Micellar liquid chromatographic green enantioseparation of racemic amino alcohols and determination of elution order.

A micellar liquid chromatographic method was developed for the green enantioseparation of racemic amino alcohols using an aqueous solution of the mixed surfactants as an alternative for organic solvents. In this study, the derivatives of the amino alcohols were synthesized using highly reactive chiral esters of (S)-levofloxacin (Lfx) under microwave conditions, and an aqueous solution of the surfactants (Brij-35 and SDS) was used for the enantioseparation of the synthesized diastereomeric derivatives (DDs) of amino alcohols using reversed-phase HPLC. The activated ester of Lfx was synthesized by reacting with N-hydroxybenzotriazole and characterized using UV, IR, 1 H NMR, high-resolution mass spectrometry, and elemental analysis. The DDs of racemic amino alcohols were separated on a C18 column using micellar LC. Chromatographic conditions were optimized by varying the concentration of the surfactants in aqueous solution and by varying the concentration and pH of the buffer. The green assessment score was calculated for the developed method (score: 82, an excellent green method). In addition, the density functional theory calculations were performed to develop the lowest energy-optimized structures of DDs. The method was validated according to the International Conference of Harmonization guidelines, and the retention factor (k), selectivity factor (α), resolution factor (RS ), limit of detection (0.198 ng mL-1 or 0.291 pM mL-1 ), and limit of quantification (0.594 ng mL-1 or 0.873 pM mL-1 ) were calculated.

High throughput solid-phase screening of bacteria with cyclic amino alcohol deamination activity for enantioselective synthesis of chiral cyclic ß-amino alcohols.

OBJECTIVES: To screening of bacteria with cyclic amino alcohol deamination activity for enantioselective synthesis of chiral cyclic ß-amino alcohols. RESULTS: A new strain named Arthrobacter sp. TYUT010-15 with the (R)-selective deamination activity of cyclic ß-amino alcohol has been isolated from nature via a high throughput solid-phase screening method. The reaction conditions of TYUT010-15 were optimized. Using the resting cell of TYUT010-15 as the catalyst, kinetic resolution of trans-2-aminocyclopentanol, trans-2-aminocyclohexanol and cis-1-amino-2-indanol was carried out to afford (1S, 2S)-trans-2-aminocyclopentanol, (1S, 2S)-trans-2-aminocyclohexanol and (1R, 2S)-cis-1-amino-2-indanol in > 99% ee and 49.6-50% conversion. Four aromatic ß-amino alcohols and two amines were also resolved, (S)-ß-amino alcohols and (R)-amines were obtained in > 99% ee. Preparation experiment was conducted with 200 mM (23.2 g L-1) racemic trans-2-aminocyclohexanol, yielding the desired (1S, 2S)-trans-2-aminocyclohexanol in 40% isolated yield, > 99% ee and 5.8 g L-1 d-1 space time yields. CONCLUSIONS: This study provides a high throughput solid-phase method for screening of bacteria with cyclic amino alcohol deamination activity and a first example for practical preparation of chiral cyclic ß-amino alcohol by Arthrobacter sp. TYUT010-15.

Optical and Fluorescent Dual Sensing of Aminoalcohols by in Situ Generation of BODIPY-like Chromophore.

A novel multifunctional aminophenylboronic acid connected to a diphenylketone gives both circular dichroism and fluorescence signals by in situ generation of a BODIPY-like chromophore in the presence of aminoalcohols. DFT calculations were used to understand the role of each functional group in the mechanism. This new sensor can distinguish different aminoalcohols and quantitatively indicate the concentration of the substrate, allowing for the convenient determination of the ee of racemic mixtures with a single probe.

Efficient biosynthesis of (R)-3-amino-1-butanol by a novel (R)-selective transaminase from Actinobacteria sp.

(R)-3-amino-1-butanol is a key intermediate of Dolutegravir for the treatment of HIV/AIDS and its green and efficient biosynthesis has attracted a great deal of attention. Transaminases are currently used as promising biocatalyst for the synthesis of chiral amines. However, many transaminases have (S)-specificity and (R)-selective transaminases were less exploited and studied, making the production of (R)-amines remain challenging. In this study, a novel transaminase from Actinobacteria sp. (As-TA) was obtained and applied for the biosynthesis of (R)-3-amino-1-butanol by transferring the amino group from isopropylamine to 4-hydroxy-2-butanone. After optimization of the reaction condition and using a substrate fed-batch strategy, the conversion of 100, 200, 300, 400 and 500 mM 4-hydroxy-2-butanone reached 100%, 94.9%, 86.1%, 76.1% and 70.9%, respectively. (R)-3-amino-1-butanol with a maximum yield of 29.6 g/L and 99.9% e.e. value was obtained. This was the first time demonstrating the successful biosynthesis of (R)-3-amino-1-butanol with transaminase as biocatalyst and the obtained As-TA enriched the enzyme pool of transaminase with (R)-specificity.

