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1.
J Biosci Bioeng ; 130(4): 437-442, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32616382

RESUMO

Fast enantiomeric separation of amino acids was studied by liquid chromatography/mass spectrometry (LC/MS) on a chiral crown ether stationary phase. A chiral crown ether bonded silica column (3 mm internal diameter (i.d.), 5 cm long) packed with 3 µm particles was employed instead of a 15 cm column packed with 5 µm particles used in our previous study. In addition, the extra-column variance, becoming more serious for smaller columns, was reduced by replacing 0.127 mm i.d. post-column tubes with shorter, smaller-diameter (0.0635 mm i.d.) tubes. The results demonstrated the benefits of using shorter columns packed with smaller particles and the reduction of the extra-column band broadening for fast enantiomeric separation. Finally, the enantiomeric separation of 18 pairs of proteinogenic amino acids was achieved within 2 min with a resolution (Rs) > 1.5 for each pair using an isocratic mobile phase of acetonitrile/water/trifluoroacetic acid (ACN/W/TFA) = 96/4/0.5, and a flow rate 1.2 mL/min at 30°C. This is the highest throughput method for simultaneous chiral separation of all proteinogenic amino acids except proline to date.


Assuntos
Aminoácidos/química , Aminoácidos/isolamento & purificação , Cromatografia Líquida/métodos , Éteres de Coroa/química , Espectrometria de Massas , Acetonitrilos/química , Aminas/química , Dióxido de Silício/química , Estereoisomerismo , Fatores de Tempo , Ácido Trifluoracético/química , Água/química
2.
J Chromatogr A ; 1624: 461235, 2020 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-32540075

RESUMO

In this study, 31 racemates of Nα-FMOC (fluorenylmethoxycarbonyl) amino acids (AAs) with different chemico-physical characteristics (neutral nonpolar, neutral polar, acidic and basic) have been successfully resolved in fast enantioselective chromatography on recently-developed zwitterionic-teicoplanin chiral stationary phases (CSPs). The CSPs were prepared by covalently bonding the teicoplanin selector on fully-porous particles of narrow dispersion particle-size distribution (particle diameter 1.9 µm) and superficially-porous particles (2.0 µm). Both the zwitterionic-teicoplanin CSPs have proved to be ideal media for the separation of this important class of compounds. In particular, the zwitterionic CSP prepared on superficially-porous particles exhibited superior enantioselectivity and resolution, compared to that made of fully porous particles, in virtue of more favorable thermodynamics. The zwitterionic nature of these CSPs allowed avoiding the annoying effect of Donnan's exclusion of enantiomers from the stationary phase. This effect, on the opposite, was frequently observed on a commercial teicoplanin CSP (Teicoshell) employed for comparative purposes. Noticeably, on the zwitterionic-teicoplanin CSPs, by using either acetonitrile- or methanol-rich mobile phases (MPs), it was possible to favor speed over enantioresolution and vice versa. This work gives further replies to the request for rapid determination of enantiomeric excess of Nα-FMOC proteinogenic (and non-proteinogenic) AAs, typically used as preferred chiral synthons in the solid-phase synthesis of therapeutic peptides.


Assuntos
Aminoácidos/isolamento & purificação , Cromatografia/métodos , Fluorenos/isolamento & purificação , Proteínas/química , Teicoplanina/química , Porosidade , Estereoisomerismo
3.
Chemosphere ; 253: 126728, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32298913

RESUMO

Nile perch wastewater was biodegraded using two Bacillus species to recover bioactive substances to enhance its reutilization value. The two Bacillus species successfully produced low-molecular-weight substances with a 47.8% degree of hydrolysis. The antioxidant activities of the Nile perch wastewater increased as the biodegradation proceeded, and the culture supernatant exhibited the highest DPPH (80.1%), ABTS (93.1%) and Fe2+ chelating (88.5%) antioxidant activities at 60 h. The antioxidant potential of the biodegraded Nile perch wastewater was found to be higher than those of other fish hydrolysates. Moreover, the biodegraded Nile perch wastewater exhibited effective antimicrobial activity against Vibrio vulnificus, exhibiting a minimal inhibitory concentration of 585 µg mL-1. Two-dimensional thin layer chromatography analysis revealed the specific amino acids responsible for the antioxidant activity, and molecular-weight cut-off ultrafiltration revealed that the <2-kDa fraction exhibited the highest antioxidant activity with the lowest IC50 values (0.43 and 0.22 mg mL-1 for DPPH and ABTS antioxidant activities, respectively). This is the first report of the reutilization of Nile perch wastewater as a natural antioxidant and antimicrobial ingredient for nutraceuticals.


