Rapid Determination of Dopamine and Its Metabolites During in vivo Cerebral Microdialysis by Routine High Performance Liquid Chromatography With Electrochemical Detection

To determine dopamine and its metabolites during in vivo cerebral microdialysis by routine high performance liquid chromatography with electrochemical detection. Methods Microdialysis probes were placed into the right striatum of Wistar rat brains and perfused with Ringer＇s solution at a rate of 1.5 pL/min. A reverse phase HPLC with electrochemistry was used to assay DA, DOPAC, and HVA after cerebral microdialysates were collected every 20 minutes from awake and freely moving rats. In order to identify the reliability of this method, its selectivity, linear range, precision and accuracy were tested and the contents of DA, DOPAC, and HVA in rat microdialysates were determined. Results The standard curve was in good linear at the concentration ranging from 74 nmol/L to 1.5 pmol/L for DOPAC （r^2= 0.9996）, from 66 nmol/L to 1.3 gmol/L for DA （r^2=l.0000） and from 69 nmol/L to 1.4 pmol/L for HVA （r^2=0.9992）. The recovery of DOPAC （0.30, 0.77, 1.49 gmol/L）, DA （0,26, 0.69, 1.32 gmol/L）, and HVA （0.27, 0.71, 1.37 gmol/L） was 82.00±1.70%, 104.00±4.00%, 98.70±3.10%; 92.30± 1.50%, 105.30±2.30%, 108.00±2.00%; 80.00±7.80%, 107.69±8.00%, and 108.66±3.10%, respectively at each concentration. Their intra-day RSD was 3.3%, 3.4%, and 2.5%, and inter-day RSD was 4.2%, 2.3%, and 5.6%, respectively. The mean extracellular concentrations of DOPAC, DA, and HVA in rat brain microdialysates were 10.7, 2.4, and 9.2 gmol/L （n=6）, respectively. Conclusion The findings of our study suggested that the simple, accurate and stable method can be applied to basic researches of diseases related to monoamines neurotransmitters by cerebral microdialysis in rats.

Trace Determination of Tamoxifen in Biological Fluids Using Hollow Fiber Liquid-Phase Microextraction Followed by High-Performance Liquid Chromatography-Ultraviolet Detection

The applicability of hollow fiber liquid-phase microextraction (HF-LPME) combined with high-performance liquid chromatography-ultraviolet detection (HPLC-UV) was evaluated for the extraction and determination of tamoxifen (TAM) in biological fluids including human urine and plasma. The drug was extracted from a 15 mL aqueous sample (source phase;SP) into an organic phase impregnated in the pores of the hollow fiber (membrane phase;MP) followed by the back-extraction into a second aqueous solution (receiving phase;RP) located in the lumen of the hollow fiber. The effects of several factors such as the nature of organic solvent, compositions of SP and RP solutions, extraction time, ionic strength and stirring rate on the extraction efficiency were examined and optimized. An enrichment factor of 360 along with substantial sample clean up was obtained under the optimized conditions. The calibration curve showed linearity in the range of 1 - 500 ng?mL–1 and the limit of detection was found to be 0.5 ng?mL–1 in aqueous medium. A reasonable relative recovery (≥89%) and satisfactory intra-assay (3.7% - 4.2%, n = 3) and inter-assay (7.5% - 7.8%, n = 3) precision illustrated good performance of the analytical procedure in spiked human urine and plasma samples.

Novel method for the determination of five carbamate pesticides in water samples by dispersive liquid-liquid microextraction combined with high performance liquid chromatography

A novel method for the determination of five carbamate pesticides （metolcarb, carbofuran, carbaryl, isoprocard and diethofencard） in water samples was developed by dispersive liquid-liquid microextraction （DLLME） coupled with high performance liquid chromatography-diode array detector （HPLC-DAD）. Some experimental parameters that influence the extraction efficiency were studied and optimized to obtain the best extraction results. Under the optimum conditions for the method, the calibration curve was linear in the concentration range from 5 to 1000 ng mL^-1 for all the five carbamate pesticides, with the correlation coefficients （r^2） varying from 0.9984 to 0.9994. Good enrichment factors were achieved ranging from 80 to 177- fold, depending on the compound. The limits of detection （LODs） （S/N = 3） were ranged from 0.1 to 0.5 ng mL^-1. The method has been successfully applied to the analysis of the pesticide residues in environmental water samples.