Chiral high-performance liquid and supercritical fluid chromatographic enantioseparations of limonene-based bicyclic aminoalcohols and aminodiols on polysaccharide-based chiral stationary phases.

Enantioseparation of limonene-based bicyclic 1,3-aminoalcohols and 1,3,5- and 1,3,6-aminodiols was performed by normal-phase high-performance liquid chromatographic and supercritical fluid chromatographic (SFC) methods on polysaccharide-based chiral stationary phases. The effects of the composition of the mobile phase, the column temperature and the structures of the analytes and chiral selectors on retention and selectivity were investigated by normal-phase LC and SFC technique. Thermodynamic parameters derived from selectivity-temperature-dependence studies were found to be dependent on the chromatographic method applied, the nature of the chiral selector and the structural details of the analytes. Enantiorecognition in most cases was enthalpically driven but an unusual temperature behavior was also observed: decreased retention times were accompanied by improved separation factors with increasing temperature, i.e. some entropically driven separations were also observed. The elution sequence was determined in all cases. The separation of the stereoisomers was optimized in both chromatographic modalities.

Enantioselective synthesis of enantiopure ß-amino alcohols via kinetic resolution and asymmetric reductive amination by a robust transaminase from Mycobacterium vanbaalenii.

Chiral ß-amino alcohols are very important chiral building block for preparing bioactive compounds for use in pharmaceutical and fine chemical industries. Synthesis of chiral ß-amino alcohols by transaminase is big challenging due to the strict substrate specificities and very low activity of the enzyme. In this work, a (R)-selective ω-transaminase (MVTA) from Mycobacterium vanbaalenii was employed as a biocatalyst for the first time for the synthesis of chiral ß-amino alcohol via kinetic resolution and asymmetric reductive amination. The enzyme was purified and characterized. Kinetic resolution of a set of racemic ß-amino alcohols including two cyclic ß-amino alcohols by MVTA was demonstrated, affording (R)-ß-amino alcohols, (1S, 2S)-trans-2-aminocyclopentanol and (1R, 2S)-cis-1-amino-2-indanols in >99% ee and 50-62% conversion. Asymmetric reductive amination of three α-hydroxy ketones (10-300 mM) by MVTA was conducted, (S)-ß-amino alcohols were obtained with >99% ee and 80-99% conversion. Preparation experiment for the reductive amination of 200 mM 2-hydroxyacetophenone by the resting cells of recombinant E. coli (MVTA) was proceeded smoothly and product (S)-2-amino-2-phenylethanol was obtained with 71% isolated yield, >99% ee and 68.6 g/L/d volumetric productivity. The current research proved that the MVTA is a robust enzyme for the preparation of chiral ß-amino alcohol with high volumetric productivity.

One pot simultaneous preparation of both enantiomer of ß-amino alcohol and vicinal diol via cascade biocatalysis.

OBJECTIVES: To investigate the efficiency of a new cascade biocatalysis system for the conversion of R, S-ß-amino alcohols to enantiopure vicinal diol and ß-amino alcohol. RESULTS: An efficient cascade biocatalysis was achieved by combination of a transaminase, a carbonyl reductase and a cofactor regeneration system. An ee value of > 99% for 2-amino-2-phenylethanol and 1-phenyl-1, 2-ethanediol were simultaneously obtained with 50% conversion from R, S-2-amino-2-phenylethanol. The generality of the cascade biocatalysis was further demonstrated with the whole-cell approaches to convert 10-60 mM R, S-ß-amino alcohol to (R)- and (S)-diol and (R)- and (S)-ß-amino alcohol in 90-99% ee with 50-52% conversion. Preparative biotransformation was demonstrated at a 50 ml scale with mixed recombinant cells to give both (R)- and (S)-2-amino-2-phenylethanol and (R)- and (S)-1-phenyl-1, 2-ethanediol in > 99% ee and 40-42% isolated yield from racemic 2-amino-2-phenylethanol. CONCLUSIONS: This cascade biocatalysis system provides a new practical method for the simultaneous synthesis of optically pure vicinal diol and an ß-amino alcohol.

