Abstract: The extensive metabolism and administration of low doses of ethinylestradiol (EE) in preclinical animal species necessitates a sensitive analytical method to quantify the drug at low picogram-per-milliliter concentrations in biological matrixes. A highly sensitive and accurate method based on the derivatization of EE with dansyl chloride coupled with liquid chromatography/tandem mass spectrometry is described. The dansyl derivatization of EE introduced a basic secondary nitrogen into the molecule that was readily ionized in commonly used acidic HPLC mobile phases. The derivative showed an intense protonated molecular ion at m/z 530 under positive turbo ion spray ionization. The collision-induced dissociation of this ion formed a distinctive product at m/z 171, corresponding to the protonated 5-(dimethylamino)naphthalene moiety. The selected reaction monitoring, based on the m/z 530 → 171 transition, was highly specific for EE, since no background signal was observed from blank plasma obtained from rhesus ...

Abstract: This survey as a sequel of two earlier reports gives an overview of recent developments, starting from 1999, in the use of derivatization protocols in capillary electrophoretic (CE) analysis. Derivatization is mainly used for enhancement of the detection sensitivity in CE, for which a combination of fluorescence labeling and laser-induced fluorescence detection is favorable. Moreover, especially in the field of saccharide assay, derivatization to introduce charge into the molecule, is common. Derivatization procedures are classified in tables, focused on precapillary, on-line, on-capillary and postcapillary arrangements and divided in sections concerning the functional group that is derivatized. The most frequently reported groups are amines and the reducing end of (oligo)saccharides, but thiols, carbonyl and carboxyl groups, steroids and inorganic ions have also been reported about. Other reasons for derivatization are to enhance chiral separation, introduction of a suitable charge into the molecule or to improve mass spectrometric detection. The use of derivatization techniques for special cases, such as the analysis of neurotransmitters, insulin antibodies and mitochondria has also been incorporated as well as a study on the adsorption of proteins onto capillary walls during CE in which derivatization plays a role.

Abstract: Part I: introduction and general considerations sampling and its connection with sample preparation overview on sample preparation for chromatography chromatography as the core step in an analytical process data processing in chromatographic methods automation in sample preparation. Part II: sample preparation techniques using physical processes phase transfer separations applied in sample preparation headspace sampling techniques in gas chromatography solvent extraction sorbent extraction chromatographic procedures as preliminary separation membrane separations electroseparation techniques in sample preparation other separation techniques in sample preparation. Part III: sample preparation techniques involving chemical modifications - chemical modifications for dissolution and fractionation purpose of derivatization in chromatographic analysis chemical reactions used in derivatization derivatization reactions for analytes with various functional groups chemical degradation of polymers and pyrolysis.

Abstract: A new sample preparation method, stir bar sorptive extraction (SBSE), has been evaluated for the enrichment of organic solutes from biological fluids such as urine and blood. In SBSE, a stir bar coated with a polydimethylsiloxane layer is stirred for a given time in the sample. After sampling the stir bar is placed in a thermal desorption unit coupled on-line to capillary gas chromatography-mass spectrometry (SBSE-TD-CGC-MS). The principle and operation of SBSE are presented. Total profiling and target compound analysis have been selected as applications to illustrate the performance of SBSE-TD-CGC-MS (MSD). It is demonstrated that a variety analytes ranging from biological markers (phenols, hormones, fatty acids) to artificial contaminants (recreational drugs, plasticizers) can be enriched with high sensitivity. For polar solutes, in-situ derivatization can enhance both recovery into the polydimethylsiloxane (PDMS) layer and chromatographic analysis. Two types of derivatization have been applied, derivatization with ethyl chloroformate and with acetic acid anhydride. Linearity, detectability, and repeatability are illustrated by the determination of 1-hydroxypyrene in a urine sample from a smoker.

Abstract: A method has been developed for quantitative determination of carbonyl disinfection by-products (DBP) from aqueous samples by derivatization with 2,4-dinitrophenylhydrazine combined with high-performance liquid chromatography (HPLC) and electrospray ionization (ESI) tandem mass spectrometry (MS–MS). The effect of excess of derivatization reagent and derivatization time, the effect of buffer and dry-gas temperature in the ESI process, and the effect of focus potential and collision energy in MS measurement are shown. Major fragment ions for compound identification on the basis of collision-induced dissociation (CID) mass spectra (MS) are given, as are common fragments for screening analyses by MS experiments such as the use of precursor ion scans. Detection limits in the µg L–1 range could be achieved by selected ion monitoring measurements without sample preconcentration. Solid-phase extraction improved the sensitivity by a factor of 25 to 250. The applicability of the method is illustrated by DBP analyses of samples from outdoor swimming pools after chlorination. Several carbonyl compounds, e.g. aldehydes, ketones, hydroxybenzaldehyde, and dicarbonyl compounds were identified.

