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Derivatizing Agent (derivatizing + agent)
Selected AbstractsIntegrated on-chip derivatization and electrophoresis for the rapid analysis of biogenic aminesELECTROPHORESIS, Issue 14 2004Nigel P. Beard Abstract We demonstrate the monolithic integration of a chemical reactor with a capillary electrophoresis device for the rapid and sensitive analysis of biogenic amines. Fluorescein isothiocyanate (FITC) is widely employed for the analysis of amino-group containing analytes. However, the slow reaction kinetics hinders the use of this dye for on-chip labeling applications. Other alternatives are available such as o -phthaldehyde (OPA), however, the inferior photophysical properties and the UV ,max present difficulties when using common excitation sources leading to a disparity in sensitivity. Consequently, we present for the first time the use of dichlorotriazine fluorescein (DTAF) as a superior in situ derivatizing agent for biogenic amines in microfluidic devices. The developed microdevice employs both hydrodynamic and electroosmotic flow, facilitating the creation of a polymeric microchip to perform both precolumn derivatization and electrophoretic analysis. The favorable photophysical properties of the DTAF and its fast reaction kinetics provide detection limits down to 1 nM and total analysis times (including on-chip mixing and reaction) of <60 s. The detection limits are two orders of magnitude lower than current limits obtained with both FITC and OPA. The optimized microdevice is also employed to probe biogenic amines in real samples. [source] Application of 31P NMR spectroscopy and chemical derivatization for metabolite profiling of lipophilic compounds in human serumMAGNETIC RESONANCE IN CHEMISTRY, Issue S1 2009M. Aruni DeSilva Abstract New methods for obtaining metabolic fingerprints of biological samples with improved resolution and sensitivity are highly sought for early disease detection, studies of human health and pathophysiology, and for better understanding systems biology. Considering the complexity of biological samples, interest in biochemical class selection through the use of chemoselective probes for improved resolution and quantitation is increasing. Considering the role of lipids in the pathogenesis of a number of diseases, in this study fingerprinting of lipid metabolites was achieved by 31P labeling using the derivatizing agent 2-chloro-4,4,5,5-tetramethyldioxaphospholane. Lipids containing hydroxyl, aldehyde and carboxyl groups were selectively tagged with 31P and then detected with good resolution using 31P NMR by exploiting the 100% natural abundance and wide chemical shift range of 31P. After standardizing the reaction conditions using representative compounds, the derivatization approach was used to profile lipids in human serum. The results show that the 31P derivatization approach is simple, reproducible and highly quantitative, and has the potential to profile a number of important lipids in complex biological samples. Copyright © 2009 John Wiley & Sons, Ltd. [source] The use of MPA amide for the assignment of absolute configuration of a sterically hindered cyclic secondary amine by ,mix and shake' NMR methodMAGNETIC RESONANCE IN CHEMISTRY, Issue 1 2008Jinhai Gao Abstract We present here a new method using methoxyphenylacetic acid (MPA) as the chiral derivatizing agent (CDA) for the assignment of absolute configuration of cyclic secondary amines. The MPA amides were prepared using the purification-free ,mix and shake' method. A detailed conformational analysis for the two diastereomeric amides was conducted by 2D NMR experiments and molecular mechanics calculations. We have established that, in the most stable conformation of each syn rotamer of MPA amides, the H-, in the MPA moiety is oriented toward the bulky substituent group at the asymmetric carbon in the chiral amine, presumably to avoid steric and/or electrostatic interactions. The observed NMR data were correlated with the conformational model to allow unambiguous assignment of absolute configuration of secondary amines. The results demonstrate that the MPA can be used as a useful CDA in the case of sterically crowded cyclic secondary amines from which the MTPA amides are usually difficult to make. Copyright © 2008 John Wiley & Sons, Ltd. [source] Determination of glutathione disulfide levels in biological samples using thiol,disulfide exchanging agent, dithiothreitolBIOMEDICAL CHROMATOGRAPHY, Issue 2 2009Burhan Ates Abstract A reverse-phase HPLC method incorporating