Liquid Chromatography-electrospray Ionization Tandem Mass Spectrometry for Simultaneous Determination of Metformin and Glimepiride in Beagle Dog Plasma and Bioequivalence Study

A sensitive and selective liquid chromatography-electrospray ionization tandem mass spectrometry（LC- ES1-MS/MS） was used for the simultaneous determination of metformin and glimepiride in beagle dog plasma with glipizide as internal standard（IS）. After simplified protein precipitation with methanol, both the analytes and IS were chromatographed on a Zorbax CN column via gradient elution with methanol（containing 5 mmol/L ammonium ace- tate） and 5 mmol/L aqueous ammonium acetate as the mobile phase. Detection was performed by multiple reaction monitoring（MRM） scanning via ESI source operated in positive ionization mode. Specificity, linearity, accuracy, pre- cision, recovery, matrix effect and stability were validated for metformin and glimepiride in beagle dog plasma. The calibration curves were linear in a concentration range of 10-10000 ng/mL for metformin and 4-4000 ng/mL for glimepiride with both correlation coefficients higher than 0.99. The recoveries obtained for the analytes and IS were all between 82.7% and 101.2%. The method exhibited excellent performance in terms of selectivity, robustness, short analytical time and simplicity of sample preparation. Finally, the proposed method was applied to a bioequivalence study of self-made bilayer tablet and commercial formulation containing 500 mg of metformin and 1 mg of glimepi- ride in beagle dogs.

High Performance Liquid Chromatography-Electrospray Ionization-Mass Spectrometric Analysis of Bilobalide and Ginkgolides in Ginkgo biloba L. Leaves

The ginkgo terpenoids including bilobalide and ginkgolides are the main pharmaceutical components in the leaves or extracts of Ginkgo biloba L. In this paper, the analysis of bilobalide and ginkgolides in leaves of Ginkgo biloba L. by high performance liquid chromatography (HPLC)-electrospray ionization (ESI)-mass spectrometry (MS) was carried out. The separation was performed on Inertsil ODS3 column with methanol-water (36:64) as mobile phase, with 1 mL·min -1 of flow rate at 35℃. Then the mass spectrum analysis was conducted by ZMD micromass electrospray ionization (ESI)-mass spectrometer (MS). The HPLC total ion chromatogram and selected ion chromatogram (with 325, 407, 423, 439 of m/z) of the sample and ESI-/MS mass spectra of the peaks in the chromatograms were obtained. So bilobalide, ginkgolide A, B, C and J in Ginkgo biloba L. leaves were identified. The method is easy and rapid, with a good accuracy.

Simultaneous determination of seven flavonoids in Epimedium by liquid chromatography-tandem mass spectrometry method

In this paper, a sensitive and specific liquid chromatography-electrospray ionisation-mass spectrometry （LC-ESI-MS） method has been developed and validated for the identification and determination of seven flavonoids, namely epimedin A, epimedin B, epimedin C, icariin, sagittatoside B, 2＂-O-rhamnosyl icariside II, and baohuoside I in Epimedium from different sources.

Probing interaction of melittin with lipid membranes using liquid secondary ion mass spectrometry

The membrane insertion mechanism of toxic protein is a very active domain in the study of molecular biology. and the “anchor state” of the membrane-hund protein on membrane is the key problem which it is difficult to solve with traditional methotls. In the present work we first studied the “anchor state” of melittin on membrane using liquid secondary ion mass spectrometry (LSIMS) in combination with proteolysis by specific enzyme. The results show that the membrane-bound melittin molecules mainly take the conformation in which the axis of α-helix lies parallel to the membrane surface and the side containing Lys7, Lys2I and Arg22 faces the outside of the lipid membrane. This discovery is very significant to the studies of membrane insertion mechanism. The results also indicate that the combination of mass spectrometry technique with the proteolysis by specific enzyme has provided a very new and effective method for the studies of the membrane insertion mechanism.

Liquid chromatography electrospray ionization tandem mass spectrometry(LC/ESI-MS/MS) method for quantitative estimation of moxifloxacin in human plasma

A rapid and sensitive liquid chromatography tandem mass spectrometry(LC-MS/MS) method has been developed and validated for the determination of moxifloxacin(MOXI) in human plasma. After a simple protein precipitation using acetonitrile, the post treatment samples were analysed on a C18 column interfaced with a Triple Quadropole Tandem Mass Spectrometer. Positive electrospray ionization was employed as the ionization source. The mobile phase consisted of 0.1% formic acid–acetonitrile(60:40, v/v). Ciprofloxacin(CIPRO) was used as an internal standard. The analyte and internal standard(CIPRO) were monitored in the multiple reaction monitoring mode(MRM). The mass transition ion-pair has been followed as m/z 402→358.2 for MOXI and 332→288.1 for CIPRO. The method was linear in the concentration range of 25–5000 ng/mL. The lower limit of quantification was 25 ng/mL. The intra- and inter-day precision(relative standard deviation) and accuracy(relative error) values were within 12.4%. Each plasma sample was analyzed within 3 min.

