基于LC−MS的樟叶越桔愈伤组织次生代谢成分分析

以樟叶越桔健康愈伤组织为研究对象,采用LC−MS联用技术,对樟叶越桔愈伤组织的甲醇提取物进行分析。结果表明:正离子模式下共检测出14类共281个化合物,以萜类、生物碱类、有机酸、杂环化合物、糖苷类和醇类为主,化合物数量分别为58、41、28、24、22个和13个,相对含量分别为16.866%、13.711%、6.108%、10.138%、12.488%和17.513%;负离子模式下检测出13类共71个化合物,以生物碱类、有机酸、糖苷类、萜类和酯类为主,化合物数量分别为14、9、9、7个和7个,相对含量分别为5.839%、72.653%、3.423%、2.062%和2.735%,其中丙酮酸的相对含量高达47.474%。研究结果可为进一步开展樟叶越桔愈伤组织中主要化学成分的生产调控等研究提供参考。

Fast and sensitive LC–MS/MS method for the simultaneous determination of lisinopril and hydrochlorothiazide in human plasma

A sensitive and rapid liquid chromatography-tandem mass spectrometry(LC–MS/MS) method has been developed for the simultaneous determination of lisinopril(LIS) and hydrochlorothiazide(HCTZ) in human plasma using their labeled internal standards(ISs). Sample pre-treatment involved solid phase extraction on Waters Oasis HLB cartridges using 100 μL of plasma, followed by liquid chromatography on Hypersil Gold C_(18)(50 mm×3.0 mm, 5 μm) column. The analytes were eluted within 2.0 min using acetonitrile-5.0 m M ammonium formate, p H 4.5(85:15, v/v) as the mobile phase. The analytes and ISs were analyzed in the negative ionization mode and quantified using multiple reaction monitoring. The method showed excellent linearity over the concentration range of 0.50–250.0 ng/m L for both the analytes. The intra-batch and inter-batch precision(% CV) was ≤5.26% and their extraction recoveries were in the range of 96.6%–103.1%. Matrix effect evaluated in terms of IS-normalized matrix factors ranged from 0.97 to 1.03 for both the analytes. The validated method was successfully applied to determine the plasma concentration of the drugs using 10 mg lisinopril and 12.5 mg hydrochlorothiazide fixed dose formulation in 18 healthy Indian volunteers.

Detecting and Identifying in vivo Metabolites of Brodimoprim via LC/ESI-MS with Data-dependent Scanning

The present article covers a simple approach to detect and subsequently identify in vivo metabolites of brodimoprim, using high performance liquid chromatography coupled to ion trap mass spectrometer（LC/ESI-MS）, which is based on a data-dependent acquisition of isotope ions and result verified by full scan mass spectrum. The distinguished advantage of data-dependent scan is rapidness because it requires minimum sample preparation, and all the necessary data can be obtained in one chromatographic run. In addition, it is highly sensitive and selective, allowing detection of trace metabolites even in the presence of complex biomatrix. As a result, four phase-Ⅰ（M1--M4） and four Phase-Ⅱ（M5--M8） metabolites of brodimoprim were identified in urine after the oral administration of hrodimoprim to Wistar rats. Their chemical structures were proposed based on the interpretation of their CID fragmentation characterizations and the metabolic pathway was exhibited in this article.

Quantification of sibutramine and its two metabolites in human plasma by LC-ESI-MS/MS and its application in a bioequivalence study

Obesity can be considered as a chronic illness of epidemic proportion and its incidents have increased exponentially in recent years.The use of anti-obesity drugs such as sibutramine is somewhat helpful.There is a need to quantify such drugs in biological samples,which is generally quite difficult.In this report,we developed and validated a simple,sensitive and specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of sibutramine (SB) and its two metabolites N-des methyl sibutramine (DSB) and N-di desmethyl sibutramine (DDSB) in human plasma.Zorbax SBC18 (4.6 mm × 75 mm,3.5 μm,80 (A)) analytical column and 5 mM ammonium formate:acetonitrile (10∶90,v/v) mobile phase were used for chromatographic separation of SB,DSB and DDSB.Multiple reaction monitoring (MRM) in the positive mode was used to detect SB,DSB and DDSB at m/z 280.3/124.9,266.3/125.3 and 252.2/124.9,respectively.Liquid liquid extraction was used for the extraction of analytes and internal standard from human plasma.This method was validated over a linear concentration range of 10.0-10,000.0 pg/mL for SB,DSB and DDSB with correlation coefficients (r) of ≥0.9997.The drug and the two metabolites were stable in plasma samples.The validated method was successfully applied in a bioequivalence and pharmacokinetic study with human volunteers under fasting condition.

