Salting-out extraction technique for pretreatment in the liquid chromatographic determination of copper(II), aluminum(III), iron(III) and manganese(II) in biological samples

A solvent extraction method for the simultaneous determination of Cu, Al, Fe and Mn by reversed phase liquid chromatography is presented. The metal chelates with 5-chloro-8-quinolinol are extracted into an acetonitrile phase with tetrabutylammonium perchlorate and ammonium sulfate as salting-out agents, followed by LC separation and determination using an ODS column. The experimental parameters such as the composition of mobile phase and concentration of the salting-out agent for phase separation have been investigated. The mobile phase is a mixture of 3:1 (v/v) acetonitrile-0.02 mol l(-1) sodium acetate solution containing 5x10(-3) mol l(-1) 5-chloro-8-quinolinol. The proposed method has been applied to the trace analysis of Cu, Al, Fe and Al in bovine liver and citrus leaves.     

Extraction-liquid chromatography with electrochemical and spectrophotometric detection for the determination of copper and iron in biological and river water samples

Reversed-phase liquid chromatography using cupferron as a precolumn derivatizing agent was developed for the determination of Cu(II) and Fe(III) in biological materials and natural water samples. In the direct method, the metal cupferronates formed in acetonitrile-water (1 + 1) are injected onto an ODS column followed by separation with a mobile phase containing acetonitrile-acetate buffer (pH 3.5) (7 + 3) and other reagents. Amperometric detection with a glassy carbon electrode at ?0.40 V vs. Ag/AgCl can be used to determine both metals simultaneously. The electrochemical detection method has better sensitivity for the determination of Fe(III) than the usual spectrophotometric detection at 375 nm. If a large volume of aqueous sample is available, concentration of the two metal ions can be made by extraction with ethyl acetate prior to the chromatographic determination. In this case, liquid chromatographic separation and determination can be performed with the ODS column using a mobile phase consisting of acetonitrile-methanol-ethyl acetate-0.02 M acetate buffer (pH 3.5) (45 + 20 + 5 + 30).     

Separation and simultaneous determination of cobalt, nickel and copper with 2-(4-methyl-2-quinolylazo)-5-diethylaminophenol by reversed-phase high-performance liquid chromatography

Performance of six quinolylazo compounds was studied as a pre-column derivatizing reagent for the separation of metal ions in the reversed-phase high-performance liquid chromatography. 2-(4-Methyl-2-quinolylazo)-5-diethylaminophenol (QADP) was suited for the determination of Co, Ni and Cu in the presence of a large amount of Fe. The detection limits under the optimized conditions [stationary phase: L-column ODS; mobile phase: acetone-water (62:38, v/v) containing 0.01 M NH(4)SCN and 0.01 M 2-morpholinoethanesulfonic acid-NaOH (pH 6); injection volume: 10 mul] were 0.24 ng for Co, 0.22 for Ni and 0.53 for Cu. This method was successfully applied to the determination of these metal ions in a standard steel sample.     

High-performance liquid chromatography of 8-hydroxyquinoline chelates of aluminium and cobalt(III)

Summary The 8-hydroxyquinoline chelates of Co(III) and Al(III) may be separated by high-performance liquid chromatography using a silica column and 5% methanol in chloroform as mobile phase. Using detection at 254 nm, the method provided detection limits of 0.9 ng of Co(III) and 17 ng of Al(III) in a 10 mm³ injection.     

Determination of Florfenicol in Freshwater, Sediments and Bryophyte Fontinalis antipyretica by HPLC with Fluorescence Detection

Novel procedures for the determination of florfenicol in freshwater, sediments and bryophyte Fontinalis antipyretica, using reversed-phase high-performance liquid chromatography are described. Liquid chromatography was performed on a 5 µm PuroSpher RP-18E^® column using methanol and 0.05 M phosphate buffer (18/82 v/v, pH 7.3) as mobile phase (0.8 ml min^?1) and fluorescence detection (excitation wavelength 265 nm and emission wavelength 295 nm). Florfenicol was determined in centrifuged freshwater samples. Florfenicol was extracted from sediments and bryophytes samples by using a solid-liquid extraction step followed by a solid phase extraction step. Linearity was confirmed over the concentration range 25–1000 ng mL^?1 water and 50-1000 ng g^?1 sediment or bryophyte. Limits of detection and quantitation were 8 and 25 ng mL^?1 water and 17 and 50 ng g^?1 sediment or bryophyte respectively. Mean extraction recoveries of florfenicol from sediments and bryophyte were from 85.9 to 109.1%.     