Probing the Proteolysis of Melittin Using Liquid Secondary Ion Mass Spectrometry

Liquid secondary ion mass spectrometry(LSIMS) was used to probe the proteolysis of melittin by trypsin and pepsin. The results showed that LSIMS is good for monitoring proteolytic reactions. It can not only identify the proteolytic products of proteins, but also be used to research the dynamics of proteolytic reactions. The proteolysis of melittin by trypsin gave seven main peptide fragments: 1, AA 1 AA 7 ; 2, AA 1 AA 21 ;3, AA 1 AA 22 ; 4, AA 1 AA 23 ; 5, AA 1 AA 24 ; 6, AA 8 AA 21 ; 7, AA 8 AA 22 . The proteolysis of melittin with pepsin gave thirteen peptide fragments: 1, AA 1 AA 3; 2, AA 4 AA 6;3, AA 1 AA 4;4, AA 1 AA 5 ;5, AA 1 AA 6 ;6, AA 1 AA 8;7, AA 7 AA 13 ;8, AA 10 AA 13 ;9, AA 14 AA 16 ;10, AA 14 AA 26 ;11, AA 15 AA 26 ;12, AA 17 AA 26 ;13, AA 20 AA 26 .

Quantitative determination of ilexgenin A in rat plasma by liquid chromatography coupled with mass spectrometry and its pharmacokinetics

A sensitive and selective high performance liquid chromatography coupled with electrospray ionization mass spectrometry （LC-MS） was developed for the quantitative determination of ilexgenin A （IA）,a major component in Radix Ilicis Pubescentis,in rat plasma.Chromatographic separation was performed on a C 18 column,with methanol-5 mM ammonium acetate （80：20,v/v） as the mobile phase.Mass spectrometer was set in negative mode with target ions at m/z 501.1→501.1 for IA and m/z 779.4→779.4 for digoxin （internal standard,IS）.Rat plasma was extracted with ethyl acetate after addition of phosphoric solution and the organic layer was evaporated and reconstituted with mobile phase for LC-MS analysis.The proposed method was validated with a linear range of 1.05-525.5 ng/mL for IA with limit of quantitation （LOQ） at 1.05 ng/mL.Intra-and inter-day precision expressed as relative standard deviation （RSD） were less than 10% at LOQ level and overall recovery was over 80%.This validated method was used successfully for the pharmacokinetic study of IA in rats after oral dosing of IA （100 mg/kg） and some main pharmacokinetic parameters of IA in rats were obtained.

湖南特色刺葡萄果皮中优势次生代谢物质分析

以‘夏黑’葡萄和‘阳光玫瑰’葡萄为对照,利用超高效液相色谱质谱联用技术广泛靶向代谢组学的方法,对果皮中的次生代谢产物进行组学分析,结合统计学方法筛选出湖南省3个刺葡萄品种果皮中的差异代谢物质。结果表明:本试验共检测到737种代谢物质,其中‘湘刺1号’检测出的代谢物质有711种,‘湘刺2号’检出666种,‘湘刺4号’检出710种;不同刺葡萄品种果皮的代谢物质有较大差异,主要表现为类黄酮化合物含量的差异,其中‘湘刺2号’的原花青素相对含量较高,‘湘刺4号’的花色苷和类黄酮物质相对含量较高,‘湘刺1号’果皮中含有较多的槲皮素-3-O-桑布双糖苷、丁香亭-3-O-(6″-乙酰)葡萄糖苷、柠檬素-3-O-葡萄糖苷、芍药花素-3-O-芸香糖苷和锦葵色素-3-O-(6″-O-乙酰)葡萄糖苷-5-O-葡萄糖苷,且香豆酰阿魏酰酒石酸和紫丁香苷仅在‘湘刺1’号中检出。