dithiothreitol (DTT) reduction for quantitative determination of oxidized glutathione (GSSG) in biological samples is described here. This method is based on our previous enzymatic reduction technique that uses N-1-(pyrenyl) maleimide (NPM) as a derivatizing agent. In our earlier method, glutathione disulfide (GSSG) was measured by first reducing it to GSH with glutathione reductase (GR) in the presence of NADPH. However, this is a very costly and time-consuming technique. The method described here employs a common and inexpensive thiol,disulfide exchanging agent, DTT, for reduction of GSSG to GSH, followed by derivatization with NPM. The calibration curves are linear over a concentration range of 25,1250 nm (r2 > 0.995). The coefficients of variations for intra-run precision and inter-run precision range from 0.49 to 5.10% with an accuracy range of 1.78,6.15%. The percentage of relative recovery ranges from 97.3 to 103.2%. This new method provides a simple, efficient, and cost-effective way of determining glutathione disulfide levels with a 2.5 nm limit of detection per 5 µL injection volume. Copyright © 2008 John Wiley & Sons, Ltd. [source] Separation and quantification of N -acetyl- l -cysteine and N -acetyl-cysteine-amide by HPLC with fluorescence detectionBIOMEDICAL CHROMATOGRAPHY, Issue 5 2006Wei Wu Abstract N- acetyl- l -cysteine (NAC) is a well-known antioxidant that is capable of facilitating glutathione (GSH) biosynthesis and replenishing intracellular GSH under oxidatively challenging circumstances. N- acetyl-cysteine-amide (NACA), the amide form of NAC, is a newly designed and synthesized thiol-containing compound which is believed to be more lipophilic and permeable through cell membranes than NAC. The metabolic and antioxidant effects of these compounds in vitro and in vivo are under investigation. However, an analytical method that can separate and quantify both compounds simultaneously is not yet available, to the best of our knowledge. Because of their structural similarities, the two compounds are difficult to separate using earlier HPLC methods which were designed for NAC quantification. Therefore, the goal of this work was to develop an HPLC method with fluorescence detection for simultaneous quantification of NAC and NACA in biological blood and tissue samples. A gradient HPLC program with fluorescence detection (,ex = 330 nm, ,em = 376 nm) using N -(1-pyrenyl)maleimide (NPM) as the derivatizing agent was developed. The calibration curves were linear over a concentration range of 25,5000 nm (r2 > 0.997). The coefficients of variation for within-run precision and between-run precision ranged from 0.67 to 5.23% and for accuracy ranged from 0.98 to 10.54%; the percentage relative recovery ranged from 94.5 to 102.8%. This new method provides satisfactory separation of NAC and NACA, along with other biological thiols, in 20 min with a 5 nm limit of detection (LOD) per 5 µL injection volume. Copyright © 2005 John Wiley & Sons, Ltd. [source] Determination and Kinetics of Producing Glucosamine Using FungiBIOTECHNOLOGY PROGRESS, Issue 5 2007Jui-Wei Hsieh This work used three fungi, Rhizopus oligosorus BCRC 31996, Monascus pilosus BCRC31527, and Aspergillussp. BCRC31742, to produce glucosamine by using submerged fermentation and flask cultures. The reaction of glucosamine with 1-naphthyl isothiocyanate as derivatizing agent was carried out in pyridine at 50 °C for 1 h. The derivative was accurately analyzed and quantified by using high performance liquid chromatography. The relative standard deviation of glucosamine determined between experimental and real values were less than 2%. The kinetic and strategy of producing glucosamine in a flask culture was investigated to achieve an optimum yield of glucosamine under different conditions including three kinds of fungi, medium, and pH values. The descending ability of producing glucosamine for the three fungi was Aspergillus sp. BCRC31742 > Monascus pilosus BCRC31527 > Rhizopus oligosorus BCRC 31996 under the conditions studied. The experimental result shows that the glucosamine concentration had an optimum value and was 3430 mg/L by using Aspergillus sp. BCRC31742 culture in glucose and peptone (GP) medium, the yield of which was the best amount using wild-type microorganisms in the past. The generation culture of fungi and the pH control played important roles in enhancing the yield of glucosamine. The specific growth rate of the microorganism and the biomass, content, yield, and productivity of glucosamine were calculated as