Ion-interaction High-performance Liquid Chromatography Research Articles

Validated Reversed-Phase Ion-Interaction High-Performance Liquid Chromatography for Quantitation of Nitrate Content of Clausena anisata (Willd.) Hook. f. ex Benth. Leaves.

This work sought to validate the reversed-phase ion-interaction high-performance liquid chromatography for quantifying the nitrate content in the extract and raw material of Clausena anisata (Willd.) Hook. f. ex Benth. leaves. Three extraction methods (i.e., decoction, infusion, and ultrasound-assisted extraction) were investigated and compared. Furthermore, the effect of the solid-to-solvent ratio and defatting was also evaluated. The validation result showed that the high-performance liquid chromatographic method had a linear response (R2 = 0.9999) in the range of 1–50 μg/mL. The limit of detection and limit of quantitation were 0.25 μg/mL and 0.75 μg/mL, respectively. In addition, the method was specific, precise, and accurate. So the validated method was suitable for determination of the nitrate content in C. anisata leaves. Infusion of a nondefatted sample using a solid-to-solvent ratio of 1 : 10 gave the highest nitrate content in the raw material, 0.251 ± 0.003%. In case of a defatted sample, decoction provided the highest nitrate content, 0.309 ± 0.001%. Increasing the solid-to-solvent ratio and defatting had a huge effect on the nitrate content of C. anisata leaves extracted from decoction. To the best of our knowledge, this is the first report that used the reversed-phase ion-interaction high-performance liquid chromatography for quantifying the nitrate content in C. anisata leaves. Furthermore, the authors suggested that nitrate could be used as a standard marker for quality control of C. anisata leaves' extract and raw material.

Investigation of the retention/pH profile of zwitterionic fluoroquinolones in reversed-phase and ion-interaction high performance liquid chromatography

The retention/pH profiles of three fluoroquinolones, ofloxacin, norfloxacin and ciprofloxacin, was investigated by means of reversed-phase high performance liquid chromatography (RP-HPLC) and reversed-phase ion-interaction chromatography (RP-IIC), using an octadecylsilane stationary phase and acetonitrile as organic modifier. Sodium hexanesulphonate and tetrabutylammonium hydroxide were used as sources of counter ions in ion-interaction chromatography. The retention/pH profiles under in RP-HPLC were compared to the corresponding lipophilicity/pH profiles. Despite the rather hydrophilic nature of the three fluoroquinolones positive retention factors were obtained while there was a shift of the retention maximum towards more acidic pH values. This behavior was attributed mainly to non-hydrophobic silanophilic interactions with the silanized silica gel material of the stationary phase. In ion-interaction chromatography the effect of counter ions over a broad pH range was found to be ruled rather by the ion pair formation in the mobile phase which led to a drastic decrease in retention as a consequence of the disruption of the zwitterionic structure and thereupon the deliberation of a net charge in the molecules. At pH values at which zwitterionic structure was not favored both the ion-exchange and ion pair formation mechanisms were assumed to contribute to the retention.

On-line preconcentration and sample clean-up system for the determination of vanadium as a 4-(2-pyridylazo) resorcinol–hydrogen peroxide ternary complex in plant tissues by ion-interaction high performance liquid chromatography

An HPLC method for the determination of vanadium using on-line preconcentration and sample clean-up has been developed employing a C 18 column. 4-(2-Pyridylazo) resorcinol (PAR) and hydrogen peroxide were used as pre-column complexing agents for the determination of vanadium. Aliquots of either standard or sample solutions (100 μL and 2 mL) containing the vanadium complex were loaded onto a guard column (C 18, 0.4 mm × 4.6 mm) with a carrier mobile phase comprising 20% (v/v) methanol, 5 mM acetic acid, 5 mM citric acid, l0 mM tetrabutylammonium bromide (TBABr) at pH 7 (eluent 1). A flow rate of 2 mL/min for 0.40 min was used for a 100 μL injection loop, and 1.65 min for the 2 mL injection loop, with the effluent being directed to waste. The vanadium complex was flushed onto the analytical column (C 18, 150 mm × 3.9 mm, 4 μm), via an automatic switching valve, using a mobile phase comprising 32% (v/v) methanol, 5 mM acetic acid, 5 mM citric acid, and 5 mM tetrabutylammonium bromide (TBABr) at pH 7, with a flow rate of 1 mL/min for 1.60 min. The switching valve was returned to its original position, and any unwanted species remaining on the guard column were eluted with methanol for 5 min. The guard column was then re-equilibrated with eluent 1 for 5 min before the next injection. The analytes were detected at 540 nm. The detection limits (determined by a signal-to-noise ratio of 3) of vanadium with on-line preconcentration using 2 mL and 100 μL injection loops were 3.1 and 67 ng/kg, respectively. The developed method was used for the determination of vanadium in plant tissue. Chinese green mustard and tomato plant samples analysed in this study were cultured hydroponically. Vanadium concentrations measured using the HPLC technique were compared with those obtained using magnetic sector ICP-MS. Vanadium levels in plant tissue samples (root, stem and leaf of Chinese green mustard) found by HPLC agreed well (>85%) with those found using magnetic sector ICP-MS.