Assuntos
Antibacterianos/isolamento & purificação , Antioxidantes/isolamento & purificação , Bacillus/metabolismo , Peptídeos/isolamento & purificação , Percas , Águas Residuárias/química , Aminoácidos/isolamento & purificação , Aminoácidos/farmacologia , Animais , Antibacterianos/farmacologia , Antioxidantes/farmacologia , Biodegradação Ambiental , Compostos de Bifenilo/química , Quelantes/isolamento & purificação , Quelantes/farmacologia , Pesqueiros , Hidrólise , Peso Molecular , Peptídeos/farmacologia , Percas/crescimento & desenvolvimento , Picratos/química , Ultrafiltração , Vibrio vulnificus/efeitos dos fármacos
4.
J Chromatogr A ; 1621: 461033, 2020 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-32188563

RESUMO

A diethylammonio-propylsulfate amphoteric ionic resin was synthesized and employed as the stationary phase for high-performance liquid chromatography-mass spectrometry (LC-MS) separation and detection of amino acids. The influence of experimental conditions such as mobile phase composition, column length and temperature upon the amino acid separations was evaluated. However, temperature, addition of water-miscible organic solvent to the mobile phase and mobile phase gradients were not effective at improving the separations. In contrast, the use of an unbuffered pure water mobile phase proved successful for the separation and detection of amino acids. The observed order of elution seems to parallel the isoelectric points of the respective amino acids and suggests that this diethylammonio-propylsulfate stationary phase column functions as an amphoteric ion-exchanger. Under optimized chromatographic conditions, the detection limits for the amino acids were in range of 0.07-0.44 pmol (pico-mole). The method was also successfully applied for the analysis of an actual commercial sample.


Assuntos
Aminoácidos/análise , Cromatografia Líquida de Alta Pressão/métodos , Espectrometria de Massas/métodos , Compostos de Enxofre/química , Aminoácidos/isolamento & purificação , Íons , Solventes/química , Água/química
5.
J Chromatogr A ; 1621: 461045, 2020 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-32201036

RESUMO

New zwitterionic (ZIC) stationary phases (SPs) are synthesized with the click and conventional bonding of tyrosine to silica gel. Infrared spectra and elemental analysis demonstrate the successful click and conventional bonding of this ZIC group on silica particles by the surface coverage including 2.36 and 0.75 µm m-2, respectively. Given the above-mentioned explanation, the present study evaluated the retention mechanism and chromatographic manners of polar compounds on these new materials under hydrophilic interaction liquid chromatography (HILIC) conditions. Based on the results, the Click-Tyrosine Stationary Phase provided good HILIC characteristics when it was applied to separate phenolic compounds, amino acids, alkaloids, and nucleobases compared to bare silica gel SP and even conventional tyrosine SPs. Further, this new Click-Tyrosine-SP represented appropriate HILIC features and column efficiency (the theoretical plate number was up to 50,000 plates m-1 for thebaine). Furthermore, the study investigated the effect of solute polarity (the number of the hydroxyl group of phenolic compounds) and hydrophobicity (the number of the side chain of aliphatic amino acids) on retention behaviors. Finally, some important factors were studied as the potential variables for guiding the retention behavior of the polar compound in HILIC condition including solvent composition, salt concentration, and the buffer pH of the mobile phase.


