Simultaneous determination of alcohols including diols and triols by HPLC with ultraviolet detection based on the formation of a copper(II) complex.

We developed a reversed-phase high-performance liquid chromatography method with ultraviolet detection using on-line complexation with Cu(II) ion for analysis of five alcohols including diols and triol (methanol, ethanol, 1,2-propanediol, 1,3-propanediol, and glycerol). The Cu(II) ion concentration in the mobile phase had a great influence on the peak areas of these alcohols, but not on their retention times. Column temperature (25-40°C) and pH of the mobile phase did not affect the separation of analytes. The optimum separation conditions were determined as 5 mM CuSO4 , 3 mM H2 SO4 , and 3 mM NaOH at 30°C. The ratio of the peak areas for three alcohols (methanol, 1,2-propanediol, and glycerol) was in good agreement with that calculated from the obtained stability constants, molar absorption coefficients for the 1:1 Cu(II) complexes with the three alcohols, and the injected molar quantities. This fact strongly suggests that the observed high-performance liquid chromatography signals resulted from formation of the 1:1 Cu(II)-alcohol complexes. Using the proposed method, these five alcohols in spirit, liquid for electronic cigarette, mouthwash, and nail enamel remover samples were successfully analyzed with only a simple pretreatment.

Separation of inorganic anions on reversed-phase C18 columns with a phosphomolybdate mobile phase.

Reversed-phase high performance liquid chromatography (RP-HPLC) is the most widely used chromatographic method. In addition to hydrophobic interactions, additional interactions such as electrostatic interactions may participate in the retention behaviour of an analyte. This makes it possible to use RP-HPLC for many types of analyte. We describe a simple method for separating inorganic anions on a C18 column, in which retention of inorganic anions is almost entirely due to electrostatic interactions. This leads to rapid separations as well as higher theoretical plate numbers. We used 2 mM phosphoric acid containing a low concentration of disodium molybdate as the mobile phase, which allows UV detection of non-UV-absorbing anions. With this method, we determined eight inorganic anions including several non-UV-absorbing anions photometrically at 220 nm. The detection limits of the examined eight inorganic anions calculated at a signal-to-noise ratio of 3 were between 0.3 and 10 µM. The detector response was linear over three orders of magnitude of inorganic anion concentration. The proposed RP-HPLC/UV method was successfully applied to determine inorganic anions in some water samples.

Determination of α-hydroxy acids and their enantiomers in fruit juices by ligand exchange CE with a dual central metal ion system.

The content of α-hydroxy acids and their enantiomers can be used to distinguish authentic and adulterated fruit juices. Here, we investigated the use of ligand exchange CE with two kinds of central metal ion in a BGE for the simultaneous determination of enantiomers of dl-malic, dl-tartaric and dl-isocitric acids, and citric acid. Ligand exchange CE with 100 mM d-quinic acid as a chiral selector ligand and 10 mM Cu(II) ion as a central metal ion could enantioseparate dl-tartaric acid but not dl-malic acid or dl-isocitric acid. Addition of 1.8 mM Sc(III) ion to the BGE with 10 mM Cu(II) ion to create a dual central metal ion system permitted the simultaneous determination of these α-hydroxy acid enantiomers and citric acid. The proposed ligand exchange CE was thus well suited for detecting adulteration of fruit juices.

Direct enantioseparation of lipoic acid in dietary supplements by capillary electrophoresis using trimethyl-ß-cyclodextrin as a chiral selector.

Lipoic acid, an antioxidant, naturally occurs as the (R)-enantiomer, while synthetic lipoic acid is racemic. It is thus of interest to know the (R)-enantiomer content of lipoic acid supplements. Here, we used capillary electrophoresis to directly enantioseparate lipoic acid in dietary supplements by using a sulfonated capillary with an effective voltage of +18 kV and direct detection at 200 nm. Factors affecting migration time and resolution of lipoic acid were investigated. The optimum background electrolyte was found to be 100 mM phosphate buffer (pH 7.0) containing 8 mM trimethyl-ß-cyclodextrin as a chiral selector at 20°C. Under the proposed conditions, direct chiral resolution of lipoic acid in dietary supplements was conducted successfully.

