Rapid, selective and direct spectrophotometeric determination of aliphatic amines with m-dinitrobenzene

Color reaction has been studied for identification and spectrophotometric determination of aliphatic amines at room temperature by m-dinitrobenzene (m-DNB) as reagent. The λ_max value ranges from 458 to 570 nm. This is a simple and rapid method for determination of aliphatic amines in the acidic, water and acetone medium. Beer's law is verified for methylamine, dimethylamine, trimethylamine and n-butylamine in the range of 0.5–8 mg l⁻¹. The effect of pH on the molar absorptivity is investigated for a representative primary amine i.e. methylamine and it was observed that molar absorptivity increases from acidic to basic pH, with a sharp increase at pH 12. The kinetic of reaction was also studied and found that reaction time has marked effect on the molar absorptivity of electron donor–acceptor (EDA) complex. The detection of methyl amine has been reported in three real samples of water.     

Effect of moisture on copolymer fibers based on 5-amino-2-(p-aminophenyl)-benzimidazole

In recent years, there has been concern in the soft body armor community that copolymer fibers based on 5-amino-2-(p-aminophenyl)-benzimidazole can release hydrochloric acid, which is present in these fibers as a by-product of the manufacturing process. The presence of acids could potentially be detrimental to other fibers that might come in contact with these materials. In an effort to examine this issue, a study was designed to investigate the release of acid in different environments from these fibers. During the first phase of the study, fibers were exposed to water and pH decreases were observed. While immersed in deionized water, two of the fiber samples studied released a sufficient amount of acid to drop the pH of the solution from approximately pH 6.0 to approximately pH 3.0 in less than 10 d at room temperature. Further ion-selective electrode studies of chloride ion released from these fibers indicated that hydrochloric acid may not be the species responsible for this pH reduction. In a second phase of the investigation, fibers were exposed to water vapor in an elevated temperature environment (conditions were 65 °C, 80% RH). While the pH reduction released by the water vapor exposure was substantially less than observed in the submersion phase, a reduction in the yarn tensile strength of some of the fibers was observed during this phase of the study. In a third phase, fibers were exposed in a dry oven (less than 5% RH) at 65 °C. Almost no pH reduction or strength reduction was observed. Molecular spectroscopy was also performed to better understand the effect of elevated temperature and moisture environments on these fibers.     

Mechanism for the reduction of the mixed-valent MnIIIMnIV[2-OHsalpn]2+ complex by tertiary amines

The mixed-valent dimanganese(III/IV) complex MnIIIMnIV(2-OHsalpn)2+, 1, is cleanly reduced in acetonitrile by aliphatic tertiary amines to give the dimanganese(III) product MnIII2(2-OHsalpn)2, 2. Thorough characterization of the organic reaction products shows that tributylamine is converted to dibutylformamide and propionaldehyde. Kinetic studies and radical trapping experiments suggest that this occurs via initial single-electron transfer from the amine to 1 coupled with C-H alpha proton transfer from the oxidized amine. EPR spectroscopy and base inhibition studies indicate that coordination of the amine to 1 is a critical step prior to the electron transfer step. Rate data and its dependence on the amine indicate that the ability of the amine to reduce 1 is correlated to its basicity rather than to its reduction potential. Weakly basic amines were unable to reduce 1 irrespective of their reduction potential. This was inferred to indicate that proton transfer from the amine radical cation is also important in the reduction of 1 by tertiary amines. Comparison of the activation energy with reaction thermodynamics indicates that proton transfer and electron transfer must be concerted to explain the rapidity of the reaction. The fate of the amine radical is dependent on the presence of oxygen, and labeling studies show that oxygen in the organic products arises from dioxygen, although incorporation from trace water was also observed. These data indicate that inhibition of the hydrolytic quenching of the amine radical in an aprotic solvent results in a different fate for the amine radical when compared to amine oxidation reactions in aqueous solution. The proposed mechanism gives new insight into the ability of amines with high reduction potential to reduce metal ions of lower potential. In particular, these data are consistent with the ability of small amines and certain amine-containing buffers to inhibit manganese-dependent oxygen evolution in photosynthesis, which arises in some cases as a result of manganese reduction and its concomitant loss from the PS II reaction center.     