A high-performance liquid chromatography with fluorescence detection method for the simultaneous quantitation of monoamine neurotransmitters and their metabolites in subregions of rat brain

Abstract： In the presem study, we simultaneously quantified the levels of monoamine neurotransmitters （MANTs） and their metabolites （levodopa, norepinephrine, epinephrine, dopamine, 5-HT, 3,4-dihydroxyphenylacetic acid, homovanillic acid and 5-hydroxyindole-3-acetic acid） in different brain subregions of rats using a newly developed simple, sensitive and selective high-performance liquid chromatography with fluorescence detection （HPLC-FLD） method. In this new HPLC-FLD method, analytes were directly extracted and separated without deriveatization step within 20 min. The FLD wavelength was set at 280 nm and 330 nm for excitation and emission, respectively. The analytes were separated on an Agilent Eclipse Plus Cls column （4.6 mm×150 mm, 5.0 μm） equipped with an Agilent XDB-C18 security guard column （4.6 mm×12.5 mm, 5.0 lam）, and the column temperature was maintained at 35 ℃. The mobile phase for elution was isocratic. The mobile phase consisted of citric acid buffer （50 mmol/L citric acid, 50 mmol/L sodium acetate, 0.5 mmol/L octane sulfonic acid sodium salt, 0.5 mmol/L Na2EDTA and 5 mmol/L triethylamine, pH 3.8） and methanol （90：10, v/v） at a flow rate of 1.0 mL/min. The detection limit （DL） was 0.9-23 nM for all the MANTs and their metabolites with a sample volume of 50 μL. The method was shown to be highly reproducible in terms of peak area （intraday, 0.08%-1.85% RSD, n = 5）. The simultaneous measurement of these MANTs and their metabolites improved our understanding of the neurochemistry in the central nervous system （CNS） in relation to different addictive drugs （methamphetamine, heroin and their mixture） in drug-addicted rat models.

Imidazolium ionic liquid as the background ultraviolet absorption reagent for determination of morpholinium cations by high performance liquid chromatography-indirect ultraviolet detection

A novel analytical method was developed for determining morpholinium cations lacking ultraviolet absorption groups.This determination was carried out by high performance liquid chromatographyindirect ultraviolet（HPLC-1UV） detection using imidazolium ionic liquid as background absorption reagents,and imidazolium ionic liquid aq.soln.-organic solvent as mobile phase by a reversed-phase C18 column.The background ultraviolet absorption reagents,imidazolium ionic liquids and organic solvents were investigated.The imidazolium ionic liquid in the mobile phase is not only the background ultraviolet absorption reagent for IUV,but also an active component to improve the separation of morpholinium cations.It was found that morpholinium cations could be adequately determined when0.5 mmol/L 1-ethyl-3-methylimidazolium tetrafluoroborate aq.soln./methanol（80：20,v/v） was used as mobile phase with an IUV detection wavelength of 210 nm.In this study,the baseline separation of Nmethyl,ethylmorpholinium cations（MEMo） and N-methyl.propylmorpholinium cations（MPMo） was successfully achieved in 8.5 min.The detection limits（S/N = 3） for MEMo and MPMo were 0.15 and0.29 mg/L,respectively.This simple and practical method has been successfully applied to the determination of two morpholinium ionic liquids synthesized by the chemistry laboratory.

