Enantiomers In Samples Research Articles

HPLC separation of panthenol enantiomers on different types of chiral stationary phases

Abstract Panthenol is a biologically active compound closely related to vitamin B5 (pantothenic acid). This work deals with the separation of panthenol enantiomers using high performance liquid chromatography. Different types of chiral stationary phases (β-cyclodextrin, isopropyl carbamate cyclofructan 6, amylose tris(3,5-dimethylphenylcarbamate)) were tested in normal phase separation mode. Effect of mobile phase composition on the resolution and retention of enantiomers was studied. Two types of detectors, low-wavelength UV and polarimetric, were used. The optimal chromatographic system includes a chiral stationary phase based on amylose and a mobile phase of hexane/ethanol (60/40, v/v) where the resolution of enantiomers reached the value R s = 2.49. Suitable chromatographic conditions were applied for the determination of panthenol enantiomers in samples of pharmaceutical preparations with the obtained recovery of more than 92 %. Linearity of the high performance liquid chromatography method with spectrophotometric detection was from 1.0 × 10−3 to 1.3 mg mL−1 (R 2 = 0.998), with the limit of detection of 0.3 × 10−3 mg mL−1 for both enantiomers.

Enzyme assay for d-amino acid oxidase using optically gated capillary electrophoresis-laser induced fluorescence detection

Because d-amino acids (AAs) play an essential role in the regulation of many processes in living cells, detection of D-AAs and assay of d-amino acid oxidase (DAAO) activity are of vital importance in bioanalytical science. However, the reliability and accuracy of DAAO assays could be interfered due to the facts that DAAO presents broad substrate activity towards different D-AAs and there could be abundant L-AA enantiomers in biological samples. In this study we presented the first application of optically gated capillary electrophoresis with LIF detection (OGCE-LIF) for efficient assay of DAAO activity. High repeatability of the OGCE-LIF assay of amino acids (AAs) was achieved with relative standard deviation (RSD) (n = 15) less than 1.5% and 2.7% for migration time and peak height, respectively. Under the optimal OGCE-LIF conditions, five pairs of D/L-AA enantiomers were efficiently separated in less than 1 min with low limit of detection of 1.3 μM. Enzymatic assays of DAAO were successfully achieved by detection the substrate consumption with OGCE-LIF, for either single or mixed AA substrates. Kinetic analysis of the parallel oxidation reactions of two different substrates was performed, which was in good agreement with the experimental results. Our study indicates OGCE-LIF can perform rapid and efficient separation of mixed pairs of AA enantiomers and is a promising method for quantitatively assaying DAAO catalyzed reaction with the presence of L-AA enantiomers in the sample. Our study would pave the way for accurate determination of D-AAs and DAAO enzymes in complicated biological samples.

Effects of the dynamic modification of stationary phases by sorbates in gas chromatography: The possibility of separating enantiomers in achiral systems

It is shown that the gas chromatographic separation of enantiomers on columns with achiral nonpolar stationary phases is principally possible as a result of the dynamic modification of stationary phases by sorbates under analysis. It is found that a number of key characteristic features is intrinsic to such separation: it can be only partial, it does not occur for all chromatographic columns, and it is observed only for some compounds and only within narrow ranges of quantities of sorbates that are close to the limits of mass overload of chromatographic systems. These characteristic features are illustrated by the examples of separating (1R,5R)-(+)- and (1S,5S)-(−)-α-pinenes on a WCOT column with an RTX-5 phase. The main characteristic feature of the separation of enantiomers as a result of the dynamic modification of stationary phases is the nonconformity of peaks in chromatograms with two individual enantiomers, compared to other ways and means for their separation; the first eluting peak belongs to the enantiomer that predominates in a mixture irrespective of its configuration, while the second peak corresponds to the racemic mixture of enantiomers; i.e., the ratio of peak areas in chromatograms does not correspond to the actual ratio of enantiomers in samples under analysis and is strongly distorted as a result of their incomplete separation. It is concluded that the separation of racemic mixtures in achiral systems is fundamentally impossible under any conditions, and this is one of the key criteria of the validity of the considered concept as a whole.

Analysis of free fractions for chiral drugs using ultrafast extraction and multi-dimensional high-performance affinity chromatography

A multi-dimensional chromatographic approach was developed to measure the free fractions of drug enantiomers in samples that also contained a binding protein or serum. This method, which combined ultrafast affinity extraction with a chiral stationary phase, was demonstrated using the drug warfarin and the protein human serum albumin.

