Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 1.212
Filtrar
1.
Arch Insect Biochem Physiol ; 102(4): e21611, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31471923

RESUMO

Seventeen species of the coleopteran series Cucujiformia are investigated for the presence and sequence of putative adipokinetic hormones (AKHs). Cucujiformia includes species from the major superfamilies, that is, Chrysomeloidea, Curculionoidea, Cucujoidea, and Tenebrionoidea. The clade Phytophaga in which the Chrysomeloidea and Curculionoidea reside, harbor very detrimental species for agriculture and forestry. Thus, this study aims not only to demonstrate the structural biodiversity of AKHs in these beetle species and possible evolutionary trends but also to determine whether the AKHs from harmful pest species can be used as lead substances for a future putative insecticide that is harmless to beneficial insects. Sequence analysis of AKHs is achieved by liquid chromatography coupled to mass spectrometry. Most of the investigated species contain AKH octapeptides in their corpora cardiaca, although previously published work also found a few decapeptides, which we comment on. The signature and sole AKH in cerambycidae Chrysomeloidea and Curculionoidea is Peram-CAH-I (pEVNFSPNW amide), which is also found in the majority of chrysomelidae Chrysomeloidea and in the one investigated species of Cucujoidea albeit in a few cases associated with a second AKH which can be either Peram-CAH-II (pELTFTPNW amide), Emppe-AKH (pEVNFTPNW amide), or Micvi-CC (pEINFTPNW amide). The most often encountered AKH in Tenebrionoidea, family Meloidae as well as family Tenebrionidae, is Tenmo-HrTH (pELNFSPNW amide) followed by Pyrap-AKH (pELNFTPNW amide) and a Tenmo-HrTH extended decapeptide (in Meloidae). Finally, we examine AKH sequences from 43 species of cucujiform beetles, including the superfamily Coccinelloidea for a possible lead compound for producing a cucujiform-specific pesticide.


Assuntos
Besouros/química , Hormônios de Inseto/química , Oligopeptídeos/química , Ácido Pirrolidonocarboxílico/análogos & derivados , Sequência de Aminoácidos , Animais , Cromatografia Líquida , Corpora Allata/química , Hormônios de Inseto/análise , Espectrometria de Massas , Oligopeptídeos/análise , Peptídeos/análise , Peptídeos/química , Ácido Pirrolidonocarboxílico/análise , Ácido Pirrolidonocarboxílico/química
2.
Anal Bioanal Chem ; 411(15): 3395-3404, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31011782

RESUMO

Short peptides are important compounds in a variety of fields, including food and nutraceutical applications, but also biomarker discovery, bioactive peptide discovery and peptide drug separation. Despite the importance of short peptides, they are currently less studied than other peptides because of the lack of dedicated methods for their characterization. The method described in this paper comprises a combination of strategies to tackle the main limitations in short peptide analysis. In particular, in this work an untargeted peptidomic approach based on ultrahigh-performance liquid chromatography coupled to high-resolution mass spectrometry was developed for the identification of short peptides in cow milk samples. After milk defatting and precipitation, the sample was purified by cotton-hydrophilic interaction liquid chromatography (HILIC) micro tip in order to avoid suppression phenomena due to contaminants present in milk, such as carbohydrates. The sample was then separated by means of two chromatographic columns, with a complementary selectivity mechanism, namely reversed-phase C18 column and porous graphitic carbon (PGC). By this approach, the method allowed the separation and characterization of di-, tri- and tetrapeptides. A total of 57 and 41 peptides were identified by using a C18 and a PGC column, respectively; in particular, 31 were exclusively identified by using the C18 column, 15 unique peptides were identified by using the PGC column, while 26 were in common between the two data sets, demonstrating that the two columns have a different selectivity mechanism. The results indicated that an integrated approach may be appropriate to improve the separation of different peptides and increase the number of identifications because of the wide range of polarity of short peptides. The method allowed the untargeted identification of short peptides in milk, a complex matrix chosen as a representative real sample for method application, and provides complementary information to that accessible by ordinary peptidomics. Graphical abstract.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Leite/química , Oligopeptídeos/análise , Espectrometria de Massas em Tandem/métodos , Sequência de Aminoácidos , Animais , Carbono/química , Bovinos , Feminino , Grafite/química , Porosidade
3.
Naturwissenschaften ; 106(5-6): 14, 2019 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-30963267