Determination of Amantadine Residue in Honey by Solid-phase Extraction and High-performance Liquid Chromatography with Pre-column Derivatization and Fluorometric Detection

Amantadine （AMA） is an anti-viral drug used in apiculture to protect honeybee against the sacbrood virus （Morator aetatulae）. This study described a reliable high-performance liquid chromatographic （HPLC） method for analyzing AMA in honey using a solid-phase extraction （SPE） cartridge （Plexa PCX） for purification, 4-fluoro-7- nitro-2,1,3-benzoxadiazole （NBD-F） as a pre-column derivatization agent, and fluorometric detection （λex =470 nm, λem=530 nm）. The chromatographic separation was performed on an XDB C18 column （150×4.6 mm i.d.） using 0.1% trifluoroacetic acid/acetonitrile （35 ; 65, V/V） as the mobile phase at a flow rate of 1.0 mLomin 1 with a run time of 20 min. Under these optimal conditions, a linear relationship was observed in the range of 0.025--1.0μg·mL-1 with a good correlation coefficient （0.998） and low limit of detection （0.0080 μg·g-1）, the recoveries were all above 90%, and the intra-day and inter-day precision （RSD） ranged from 3.4%--5.1%.

Separation and identification of moxifloxacin impurities in drug substance by high-performance liquid chromatography coupled with ultraviolet detection and Fourier transform ion cyclotron resonance mass spectrometry

In this paper, a high-performance liquid chromatography coupled with ultraviolet detection and Fourier transform-ion cyclotron resonance mass spectrometry （HPLC-UV/FrICRMS） method was described for the investigation of impurity profile in moxifloxacin （MOX） drug substance and chemical reference substance. Ten impurities were detected by HPLC-UV, while eight impurities were identified by using the high accurate molecular mass combined with multiple-stage mass spectrometric data and fragmentation rules. In addition, to our knowledge, five impurities were founded for the first time in MOX drug substance.

菊苣中酚类化合物的HPLC-ECD指纹图谱及抗氧化活性谱-效关系

建立不同产地菊苣的高效液相色谱-电化学检测法(High Performance Liquid Chromatography-Electrochemical Detection,HPLC-ECD)指纹图谱,采用DPPH、ABTS+自由基清除能力和铁离子还原能力来评价菊苣的体外抗氧化活性,并应用皮尔逊分析、灰色关联度分析和偏最小二乘法回归分析研究菊苣指纹图谱与抗氧化活性之间的“谱-效”关系,探寻菊苣的抗氧化活性成分。该研究从建立的21批不同产地的菊苣指纹图谱中,指认出13个酚类化学成分。菊苣提取物的最高总酚含量达30.76 mg GAE/g,其峰面积与抗氧化活性的相关系数大于0.92,“谱-效”关系研究表明对抗氧化活性贡献较大的有菊苣酸、单咖啡酰酒石酸、异绿原酸B、异绿原酸A、1,5-二咖啡酰奎宁酸、异绿原酸C,表明其化学成分与抗氧化活性之间具有显著的相关性(P<0.01)。该研究建立的HPLC-ECD的菊苣指纹图谱结合统计分析方法研究“谱-效”,既体现样品化学成分的差异性,又可直接反映样品的抗氧化活性,该方法简单、可靠,为菊苣的质量评价及产品开发提供参考。

高效液相色谱及其串联质谱技术在牙膏风险物质分析中的应用

牙膏是日常生活中必不可少的口腔清洁护理用品,牙膏的基料较为复杂,牙膏中潜在的风险物质对人体存在一定危害,因此牙膏产品的安全性已成为当今社会关注的焦点之一。利用现代先进的检验检测技术对牙膏中潜在的风险物质进行分析、研究,将有利于实现对牙膏产品的安全监管,本文综述了近几年来高效液相色谱及其串联质谱技术在牙膏产品检验标准和检验方法开发等方面的研究以及应用进展,一定程度上能为牙膏产品的质量控制和科学监管提供参考。

高效液相荧光色谱法同时测定六神曲中黄曲霉毒素、玉米赤霉烯酮和赭曲霉毒素A

目的建立一种用免疫亲和柱净化-柱后光化学衍生-高效液相色谱同时检测六神曲中黄曲霉毒素、玉米赤霉烯酮和赭曲霉毒素A的方法。方法样品采用60%乙腈超声提取,免疫亲和柱净化,采用XBridge^(®)Phenyl苯基色谱柱(4.6 mm×250 mm,5μm),以乙腈和0.1%磷酸溶液为流动相,梯度洗脱,光化学衍生仪衍生,通过切换荧光波长检测。结果黄曲霉毒素、玉米赤霉烯酮和赭曲霉毒素A标准曲线的线性范围分别为0.006~0.108、0.500~2.500和0.202~1.009 ng,6种毒素的线性关系在0.9994以上,检出限分别为1.2、100.0和40.3 ng/ml,平均加标回收率为73.2%~92.3%,相对偏差为2.1%~5.4%。结论该方法具有专属性强、操作方便等特点,能够有效用于六神曲中3种毒素的同时检测和安全质量控制。

