Chiral Speciation Research Articles

Chiral speciation and determination of selenomethionine enantiomers in selenized yeast by ligand-exchange micellar electrokinetic capillary chromatography after solid phase extraction

A new phenylalanine derivative (l-N-(2-hydroxy-propyl)-phenylalanine, l-HP-Phe) was synthesized and its chelate with Cu(II) (Cu(II)–(l-HP-Phe)2) was used as the chiral selector for the ligand-exchange (LE) chiral separation of d,l-selenomethionine (SeMet) in selenized yeast samples by micelle electrokinetic capillary chromatography (MEKC). In order to improve the sensitivity of MEKC–UV, two-step preconcentration strategy was employed, off-line solid phase extraction (SPE) and on-line large volume sample stacking (LVSS). d,l-SeMet was first retained on the Cu(II) loaded mesoporous TiO2, then eluted by 0.1mL of 5molL−1 ammonia, and finally introduced for MEKC–UV analysis by LVSS injection after evaporation of NH3. With the enrichment factors of 1400 and 1378, the LODs of 0.44 and 0.60ngmL−1 for l-SeMet and d-SeMet was obtained, respectively. The developed method was applied to the analysis of d,l-SeMet in a certified reference material of SELM-1 and a commercial nutrition yeast, and the results showed that most of SeMet in the SELM-1 selenized yeast was l isomer and the recovery for l and d isomers in the spiked commercial nutrition yeast was 96.3% and 103%, respectively. This method is featured with low running cost, high sensitivity and selectivity, and exhibits application potential in chiral analysis of seleno amino acids in real world samples.

Chiral speciation in terrestrial pulmonate snails.

On the basis of data in the literature, the percentages of dextral versus sinistral species of snails have been calculated for western Europe, Turkey, North America (north of Mexico), and Japan. When the family of Clausiliidae is represented, about a quarter of all snail species may be sinistral, whereas less than one per cent of the species may be sinistral where that family does not occur. The number of single-gene speciation events on the basis of chirality, resulting in the origin of mirror image species, is not closely linked to the percentage of sinistral versus dextral species in a particular region. Turkey is nevertheless exceptional by both a high percentage of sinistral species and a high number of speciation events resulting in mirror image species. Shell morphology and genetic background may influence the ease of chirality-linked speciation, whereas sinistrality may additionally be selected against by internal selection. For the Clausiliidae, the fossil record and the recent fauna suggest that successful reversals in coiling direction occurred with a frequency of once every three to four million years.

Enantioselective determination of thyroxine enantiomers by ligand‐exchange CE with UV absorbance and ICP‐MS detection

A simple CE method has been developed for the separation and determination of thyroxine (T4) enantiomers in pharmaceutical formulations. The method was based on ligand-exchange mechanism using a Cu(II)/L-proline complex as chiral selector. The effects of different parameters affecting separation such as chiral selector concentration, organic additive, buffer pH and temperature were investigated. A baseline separation of the two enantiomers was obtained at a Cu(II)/L-proline ratio of 1:8 in a borate buffer (15 mmol/L, pH 9.6) containing 10% v/v acetonitrile. Under the optimized conditions, precision linearity range and detection limits of the developed enantioselective CE method were evaluated and compared using two different detection systems: conventional UV detection at 226 nm and iodine (127I)specific detection ("chiral speciation") with ICP-MS. Both methodologies show adequate analytical performance characteristics with detection limits around 0.30 microg/mL for each enantiomer of T4. Finally, a levothroid pharmaceutical formulation sample was successfully analyzed using both developed methods CE-UV and CE-ICP-MS.

First approach of a methodological set‐up for selenomethionine chiral speciation in breast and formula milk using high‐performance liquid chromatography coupled to atomic fluorescence spectroscopy

AbstractThe chiral speciation of selenomethionine in breast and formula milk based on species separation by high‐performance liquid chromatography followed by online microwave‐assisted digestion and detection with hydride generation atomic fluorescence spectrometry (HPLC‐MAD‐HG‐AFS) requires severe sample manipulation to avoid matrix influence. Sample clean‐up for fat and protein elimination using centrifugation and ultrafiltration was optimized, and selenomethionine preconcentration based on cation exchange solid‐phase extraction was studied and optimized. The resulting procedure is suitable for chiral selenium speciation in infant milk with detection limits of 3.1 and 3.5 ng ml−1 as Se for L‐selenomethionine and D‐selenomethionine, respectively. The time necessary for the analysis, about 90 min, including sample clean‐up, analyte preconcentration and chromatographic separation, makes the approach suitable for routine analysis. Copyright © 2007 John Wiley & Sons, Ltd.

