Synthesis of new reagents for the fluorescence derivatisation of thiols and alcohols

The synthesis of two selective thiol derivatisation reagents (3, 4b) is described, starting from readily available 2-(4-aminophenyl)-6-methyl-benzothiazole (1). The isocyanate6, being a known alcohol derivatisation reagent, is obtained from1 in a two step synthesis, in which the usual isocyanate synthesis using phosgen is avoided.     

Polymer-supported diimine molybdenum carbonyl complexes as highly reusable and efficient pre-catalysts in epoxidation of alkenes

By reaction of aldehydic polystyrene and ethylene diamine, polystyrene–imine–amine reagent was produced. Reaction of this reagent with benzaldehyde and 4-nitrobenzaldehyde resulted in polystyrene-diimines (3a and 3b). These reagents were used for the immobilization of molybdenum hexacarbonyl. The functionalized polystyrene and supported-diimine molybdenum carbonyl catalysts were characterized by FT-IR spectrum and CHN analysis. The molybdenum content of catalysts was determined by neutron activation analysis. Supported-diimine molybdenum carbonyl pre-catalysts (3aM and 3bM) were used in epoxidation of cyclooctene, and the reaction parameters such as solvent and oxidant were optimized and the epoxidation of different alkenes was investigated in optimizing these conditions. The obtained results in the presence of polymer-supported diimine molybdenum carbonyl pre-catalysts (3aM and 3bM) showed that they were very active and selective in the epoxidation of a wide range of alkenes. The reusability of the supported pre-catalysts was also studied. The results showed that they were highly reusable in epoxidation of alkenes.     

Protected Amine Labels: A Versatile Molecular Scaffold for Multiplexed Nominal Mass and Sub-Da Isotopologue Quantitative Proteomic Reagents

We assemble a versatile molecular scaffold from simple building blocks to create binary and multiplexed stable isotope reagents for quantitative mass spectrometry. Termed Protected Amine Labels (PAL), these reagents offer multiple analytical figures of merit including, (1) robust targeting of peptide N-termini and lysyl side chains, (2) optimal mass spectrometry ionization efficiency through regeneration of primary amines on labeled peptides, (3) an amino acid-based mass tag that incorporates heavy isotopes of carbon, nitrogen, and oxygen to ensure matched physicochemical and MS/MS fragmentation behavior among labeled peptides, and (4) a molecularly efficient architecture, in which the majority of hetero-atom centers can be used to synthesize a variety of nominal mass and sub-Da isotopologue stable isotope reagents. We demonstrate the performance of these reagents in well-established strategies whereby up to four channels of peptide isotopomers, each separated by 4 Da, are quantified in MS-level scans with accuracies comparable to current commercial reagents. In addition, we utilize the PAL scaffold to create isotopologue reagents in which labeled peptide analogs differ in mass based on the binding energy in carbon and nitrogen nuclei, thereby allowing quantification based on MS or MS/MS spectra. We demonstrate accurate quantification for reagents that support 6-plex labeling and propose extension of this scheme to 9-channels based on a similar PAL scaffold. Finally, we provide exemplar data that extend the application of isotopologe-based quantification reagents to medium resolution, quadrupole time-of-flight mass spectrometers.

A fluorous tag-bound fluorescence derivatization reagent, F-trap pyrene, for reagent peak-free HPLC analysis of aliphatic amines

We have developed a novel pre-column fluorescence derivatization reagent for amines, F-trap pyrene. This reagent comprises a fluorescent pyrene moiety, an amine-reactive Marshall linker, and a fluorophilic perfluoroalkyl group known as fluorous tag. When the reagent reacts with aliphatic amines and amino acids to give fluorescent derivatives, the fluorous tag in the reagent is eliminated simultaneously. Therefore, excess unreacted reagents in the derivatization reaction solution still have the fluorous tag and could be removed by fluorous solid-phase extraction selectively before high-performance liquid chromatography (HPLC) analysis. By using this reagent, 13 kinds of aliphatic amine (C₂–C₁₆) derivatives can be separated within 40 min by reversed-phase HPLC with gradient elution. In this chromatogram, unreacted reagents peak at around 28 min, greatly decrease after fluorous solid-phase extraction, and do not interfere with the quantification of each amine. The detection limits (S/N = 3) for examined aliphatic amines are 3.6–25 fmol per 20 μL injection. We have also applied this reagent successfully to the amino acid analysis.

