首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 125 毫秒
1.
建立了以12种同位素标记氨基酸做内标,采用衍生化方法,LC-MS/MS采用MRM方式同时定量30种氨基酸的检测方法。生物样本加入含有同位素内标的甲醇溶液,沉淀蛋白后氮气吹干加入衍生试剂,样本氨基酸与同位素氨基酸内标同步衍生,减小实验误差。选用TC-C18色谱柱,以水-乙腈(均含有0.01%七氟丁酸和0.1%甲酸)为流动相,采用梯度洗脱进行分离,30种氨基酸及其中的同分异构体都能基线分离。选用3200QTRAP型质谱仪的多重反应监测(MRM)扫描方式进行检测。本方法用于实时监测恶性血液病患者血浆氨基酸谱的变化,可以直接反映机体的代谢及营养状况,对患者的移植成功率及改善移植后患者的生存质量均有重要意义。  相似文献   

2.
NEWS     
《分析化学》2014,(11):1715
稳定同位素标记-液相色谱-质谱联用技术被广泛应用于代谢组学研究中,它克服了部分化合物内标获取困难的问题,可以实现对非目标性代谢物的定性和定量分析。  相似文献   

3.
利用蛋白质组学方法,将大鼠肝微粒体样品进行胰蛋白酶水解;再利用液相色谱-串联质谱法(LCMS/MS),采用多反应监测模式(MRM),通过测定蛋白质水解后产生的特征酶切肽段,实现同时对大鼠肝微粒体内药物代谢酶P450和UGT的绝对定量。本实验首先建立标准工作曲线,对肝微粒体样品中P450和UGT进行定量,在线性范围内,相关系数r>0.995,线性关系良好,定量限≤10 nmol/L;以合成的稳定同位素标记特征肽段作为内标,对UGT1A1进行定量分析。结果表明,同位素标记特征肽段与未标记肽段色谱行为与质谱响应一致,在基质溶液中同位素标记肽段线性关系良好,利用标准曲线法和稳定同位素稀释法测得UGT1A1含量分别为17.30和18.23 nmol/g,两种方法所得结果基本一致,但稳定同位素稀释法操作简便,更适用于复杂样品的高通量测定。  相似文献   

4.
建立了精氨酸、赖氨酸、丝氨酸、缬氨酸、亮氨酸等15种氨基酸的气相色谱-质谱联用(GC-MS)检测方法。以硅烷化试剂N-甲基-N-(三甲基硅烷)三氟乙酰胺(MSTFA)为衍生化试剂对氨基酸进行衍生化,在优化的色谱-质谱条件下,15种氨基酸均达到基线分离。结合混合阳离子(MCX)固相萃取柱对氨基酸发酵液实际样品进行净化,将所建立的方法用于~(15)N标记精氨酸发酵液中的氨基酸组成(包括亮氨酸-~(15)N、脯氨酸-~(15)N、精氨酸-~(15)N_4、谷氨酰胺-~(15)N和赖氨酸-~(15)N_2)分析,并根据EI图谱的离子碎片信息对~(15)N标记精氨酸发酵液中的氨基酸同位素丰度进行计算。  相似文献   

5.
建立了同时定性与定量分析动物饲料种24中禁用药物的超高效液相色谱串联质谱(UHPLC-MS/MS)分析方法。样品经Na2HPO4与饱和NaCl溶液共同盐析,乙腈提取,混合阳离子交换固相萃取柱(PCX)净化,乙腈和0.3%甲酸溶液梯度洗脱,经BEH C18色谱柱分离,串联四级杆质谱动态多反应监测正离子模式监测,稳定同位素标记内标法定量。在最佳实验条件下,24种禁用兽药在各自的线性范围内线性关系良好,相关系数(r)均大于0.9960,检出限为1.0μg/kg,定量限为2.0μg/kg,加标回收率在77%~115%之间,相对标准偏差小于10%。本方法的样品前处理过程简单,净化效果好,有效解决了基质效应问题,灵敏度高,适用于各种饲料中多组分禁用兽药的快速定性与定量分析。  相似文献   

