对酞内酰胺苯甲酰氯柱前衍生反应HPLC分析痕量多胺

本文提出了一种快速、简便、灵敏检测痕量多胺的新方法。用对酞内酰胺苯甲酰氯同多胺进行柱前衍生反应,以反相高效液相色谱定量测定多胺含量。在4～500 pmol范围内有良好的定量线性关系,腐胺、精脒和精胺的检测限均为0.1 pmol。     

对酞内酰胺苯甲酰氯柱前衍生反应HPLC法检测痕量芳香胺

建立了一种快速、灵敏检测痕量芳香胺的新方法。对苯胺、邻甲苯胺和间甲苯胺与对酞内酰胺苯甲酰氯（简称ＰＩＢ－Ｃｌ）的衍生反应条件、衍生物色谱分离及定量检测条件等进行了研究。衍生试剂在有机溶剂中与３种芳香胺反应迅速，衍生物用乙腈＋水（４８＋５２）作流动相，在反相色谱系统中得到了良好的分离。３种芳香胺的检出限分别为：苯胺１．５ｐｍｏｌ，邻甲苯胺和间甲苯胺均为２．０ｐｍｏｌ。     

化学衍生法应用于生物多胺的HPLC荧光检测

介绍了在高效液相色谱荧光衍生法分析多胺中常用的几类荧光衍生试剂及其应用,对近年多胺分析中的高效液相色谱荧光衍生方法进行了综述。     

高效液相色谱在生物医药研究中的应用第七讲生物胺的分离和测定（下）：多胺及有关化合物的分离和测定

§7-6 多胺及有关化合物的分离 多胺是具有两个以上氨基的脂肪族化合物,四种重要的多胺及有关化合物的名称和结构见表1。此类化合物无紫外吸收,又无荧光和电化学活性,检测困难,必须衍生后才能检测,因此多胺的分离方法与其衍生有关。     

氯甲酸芴甲酯柱前衍生高效液相色谱法测定植物中多胺

建立了一种采用氯甲酸芴甲酯进行柱前衍生，高效液相色谱分离和测定植物中的游离态和结合态多胺的新方法。腐胺、精脒和精胺的分离在１６ｍｉｎ内完成；检出限分别达到０．０３０、０．０１８和０．０２３ｐｍｏｌ线性动态范围至少是３个数量级。研究了反应温度和时间对荧光衍生物产率的影响，指出在２５℃下反应２０ｍｉｎ可获得较高产率。多胺总量与衍生试剂的摩尔比超过１：８时，荧光衍生物产率恒定。本方法用于要和番茄样品的测     

对甲氧基苯磺酰氯柱前衍生生物胺的高效液相色谱分离

采用对甲氧基苯磺酰氯作为柱前衍生试剂,RP-HPLC为分析模式,建立了一种新的生物胺衍生化方法,并对葡萄酒中生物胺含量进行检测。通过液质联用对产物进行定性,研究并确定了最适衍生化条件:衍生温度50℃,缓冲液pH 9.0,衍生时间15 min。实验建立了7种生物胺的HPLC分离方法:Beckman ODS柱;流动相A为10 mmol/L的NH4Ac溶液(pH 6.37),B相为乙腈;采用梯度洗脱;流速1 mL/min;检测波长240 nm,室温。     

应用固相萃取技术分析主流烟气中的芳香胺

建立了一种定量检测卷烟主流烟气中的芳香胺的新方法:应用第一柱为阳离子交换保留机理而第二柱为吸附保留机理的固相萃取(SEE)净化程序来处理样品,从第一柱上洗脱的芳香胺用五氟丙酸酐进行衍生化后,进入第二柱吸附色素和其它杂质,利用GC/MS进行分析测定.该方法具有灵敏度高、准确度高和易于自动化等优点,检出限为0.09ng/cig.～1.26ng/cig.,回收率为76.4%～103.1%.用该方法测定了某品牌卷烟烟气总粒相物中的14种芳香胺.     