Synthesis and structure of iron(III) complexes with a new ligands based on Girard’s reagent

Condensation of 5-bromosalicylaldehyde with Girard’s reagent T yields a new ligand in the form of a salt, 5-bromosalicylaldehyde (carboxymethyl)trimethylammonium chloride hydrazone (5-BrH₂SalGT)Cl (I). Ligand I is readily soluble in water and reacts with iron chloride to give the complex [Fe(5-BrSalGT)Cl₂] (II). Treatment of II with KCNS leads to the compound [Fe(5-BrSalGT)(NCS)₂(H₂O)] (III). At any ratio of the initial reagents, only complexes with the ratio metal: ligand = 1: 1 are isolated. Comparison of the structural data for compounds I–III shows that ligand I is deprotonated in the course of complex formation and is coordinated in the anionic form. Its conformational rearrangement is minimal and involves only a change in the orientation of the terminal group N(CH₃)₃. In complexes II and III, ligand I is coordinated to the metal ion through the ONO donor atoms. The structures of the complexes with different acido ligands are significantly different. Although the complexes contain each two inorganic anions, their coordination polyhedra differ from each other. In II, the iron atom is at the center of a trigonal bipyramid, whereas in III the iron atom has a tetragonal-bipyramidal environment due to the extra coordination of a water molecule. In both complexes, the iron atom is in the high-spin state: at room temperature, μ_eff is 5.86 and 5.81 μ_B for II and III, respectively. Complexes II and III are ordinary paramagnets down to 2 K.     

Substituted quinolinones. 18. 3-Acetyl-4-methylthioquinolin-2(1H)-one as useful synthon intermediate for synthesis of some new quinolinones

3-Acetyl-4-methylthioquinolin-2(1H)-one (3) has been prepared and its reactivity towards treatment with different reagents, dilute sulfuric acid, aqueous sodium hydroxide, and hydrogen peroxide, is described. The reactions of compound 3 with benzylamine, benzaldehyde, DMF-DMA, Vilsmeier-Haack reagent, 2-chloro-3-formyl-pyridopyrimidone and malononitrile have been carried out under convenient conditions. Compound 3 underwent heterocyclization reactions with some binucleophiles, hydrazine, hydroxylamine, urea, thiourea, semicarbazide, and thiosemicarbazide, furnishing some known five, six, and seven heterocyclic annellated quinolines. All the new compounds have been characterized using different spectral and analytical tools.     

Charge Site Mass Spectra: Conformation-Sensitive Components of the Electron Capture Dissociation Spectrum of a Protein

A conventional electron capture dissociation (ECD) spectrum of a protein is uniquely characteristic of the first dimension of its linear structure. This sequence information is indicated by summing the primary c ^m+ and z ^m+? products of cleavage at each of its molecular ion’s inter-residue bonds. For example, the ECD spectra of ubiquitin (M?+?nH)ⁿ⁺ ions, n?=?7–13, provide sequence characterization of 72 of its 75 cleavage sites from 1843 ions in seven c ^(1–7)+ and eight z ^(1–8)+? spectra and their respective complements. Now we find that each of these c/z spectra is itself composed of “charge site (CS)” spectra, the c ^m+ or z ^m+? products of electron capture at a specific protonated basic residue. This charge site has been H-bonded to multiple other residues, producing multiple precursor ion forms; ECD at these residues yields the multiple products of that CS spectrum. Closely similar CS spectra are often formed from a range of charge states of ubiquitin and KIX ions; this indicates a common secondary conformation, but not the conventional α-helicity postulated previously. CS spectra should provide new capabilities for comparing regional conformations of gaseous protein ions and delineating ECD fragmentation pathways.