Enantioselective Degradation of Indoxacarb in Cabbage and Soil under Field Conditions

The enantioselective degradation of indoxacarb in cabbage and soil has been investigated in Beijing and Anhui under open conditions. Indoxacarb enantiomers in samples were extracted with acetonitrile, cleaned up by florisil SPE column, separated on high performance liquid chromatography with a cellulose-tris-(3, 5-dimethylphenylcarbamate)-based chiral stationary phase (CDMPC-CSP), and determined by a photodiode array detector. The validation of the developed method by fortification rac-indoxcarb in cabbage and soil showed good accuracy and precision. The results of field trials indicated that the dissipation of indoxacarb enantiomers followed pseudo-first-order kinetics or first-order kinetics in cabbage and soil at two locations. The half-lives of two enantiomers in cabbage ranged from 2.8 to 4.6 d which were shorter than those in soil ranging from 23 to 35 d. The changes of enantiomeric fraction values proved that enantioselective degradation of indoxacarb happened in cabbage and soil. The (-)-indoxacarb showed faster degradation in the Beijing cabbage, whereas in the Anhui cabbage, (+)-indoxacarb preferentially degraded. In soil, preferential degradation of (+)-indoxacarb was observed at two locations.

Determination of the enantiomers of α‐hydroxy‐ and α‐amino acids in capillary electrophoresis with contactless conductivity detection

The enantiomers of the anions of five alpha-hydroxy acids, namely lactic acid, alpha-hydroxybutyric acid, 2-hydroxycaproic acid, 2-hydroxyoctanoic acid and 2-hydroxydecanoic acid, as well as the two alpha-amino acids aspartic acid and glutamic acid, were baseline separated and detected by CE with contactless conductivity detection. Vancomycin was employed as chiral selector and could be used with conductivity detection without having to resort to a partial filling protocol as needed when this reagent is used with UV absorbance measurements. The procedure was successfully applied to the determination of the lactic acid enantiomers in samples of milk and yogurt. Linearity was achieved in the concentration range of 10-500 micromol/L with good correlation coefficients (0.9993 and 0.9990 for L- and D-lactic acid, respectively). The LODs (3 S/N) for L- and D-lactic acid were determined as 2.8 and 2.4 micromol/L, respectively.

Spatial and Temporal Trends of Chiral Organochlorine Signatures in Great Lakes Air Using Passive Air Samplers

Passive air samples (PAS) were collected and analyzed to assess the spatial and temporal trends of chiral organochlorine signatures in the Laurentian Great Lakes. Samples were collected from 15 sites and analyzed for the concentrations and enantiomer signature of chlordanes and alpha-hexachlorocyclohexane (alpha-HCH). Levels of the chlordanes were typically 4 times higher in urban areas than what were observed at rural and remote locations, exhibiting strong urban-rural gradients. Near racemic residues were seen for the chlordane enantiomers in samples collected from sites located in Toronto and Chicago, which can be attributed to continued emissions of historical use of the technical chlordane mixture, while the chiral signature observed at sites located in rural and remote locations was indicative of an aged source. Knowledge of the spatial and temporal distribution of the enantiomer signatures of chlordane and alpha-HCH in air is useful for distinguishing sources of these compounds to ambient air. Results suggest that potential sources, such as those associated with Toronto and Chicago, have limited influence over the levels at rural and remote sites within the Great Lakes. Sources that are relatively close to sample sites, however, have a strong influence on levels observed at those sites. For instance, results indicate that Lake Superior continues to act as a source of alpha-HCH to sites located on its shores. Generally, it appears that during the warmer months, local enhanced surface-air exchange influences air concentrations and that during the cooler periods of the year, levels in the atmosphere are more strongly influenced by advective transport from source regions.

Analytical and semi-preparative high-performance liquid chromatographic separation and assay of hydroxychloroquine enantiomers

(±)-Hydroxychloroquine (HCQ) is an antimalarial and anti-arthritic drug which is administered as the racemate. An accurate, precise and sensitive high-performance liquid chromatographic assay was developed for the determination of HCQ enantiomers in samples from human plasma, serum, whole blood, and urine. After addition of (±)-chloroquine (internal standard), samples of blood component (0.5 ml) or urine (0.1 ml) were alkalinized and extracted with 5 ml of diethyl ether. After solvent evaporation the residues were derivatized with (+)-di-O-acetyl- l-tartaric anhydride at 45°C for 30 min. The resulting diastereomers were then resolved using a C 8 analytical column with a mobile phase consisting of 0.05 M KH 2PO 4 (pH 3)—methanol—ethanol—triethylamine (78:22:1:0.08). The ultraviolet detection wavelength was set at 343 nm. The derivatized HCQ enantiomers eluted in less than 40 min, free of interfering peaks. Excellent linear relationships ( r 2 > 0.997) were obtained between the area ratios and the corresponding plasma concentrations over a range of 12.5–500 ng/ml. The diastereomers could be hydrolysed using microwave energy and neutral pH, which enabled us to resolve the enantiomers on a semi-preparative (C 18 column) scale. The method was suitable for the analysis and semi-preparative separation of HCQ enantiomers.