A simplex-optimized chromatographic separation of fourteen cosmetic preservatives: analysis of commercial products.

An ion-interaction high-performance liquid chromatography (HPLC)-diode-array detection method is developed and optimized for the separation of typical antimicrobial agents used in cosmetics and hygiene products. The most used preservatives contain different molecular structures, different functionalities, and are characterized by different chemical properties. Organic acids, alkyl esters of benzoic acids, alkyl p-hydroxy benzoic acids (parabens), phenol derivatives, and carbanilides represent the most used preservatives, and are often present in multicomponent mixtures. In order to develop a multicomponent method to be used in quality control analysis, the ion-interaction reagent reversed-phase HPLC technique seems to be particularly suitable, because it allows for the simultaneous separation of acidic, basic, and neutral species. The experimental conditions of the method are developed by OVAT (one variable at a time) treatment and further optimized by a multivariate approach based on a Simplex algorithm that works on a desirability function targeted to maximize the resolution in a multicomponent mixture. The new method proposed that is able to simultaneously separate fourteen preservatives is applied in the analysis of commercial products.

A survey of the potential of the high performance liquid chromatography and capillary zone electrophoresis for the determination of platinum and platinum group metals

Various platinum group metals (PGMs) can be separated in the form of complexes with monodentate ligands, ion associates or chelates by high performance liquid chromatography (HPLC) or capillary zone electrophoresis (CZE). Ion associates of anionic monodentate PGM complexes with low and high-molecular-mass ammonium cations give satisfactory results in ion interaction HPLC on reversed phase silica in the presence of CH3CN or CH3OH modifiers. PGM chelates often show considerable shortcomings due to kinetic hindrance during their formation and must be formed prior to HPLC application. Capillary zone electrophoresis (CZE) brings some new aspects to the separation of PGMs in addition to increasing separation speed and miniaturization of the system. Both methods can be successfully used for the treatment of platinum anti-cancer drugs, their organic impurities, hydrolytic products or metabolites in body fluids. Representative examples are discussed in this paper.

On-line preconcentration of niobium(V) and tantalum(V) as 4-(2-pyridylazo) resorcinol–citrate ternary complexes in geological samples by ion interaction high-performance liquid chromatography

4-(2-Pyridylazo) resorcinol (PAR) and citrate were used as pre-column complexing agents for the determination of Nb(V) and Ta(V) as ternary complexes in geological samples. Aliquots of 2 ml of the standard and sample solutions containing the Nb(V) and Ta(V) complexes were loaded onto a concentrator column (C 18, 0.4 cm×4.6 mm) with a carrier mobile phase comprising 20% (v/v) methanol and containing 5 m M acetic acid, 5 m M citric acid and 10 m M tetrabutylammonium bromide (TBABr), pH 6.5 at 2 ml/min for 2 min, with the effluent being directed to waste. An automatic switching valve was then switched to flush both complexes from the concentrator column onto a C 18 analytical column using a mobile phase comprising 32% (v/v) methanol and containing 5 m M acetic acid, 5 m M citric acid and 3 m M TBABr, pH 6.5 for 2.5 min. The switching valve was then switched back to the original position, and cleaned with methanol for 7 min to eliminate unwanted species still adsorbed to the concentrator column. This procedure prevented later eluting compounds from reaching the analytical column, which reduced the overall run time. The detection limits of Nb(V) and Ta(V) (determined at a signal-to-noise ratio of 3, detection wavelength of 540 nm and a 2-ml sample volume) were 0.012 and 0.039 ppb for Nb(V) and Ta(V), respectively. Recoveries of Nb(V) and Ta(V) were 99.4 and 96.2%, respectively. The HPLC results obtained from the reference granite and basalt samples agreed well with inductively coupled plasma MS and certified values, but the HPLC method yielded slightly low values of the Nb/Ta ratio.