UPLC-MS/MS法检测3种食品中松仁过敏原

基于食品基质中松仁过敏原Pin k 2建立了一种超高效液相色谱-串联质谱法。将松仁经过研磨、脱脂、浸提、酶解后经Easy-nLC 1000-QExactive高分辨质谱仪进行分离分析,结合Uniprot蛋白数据库以及ProteinPilotTM软件对质谱图进行数据处理,经BLAST验证特异性,最终筛选3条松仁特异性肽段。方法学验证结果表明,方法在0.001~50mg/mL范围内线性关系良好,定量限为1mg/kg;在饼干、巧克力和饮料3种空白基质中的平均回收率为88.50%~107.57%,相对标准偏差不高于6.08%,基质效应为89.77%~96.13%。该方法具有灵敏度高、特异性好的优势,可应用于饼干、巧克力、饮料等食品样品中松仁过敏原的检测,为我国食品标签真实性检验及食品中隐性过敏原的检测提供技术支持。

溶剂提取-高效液相色谱法测定大气颗粒物中18种多环芳烃

建立大气颗粒物PM2.5中18种多环芳烃的样品快速溶剂提取结合高效液相色谱的测定方法。将1/2玻璃滤膜剪碎后加入2.0 mL乙腈,于20℃水浴超声提取30 min,提取液经0.22μm有机相滤膜过滤,多环芳烃C18专用柱分离,以乙腈-水作为流动相进行梯度洗脱,采用紫外串联荧光检测器-高效液相色谱仪检测,以色谱峰面积外标法定量。18种PAHs的质量浓度在0.01~0.50μg/mL范围内与色谱峰面积线性关系良好,相关系数不小于0.999,检出限为0.03~0.47 ng/m^(3),样品加标平均回收率为75.6%~114.7%,相对标准偏差为0.09%~3.78%(n=7)。该方法简便、快速、准确、灵敏,可应用于大气颗粒物PM2.5中18种多环芳烃的测定。

九蒸九制对鸡头黄精理化性质及抗氧化性的影响

为了探究蒸制处理对鸡头黄精有效成分及代谢物种类和含量影响,采用低场核磁、苯酚-浓硫酸法、高效液相色谱-蒸发光散射检测法(HPLC-ELSD)和液相色谱-质谱技术(LC-MS)等对其中的水分分布、多糖含量、单糖组成和代谢产物等进行分析。结果表明,多次蒸制后,鸡头黄精结合水的含量、多糖含量逐渐降低,水提液pH值逐渐降低呈弱酸性达到4.07,还原糖、总酚、黄酮含量逐渐升高分别达到28.69%、10.02 mg/g和0.69%,抗氧化性逐渐增强,ABTS抗氧化能力在7制达到最高为0.73 mmol/L,较1制增加0.35 mmol/L,DPPH自由基清除率和FRAP值在8制达到最高分别为81.95%,1.97 mmol/L,较1制分别增加50.92%,1.72 mmol/L;同时蔗糖逐渐水解从18.53mg/g到7.62mg/g,葡萄糖和果糖含量提高,分别由1制0.00和11.30mg/g,达到9制17.25和230.89 mg/g。选取一制与九制黄精进行代谢物差异分析,在正离子模式下共检测到1310种代谢物,差异代谢物有176种(按其特性分为38类),在负离子模式下共检测到1841种代谢物,差异代谢物有148种(按其特性分为26类)。黄精经蒸制后有效成分差异性显著,对鸡头黄精炮制加工提供科学依据。