Selenium speciation studies in Se-enriched chives ( Allium schoenoprasum) by HPLC-ICP–MS

In this study, three liquid chromatographic techniques were employed to better understand selenium species distribution in Chives ( Allium schoenoprasum) separately grown in three different supplementation media Se(IV), Se(VI), and SeMet. The highest selenium accumulation up to 700 μg Se g −1 was observed in the case of the Se(VI)-enriched samples on the basis of total selenium measurements. Size-exclusion chromatography (SEC-HPLC) was performed for investigation of selenium containing proteins in chives. For speciation of selenium containing amino acids, reversed-phase ion-pairing chromatography (RP-IP-HPLC) and for enantiomeric separations, a crown ether column was used. In all three cases online detection with inductively coupled plasma mass spectrometry was performed for selenium specific detection. Two extractions (perchloric acid–ethanol and enzymatic) were carried out on chive samples. Speciation analysis on the chives grown in three different media revealed that selenium distribution among different forms of amino acids in the sample strongly depends on the type of enrichment employed. Enrichment with Se(VI) leads to accumulation of selenium in inorganic forms, while in case of Se(IV) and SeMet-enriched samples, methyl-selenocysteine and selenocystine were found to be present. Not surprisingly, chiral speciation revealed the presence of the l-enantiomeric forms of selenoamino acids in the sample. The major enantiomer found in the perchloric acid–ethanol extracts was l-MeSeCys, while in the enzymatic extracts l-SeMet was also detected.

SPME-enantioselective gas chromatography with ECD and ICP-MS detection for the chiral speciation of the pesticide ruelene in environmental samples

Most chiral pesticides are used as racemates despite the fact that the pesticidal activity is generally due to just one enantiomer while the other one may have toxic effects for nontarget organisms. Accordingly, a separation of the two enantiomers of the organophosphorus pesticide ruelene by capillary gas chromatography (CG) has been developed, using a commercial chiral column (CP-Chirasil-Dex CB) and a temperature program from 50 °C (held for 1 min) to 190 °C at 40 °C min−1 with He as carrier gas. This enantioselective gas chromatographic separation of ruelene can be enhanced by a previous solid-phase microextraction (SPME) sample preparation. Experimental SPME conditions such as extraction time, extraction temperature, salt additives and pH have been optimised. The developed SPME-enantioselective GC procedure has been evaluated using two different detection systems: a classical electron capture detection (ECD) and a P-specific detection with inductively coupled plasma mass spectrometry (ICP-MS). The ICP-MS detector has proved to be superior to the ECD in terms of selectivity and sensitivity with excellent detection limits of 27 ng L−1 (3 ng L−1 as P) for each enantiomer of ruelene. The final determination of enantiomers of ruelene in different spiked samples (river water, red wine, orange and tomato juices) has been worked out. The general advantages of ICP-MS detection for chiral pesticides determinations in complex environmental samples are discussed.

Chiral Speciation and Determination of dl-Selenomethionine Enantiomers on a Novel Chiral Ligand-Exchange Stationary Phase

A new type of chiral ligand-exchange stationary phase (CLES) was successfully synthesized by treating silica gel with beta-(3,4-epoxycyclohexyl)ethyltrimethoxy silane and opening the epoxy ring by L-isoleucine. The chiral speciation of DL-selenomethionine (DL-SeMet) by high-performance liquid chromatography (HPLC) with UV absorbance on the CLES column was studied. The influences of the contents of copper ion and methanol as well as the pH value in the mobile phase and temperature of the column on the efficiency of resolution of DL-SeMet were investigated in detail. DL-SeMet could be completely resolved within 40 min under the optimal operating conditions of 0.1 mmol/L Cu2+ at 0.05 mol/L KH2PO4 buffer (pH = 5.5) and 35 degrees C temperature of the column. The limits of detection of D- and L-SeMet were 255 ppb and 286 ppb, respectively. This method was applied to determine the D- and L-enantiomers of DL-SeMet in real samples, such as selenized yeast powder and garlic.