RESUMO

Livestock can die from grazing in areas where larvae of certain Argidae or Pergidae species containing toxic peptides occur in mass. However, it remains unknown whether other stages also contain these compounds. Here, single specimens of larvae, prepupae, and adults of Arge berberidis, plus samples of its cocoons and larval feces, were analyzed by liquid chromatography-tandem mass spectrometry. The four peptides, pergidin (Perg), 4-valinepergidin (VPerg), dephosphorylated pergidin (dpPerg), and lophyrotomin (LGln), were detected in each of the three stages. Peptide concentrations, in percentage fresh weight, increased from larval up to adult stages, with mean values from 0.044 to 0.125% for Perg, 0.008 to 0.023% for VPerg, and 0.064 to 0.116% for LGln, whereas dpPerg never exceeded 0.001%. The concentrations of this latter peptide averaged 0.002% in the cocoon built by the prepupa, and nearly no peptides were detected in larval feces. Moreover, the concentrations of the three main peptides (Perg, LGln, and VPerg) tended to be correlated with each other in larvae and especially in adults. It is likely that peptide production, purportedly by an endosymbiont, stops at prepupal stage and that concentration of the peptides increases from prepupa to adult due to a decrease of body weight.


Assuntos
Himenópteros/química , Peptídeos/análise , Toxinas Biológicas/análise , Animais , Cromatografia Líquida , Fezes/química , Larva/química , Oligopeptídeos/análise , Pupa/química , Espectrometria de Massas em Tandem
4.
Cell Prolif ; 52(2): e12556, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30710394

RESUMO

OBJECTIVES: Nanocarriers can greatly enhance the cellular uptake of therapeutic agents to regulate cell proliferation and metabolism. Nevertheless, further application of nanocarriers is often limited by insufficient understanding of the mechanisms of their uptake and intracellular behaviour. MATERIALS AND METHODS: Fluorescent polymer dots (Pdots) are coated with synthetic octaarginine peptides (R8) and are analysed for cellular uptake and intracellular transportation in HeLa cervical cancer cells via single particle tracking. RESULTS: Surface modification with the R8 peptide efficiently improves both cellular uptake and endosomal escape of Pdots. With single particle tracking, we capture the dynamic process of internalization and intracellular trafficking of R8-Pdots, providing new insights into the mechanism of R8 in facilitating nanostructure-based cellular delivery. Furthermore, our results reveal R8-Pdots as a novel type of autophagy inducer. CONCLUSIONS: This study provides new insights into R8-mediated cellular uptake and endosomal escape of nanocarriers. It potentiates biological applications of Pdots in targeted cell imaging, drug delivery and gene regulation.


Assuntos
Endocitose , Corantes Fluorescentes/metabolismo , Oligopeptídeos/metabolismo , Polímeros/metabolismo , Transporte Biológico , Endossomos/metabolismo , Corantes Fluorescentes/análise , Células HeLa , Humanos , Microscopia Confocal , Nanoestruturas/análise , Oligopeptídeos/análise , Imagem Óptica , Polímeros/análise
5.
Biosci Biotechnol Biochem ; 83(2): 357-364, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30295131

RESUMO

Three new peptides: (pGlu)L-ethyl, (pGlu)LFGP-ethyl and (pGlu)LFNP-ethyl, were identified in the search for pyroglutamyl oligopeptide ethyl esters in sake. The ethyl esterified peptides in sake were quantitated using stable isotope dilution analysis and additional quantitation of (pGlu)L was performed using an external standard method. The concentrations of (pGlu)L-ethyl and (pGlu)L in 33 commercial sake samples ranged from 0.16 to 1.57 mg/L and 1.49 to 7.55 mg/L, respectively. The sensory properties of the pyroglutamyl oligopeptide ethyl esters and corresponding non-esterified peptides were examined: the estimated difference threshold of (pGlu)L (2.0 mg/L) and (pGlu)L-ethyl (0.267 mg/L) was exceeded in 32 and 26 samples, respectively. Estimated thresholds of (pGlu)LFGP-ethyl and (pGlu)LFNP-ethyl were often lower than the levels in quantitated sake samples. The sensory effects of these pyroglutamyl dipeptides on a model sake quality may be negative because of their unpleasant taste, however, (pGlu)LFNP-ethyl may be positive because of its mild taste.


