Mechanochemical Synthesis and Isomerization of N-Substituted Indole-3-carboxaldehyde Oximes †.

Performing solution-phase oximation reactions with hydroxylamine hydrochloride (NH2OH·HCl) carries significant risk, especially in aqueous solutions. In the present study, four N-substituted indole-3-carboxaldehyde oximes were prepared from the corresponding aldehydes by solvent-free reaction with NH2OH·HCl and a base (NaOH or Na2CO3) using a mechanochemical approach, thus minimizing the possible risk. In all cases, the conversion to oximes was almost complete. The focus of this work is on 1-methoxyindole-3-carboxaldehyde oxime, a key intermediate in the production of indole phytoalexins with useful antimicrobial properties. Under optimized conditions, it was possible to reach almost 95% yield after 20 min of milling. Moreover, for the products containing electron-donating substituents (-CH3, -OCH3), the isomerization from the oxime anti to syn isomer under acidic conditions was discovered. For the 1-methoxy analog, the acidic isomerization of pure isomers in solution resulted in the formation of anti isomer, whereas the prevalence of syn isomer was observed in solid state. From NMR data the syn and anti structures of produced oximes were elucidated. This work shows an interesting and possibly scalable alternative to classical synthesis and underlines environmentally friendly and sustainable character of mechanochemistry.

Effect of oximation reagents on gas chromatographic separation of eight different kinds of mono- and di-saccharides.

The objective of this study was to investigate the effect of oximation reagents in simultaneous analysis of mono and di-saccharides using gas chromatography. Sugar oximation with O-ethylhydroxylamine separated all the mono- and di-saccharides while hydroxylamine and O-benzylhydroxylamine could make most of the saccharides separable except for xylose and arabinose. Resolution of xylose: arabinose, galactose: glucose, and fructose: galactose oximated by O-ethylhydroxylamine in DB-1ms column were 1.66, 2.15, and 6.19, respectively, which were above 1.5 and were officially acceptable for quantitative analysis according to the AOAC guideline. The applied method was then verified by the method validation parameters; LOD (0.011-0.02 mg/100 g), LOQ (0.032-0.061 mg/100 g), linearity (R2 = 0.9991-1.0000) and precision (repeatability RSD: 1.4-3.3%, reproducibility RSD: 1.7-7.6%). The greatest amounts of xylose (19.03 ± 0.38 mg/100 g), maltose (6,274.48 ± 173.59 mg/100 g) were found in the oyster sauce, and the contents of glucose (10,565.00 ± 125.31 mg/100 g), galactose (170.40 ± 4.62 mg/100 g) were greatest in soybean paste.

Trimethylsilyl speciations of cathine, cathinone and norephedrine followed by gas chromatography mass spectrometry: Direct sample preparation and analysis of khatamines.

A literature criticism is given on methods using currently gas chromatography mass spectrometry (GC/MS) to determine cathine (CAT), cathinone (CTN) and norephedrine (NE), jointly khatamines. In this study, khatamines' oximation, trimethylsilylation and mass fragmentation properties-applying N-Methyl-N-(trimethylsilyl)trifluoroacetamide (MSTFA), its trimethyliodosilane (TMIS) catalyst containing version (MSTFA(TMIS)), N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) and hexamethyldisilazane (HMDS)-was highlighted, at first. Derivatization, mass fragmentation and quantitation related, optimized model investigations have been carried out as a function of the reaction times and conditions. Special emphasis was put (i) on the stability of the primarily formed (CAT-2TMS, NE-2TMS, CTN-TMS(TMS-oximes)1,2), then transformed, fully derived (CAT-3TMS, NE-3MTS, CTN-2TMS(TMS-oximes)1,2) species, and, (ii) on the proportionally formed stable products, suitable to selective quantitation of all three natural amines, simultaneously. Results, as novelty to the field confirmed that (i) TMIS catalyzed trimethylsilyation triggers to form fully derivatized species unfortunately, in part only; while, (ii) khatamines' simultaneous quantitation needs to be carried out in a two steps derivatization process consisting of oximation (1st step, hydroxylamine in pyridine) and trimethylsilylation (2nd step, MSTFA), to the CAT-2TMS, NE-2TMS, CTN-TMS(TMS-oximes)1,2. These species were characterized with their retention, mass fragmentation and analytical performance properties, in model solutions and in the presence of plant tissues, as well: R(2), limit of quantitation (LOQ) data, expressed in pg/1µL injection basis, proved to be 62.5pg (CAT), 20pg (NE) and 62.5pg (CTN), respectively. The practical utility of proposal was enormously enhanced by the novel, direct sample preparation method. In this process, the freshly harvested, freeze-dried, then pulverized leaves of Catha edulis FORKS were directly derivatized, in the presence of the matrix. Reproducibility (in average 2.07 RSD% varying between 0.15 and 5.5 RSD%), linearity (0.9990-0.9994) and recovery (95.7-99.1%) values of the new sample preparation protocol was confirmed by the standard addition method for CAT, NE and CTN equally. From plant leaf, 0.061w/w% CAT and 0.014w/w% NE contents were obtained. In this tissue CTN was not found. Very likely attributable to the unfavorable climate for the plant: grown in Hungary of temperate zone and naturalized in the tropical Africa.