Comparative study of the instrumental couplings of high performance liquid chromatography with microwave-assisted digestion hydride generation atomic fluorescence spectrometry and inductively coupled plasma mass spectrometry for chiral speciation of selenomethionine in breast and formula milk

A comparison of chiral separation and analysis of selenomethionine in breast and formula milk, using high performance liquid chromatography (HPLC) on a glycopeptide teicoplanin-based chiral stationary phase (Chirobiotic T), coupled to atomic fluorescence spectrometry (AFS) and inductively coupled plasma (ICP) MS detectors has been performed. The coupling HPLC-microwave-assisted digestion hydride generation requires on-line post-column analytes treatment, and a severe sample clean-up for fat and proteins elimination using centrifugation and ultrafiltration. Underivatized dl-selenomethionine enantiomers were completely resolved in 10 min using unbuffered water mobile phase at 1 ml min −1 flow. Good selectivity and sensitivities (detection limits 3.1 and 3.5 ng ml −1 as Se for l- and d-selenomethionine, respectively) were obtained, and method robustness and simplicity, together to the low cost of AFS detector, makes it suitable for infant milk routine analysis. HPLC–ICP-MS coupling exhibits very low detection limits (0.9 ng ml −1, as Se) for each dl-selenomethionine enantiomers, but the method suffers from matrix influence, that produces a poor S/N ratio and low reliability. The methods were applied to breast and formula milk samples with recoveries of 80% of the total selenium presence, which is attributable to the existence of other unknown species. l-Selenomethionine was the only isomer present in breast milk, but a 30% of d-selenomethionine was also detected in formula milk.

Chiral trace-element speciation in biological samples: present importance and application to speciation for seleno-amino acids

Chiral trace-element speciation, the recognition of the particular enantiomer where the trace element sought is present, is restricted so far virtually to speciation of seleno-amino acids. The present importance of such studies, in general, and of chiral speciation, in particular, is highlighted in this article by comparing existing chiral speciation methods for seleno-amino acids. Usually these methods rely on hybrid techniques, where an adequate separation technique, such as high-performance liquid chromatography (HPLC) or gas chromatography (GC), is coupled on-line with element (selenium)-specific detection by inductively coupled plasma mass spectrometry (ICP-MS). Reported chiral speciation studies of seleno-amino acids in real biological samples, such as selenized yeast, are reviewed.

Chiral speciation of Marfey's derivatizedDL-selenomethionine using capillary electrophoresis with UV and ICP-MS detection

The chiral speciation of DL-selenomethionine by capillary electrophoresis (CE) with UV absorbance and inductively coupled plasma mass spectrometry (ICP-MS) is described. Chiral derivatization of DL-selenomethionine with 1-fluoro-2,4-dinitrophenyl-5-L-alanine amide (Marfey's reagent) forms diastereomers that are separated with CE in reversed polarity. Three CE buffer systems were evaluated using UV absorbance detection. A 30 mM ammonium phosphate buffer at pH 3.3 was chosen for good resolution of the D- and L-forms in under 14 min. Limits of detection for D- and L-selenomethionine were 250 ppb (ng mL−1) using CE-UV, and 50 ppb using CE-ICP-MS. The migration time reproducibility of both detection limits was comparable (∼2%). This method was applied to the qualitative chiral identification of selenomethionine in selenized yeast digested with proteinase K (enzymatic hydrolysis).

Hybridation of different chiral separation techniques with ICP‐MS detection for the separation and determination of selenomethionine enantiomers: chiral speciation of selenized yeast

Enantioseparation and determination of selenomethionine enantiomers in selenized yeast was investigated using chiral separation techniques based on different principles, coupled on-line to inductively coupled plasma mass spectrometry (ICP-MS) for selenium-specific detection. High performance liquid chromatography (HPLC) on a beta-cyclodestrin (beta-CD) column, cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC), gas chromatography (GC) on a Chirasil-L-Val column, and HPLC on a Chirobiotic T column have been investigated as the chiral separation techniques. For HPLC separation on the beta-CD column, and also for CD-MEKC, selenomethionine enantiomers were derivatized with NDA/CN(-). For chiral separation by GC, selenomethionine enantiomers were converted into their N-trifluoroacetyl (TFA)-O-alkyl esters. The developed hybridation methodologies are compared with respect to enantioselectivity, sensitivity and analysis time. The usefulness of the best-suited method [HPLC (Chirobiotic T)-ICP-MS] was demonstrated by its application to the successful chiral speciation of selenium and D-and L-selenomethionine content determination in selenized yeast.

