Plasma free amino acid profiling of esophageal cancer using high-performance liquid chromatography spectroscopy

AIM:To perform plasma free amino acid(PFAA)profiling of esophageal squamous cell carcinoma(ESCC)patients at different pathological stages and healthy subjects.METHODS:Plasma samples from ESCC patients(n=51)and healthy control adults(n=60)were analyzed by high-performance liquid chromatography(HPLC).The ESCC patients included moderate/poorly-differentiation(n=24),lymph node metastasis(n=17)and clinical stage>Ib2(n=36).Partial least squares discriminant analysis was performed to demonstrate that the PFAA metabolic patterns enabled discrimination between ESCC patients and controls,and the Student t test was applied to assess significant differences in PFAA concentrations between the two groups.RESULTS:There were significant differences in the PFAA profiles between controls and ESCC patients.Compared with healthy controls,the levels of Asp,Glu,Gly,His,Thr,Tau,Ala,Met,Ile,Leu,and Phe were decreased in ESCC patients,but Cys was increased.There exists a strong correlation between PFAA profiles and clinicopathological characteristics in ESCC patients.The levels of many PFAAs(i.e.,Glu,Asp,Ser,Gly,Tau,Ala,Tyr,Val,Ile,and Leu)were related to pathological grading,lymph node metastasis,and ESCC clinical stage.Very good discrimination between ESCC patients and control subjects was achieved by multivariate modeling of plasma profiles.CONCLUSION:HPLC-based plasma profiling analysis was shown to be an effective approach to differentiate between ESCC patients and controls.PFAA profiles may have potential value for screening or diagnosing ESCC.

Determination of Trace Amount of Polycyclic Aromatic Hydrocarbons in Urban Sewage by Solid-phase Extraction Coupled with High Performance Liquid Chromatograph

[Method] This study aimed to determine trace amount of polycyclic aromatic hydrocarbons(PAHs) in urban sewage by using solid-phase extraction(SPE) coupled with high performance liquid chromatograph(HPLC).[Method] From the aspects of solid-phase extraction column,elution solvent,elution volume,elution speed and so forth,the test conditions of SPE-HPLC method were optimized,and trace amount of PAHs in urban sewage was determined.[Result] The optimized solid-phase extraction conditions were SUPELCLEAN LC-18 solid-phase extraction column,methylene dichloride as elution solvent,15 ml elution volume,2 ml/min elution speed,5 ml/min loading speed,1 000 ml water with 200 ml methanol loading volume.Under the optimized extraction conditions,the recovery was high,namely 76.3%-105.2%;relative standard deviation was 3.8%-6.0%,showing good precision;detection limit was low,only 0.000 8-0.048 0 μg/L.[Conclusion] This method is user-friendly,with high sensitivity and good precision,and suitable for continuous determination of a large volume of water samples.

Determination of Organic Acids in Root Exudates by High Performance Liquid Chromatography:Ⅱ.Influence of Several Testing Conditions

Effects of column temperature and flow rate on separation of organic acids were studied by determining nine low-molecular-weight organic acids on reversed- phase C18 column, using high performance liquid chromatography (HPLC) with a wavelength of UV (ultraviolet) 214 urn and a mobile phase of 18 mmol L-1 KH2PO4 buffer solution (pH 2.1). The thermal stability of organic acids was determined by comparing the recoveries of organic acids in different temperature treatments. The relationships between column temperature, flow rate or solvent pH and retention time were analyzed. At low solvent pH, separation efficiency of organic acids was increased by raising the flow rate of the solvent because of lowering the retention time of organic acids. High column temperature was unfavorable for the separation of organic acids. The separating effect can be enhanced through reducing column temperature in organic acid determination due to increasing retention time. High thermal stability of organic acids with low concentrations was observed at temperature of 40 ℃-45℃. Sensitivity and separation effect of organic acid determination by HPLC were clearly improved by a combination of raising flow rate and lowering column temperature at low solvent pH.