Assuntos
Cromatografia Líquida/métodos , Tirosina/química , Alcaloides/análise , Alcaloides/isolamento & purificação , Aminoácidos/análise , Aminoácidos/isolamento & purificação , Química Click , Interações Hidrofóbicas e Hidrofílicas , Fenóis/análise , Fenóis/isolamento & purificação , Sílica Gel/química
6.
J Chromatogr A ; 1619: 460937, 2020 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-32063276

RESUMO

An ethylenediamine dicarboxyethyl diacetamido-bridged bis(ß-cyclodextrin) was firstly synthesized through the reaction of 6-deoxy-6-amino-ß-cyclodextrin (NH2-CD) with ethylenediaminetetraacetic dianhydride. Then it was bonded onto the surface of silica gel SBA-15 to obtain an ethylenediamine dicarboxyethyl diacetamido-bridged bis(ß-CD)-bonded chiral stationary phase (EBCDP). The structures of the bridged bis(ß-CD) and EBCDP were characterized by infrared spectroscopy, mass spectrometry, elemental analysis and thermogravimetric analysis, accordingly. The chiral chromatographic performances of EBCDP were systematically evaluated by separating 28 racemic analytes in the reversed-phase or polar organic mode, including eight flavanones, eight bolckers, five dansyl-amino acids, three DL-amino acids and four other common drugs. As a result, the relatively high enantioselectivity of EBCDP was observed in comparison with a native ß-CD-CSP (CDSP). All selected analytes were separated on EBCDP, of which 20 analytes had resolutions up to baseline, 2'-hydroxyflavanone and arotinolol had resolutions up to 4.35 and 2.05 in about 30 min, respectively, whereas CDSP only separated 11 analytes with low resolutions (0.55~1.69). Moreover, EBCDP was able to utilize the complexation of the bridging linker (ethylenediamine dicarboxyethyl diamide group, EDTA-based) to realize direct separations of DL-amino acids with a mobile phase containing copper ion (Cu2+), which was similar to the chiral ligand exchange chromatography. Unlike the native cyclodextrin with small cavity (~242 Å3), the bridged bis(ß-CD) combined two ß-CD units with a bridging linker, having a well-organized "pseudo-cavity" as an organic whole to encapsulate more analytes, which made EBCDP have broad-spectrum applications in chiral separations.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Cromatografia Líquida de Alta Pressão/normas , beta-Ciclodextrinas/química , Aminoácidos/isolamento & purificação , Flavanonas/química , Dióxido de Silício/química , Estereoisomerismo , beta-Ciclodextrinas/síntese química , beta-Ciclodextrinas/normas
7.
Mar Drugs ; 18(2)2020 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-31991623

RESUMO

UV-absorbing compounds, such as mycosporine-like amino acids (MAAs), are a group of secondary metabolites present in many marine species, including red seaweeds. In these organisms, the content and proportion of the composition of MAAs vary, depending on the species and several environmental factors. Its high cosmetic interest calls for research on the content and composition of MAAs, as well as the dynamics of MAAs accumulation in seaweeds from different latitudes. Therefore, this study aimed to survey the content of UV-absorbing MAAs in three Subantarctic red seaweeds during a seasonal cycle. Using spectrophotometric and HPLC techniques, the content and composition of MAAs of intertidal Iridaea tuberculosa, Nothogenia fastigiate, and Corallina officinalis were assessed. Some samples were also analyzed using high-resolution mass spectrometry coupled with HPLC-ESI-MS in order to identify more precisely the MAA composition. I. tuberculosa exhibited the highest MAA values (above 1 mg g-1 of dried mass weight), while C. officinalis showed values not exceeding 0.4 mg g-1. Porphyra-334 was the main component in N. fastigiata, whereas I. tuberculosa and C. officinalis exhibited a high content of palythine. Both content and composition of MAAs varied seasonally, with high concentration recorded in different seasons, depending on the species, i.e., winter (I. tuberculosa), spring (N. fastigiata), and summer (C. officinalis). HPLC-ESI-MS allowed us to identify seven different MAAs. Two were recorded for the first time in seaweeds from Subantarctic areas (mycosporine-glutamic acid and palythine-serine), and we also recorded an eighth UV-absorbing compound which remains unidentified.