Chiral speciation of trace elements: approaches to the speciation of selenoaminoacid enantiomers in biological samples

The “state of the art” of chiral trace element speciation, that is, the recognition of the particular enantiomer in which the trace element sought is occurring, is given here. The importance of such studies and speciation information is highlighted. Recent advances and trends in the development and applications of analytical approaches for chiral speciation of trace elements in biological material are also reviewed. A comparative overview is given of existing selenoaminoacids chiral speciation methods, based on hybridization of an adequate chiral separation technique (e.g., high-performance liquid chromatography, gas chromatography or capillary electrophoresis) coupled on-line with selenium specific detection by inductively coupled plasma mass spectrometry (ICP-MS). Applications of these hybrid methodologies, reported so far in the literature, to selenoaminoacids chiral speciation in biological real samples are also reviewed.

Chiral speciation and determination of selenomethionine enantiomers in selenized yeast by HPLC-ICP-MS using a teicoplanin-based chiral stationary phase

Chiral separation of underivatized selenoaminoacids enantiomers by high performance liquid chromatography (HPLC) on a glycopeptide antibiotic teicoplanin-based chiral stationary phase (Chirobiotic T) was investigated. The effect of methanol content, pH and ionic strength of the mobile phase on retention and resolution of the enantiomers was examined to optimize the chromatographic conditions. Underivatized DL-selenomethionine and DL-selenoethionine enantiomers were completely resolved within 8 min using a 2% (v/v) methanol–water (unbuffered) mobile phase delivered at 1 mL min−1. Very good selectivity and excellent detection limits of 1.9 µg L−1 (0.8 µg L−1 as Se) for each DL-selenomethionine enantiomer were obtaining by on-line coupling the chiral HPLC separation with selenium-specific detection by inductively coupled plasma quadrupole mass spectrometry (ICP-MS). The HPLC-ICP-MS method was successfully applied to the chiral speciation and determination of D- and L- selenomethionine enantiomers in real samples of selenized yeast.

Chiral speciation of selenoamino acids and selenium enriched samples using HPLC coupled to ICP-MS

The chiral separation of nine selenoamino acids using a crown ether HPLC column coupled to ICP-MS for detection is described. Three different temperatures were investigated for the separation of the enantiomers of individual amino acids and for a mixture of eight selenoamino acids. Although complete resolution was not achieved, better resolution of the mixture was obtained at higher temperatures. Selenium enriched onion, garlic and yeast were also analyzed. Two different extraction methods for the samples are compared; water extraction and a pepsin enzymatic extraction. Some of the selenoamino acid enantiomers are identified for the different extraction methods. Comparison is also made with earlier reported ion pairing separations for some of these selenoamino acids.

Selenomethionine chiral speciation in yeast and parenteral solutions by chiral phase capillary gas chromatography-ICP-MS

Chiral resolution and speciation of DL-selenomethionine enantiomers by capillary gas chromatography (GC) as N-trifluoroacetyl (TFA)-O-isopropyl derivatives using an l-valine-tert-butylamide modified polydimethylsiloxane chiral stationary phase (Chirasil-L-Val) were investigated. Good resolution was achieved using a temperature program from 100 °C (held for 3 min) to 160 °C at 1.5 °C min –1 and He as carrier gas. Very good selectivity and excellent detection limits of 0.25 µg L –1 (0.1 µg L –1 as Se) for each enantiomer were obtained by on-line coupling of the chiral capillary column with selenium-specific detection by inductively coupled plasma mass spectrometry (ICP-MS). Experimental parameter optimization is described in detail. This optimised GC-ICP-MS method was successfully applied to the determination of the optical purity of l-selenomethionine in commercial samples, to the determination of the enantiomeric ratio of selenomethionine in parenteral solutions used for human use and also to chiral Se speciation in selenized yeast.