Freshwater algae chemotaxonomy by high-performance liquid chromatographic(HPLC)analysis

The study of community composition of algae is essential for understanding the structure and dynamics of the aquatic ecosystem and for evaluating the eutrophic level of the water body.A high-performance liquid chromatographic(HPLC)method based on a reversephase C_(18) nonpolar column was developed for the main algal taxa,which includes cyanophytes,bacillariophytes,euglenophytes,dinophytes,and chlorophytes.Based on the elution order using HPLC,19 pigments were identified,and they were chlorophyllide a,19′-butanoyloxyfucoxanthin,chlorophyll c_(1)+c_(2),phephorbides a,peridinin,methyl-chlorophyllide a,fucoxanthin,neoxanthin,violaxanthin,myxoxanthophyll,diadinoxanthin,diatoxanthin,lutein,zeaxanthin,chlorophyll b allomer,chlorophyll b,chlorophyll a allomer,chlorophyll a,andβ,β-carotene.A comparison study of cell microscopic counts and accessory pigment analysis indicated that HPLC analysis could be a useful tool for monitoring phytoplankton communities and their abundance.

HPLC法测定云南省不同辣椒品种中辣椒碱及二氢辣椒碱的含量

利用超声提取法对云南省不同地区不同品种辣椒样品进行预处理,采用高效液相色谱法(HPLC)测定样品中辣椒碱和二氢辣椒碱的含量。具体方法为:采用色谱柱HanbonSci&Tech.LichrospherC18(250×4.6mm,5μm)分离,甲醇-3‰磷酸水溶液梯度洗脱(65%～90%,0～30min),检测波长280nm,辣椒碱和二氢辣椒碱的相对标准偏差都小于1%,平均回收率都在99%以上。该方法具有较好的分离效果和较高的稳定性,适用于云南省不同地区不同品种中辣椒碱和二氢辣椒碱含量的测定。测定结果显示云南省不同地区不同品种不同成熟度的辣椒中辣椒碱和二氢辣椒碱含量差异很大。

高效液相色谱法(HPLC)测定大豆异黄酮含量的研究

建立检测大豆异黄酮5种组分（染料木素、染料木苷、大豆苷元、大豆苷和黄豆黄苷）含量的高效液相色谱法,并且测定了大豆各组织和不同时期胚的异黄酮含量。色谱条件：Phenomenex C18色谱柱（150 mm×4.6 mm,5.0μm）;流动相：甲醇-水（30∶70,v/v）;流速：1 mL.min-1,检测波长：254 nm;柱温：40℃,进样量：10μL。试验建立的色谱条件能使大豆异黄酮主要组分得到很好分离,线性关系R2为0.9994~0.9998,回收率为99.29%~100.76%且变异系数小于3%,适用于大豆异黄酮不同组分的检测。不同大豆品种异黄酮含量表现为：吉林32〉GR8836〉吉大豆2号〉平安8〉小黑豆〉吉林47〉吉林35〉吉大豆1号;吉林32各组分大豆异黄酮含量表现为：大豆籽粒〉60 d胚〉50 d胚〉40 d胚〉花〉荚〉20 d胚〉30 d胚〉叶〉根〉茎。

HPLC-DAD法测定粗粮馒头中6种合成色素

建立一种简便、快速、重复性好的高效液相色谱-二极管阵列检测器测定粗粮馒头中的柠檬黄、苋菜红、胭脂红、诱惑红、日落黄、亮蓝6种合成色素的含量。选用无水乙醇-氨水-水(7∶2∶1,V/V)为样品的提取溶剂,色谱柱为Waters Symmetry?C18柱(250 mm×4.6 mm,5μm),流速1.0 mL/min,柱温35℃,多波长同时测定6种的合成色素,检测波长分别为425、483、520、620 nm。流动相以甲醇为A相,以含0.02 mol/L乙酸铵、1%冰醋酸和0.2%三乙胺溶液(pH 4)为B相,进行梯度洗脱。结果表明:6种合成色素均达到基线分离,具有良好的线性关系(r=0.999 9);样品的柠檬黄、苋菜红、胭脂红、诱惑红、日落黄、亮蓝加标回收率分别为97.94%、96.40%、96.28%、97.63%、98.80%、100.40%,相对标准偏差均小于2.0%。该方法可用于甄别是否由染色而成的粗粮馒头。