Reagent Cluster Anions for Multiple Gas-Phase Covalent Modifications of Peptide and Protein Cations

Multiple gas phase ion/ion covalent modifications of peptide and protein ions are demonstrated using cluster-type reagent anions of N-hydroxysulfosuccinimide acetate (sulfo-NHS acetate) and 2-formyl-benzenesulfonic acid (FBMSA). These reagents are used to selectively modify unprotonated primary amine functionalities of peptides and proteins. Multiple reactive reagent molecules can be present in a single cluster ion, which allows for multiple covalent modifications to be achieved in a single ion/ion encounter and at the ‘cost’ of only a single analyte charge. Multiple derivatizations are demonstrated when the number of available reactive sites on the analyte cation exceeds the number of reagent molecules in the anionic cluster (e.g., data shown here for reactions between the polypeptide [K₁₀ + 3H]³⁺ and the reagent cluster [5R_5Na – Na]^–). This type of gas-phase ion chemistry is also applicable to whole protein ions. Here, ubiquitin was successfully modified using an FBMSA cluster anion which, upon collisional activation, produced fragment ions with various numbers of modifications. Data for the pentamer cluster are included as illustrative of the results obtained for the clusters comprised of two to six reagent molecules.

Chemical Derivatization in Flow Analysis

Chemical derivatization for improving selectivity and/or sensitivity is a common practice in analytical chemistry. It is particularly attractive in flow analysis in view of its highly reproducible reagent addition(s) and controlled timing. Then, measurements without attaining the steady state, kinetic discrimination, exploitation of unstable reagents and/or products, as well as strategies compliant with Green Analytical Chemistry, have been efficiently exploited. Flow-based chemical derivatization has been accomplished by different approaches, most involving flow and manifold programming. Solid-phase reagents, novel strategies for sample insertion and reagent addition, as well as to increase sample residence time have been also exploited. However, the required alterations in flow rates and/or manifold geometry may lead to spurious signals (e.g., Schlieren effect) resulting in distorted peaks and a noisy/drifty baseline. These anomalies can be circumvented by a proper flow system design. In this review, these aspects are critically discussed mostly in relation to spectrophotometric and luminometric detection.     

Syntheses of perfluorinated epoxides,diepoxides, and a novel rearrangement

t-Butylhydroperoxide/butyl-lithium is a good reagent for synthesis of epoxides and diepoxides from polyfluorinated-alkenes and-polyenes. This reagent and calcium hypochlorite both give a novel diepoxide 5 from perfluoro-3,4-dimethyl-2,4-diene 1; diepoxide 5 undergoes a novel rearrangement, at 200°C, to the corresponding 1,4-dioxine derivative 18. Possible mechanisms are discussed.     

Development of chlorotriazine polymer dehydrocondensing reagents (Poly-Trzs)

Polymer-type dehydrocondensing reagents comprising of a triazine dehydrocondensing reagent, itself in a polymerized form (Poly-Trz—MMs'), have been developed by exploiting the chemical properties of cyanuric chloride that readily binds to alcohol or amines. A chlorotriazine polymer bearing two alkoxy substituents at the 4- and 6- positions (Poly-O-Trz-Cl) was prepared by alternating copolymerization between cyanuric chloride and tetra(ethylene glycol). Similarly, polymers bearing both alkoxy and amino substituents (Poly-N-Trz-Cl) were synthesized from tetra(ethylene glycol) bis(4,6-dichlorotriazin-2-yl) ether 6 and tris(2-aminoethyl)amine 7 and/or ethylenediamine 8. All the polymers were shown to be good reagents for dehydrocondensation of carboxylic acids and amines in the presence of NMM, compared to the corresponding monomeric dehydrocondensing reagent (DMT-MM). The advantages of the polymeric reagents are as follows: (1) both the reaction and isolation procedure of dehydrocondensation can be greatly simplified, (2) the dehydrocondensation can be conducted in protic solvents as well as in common organic solvents, (3) the reagents can be efficiently prepared at a low cost, and (4) these reagents are considered eco-friendly, generating a lower amount of waste compared to conventionally related reagents because of high loading of the dehydrocondensing activity (ca. 3 mequiv/g).     