Application of an LC–MS/MS method for the analysis of amlodipine,valsartan and hydrochlorothiazide in polypill for a bioequivalence study

A sensitive and selective method has been proposed for the simultaneous determination of amlodipine(AML),valsartan(VAL) and hydrochlorothiazide(HCTZ) in human plasma by liquid chromatography–tandem mass spectrometry(LC–MS/MS). The analytes and their deuterated analogs were quantitatively extracted from100 μL human plasma by solid phase extraction on Oasis HLB cartridges. The chromatographic separation of the analytes was achieved on a Chromolith RP18 e(100 mm × 4.6 mm) analytical column within 2.5 min. The resolution factor between AML and VAL, AML and HCTZ, and VAL and HCTZ was 2.9, 1.5 and 1.4, respectively,under isocratic conditions. The method was validated over a dynamic concentration range of 0.02–20.0 ng/m L for AML, 5.00–10,000 ng/m L for VAL and 0.20–200 ng/m L for HCTZ. Ion-suppression/enhancement effects were investigated by post-column infusion technique. The mean IS-normalized matrix factors for AML, VAL and HCTZ were 0.992, 0.994 and 0.998, respectively. The intra-batch and inter-batch precision(% CV) across quality control levels was ≤ 5.56% and the recovery was in the range of 93.4%–99.6% for all the analytes. The method was successfully applied to a bioequivalence study of 5 mg AML + 160 mg VAL + 12.5 mg HCTZ tablet formulation(test and reference) in 18 healthy Indian males under fasting. The mean log-transformed ratios of C max, AUC0–120 h and AUC0-inf and their 90% CIs were within 90.2%–102.1%. The assay reproducibility was demonstrated by reanalysis of 90 incurred samples.

LC and LC–MS/MS studies for the identification and characterization of degradation products of acebutolol

The aim of the present investigation was to demonstrate an approach involving use of liquid chromatography(LC) and liquid chromatography-mass spectrometry(LC–MS) to separate, identify and characterize very small quantities of degradation products(DPs) of acebutolol without their isolation from the reaction mixtures. The drug was subjected to oxidative, hydrolytic, thermal and photolytic stress conditions as per International Conference on Harmonization(ICH) guideline Q1 A(R2). Among all the stress conditions the drug was found to be labile in hydrolytic(acidic & basic) and photolytic stress conditions, while it was stable in water-induced hydrolysis, oxidative and thermal stress conditions. A total of four degradation products were formed. A C18 column was employed for the separation of all the DPs on a gradient mode by using high-performance liquid chromatography(HPLC). All the DPs were characterized with the help of their fragmentation pattern and the masses obtained upon LC–MS/MS and MSnanalysis. All the hitherto unknown degradation products were identified as 1-(2-(2-hydroxy-3-(isopropylamino)propoxy)-5-(amino)phenyl)ethanone(DP-I), N-(4-(2-hydroxy-3-(isopropylamino)propoxy)-3-acetylphenyl)acrylamide(DP-II), 1-(2-(2-hydroxy-3-(isopropylamino)propoxy)-5-(hydroxymethylamino)phenyl)ethanone(DP-III) and 1-(6-(2-hydroxy-3-(isopropylamino)propoxy)-2,3-dihydro-2-propylbenzo[d]oxazol-5-yl)ethanone(DP-IV). Finally the in-silico carcinogenicity and hepatotoxicity predictions of the drug and all the DPs were performed by using toxicity prediction softwares viz., TOPKAT, LAZAR and Discovery Studio ADMET. The results of in-silico toxicity studies revealed that acebutolol(0.967) and DP-I(0.986) were found to be carcinogenic, while acebutolol(0.490) and DP-IV(0.437) were found to be hepatotoxic.