Determination of aromatic amines formed from azo colorants in toy products

A study for the determination of the aromatic amines formed after reduction of the azo colorants mostly used in toys was conducted. Sodium dithionite was used in the reductive cleavage of the azo group for the dyes, and the released amines were subsequently analysed by high-performance liquid chromatography with UV detection. The influence of different variables related to the reduction process was investigated by the use of a full-level factorial design, where most significant parameters as well as order interactions were studied. Reduction profiles for each colorant were obtained by studying the changes in the amount of amine obtained with different dithionite/colorant ratios. The expected aromatic amines forming azo colorants were detected, and in the presence of a nitro group a further reduction was observed. The yield of the total reduction process was determined by using standard addition of different quantities of amines to the colorants.     

Chromatographic methods for the analysis of heterocyclic amine food mutagens/carcinogens.

A series of potent heterocyclic amines that are mutagenic and carcinogenic have been discovered that are formed in some heated foods, most notably, meats derived from muscle. Determining the heterocyclic amine content in foods and food products is required for toxicological research, industry quality control, and possibly in the future, regulatory control. The contents of food needs to be determined using reliable analytical techniques. Since heterocyclic amines are present in foods at ng/g levels, a variety of liquid-liquid or solid-phase purification techniques are required, followed by gas or high-performance liquid chromatography. Peak detection has been successful using UV, fluorescence, and mass spectrometric detection, and biological activity using the Ames/Salmonella test. The low levels present require that chromatographic efficiency, and both detector sensitivity and selectivity be optimized. The cartridge solid-phase extraction and high-performance liquid chromatography method have been used to measure the known food-derived heterocyclic amines for several types of food, and to the authors knowledge, this is the only method undergoing intralaboratory comparison and validation. Our analysis of the literature shows that chromatographic analysis of the heterocyclic amines by high-performance liquid chromatography or gas chromatography (with derivatization) is satisfactory for heterocyclic amine analysis in foods although the methods are just now being optimized for routine use. The biggest improvements in speed and accuracy will probably come from improved extraction methods as analysis of complex food samples for heterocyclic amines will always be a challenge.     

Monitoring of aromatic amines by HPLC with electrochemical detection: comparison of methods for destruction of carcinogenic aromatic amines in laboratory wastes

A new chemical method for destruction of carcinogenic aromatic amines in laboratory wastes has been developed. The method is based on enzymatic oxidation of the amines in solution (with hydrogen peroxide and horseradish peroxidase), followed by oxidation of the solid residues with permanganate in sulphuric acid medium. To monitor the efficiency of destruction, a reversed-phase HPLC system has been developed, with voltammetric detection with a carbon-fibre detector, which is substantially more sensitive (detection limits from a few ng down to a few pg of amine) than the commonly used ultraviolet photometric detection. It has been demonstrated that the proposed method of destruction is highly efficient (99.8% destruction).     

pH-dependent spectral properties of para-aminobenzoic acid and its derivatives

The local environment dictates the structural and functional properties of many important chemical and biological systems. The impact of pH on the photophysical properties of a series of para-aminobenzoic acids is examined using a combination of experimental spectroscopy and quantum chemical calculations. Following photoexcitation, PABA derivatives may undergo an intramolecular charge transfer (ICT) resulting in the formation of a zwitterionic species. The thermodynamics of the excited state reaction and temperature-dependence of the radiative emission processes are evaluated through variable temperature fluorescence spectroscopy carried out in a range of aqueous buffers. Quantum chemical calculations are used to analyze structural changes with modifications at the amine position and different protonation states. The ICT is only observed in the tertiary amine, which calculations show has more sp(2) character than the primary or secondary amines. Thermodynamic analysis indicates the ICT reaction is driven by entropy.     

Deamination of protonated amines to yield protonated imines

Primary and secondary amines, when examined in atmospheric pressure chemical ionization, electrospray ionization, or chemical ionization, display protonated imines in their mass spectra. These products arise formally by nucleophilic substitution at the α-carbon with loss of both ammonia and molecular hydrogen. Collision-induced dissociation (CID) is used to characterize the product ions by comparison with authentic protonated imines. Gas-phase ion/molecule reactions of protonated amines with neutral amines also yield products that correspond to protonated imines (deamination and dehydrogenation), as well as providing simple deamination products. The reaction mechanism was investigated further by reacting the deamination product, the alkyl cation, with a neutral amine. The observed dehydrogenation of the nascent protonated secondary amine indicates that the reaction sequence is loss of ammonia followed by dehydrogenation even though the isolated protonated secondary amines did not undergo dehydrogenation upon CID. Formation of the deamination products in the protonated amine/amine reaction is competitive with proton-bound dimer formation. The proton-bound dimers do not yield deamination products under CID conditions in the ion trap or in experiments performed using a pentaquadrupole instrument. This demonstrates that the geometry of the proton-bound dimer, in which the α-carbons of the alkylamines are well separated [C _a -N-H-N-C _a ], is an unsuitable entry point on the potential energy hypersurface for formation of the imine [C _a -N-C _a ]. Isolation of the proton-bound dimers in the quadrupole ion trap is achieved with low efficiency and this characteristic can be used to distinguish them from their covalently bound isomers.     