Evaluation of a series of prolylamidepyridines as the chiral derivatization reagents for enantioseparation of carboxylic acids by LC–ESI–MS/MS and the application to human saliva

Mass spectrometry has become a popular analytical tool because of its high sensitivity and specificity. The use of a chiral derivatization reagent for the mass spectrometry (MS) detection seems to be efficient for the enantiomeric separation of racemates. However, the number of chiral reagents for the liquid chromatography (LC)–MS/MS analysis is very limited. According to these observations, we are currently in the process of developing novel labeling reagents for chiral molecules in MS/MS analysis. The derivatization reagent that is effective for enhancing not only the electrospray ionization–MS/MS sensitivity but also the reversed-phase LC resolution of carboxylic acid enantiomers should have a highly proton-affinitive moiety and an asymmetric structure near the reactive functional group. Furthermore, the resulting derivative has to provide a characteristic product ion suitable for the selected reaction monitoring. Based upon these considerations, a series of prolylamidepyridines ((S)-N-pyrrolidine-2-carboxylic acid N-(pyridine-2-yl)amide (PCP2), (S)-N-pyrrolidine-2-carboxylic acid N-(pyridine-3-yl)amide, and (S)-N-pyrrolidine-2-carboxylic acid N-(pyridine-4-yl)amide) was synthesized as ideal labeling reagents for the enantioseparation of chiral carboxylic acids and evaluated in terms of separation efficiency and detection sensitivity by ultra-performance LC (UPLC)–MS/MS. Among the synthesized reagents, PCP2 was the most efficient chiral derivatization reagent for the enantioseparation of carboxylic acid. The Rs values and the detection limits of the derivatives of non-steroidal anti-inflammatory drugs, which were selected as the representative carboxylic acids, were in the range of 2.52–6.07 and 49–260 amol, respectively. The sensitive detection of biological carboxylic acids (detection limits, 32–520 amol) was also carried out by the proposed method using PCP2 and UPLC–MS/MS. The PCP2 was applied to the determination of carboxylic acids in human saliva. Several biological carboxylic acids, such as lactic acid (LA), 3-hydroxybutylic acid, maric acid, succinic acid, α-ketoglutalic acid, and citric acid, were clearly identified in the saliva of healthy persons and diabetic patients. Furthermore, the ratio of d-LA in diabetic patients was higher than that in normal subjects. Judging from these results, PCP2 seems to be a useful chiral derivatization reagent for the determination not only of chiral, but also achiral, carboxylic acids in real samples.

Soft Supercharging of Biomolecular Ions in Electrospray Ionization Mass Spectrometry

The charge states of biomolecular ions in ESI-MS can be significantly increased by the addition of low-vapor supercharging (SC) reagents into the spraying solution. Despite the considerable interest from the community, the mechanistic aspects of SC are not well understood and are hotly debated. Arguments that denaturation accounts for the increased charging observed in proteins sprayed from aqueous solutions containing SC reagent have been published widely, but often with incomplete or ambiguous supporting data. In this work, we explored ESI MS charging and SC behavior of several biopolymers including proteins and DNA oligonucleotides. Analytes were ionized from 100 mM ammonium acetate (NH₄Ac) aqueous buffer in both positive (ESI+) and negative (ESI–) ion modes. SC was induced either with m-NBA or by the elevated temperature of ESI capillary. For all the analytes studied we, found striking differences in the ESI MS response to these two modes of activation. The data suggest that activation with m-NBA results in more extensive analyte charging with lower degree of denaturation. When working solution with m-NBA was analyzed at elevated temperatures, the SC effect from m-NBA was neutralized. Instead, the net SC effect was similar to the SC effect achieved by thermal activation only. Overall, our observations indicate that SC reagents enhance ESI charging of biomolecules via distinctly different mechanism compared with the traditional approaches based on analyte denaturation. Instead, the data support the hypothesis that the SC phenomenon involves a direct interaction between a biopolymer and SC reagent occurring in evaporating ESI droplets.