Precolumn derivatization LC-MS/MS method for the determination and pharmacokinetic study of glucosamine in human plasma and urine

A selective precolumn derivatization liquid chromatography–tandem mass spectrometric (LC–MS/MS) method for the determination of glucosamine in human plasma and urine has been developed and validated. Glucosamine was derivatized by o-phthalaldehyde/3-mercaptopropionic acid. Chromatographic separation was performed on a Phenomenex ODS column (150 mm 4.6 mm, 5 mm) using linear gradient elution by a mobile phase consisting of methanol (A), and an aqueous solution containing 0.2% ammonium acetate and 0.1% formic acid (B) at a flow rate of 1 mL/min. Tolterodine tartrate was used as the internal standard (IS). With protein precipitation by acetonitrile and then the simple one-step derivatization, a sensitive bio-assay was achieved with the lower limit of quantitation (LLOQ) as low as 12 ng/mL for plasma. The standard addition calibration curves suitable for clinical sample analysis showed good linearity over the range of 0.012–8.27 mg/mL in plasma and 1.80–84.1 mg/mL in urine. The fully validated method has been successfully applied to a pharmacokinetic study of compound glucosamine sulfate dispersible tablets in health Chinese volunteers receiving single oral doses at 500, 1000 and 1500 mg of glucosamine sulfate, as well as multiple oral doses of 500 mg t.i.d. for 7 consecutive days.

Identification,synthesis and characterization of an unknown process related impurity in eslicarbazepine acetate active pharmaceutical ingredient by LC/ESI-IT/MS,~1H,^(13)C and ~1H-~1H COSY NMR

A new impurity was detected during high performance liquid chromatographic （HPLC） analysis of eslicarbazepine acetate active pharmaceutical ingredient. The structure of unknown impurity was postulated based on liquid chromatography mass spectrometry using electrospray ionization and ion trap analyzer （LC/ESI-IT/MS） analysis. Proposed structure of impurity was unambiguously confirmed by synthesis followed by characterization using 1H, 13C nuclear magnetic resonance spectrometry （NMR）, 1H-1H correlation spectro-scopy （COSY） and infrared spectroscopy （IR）. Based on the spectroscopic and spectrometric data, unknown impurity was characterized as 5-carbamoyl-10,11-dihydro-5H-dibenzo[b,f]azepin-10-yl propionate.

Pharmacokinetic study of inosiplex tablets in healthy Chinese volunteers by hyphenated HPLC and tandem MS techniques

Inosiplex is a compound formulation composed of inosine and p-acetaminobenzoic acid （PABA） salt of N,N-dimethylamino-2-propanol （DIP）. This study was to investigate the clinical plasma pharmacokiuetic properties of DIP and PABA after single and multiple oral doses of inosiplex tablets in healthy Chinese volunteers. The established LC/MS/MS method for plasma DIP determination had a linear range of 0.02-10 pg/mL, and the HPLC method for plasma PABA determination had a linear range of 0.0540 pg/mL. Linear pharmacokinetic characteristics were found with single oral doses of 0.5, 1.0 and 2.0 g. No obvious accumulation effects were observed for DIP and PABA.

基于LC3的长时后置滤波器研究及其FPGA实现

基于LC3编码协议,详细探讨了长时后置滤波器(LTPF)的硬件设计与实现。研究内容包括LTPF的基本原理、硬件设计架构及其在FPGA上的实现和测试。通过使用Altera(已被英特尔收购)MAX 10开发板进行了验证,结果显示,硬件实现显著提高了处理效率,并在较低资源消耗下实现了LTPF的硬件加速功能。此外,本文还将硬件实现与STM32平台上的C语言定点程序进行了对比,展示了硬件设计在处理速度和资源利用上的优势。研究结果表明,尽管当前设计已优于软件架构,但未来通过逻辑重组或流水线技术对设计进行优化,系统性能还有提升空间。

Comparison of ESI–and APCI–LC–MS/MS methods:A case study of levonorgestrel in human plasma