Abstract: Quantitation of amino acids in complex matrixes without derivatization is advantageous; however, difficulties exist in both the separation and the detection of those compounds. A validated method that is based on the use of volatile ion-pair liquid chromatography coupled to stable isotope dilution tandem mass spectrometry has been developed for the simple and accurate quantitation of underivatized amino acids in biological samples. Sufficient separation of 22 underivatized amino acids was achieved on a C18 column in 36 min using perfluoroheptanoic acid (PFHA) and trifluoroacetic acid (TFA) as mobile phase modifiers. The collisionally activated dissociation spectra of the amino acids were investigated and the transitions of [M + H]+ → [M + H − 46]+, which are specific to α-amino acids, were used for the detection of most amino acids and their stable isotopes. The calibration curves were linear over the range of 0.10−100 μg/mL, and the detection limits were 0.03−20 pmol on column. The quantitative results b...

Abstract: Plasma-assisted polymerization of maleic anhydride has been investigated under different experimental conditions. Significant variations in the film chemical structure and the film properties were obtained using pulsed plasma depositions operated at different duty cycles. The film chemical structures were obtained using X-ray photoelectron spectroscopy (XPS) and Fourier transform infra red spectroscopy (FT-IR). Surface derivatization reactions using decylamine and benzylamine were used to demonstrate their surface reactivity toward nucleophilic moieties and to change the surface free energy of the plasma polymer films, all of which are of particular interest for future applications in the attachment of biological molecules and cells. A method of substrate pretreatment was developed to ensure reliable binding between the substrate and the plasma polymer film in aqueous solution. Impedance spectroscopy was used to monitor polymer film changes in aqueous media. The hydrated films showed some resemblance to p...

Abstract: A strategy for generating potential galectin inhibitors was devised based on derivatization at the C-3' atom in 3'-amino-N-acetyllactosamine by using structural knowledge of the galectin carbohydrate recognition site A collection of 12 compounds was prepared by N-acylations or N-sulfonylations Hydrophobic tagging of the O-3 atom in the N-acetylglucosamine residue with a stearic ester allowed rapid and simple product purification The compounds were screened in a galectin-3 binding assay and three compounds with significantly higher inhibitory activities compared to the parent N-acetyllactosaminide were found These three best inhibitors all carried an aromatic amide at the C-3' position of the galactose moiety, which indicates that favorable interactions were formed between the aromatic group and galectin-3 The best inhibitor had an IC50 value (44 microM) about 50 times better than the parent N-acetyllactosaminide, which implies that it has potential as a valuable tool for studying galectin-3 biological functions and also as a lead compound for the development of galectin-3-blocking pharmaceuticals

Abstract: The most commonly used method for analysis of airborne carbonyls is to collect the analytes on solid sorbents coated with a suitable derivatization agent, followed by solvent desorption and liquid injection for analysis by high-pressure liquid chromatography. We have explored a new approach by combining on-sorbent derivatization and thermal desorption to measure airborne carbonyls. More specifically, carbonyls in the air are collected onto an O-(2,3,4,5,6−pentafluorobenzyl)hydroxylamine (PFBHA)-coated Tenax sorbent packed in a tube with dimensions identical to those of a gas chromatography (GC) injector liner. The derivatives are then released by in-injection port thermal desorption to a GC column for analysis. Gaseous carbonyls, including formaldehyde, acetaldehyde, benzaldehyde, glyoxal, and methylglyoxal, at ppbv levels are shown to be effectively collected (≥92% collection efficiency) onto the sampling tubes at a flow rate of 20 mL/min under both <1% and 71% relative humidity. The collection efficienc...