Assuntos
Bebidas Alcoólicas/análise , Oligopeptídeos/análise , Ácido Pirrolidonocarboxílico/química , Cromatografia Líquida de Alta Pressão , Limite de Detecção , Ácido Pirrolidonocarboxílico/normas , Padrões de Referência , Espectrometria de Massas por Ionização por Electrospray
6.
Drug Test Anal ; 11(1): 162-167, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30051972

RESUMO

Growth hormone releasing peptides (GHRPs) are synthetic peptides with the ability to stimulate human growth hormone (hGH) secretion. Several GHRPs have been developed as drug candidates; however, only one of them, GHRP-2 (Pralmorelin), has received a clinical approval. Nevertheless, they are distributed on the black market and misused by cheating athletes, due to their performance-enhancing effects. Hence, GHRPs have been included in the World-Anti-Doping-Agency's Prohibited List as forbidden substances in sport. Predominantly, analytical methods for detection and unequivocal identification of doping substances are based on mass spectrometry. Therefore, in the present work, a qualitative analysis by liquid chromatography coupled to high-resolution tandem mass spectrometry with a quadrupole time-of-flight analyzer was performed to identify a new heptapeptide (MW = 874.02 Da) - a glycine analogue of GHRP-2. Structure determination using de novo sequencing is described here in detail. The results of this study may indicate a new approach to circumvent a detection of doping practices.


Assuntos
Emigração e Imigração/legislação & jurisprudência , Oligopeptídeos/análise , Substâncias para Melhoria do Desempenho/análise , Detecção do Abuso de Substâncias/métodos , Doping nos Esportes/legislação & jurisprudência , Doping nos Esportes/prevenção & controle , Humanos , Espectrometria de Massas por Ionização por Electrospray/métodos , Espectrometria de Massas por Ionização por Electrospray/normas , Detecção do Abuso de Substâncias/normas , Espectrometria de Massas em Tandem/métodos , Espectrometria de Massas em Tandem/normas
7.
Drug Test Anal ; 11(2): 350-354, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30136411

RESUMO

A number of unknown pharmaceutical preparations seized by Danish customs authorities were submitted for liquid chromatography-high resolution mass spectrometry (LC-HRMS) analysis. Comparison with reference standards unequivocally identified the content of the powders as analogs of the growth hormone secretagogues GHRP-2 (Pralmorelin), GHRP-6, Ipamorelin, and modified growth hormone releasing factor (modified GRF 1-29), which can be used as performance-enhancing substances in sports. In all cases, the detected modification involved the addition of an extra glycine amino acid at the N-terminus, and analytical methods targeting growth hormone secretagogues should hence be updated accordingly.


Assuntos
Doping nos Esportes , Glicina/química , Hormônio Liberador de Hormônio do Crescimento/análise , Oligopeptídeos/análise , Fragmentos de Peptídeos/análise , Substâncias para Melhoria do Desempenho/análise , Secretagogos/análise , Detecção do Abuso de Substâncias/métodos , Hormônio Liberador de Hormônio do Crescimento/química , Oligopeptídeos/química , Fragmentos de Peptídeos/química , Pós , Padrões de Referência , Secretagogos/química
8.
Arch Oral Biol ; 99: 15-21, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30590229

RESUMO

OBJECTIVES: Opiorphin is a recently discovered peptide shown to inhibit the enkephalin-degrading enzymes and prolong the effects of enkephalins. Although opiorphin is found in high concentrations in saliva, the relationship between salivary opiorphin and orofacial pains is not yet fully understood. We aimed to determine salivary opiorphin concentrations in dental pain related to symptomatic irreversible pulpitis (SIP), and symptomatic apical periodontitis (SAP). DESIGN: 39 patients participated in this study. The participants were categorized into SIP and SAP based on their diagnosis. All the patients were treated with root canal treatment. Saliva specimens were collected, and pain levels were recorded at pre-treatment, 7 days post-treatment and 30 days post-treatment. Saliva opiorphin levels were measured using a commercially available ELISA kit. Pre-treatment and post-treatment opiorphin levels were evaluated using repeated measures ANOVA. Correlations between VAS scores, opiorphin levels and age were evaluated using Spearman's Rank Correlation. RESULTS: The average saliva opiorphin level pre-treatment, 7 days post-treatment and 30 days post-treatment were 31.28 ± 7.10 ng/ml, 20.41 ± 2.67 ng/ml and 18.61 ± 2.05 ng/ml respectively. Significantly higher pre-treatment opiorphin levels were observed in the SIP group compared to the SAP group. A strong correlation was observed between the pre-treatment pain levels and the saliva opiorphin concentrations. CONCLUSIONS: Our findings indicate that saliva opiorphin levels increase in inflammation related dental pain. The level of salivary opiorphin is strongly correlated with the reported level of pain. The extent of the inflammation (pulpal vs. periodontal) also affects the opiorphin level.