Separation and determination of niobium(V) and tantalum(V) as 2-(5-bromo-2-pyridylazo)-5-[ N-propyl- N-(3-sulfopropyl)amino]phenol citrate ternary complexes in geological samples using ion interaction high-performance liquid chromatography

A method for the simultaneous separation and determination of Nb(V) and Ta(V) as ternary complexes formed with 2-(5-bromo-2-pyridylazo)-5-[ N-propyl- N-(3-sulfopropyl)amino]phenol (PAPS) and citrate was developed using ion interaction reversed-phase high-performance liquid chromatography (HPLC) on a C18 column. An artificial neural network (ANN) in combination with a reduced factorial experimental design was utilised to optimise the mobile phase compositions and the conditions used for pre-column complex formation. Under the optimum conditions determined by the ANN, the Nb(V) and Ta(V) complexes were eluted within 5 min. The detection limits (signal to noise ratio = 3, detection wavelength 600 nm) for Nb(V) and Ta(V) were 0.03 and 0.40 μg/l, respectively, which correspond to 0.03 and 0.40 μg/g in the solid samples. The method was applied to the analysis of geological samples and the results obtained by the HPLC method agreed well with the results obtained by ICP-MS and also with the certified values.

Determination of sulphonated dyes in water by ion-interaction high-performance liquid chromatography

An ion-interaction high-performance liquid chromatography method for quick separation and determination of the sulphonated dye Acid Yellow 1, and the sulphonated azo dyes Acid Orange 7, Acid Orange 12, Acid Orange 52, Acid Red 2, Acid Red 26, Acid Red 27 and Acid Red 88 has been developed. An RP-ODS stationary phase is used, and the mobile phase contains an acetonitrile–phosphate buffer (27:73, v/v) mixture at pH 6.7, containing 2.4 m M butylamine as ion-interaction reagent. Good separations were obtained using isocratic elution and spectrophotometric detection at 460 nm. The detection limits for the eight dyes ranged from 7 to 28 μg/l for an injection volume of 100 μl. Spiked tap water samples (100 ml), containing different concentration levels (0.3–1.2 μg/l) of the dyes were analyzed after acidification (pH 3) and preconcentration in disposable solid-phase extraction C 18 cartridges.

Determination of Niobium(V) and Tantalum(V) as 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol (Br-PADAP)-citrate ternary complexes in geological materials using ion-interaction reversed-phase high-performance liquid chromatography

A method for the simultaneous separation and determination of Nb(V) and Ta(V) as ternary complexes with 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol (Br-PADAP) and citrate using ion-interaction reversed-phase high performance liquid chromatography on a C 18 column has been developed. The mobile phase compositions and pre-column complex formation conditions were optimised. Under the optimum conditions, the Nb(V) and Ta(V) complexes were eluted within 4 min with a mobile phase of methanol-water (50% v/v) containing 12 mM tetrabutylammonium bromide and 12 mM citrate buffer at pH 7, with absorbance detection at 600 nm. The resulting % relative standard deviation of retention times, peak area and peak height for Nb(V) and Ta(V) complexes were 0.007, 0.000, 0.67, 2.69, 0.44 and 2.17, respectively. The detection limits (signal to noise ratio = 3) for Nb(V) and Ta(V) were 0.03 ppb and 0.41 ppb, respectively. The method was applied to the analysis of geological samples (granite, basalt and andesite) and the results for Nb(V) obtained by the HPLC method using standard addition calibration agreed well with ICP-MS analysis and with certified values for all materials. For Ta(V), good agreement with certified values was obtained for granite samples using standard addition, but interferences were observed for basalt and andesite which prohibited the determination of Ta(V) in these samples.

Optimization of the separation of mono- and dichloroanilines in ion interaction high-performance liquid chromatography

To optimize the ion-interaction chromatographic separation of nine chloroaniline isomers, the effect on retention of six experimental parameters is investigated by means of multivariate analysis. The factors considered are the organic modifier concentration in the mobile phase; the length of the alkyl chain of the alkylammoniumion salts used as the ion-interaction reagents (IIRs); the concentration of IIRs; the pH of the mobile phase, the flow-rate and the ionic strength. The use of fractional factorial and star designs allowed one to draw out useful information on the retention mechanism involved and to build a model characterized by both descriptive and predictive ability. Concerning descriptions, the results suggest a retention mechanism mainly based on reversed-phase partition, while the main role of the alkylamine (used as IIR) seems to mask the activity of the residual silanol groups on the stationary phase. As a result efficiency is improved. For prediction purposes, the regression models allow the optimization of the chromatographic separation, as regards both resolution and total analysis time. The study allowed one to develop a method able to separate the nine mono- and dichloroanilines in a total analysis time within 66 min and with detection limits ranging from 4.0 to 21.0 μg/l.