Enantioseparation of α-hydroxy acids by chiral ligand exchange CE with a dual central metal ion system.

Using two kinds of central metal ions in a background electrolyte, ligand exchange CE was investigated for the simultaneous enantioseparation of dl-malic, dl-tartaric, and dl-isocitric acids. Ligand exchange CE with 100 mM d-quinic acid as a chiral selector ligand and 10 mM Cu(II) ion as a central metal ion could enantioseparate dl-tartaric acid but not dl-malic acid or dl-isocitric acid. A dual central metal ion system containing 0.5 mM Al(III) ion in addition to 10 mM Cu(II) ion in the background electrolyte enabled the simultaneous enantioseparation of the three α-hydroxy acids. These results suggest that the use of a dual central metal ion system can be useful for enantioseparation by ligand exchange CE.

Metal(II)-ligand molar ratio dependence of enantioseparation of tartaric acid by ligand exchange CE with Cu(II) and Ni(II)-D-quinic acid systems.

Enantioseparation of tartaric acid by ligand exchange CE with a Cu(II)-D-quinic acid system was studied. Racemic tartaric acid was enantioseparated by ligand exchange CE using BGEs containing relatively low Cu(II)-D-quinic acid molar ratios ranging from 1:1 to 1:3 and high molar ratios ranging from 1:8 to 1:12 but was not enantioseparated using BGEs with medium molar ratios ranging from 1:4 to 1:6. While the migration order of D-tartaric acid was prior to L-tartaric acid at the lower Cu(II)-D-quinic acid molar ratios, the enantiomer migration order was reversed at the higher molar ratios. These results were compared with those for Ni(II)-D-quinic acid system. The molar ratio dependence of enantiomer migration order can be attributed to a change in the coordination structure of Cu(II) ion with D-quinic acid.

Enantioseparation of DL-isocitric acid by a chiral ligand exchange CE with Ni(II)-D-quinic acid system.

The ratio of citric acid to D-isocitric acid can be used to distinguish authentic and adulterated fruit juices. To separate DL-isocitric acid enantiomers, we used ligand exchange CE. D-Quinic acid was used as a chiral selector ligand and Mn(II), Fe(III), Co(II), Ni(II), Cu(II), and Zn(II) ions were used as the central ions of the chiral selector in the BGE. DL-Isocitric acid was found to be enantioseparated with the above metal ions except for Mn(II) ion. The optimum running conditions for the analysis of D- and L-isocitric acids along with citric acid, an isomer of isocitric acid, were found to be a BGE (pH 5.0) containing 30% ACN, 20 mM acetic acid, 20 mM NiSO(4), and 80 mM D-quinic acid. Under these conditions, DL-isocitric and citric acids in fruit juices were analyzed successfully.

Separation of adenine, adenosine-5'-monophosphate and adenosine-5'-triphosphate by fluidic chip with nanometre-order diameter columns inside porous anodic alumina using an aqueous mobile phase.

Adenine, adenosine-5'-monophosphate and adenosine-5'-triphosphate were separated by fluidic chip with PAA membranes using an aqueous mobile phase.

Enantioseparation of nicotine alkaloids in cigarettes by CE using sulfated beta-CD as a chiral selector and a capillary coated with amino groups.