Assuntos
Aminoácidos/isolamento & purificação , Rodófitas/química , Alga Marinha/química , Protetores Solares/isolamento & purificação , Aminoácidos/metabolismo , Aminoácidos/efeitos da radiação , Cromatografia Líquida de Alta Pressão , Espectrometria de Massas , Rodófitas/metabolismo , Estações do Ano , Alga Marinha/metabolismo , Metabolismo Secundário/efeitos da radiação , Protetores Solares/metabolismo , Raios Ultravioleta/efeitos adversos
8.
Molecules ; 25(1)2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31906356

RESUMO

The crude drug ysypó hû (Adenocalymma marginatum DC., Bignoniaceae) is used traditionally by the Guarani of Eastern Paraguayan as a male sexual enhancer. The aim of the present study was to identify the main constituents of the crude drug and to evaluate the in vitro inhibitory activity towards the enzyme phosphodiesterase-5 (PDE-5). The main compounds were isolated by counter-current chromatography (CCC). The metabolites were identified by spectroscopic and spectrometric means. The chemical profiling of the extracts was assessed by high-performance liquid chromatography coupled to mass spectrometry (HPLC-MS/MS). The crude extract and main isolated compounds were tested for their PDE-5 inhibitory activity using commercial kits. The iridoid theviridoside and 4-hydroxy-1-methylproline were isolated as the main constituent of the crude drug. Four chlortheviridoside hexoside derivatives were detected for the first time as natural products. Chemical profiling by HPLC-MS/MS led to the tentative identification of nine iridoids, six phenolics, and five amino acids. The crude extracts and main compounds were inactive towards PDE-5 at concentrations up to 500 µg/mL. Iridoids and amino acid derivatives were the main compounds occurring in the Paraguayan crude drug. The potential of ysypó hû as a male sexual enhancer cannot be discarded, since other mechanisms may be involved.


Assuntos
Bignoniaceae/química , Iridoides/química , Inibidores da Fosfodiesterase 5/química , Extratos Vegetais/química , Aminoácidos/análise , Aminoácidos/química , Aminoácidos/isolamento & purificação , Bignoniaceae/metabolismo , Cromatografia Líquida de Alta Pressão , Misturas Complexas , Distribuição Contracorrente , Glicosídeos Iridoides , Iridoides/análise , Iridoides/isolamento & purificação , Paraguai , Fenóis/análise , Fenóis/química , Fenóis/isolamento & purificação , Inibidores da Fosfodiesterase 5/metabolismo , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/metabolismo , Espectrometria de Massas em Tandem
9.
J Chromatogr A ; 1609: 460446, 2020 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-31420178

RESUMO

Two new copolymer-grafted silica stationary phases were prepared and employed in hydrophilic interaction chromatography (HILIC). 2-(Dimethylamino)ethyl methacrylate (DMAEMA) are copolymerized with itaconic acid (IA) and acrylic acid (AA) respectively, via thiol-ene click reaction on silica surface with deep eutectic solvents (DES) as new solvents. The obtained poly(DMAEMA-co-itaconic acid)-grafted silica (Sil-PDM-PIA) and poly(DMAEMA-co-acrylic acid)-grafted silica (Sil-PDM-PAA) were characterized by Fourier transform infrared spectroscopy, elemental analysis and solid-state 13C NMR spectra. Their hydrophilic interaction performances were evaluated by separating nucleosides, nucleobases, saccharides, and amino acids. Compared with previous reported poly(itaconic acid)-grafted silica (Sil-PIA) and poly(acrylic acid)-grafted silica (Sil-PAA) stationary phases, these two new copolymer-grafted silica performed higher selectivity and better separation for polar analytes in HILIC.


Assuntos
Cromatografia/métodos , Química Click/métodos , Interações Hidrofóbicas e Hidrofílicas , Polímeros/química , Dióxido de Silício/química , Solventes/química , Compostos de Sulfidrila/química , Acetonitrilos/química , Aminoácidos/isolamento & purificação , Entropia , Concentração de Íons de Hidrogênio , Espectroscopia de Ressonância Magnética , Metacrilatos/química , Nucleosídeos/isolamento & purificação , Nylons/química , Reprodutibilidade dos Testes , Sais/química , Espectroscopia de Infravermelho com Transformada de Fourier , Succinatos/química , Temperatura
10.
J Chromatogr A ; 1614: 460729, 2020 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-31785894