肉鸭表皮组织中脱氢枞酸残留的SPE-HPLC检测方法

建立经松香脱毛处理的肉鸭表皮组织中残留脱氢枞酸的固相萃取-高效液相色谱检测方法。肉鸭表皮组织中的脱氢枞酸用乙腈提取,经C18固相萃取小柱净化,以0.003 mol/L磷酸溶液-甲醇(13:87,v/v)为流动相,采用反相C18(250 mm×4.6 mm,5 μm)色谱柱分离,以紫外检测器(检测波长212 nm)进行检测。结果表明:该方法检测脱氢枞酸线性范围为0.1~10 mg/L,检测限为0.10 μg/g,定量限为0.33 μg/g。在低中高3个添加量范围内回收率为80.8%~91.8%。实际样品分析显示,肉鸭表皮中脱氢枞酸含量最高达10.06 μg/g。所建立的分析方法简便快捷、具有较好的灵敏度和可靠性,可用于肉鸭表皮组织中脱氢枞酸残留量的分析。

昆虫体内多胺的高效液相色谱(HPLC)测定

建立丹磺酰氯柱前衍生HPLC快速测定昆虫体内多胺含量的方法。以C18(250mm×4.6mm,5μm)为固定相,甲醇和水为流动相,梯度洗脱,柱温40℃,流速1mL/min,荧光检测波长激发波长(Ex)280nm,发射波长(Em)515nm,测得腐胺(put)、亚精胺(spd)和精胺(spm)三者回收率分别为98.7%,99.2%和97.8%,回归方程线性良好(r值均大于0.99),分析时间为16min。该法简洁、快速、灵敏度高、重现性好,可有效分析昆虫及其他生物样品中微量多胺的含量。

大豆异黄酮组分HPLC快速分析技术及其在豆腐加工中的应用

大豆异黄酮育种与大豆制品加工研究需要进行大批量样品12种异黄酮组分的快速定量分析。在前人研究基础上利用Agilent 1100高效液相色谱（HPLC）系统,以大豆品种南农95C-13为材料,研究大豆苷元、大豆苷、乙酰基大豆苷、丙二酰基大豆苷、染料木苷元、染料木苷、乙酰基染料木苷、丙二酰基染料木苷、黄豆苷元、黄豆苷、乙酰基黄豆苷、丙二酰基黄豆苷等12种标准品外标法快速定量技术。从样品制备与色谱条件对分析12种异黄酮组分的准确度和分离度入手,确定分析流程,以（科丰1号×南农1138-2）的184个重组自交系（NJRIKY）为材料,研究豆腐加工中总量和各组分的变化特点。（1）样品以80%甲醇水溶液50℃超声1 h提取;色谱条件为,检测波长254 nm,柱温36℃,流速2.0 mL min^-1,进样量10μL,流动相0.1%（V/V）乙酸水溶液（A）和100%甲醇（B）,02 min,27%B（V/V）→2-3 min,27%-38%B→310 min,38%B→1012 min,38%-39%B→1214 min,39%B→1415 min,39-27%B梯度洗脱;在15 min内将12个组分良好分离,各组分峰面积与其相应浓度均呈良好线性关系（R^2为0.99760.9999）;加标回收率均大于99%,变异系数低于2%。（2）NJRIKY群体籽粒、豆乳、豆腐异黄酮总量和组分的大量分析验证了HPLC快速技术的效果。籽粒异黄酮总含量（3 695.00μg g^-1）在豆乳加工中平均85.15%转入豆乳（3 146.12μg g^-1）,14.85%（548.88μg g^-1）进入豆渣。传统豆腐加工通过硫酸钙絮凝,只有17.32%（639.89μg g^-1）转入豆腐,67.83%（2 506.23μg g^-1）留在黄浆水中。豆乳中12种组分含量比籽粒中稍低,均以丙酰基染料木苷含量最高;而豆腐中乙酰基染料木苷和乙酰基黄豆苷缺失,以染料木苷元与大豆苷元含量较高。大豆科丰1号与南农1138-2杂交重组后家系间的异黄酮遗传变异增大,增加了对其遗传改良的潜力。