Nicotine (NC) and its related compounds (cotinine (CN), nornicotine (NN), anatabine (AT) and anabasine (AB)) were simultaneously enantioseparated by CE using a capillary with amino groups and sulfated beta-CD as a chiral selector. The optimum running conditions were found to be 30 mM acetate buffer (pH 5.0) containing 8% sulfated beta-CD with an applied voltage of +15 kV at 30 degrees C using direct detection at 260 nm. Using a capillary coated with amino groups, the EOF migrates toward the positive pole. However, when sulfated beta-CD was added to the BGE, it was found that the EOF migrated toward the negative pole due to ionic adsorption of sulfated beta-CD to amino groups on the capillary inner wall. All the cationic analytes migrated as anions, suggesting that they formed stable anionic complexes with sulfated beta-CD. With this system and a simple pretreatment with mini-cartridges, NC alkaloids in five cigarette samples were enantioseparated. As a result, each of the compounds except for CN was detected. In the case of NC, only (S)-NC was detected (more than 99.9%), but in the case of NN, AT and AB, the ratios of (S)-isomer to total isomers were in the ranges 58-70, 81-85 and 59-65%, respectively. On the other hand, only NC was detected in cigarette smoke and the ratio of (S)- and (R)-NCs was 96:4. The amounts of NC alkaloids in cigarettes suggest that the production of (R)-NC resulted from racemization due to the high temperature/burning of the cigarette.

Study of solid-phase extraction for the determination of sequestering agents in river water by high-performance liquid chromatography.

Anion-exchange solid-phase extraction accompanied with high-performance liquid chromatography has been developed for the determination of six kinds of aminopolycarboxylic acids (APCAs) in river water [N-(2-hydroxyethyl)ethylenediaminetriacetate (HEDTA), ethylenediaminetetraacetate (EDTA), 1,3-propanediaminetetraacetate (PDTA), diethylenetriaminepentaacetate (DTPA), 1,2-propanediaminetetraacetate (MeEDTA), and O,O'-bis(2-aminoethyl)ethyleneglycoltetraacetate (GEDTA)]. The enrichment of APCAs using an anion-exchange cartridge was successfully done by the removal of anions, which competed with APCAs in anion-exchange processes. Barium chloride solution was added to river water and the mixture was passed through On Guard II Ag and H cartridges and then a Bond Elut Jr.SAX cartridge to enrich APCAs. After elution, APCAs were analyzed on two reversed phase C30 columns connected in series and detected with ultraviolet detection. The enrichment using solid-phase extraction permitted the determination of APCAs in river water at concentrations as low as 1nM. Good recoveries (83-111%) were obtained for each APCA by the standard addition method on three river water samples with high accuracy (RSD 1.8-9.5%). Applying this method, two kinds of APCAs, EDTA and DTPA, were determined in samples from the Oyabe and Senbo Rivers in Japan.

Reversed phase ion-pair chromatographic separation of sugar alcohols by complexation with molybdate ion.

In this study, we developed a simple and sensitive reversed phase ion-pair chromatographic method for the analysis of C4-C6 sugar alcohols. The method is based on the on-line complexation of sugar alcohols with molybdate ion. The resulting dinuclear anionic complexes can be separated on a reversed-phase C18 column using tetrabutylammonium chloride as an ion-pairing reagent. The mobile phase (pH 3.1) consisted of 0.1 mM disodium molybdate, 1 mM hydrochloric acid and 0.4 mM tetrabutylammonium chloride - 10% v/v methanol. By complexing with molybdate ion, sugar alcohols can be detected by their UV absorption at 247 nm with high resolution and sensitivity. The quantification limits of the examined sugar alcohols calculated at S/N = 10 were 0.1 mM for erythritol and xylitol and 0.01 mM for arabitol, sorbitol, mannitol and dulcitol. The detector response was linear over three orders of magnitude of sugar alcohol concentration. The proposed method was successfully applied to measure sugar alcohols in health drinks, eyedrops and mouthwashes.

Use of porous anodic alumina membranes as a nanometre-diameter column for high performance liquid chromatography.

The possibility of using porous anodic alumina membranes as a column for normal-phase high performance liquid chromatography was evaluated using phenol and toluene with mobile phases having different solvent compositions.