Abstract: A simple, rapid, and reliable reversed-phase high-performance liquid chromatographic method for the analysis of 16 amino acids of main interest in commercial fruit juices (pear, orange, grapefruit, pineapple, peach, and apricot) is described. No sample cleanup is required. The pH of the fruit juices is adjusted to alkaline value (8.5) using 200 mM borate buffer, then amino acid is converted to stable derivatives using 9-fluorenylmethyl-chloroformate. The excess of derivatization reagent is removed by a hydrophobic amine, 1-amino-adamantane hydrochloride. The derivatization procedure is simple, fast, and described in detail. Amino acids are detected at 263 nm and eluted within 35 min. The calibration, precision (< or = 6.1%), and recovery (102% +/- 4%) of the method are reported. The conditions of separation are optimized; however, serine partially overlapped with aspartic acid. The amino acid profile of fruit juices is consistent with data from the literature.

Abstract: The most commonly used method for analysis of airborne carbonyls is to collect the analytes on solid sorbents coated with a suitable derivatization agent, followed by solvent desorption and liquid injection for analysis by high-pressure liquid chromatography. We have explored a new approach by combining on-sorbent derivatization and thermal desorption to measure airborne carbonyls. More specifically, carbonyls in the air are collected onto an O-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine (PFBHA)-coated Tenax sorbent packed in a tube with dimensions identical to those of a gas chromatography (GC) injector liner. The derivatives are then released by in-injection port thermal desorption to a GC column for analysis. Gaseous carbonyls, including formaldehyde, acetaldehyde, benzaldehyde, glyoxal, and methylglyoxal, at ppbv levels are shown to be effectively collected (> or = 92% collection efficiency) onto the sampling tubes at a flow rate of 20 mL/min under both < 1% and 71% relative humidity. The collection efficiency drops as the sampling flow rate increases, and the degree of decrease is compound dependent. The derivatization agent, at a level of approximately 127 nmol/tube, is thermally desorbed and eluted from the GC column without compromising the determination of any carbonyl-PFBHA derivatives. Detection limits of low ppbv to sub-ppbv are achieved for a sample air volume of 4.8 L. Using this new method, we have measured formaldehyde, acetaldehyde, benzaldehyde, glyoxal, and methylglyoxal to be 4.9-16.3, 0.6-8.2, <5.9, 0.5-4.1, and <2.4 ppbv, respectively, in the ambient atmosphere at the university bus stop. This method is less labor intensive than the solvent desorption technique and avoids use of organic solvents. Other classes of airborne polar species can be measured through the same approach by selecting an appropriate derivatization agent.

Abstract: Determination of amino acids in a complex matrix without derivatization is advantageous, however, difficulties are found in both the detection and the separation of those compounds. In this study, a rapid and reliable LC-MS-MS method for the quantitation of underivatized amino acids in exocellular media was established. Injections were made directly after centrifugation of the samples, without further preparation. The separation of seven underivatized amino acids was achieved on a reversed-phase C18 column with pentadecafluorooctanoic acid as a volatile ion-pair reagent, and the specific detection of most amino acids was achieved by MS-MS of the specific transitions [M + H]+→[M + H − 46]+. The calibration curves of all analytes were linear over the range of 1.0–1000 μg ml−1 and the detection limits ranged from 0.1 to 5 ng ml−1, with an injection volume of 20 μl. The inter-day and intra-day precisions ranged from 2.6 to 5.7% and 4.8 to 8.2%, respectively; the mean recoveries of the seven analytes were 81–104%, 91–107% and 93–101% respectively at the spiked level of 10, 40 and 200 μg ml−1. A large number of fermentation samples were analysed using this method. The technique is simple, rapid, selective and sensitive, and shows potential for the high-throughput quantitation of amino acids from other biological matrices.

Abstract: Background: Enantioselective analysis of amphetamine (AM) or methamphetamine (MA) in urine is already a well-established tool for differentiation of illicit from therapeutic ingestion of AM or MA derivatives. However, because of the increasing importance of plasma or serum in analytical toxicology, a method for enantioselective analysis of AM and MA in these matrices is needed. Methods: AM and/or MA were extracted from 0.2 mL of blood plasma or serum by mixed-mode solid-phase extraction. After derivatization with S-(−)-heptafluorobutyrylprolyl chloride, the resulting diastereomers were separated by gas chromatography on a HP-5MS column during a 15-min program and detected by mass spectrometry in the negative-ion chemical ionization mode (NICI-GC-MS). The method was fully validated and applied to >50 samples from authentic toxicology cases. Results: The derivatized AM and MA enantiomers were well separated and sensitively detected. The method was linear from 5 to 250 μg/L per enantiomer with analytical recoveries, accuracy, and within- and between-run precision well within required limits. Extraction yields were 88.9–98.6%. Implications of concentrations and enantiomeric composition of AM and MA in the authentic samples were considered. Conclusions: This sensitive, reliable, rapid NICI-GC-MS assay is suitable for enantioselective determination of AM and MA in blood plasma or serum samples.

