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Benzoin is one of the most commonly used photoinitiators to induce free radical polymerization. Here, improved benzoin properties could be accomplished by the introduction of two methoxy substituents, leading to the formation of 3',5'-dimethoxybenzoin (DMB) which has a higher photo-cleavage quantum yield (0.54) than benzoin (0.35). To elucidate the underlying reaction mechanisms of DMB and obtain direct information of the transient species involved, femtosecond transient absorption (fs-TA) and nanosecond transient absorption (ns-TA) spectroscopic experiments in conjunction with density functional theory/time-dependent density functional theory (DFT/TD-DFT) calculations were performed. It was found that the photo-induced α-cleavage (Norrish Type I reaction) of DMB occurred from the nπ* triplet state after a rapid intersystem crossing (ISC) process (7.6 ps), leading to the generation of phenyl radicals on the picosecond time scale. Compared with Benzoin, DMB possesses two methoxy groups which are able to stabilize the alcohol radical and thus result in a stronger driving force for cleavage and a higher quantum yield of photodissociation. Two stable conformations (cis-DMB and trans-DMB) at ground state were found via DFT calculations. The influence of the intramolecular hydrogen bond on the α-cleavage of DMB was elaborated.

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Twelve chiral compounds were enantiomerically resolved on bovine serum albumin chiral stationary phase (BSA-CSP) by high-performance liquid chromatography (HPLC) in reversed-phase modes. Chromatographic conditions such as mobile phase pH, the percentage of organic modifier, and concentration of analyte were optimized for separation of enantiomers. For N-(2, 4-dinitrophenyl)-serine (DNP-ser), the retention factors (k) greatly increase from 0.81 to 6.23 as the pH decreasing from 7.21 to 5.14, and the resolution factor (R(s)) exhibited a similar increasing trend (from 0 to 1.34). More interestingly, the retention factors for N-(2, 4-dinitrophenyl)-proline (DNP-pro) decrease along with increasing 1-propanol in mobile phase (3%, 5%, 7% and 9% by volume), whereas the resolution factor shows an upward trend (from 0.96 to 2.04). Moreover, chiral recognition mechanisms for chiral analytes were further investigated through thermodynamic methods.

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Capillary electrochromatography (CEC) is a micro-separation technique that combines the advantages of capillary zone electrophoresis with those of high-performance liquid chromatography. Accordingly, it has attracted extensive attention over the last decade. Among the stationary phases for CEC, monolithic stationary phase has been regarded as the most suitable stationary phase for CEC because of its simple preparation, the elimination of frits, and its excellent performance. In this chapter, procedures for preparing CEC monolithic columns with an improved configuration, in which there are stationary phases at both sides of detection window and no stationary phase at detection window, are presented. The separation of acidic and basic compounds on such monolithic columns is used as an example to demonstrate CEC separation protocol. Additionally, an on-line concentration technique in CEC is presented. As a result of the coexistence of stationary phase and electric field in a CEC column, it is possible to employ chromatographic zone sharpening and field-amplified sample stacking effects simultaneously to improve CEC detection sensitivity.

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A method involving self-concentration, on-column enrichment and field-amplified sample stacking for on-line concentration in capillary electrochromatography with a polymer monolithic column is presented. Since monolithic columns eliminate the frit fabrication and the problems associated with frits, the experimental conditions could be more flexibly adjusted to obtain higher concentration factor in comparison with conventional particulate packed columns. With self-concentration effect, the detection sensitivity of benzene and hexylbenzene is improved by a factor of 4 and 8, respectively. With on-column enrichment and ultralong injection, improvement as high as 22,000 times in detection sensitivity of benzoin is achieved. Furthermore, a combination of the three above-mentioned methods yields up to a 24,000-fold improvement in detection sensitivity for caffeine, a charged compound. Parameters affecting the efficiency of on-line concentration are investigated systematically. In addition, equations describing on-line concentration process are deduced.

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The diketone compound, benzil is reduced to (S)-benzoin with living Bacillus cereus cells. Recently, we isolated a gene responsible for benzil reduction, and Escherichia coli cells in which this gene was overexpressed transformed benzil to (S)-benzoin. Although this benzil reductase showed high identity to the short-chain dehydrogenase/reductase (SDR) family, enzymological features were unknown. Here, we demonstrated that many B. cereus strains had benzil reductase activity in vivo, and that the benzil reductases shared 94-100% amino acid identities. Recombinant B. cereus benzil reductase produced optically pure (S)-benzoin with NADPH in vitro, and the ketone group distal to a benzene ring was asymmetrically reduced. B. cereus benzil reductase showed 31% amino acid identity to the yeast open reading frame YIR036C protein and 28-30% to mammalian sepiapterin reductases, sharing the seven residues consensus for the SDR family. We isolated the genes encoding yeast YIR036C protein and gerbil sepiapterin reductase, and both recombinant proteins also reduced benzil to (S)-benzoin in vitro. Green fluorescent protein-tagged B. cereus benzil reductase distributed in the bipolar cytoplasm in B. cereus cells. Asymmetric reduction with B. cereus benzil reductase, yeast YIR036C protein and gerbil sepiapterin reductase will be utilized to produce important chiral compounds.

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With the deposition of cellulose tris(3,5-dimethylphenylcarbamate) (CDMPC) on aminopropylated silica gel (mean particle size, 5 microns; pore size, 13 nm; surface area, 110 m2/g) by using two different methods (evaporation and precipitation), two chiral stationary phases (CSP1 and CSP2) characterized by elemental analysis and scanning electron micrography were obtained. They were also evaluated by using seven racemic compounds with n-hexane/ethanol(95/5, V/V) and n-hexane/2-propanol (90/10, V/V) as mobile phases. The results showed that the chiral stationary phase CSP1 obtained by the evaporation method had better efficiency and chiral resolution ability than CSP2 by the precipitation method.

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Many selectors are used both in pressure-driven liquid chromatography (LC) and in electrokinetic chromatography (EKC), particularly chiral species such as cyclodextrins and proteins. It should be possible to readily apply information gleaned using one technique to the other, since in both techniques the underlying molecular interactions which lead to separations are expected to be the same. Superficially this may be the case, but an exact transfer of operating conditions, i.e., background electrolyte (BGE) composition/mobile phase composition, assuming that these meet certain minimum requirements for each technique, is not often possible. To investigate the reason for this we have measured retention (k') of a neutral solute (racemic benzoin) in HPLC and EKC using an identical range of BGE/mobile phase conditions in both techniques. The selector used was human serum albumin. The k' measurements obtained for each benzoin enantiomer were consistently higher in HPLC than in EKC. This can be explained very simply if one considers that retention in both systems is related to the selector concentration [S], by the expression k'=K[S], where K is the affinity constant. In EKC, [S] is simply the concentration of free selector in the BGE, while in LC, [S] = m(p)/Vo, where m(p) is the number of moles of accessible selector, and Vo is the column void volume. In LC, [S] is generally considerably higher than in EKC, leading to larger values of k'.

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A liquid chromatographic method for the individual determination of benzoic and cinnamic acids in 2 benzoin preparations is presented. The method specifies a reverse phase column and 0.01M KH2PO4-methanol (85 + 15) as mobile phase at a flow rate of 1.8 mL/min, with detection at 254 nm. The method has been applied to 2 benzoin preparations and the results were compared with those from the British Pharmacopoeia method.

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