always less than 10.6%. The recovery of anthracyclines was assessed at two concentrations of the range tested (0.1 and 2.0 microg/L) and it ranged from 87.7% (daunorubicin) to 102.0% (doxorubicin) and from 79.1% (daunorubicin) to 90.7% (idarubicin) for the lower and the higher level, respectively, with a RSD always less than 9.1%. The uncertainty of the present assay was also evaluated and the combined uncertainty was always less than 20% over all the days of the validation study. This is the first method that makes use of LC/MS/MS for the biological monitoring of occupational exposure to anthracyclines.     

Quantification of tizanidine in human plasma by liquid chromatography coupled to tandem mass spectrometry

A simple, sensitive and rapid high-performance liquid chromatography/positive ion electrospray tandem mass spectrometry (MS/MS) method was developed and validated for the assay of tizanidine in human plasma. Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reversed-phase column and analyzed by MS/MS in the selected reaction monitoring mode. The assay exhibited a linear dynamic range of 50-5000 pg/mL for tizanidine in human plasma. The lower limit of quantification was 50 pg/mL with a relative standard deviation of less than 13%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. A run time of 2.5 min for each sample made it possible to analyze more than 300 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies.     

Highly sensitive method for the determination of omeprazole in human plasma by liquid chromatography-electrospray ionization tandem mass spectrometry: application to a clinical pharmacokinetic study

A highly sensitive, rapid assay method has been developed and validated for the estimation of omeprazole (OPZ) in human plasma with liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the positive-ion mode. The assay procedure involves alkalinization of plasma followed by simple liquid-liquid extraction of OPZ and lansoprazole (internal standard, IS) from human plasma with acetonitrile. Chromatographic separation was achieved with 0.01 M ammonium acetate:acetonitrile (40:60, v/v) at a flow rate of 0.25 mL/min on an Inertsil ODS 3 column with a total run time 2.5 min. The MS/MS ion transitions monitored were 346.1 --> 198.1 for OPZ and 370.1 --> 252.1 for IS. Method validation and clinical sample analysis were performed as per FDA guidelines and the results met the acceptance criteria. The lower limit of quantitation achieved was 0.05 ng/mL and the linearity was observed from 0.05 to 10.0 ng/mL. The intra-day and inter-day precisions were in the ranges 2.09-8.56 and 5.29-8.19%, respectively. This novel method has been applied to a pharmacokinetic study of OPZ in humans.     

A liquid chromatography/electrospray ionization-tandem mass spectrometry method for quantification of specific organophosphorus pesticide biomarkers in human urine

Organophosphorus pesticides are commonly used in both agricultural and residential settings. The widespread use of these chemicals makes it almost impossible for humans to avoid exposure. In order to determine background human exposure, there is a need for fast, reliable, and sensitive analytical methods. We have developed a sensitive method to quantify specific biomarkers of the organophosphorus pesticides acephate, azinphos, chlorpyrifos, coumaphos, diazinon, isazofos, malathion, methamidophos, parathion and pirimiphos or their O,O-dimethyl analogues in human urine, as their selective metabolites or as the intact pesticide. Isotopically labeled internal standards were used for eight of the analytes. The use of labeled internal standards in combination with high-performance liquid chromatography electrospray ionization–tandem mass spectrometry provided a high degree of specificity. Repeated analysis of urine samples fortified with high and low concentrations of the analytes gave relative standard deviations (RSD) of less than 10% for the analytes with an isotopically labeled standard. Analytes without isotopically labeled standards had higher RSD. For all compounds except methamidophos and acephate, the recoveries were greater than 70%. The limits of quantification for most of the analytes were in the range of 0.1 to 1 ng/mL. We detected concentrations of most of these pesticides and/or their metabolites in urine samples from non-occupationally exposed persons using our method. Our frequencies of detection for the analytes measured ranged from 1% to 98%.     

LC Determination of Ketorolac in Human Plasma and Urine for Application to Pharmacokinetic Studies

A simple, specific and sensitive liquid chromatographic method has been developed for the assay of ketorolac in human plasma and urine. The clean-up of plasma and urine samples were carried out by protein precipitation procedure and liquid–liquid extraction, respectively. Separation was performed by a Waters sunfire C₁₈ reversed-phase column maintained at 35 °C. The mobile phase was a mixture of 0.02 M phosphate buffer (pH adjusted to 4.5 for plasma samples and to 3.5 for urine samples) and acetonitrile (70:30, v/v) at a flow rate of 1.0 mL min^?1. The UV detector was set at 315 nm. Nevirapine was used as an internal standard in the assay of urine sample. The method was validated over the concentration range of 0.05–8 and 0.1–10 μg mL^?1 for ketorolac in human plasma and urine, respectively. The limits of detection were 0.02 and 0.04 μg mL^?1 for plasma and urine estimation at a signal-to-noise ratio of 3. The limits of quantification were 0.05 and 0.1 μg mL^?1 for plasma and urine, respectively. The extraction recoveries were found to be 99.3 ± 4.2 and 80.3 ± 3.7% for plasma and urine, respectively. The intra-day and inter-day standard deviations were less than 0.5. The method indicated good performance in terms of specificity, linearity, detection and quantification limits, precision and accuracy. This assay demonstrated to be applicable for clinical pharmacokinetic studies.     