HPLC-MS/MS法分析大鼠体内淫羊藿素葡糖醛酸结合物

应用高效液相色谱-串联质谱(HPLC-MS/MS)法鉴别大鼠血浆及尿粪中淫羊藿素葡糖醛酸结合物。生物样品经固相萃取法处理后,以V(乙腈)∶V(水)∶V(乙酸)=45∶55∶1.5为流动相,通过Dikma C18柱分离,采用电喷雾离子四极杆串联质谱在负离子检测方式下进行一级和二级全扫描质谱分析。通过提高离子源内的DP电压,使待测物发生源内裂解产生碎片离子,再对产生的碎片离子进行子离子扫描,可获得相当于三级质谱分析的结果。根据待测物的色谱和质谱信息,发现淫羊藿素在大鼠体内可代谢为1种双葡糖醛酸结合物M1和2种单葡糖醛酸结合物M2、M3。大鼠灌胃给予淫羊藿素后,在血浆和尿样中可检测到M1、M2和M3,在粪便中仅检测到M2和M3。

灯笼果中噻菌灵的RP-HPLC-PDA残留分析

目的:建立反相高效液相色谱-二极管阵列检测器测定灯笼果中杀菌剂噻菌灵残留的分析法。方法:色谱柱为Shim-pack VP-ODS(150mm×4.6mm),检测波长为299nm,流动相为pH6.8的乙腈-0.02mol/L磷酸缓冲溶液(70:30,V/V),流速为0.6mL/min,进样量20μL。结果:噻菌灵在0.10～90.0μg/mL范围内与峰面积成良好线性关系(r=0.9995),检测限为0.05μg/mL,噻菌灵的回收率为87.9%～102.6%,相对标准偏差为0.98%～2.79%。结论:该方法操作简便快速,可作为灯笼果中噻菌灵含量监测的控制方法。

黄芪超微粉HPLC指纹图谱的建立

目的:建立黄芪超微粉高效液相色谱(HPLC)的指纹图谱,为其质量控制和科学评价提供依据。方法:采用Inertsil ODS-SP(4.6mm×250mm,5μm)色谱柱,甲醇-乙腈(1:1,V/V)为流动相A,超纯水为流动相B,进行梯度洗脱,流速0.8mL/min,柱温25℃,检测波长260nm。采用中药色谱指纹图谱相似度评价系统2004A进行相似度分析,并进行聚类分析。结果:建立了黄芪超微粉的指纹图谱,确定12个共有峰。聚类分析表明10批样品存在明显的南北地域差异。结论:此方法简便、准确、重复性好,可用于黄芪超微粉质量控制与评价。

反相高效液相色谱法(RP-HPLC)测定丙酮酸发酵液中丙酮酸

提出一种利用反向高效液相色谱法(RP-HPLC)测定丙酮酸发酵液中丙酮酸的方法。应用反相C18色谱柱 ODS-2 HYPERSIL 250 mm×4．6 mm 5μm,以0．05 mol·L-1NH4H2PO4(用H3PO4调pH2．6)为流动相,发酵液经稀硫酸预处理后直接进样分离定量,每一样品的分析过程不超过5 min,可及时了解丙酮酸发酵过程中丙酮酸的变化,适于发酵液中丙酮酸的检测。

RP-HPLC-DAD法测定茶叶籽油酚类化合物

建立同时分离、检测茶叶籽油中酚类化合物组分和含量的高效液相色谱法。比较了流动相组成的影响,选择合适的乙酸加入量,建立最优梯度洗脱条件。结果表明,14种酚类物质在该条件下能够得到较好的分离,各物质的线性相关系数均在0.99以上。运用该方法从茶叶籽油中共检测到13种酚类化合物,包括5种酚酸、3种黄酮类和5种儿茶素类,含量分别为46.8、21.72、40.16μg/g;其酚酸类主要为肉桂酸和咖啡酸,黄酮类主要为山奈酚、槲皮素和芦丁,儿茶素类主要为儿茶素。该方法准确、可靠,适用于茶叶籽油中酚类化合物分析。