Efficient phenolic oxidations using μ-oxo-bridged phenyliodine trifluoroacetate

The excellent oxidizing behavior of the μ-oxo-bridged phenyliodine trifluoroacetate 1 is revealed during the phenolic oxidations mediated by hypervalent iodine(III) reagents. The use of the μ-oxo-bridged compound 1 instead of PhI(OAc)₂ (PIDA) and PhI(OCOCF₃)₂ (PIFA) during the oxidative cyclization of phenols involving carbon-oxygen, carbon-nitrogen, and carbon-carbon bond formations could produce spirocyclized cyclohexadienones in comparable or somewhat better yields. Thus, we have concluded that the unique reagent 1 is a promising alternative to PIDA and PIFA, and the use of reagent 1 as a reasonable choice is recommended for the hypervalent iodine(III)-mediated phenolic oxidations as well as other transformations.     

Silicon photonic sensors incorporated in a digital microfluidic system

Label-free biosensing with silicon nanophotonic microring resonator sensors has proven to be an excellent sensing technique for achieving high-throughput and high sensitivity, comparing favorably with other labeled and label-free sensing techniques. However, as in any biosensing platform, silicon nanophotonic microring resonator sensors require a fluidic component which allows the continuous delivery of the sample to the sensor surface. This component is typically based on microchannels in polydimethylsiloxane or other materials, which add cost and complexity to the system. The use of microdroplets in a digital microfluidic system, instead of continuous flows, is one of the recent trends in the field, where microliter- to picoliter-sized droplets are generated, transported, mixed, and split, thereby creating miniaturized reaction chambers which can be controlled individually in time and space. This avoids cross talk between samples or reagents and allows fluid plugs to be manipulated on reconfigurable paths, which cannot be achieved using the more established and more complex technology of microfluidic channels where droplets are controlled in series. It has great potential for high-throughput liquid handling, while avoiding on-chip cross-contamination. We present the integration of two miniaturized technologies: label-free silicon nanophotonic microring resonator sensors and digital microfluidics, providing an alternative to the typical microfluidic system based on microchannels. The performance of this combined system is demonstrated by performing proof-of-principle measurements of glucose, sodium chloride, and ethanol concentrations. These results show that multiplexed real-time detection and analysis, great flexibility, and portability make the combination of these technologies an ideal platform for easy and fast use in any laboratory.

Schiff base compounds derived from (R)-3-phenyl-2-phthalimidopropionic acid: photochromism, solvatochromism, and fluorescence

The photochromic behaviors of four Schiff bases derived from (R)-3-phenyl-2-phthalimidopropionic acid were studied to reveal the substituent effect on the photosensitivity. Upon ultraviolet light radiation, all of compounds 1–4 exhibit photochromic behavior in solution through intramolecular hydrogen atom transfer. In solid state, only compound 2 is photochromic, which may be due to the presence of meta-site methoxyl. In solution, the photochromic behavior of compound 3 is remarkable than the other compounds, which may be ascribed to the presence of para-site hydroxyl. Only compound 4 exhibits solvatochromism, which may be ascribed to the large dissociation tendency of the naphthol hydroxyl. The influences of acidity on the UV–Vis absorption spectra of the title compounds were also studied.     