8-Phenyl-(4-oxy-acetic acid N-hydroxysuccinimidyl ester)-4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene as a new highly fluorescent-derivatizing reagent for aliphatic amines in disease-related samples with high-performance liquid chromatography

A novel fluorescent-activated ester, 8-phenyl-(4-oxy-acetic acid N-hydroxysuccinimidyl ester)-4,4-difluoro-1,3,5,7-tetramethyl-4-bora-3a,4a-diaza-s-indacene (TMPAB-OSu) has been designed and synthesized for amine labeling in HPLC. Being used 11 aliphatic amines as the models, the derivatization conditions were optimized. In 0.2 mol/l borate buffer (pH 8.8), amines reacted with TMPAB-OSu at 30 °C to form the derivatives in 10 min. The fluorescent quantum yield of TMPAB-OSu and its amine derivatives are high even compared with fluorescein. The separation of these amine derivatives was achieved with a C₈ column and gradient elution by using 0.1 mol/l sodium acetate buffer (pH 5.0) and methanol. With fluorescence detection at an emission wavelength of 509 nm and an excitation wavelength of 497 nm, the detection limits of aliphatic amines were 2–18 fmol, at a signal-to-noise ratio of 3:1. The proposed TMPAB-OSu-based HPLC method has been applied to the analysis of urine samples of health, hepatic and renal patients and lake water. Recoveries from different matrices are from 96 to 104%, depending on the sample investigated.     

Determination of biogenic amines in wines by pre-column derivatization and high-performance liquid chromatography coupled to mass spectrometry

A new HPLC method for determining biogenic amines in wines is developed. This method is based on pre-column amine derivatization, further separation of derivatives and on-line hyphenation of HPLC to atmospheric pressure chemical ionization mass spectrometry (APCI-MS). Biogenic amines have been derivatized with 1,2-naphthoquinone-4-sulfonate at 65 °C and pH 9.2 for 5 min. The separation of derivatives has been accomplished in a C₁₈ analytical column using an elution gradient based on increasing the percentage of methanol. Derivatives have been ionized in positive mode and detected by selected ion monitoring. The operating conditions of the APCI-MS system (voltages, temperatures and gases) have been thoroughly optimized to obtain the maximum sensitivity for all analytes. In the selected conditions, APCI-MS spectra display little fragmentation and good signal-to-noise ratio. Depending on the amine characteristics, the main spectral peaks are due to mono- and di-derivative products. Figures of merit of the method have been established under the selected conditions using red wine samples. Recoveries ranging from 94% to 106% have been obtained which prove excellent accuracy of the method in the determination of histamine, putrescine, cadaverine, tryptamine, phenylethylamine, tyramine and serotonin in red wines. The proposed method has been applied to the analysis of commercial wines from different Spanish regions.     

Characterisation of leather candidate certified reference materials for their mass fractions of aromatic amines

Three batches of leather samples were coloured with nine azo dyes that can yield eight proven or suspected carcinogenic aromatic amines under reduction conditions. The samples were milled to grains and bottled in jars. A group of five laboratories has established the mass fraction of the amines in a ring test using different analytical methods. The methods included a reduction step in order to cleave the azo dyes into the aromatic amines. Quantification was by standard addition: sub-samples of the leathers were spiked with known amounts of azo dyes of known purity. It was possible to establish the mass fractions of six of the eight aromatic amines in three of the leather samples.     

Direct reductive amination of aldehydes using lithium-arene(cat.) as reducing system. A simple one-pot procedure for the synthesis of secondary amines

A simple one-pot procedure for the direct reductive amination of aldehydes using lithium powder and a catalytic amount of 4,4′-di-tert-butylbiphenyl (DTBB) or a polymer supported naphthalene as reducing system is described. The direct reductive amination of a variety of aldehydes with primary amines was achieved simply by adding a mixture of the corresponding carbonyl compound and the amine, over a solution of the lithium arenide in THF at room temperature. For most of the substrates tested the main reaction products were the secondary amines along with variable amounts of the corresponding alcohol and/or imine products. Theoretical DFT calculations have been applied in order to explain the differences in reactivity observed for aromatic substrates.     