A chiral, porous, organic cage-based, enantioselective potentiometric sensor for 2-aminobutanol.

A new enantioselective potentiometric sensor containing R-type chiral porous organic cage CC9 as the chiral selector was designed for the assay of 2-aminobutanol. Optimized membrane electrodes displayed a linear dynamic range from 10-3  ~ 10-1  mol·L-1 with a detection limit of 2.5 × 10-4  mol·L-1 and a Nernstian response of 27 ± 0.5mV·decade-1 toward S-2-aminobutanol within the pH range 7.0-10.0. The potentiometric enantioselectivity coefficient ( logKS,RPot) of this sensor was -1.333, indicating that the porous organic cage-based electrode exhibited good discrimination toward S-2-aminobutanol over R-2-aminobutanol.

Efficient Routes to a Diverse Array of Amino Alcohol-Derived Chiral Fragments.

Efficient syntheses of chiral fragments derived from chiral amino alcohols are described. Several unique scaffolds were readily accessed in 1-5 synthetic steps leading to 45 chiral fragments, including oxazolidinones, morpholinones, lactams, and sultams. These fragments have molecular weights ranging from 100 to 255 Da and are soluble in water (0.085 to >15 mM).

Fragmentation study of aminoalcohol-diterpenoid alkaloids by electrospray ionization time-of-flight mass spectrometry.

RATIONALE: Aminoalcohol-diterpenoid alkaloids were found to be a group of cardioactive substances in the lateral roots of Aconitum carmichaeli Debx. Studies on the fragmentation behaviors and features of these alkaloids in mass spectrometry would be important for their structural identification and in vivo metabolic research, which has not received much attention thus far. METHODS: In this study, the fragmentation behaviors of 14 aminoalcohol-diterpenoid alkaloids were investigated by utilizing high-resolution time-of-flight tandem mass spectrometry. By analysis of the obtained MS(2) data, we summarized the fragmentation features of the corresponding alkaloids under different collision energy. RESULTS: The dissociation of functional groups from the skeleton was observed as the main fragmentation way in electrospray ionization (ESI) mode. The order of fragmentation sites was C1/C3 > C16 > C15 > C6 > N, with loss of one or more CH3OH, H2O, C2H4 (substituent on N atom) or CO (at C15 ) groups. CONCLUSIONS: The first systematic investigations on the fragmentation of aminoalcohol-diterpenoid alkaloids are described in this paper, setting the stage for an in-depth identification and study of the corresponding components in complex systems.

Magnetic graphene - polystyrene sulfonic acid nano composite: A dispersive cation exchange sorbent for the enrichment of aminoalcohols and ethanolamines from environmental aqueous samples.

Present study aimed at graphene surface modification to achieve selective analyte binding in dispersive solid phase extraction. Magnetic graphene - polystyrene sulfonic acid (MG-PSS) cation exchange nano-composite was prepared by non-covalent wrapping method. Composite was characterized by FT-IR and zeta potential. Material exhibited good dispersion in water and high exchange capacity of 1.97±0.16mMg(-1). Prepared nano-sorbent was then exploited for the cation exchange extraction and gas chromatography mass spectrometric analysis of Chemical Weapons Convention relevant aminoalcohols and ethanolamines from aqueous samples. Extraction parameters such as sorbent amount, extraction time, desorption conditions and sample pH were optimized and effect of common matrix interferences such as polyethylene glycol and metal salts was also studied. Three milligram of sorbent per mL of sample with 20min of extraction time at room temperature afforded 70-81% recoveries of the selected analytes spiked at concentration level of 1µgmL(-1). Method showed good linearity in the studied range with r(2)≥0.993. The limits of detection and limits of quantification ranged from 23 to 54ngmL(-1) and 72 to 147ngmL(-1), respectively. The relative standard deviation for intra- and inter-day precision ranged from 4.6 to 10.2% and 7.4 to 14.8% respectively. Applicability of the method to different environmental samples as well as the proficiency tests conducted by the Organization for the Prohibition of Chemical Weapons (OPCW) was also ascertained.

Whole-body imaging with single-cell resolution by tissue decolorization.