Determination of sequestering agents in cosmetics and synthetic detergents by high-performance liquid chromatography with ultraviolet detection.

Using a fast reversible reaction of aminopolycarboxylic acids (APCAs) into Fe(III)-APCA complexes in the presence of Fe(III) ions, seven kinds of APCAs [nitrilotriacetate (NTA), N-(2-hydroxyethyl)ethylenediamine-triacetate (HEDTA), ethylenediamine-tetraacetate (EDTA), 1,3-propanediamine-tetraacetate (PDTA), diethylenetriamine-pentaacetate (DTPA), 1,2-diaminopropane-tetraacetate (MeEDTA), and O,O'-bis(2-aminoethyl)ethyleneglycol-tetraacetate (GEDTA)] in cosmetics and synthetic detergents were separated on two reversed-phase C30 columns connected in series and detected with ultraviolet detection. Simple pretreatment, consisted of thousand times dilution of samples and addition of 100 microl of the Fe(III) solution containing 10 mM Fe(III) chloride and 0.5 M sulfuric acid to 10 ml of diluted samples, permitted the determination of APCAs in cosmetics and synthetic detergents at concentration level of 0.1 mM, except 0.3 mM for GEDTA. APCAs except GEDTA could be detected at concentration level of 0.03 mM and GEDTA could be detected at concentration level of 0.09 mM. Good recoveries (95-110%) were obtained for each APCA by the standard addition method on two diluted samples with high accuracy (RSD 0.2-9.1%). Three APCAs (EDTA, HEDTA and NTA) were detected in various concentrations in cosmetics and synthetic detergents and the other APCAs were not detected in any of the samples. This method requires no tedious pretreatment and takes only 15 min for one analysis, so it is useful for determination of APCAs.

Enantioseparation of isoxanthohumol in beer by hydroxypropyl-gamma-cyclodextrin-modified micellar electrokinetic chromatography.

Chiral resolution of isoxanthohumol (IX) in beer samples was performed by hydroxypropyl-gamma-cyclodextrin-modified micellar electrokinetic chromatography. The optimum running conditions were found to be 20 mM phosphate buffer (pH 7.0) containing 45 mM hydroxypropyl-gamma-cyclodextrin and 100 mM sodium dodecyl sulfate with an effective voltage of +20 kV at 20 degrees C using direct detection at 210, 295, and 370 nm. IX was detected in 12 beer samples and the total levels of (+)- and (-)-IX ranged from 0.15 to 1.4 mg/L. But the amount of xanthohumol (XN) was below the detection limit (0.017 mg/L). Each level of (-)-IX was almost the same as that of (+)-IX, suggesting that IX was a racemic mixture in these beer samples. The racemization of IX in beer could be attributed to the production of a racemic mixture from XN during boiling and to the fact that IX enantiomers were easily interconverted.

Determination of hexitols by reversed phase liquid chromatography using on-line complexation with molybdate ion.

A new approach is proposed to determine three hexitols (mannitol, sorbitol and dulcitol) by reversed phase liquid chromatography and ultraviolet detection. When molybdate ion is added to the mobile phase, it forms complexes with hexitols that can be separated on a reversed phase C30 column and detected by UV absorption at 247 nm. The mobile phase (pH 3.1) consisted of 0.1 mM disodium molybdate and 1 mM phosphoric acid. Other sugar alcohols, such as erythritol and xylitol, and glucose could not be detected under these conditions. The quantification limits of the examined three hexitols calculated at S/N = 10 were 0.001 mM and the detector response was linear in the range 0.001-0.3 mM. The method successfully measured these hexitols in candy samples, and the results obtained by the proposed method agreed well with those obtained by enzymatic methods.

Permeation flux of organic molecules through silica-surfactant nanochannels in a porous alumina membrane.