RESUMO

Most HPLC enantiomer separations are performed with columns packed with a chiral stationary phase (CSP) operated with an achiral mobile phase. The intrinsically limited chemical selectivity of most CSPs to the simultaneous resolution of several pairs of enantiomers means that complex mixtures of diverse pairs of enantiomers cannot be resolved in a single run due to peak overlapping. Moreover, some drawbacks remain when the analyte is present in very complex samples containing other achiral compounds which can co-elute with the enantiomer peaks. Multidimensional chromatography becomes an option to increase peak capacity and resolve these samples. The aim of this work was to study an online fully comprehensive 2D-LC mode utilizing a combination of a chiral column in the first dimension and an achiral column in the second dimension. The 2D-LC system was built with an active flow splitter pump in order to easily adjust the volume of sample transferred into the second dimension and to independently optimize the flow rate in the first dimension. The present LCxLC method was optimized for the separation of amino acids present in honey samples, taking into account key parameters that influence the bidimensional peak capacity (orthogonality, sampling frequency, etc.). The amino acids have been preconcentrated on a cation-exchange column followed by derivatization. Several amino acids present in different honey samples have been identified and the data generated has been analyzed by principal component analysis.


Assuntos
Aminoácidos/isolamento & purificação , Cromatografia Líquida , Análise de Alimentos/métodos , Mel , Aminoácidos/química , Mel/análise , Análise de Componente Principal , Estereoisomerismo
11.
Nat Commun ; 10(1): 5813, 2019 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-31862950

RESUMO

Gut microbiota has been implicated in major diseases affecting the human population and has also been linked to triglycerides and high-density lipoprotein levels in the circulation. Recent development in metabolomics allows classifying the lipoprotein particles into more details. Here, we examine the impact of gut microbiota on circulating metabolites measured by Nuclear Magnetic Resonance technology in 2309 individuals from the Rotterdam Study and the LifeLines-DEEP cohort. We assess the relationship between gut microbiota and metabolites by linear regression analysis while adjusting for age, sex, body-mass index, technical covariates, medication use, and multiple testing. We report an association of 32 microbial families and genera with very-low-density and high-density subfractions, serum lipid measures, glycolysis-related metabolites, ketone bodies, amino acids, and acute-phase reaction markers. These observations provide insights into the role of microbiota in host metabolism and support the potential of gut microbiota as a target for therapeutic and preventive interventions.


Assuntos
Microbioma Gastrointestinal/fisiologia , Interações entre Hospedeiro e Microrganismos/fisiologia , Metaboloma/fisiologia , Proteínas da Fase Aguda/isolamento & purificação , Proteínas da Fase Aguda/metabolismo , Adulto , Aminoácidos/sangue , Aminoácidos/isolamento & purificação , Aminoácidos/metabolismo , Bactérias/genética , Bactérias/isolamento & purificação , Estudos de Coortes , DNA Bacteriano/isolamento & purificação , Fezes/microbiologia , Feminino , Glicólise/fisiologia , Humanos , Lipoproteínas HDL/sangue , Lipoproteínas HDL/isolamento & purificação , Lipoproteínas HDL/metabolismo , Masculino , Metabolômica/métodos , Pessoa de Meia-Idade , Países Baixos , Espectroscopia de Prótons por Ressonância Magnética , RNA Ribossômico 16S/genética , Análise de Regressão , Triglicerídeos/sangue , Triglicerídeos/isolamento & purificação , Triglicerídeos/metabolismo
12.
J Photochem Photobiol B ; 201: 111684, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31733505

RESUMO

Since the beginning of life on Earth, cyanobacteria have been exposed to natural ultraviolet-A radiation (UV-A, 315-400 nm) and ultraviolet-B radiation (UV-B, 280-315 nm), affecting their cells' biomolecules. These photoautotrophic organisms have needed to evolve to survive and thus, have developed different mechanisms against ultraviolet radiation. These mechanisms include UVR avoidance, DNA repair, and cell protection by producing photoprotective compounds like Scytonemin, carotenoids, and Mycosporine-like amino acids (MAAs). Lyngbya marine species are commercially important due to their secondary metabolites that show a range of biological activities including antibacterial, insecticidal, anticancer, antifungal, and enzyme inhibitor. The main topic in this review covers the Lyngbya sp., a cyanobacteria genus that presents photoprotection provided by the UV-absorbing/screening compounds such as MAAs and Scytonemin. These compounds have considerable potentialities to be used in the cosmeceutical, pharmaceutical, biotechnological and biomedical sectors and other related manufacturing industries with an additional value of environment friendly in nature. Scytonemin has UV protectant, anti-inflammatory, anti-proliferative, and antioxidant activity. MAAs act as sunscreens, provide additional protection as antioxidants, can be used as UV protectors, activators of cell proliferation, skin-care products, and even as photo-stabilizing additives in paints, plastics, and varnishes. The five MAAs identified so far in Lyngbya sp. are Asterina-330, M-312, Palythine, Porphyra-334, and Shinorine are capable of dissipating absorbed radiation as harmless heat without producing reactive oxygen species.