6.
采用稳定同位素标记的多氯萘(PCNs)同类物为内标,建立了同位素稀释气相色谱/三重四极杆串联质谱技术测定环境样品中20种高关注的PCNs同类物的方法。结果表明:PCNs同类物的校正曲线在0.5~200 μg/L范围内线性良好(R2>0.99),检出限(LOD)为0.04~0.48 μg/L,相对标准偏差(RSD)小于15%。采用基质加标法评价该方法对实际环境样品中PCNs测定的回收率为45.2%~87.9%。为验证方法的适用性,以河流沉积物和再生铝冶炼排放的烟道气样品为对象,利用所建立的方法测定了20种PCNs同类物,并将结果与高分辨气相色谱/高分辨质谱方法的测定结果进行了比对,两种方法测定结果的RSD为0.5%~41.4%,表明所建立的同位素稀释气相色谱/三重四极杆串联质谱方法可用于实际环境样品中PCNs的定性、定量分析。  相似文献   

7.
提出了同位素稀释-气相色谱-质谱法测定白酒中23种邻苯二甲酸酯类化合物含量的方法。样品用正己烷-乙酸乙酯(1+1)混合液提取后,经DB-5MS毛细管色谱柱分离,采用电子轰击离子源选择离子反应监测模式进行质谱测定,同位素内标法进行定量分析。23种邻苯二甲酸酯类化合物的线性范围在0.2~3.0mg·L-1之间,测定下限(10S/N)在0.05~0.1mg·kg-1之间。加标回收率在73.2%~122%之间,相对标准偏差(n=6)在0.49%~11%之间。  相似文献   

8.
建立了准确、快速测定痕量人体生长激素的同位素稀释质谱法。选取同位素标记脯氨酸、缬氨酸和苯丙氨酸为内标物,将内标物以重量法与待测样品准确混合,利用Kinetex C18色谱柱分离,以电喷雾三重串联四级杆质谱多反应监测模式测定,建立了人体生长激素的液相色谱–同位素稀释质谱联用定量方法。人体生长激素溶液标准物质的含量测定结果为(1.82±0.04)mg/g,相对标准偏差为0.43%(n=6)。该方法简易、实用、准确、可靠,可作为人体生长激素溶液标准物质的定值方法,并为人体生长激素的日常检测提供参考。  相似文献   

9.
氨基酸稳定氮同位素(δ15 N)分析能准确有效地评估生物体的营养级以及氮在食物链中的流动.本研究优化了氨基酸氮同位素的分析方法:样品在酸性条件下水解后,释放出的蛋白质氨基酸经阳离子交换树脂纯化后,衍生为对应的N-新戊酞基,O-异丙醇(N-pivaloyl-isopropyl,NPP)酯,利用气相色谱-燃烧-同位素比值质谱仪(Gas chromatography-combustion-isotope ratio mass spectrometry,GC-C-IRMS)测定其δ15 N.经非极性气相色谱柱DB-5ms分离后,13种氨基酸NPP酯衍生物均可得到良好的基线分离.在样品量不低于20 ng N条件下,GC-C-IRMS方法的精密度优于1‰,测得的δ15 N值与EA-IRMS法测得的δ15 N值没有明显差异.阳离子树脂纯化前后各氨基酸δ15 N值差异低于1‰,表明没有产生明显的同位素分馏.采用本方法成功地估算了阿哈湖生态系统中常见水生生物的营养级,可作为研究氨基酸代谢以及生态系统特征的新方法.  相似文献   

10.
建立同位素内标稀释,超高效液相色谱串联质谱(Uhra performance liquid chromatography-tandem mass sepctrometry,UPLC-MS/MS)同时测定尿中苯系物和三氯乙烯代谢产物的方法,并用于孕妇尿样的分析.尿样过经含同位素内标的醋酸铵缓冲液稀释10倍后,过滤,取滤液进样分析.采用C18色谱柱,梯度洗脱进行UPLC分离,电喷雾负离子扫描多反应监测(Multiple reaction monitoring,MRM)模式检测.方法检出限在0.0350~ 1.75 μg/L之间,方法的日内和日间精密度分别为1.2%~9.1%和2.0%~9.7%;用所建立的方法测定了650个孕妇尿液样品,并统计分析了其地区差异,加标回收率为85.0%~104%.所建立的方法快速灵敏,适合尿液样品的批量分析.  相似文献   