p-Tolyl isocyanate derivatization for analysis of CWC-related polar degradation products by mass spectrometry

Most of the precursors and/or degradation products related to the Chemical Weapons Convention (CWC) are polar. Identification of these molecules in environmental samples provides clues regarding the alleged usage and/or synthesis of the parent toxic chemicals. Such polar compounds need to be derivatized in order to analyze them by gas chromatography–mass spectrometry (GC–MS). In this study, we developed a new derivatizing reagent, para-tolyl isocyanate (PTI), for derivatization of polar CWC-related compounds. The PTI reagent selectively derivatizes the –OH and/or?SH functional groups with high efficiency, but does not react with carboxylic acid (?COOH) or phosphonic acid (?(O)P(OH)₂) groups. The PTI derivatives of dialkyl aminoethanols, dialkyl aminoethanol-N-oxides, and 3-quinuclidinol were successfully eluted through GC, and their electron ionization (EI) mass spectra were distinct and provided the structure information by which the isomeric compounds can be easily distinguished. We also calculated the GC-retention index values that can be used for further confirmation of the target compounds. All the studied PTI derivatives can be analyzed by EI-MS with direct insertion probe and/or by direct electrospray ionization mass spectrometry (ESI-MS) together with the MS–MS data; both sets of data provide full structure information. The PTI reagent was found to be better in some respects than the conventional bistrimethylsilyl trifluoroacetamide (BSTFA), a trimethyl silylating reagent. The PTI reagent is commercially available, and the PTI derivatives are highly stable for months and are not sensitive to moisture. The applicability of the PTI derivatization for trace-level determination of the target CWC-related polar compounds in environmental matrices and in human plasma samples is also evaluated.

Possibilities of inductively coupled plasma atomic emission spectrometry with electrothermal vaporization in the analysis of high-purity reagents

Possibilities of electrothermal sample vaporization in inductively coupled plasma atomic emission spectrometry (ETV-ICP-AES) in the analysis of high-purity reagents were studied on an example high-purity waters, acid solutions, and trace impurity concentrates. The analytical and background signals in the injection of solutions into inductively coupled plasma (ICP) by pneumatic nebulization and electrothermal vaporization were compared and the of limits of detection in the analysis of high-purity reagents with impurity preconcentration by evaporation were estimated and compared.     

Multi-step synthesis of nanoparticles performed on millisecond time scale in a microfluidic droplet-based system

This paper reports a plug-based, microfluidic method for performing multi-step chemical reactions with millisecond time-control. It builds upon a previously reported method where aqueous reagents were injected into a flow of immiscible fluid (fluorocarbons)(H. Song et al., Angew. Chem. Int. Ed., 2003, 42, 768). The aqueous reagents formed plugs--droplets surrounded and transported by the immiscible fluid. Winding channels rapidly mixed the reagents in droplets. This paper shows that further stages of the reaction could be initiated by flowing additional reagent streams directly into the droplets of initial reaction mixture. The conditions necessary for an aqueous stream to merge with aqueous droplets were characterized. The Capillary number could be used to predict the behavior of the two-phase flow at the merging junction. By transporting solid reaction products in droplets, the products were kept from aggregating on the walls of the microchannels. To demonstrate the utility of this microfluidic method it was used to synthesize colloidal CdS and CdS/CdSe core-shell nanoparticles.     

Extraction of transplutonium elements with diphenyl (alkyl)dialkyl-carbamoylmethyl phosphine oxides

Polydentate neutral organophosphorus compounds attract great interest as efficient extractants of transplutonium elements(TPE) from acid waste nuclear fuel element solution. With the view of a further search, reagents R′R″-PO-CH₂-CNO-R ₂ ^″' were synthetized and investigated. The solubility of R ₂ ^′ POCH₂CNOR ₂ ^″' vas investigated in organic solvents and in nitric acid solutions. This reagent was found to be the best one.