高效液相色谱-荧光分析法同时测定人体尿液中黄蝶呤与异黄蝶呤

建立了高效液相色谱-荧光法同时测定癌症病人尿液中黄蝶呤及异黄蝶呤的新方法。选择荧光检测波长λex=345nm,λem=420nm。以磷酸盐缓冲溶液(pH=7.5)-甲醇(体积比为98∶2)为流动相,流速1.0mL/min,黄蝶呤与异黄蝶呤含量分别在0.0013～0.945μg/mL及0.00017～0.118μg/mL范围内与色谱峰面积呈良好的线性关系,线性相关系数分别为0.9999和0.9996,检出限分别为0.5ng/mL和0.05ng/mL,加标平均回收率在86.2%～107.5%之间。方法应用于癌症病人尿样分析,取得了较好的结果。     

Determination of organic and inorganic mercury compounds by reverse-phase high-performance liquid chromatography after extraction of the compounds as their dithizonates

A sensitive and selective method is described for the simultaneous determination of inorganic and organic mercury compounds. The mercury compounds are extracted into toluene or chloroform with dithizone, and the dithizonates are separated by liquid chromatography on an ODS column. Complete resolution was obtained between methylethyl-, phenyl- and inorganic mercury with a mobile phase of THF/methanol (2:1) with 0.05 M acetate buffer pH 4 (62 + 38), containing 50 μM EDTA. The mercury chelates were detected spectrophotometrically at 475 nm. The detection limits were at the subnanogram level. The method is applicable to human urine, tap water and tomatoes.     

Determination of catecholamines as aminochromes by micellar liquid chromatography with thermal lens spectrophotometric detection

Summary The determination of catecholamines (CAs) using micellar liquid chromatography with thermal lens spectrophotometric detection has been studied. CAs are oxidized with hexacyanoferrate(III) to aminochromes which are separated with a mobile phase of 0.05 M sodium dodecyl sulphate, 7% propanol and 0.03 M citrate buffer, pH 4.8, on a partially endcapped C₁₈ column. The aminochrome-micelles and aminochrome-stationary phase association constants are evaluated. Using the 488 nm line of an Ar⁺ laser with 250 mW pump power the limits of detection are about 4 ng mL^–1. The technique is applied to the determination of unconjugated CAs in urine using isoproterenol as internal standard.     

Determination of climbazol in shampoos by reversed-phase liquid chromatography

A method based on ior-pair reversed-phase liquid chromatography is reported for the determination of climbazol, a preservative used in cosmetic products. The column was packed wit LiChrosob RP-18, the mobile phase was acetonitrile-sodium perchlorate (0.05 M, pH 3.0) (40 + 60) and detection was at 224 nm. The method was applied to the analysis of commercial shampoos. The detection limit was 10 ng injected and calibration linearity was observed up to 6 μg.     

Comparison of Micellar and Reversed-Phase Liquid Chromatography for Determination of Sulfamethoxazole and Trimethoprim

A simple, sensitive, and precise micellar liquid chromatographic method for simultaneous analysis of sulfamethoxazole and trimethoprim, with ultraviolet detection at 245 nm, has been developed, validated, and used for determination of the compounds in commercial pharmaceutical products. The compounds were well separated on a Hypersil ODS reversed-phase column at 35°C by use of a mobile phase consisting of 0.1M sodium dodecyl sulfate in a 2:98 (V/V) mixture of 1-butanol and pH 3.0 phosphate buffer solution at a flow rate of 1.0 mL min^?1. A comparative study of the performance of reversed-phase liquid chromatography with aqueous-organic or micellar-organic mobile phases for separation of sulfamethoxazole and trimethoprim is reported. The study showed that micellar liquid chromatography (MLC) and reversed-phase liquid chromatography (RP HPLC) are of similar efficiency, sensitivity, and selectivity for determination of sulfamethoxazole and trimethoprim.     

A spectrometric,separation and voltammetric study of the complexation reactions of bromazepam with iron(ii),copper(ii) and cobalt(ii)

Bromazepam, in the form of a cationic iron(II) chelate, can be determined spectrophotometrically at 588 nm with a limit of detection of ca. 10^-6 M. When this chelate is ion-paired with perchlorate, it can be extracted into organic solvents such as 1,2-dichloroethane and 4-methyl-2-pentanone, and determined by atomic absorption spectrometry with a limit of detection of 1.5 × 10^-5 M bromazepam at the iron resonance 248.3-nm line. Ion-pairs involving the Fe(II), Cu(II) and Co(II) chelates and perchlorate can be separated by h.p.l.c. using a C₁₈ reverse-phase column and a mobile phase of 4:1 water—methanol, with a u.v. detector at 242 nm. This approach allowed for the determination of iron(II) ions in aqueous solution with a limit of detection of 10^-8 M. The h.p.l.c. method could also be used to quantify bromazepam spiked in plasma in the concentration range 1–10 μg ml^-1, following extraction of bromazepam from plasma and subsequent formation of the iron(II) ion-pair. Copper(II) forms a labile chelate with bromazepam in pH 4.8 acetate buffer which, when subjected to differential pulse voltammetry at the hanging mercury drop electrode, gives rise to a catalytic phenomenon which can be utilised for the determination of bromazepam in the concentration range 10^-5–10^-9 M.     