Assuntos
Aminoácidos/química , Cianobactérias/metabolismo , Cicloexanóis/química , Indóis/química , Fenóis/química , Protetores Solares/química , Raios Ultravioleta , Aminoácidos/isolamento & purificação , Antioxidantes/química , Cicloexanóis/isolamento & purificação , Indóis/isolamento & purificação , Fenóis/isolamento & purificação , Protetores Solares/metabolismo
13.
Mikrochim Acta ; 186(9): 636, 2019 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-31432257

RESUMO

The preparation of an organic polymer monolithic column modified with an amino acid ionic liquid and graphene oxide (AAIL-GO) and its application to capillary electrochromatography (CEC) was described. The AAIL tetramethylammonium-L-arginine was bonded to a monolithic column that was previously modified with graphene oxide by using an hydrochloride/N-hydroxysuccinimide coupling reaction. The morphology of a poly(glycidyl methacrylate-co-ethylene dimethacrylate) monolith was examined by scanning electron microscopy. The incorporation of AAIL and graphene oxide was detected by infrared spectroscopy and elemental analysis. The resulting monolithic column produced a strong and stable electroosmotic flow from the anode to the cathode in the pH range from 3 to 9. Compared with a column modified with AAIL or graphene oxide only, the AAIL-GO-modified column has a better separation ability for amino acids, ß-blockers, and nucleotides (the resolution of three amino acids: 2.231 and 2.036, ß-blockers: 2.779 and 2.470 and nucleotides: 8.345 and 3.321). Molecular modeling was applied to demonstrate the separation mechanism of small molecules which showed a good support for experimental results. Graphical abstract Schematic representation of capillary electrochromatography (CEC) systems with an amino acid ionic liquid-graphene oxide modified organic polymer monolithic column as stationary phases for separation of amino acids, ß-blockers, and nucleotides.


Assuntos
Antagonistas Adrenérgicos beta/isolamento & purificação , Aminoácidos/química , Eletrocromatografia Capilar , Grafite/química , Líquidos Iônicos/química , Nucleotídeos/isolamento & purificação , Polímeros/química , Antagonistas Adrenérgicos beta/química , Aminoácidos/isolamento & purificação , Estrutura Molecular , Nucleotídeos/química , Tamanho da Partícula , Propriedades de Superfície
14.
Ultrason Sonochem ; 58: 104603, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31450354

RESUMO

As part of our efforts to develop a new method for chiral resolution of amino acids with sonochemically modified proteins, we present result that indicates how ovalbumin microspheres (OAMS) interact specifically with l-amino acids from a racemate in solution, leaving an excess of d-enantiomer in the permeate solution. Among different amino acids that interacted with the OAMS, tryptophan (Trp) was the most successfully resolved with 65% enantiomeric excess. A control experiment with native ovalbumin in solution did not show any chiral resolution of amino acids. Interestingly, when the OAMS were pretreated with racemic lysine (Lys) solution and then used for resolution of tryptophan the enantiomeric enrichment of d-tryptophan was raised to 98%. This unanticipated positive effect is discussed in terms of the structural correlation between Trp and Lys, which is less apparent in other amino acids such as phenylalanine.


Assuntos
Aminoácidos/química , Ovalbumina/química , Sonicação , Aminoácidos/isolamento & purificação , Animais , Modelos Moleculares , Conformação Molecular , Estereoisomerismo
15.
Sensors (Basel) ; 19(17)2019 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-31443493