低盐雪菜中呈味核苷酸关联物的检测及贮藏过程中的变化

为探明低盐雪菜中呈味核苷酸关联物在贮运期间的变化规律,建立高效液相色谱检测方法,分析低盐雪菜在(25±5)℃贮藏(J-25)、(4±1)℃贮藏(J-4)、灭菌后(25±5)℃贮藏(M-25)和灭菌后(4±1)℃贮藏(M-4)期间感官品质和呈味核苷酸关联物含量的变化。结果表明,12种呈味核苷酸关联物在33 min内有效分离。灭菌和冷藏均能抑制雪菜贮藏期间感官品质和呈味核苷酸关联物的劣化,且灭菌冷藏组合的效果最佳,M-4组雪菜中三磷酸腺苷(adenosine triphosphate,ATP)和肌苷酸含量在第60天仍分别是初始值的29.7%和66.4%,且感官品质合格。雪菜在(25±5)℃贮藏期间与ATP相关的核苷酸关联物总量呈先增加后迅速降低的趋势,其中J-25组雪菜仅有9 d的感官合格期。灭菌和冷藏能够有效抑制雪菜中核酸代谢的呈味核苷酸(腺苷酸、鸟苷酸、胞苷酸、尿苷酸)在贮藏期内的变化,M-4组雪菜中这4种核苷酸总量在贮藏末期仍有4.30 mg/100 g。低盐雪菜中发酵微生物的代谢和较高的温度加快了呈味核苷酸关联物在贮藏期间的劣变,灭菌和冷藏有利于呈味核苷酸关联物的保持,研究结果可为低盐雪菜的加工和贮藏提供一定的理论支撑。

A high-performance liquid chromatography with circular dichroism detector for determination of stereochemistry of 6, 9-oxygen bridge dibenzocyclooctadiene lignans from kadsura coccinea

The stereochemistry of two 6, 9-oxygen bridge dibenzocyclooctadiene lignans from Kadsura coccinea, are difficult to separate and very unstable. The present study was designed to develop a high-performance liquid chromatography using circular dichroism detection for the analysis of the stereochemistry. A new 6, 9-oxygen bridge dibenzocyclooctadiene lignans named Kadsulignan Q was firstly found with an S-biphenyl configuration. The other compound was identified as Kadsulignan L with an R- biphenyl configuration. In order to obtain kinetic data on their reversible interconversion, the stability was measured at different deuterated solvents such as deuterated methanol, deuterated chloroform and deuterated dimethylsulfoxide. The lignans were more unstable and converted more easily in deuterated methanol than in deuterated chloroform and deuterated dimethylsulfoxide.

植物中1⁃脱氧野尻霉素特异性存在的考证

1⁃脱氧野尻霉素(DNJ)是桑树中最具有特征性、标志性的生物碱,具有调节糖脂代谢、抗菌、抗病毒等药理活性,是一种极具开发价值的天然活性产物。目前DNJ检测方法主要为柱前衍生高效液相色谱⁃荧光检测法,该方法能够检测到所有与荧光剂结合生成的络合物,但对被检测的物质无法进行定性,往往造成假阳性。通过对桑科植物和已报道含有DNJ的非桑科植物进行检测分析,利用柱前衍生高效液相色谱⁃荧光检测法与高效液相色谱⁃质谱(HPLC⁃MS)检测法相结合,挖掘并考证含有DNJ的植物种类,为DNJ的开发利用提供更广泛的原料素材。检测发现,除了桑树,其他常见桑科植物菠萝蜜、全叶无花果、小叶榕、裂叶无花果、构树、大叶榕、橡皮树、柳叶榕中均不含有DNJ;非桑科植物也只有风信子中含有DNJ;常规桑品种强桑1号的桑叶DNJ含量是风信子叶片DNJ含量的212倍。结果表明,对于植物中DNJ的检测,应先通过HPLC⁃MS进行定性研究,然后再通过柱前衍生高效液相色谱⁃荧光检测法进行定量检测,以避免出现假阳性。