Structure-related related new approach in the gas chromatography/mass spectrometry analysis of cathinone type synthetic drugs.

A novel, structure-related derivatization principle has been developed in order to quantify cathinone-type synthetic drugs (CTSDs), focusing on the most common pentedrone (PENT), including also 4-fluoromethcathinone (4-FMC), methcathinone (MCTN), 4-methylethcathinone (4-MEC), 3,4-dimethylmethcathinone (3,4-DMMC), and 4-ethylmethcathinone (4-EMC). Firstly, oximated and, secondly, trimethylsilylated CTSD derivatives were characterized by mass fragmentation patterns using GC/MS that led to the development of a harmonized, quantitative, two-steps derivatization methodology. The two-step process involved i) oximation with hydroxylamine hydrochloride; and ii) trimethylsilylation with N-methyl-N-(trimethylsilyl)-trifluoroacetamide (MSTFA). Next, the oximated-trimethylsilylated species were characterized by retention and mass fragmentation properties. Due to α-cleavage decomposition at their C1C2 bonds without exception, CTSDs uniformely exhibited a structure-related fragmentation pattern. The practical utility of this newly recognized mechanism was validated in urine samples employing an extraction-free and time-, work-, cost- and solvent-effective protocol in accordance with Green Chemistry. The centrifuged urines (10-40µL) were evaporated to dryness, followed by derivatization. The analytical performance of the methodology was characterized by repeatability (RSD%, varying between 1.43% and 5.44%), limit of quantitation (LOQ, 15-24µg/mL), linearity (R2, 0.9976-0.9998) and recovery (97-99%) values. The new principle was tested on drug users' urine: one specimen provided 56.8µg/mL PENT (3.8 RSD%). Simple trimethylsilylation of CTSDs confirmed their special fragmentation patterns, not yet described. The instantenous combination of the primarily formed, characteristic fragments with the mass m/z 44, led to the special equilibrium of products: confirming that direct trimethylsilylation of CTSDs is not suitable for their quantitation.

Versatile preparation of intracellular-acidity-sensitive oxime-linked polysaccharide-doxorubicin conjugate for malignancy therapeutic.

Recently, chemotherapy has been one of the most important therapeutic approaches for malignant tumors. The tumor tissular or intracellular microenvironment-sensitive polymer-doxorubicin (DOX) conjugates demonstrate great potential for improved antitumor efficacy and reduced side effects. In this work, the acid-sensitive dextran-DOX conjugate (noted as Dex-O-DOX) was synthesized through the versatile efficient oximation reaction between the terminal aldehyde group of polysaccharide and the amino group in DOX in the buffer solution of sodium acetate/acetic acid. The insensitive one, i.e., Dex-b-DOX, was prepared similarly as Dex-O-DOX with a supplemented reduction reaction. The DOX release from Dex-O-DOX was pH-dependent and accelerated by the decreased pH. The efficient intracellular DOX release from Dex-O-DOX toward the human hepatoma HepG2 cells was further confirmed. Furthermore, Dex-O-DOX exhibited a closer antiproliferative activity to free DOX·HCl as the extension of time. More importantly, compared with Dex-b-DOX, Dex-O-DOX exhibited higher antitumor activity and lower toxicity, which were further confirmed by the systemic histological and immunohistochemical analyses. Hence, the facilely prepared smart polysaccharide-DOX conjugates, i.e., Dex-O-DOX, exhibited great potential in the clinical chemotherapy of malignancy.

Site-Specific Protein Bioconjugation via a Pyridoxal 5'-Phosphate-Mediated N-Terminal Transamination Reaction.

The covalent attachment of chemical groups to proteins is a critically important tool for the study of protein function and the creation of protein-based materials. Methods of site-specific protein modification are necessary for the generation of well defined bioconjugates possessing a new functional group in a single position in the amino acid sequence. This article describes a pyridoxal 5'-phosphate (PLP)-mediated transamination reaction that is specific for the N-terminus of a protein. The reaction oxidizes the N-terminal amine to a ketone or an aldehyde, which can form a stable oxime linkage with an alkoxyamine reagent of choice. Screening studies have identified the most reactive N-terminal residues, facilitating the use of site-directed mutagenesis to achieve high levels of conversion. Additionally, this reaction has been shown to be effective for a number of targets that are not easily accessed through heterologous expression, such as monoclonal antibodies. Curr. Protoc. Chem. Biol. 2:125-134 © 2010 by John Wiley & Sons, Inc.