Abstract: A new capillary electrophoresis (CE) method was developed for the rapid, simple and selective determination of thiosulfate, sulfide and sulfite species. The proposed method is based on the in-capillary derivatization of separated sulfur anions by mixing their zones with the iodine zone during the electrophoretic migration and direct UV detection of iodide formed. The optimal conditions for the separation and derivatization reaction were established by varying electrolyte pH, electrolyte counter-ion, concentration of iodine, and applied voltage. The optimized separations were carried out in 20 mmol/L Tris-chloride electrolyte (pH 8.5) using direct UV detection at 214 nm. All three sulfur species were well resolved in less than 4 min. The method gives repeatability comparable or even better than this obtained for sulfur anions using standard CE technique. The proposed CE system was applied to the monitoring of sulfur anions in spent fixing solutions during the electrolytic oxidation.

Abstract: We have demonstrated that precolumn derivatization and capillary electrophoresis separation on a poly(dimethylsiloxane) (PDMS) microchip can be realized as efficient as those on glass microchips In an optimized condition of micellar electrokinetic chromatography (MEKC), using 25 mM sodium borate buffer (pH 100) with 25 mM sodium dodecyl sulfate (SDS) and 5% v/v methanol, the electroosmotic flow in an oxidized PDMS microchip is stabilized within 3% for days By employing a fluorometric derivatization with o-phthaldialdehyde (OPA) in an optimally designed reaction chamber, four most important biogenic amines occurring in foods, histamine, tyramine, putrescine, and tryptamine, are quantitatively determined in less than 1 min at the levels applicable to real samples The migration behaviors of anionic OPA-derivatized biogenic amines under the MEKC conditions are analyzed, and it has been found that under our separation conditions, the electrophoretic mobility of the SDS micelles is significantly greater than those of the anions in the aqueous phase The channel manifold in a PDMS substrate is fabricated using replica molding against a thick photoresist, SU-8, pattern generated by photolithography The plate with the microchannel pattern is strongly, irreversibly bonded to another PDMS plate by using a new bonding technique, which employs surface oxidation by corona discharge generated from a cheap, handy source, Tesla coil

Abstract: An in-sample derivatization headspace solid-phase microextraction method has been developed for the simultaneous determination of nonylphenol, nonylphenol mono- and diethoxylates (NP, NP1EO, NP2EO), and their acidic metabolites (NP1EC, NP2EC) in water. The analytical procedure involves derivatization of NPEOs and NPECs to their methyl ethers−esters with dimethyl sulfate/NaOH and further headspace (HS) solid-phase microextraction (SPME) and gas chromatography/mass spectrometry (GC/MS) determination. Parameters affecting both derivatization efficiency and headspace SPME procedure, such as the selection of the SPME coating, derivatization−extraction time, temperature and ionic strength were optimized. The commercially available Carbowax−divinylbenzene (CW−DVB) fiber appears to be the most suitable for the simultaneous determination of both NPEOs−NPECs. Run-to-run precision of the in-sample derivatization/HS-SPME-GC/MS method gave relative standard deviations between 8 and 18%. The method was linear for NP ov...

Abstract: This paper describes a fully automated procedure using alkaline hydrolysis and headspace solid-phase microextraction (HS-SPME) followed by on-fiber derivatization and gas chromatographic-mass spectrometric (GC-MS) detection of cannabinoids in human hair samples. Ten milligrams of hair was washed with deionized water, petroleum ether, and dichloromethane. After the addition of deuterated internal standards the sample was hydrolyzed with sodium hydroxide and directly submitted to HS-SPME. After absorption of analytes for an on-fiber derivatization procedure the fiber was directly placed into the headspace of a second vial containing N-methyl-N-trimethylsilyltrifluoroacetamide (MSTFA) before GC-MS analysis. The limit of detection was 0.05 ng/mg for delta9-tetrahydrocannabinol (THC), 0.08 ng/mg for cannabidiol (CBD), and 0.14 ng/mg for cannabinol (CBN). Absolute recoveries were in the range between 0.3 and 7.5%. Linearity was proved over a range from 0.1 to 20 ng/mg with coefficients of correlation from 0.998 to 0.999. Validation of the whole procedure revealed excellent results. In comparison with conventional methods of hair analysis this automated HS-SPME-GC-MS procedure is substantially faster. It is easy to perform without use of solvents and with minimal sample quantities, but with the same degree of sensitivity and reproducibility. The applicability was demonstrated by the analysis of 25 hair samples from several forensic cases. The following concentration ranges were determined: THC 0.29-2.20 (mean 1.7) ng/mg, CBN 0.55-4.54 (mean 1.2) ng/mg, and CBD 0.53-18.36 (mean 1.3) ng/mg. 11-nor-Delta9-tetrahydrocannabinol-9-carboxylic acid could not be detected with this method.