LC Determination of Ketorolac in Human Plasma and Urine for Application to Pharmacokinetic Studies

A simple, specific and sensitive liquid chromatographic method has been developed for the assay of ketorolac in human plasma and urine. The clean-up of plasma and urine samples were carried out by protein precipitation procedure and liquid–liquid extraction, respectively. Separation was performed by a Waters sunfire C₁₈ reversed-phase column maintained at 35 °C. The mobile phase was a mixture of 0.02 M phosphate buffer (pH adjusted to 4.5 for plasma samples and to 3.5 for urine samples) and acetonitrile (70:30, v/v) at a flow rate of 1.0 mL min⁻¹. The UV detector was set at 315 nm. Nevirapine was used as an internal standard in the assay of urine sample. The method was validated over the concentration range of 0.05–8 and 0.1–10 μg mL⁻¹ for ketorolac in human plasma and urine, respectively. The limits of detection were 0.02 and 0.04 μg mL⁻¹ for plasma and urine estimation at a signal-to-noise ratio of 3. The limits of quantification were 0.05 and 0.1 μg mL⁻¹ for plasma and urine, respectively. The extraction recoveries were found to be 99.3 ± 4.2 and 80.3 ± 3.7% for plasma and urine, respectively. The intra-day and inter-day standard deviations were less than 0.5. The method indicated good performance in terms of specificity, linearity, detection and quantification limits, precision and accuracy. This assay demonstrated to be applicable for clinical pharmacokinetic studies.

     

High-performance liquid chromatographic assay for dilevalol in human plasma and urine using a PRP-1 column and fluorimetric detection

A single high-performance liquid chromatographic (HPLC) assay for the quantitative determination of dilevalol, the R,R isomer of labetalol, was developed for both plasma and urine. A significantly improved limit of detection for dilevalol in plasma was accomplished by extensive modification of an HPLC assay originally developed in our laboratory for labetalol. This simplified method is readily adaptable to urine and represents the first reported HPLC assay for the quantitative determination of dilevalol in this biofluid. Drug was recovered from plasma or urine by partition into diethyl ether under mildly alkaline conditions and back-extraction into dilute acid. Reversed-phase separation of dilevalol and the internal standard was accomplished on a 150 X 4.1 mm column commercially packed with a spherical (5 micron) macroporous copolymer (PRP-1). No interferences were observed in extracts obtained from drug-free plasma or urine. Selectivity for dilevalol in the presence of other beta-blockers was established. This method demonstrated a linear detector response to concentrations of unchanged drug typically observed in urine and plasma following once-a-day treatment with dilevalol hydrochloride (100-800 mg). The lowest limit of reliable quantitation was established at 1 ng/ml in plasma. The intra-assay precision (coefficient of variation) remained less than 6% at all concentrations evaluated from 1 to 800 ng/ml. In urine, the lowest limit of quantitation was validated to 20 ng/ml where the intra-assay precision (coefficient of variation) for unchanged drug was less than 4% at all concentrations evaluated up to 400 ng/ml. This method is suitable for routine quantitation of unchanged drug in human plasma and urine following the administration of therapeutically effective doses of dilevalol hydrochloride.     

High-performance liquid chromatographic determination of glipizide in human plasma and urine

A sensitive and selective high-performance liquid chromatographic method for determination of intact glipizide in human plasma or urine has been developed. The plasma and urine samples were acid-buffered, before tolbutamide was added as internal standard. The samples were extracted with benzene, and the organic layer was evaporated to dryness. The residue was dissolved in equilibrated mobile phase (acetonitrile-0.01 M phosphate buffer pH 3.5, 35:65), and an aliquot of 20 microliters was chromatographed on a Spherisorb ODS reversed-phase column. Quantitation was achieved by monitoring the ultraviolet absorbance at 275 nm. The response was linear (0-1000 ng/ml) and the detection limit was 5-10 ng/ml in plasma or urine. The within-assay variation was less than or equal to 10%. No interferences from metabolites or endogenous constituents were observed. The utility of the assay was demonstrated by determining glipizide in samples from a diabetic subject receiving a therapeutic dose of 5 mg of the drug.     