Insertion of Chemical Handles into the Backbone of DNA during Solid-Phase Synthesis by Oxidative Coupling of Amines to Phosphites

Conjugation of molecules or proteins to oligonucleotides can improve their functional and therapeutic capacity. However, such modifications are often limited to the 5′ and 3′ end of oligonucleotides. Herein, we report the development of an inexpensive and simple method that allows for the insertion of chemical handles into the backbone of oligonucleotides. This method is compatible with standardized automated solid-phase oligonucleotide synthesis, and relies on formation of phosphoramidates. A unique phosphoramidite is incorporated into a growing oligonucleotide, and oxidized to the desired phosphoramidate using iodine and an amine of choice. Azides, alkynes, amines, and alkanes have been linked to oligonucleotides via internally positioned phosphoramidates with oxidative coupling yields above 80 %. We show the design of phosphoramidates from secondary amines that specifically hydrolyze to the phosphate only at decreased pH. Finally, we show the synthesis of an antibody-DNA conjugate, where the oligonucleotide can be selectively released in a pH 5.5 buffer.     

C-N coupling of 3-methylcatechol with primary amines using native and recombinant laccases from Trametes versicolor and Pycnoporus cinnabarinus

Five laccase genes from Pycnoporus cinnabarinus and Trametes versicolor encoding for different isoenzymes have been cloned, recombinantly expressed and characterized. Following C-N coupling of primary linear, branched-chained and cyclic amines to 3-methylcatechol was mediated by native and recombinant laccases yielding the corresponding secondary amines. Formation of C5-monoaminated ortho-methylquinones occurred within 1-2 h; prolonged incubation led to the formation of high-molecular mass products. No difference between the use of native or recombinant isoenzymes from P. cinnabarinus or T. versicolor was observed. Optimization of the reaction conditions included variation of amine donor ratios, pH, amount and type of enzyme preparations. The formation of by-products could be suppressed at pH values corresponding to the enzymes optima (pH 4-5). A total of 10 secondary amines were synthesized with product formations of up to 80%. Furthermore, all purified secondary amines were characterized by NMR-, LC-MS- and HRMS-analysis and log P values were determined.     

Fate of tris(2-chloroethyl)amine in water and alkaline environment determined by thin-layer chromatography and gas chromatography–mass spectrometry

Tris(2-chloroethyl)amine is a chemical warfare agent which is considered to be a persistent contaminant highly resistant to decontamination. The time dependence of tris(2-chloroethyl)amine degradation on the water pH value is observed to determine decontamination options using the decontamination of hydroxide-based mixtures which are used by a number of armed forces. The kinetics was observed using thin-layer chromatography and gas chromatography. The time development of tris(2-chloroethyl)amine concentration decrease in the environment with the pH ranging from 7.5 to 12.5 was recorded. The reaction rate constants were established at all observed pH values, half-lifes of reactions and the influence of temperature on the reaction rate at a slightly alkaline pH of 8.5. The equation to estimate the rate constant of the first step of tris(2-chloroethyl)amine hydrolysis at various temperatures was formed. The rate of triethanolamine formation in the samples of neutral to alkaline pH was recorded. The results indicate the substantial influence of pH on tris(2-chloroethyl)amine hydrolysis. The rate constant was significantly increased from a neutral environment to the pH of 12.5 reaching 500% of the original value using alkalisation. The hydrolysis in a neutral environment was also influenced by the analyte concentration. The concentrated solutions hydrolysed more slowly as a result of hydrochloric acid release as the acid lowered the pH value of the environment. The results demonstrate that there is an important kinetic difference between sulphur and nitrogen mustard hydrolysis in an alkaline environment.

     

The determination and characterisation of 2-naphthyloxycarbonyl chloride derivatised biogenic amines in pet foods

The need to monitor biogenic amines levels is essential for many areas of the food industry for two main reasons: the caustic nature and potential toxicity of these amines, and the potential to use amine levels as markers for freshness and quality in foodstuffs. Optimised analysis conditions used for the determination of biogenic amines derivatised with 2-napthyloxycarbonyl chloride has been applied to different pet food samples to assess the effectiveness of this method for complex sample matrices. Further to this, the use of high-resolution mass spectrometry has enabled the previously unconfirmed derivatised form of seven biogenic amines to be established. The derivatised forms identified include as mono substituted (tryptamine and histamine), bisubstituted (putrescine, cadaverine and tyramine), trisubstituted (spermidine) and tetrasubstituted (spermine). The methodology of biogenic amine determination was performed successfully to a range of pet food products highlighting the applicability to a variety of complex sample matrices.     