Ion-interaction high-performance liquid chromatography and micellar electrokinetic capillary chromatography: two complementary techniques for the separation of aromatic sulfonated compounds

Aromatic sulfonates and their amino and/or hydroxyl derivatives are widespread environmental pollutants. Since they are commonly present as complex mixtures, with the presence of many isomers, their separation is still a critical analytical step. Due to the lack of a standard reference method, recommendations are made to always use two independent methods in order to compare and confirm the results. Due to the ionic nature of these compounds ion-interaction HPLC and micellar electrokinetic capillary chromatography were chosen to solve this separative problem. With the use of both techniques the separation of 22 aromatic sulfonates was optimized, and the effect of several experimental parameters evaluated. The results obtained were discussed in terms of resolution, sensitivity and precision of the methods. An industrial effluent was analyzed employing the proposed procedures.

Photodiode Array Detection as a Fingerprinting Tool for Metal Cyanide Complexes in Gold Processing Solutions

Reversed-phase ion-interaction high-performance liquid chromatography with photodiode array detection (PDAD) was used to simultaneously identify and quantify mental cyanide complexes present in gold processing solutions. Identification was achieved using the “fingerprint” or full-scan UV spectrum (190 - 350 nm) obtained from the PDAD. The complexes identified so far were; Cu(CN)4 3− Fe(CN)6 4− Fe(CN)6 3− Co(CN)6 3− Ni(CN)4 2− Au(CN)2 − and Cr(CN)6 3−. Thiocyanate (SCN) was also detected and identified by this technique.

Monitoring the cyanidation of gold–copper ores with ion-interaction chromatography: Determination of the CN −:Cu(I) molar ratio

The use of reversed-phase ion-interaction high-performance liquid chromatography is reported for process monitoring of cyanide, thiocyanate and metallo–cyanide species in the gold cyanidation process. Thiocyanate and the metallo–cyanide complexes were monitored with a programmable, variable-wavelength UV detector mounted directly after the column, while cyanide, thiocyanate and the Cu(I) and Ag(I) cyanide complexes were monitored with a post-column reaction (PCR) detection system mounted after the UV detector. The eluent composition was optimised to enable separation of the major components of process samples in less than 10 min on a 5 cm column, but a 15 cm column was required for samples that contained Cu(I) at concentrations in excess of 300 ppm. The major emphasis during development of the eluent was to provide baseline resolution of thiocyanate and the Cu(I)–cyanide peaks and the optimal eluent composition consisted of 25% acetonitrile, 10 m M tetrabutylammonium hydroxide, 10 m M KH 2PO 4, 5 m M H 2SO 4 and 80–160 μ M NaCN, adjusted to pH 8.0. The level of cyanide present in the eluent was determined by the degree of separation required between thiocyanate and Cu(I). The PCR detection system enabled the cyanide:copper molar ratio, R, in process samples to be monitored routinely. R was determined from the peak area ratio of the cyanide and Cu(I)–cyanide peaks. The use of the peak ratio eliminated drift in the PCR detection system, providing a stable calibration for R. Good agreement was observed between standard analytical methods and the chromatographic method for determining R. Controlled cyanide leach tests showed that all the cyanide in the samples could be accounted for by the various cyano species detected. The proposed method has been utilised under field conditions on a working gold mine and has been shown to be very useful for determining cyanide losses and for monitoring the gold leaching process when treating cupriferous and pyritic gold ores.

Neural network and experimental design to investigate the effect of five factors in ion-interaction high-performance liquid chromatography

The effect of five experimental parameters on the ion-interaction chromatographic retention of pesticides characterized by different polarity was investigated by means of experimental design and artificial neural network treatments. The factors considered were: (1) the mobile phase pH; (2) N, the alkyl-chain length of the IIR (ion-interaction reagent); (3) CM, the organic modifier concentration in the mobile phase (4) CR, the concentration of IIR and (5) F, the flow-rate. The use of fractional design and Hoke design allowed useful information to be drawn about the retention mechanism involved and to build, through artificial neural network treatment (ANN), a model characterised by both descriptive and predictive ability. Four neurons and a bias unit were employed. The ANN proved to be a useful instrument in the optimisation of the chromatographic separation, as regards resolution and total analysis time: the experimental retention obtained in the optimal conditions always differed within 14% from the predicted ones.