超高效液相色谱-三重四极杆/复合线性离子阱质谱法同时快速测定血浆和尿液中84种有毒植物成分

采用超高效液相色谱-三重四极杆/复合线性离子阱的质谱联用技术,建立了同时快速测定血浆和尿液中84种有毒植物成分的方法。血浆样品经乙腈沉淀去蛋白和除磷脂、尿液样品经甲醇稀释后直接进样,以含0.1%（体积分数,下同）甲酸和2 mmol/L甲酸铵的97%乙腈水溶液、含0.1%甲酸的2 mmol/L甲酸铵水溶液作为流动相进行梯度洗脱,在Acquity BEH C₁₈色谱柱上实现分离,在电喷雾正离子多离子监测触发的增强子离子扫描（MRM-IDA-EPI）模式下检测,基质工作曲线内标法定量。血浆和尿液中84种待测物在相应的浓度范围内线性关系良好,相关系数均大于或等于0.9911,血浆和尿液中的检出限（S/N=3）分别为0.01~1和0.03~2 μg/L,准确度（平均加标回收率）为70.6%~124.5%,日内和日间精密度分别为0.7%~18.4%和1.1%~18.5%。该法简单、快速、灵敏、准确,可用于血浆和尿液中84种有毒植物成分的中毒检测。     

Determination of polyamine metabolome in plasma and urine by ultrahigh performance liquid chromatography–tandem mass spectrometry method: Application to identify potential markers for human hepatic cancer

To evaluate the potential relationship between cancer and polyamine metabolome, a UHPLC–MS/MS method has been developed and validated for simultaneous determination of polyamine precursors, polyamines, polyamine catabolite in human plasma and urine. Polyamine precursors including l-ornithine, lysine, l-arginine and S-adenosyl-l-methionine; polyamines including 1,3-diaminopropane, putrescine, cadaverine, spermidine, spermine, agmatine, N-acetylputrescine, N-acetylspermine and N-acetylspermidine; polyamine catabolite including γ-aminobutyric acid had been determined. The analytes were extracted from plasma and urine samples by protein precipitation procedure, and then separated on a Shim-pack XR-ODS column with 0.05% heptafluorobutyric acid (HFBA) in methanol and 0.05% HFBA in water. The detection was performed on UHPLC–MS/MS system with turbo ion spray source in the positive ion and multiple reaction-monitoring mode. The limits of quantitation for all analytes were within 0.125–31.25 ng mL⁻¹ in plasma and urine. The absolute recoveries of analytes from plasma and urine were all more than 50%. By means of the method developed, the plasma and urine samples from hepatic cancer patients and healthy age-matched volunteers had been successfully determined. Results showed that putrescine and spermidine in hepatic cancerous plasma were significant higher than those in healthy ones, while spermidine, spermine and N-acetylspermidine in hepatic cancerous urine were significant higher than those in healthy ones. The methods demonstrated the changes of polyamine metabolome occurring in plasma and urine from human subjects with hepatic cancer. It could be a powerful manner to indicate and treat hepatic cancer in its earliest indicative stages.     

Simultaneous analysis of cocaine and its metabolites in urine by capillary electrophoresis-electrospray mass spectrometry using a pressurized liquid junction nanoflow interface

A new method for the simultaneous separation of cocaine and four metabolites in urine by CE-ESI-MS via a pressurized nanoliquid junction interface was developed. The resolution of cocaine, cocaethylene, benzoylecgonine, norcocaine, and ecgonine methyl ester was achieved in a polyvinyl-alcohol-coated capillary with 75 μm id × 50 cm total length, using a 15 mM ammonium formate electrolyte solution (pH 9.5) in less than 15 min. In addition, to enhance sensitivity, a field-amplified sample injection (FASI) was evaluated in terms of injection time and sample solvent composition. The limits of detection achieved with the FASI method ranged from 1.5 to 10 ng/mL for all the compounds. The detection of the studied compounds was performed using an ion-trap mass spectrometer in a positive ionization mode. A mixture of methanol:water (80:20 v/v) containing 0.1% v/v of formic acid was employed as spray liquid and delivered at ~200 nL/min. Under optimal CE-MS conditions, linearity was assessed in the concentration range of interest for all analytes with correlation coefficients r2 ≥ 0.9913. Intra- and inter-day precision provided a relative standard deviation lower than 1.54% for migration times and lower than 12.15% for peak areas. Finally, urine samples, spiked with the standard mixture, were extracted using a solid-phase extraction procedure and injected under FASI conditions, providing recoveries from 80% to 94% for all analytes.