Voltammetric analysis of the possibility of derivatization of levodopa in the presence of aniline derivatives

Electrochemical oxidation of levodopa (LD) as one of the most well-known neurotransmitters has been studied in the presence of some aniline derivatives. The electron transfer of LD is followed by two competitive reactions in the presence of these amines. The reactions are the Michael additions of side chain amine group of LD and/or aromatic amines to electrochemically generated o-quinone. There are two ECE mechanisms for both pathways and the competition between these inter and intramolecular reactions drastically depends on the pH of the medium. The pH dependence of reactions has been studied and the observed homogeneous rate constants of the reactions were estimated by digital simulation of cyclic voltammograms. The effect of aniline substituents was also studied with regard to their reactivities toward o-quinone of LD and the competitive reactions. Based on the obtained results, the products of intermolecular reactions are electroactive diphenylamine derivatives and their half-wave potentials depend on the nature of the aniline substituent.     

Mechanisms of amine-catalyzed organosilicate hydrolysis at circum-neutral pH

Mono- and polyamines can catalyze the hydrolysis and condensation of organosilicate starting materials in biomimetic silica synthesis pathways at circum-neutral pHs and room temperature. Our study is focused on understanding the mechanistic role of amines in catalyzing the hydrolysis process that precedes condensation. We have conducted (29)Si NMR experimental studies over a range of temperature and pHs for the hydrolysis rates of trimethylethoxysilane (TMES), a model compound with only one hydrolyzable bond, combined with quantum mechanical hybrid density functional theory calculations of putative intermediate and transition-state structures for TMES and tetramethyl orthosilicate (TMOS). Comparison of calculated energies with experimentally determined activation energies indicates that amine catalysis of TMES is primarily a consequence of the amine's acidity at neutral pH. The proton released by the amine is transferred to the organosilicate, producing a protonated ethoxy leaving group that can be displaced by water in an S(N)2 reaction. For TMOS, the activation energy of proton-transfer coupled with S(N)2 substitution is comparable to that for Corriu's nucleophile-activated nucleophilic displacement, such that the mechanism of amine-catalyzed hydrolysis is dependent mostly on the ambient pH conditions as well as the type of amine. The relevance of our results to biological silica precipitation is discussed.     

Determination of trace biogenic amines with 1,3,5,7-tetramethyl-8-(N-hydroxysuccinimidyl butyric ester)-difluoroboradiaza-s-indacene derivatization using high-performance liquid chromatography and fluorescence detection

A reversed-phase high-performance liquid chromatographic method based on chemical derivatization with fluorescence detection has been developed for analyzing biogenic amines in food and environmental samples. A BODIPY-based fluorescent reagent, 1,3,5,7-tetramethyl-8-(N-hydroxysuccinimidyl butyric ester)-difluoroboradiaza-s-indacene (TMBB-Su), was employed for the derivatization of these biogenic amines at 20 °C for 20 min in pH 7.20 borate buffer after careful investigation of the derivatization conditions including reagent concentration, buffer solution, reaction temperature and reaction time. Separation of biogenic amines with gradient elution was conducted on a C8 column with methanol-tetrahydrofuran-water as mobile phase. The detection limits were obtained in the range from 0.1 to 0.2 nM (signal-to-noise=3). This procedure has been validated using practical samples. The study results demonstrated a potential of employing high-performance liquid chromatography (HPLC) with 1,3,5,7-tetramethyl-8-(N-hydroxysuccinimidyl butyric ester)-difluoroboradiaza-s-indacene labeling as a tool for quantitative analysis of biogenic amines involved in various matrices.     

A new route to poly(N-isopropylacrylamide) microgels supporting a polyvinylamine corona

Poly(N-isopropylacrylamide) thermoresponsive microgel particles with an amine-rich corona were prepared by the copolymerization of N-isopropylacrylamide with N-vinylformamide, NVF. Hydrolysis above the volume phase transition temperature converted the surface formamide moieties to the corresponding amine. The surface amine concentration was enriched by coupling iodine-terminated polyNVF oligomers (DP=7) to the microgel amines, followed by a second hydrolysis to give the corresponding polyvinylamine. Microgel swelling and electrophoretic mobility values as functions of pH and temperature were consistent with published results for amine-containing microgels.