Capillary electrophoretic separation between Fe(II) and Ni(II) precomplexed with 1,10-phenanthroline in a high concentration buffer of n-butyric acid/ n-butyrate and its application

The capillary electrophoretic separation was accomplished for Fe(II) and Ni(II) precomplexed with 1,10-phenanthroline (phen) in 2 M n-butyric acid/ n-butyrate buffer at pH 4.5 with direct UV detection at 260 nm. The applied voltage was 5 kV. The high concentration buffer of the n-butyrate resulted in a similar separation mechanism to that of ion-pair reversed-phase high-performance liquid chromatography. The separation would be due to the hydrophobic interaction between the ionic associates, [Fe(phen)(3)]( n-butyrate)(+) and [Ni(phen)(3)]( n-butyrate)(+), with the n-butyrate ion and n-butyric acid as background electrolyte. Linear calibration ranges were obtained for Fe(II) and Ni(II) from 100 to 500 ng ml(-1). The relative standard deviations ( n=10) for 3 g mL(-1) Fe(II) and Ni(II) were 0.090 and 0.086, respectively. Detection limits ( S/ N=3) for Fe(II) and Ni(II) were 20 ng mL(-1). The method was applied to the determination of nickel in aluminium and duralumin alloys.     

Selective and Sensitive LC-MS Determination of Avenanthramides in Oats

A new liquid chromatographic – mass spectrometric (LC-MS) method suitable for the determination of avenanthramides in oats has been developed. The avenanthramides were detected using positive ion electrospray with selective ion monitoring of protonated ions [M+H]⁺. The separation of avenanthramides was achieved on Genesis C₁₈ column by using acetonitrile – aqueous formic acid as mobile phase and gradient elution. The effects of buffer nature and concentration on separation, peak shape and intensity of mass spectrometric signal were investigated. The detection limits ranged between 10 and 30 ng mL^?1 for standard solutions when using 1 μL injection. The developed LC-MS method was found to be superior over commonly used liquid chromatographic methods with UV detection in terms of selectivity and sensitivity. The applicability of the method to real samples (oat groats) was demonstrated.     

土壤和植物中乙二胺二琥珀酸的反相液相色谱法测定

乙二胺二琥珀酸(EDDS)可应用于重金属污染土壤的强化植物修复或是土壤淋洗修复。但是现有的检测方法或过于复杂,或检出限太高。本研究建立了应用超声波萃取、硫酸铜衍生化和液相色谱,紫外检测器测定土壤、植物中EDDS含量的方法。其回收率为93%~118%,灵敏度较高;检出限达1.5μmol/L;重现性好,不受其它多种共存离子的干扰。     

Improved high-performance liquid chromatographic procedure for the determination of chlormethiazole levels following solid-phase extraction from plasma

An improved high-performance liquid chromatographic method for the determination of chlormethiazole levels in plasma is described. The drug is extracted from plasma using commercially available reversed-phase extraction columns; recovery values obtained using Sep-Pak C18 and Bond Elut C1, C2, C4, C6, C8, C18 columns are compared. Separation was achieved by reversed-phase chromatography, using a mobile phase consisting of 0.025 M sodium acetate buffer, pH 4.5-acetonitrile (67:33) at a flow-rate of 1.6 ml/min in conjunction with a 15-cm Jones Chromatography Apex ODS column. The analytical column was protected by a Waters Assoc. Guard-Pak module containing a Guard-Pak CN insert. Using ultraviolet detection at 254 nm chlormethiazole levels in the region of 50 ng/ml can be measured with only 500 microliter of plasma.     

高效液相色谱光度法测定痕量铜、锌、铁、锰的磺化四苯基卟啉配合物

本文用磺化四苯基卟啉作柱前衍生试剂,研究了试剂与铜(Ⅱ)、锌(Ⅱ)、铁(Ⅲ)及锰(Ⅱ)的络合反应。在ODS柱上,用乙腈-水(40:60,V/V)作流动相,溴化四丁基铵作离子对试剂,在420nm波长处进行光度检测。提出了离子对反相高效液相色谱快速分离及测定痕量铜、锌、铁及锰的新方法。检测下限为(×10^-3ppm):Mn 3.6、Fe 1.8 、Zn 0.93、Cu 0.70。应用于植物样品中痕量铜、锌、铁及锰的测定,分析结果与标准参考值相符。     