The permeation fluxes of phenol, benzene sulfonate (BS) and benzene disulfonate (BDS) through a porous anodic alumina membrane with the perpendicularly oriented silica-surfactant nanochannel assembly membrane (NAM) were measured in water-ethanol mixture media. The permeation flux depended on solute charges and on solvent composition. As the ethanol ratio increased, the fluxes of BS and BDS increased and the flux of phenol decreased. The results of extraction/elution experiments also depended on the solute charges and the solvent composition. Chromatographic experiments in n-hexane showed that dipole and hydrophobic interactions affect the retention of solutes. Permeation of the solute across the NAM in water-ethanol mixture is likely to be determined by various factors such as dipole interaction, hydrophobic interaction, solvation, and anion-exchange efficiencies.

Reversed phase liquid chromatographic determination of organic acids using on-line complexation with copper(II) ion.

We describe a simple and sensitive liquid chromatographic method for the analysis of organic acids using on-line complexation with copper(II) ion. Organic acids complexed with copper(II) ion were separated on a reversed-phase C18 column and detected by UV absorption at 240 nm. The copper(II) ion concentration in the mobile phase had a great influence on separation and sensitivity. A mobile phase consisting of 10 mM copper(II) sulfate in 5 mM sulfuric acid (pH 2.3) was used to separate nine organic acids (tartaric, malic, malonic, lactic, acetic, citric, maleic, succinic and fumaric acids). The detection limits of the examined organic acids calculated at S/N = 3 ranged from 0.6 to 100 µM. The detector signal was linear over three orders of magnitude of organic acid concentration. The method successfully measured organic acids in juice and vinegar samples.

Simultaneous and sensitive analysis of aliphatic carboxylic acids by ion-chromatography using on-line complexation with copper(II) ion.

A new approach to ion chromatography is proposed to improve the UV detection of aliphatic carboxylic acids separated by anion-exchange chromatography. When copper(II) ion added to the mobile phase, it forms complexes with carboxylic acids that can be detected at 240 nm. The absorbance was found to increase with increasing copper(II) ion concentration. The retention times of α-hydroxy acids were also found to depend on the copper(II) ion concentration. Addition of acetonitrile to the mobile phase improved the separation of aliphatic carboxylic acids. The detection limits of the examined carboxylic acids (formate, glycolate, acetate, lactate, propionate, 3-hydroxypropionate, n-butyrate, isobutyrate, n-valerate, isovalerate, n-caproate) calculated at S/N=3 ranged from 0.06 to 3 µM. The detector signal was linear over three orders of magnitude of carboxylic acid concentration. The proposed method was successfully applied to analyze aliphatic carboxylic acids in rainwater and bread.

Low-cost Methods for Making 3D Fluidic Polymer and Glass Chips Using Metal Templates.

Microfluidics is a rapidly growing field in which small volumes of liquid are moved through channels in a large variety of applications. Fabricating such channels can be expensive. Here, we describe an inexpensive method for making 3D channels in fluidic chips by using a sacrificial template made of coated metal wire or metal tubes. A 3D template is embedded in polymer or glass and then dissolved, leaving channels in the chip, without the need for expensive instruments. By changing the mold, chips of various shapes can be made.

Determination of ethylenediaminetetraacetic acid in foods by reversed-phase high-performance liquid chromatography.

A convenient HPLC method for the quantitative determination of EDTA in foods was developed. EDTA in food samples was easily extracted with water by ultrasonication. After converting to Fe(III) complex in the presence of Fe(III) ions, EDTA was separated on a reversed-phase C30 column and detected with ultraviolet detection (260 nm). Citrate and malate, which are present in many foods, also formed Fe(III) complexes but they did not interfere the chromatographic detection of EDTA. The method allowed determination of EDTA in foods at concentrations as low as 0.01 mmol/kg. Good recoveries (95.2-101%) were obtained by the standard addition method on four samples with high repeatability (RSD, 0.8-3.4%). The method was successfully applied to the analysis of EDTA in carbonated drinks, jellies, canned beans, canned corn and food supplements.