11.
Very often, the accuracy of quantitative analytical methods for the determination of mycotoxins by liquid chromatography (LC)-mass spectrometry (MS) and LC-MS/MS is limited by matrix effects during the ionization process in the MS source. Stable isotope labeled standards are best suited to correct for matrix effects and to improve both the trueness and the precision of analytical methods employing LC-MS and LC-MS/MS. This paper describes the successful use of fully 13C isotope labeled deoxynivalenol [(13C15)DON] as an internal standard (IS) for the accurate determination of DON in maize and wheat by LC electrospray ionization MS/MS. To show the full potential of (13C15)DON as IS, maize and wheat extracts were analyzed without further cleanup. Subsequent to calibration for the LC-MS end determination, DON was quantified in matrix reference materials (wheat and maize). Without consideration of the IS, apparent recoveries of DON were 29±6% (n=7) for wheat and 37±5% (n=7) for maize. However, the determination of DON in the reference materials yielded 95±3% (wheat) and 99±3% (maize) when (13C15)DON was used as an IS for data evaluation.  相似文献   

12.
SILAC is a widely accepted approach for quantitative proteomics in which proteins are labeled with stable isotopes during cell culture. A major drawback of this technique is the metabolic conversion of labeled amino acids that may hamper accurate quantification. A paradigmatic example of this phenomenon is the generation of labeled proline from arginine, known to occur in a good number of biological models. We propose a novel methodology to identify and quantitate metabolic conversions as well as to evaluate labeling efficiency in SILAC experiments. In this approach, labeled proteins are reduced to amino acids by acid hydrolysis before LC-MS/MS analysis. Since it is carried out at the amino acid level, tracking the fate of the isotope label is straightforward and can be performed for each amino acid independently. After applying this method to mammalian cells, grown in the presence of heavy arginine and lysine, labeling efficiency and amino acid conversions could be accurately evaluated. Only undesirable labeling of proline was found to occur at a significant extent, varying greatly among cell lines. Finally, increasing proline concentration in the growing medium was shown to be effective at preventing arginine conversion without any noticeable side effect.  相似文献   

13.
A precolumn derivatization method for the determination of amino acids using 6-aminoquinolyl-N-hyroxy-succinimidyl carbamate (AQC) followed by high-performance liquid chromatography is described. Ultraviolet detection was used for the assay of AQC derivatives of amino acids with the detection wavelength set at 248 nm. The reagent peak interference was minimized by optimizing the pH of the eluent and the gradient elution profile to improve the resolution between the reagent peak and amino acid derivatives. All nineteen amino acids were separated in 35 min with resolutions 1.6. The correlation coefficients of the calibration graphs for seventeen amino acids were fairly good (r 0.9999) at concentrations of 25–500 μM. The detection limits for all common amino acids including cystine and trytophan were at the range of 0.07–0.3 pmol. Good reproducibility and accuracy of the method were demonstrated by the determination of amino acids in three typical kinds of samples (protein, peptide and feed.) The average relative standard deviations for bovine serum albumin (BSA) and neuromedin were 0.86% and 1.36, respectively, and the average relative errors were 3.2% and 2.3%, respectively. The results of the analysis of feed hydrolysates agreed with those obtained by an ion-exchange method and the average recovery of the method for feed hydrolysates was 98%.  相似文献   