well. [source] Isomerization of delta-9-THC to delta-8-THC when tested as trifluoroacetyl-, pentafluoropropionyl-, or heptafluorobutyryl- derivatives,,JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 5 2008Justin M. Holler Abstract For GC,MS analysis of delta-9-tetrahydrocannabinol (delta-9-THC), perfluoroacid anhydrides in combination with perfluoroalcohols are commonly used for derivatization. This reagent mixture is preferred because it allows simultaneous derivatization of delta-9-THC and its acid metabolite, 11-nor-delta-9-THC-9-carboxylic acid present in biological samples. When delta-9-THC was derivatized by trifluoroacetic anhydride/hexafluoroisopropanol (TFAA/HFIPOH) and analyzed by GC,MS using full scan mode (50,550 amu), two peaks (P1 and P2) with an identical molecular mass of 410 amu were observed. On the basis of the total ion chromatogram (TIC), P1 with a shorter retention time (RT) was the major peak (TIC 84%). To identify the peaks, delta-8-THC was also tested under the same conditions. The RT and spectra of the major peak (TIC 95%) were identical with that of P1 for delta-9-THC. A minor peak (5%) present also correlated well with the latter peak (P2) for the delta-9-THC derivative. The fragmentation pathway of P1 was primarily demethylation followed by retro Diels-Alder fragmentation (M , 15,68, base peak 100%) indicating P1 as a delta-8-THC-trifluoroacetyl compound. This indicated that delta-9-THC isomerized to delta-8-THC during derivatization with TFAA/HFIPOH. Similar results were also observed when delta-9-THC was derivatized with pentafluoropropionic anhydride/pentafluoropropanol or heptafluorobutyric anhydride/heptafluorobutanol. No isomerization was observed when chloroform was used in derivatization with TFAA. In this reaction, the peaks of delta-8-THC-TFA and delta-9-THC-TFA had retention times and mass spectra matching with P1 and P2, respectively. Because of isomerization, perfluoroacid anhydrides/perfluoroalcohols are not suitable derivatizing agents for analysis of delta-9-THC; whereas the TFAA in chloroform is suitable for the analysis. Published in 2008 by John Wiley & Sons, Ltd. [source] Substituent effects on conformational preference in , -substituted , -fluorophenylacetic acid methyl ester model systems for chiral derivatizing agentsJOURNAL OF PHYSICAL ORGANIC CHEMISTRY, Issue 10 2009Riadh Sahnoun Abstract In connection with study of chiral derivatizing agents (CDAs) for NMR determination of absolute configuration of organic compounds, factors controlling the conformational preference between syn - and anti -forms in , -substituted , -fluorophenylacetic acid methyl ester (FC(X)(Ph)COOMe) model systems were theoretically investigated. Substituents X at the stereogenic carbon atom were X,=,H, CCH and CH3, the electronic and steric properties of which were significantly different from each other. The model system with X,=,CCH and that with X,=,CH3 were found to be possible candidates for fluorine-containing CDAs. The syn conformation is stable compared with the anti one by 0.7,kcal,mol,1 for the ester with X,=,CCH. On the other hand, the anti conformation is stable compared with the syn one by 0.5,kcal,mol,1 for the ester with X,=,CH3. Both natural bond orbital (NBO) analysis and deletion of selected orbitals based on the donor,acceptor NBO scheme were adopted for semi-quantitative estimation of factors responsible for the conformational preference as well as a qualitative inspection of occupied canonical molecular orbitals (MOs). It was shown that [,,(,*,+,,*)(CO)] and [,,,*(Ph) and ,(Ph),,*] hyperconjugations are the main factors controlling the conformational preferences between the syn and anti conformations. Other types of effects such as electrostatic effects were also investigated. The role of the fluorine atom was also clarified. Copyright © 2009 John Wiley & Sons, Ltd. [source] Determination of the enantiomeric excess of chiral carboxylic acids by 31P NMR with phosphorylated derivatizing agents from C2 -symmetrical diamines containing the (S)-,-phenylethyl groupCHIRALITY, Issue 6 2007Virginia M. Mastranzo Abstract The use of P(III) and P(V) organophosphorus derivatizing agents prepared from C2 symmetrical (1R,2R)- and (1S,2S)- trans - N,N,-bis-[(S)-,-phenylethyl]-cyclohexane-1,2-diamines 1 and 2, as well as (1R,2R)- and (1S,2S)- trans - N,N,-bis-[(S)-,-phenylethyl]-4-cyclohexene-1,2-diamines 3 and 4 for the determination of enantiomeric composition of chiral carboxylic acids by 31P NMR, is described. Chirality, 2007. © 2007 Wiley-Liss, Inc. [source] | |||||||||||||