基于质谱的单细胞分析技术研究新进展

细胞是生物体最基本的结构和功能单位。近年来,人们对细胞异质性的关注越来越多。为了分析细胞的这种异质性,避免单细胞信息被群体细胞的平均值所淹没,单细胞分析技术不断发展起来。由于单细胞的体积小,内含物质含量低且种类繁多,某些物质在细胞内的变化迅速,分析过程中不同物质之间相互干扰,造成了单细胞分析极具挑战性。基于质谱的单细胞分析技术具有通用性强、无需标记、高灵敏度、高分辨率、高选择性的优点,已逐渐在单细胞分析技术中脱颖而出,成为单细胞分析的理想工具。本文系统地总结了近5年来基于质谱的单细胞分析技术的最新进展,包括电喷雾电离质谱、激光解吸电离质谱、二次离子质谱、电感耦合等离子体质谱,并对未来的研究和技术方向进行展望,希望为单细胞质谱分析技术的研究提供参考。

青刺果ACE抑制肽的分离纯化、结构鉴定及其体外活性评价

以具有血管紧张素转化酶(ACE)抑制活性的青刺果蛋白酶解物为研究对象,采用超滤、强阴离子交换层析分离纯化ACE抑制肽,液相色谱-串联质谱鉴定其肽序列。采用傅里叶红外光谱、酶反应抑制剂动力学、MTT试验和分子对接技术解析其二级结构特征、体外活性以及与ACE的结合机制。结果表明,分子质量<3 ku的超滤组分活性较好(IC50=0.380 mg/mL),离子交换层析后以F-a组分活性最好(IC50=0.159 mg/mL)。质谱鉴定出4条肽序列,通过生物信息学分析确定肽PGDVF为潜在的ACE抑制肽[IC50=(0.56±0.1)mmol/L]。二级结构分析表明PGDVF由α-螺旋(20.28%)、β-折叠(6.21%)、β-转角(31.55%)和无规则卷曲(41.85%)构成,其抑制模型为非竞争性抑制,肽质量浓度小于1 mg/mL时,对HepG2细胞无毒性。分子对接显示PGDVF可通过氢键、疏水作用与ACE紧密结合,从而有效抑制ACE活性。研究结果可为青刺果降压肽的开发利用提供参考。

基于广泛靶向代谢组学的主产区素心蜡梅代谢产物比较分析

[目的]探讨主要产区素心蜡梅次生代谢物组分,通过筛选和分析河南鄢陵和重庆北碚素心蜡梅的差异次生代谢物及其差异代谢途径,为蜡梅的道地产区的科学种植提供理论依据。[方法]采用基于超高效液相色谱串联质谱的广泛靶向代谢组学,对两个产地盛花期的样本进行代谢组分析[结果]1)两个产地的样本中共鉴定出2175种次生代谢物,按化学结构主要分为13类,其中黄酮类物质种类占比最大,共387种,其次是酚酸类、脂质、生物碱、萜类、氨基酸及其衍生物、木脂素和香豆素、有机酸、核苷酸及其衍生物、醌类、鞣质和甾体。2)相比于重庆素心蜡梅,河南素心蜡梅共有613种显著差异代谢物,其中346种代谢物的含量显著上调,包括79种酚酸类,52种黄酮,38种生物碱,32种有机酸,27种氨基酸及其衍生物,24种萜类,8种脂质,7种醌类,7种核苷酸及其衍生物和38种其他类。3)根据KEGG注释,129种差异代谢物被注释至83条代谢途径。其中,苯丙烷生物合成、黄酮类生物合成、氨基酸生物合成、ABC转运体等通路的差异代谢物富集较多,苯丙烷的生物合成、苯丙氨酸代谢、二萜生物合成、黄酮类化合物生物合成、泛醌和其他萜类-醌生物合成5条代谢通路富集显著。[结论]两个主产区素心蜡梅中共检测到2175种次生代谢物,不同产区素心蜡梅在次生代谢物质方面表现出显著的差异性,包括613种显著差异代谢物,主要集中在酚酸类和黄酮类。其中,杜仲脂素A和七叶树碱是区分两个主产区素心蜡梅的关键标志物。苯丙烷生物合成、黄酮类生物合成、氨基酸生物合成、ABC转运体等通路的差异代谢物较多,苯丙烷的生物合成、苯丙氨酸代谢和二萜生物合成的差异代谢物富集显著。

基于原位液相飞行时间二次离子质谱的电化学固液界面研究

电化学界面是能源转换与存储、生物化学、传感器和腐蚀等领域的核心,研究电化学界面结构与性能的关系一直是电化学和化学测量学的热点与难点。本文重点介绍了基于高真空兼容微流控装置的原位液相飞行时间二次离子质谱(ToF-SIMS)技术的原理、特性,及其在电化学固液界面实时、原位探测方面的研究进展。随着微流控装置的进一步创新以及ToF-SIMS仪器的不断发展,原位液相ToF-SIMS将为电催化、电池等领域的复杂电化学固液界面研究提供重要手段。