14.
A rapid analytical method for amines and amino acids was developed, involving derivatization with the novel reagent 3‐aminopyridyl‐N‐hydroxysuccinimidyl carbamate (APDS), followed by reversed‐phase high‐performance liquid chromatography and electrospray ionization tandem mass spectrometry (HPLC/ESI‐MS/MS). More than 100 different analytes with amino groups, including amino acids in biological fluids such as mammalian plasma, could be measured within 10 min. The analytes were easily derivatized with APDS under the mild conditions. Selective reaction monitoring of ESI‐MS/MS in positive mode was carried out to include the transitions of all of the protonated molecular ions of analytes derivatized with APDS to the common fragment at m/z 121, which was derived from the amino pyridyl moiety of the reagent. We evaluated the retention time precision, the quantification limits, the linearity, the intra‐ and inter‐day precisions and the accuracy of 22 typical amino acids found in biological fluids, by analyzing a standard amino acid mixture and rat plasma. The intra‐day relative standard deviations (RSDs) of the retention times of the 22 amino acids and their internal standards were within 0.9% and the inter‐day RSDs were less than 1.1%, except for asparagines, with an RSD of 1.9%. The intra‐day and inter‐day RSDs of amino acid analyses in rat plasma were within 8.0% and 4.5%, respectively. The method, which facilitates the amino acid analysis of more than 100 samples in a day, represents an alternative to traditional amino acid analysis techniques, such as chromatography using postcolumn derivatization by ninhydrin. Copyright © 2009 John Wiley & Sons, Ltd.  相似文献   

15.
建立了饲料中8种脂溶性着色剂(对位红、苏丹红Ⅰ、苏丹红Ⅱ、苏丹红Ⅲ、苏丹红Ⅳ、苏丹红7B、苏丹红G、苏丹黄)含量的液相色谱-串联质谱测定方法。饲料样品中脂溶性着色剂经乙腈提取,离心后上清液采用分散固相萃取净化,净化液稀释后进行LC-MS/MS分析。样品测定时采用Acquity BEH C18色谱柱进行色谱分离,以0.2%甲酸溶液-乙腈作为流动相进行梯度洗脱,电喷雾正离子(ESI+)模式电离,多反应监测(MRM)模式检测,同位素稀释内标法定量。8种脂溶性着色剂在1.0~200μg/L范围内线性关系良好,相关系数(r~2)均大于0.998;在饲料中的方法检出限为5.0μg/kg,定量下限为10μg/kg。在10,50,500μg/kg加标浓度下8种脂溶性着色剂的回收率为102%~111%,批内相对标准偏差(RSD)为2.8%~8.0%,批间RSD为2.8%~7.8%。该方法能满足饲料样品中脂溶性着色剂监控的需要。  相似文献   

16.
An isotope-dilution liquid chromatography-tandem mass spectrometry (ID LC-MS/MS) measurement procedure was developed to accurately quantify amino acid concentrations in National Institute of Standards and Technology (NIST) Standard Reference Material (SRM) 2389a—amino acids in 0.1 mol/L hydrochloric acid. Seventeen amino acids were quantified using selected reaction monitoring on a triple quadrupole mass spectrometer. LC-MS/MS results were compared to gravimetric measurements from the preparation of SRM 2389a—a reference material developed at NIST and intended for use in intra-laboratory calibrations and quality control. Quantitative mass spectrometry results and gravimetric values were statistically combined into NIST-certified mass fraction values with associated uncertainty estimates. Coefficients of variation (CV) for the repeatability of the LC-MS/MS measurements among amino acids ranged from 0.33% to 2.7% with an average CV of 1.2%. Average relative expanded uncertainty of the certified values including Types A and B uncertainties was 3.5%. Mean accuracy of the LC-MS/MS measurements with gravimetric preparation values agreed to within |1.1|% for all amino acids. NIST SRM 2389a will be available for characterization of routine methods for amino acid analysis and serves as a standard for higher-order measurement traceability. This is the first time an ID LC-MS/MS methodology has been applied for quantifying amino acids in a NIST SRM material.  相似文献   