HPLC法测定消瘿平亢颗粒中栀子苷含量

目的:用HPLC法测定消瘿平亢颗粒中栀子苷含量。方法:色谱柱为ODS(Diamonsil,250 mm×4．6 mm, 5μm),流动相为乙腈-水(15:85),流速为0．8 mL·min^(-1),检测波长238 nm。结果:栀子苷在0．12～0．24μg进样量范围内栀子苷进样量与色谱峰面积呈良好的线性关系(r=0．9992);平均回收率为96．79%(n=5),RSD为1．1%。结论:本法可以用于消瘿平亢颗粒中栀子苷含量的测定。

反相HPLC同时检测乳酸及乳酸甲酯含量

本实验采用反相高效液相色谱法实现了对乳酸、乳酸甲酯的分离并对其定量分析效果进行研究。HPLC分析条件为:Purospher STAR C18反相柱,二极管阵列检测器,以0.005mol/LH2SO4与甲醇混合溶液(9:1,V/V)为流动相,流速为1.0ml/min,进样量20μl,检测波长210nm。该方法快速、简便、灵敏、重现性好。

HPLC法测定清肺消炎颗粒中葛根素血浆浓度及其在小鼠体内的药动学研究

目的：建立HPLC法测定清肺消炎颗粒水溶液灌胃给药后小鼠血浆中葛根素的含量,并研究其在小鼠血浆中的药动学行为。方法：采用Hypersil BDS C18色谱柱（250 mm×4.6 mm,5μm）,流动相∶甲醇-水（25∶75）,流速：1.0mL/min;检测波长：250nm;柱温：30℃。血浆样品采用甲醇直接沉淀蛋白后,测定小鼠血浆中葛根素的含量并计算药动学参数。结果：小鼠血浆样品中葛根素的线性范围为0.6563~42.00μg/mL,日内与日间精密度均〈10%,平均提取回收率为（97.39±1.44）%。灌胃给药后葛根素在小鼠体内的药-时曲线符合一室模型,主要药动学参数Cmax为1.575μg.L-1,tmax为0.9901 h,t1/2α为0.3281 h,AUC为6.039μg·L-1.h-1。结论：该法准确快速,灵敏度高,可用于小鼠血浆中葛根素的含量测定及体内药动学研究。

超声波辅助-反相HPLC法测定郑州市火棘果中的有机酸

用高效液相色谱法对郑州地区火棘果中有机酸的种类及含量进行了测定.色谱柱为HypersilODSC1(84.6mm×250mm,5μm);流动相:磷酸二氢钠-磷酸缓冲液(pH2.6);流速:0.8mL/min;检测波长:210nm.实验结果表明:火棘果果实中主要含柠檬酸9.243mg/kg,苹果酸10.244mg/kg,琥珀酸7.215mg/kg;在0.1～200.0μg/mL范围内,线性关系良好,相关系数分别为0.9999,0.9999,0.9998;最低检测限为0.01～0.04μg/mL.本法简便、准确、快捷,用于郑州地区火棘果中有机酸的测定,为火棘资源的质量检测与开发利用提供了科学依据.

进口西拉葡萄酒的HPLC-DAD-MS特征指纹图谱

建立进口西拉葡萄酒的高效液相色谱-二极管-离子阱质谱指纹图谱分析方法。样品采用乙酸乙酯萃取,旋转蒸发仪浓缩,甲醇溶解,高效液相色谱-离子阱质谱分析测定。色谱柱采用MGⅡC18柱(150 mm×2.0 mm,5μm),以pH 3.0水-乙腈为流动相,梯度洗脱,流速0.25 mL/min,多波长检测,离子阱质谱采用负离子模式,扫描范围m/z 50~1 200,可对没食子酸等11个组分进行定性和定量。维拉西拉葡萄酒的15个共有峰在60 min内得到良好分离,通过与标准品比较,对其中6个峰进行了确证,10批次维拉西拉葡萄酒的相似度均大于0.9。方法简便、灵敏、稳定、精确,生成的葡萄酒指纹图谱能够用于进口西拉葡萄酒的质量评价。