双冠醚的研究——Ⅳ.N,N-氨二乙酸类化合物与苯并-15-冠-5在多聚磷酸中生成亚甲基双(4'-苯并-15-冠-5)的反应

在多聚磷酸中,苯并冠醚与一元酸或二元酸反应,缩合而生成相应的酰基衍生物。本文用氨或胺衍生的N,N-胺二乙酸类化合物与苯并-15-冠-5和多聚磷酸相反应,探讨了这种体系的反应特点。所用的N,N-胺二乙酸为分析化学上几种常用的氨羧络合剂     

柱前衍生-荧光检测-高效液相色谱-质谱法测定尿中游离多胺

用新型荧光试剂1,2-苯并-3,4-二氢咔唑-9-乙基氯甲酸酯作为柱前衍生化试剂,在乙腈中,以pH 9的硼酸钠缓冲溶液为催化剂,40℃下衍生反应10 min后获得稳定的荧光产物.在Eclipse XDB-C8色谱柱上,通过梯度洗脱对5种多胺进行了分离和在线质谱定性.激发波长(λex)和发射波长(λem)分别为333 nm和390 nm.采用大气压化学电离源(APCI)正离子模式,实现了人尿中游离多胺的质谱定性及荧光定量测定.检出限(3S/N)在4.98～9.31 fmol.此方法应用于尿样中多胺的测定,并测得回收率在94%～98%之间.     

专利介绍

专利申请号 :98117772　公开号 :12 10 2 6 2专利名称 :胺羟酰氯色谱衍生试剂及其制备工艺文摘 :本发明涉及一种高灵敏度紫外及荧光检测液相色谱衍生化试剂 胺羟酰氯及其制备工艺。胺羟酰氯化学名称为 2 硝基 4 (2 邻苯二甲酰基 )苯甲酰氯。本发明主要解决的技术问题是如何用简便、经济的工艺条件来制备反应活性好、检测灵敏度高、易于保存的酰氯类色谱衍生试剂。本发明的主要工艺技术特征在于 :在乙醚介质中 ,邻苯二甲醛和 2 硝基 4 氨基苯甲酸缩合成 2 硝基 4 (2 邻苯二甲酰基 )苯甲酸。然后 ,在氯仿介质中以亚硫酰氯进行酰氯…     

Determination of polyamines in hydrolysates of uremic plasma by high-performance cation-exchange column chromatography

The levels of putrescine, cadaverine, spermidine and spermine in uremic plasma were determined with an automatic polyamine analyzer with a 7.5 X 0.2 cm I.D. cation-exchange column using a stepwise sodium chloride gradient. All four polyamines were higher in ten patients with chronic renal failure than in eight normal subjects. The total polyamine content was also measured in the patients' plasma before and after maintenance dialysis; putrescine and spermidine levels were significantly lowered by the procedure.     

Polyamine distribution in the rat intestinal mucosa

As the first step in a study of mucosal polyamine metabolism during intestinal adaptation, we have measured mucosal polyamine concentrations at different sites along the normal rat intestine. Putrescine, spermidine, spermine and cadaverine were measured by spectrofluorometric analysis after thin-layer chromatography of their dansylated derivatives. Spermidine was present in the largest amounts at each of the sampling sites. The ratio of the concentration of spermidine to that of spermine paralleled the established pattern of cellular proliferation in the normal intestine as did the putrescine concentration (nmol per 10 cm) which decreased from duodenum to colon. These results provide the essential background to an assessment of the role of polyamines in the intestinal adaptive response.     

Determination of polyamine metabolome in plasma and urine by ultrahigh performance liquid chromatography–tandem mass spectrometry method: Application to identify potential markers for human hepatic cancer