17.
建立了固相萃取-亲水相互作用色谱/串联质谱同时测定食品中三聚氰胺和三聚氰酸残留量的方法。采用乙腈和水提取试样中残留的三聚氰胺和三聚氰酸,正己烷脱脂,提取液经亲水性键合硅胶和阳离子交换树脂复合填料柱(MCT柱)净化。采用亲水相互作用色谱柱进行分离,质谱采用正、负离子切换模式电离,多反应监测模式检测,同位素内标法定量。三聚氰胺和三聚氰酸在10~2500 μg/L范围内呈线性相关,相关系数(r)均大于0.99,定量限分别为25和50 μg/kg。本方法在动物源性食品、植物源性食品、乳及乳制品等不同样品中的三聚氰胺和三聚氰酸高、中、低3个添加水平的回收率分别在70.0%~129.6%和70.0%~128.6%之间,相对标准偏差分别在1.4%~23.3%和2.8%~18.7%之间。该方法可满足食品中三聚氰胺和三聚氰酸同时定量测定的需要。  相似文献   

18.
建立了不同鱼贝类肌肉组织中以氘代同位素为内标测定环丙氟哌酸残留量的液相色谱-串联质谱(LC-MS/MS)方法。样品加入内标环丙氟哌酸-D8和磷酸盐缓冲溶液(pH 7.0)后进行匀质并用乙腈超声提取,经正己烷脱脂后采用Waters Oasis MAX小柱净化,在Cloversil-C18柱上,以乙腈-0.05%三氟醋酸(体积比为25∶75)为流动相,采用多反应监测(MRM)模式,液相色谱-电喷雾质谱法测定。根据环丙氟哌酸和氘代内标物的定量离子质量色谱图的峰面积比值,采用内标法定量。结果表明,环丙氟哌酸和内标的定量离子峰面积比值与环丙氟哌酸浓度在0.1~50.0 μg/kg范围内呈现良好的线性关系,相关系数为0.9991,方法的定量检测限为0.1 μg/kg,回收率为92.5%~98.1%,相对标准偏差(RSD)小于4.3%。将该方法用于市场上10种鱼和贝类样品的检测,结果表明该法具有灵敏、准确的优点,完全满足残留分析的确证检测要求。  相似文献   

19.
A derivatization system coupled to capillary electrophoresis for the determination of amino acids using 1,2-naphthoquinone-4-sulfonate as a labeling agent is described. In this system, amino acids are derivatized on-line in a three-channel flow manifold for sample, reagent and buffer solutions. The reaction takes place in a PTFE coil heated at 80 degrees C. The resulting solution, which contains the amino acid derivatives, is introduced into the electrophoretic system by means of an appropriate interface. Subsequently, amino acid derivatives are separated at 25 kV using a 40 mM sodium tetraborate aqueous solution with 30% (v/v) isopropanol solution as a running buffer. The electropherograms are monitored spectrophotometrically at 230 nm. The method has been applied to the determination of amino acids in feed samples and pharmaceutical preparations. A good concordance of the predicted values with those given by a standard amino acid analyzer is shown.  相似文献   

20.
N-acetylaspartic acid (NAA) is a specific urinary marker for Canavan disease, an autosomal recessive leukodystrophy. We developed a 'dilute and shoot' stable isotope dilution liquid chromatography tandem mass spectrometry (LC-MS/MS) method for determination of NAA in urine. Deuterated internal standard d(3)-NAA was added to untreated urine and the mixture was injected into the LC-MS/MS system operated in the negative ion mode. Chromatography was carried out on a C(8) minibore column using 50% acetonitrile solution containing 0.05% formic acid at a flow rate of 0.25 mL/min. The retention time was 1.6 min and the turnaround time was 2.2 min. NAA and d(3)-NAA were analyzed in multiple reaction monitoring mode. Calibrators and quality control samples were prepared in pooled control urine. The assay was linear up to 2000 micromol/L with limit of quantification at 1 micromol/L (S/N = 12). Interassay and intraassay coefficients of variation were less than 7% and recovery at three different concentrations was 98.9-102.5%. The LC-MS/MS method for NAA as described involves no extraction and no derivatization, showed no interference and gave excellent recovery with low variability and short analytical time. The method was successfully applied for the retrospective analysis of urine from 21 Canavan disease cases.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号