Electrospray ionization(ESI) and atmospheric pressure chemical ionization(APCI) techniques for liquid chromatography–tandem mass spectrometry(LC–MS/MS) determination of levonorgestrel were evaluated.In consideration of difference in ionization mechanism,the two ionization sources were compared in terms of LC conditions,MS parameters and performance of method.The sensitivity for detection of levonorgestrel with ESI was 0.25 ng/m L which was lower than 1 ng/m L with APCI.Matrix effects were evaluated for levonorgestrel and canrenone(internal standard,IS) in human plasma,and the results showed that APCI source appeared to be slightly less liable to matrix effect than ESI source.With an overall consideration,ESI was chosen as a better ionization technique for rapid and sensitive quantification of levonorgestrel.The optimized LC–ESI–MS/MS method was validated for a linear range of 0.25–50 ng/m L with a correlation coefficient ≥0.99.The intra-and inter-batch precision and accuracy were within 11.72% and 6.58%,respectively.The application of this method was demonstrated by a bioequivalence study following a single oral administration of 1.5 mg levonorgestrel tablets in 21 Chinese healthy female volunteers.

A GLOBALLY AND SUPERLINEARLY CONVERGENT TRUST REGION METHOD FOR LC^1 OPTIMIZATION PROBLEMS

A new trust region algorithm for solving convex LC 1 optimization problem is presented.It is proved that the algorithm is globally convergent and the rate of convergence is superlinear under some reasonable assumptions.

LC-UV and LC-MS evaluation of stress degradation behavior of desvenlafaxine

The objective of current study was to develop a validated specific stability indicating reversed-phase liquid chromatographic method for the quantitative determination of desvenlafaxine in bulk sample and pharmaceutical dosage form in the presence of degradation products. Forced degradation studies were performed on bulk sample of desvenlafaxine as per ICH prescribed stress conditions using acid, base, oxidative and photolytic degradation to show the stability indicating power of the method. Significant degradation was observed under acidic stress condition and the degradation product formed was identified by LC-MS and a degradation pathway for drug has been proposed. Successful separation of drug from degradation products formed under stress conditions was achieved on a SymmetryShield column C 18 (5 mm, 250 mm 4.6 mm, i.d.) using the mobile phase consisting of a mixture of 0.2% (v/v) triethylamine in ammonium acetate (0.05 M; pH 6.5) and methanol using isocratic gradient.

Identification and characterization of related substances in EVT-401 by hyphenated LC–MS techniques

A sensitive and selective method was developed for the separation and characterization of related substances(RSs) in EVT-401 by hyphenated LC–MS techniques. Complete separation of the RSs was achieved with an Inertsil ODS-SP column(250 mm×4.6 mm, 5 μm) by linear gradient elution using a mobile phase consisting of 0.2% formic acid solution, methanol and acetonitrile. EVT-401 was found to be susceptible to acid, alkaline and oxidative stresses, while relatively stable under photolytic and thermal dry stress conditions. Fourteen RSs including six process-related substances and eight degradation products were detected and identified in EVT-401 with positive ESI high-resolution TOF-MS analysis of their parent ions and the corresponding product mass spectra elucidation, and some of them were further verified by chemical synthesis and NMR spectroscopy. The specific LC–MS method developed for separation, identification and characterization of RSs is valuable for EVT-401 manufacturing process optimization and quality control.

IDENTIFICATION OF ALTERNARIOL, ALTERNARIOL MONOMETHYL ETHER AND ZEARALENONE BY PARTICLE BEAM LC/MS

A method for simultaneously analyzing altemariol(AOH), altemariol monomethyl ether (AME) and zearalenone(ZEA) by particle beam LC/MS was established, LC separation was accompiished with a solvent system of methanol and water (80:20 v/v). The followed particle beam LC/MS analysis gave searchable spectra of AOH. AME and ZEA. Application of this technique to analysis of an alternaria culture confirmed the presence of AOH and AME.