The development of whole-body imaging at single-cell resolution enables system-level approaches to studying cellular circuits in organisms. Previous clearing methods focused on homogenizing mismatched refractive indices of individual tissues, enabling reductions in opacity but falling short of achieving transparency. Here, we show that an aminoalcohol decolorizes blood by efficiently eluting the heme chromophore from hemoglobin. Direct transcardial perfusion of an aminoalcohol-containing cocktail that we previously termed CUBIC coupled with a 10 day to 2 week clearing protocol decolorized and rendered nearly transparent almost all organs of adult mice as well as the entire body of infant and adult mice. This CUBIC-perfusion protocol enables rapid whole-body and whole-organ imaging at single-cell resolution by using light-sheet fluorescent microscopy. The CUBIC protocol is also applicable to 3D pathology, anatomy, and immunohistochemistry of various organs. These results suggest that whole-body imaging of colorless tissues at high resolution will contribute to organism-level systems biology.

A convenient fluorescent method to simultaneously determine the enantiomeric composition and concentration of functional chiral amines.

A 1,1'-bi-2-naphthol (BINOL)-based chiral aldehyde in combination with Zn(II) shows a highly enantioselective fluorescent response toward functional chiral amines at λ>500 nm. However, the combination of salicylaldehyde and Zn(II) gives the same fluorescent enhancement for both enantiomers of a functional chiral amine at λ=447 nm. By using the fluorescent responses of the combination of the BINOL-based chiral aldehyde, salicylaldehyde and Zn(II) at the two emission wavelengths, both the concentration and enantiomeric composition of functional chiral amines such as amino alcohols, diamines, and amino acids can be simultaneously determined by a single fluorescent measurement. This work provides a simple and convenient method for chiral assay.

Cyclodextrin-mediated enantioseparation of phenylalanine amide derivatives and amino alcohols by capillary electrophoresis-role of complexation constants and complex mobilities.

The separation of the enantiomers of phenylalanine amide and N-methyl derivatives as well as some amino alcohols was studied by CE in acidic BGEs using CDs as chiral selectors. The native CDs displayed only low chiral recognition ability at a concentration of 15 mg/mL in 20 mM sodium phosphate buffer, pH 2.5. In contrast, the analyte enantiomers were separated in the presence of randomly sulfated CDs under reversed polarity of the applied voltage. Sulfated ß-CD proved to be the most universal selector resulting in the enantioseparation of all analytes. Opposite enantiomer migration order depending on the size of the CD cavity was observed for phenylalanine amide and 2-amino-2-phenylethanol. The R-enantiomers migrated first in the presence of sulfated α-CD and γ-CD while the S-enantiomers were detected first in the presence of sulfated ß-CD. The enantioseparations could be rationalized based on analyte complexation by the respective CDs except for 2-amino-2-phenylethanol and sulfated ß-CD where essentially equal complexation constants were derived for the enantiomers. In this case, the migration behavior could be attributed to the mobilities of the enantiomer-CD complexes adding another example to the CE-specific phenomenon of enantioseparations based primarily on complex mobilities.

Lanthanide tris(ß-diketonates) as useful probes for chirality determination of biological amino alcohols in vibrational circular dichroism: ligand to ligand chirality transfer in lanthanide coordination sphere.

A series of lanthanide tris(ß-diketonates) functioned as useful chirality probes in the vibrational circular dichroism (VCD) characterization of biological amino alcohols. Various chiral amino alcohols induced intense VCD signals upon ternary complexation with racemic lanthanide tris(ß-diketonates). The VCD signals observed around 1500 cm(-1) (ß-diketonate IR absorption region) correlated well with the stereochemistry and enantiomeric purity of the targeted amino alcohol, while the corresponding monoalcohol, monoamine, and diol substrates induced very weak VCD signals. The high-coordination number and dynamic property of the lanthanide complex offer an effective chirality VCD probing of biological substrates.

Qualitative and quantitative analysis of enantiomers by mass spectrometry: application of a simple chiral chloride probe via rapid in-situ reaction.

A tandem mass spectrometry method for high-sensitivity qualitative and quantitative discrimination of chiral amino compounds is conducted. The method is based on a chemical derivation process that uses a simple reagent, L-1-(phenylsulfonyl)pyrrolidine-carbonyl chloride, as the probe. The method is applicable in both organic solutions and biological conditions. Twenty-one pairs of enantiomer containing amino acids, amino alcohols, and amines are used to produce diastereomers using the probe via in situ reaction for 20 s at room temperature. The resulting diastereomers are successfully recognized based on the relative peak intensities of their fragments in positive mode, with the chiral recognition ability values ranging from 0.35 to 3.83. The L/D ratio of Pro spiked at different concentrations (enantiomeric excess) in both acetonitrile and dog plasma is determined by establishing calibration curves. This method achieves a lower limit of quantification of 50 pmol in analyzing amino acids using an extract ion chromatograph. The relative standard deviation for both qualitative and quantitative results is <5%. Thus, the present method is demonstrated as a new and practical technique of rapidly and sensitively determining enantiomers of amino compounds.