Abstract: In matrix-assisted laser desorption/ionization (MALDI) analyses of small oligosaccharides a very large increase in sensitivity (by a factor of 1000) may be obtained by introducing a quaternary ammonium center ('quaternization'). Such a quaternary ammonium center may be introduced into the saccharide by reaction with commercially available glycidyltrimethylammonium chloride (GTMA), or by using Girard's reagent T (Naven and Harvey, Rapid Commun. Mass Spectrom. 1996; 10: 829). GTMA reacts with alcohol functionalities, whereas Girard's reagent T is specific for aldehyde and keto groups. Thus reducing saccharides can be derivatized by both GTMA and Girard's reagent T. For example, glucose or cellobiose having a stock concentration of 3 x 10(-5) M (5 microg/mL) produces no sugar-derived signals in conventional MALDI, but their quaternized derivatives, also at 3 x 10(-5) M, yield intense signals, with the matrix-derived signals only being weak. Similar results were obtained for glucosamine. Non-reducing saccharides as well as sugar alcohols can be derivatized using GTMA; thus, although sucrose, raffinose and sorbitol do not react with Girard's reagent T, they all produce intense signals after derivatization with GTMA. An example of the application of these derivatization reactions is provided by the analysis of oligosaccharides in beer.

Abstract: The screening and quantitation of methamphetamine (MP) in urine using dansyl chloride (DNC) as the derivatization reagent were studied. Urinary MP derivatized with DNC could be detected by visual observation of the fluorescence in a solid-phase extraction column such as a Sep-Pak C18 cartridge to which the whole reaction solution was applied. The DNC-derivatized MP was eluted from the cartridge and then identified and quantitated by gas chromatography-mass spectrometry (GC-MS) and high-performance liquid chromatography (HPLC). In the GC-MS analysis with the MS detector in the electron-impact mode, DNC-derivatized MP and amphetamine (AP), exhibited diagnostic molecular ion peaks. The intensities of the molecular ions were 15% (DNC-MP) and 35% (DNC-AP) of the base peak (a fragment ion because of the loss of dimethylnaphthalene from M+), demonstrating that this method of derivatization has a major advantage for confirming APs by GC-MS. MP derivatized with DNC could be determined by HPLC with ultraviolet detection. Because a good correlation (r = 0.95) between the GC-MS and HPLC method for urinary MP was confirmed, both HPLC and GC-MS appear to be useful tools for determining urinary MP. The intensity of the cartridge fluorescence due to DNC-derivatized MP was approximately related to the urinary content of MP determined by HPLC or GC-MS, although a false positive in the visual fluorescence was observed in some urinary specimens from healthy volunteers. From these results, screening and confirmation/determination following DNC derivatization is proposed as a suitable method for the analysis of MP.

Abstract: This paper describes a general approach for the in-capillary derivatization of amino compounds and the subsequent sensitive determination of the derivatives by micellar electrokinetic chromatography (MEKC) or capillary zone electrophoresis (CZE) with laser-induced fluorescence (LIF) detection. Amino acids, biogenic amines and amino phosphonic acid-herbicides were chosen as model analytes to evaluate the analytical potential of this approach. Fulfilment of the in-capillary reaction of the analytes using LIF detection hinged on the excellent labeling chemistry of 5-(4,6-dichloro-s-triazin-2-ylamino)fluorescein (DTAF) and the good resolution achieved in the separation of derivatized analytes. Careful optimization of the electrophoretic conditions in the mixing step of this protocol allowed the determination of amino acids, biogenic amines and phosphorus-containing amino acid-herbicides with concentration limits of detection at the nug/L level and relative standard deviations from 3.5 to 5.8%. The whole analysis is carried out within 20 min, resulting in a very simple, fast and practical approach for the fully automated analysis of amino acids and related compounds in low-volume and low-concentration samples.