反相高效液相色谱法同时测定镉、铅、铜和锌

 研究了meso-四（对羟基苯基）卟啉为柱前衍生化试剂与Cd2+,Pb2+,Cu2+和Zn2+离子的配合反应条件及配合物在C18色谱柱上的分离条件,建立了反相高效液相色谱快速分离光度检测Cd2+,Pb2+,Cu2+和Zn2+的新方法。配合物和试剂在15 min内出峰完毕。4种离子的检出限为: Cd2+0.02 ng,Pb2+0.02 ng, Cu2+0.02 ng,Zn2+0.12 ng;线性范围为:Cd2+0.8 μg/L～150 μg/L,Pb2+0.8 μg/L～300 μg/L,Cu2+0.8 μg/L～500 μg/L,Zn2+5.0 μg/L～1 000 μg/L;方法的日内相对标准偏差为:2.8%～4.8%,测定低、中、高3个浓度的日间相对标准偏差为3.7%～9.7%。     

The use of 3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine as a precolumn derivatizing reagent in HPLC determination for Fe(II) in natural samples.

3-(2-Pyridyl)-5,6-diphenyl-1,2,4-triazine (PDT) was used for the first time as a precolumn derivatizing reagent in the high-performance liquid chromatography (HPLC) method with UV absorbance detection for the Fe(II) determination. The Fe(II) reacts with PDT to form a magenta colored chelate in the presence of sodium dodecyl sulfate (SDS) and acetic acid-sodium acetate buffer solution medium of pH 4.65. The selection of maximum absorbance detection wavelength and the optimum composition of the organic modifier in the mobile phase were investigated in detail for the quantitative determination of Fe(II) using HPLC system. The formed Fe(II)-PDT chelate was satisfactorily separated from PDT on an Agilent Shim-pack ODS column (Eclipse XDB-C8, 4.6 x 150 mm) by isocratic elution with acetonitrile and 0.02 mol L(-1) acetic acid-sodium acetate buffer solution (pH 4.65, containing 0.02% of SDS and 60 x 10(-3) mol L(-1) NaClO(4)) as mobile phase at a flow rate of 1 mL min(-1), and monitored with a multiple wavelength detector. The detection limit (S/N = 3) is 0.35 ng mL(-1). Due to the excellent separation ability of HPLC, the innovative introduction of PDT as the precolumn derivatizing reagent, and the proper selection of the detect wavelength, the sensitivity of our newly developed HPLC method was enhanced remarkably compared to those of the common spectrophotometric methods. The developed HPLC method was successfully applied to the determination of Fe(II) in lake water samples.     

Determination of trace amounts of vanadium in air samples with 2-(8-quinolylazo)-5-N,N-diethylaminophenol by reversed-phase liquid chromatography

A reversed-phase liquid chromatographic method for the determination of trace amounts of vanadium is described. Metal ions are converted into 2-(8-quinolylazo)-5-N,N-diethylaminophenol chelates in an off-line system. The chelates are injected onto a Zorbax CN column and separated with an aqueous acetonitrile mobile phase containing no chromogenic reagent. Unter these conditions, only vanadium(V) is spectrophotometrically detected at 540 nm among the metal ions Al(III), Ba(II), Ca(II), Cd(II), Co(II), Cr(III), Cu(II), Fe(III), Ga(III), Hg(II), Mg(II), Mn(II), Ni(II), Pb(II), V(V) and Zn(II). Amounts of 8.0–200 pg of vanadium(V) in 100-μl injections can be determined without interference from 10-fold molar excesses of many cations. At 0.001 a.u.f.s., the detection limit (twice the peak-to-peak noise) for vanadium(V) is 8.0 pg in 100 μl of injected solution and the relative standard deviation at 120 pg of vanadium(V) in a 100-μl injection is 3.5%. The proposed method is applied to the determination of vanadium in rain water and airborne particulates.     

Determination of clomipramine and its hydroxylated and demethylated metabolites in plasma and urine by liquid chromatography with electrochemical detection

A procedure for the determination of clomipramine and its 8-hydroxy, demethyl, 8-hydroxydemethyl and didemethyl metabolites in plasma and urine by high-performance liquid chromatography with electrochemical detection is described. A 1-ml plasma or urine sample is made alkaline with a carbonate buffer (pH 9.8) and extracted with 20% ethyl acetate in n-heptane. After back-extraction into an acid phosphate buffer (pH 2.4), an aliquot is injected into a 5-microns ion-paired reversed-phase column and eluted with a mobile phase containing a phosphate buffer with tetramethylammonium chloride-acetonitrile (57:43). The detection is coulometric with a first cell at +0.40 V, a second at +0.73 V and a guard cell set at 0.75 V for oxidation of the mobile phase. The method provides recoveries in the general range of 80-110% and a day-to-day precision of 3.7-8.8%, depending on the compound. The minimum quantifiable level for all compounds was 0.2 ng/ml with a 20-microliters injection. Steady-state plasma concentration data and urinary levels are reported for 24 depressed patients receiving daily either 75-150 mg orally or 50-75 mg by infusion.