LC无源无线局部放电传感器

为了实现气体绝缘开关设备(GIS)内部局部放电(PD)现象的高精度检测,针对现有局部放电在线监测方法存在的一些问题,如绝缘气体泄露风险、影响GIS内部电场分布、不能准确进行放电量测量等,提出了无源无线局部放电传感器。该传感器由螺旋电感构成并安装于GIS观察窗内部。位于GIS外部的读出线圈以无线方式读取传感器信号。所提出的方案具有可行性,且未引起GIS内部电场过度畸变。通过实验室中建立的测试系统,将该传感器与商用特高频传感器进行对比,所提出的传感器信号峰值约为UHF传感器的2倍,并且LC局部放电传感器精度高达1.12 pC。实验结果验证了所提出方法的有效性,并展示了LC无源无线局部放电传感器的出色性能。

Highly sensitive LC–MS/MS method to estimate doxepin and its metabolite nordoxepin in human plasma for a bioequivalence study

A selective, sensitive and rugged liquid chromatography–tandem mass spectrometry(LC–MS/MS) assay has been developed for the simultaneous determination of doxepin(Dox) and its pharmacologically active metabolite, nordoxepin(NDox) in human plasma. The analytes and their internal standards(IS)were extracted from 500 m L of human plasma by liquid-liquid extraction using methyl tert-butyl ether.Chromatographic separation was achieved on Hypurity C8 column(100 mm ? 4.6 mm, 5 mm) using a mixture of acetonitrile-methanol(95:5, v/v) and 2.0 mM ammonium formate in 93:7(v/v) ratio. Detection was accomplished by tandem mass spectrometry in the positive ionization and multiple reaction monitoring acquisition mode. The protonated precursor to product ion transitions studied for Dox, NDox,and their corresponding ISs, propranolol and desipramine, were m/z 280.1-107.0, 266.0-107.0,260.1-116.1 and 267.1-72.1, respectively. A linear dynamic range of 15.0–3900 pg/mL for Dox and 5.00–1300 pg/mL for NDox was established with mean correlation coefficient(r2) of 0.9991 and 0.9993, respectively. The extraction recovery ranged from 86.6%–90.4% and 88.0%–99.1% for Dox and NDox, respectively. The intra-batch and inter-batch precision(% CV) across quality control levels was r 8.3% for both the analytes. Stability evaluated under different storage conditions showed no evidence of degradation and the % change in stability samples compared to nominal concentration ranged from 4.7% to12.3%. The method was successfully applied to a bioequivalence study of 6 mg doxepin hydrochloride orally disintegrating tablet in 41 healthy Indian subjects under fasting and fed conditions.

A stability-indicating LC–MS/MS method for zidovudine: Identification,characterization and toxicity prediction of two major acid degradation products

Zidvovudine(AZT) is a nucleoside analogue reverse transcriptase inhibitor(NRTI), a class of anti-retroviral drug. A stability-indicating assay method for AZT was developed in line with ICH guideline. Successful separation of AZT and its degradation products was achieved by gradient elution mode on reverse phase C_(18) column using 10 mM ammonium acetate: acetonitrile as the mobile phase at 0.8 mL/min flow rate, 25 μL injection volume, 30 °C column temperature and 285 nm detection wavelength. Two major acid degradation products were identified and characterized by liquid chromatography–electrospray ionization mass spectrometry(LC–ESI/MS/MS) and accurate mass measurements. The probable mechanisms for the formation of degradation products were identified based on a comparison of the fragmentation pattern of the [M + H]^+ions of AZT and its degradation products. One of the degradation products, DP-1, was isolated by semi-preparative high performance liquid chromatography(HPLC) using Waters XBridge Prep C_(18)(250 mm×10 mm, 5 μm).Degradation products showed higher toxicity compared to the drug in some models assessed by TOPKAT software. The method validation was performed with respect to robustness, specificity, linearity, precision and accuracy as per ICH guideline Q2(R1).

Application of LC–MS/MS for quantitative analysis of glucocorticoids and stimulants in biological fuids

Liquid chromatography tandem mass chromatography （LC-MS/MS） is an important hyphenated technique for quantitative analysis of drugs in biological fluids. Because of high sensitivity and selectivity, LC-MS/MS has been used for pharmacokinetic studies, metabolites identification in the plasma and urine. This manuscript gives comprehensive analytical review, focusing on chromatographic separation approaches （column packing materials, column length and mobile phase） as well as different acquisition modes （SIM, MRM） for quantitative analysis of glucocorticoids and stimulants. This review is not meant to be exhaustive but rather to provide a general overview for detection and confirmation of target drugs using LC-MS/MS and thus useful in the doping analysis, toxicological studies as well as in pharmaceutical analysis.