To evaluate the potential relationship between cancer and polyamine metabolome, a UHPLC–MS/MS method has been developed and validated for simultaneous determination of polyamine precursors, polyamines, polyamine catabolite in human plasma and urine. Polyamine precursors including l-ornithine, lysine, l-arginine and S-adenosyl-l-methionine; polyamines including 1,3-diaminopropane, putrescine, cadaverine, spermidine, spermine, agmatine, N-acetylputrescine, N-acetylspermine and N-acetylspermidine; polyamine catabolite including γ-aminobutyric acid had been determined. The analytes were extracted from plasma and urine samples by protein precipitation procedure, and then separated on a Shim-pack XR-ODS column with 0.05% heptafluorobutyric acid (HFBA) in methanol and 0.05% HFBA in water. The detection was performed on UHPLC–MS/MS system with turbo ion spray source in the positive ion and multiple reaction-monitoring mode. The limits of quantitation for all analytes were within 0.125–31.25 ng mL⁻¹ in plasma and urine. The absolute recoveries of analytes from plasma and urine were all more than 50%. By means of the method developed, the plasma and urine samples from hepatic cancer patients and healthy age-matched volunteers had been successfully determined. Results showed that putrescine and spermidine in hepatic cancerous plasma were significant higher than those in healthy ones, while spermidine, spermine and N-acetylspermidine in hepatic cancerous urine were significant higher than those in healthy ones. The methods demonstrated the changes of polyamine metabolome occurring in plasma and urine from human subjects with hepatic cancer. It could be a powerful manner to indicate and treat hepatic cancer in its earliest indicative stages.     

Monitoring of biologically active amines in cereals and cereal based food products by HPLC

Summary Biologically active amines (putreanine sulphate, N-acetyl putrescine, putrescine, cadaverine, histamine, agmatine, N-acetyl spermidine, spermidine, spermine) were separated and quantified in cereal flour and cereal products by a liquid chromatographic method. The method consists of the separation of ion pairs formed between biologically active amines and octanesulphonic acid on a reversed-phase column, postcolumn derivatization with o-phtalaldehyde-2-mercapthoethanol and spectrofluorometric detection. Results of the reliability study were satisfactory. The method was linear for each amine at 1–10 mg L⁻¹. Putrescine and spermidine were the only amines always detected in cereal flour and cereal products, ranging from 2.45 to 47.83 mg kg⁻¹ for putrescine and 3.27 to 37.14 mg kg⁻¹ for spermidine. The most important differences among types of samples were found in polyamine derivatives. Presented at: Balaton Symposium on High-Performance Separation Methods, Siófok, Hungary, September 3–5, 1997.     

Sensitive fluorimetric method for the determination of putrescine, spermidine and spermine by high-performance liquid chromatography and its application to human blood

A fast and sensitive method for the determination of putrescine, spermidine and spermine by high-performance liquid chromatography is described. These compounds are converted to their fluorescent dansyl derivatives and are separated by a reversed-phase chromatographic system (Micropak CH-10) with water and acetonitrile as mobile phase. The sensitivity of the method is 30 pmoles. The application of the method to the determination of polyamines in blood is described. It was found that most of the polyamines circulating in blood are localized in the erythrocytes, their content in normal human blood being spermidine 14.1 +/- 3.1, and spermine 8.4 +/- 2.8 nmoles/ml packed erythrocytes. The polyamine level in serum is less than 0.1 nmole/ml. The polyamine content of the erythrocytes from patients with malignant neoplasms was significantly elevated.     

The polyamines of Xanthium strumarium and their response to photoperiod

Flowering plants of Xanthium strumarium L., grown in 8 h photoperiods, were analysed for polyamines. Putrescine, spermidine and spermine were found throughout the plant in three forms: (a) as free polyamines; (b) conjugates soluble in 5% trichloracetic acid (TCA); and (c) bound to the TCA-insoluble precipitate. On a fresh weight basis, total polyamines are most abundant in young leaves and buds, especially flower buds. Spermidine predominates in the free polyamine fractions, while spermine is dominant in the conjugated fraction. Transfer of vegetative plants from 16 h photoperiods to 1, 2, 3, or 4 inductive cycles (8 h light + 16 h uninterrupted dark) caused rapid and marked changes in the polyamine titer of the leaves and ultimately, floral initiation. The titer of free putrescine per mg protein declined progressively with induction in all leaf sizes, while the titers of free spermidine and spermine rose during days 2 and 3 in small and expanding leaves. Conjugated putrescine, spermidine and spermine rose sharply after only 1 inductive cycle, especially in small and expanding leaves, and maintained the higher level for at least several cycles. In plants given 4 inductive cycles, buds harvested after 4 additional days had sharply elevated levels of conjugated polyamines, especially spermine, on a protein basis.     