RESUMO

Quantum dots (QDs) are very attractive nanomaterials for analytical chemistry, due to high photostability, large surface area featuring numerous ways of bioconjugation with biomolecules, usually high quantum yield and long decay times. Their broad absorption spectra and narrow, sharp emission spectra of size-tunable fluorescence make them ideal tools for pattern-based sensing. However, almost always they are applied for specific sensing with zero-dimensional (0D) signal reporting (only peak heights or peak shifts are considered), without taking advantage of greater amount of information hidden in 1D signal (emission spectra), or huge amount of information hidden in 2D fluorescence maps (Excitation-Emission Matrixes, EEMs). Therefore, in this work we propose opposite strategy-non-specific interactions of QDs, which are usually avoided and regarded as their disadvantage, were exploited here for 2D fluorescence fingerprinting. Analyte-specific multivariate fluorescence response of QDs is decoded with the use of Partial Least Squares-Discriminant Analysis. Even though only one type of QDs is studied, the proposed pattern-based method enables to obtain satisfactory accuracy for all studied compounds-various neurotransmitters, amino-acids and oligopeptides. This is a proof of principle of the possibility of the identification of various bioanalytes by such fluorescence fingerprinting with the use of QDs.


Assuntos
Aminoácidos/isolamento & purificação , Técnicas Biossensoriais/métodos , Neurotransmissores/isolamento & purificação , Oligopeptídeos/isolamento & purificação , Aminoácidos/química , Neurotransmissores/química , Oligopeptídeos/química , Imagem Óptica , Pontos Quânticos/química
16.
Methods Mol Biol ; 2030: 315-326, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31347128

RESUMO

Whole blood and/or plasma amino acids are useful for monitoring whole-body protein and amino acid metabolism in an organism under various physiological and pathophysiological conditions. Various methodological procedures are in use for their measurement in biological fluids. From the time when capillary electrophoresis was introduced as a technology offering rapid separation of various ionic and/or ionizable compounds with low sample and solvent consumption, there were many attempts to use it for the measurement of amino acids present in physiological fluids. As a rule, these methods require derivatization procedures for detection purposes.Here, we present two protocols for the analysis of free amino acids employing free zone capillary electrophoresis. Main advantage of both methods is an absence of any derivatization procedures that permits the analysis of free amino acid in physiological fluids. The method using direct detection and carrier electrolyte consisting of disodium monophosphate (10 mM at pH 2.90) permits determination of compounds that absorb in UV region (aromatic and sulfur containing amino acids, as well as some peptides such as carnosine, reduced, and oxidized glutathione). The other method use indirect absorbance detection, employing 8 mM p-amino salicylic acid buffered with sodium carbonate at pH 10.2 as running electrolyte. It permits quantification of 30 underivatized physiological amino acids and peptides. In our experience factorial design represents a useful tool for final optimization of the electrophoretic conditions if it is necessary.


Assuntos
Aminoácidos/isolamento & purificação , Eletroforese Capilar/métodos , Peptídeos/isolamento & purificação , Plasma/química , Aminoácidos/química , Eletrólitos/química , Eletroforese Capilar/instrumentação , Estudos de Viabilidade , Humanos , Peptídeos/química , Fosfatos/química , Espectrofotometria Ultravioleta , Raios Ultravioleta
17.
Methods Mol Biol ; 2030: 351-364, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31347130

RESUMO

This chapter describes improvements in a sequential injection method to automate the fluorimetric determination of amino acids by pre-column derivatization with o-phthaldialdehyde in presence of 2-mercaptoethanol. Separation is achieved by reversed-phase liquid chromatography in a 50 × 4.6 mm C18 silica-based monolithic column. The method is low-priced, and the separation is performed by stepwise gradient elution using six mobile phases. The mobile phase used for the first elution step is composed of methanol/tetrahydrofuran/10 mM phosphate buffer (pH 7.2) at volumetric ratio 8:1:91. Additional elution steps use mobile phases containing methanol and 10 mM phosphate buffer at volumetric ratios of 17.5:82.5, 25:75, 35:65, 50:50, and 65:35. Nineteen chromatographic peaks are observed in a mixture of twenty amino acids. The only complete co-elution is between tryptophan and methionine. The entire cycle of amino acid derivatization, chromatographic separation, and column conditioning at the end of separation lasts for 30 min. The method is successfully applied to quantify the major intracellular dissolved free amino acids in the marine microalgae Tetraselmis gracilis, Phaeodactium tricornutum, and Synechococcus elongatus.