酸奶中乙酰磺胺酸钾、苯甲酸、山梨酸和糖精钠的检测方法

[目的]建立使用高效液相色谱仪同时检测酸奶中的乙酰磺胺酸钾等4种食品添加剂的方法。[方法]酸奶样品经过蛋白沉淀剂沉淀蛋白后高速离心,经0.22μm微孔滤膜过滤后上机测定。仪器使用C18色谱柱,流动相采用甲醇与0.02 mol/L乙酸铵溶液(不需调节pH值),检测波长为227 nm。[结果]4种添加剂线性范围1.00~50.0 mg/L,相关系数均在0.999 9以上,回收率91.2%~104.8%。[结论]该方法操作简单、快速,可用于同时检测酸奶中的乙酰磺胺酸钾、苯甲酸、山梨酸和糖精钠4种食品添加剂。

高效液相色谱法同时检测特殊用途饮料中常见的咖啡因等6种食品添加剂

目的建立使用高效液相色谱仪同时检测特殊用途饮料中的咖啡因、糖精钠、安赛蜜、柠檬黄、胭脂红、日落黄等6种食品添加剂的方法。方法仪器使用C18色谱柱,流动相采用甲醇与0.02mol/L乙酸铵溶液(不需要调节pH值),经梯度洗脱,通过四个检测波长通道同时采集色谱图和光谱图进行检测分析。结果上述6种食品添加剂线性范围在1.00 mg/L~100.0mg/L,相关系数均在0.9999以上,回收率为91.0%~108.2%。结论本方法操作简单、快速,可用于同时检测特殊用途饮料中的咖啡因等6种食品添加剂。

高效液相色谱-紫外检测法同时测定类鼻疽患者血浆中亚胺培南、美罗培南和头孢他啶的浓度

目的建立一种快速、经济的高效液相色谱-紫外检测法(HPLC-UV),用于同时测定人血浆中亚胺培南、美罗培南和头孢他啶的药物浓度。方法采用Millipore 10KD超滤离心去蛋白,使用互为内标法,色谱柱为SunFire-C18柱(4.6 mm×250 mm,5µm);预柱为C18柱(4.0 mm×2.1 mm,5µm),流动相为0.1 mol·L^(-1)的3-(N-吗啡啉)丙磺酸(pH=7.0)和乙腈,梯度洗脱,检测波长299nm,流速1.0mL·min^(-1),运行时间30min,进样体积30µL。结果亚胺培南、美罗培南和头孢他啶的保留时间分别为6.699 min、10.795 min和8.722 min。血浆内源性物质对样品的测定无干扰,峰形良好。该方法具有良好的线性、准确度和精密度。不同浓度的样品在–20℃反复冻融、–20℃长期冷冻和4℃(48 h)下均表现出了较高的稳定性。结论该方法操作成本低、专属性强、分离效果好,可用于类鼻疽患者血浆中亚胺培南、美罗培南和头孢他啶治疗药物浓度的测定。

高效液相色谱-串联质谱法同时测定饲料中乙氧基喹啉及其伴生物质

本研究旨在建立一种应用高效液相色谱-串联质谱(HPLC-MS/MS)技术同时测定饲料中乙氧基喹啉(EQ)、对氨基苯乙醚(PD)、非那西丁(PC)、乙氧基喹醌亚胺(EQI)和乙氧基喹啉二聚体(EQDM)含量的方法。饲料样品用乙腈提取,经BONDESIL-SI多重杂质吸附材料净化,采用HPLC-MS/MS测定,以0.1%甲酸水溶液-0.1%甲酸乙腈溶液作为流动相,梯度洗脱,Waters BEH C_(18)色谱柱分离。结果表明:通过饲料基质匹配绘制标准曲线,EQ、PD、PC、EQI和EQDM在5~500μg/L范围内线性关系良好[决定系数(R^(2))>0.9950]。方法中EQ、PD和PC的检出限和定量限分别为3.0和10.0μg/kg,EQI和EQDM的检出限和定量限分别为1.5和5.0μg/kg。EQ、PD、PC、EQI和EQDM在3个添加水平(20、50、100μg/kg)的平均回收率为73.77%~98.77%,批内相对标准偏差(RSD,n=6)<10%,批间变异系数<12%。该方法前处理简单,操作方便,适用于饲料中EQ及其伴生物质的同步测定。