Rapid and Simple Technique for the Quantitation of Polyamines in Biological Samples

Abstract

A rapid and simple technique has been developed to quantify putrescine, spermidine, and spermine in biological tissue. The method, based upon several published procedures, involves protein precipitation with perchloric acid followed by dansylation with 5-dimethylamino-1-naphthalenesulfonyl chloride (dansyl chloride). After extraction on a Waters C₁₈ Sep-Pak cartridge, the samples are analyzed by high pressure liquid chromotography using a step solvent change and a 3μ C₁₈ reverse phase column. The chromotographic conditions allowed complete analysis of the three polyamines within 10 min with a total run time of 13 min (sample injection and re-equilibrium of column). Standard curves were linear up to 1 μg polyamine and the coefficient of variation for the assay ranged from 4% at l μg polyamine per sample to 11% at 50 ng polyamine per sample. The assay is therefore both rapid and simple. Moreover, unlike other available methods, the present technique does not require duel pumps, ion pairing agents, solvent extraction or a gradient control system. The concentrations of putrescine, spermidine and spermine in rat lung, liver and kidney are reported.     

High performance liquid chromatography of biological polyamines using immobilized enzyme as post-column reactor followed by electrochemical detection

A novel analytical method for biological polyamines (putrescine, spermidine and spermine) was developed. Polyamines were separated by ion-pair reversed phase chromatography using a polymer-based octadecyl bonded column. A polyamine oxidase immobilized column worked effectively as a post-column reactor to convert polyamines to hydrogen peroxide which was eventually detected by electrochemical oxidation on platinum electrode. This method required neither tedious derivatization nor gradient elution, permitting us to perform simple and rapid analysis of polyamines. The detection limits were 0.3, 0.6, and 4 pmol injected for putrescine, spermidine, and spermine, respectively with a linear range of two to three orders of magnitude. Chromatograms obtained with samples from human urine and rat brain homogenates demonstrated the high sensitivity and selectivity of the method.     

Simplified micro-method for the quantitative analysis of putrescine, spermidine and spermine in urine

A simplified micro-method for the quantitative analysis of urinary polyamines is described. After acid hydrolysis of urine, the polyamines are converted to fluorescent 1-dimethylaminonaphthalene-5-sulfonyl (Dns; dansyl) derivatives and separated by means of thin-layer chromatography. Dns-NH2, which has been reported to interfere with the determination of putrescine, is well separated from di-Dns-putrescine. Putrescine, spermidine and spermine are quantitated by in situ scanning of their fluorescent spots on the chromatogram. The present method is both sensitive and reproducible. It eliminates a number of time-consuming steps and thus reduces preparative losses. Yet an adequate chromatographic resolution is obtained. Representative polyamine analyses of urine from normal volunteers and from cancer patients are reported. Elevated levels occur in the urines of pregnant women and of patients with various types of cancer.     

The use of selected ion flow tube mass spectrometry to detect and quantify polyamines in headspace gas and oral air

Polyamines are a class of aliphatic compounds which include putrescine, cadaverine, spermine and spermidine. They are involved in a variety of cellular processes and have been implicated in a number of different pathophysiological mechanisms. Polyamines are volatile compounds having a distinctive odour normally perceived as being unpleasant. The measurement of their abundance has, however, been restricted to compounds present in the aqueous phase. Using selected ion flow tube mass spectrometry (SIFT‐MS) we have shown that the polyamines react with the ions H₃O⁺, NO⁺ and O to form distinctive product ions allowing their levels to be quantified in the vapour phase. The low volatility of spermine did not allow extensive analysis of this compound by SIFT‐MS while the adherent properties of cadaverine and putrescine required the use of PTFE transfer lines and couplers. Our data suggested the presence of cadaverine and putrescine in both oral air and the headspace of putrefying bovine muscle, while product ions corresponding to putrescine and spermidine were found in the headspace of human semen. SIFT‐MS therefore appears to be a practical means of measuring vapour‐phase polyamine levels, having applications in biology, medicine and dentistry, and food science. Copyright © 2009 John Wiley & Sons, Ltd.