Combination of two different stationary phases for on-line pre-concentration and separation of basic drugs by using nano-liquid chromatography.

Capillary columns were packed firstly with silica modified-teicoplanin (teico-CSP) particles for a short zone (1-5 cm) and then with a Cogent Bidentate C18 silica phase (25 cm). The first part of the column (inlet) was intended for focusing the sample model, consisted of selected basic compounds, while the second zone, containing RP18 particles, was used for their separation. For method optimization, some important experimental parameters were studied including the sample solvent, injected volume and teico-CSP particles length. 3 cm teico-CSP resulted to be effective for the on-line pre-concentration, before the separation, of acebutolol, alprenolol, nadolol, oxprenolol and terbutaline with limit of detection at levels of few ng/mL. The comparison of the data obtained in absence of the chiral stationary phase revealed that the use of this chiral short sector into the capillary allowed the increase of the sensitivity of 5-12 times. Injection of larger sample volumes were easily done using higher length of the teico-CSP into the capillary, however the use of 5 cm length was not appropriate because caused the partial chiral separation of some studied compounds.

Enantioseparations of primary amino compounds by high-performance liquid chromatography using chiral crown ether-based chiral stationary phase.

Liquid chromatographic resolution of racemic compounds containing a primary amino group has been known to be most successful when chiral crown ether-based chiral stationary phases (CSPs) are used. Among various crown ether-based CSPs, the stationary phase based on (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid covalently bonded to silica gel has been successfully applied in the resolution of various racemic compounds containing primary amino groups. In this chapter, the preparation of the CSP based on (+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid covalently bonded to silica gel and examples for the application to the enantioseparation of racemic compounds including α-amino acids, cyclic amines, amino alcohols, and chiral drugs are described.

Enantioseparation by chiral ligand-exchange capillary electrophoresis.

Chiral ligand-exchange capillary electrophoresis is a relatively cheap and flexible format of capillary electrophoresis. It separates chiral analytes with a metal complex chiral selector that is composed of a metal ion and either homogeneous or heterogeneous chiral ligands (in respect of analytes). The analytes to be separated are required to possess the ability to coordinate with the metal ion, commonly containing N and/or O atoms, for example, amino acids or amino alcohols. In present chapter we take the separation of dansyl dl-amino acids as a key example to guide through the setup of a chiral ligand-exchange capillary electrophoresis enantioseparation protocol. The critical working conditions emphasized include the selection of metal ion, pH window, the ligand-to-metal ratio and/or the concentrations of the complex and electrolytes. In addition, trouble-shooting advice is given such as the way to overcome baseline drifting.

A quantitative liquid chromatography tandem mass spectrometry method for metabolomic analysis of Plasmodium falciparum lipid related metabolites.

Plasmodium falciparum is the causative agent of malaria, a deadly infectious disease for which treatments are scarce and drug-resistant parasites are now increasingly found. A comprehensive method of identifying and quantifying metabolites of this intracellular parasite could expand the arsenal of tools to understand its biology, and be used to develop new treatments against the disease. Here, we present two methods based on liquid chromatography tandem mass spectrometry for reliable measurement of water-soluble metabolites involved in phospholipid biosynthesis, as well as several other metabolites that reflect the metabolic status of the parasite including amino acids, carboxylic acids, energy-related carbohydrates, and nucleotides. A total of 35 compounds was quantified. In the first method, polar compounds were retained by hydrophilic interaction chromatography (amino column) and detected in negative mode using succinic acid-(13)C(4) and fluorovaline as internal standards. In the second method, separations were carried out using reverse phase (C18) ion-pair liquid chromatography, with heptafluorobutyric acid as a volatile ion pairing reagent in positive detection mode, using d(9)-choline and 4-aminobutanol as internal standards. Standard curves were performed in P. falciparum-infected and uninfected red blood cells using standard addition method (r(2)>0.99). The intra- and inter-day accuracy and precision as well as the extraction recovery of each compound were determined. The lower limit of quantitation varied from 50pmol to 100fmol/3×10(7)cells. These methods were validated and successfully applied to determine intracellular concentrations of metabolites from uninfected host RBCs and isolated Plasmodium parasites.