车用气雾剂中VOCs检测方法的研究

为准确测量气雾剂产品中挥发性有机化合物(VOCs)的成分及含量,设计了特定装置将气雾剂中推进剂和非推进剂进行分离,利用采集袋收集气雾剂中的推进剂,通过液液萃取法对非推进剂中的VOCs进行提取。以所建立的气相色谱-质谱/氢火焰离子化检测器(GC-MS/FID)联用方法对样品进行分析,其中推进剂气体组分含量用归一化法进行测定,非推进剂的VOCs依据检出种类,选择8种VOCs建立标准曲线。各物质在对应质量浓度范围呈良好的线性关系,相关系数(r^(2))均大于0.99;检出限和定量下限分别为0.25~0.50 mg/L和0.50~1.00 mg/L;回收率为90.5%~105%,相对标准偏差(RSD)为1.8%~9.3%。应用建立的方法分析6种不同功能的气雾剂产品,并计算其臭氧生成潜势。结果表明,推进剂部分主要检出丙烷、丁烷、二甲醚,非推进剂部分检出苯系物、烷烃、环烷烃、单萜类、羧酸衍生物、醚类,其中苯系物和单萜类含量较高。该方法可为企业改进产品配方提供参考,也可为监管部门提供技术支持。

超高效液相色谱-串联质谱-电喷雾离子源测定柑橘中灭螨醌和羟基灭螨醌残留

目的建立超高效液相色谱-串联质谱-电喷雾离子源(ultra performance liquid chromatography-tandem mass spectrometry-electrospray ionization,UPLC-MS/MS-ESI)检测柑橘中灭螨醌及其代谢物羟基灭螨醌残留的方法。方法样品经0.5%甲酸乙腈提取,氟罗里硅土吸附剂和无水硫酸镁净化,以0.1%甲酸溶液和甲醇为流动相进行梯度洗脱,经ACQUITYUPLC BEH C_(18)(100 mm×2.1 mm,1.7µm)色谱柱分离。采用电喷雾正离子模式,多重反应监测(multiple reaction monitoring,MRM)模式进行检测,基质匹配外标法定量。结果灭螨醌及羟基灭螨醌在0.0025~0.0500 mg/L质量浓度范围内线性关系良好,相关系数大于等于0.9984。在0.005~0.500 mg/kg加标水平下,灭螨醌及羟基灭螨醌在柑橘全果和果肉中的平均回收率为70%~93%,最大相对标准偏差(relative standard deviation,RSD)为12.0%,方法定量限(limit of quantitation,LOQ)为0.005 mg/kg。结论基于改良QuEChERS法,采用UPLC-MS/MS-ESI检测,该方法更方便高效且准确度、精密度和灵敏度均较好,能够满足柑橘中灭螨醌和羟基灭螨醌的残留分析要求。

超高效液相色谱质谱法测定饼干中3种大麻酚

使用超高效液相色谱-串联质谱法,建立饼干中大麻酚、大麻二酚和四氢大麻酚的测定方法。样品经乙腈醋酸(99+1)提取,加入醋酸体系盐包超声离心后,再加入吸附剂离心。采用0.1%甲酸水溶液和甲醇为流动相,梯度洗脱,通过C18色谱柱分离,大气压化学电子源(atmospheric pressure chemical ionization,APCI)正离子多反应模式监测。通过考察不同种类提取溶剂、超声时间、萃取盐包组成、吸附剂组成的目标化合物回收率,确定最优前处理方式。结果表明:大麻酚、大麻二酚、四氢大麻酚在2~100 ng/mL时呈现良好线性关系,相关系数(R2)均大于0.999,方法检出限0.01 mg/kg,定量限0.02 mg/kg。低、中、高3个浓度加标回收试验,大麻酚回收率为80.3%~91.1%,相对标准偏差(relative standard deviation,RSD)为2.4%~6.5%,大麻二酚回收率为78.4%~88.5%,RSD为2.9%~7.5%,四氢大麻酚回收率为81.5%~91.3%,RSD为2.6%~6.8%。该方法适用于饼干中大麻酚、大麻二酚和四氢大麻酚的检测。