Identification and determination of red dyes in confectionery by ion-interaction high-performance liquid chromatography

In this paper an ion-interaction HPLC method for the separation and determination of red dyes used to decorate home-made sweets is presented. In particular E122, E123 and E124 dyes are considered, with special interest for E123 (amaranth), prohibited by the US Food and Drug Administration in 1976 and by Italian legislation in 1977 and then again permitted (with restrictions) in the European Community in 1994, (94/36 EEC Directive). A RP-ODS stationary phase is used and an ion-interaction reagent (octylammonium phosphate) is added to the hydro-organic eluent [water-acetonitrile (30:70, v/v), pH = 6.4]. Spectrophotometric detection at 520 nm was employed and detection levels below 12 μg/l for the three dyes were achieved. The method is applied to two commercial products, respectively produced in France and in Italy, and the results are discussed in the light of the recent EEC Directive concerning food dyes.

Determination of creatinine in human serum. Statistical intercalibration of methods.

An intercalibration of methods was performed in the determination of creatinine in human plasma. Jaffè, enzymatic creatinine-deaminase (CRDI) and ion-interaction reagent high-performance liquid chromatography (HPLC) methods were employed and the results were compared using a parametric t-test for multiple samples and non-parametric Mann-Whitney, Wilcoxon and Friedman tests. The comparison did not show a statistically significant difference in the use of the three methods. The ion-interaction HPLC procedure is proposed as a reference method for creatinine assays.

Separation of metal-cyanide complexes by reversed-phase ion-interaction high-performance liquid chromatography

Reversed-phase ion-interaction chromatography has been used to study the separation of Cu(I), Ag(I), Ni(II), Au(I), Co(III), Fe(III), and Fe(II) cyano complexes on silica and carbon-based reversed-phases with UV detection at 215 nm. Separation of the metal-cyanide complexes was affected by a number of experimental factors including the nature and concentration of the organic modifier, nature and concentration of the ion-pairing reagent, pH, and ionic strength of the mobile phase. Differences between the elution order of metallo-cyanides observed in the present work and in other investigations under similar experimental conditions are explained by a difference in the ionic strength of the mobile phase. Finally, this technique is used to analyze metal-cyanide complexes in a gold mill solution.

Separation of food-related biogenic amines by ion-interaction reversed-phase high performance liquid chromatography. Tyramine, histamine, 2-phenylethylamine, tryptamine and precursor aminoacids. Application to red wine

Methods for the separation of food-related biogenic amines (histamine, tyramine, 2-phenylethylamine and tryptamine) have been developed based on ion-interaction reversed-phase liquid chromatography.

Reversed-phase ion-interaction chromatography of leukotrienes, lipoxins and related compounds

A novel mobile phase containing heptafluorobutyric acid has been used for ion-interaction high-performance liquid chromatography of leukotrienes, lipoxins and related compounds on octadecyl-silane silica columns. The use of a hydrophobic perfluorinated carboxylic acid as an ion-interaction agent at optimized concentration and pH permitted rapid, isocratic separation of leukotrienes, lipoxins and monohydroxyeicosatetraenoic acids. A completely volatile mobile phase suitable for preparative chromatography was obtained by using triethylamine as the base for pH adjustment. With this novel mobile phase, leukotrienes and monohydroxyeicosatetraenoic acids were completely separated in less than 15 min.

Determination of metal-cyano complexes by reversed-phase ion-interaction high-performance liquid chromatography and its application to the analysis of precious metals in gold processing solutions

A chromatographic procedure is described for the separation of metal-cyano complexes using reversed-phase ion-interaction high-performance liquid chromatography. Cyano complexes of copper(I), silver(I), iron(II), cobalt(III), iron(III), gold(I), palladium(II) and platinum(II) may be resolved on a C 18 column using a mobile phase of acetonitrile-water (23:77, v/v) containing 0.005 M tetramethylammonium hydroxide, with UV detection at 214 nm. Alternatively, cyano complexes of cobalt(III), copper(I), silver(I), nickel(II), palladium(II), gold(I) and platinum(II) may be resolved on a cyano column using a mobile phase of acetonitrile-water (8:92, v/v) containing 0.005 M tetramethylammonium hydroxide. The influence of variation of the nature of the ion-interaction reagent is discussed. The method is applied to the analysis of gold in processing solutions taken from the carbon-in-pulp process and excellent agreement between the chromatographic method and inductively coupled plasma atomic emission spectroscopic analysis is reported. Detection limits for precious metals were 1.9, 2.6, 0.9 and 0.8 ng for silver(I), gold(I), platinum(II) and palladium(II), respectively. Precision of the method for gold at the 0.1-μg level was 1.00% for peak area measurements and the calibration plot for gold was linear for injections containing up to 0.3 μg of gold.