Assuntos
Aminoácidos/isolamento & purificação , Cromatografia de Fase Reversa/métodos , Fluorometria/métodos , Microalgas/química , Aminoácidos/química , Tampões (Química) , Cromatografia de Fase Reversa/instrumentação , Fluorometria/instrumentação , Furanos/química , Metanol/química , Synechococcus/química , o-Ftalaldeído/química
18.
Methods Mol Biol ; 2030: 429-438, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31347136

RESUMO

The amino acid profile obtained from a fingerprint may provide valuable information on its donor. Unfortunately, the collection of chemicals from the fingerprint is often destructive to the fingerprint ridge detail. Herein we detail the use of cross-linkable solutions of dextran-methacrylate to form hydrogels capable of collecting amino acids from surfaces followed by extraction and quantification with UPLC-MS. This method allows for the amino acid profile analysis of fingerprints while allowing for their increased visualization at a later stage using the standard method of cyanoacrylate fuming followed by basic-yellow dyeing.


Assuntos
Aminoácidos/isolamento & purificação , Reagentes para Ligações Cruzadas/química , Dermatoglifia , Espectrometria de Massas/métodos , Adsorção , Aminoácidos/química , Cromatografia Líquida de Alta Pressão/instrumentação , Cromatografia Líquida de Alta Pressão/métodos , Dextranos/química , Estudos de Viabilidade , Humanos , Hidrogéis/química , Espectrometria de Massas/instrumentação , Metacrilatos/química
19.
Methods Mol Biol ; 2030: 439-450, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31347137

RESUMO

UPLC-MS is a commonly used technique to first separate complex samples and subsequently quantify molecules of interest. Herein we describe the use of UPLC-MS using an amide stationary phase to quantify non-derivatized amino acids extracted from fingerprints. As detector either a triple-quadrupole MS/MS or a TOF-MS detector was used. This method allows for a simple and fast sample preparation, which facilitates the analysis of large amounts of samples.


Assuntos
Aminoácidos/isolamento & purificação , Reagentes para Ligações Cruzadas/química , Dermatoglifia , Espectrometria de Massas em Tandem/métodos , Adsorção , Amidas/química , Aminoácidos/química , Cromatografia Líquida de Alta Pressão/instrumentação , Cromatografia Líquida de Alta Pressão/métodos , Dextranos/química , Estudos de Viabilidade , Metacrilatos/química , Espectrometria de Massas em Tandem/instrumentação
20.
Blood Purif ; 48(4): 321-329, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31291614

RESUMO

BACKGROUND/OBJECTIVES: During continuous venovenous hemofiltration (CVVH), there is unwanted loss of amino acids (AA) in the ultrafiltrate (UF). Solutes may also be removed by adsorption to the filter membrane. The aim was to quantify the total loss of AA via the CVVH circuit using a high-flux polysulfone membrane and to differentiate between the loss by ultrafiltration and adsorption. METHODS: Prospective observational study in ten critically ill patients, receiving predilution CVVH with a new filter, blood flow 180 mL/min, and predilution flow 2,400 mL/h. Arterial blood, postfilter blood, and UF samples were taken at baseline, and 1, 8, and 24-h after CVVH initiation, to determine AA concentrations and hematocrit. Mass transfer calculations were used to determine AA loss in the filter and by UF, and the difference between these 2. RESULTS: The median AA loss in the filter was 10.4 g/day, the median AA loss by UF was 13.4 g/day, and the median difference was -2.9 g/day (IQR -5.9 to -1.4 g/day). For the individual AA, the difference ranged from -1 g/day to +0.4 g/day, suggesting that some AA were consumed or adsorbed and others were generated. AA losses did not significantly change over the 24-h study period. CONCLUSION: During CVVH with a modern polysulfone membrane, the estimated AA loss was 13.4 g/day, which corresponds to a loss of about 11.2 g of protein per day. Adsorption did not play a major role. However, individual AA behaved differently, suggesting complex interactions and processes at the filter membrane or peripheral AA production.


Assuntos
Aminoácidos/isolamento & purificação , Terapia de Substituição Renal Contínua/métodos , Estado Terminal/terapia , Adsorção , Adulto , Idoso , Aminoácidos/sangue , Terapia de Substituição Renal Contínua/instrumentação , Feminino , Humanos , Masculino , Membranas Artificiais , Pessoa de Meia-Idade , Polímeros/química , Estudos Prospectivos , Sulfonas/química , Adulto Jovem
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