低共熔溶剂-超高效液相色谱-大气压化学离子化-串联质谱法测定火腿肠中9种N-亚硝胺

目的建立低共熔溶剂(deep eutectic solvent,DES)-超高效液相色谱-大气压化学离子化-串联质谱法快速测定火腿肠中9种N-亚硝胺污染物的分析方法。方法样品经水稀释,微量DES分散萃取,50%甲醇水复溶,Agilent Infinitylab Poroshell 120 EC-C_(18)(2.1 mm×150 mm,2.7μm)色谱柱分离,大气压化学电离源电离,多反应监测模式检测,内标法定量分析。结果9种N-亚硝胺在5.0~60.0ng/mL范围内线性关系良好,线性相关系数(r)均大于0.999。方法检出限为0.1~0.3μg/kg,定量限为0.3~1.0μg/kg,回收率为72.1%~111.6%,相对标准偏差为0.9%~8.4%。结论低共熔溶剂对N-亚硝胺的萃取效果佳,可以解决传统操作步骤烦琐的问题。本方法有机试剂用量少、绿色环保,适用于火腿肠中9种N-亚硝胺的快速、精准检测。

皮革中19种含氯苯酚的快速测定方法研究

建立了高效液相色谱串联质谱(HPLC-MS/MS)法同时测定皮革中19种含氯苯酚。样品采用甲醇(10%乙酸铵)混合溶液超声萃取,经色谱柱ZORBAX RRHD StableBond C18(21 mm×100 mm,18μm)分离,HPLC-MS/MS测定。结果表明,19种含氯苯酚在1~50μg/L浓度范围内呈现线性(R2>099),3个不同添加水平的平均回收率在813%~1237%,精密度(n=7)为08%~97%。本方法简化了前处理过程,灵敏度高,重现性好,可用于皮革中19种含氯苯酚的快速检测。

固相萃取-高效液相色谱-串联质谱法测定有机肥料中四环素类抗生素

建立了固相萃取-高效液相色谱-串联质谱法测定有机肥料中四环素类抗生素的分析方法。样品经过Na_2EDTAMcllvaine-甲醇提取液提取,提取液用SAX串联HLB固相萃取小柱净化。采用ZORBAX Eclipse Plus C_(18)色谱柱(4.6 mm×100 mm,3.5μm)进行分离,电喷雾正离子模式多反应监测(MRM)模式定量分析。化合物在5~100μg·L^(-1)范围中具有良好的线性,线性相关系数r>0.999,检出限为9~10μg·kg^(-1),加标回收率为59.1%~70.7%。实验结果表明,该方法效率高、灵敏度高,准确度好,能够满足有机肥料中四环素类抗生素的检测需求。

固相萃取-超高效液相色谱-电喷雾串联质谱法同时测定地表水中16种全氟有机化合物

建立了固相萃取-超高效液相色谱-电喷雾串联三重四极杆质谱联用技术分析水中16种全氟有机化合物的高通量检测方法。样品经 WAX 固相萃取柱富集和净化后，采用 Waters ACQUITY UPLCTM BEH C18色谱柱，含2 mmol/L乙酸铵的甲醇和含2 mmol/L乙酸铵的水作为流动相进行梯度洗脱，质谱采用电喷雾负离子电离，多反应监测模式检测。16种全氟有机化合物在0.5~100μg/L或1.0~100μg/L 浓度范围内线性关系良好，相关系数为0.9987~0.9999，方法的检出限为0.06~0.46 ng/L；高、中、低3个添加水平的回收率为67.6％~103％，相对标准偏差为2.94％~12.0％。结果表明，该方法灵敏、准确且检测范围广，分析速度快，是一种适于实际水样中全氟有机化合物检测分析的方法。

液相色谱-电喷雾串联质谱法测定食品中的伏马菌素

通过固相萃取富集伏马菌素,采用液相色谱-电喷雾串联质谱法测定玉米和芦笋中的伏马菌素B1(FB1)、伏马菌素B2(FB2).在最佳色谱条件下,伏马菌素B1、伏马菌素B2的保留时间分别为5.2 min 和8.8 min,样品空白无干扰.离子化技术采用电喷雾正离子方式,以多反应监测(MRM)方式进行检测.该法检出限为80 pg;线性定量范围为0.01～10mg/L;回收率为78.3%～104.9%;相对标准偏差为2.2%～14.2%.利用ESI/MS/MS对伏马菌素B1、伏马菌素B2进行了质谱解析,分别选择特征离子峰m/z 352、528和m/z 336、512作为伏马菌素B1、伏马菌素B2准确定性的依据.方法灵敏度高,选择性好,准确度高,可为食品中伏马菌素的风险评估提供灵敏、准确的分析方法.