Assuntos
Biomarcadores/análise , Oligopeptídeos/análise , Saliva/química , Proteínas e Peptídeos Salivares/análise , Odontalgia/induzido quimicamente , Adulto , Análise de Variância , Ensaio de Imunoadsorção Enzimática , Dor Facial/induzido quimicamente , Feminino , Humanos , Inflamação , Masculino , Pessoa de Meia-Idade , Nociceptividade , Medição da Dor , Periodontite Periapical/induzido quimicamente , Periodontite Periapical/diagnóstico , Pulpite/induzido quimicamente , Pulpite/diagnóstico , Dente não Vital , Turquia , Adulto Jovem
9.
Food Chem ; 266: 420-426, 2018 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-30381207

RESUMO

The objective of this study was to investigate the antioxidant activity and the homology of peptides extracted from dry-cured Xuanwei and Jinhua ham. The antioxidant activity of crude peptides was assessed by ORAC and ABTS assays and the scavenging effects on DPPH and O2- free radicals. LC-ESI-Q-TOF-MS/MS was used to analyze the peptide composition. Based on the identified peptides, homologous proteins were matched by the Peaks software. Overall, 243 and 213 peptides were identified with their parent proteins in Xuanwei and Jinhua dry-cured hams. Based on the ORAC and TEAC values, XHP showed higher antioxidant ability than JHP (P < 0.05). After further purification by G-15 chromatography, the results indicate that the oligopeptides with less than 1000 Da had greater antioxidant activity than the other two fractions. Homology-based proteomics showed that the majority of peptides originated from myosin which accounted for 26% in XHP and 32% in JHP respectively.


Assuntos
Antioxidantes/química , Produtos da Carne/análise , Peptídeos/química , Proteômica , Sequência de Aminoácidos , Animais , Antioxidantes/análise , Cromatografia Líquida de Alta Pressão , Peso Molecular , Oligopeptídeos/análise , Oligopeptídeos/química , Oligopeptídeos/isolamento & purificação , Peptídeos/análise , Peptídeos/isolamento & purificação , Suínos , Espectrometria de Massas em Tandem
10.
J Chem Ecol ; 44(12): 1139-1145, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30276758

RESUMO

Determination of the safety of agents prior to release is one of the most important research goals in biological control. In addition to concerns for the safety of non-target plants, determination of the potential toxic properties of new agents needs to be assessed. Numerous phytophagous insects are defended by chemicals against the attack of natural enemies. Some of these defensive compounds could pose an environmental risk if an agent is released. Here, larval populations of two pergid sawflies, Heteroperreyia hubrichi and H. jorgenseni, were analyzed by LC-MS/MS to investigate whether they contain alleged toxic peptides. The first species is a potential candidate for biological control of the invasive weed Brazilian peppertree in Florida and Hawaii. The chemical analyses revealed the presence of the peptides pergidin (Perg), 4-valinepergidin (VPerg), dephosphorylated pergidin (dpPerg), lophyrotomin (LGln and LGlu). The effect of sawfly population for each species was significantly influencing peptide concentration. All peptides occurred at lower concentrations compared with purportedly toxic species of this sawfly family. However, the concentrations of the peptides are of concern for the welfare of wildlife and livestock that would be exposed to these species. These results demonstrate that release of this biological control agent in the invaded range may pose an environmental threat.


Assuntos
Anacardiaceae/metabolismo , Agentes de Controle Biológico/análise , Peptídeos/análise , Animais , Agentes de Controle Biológico/farmacologia , Cromatografia Líquida de Alta Pressão , Himenópteros/crescimento & desenvolvimento , Himenópteros/metabolismo , Larva/efeitos dos fármacos , Larva/metabolismo , Oligopeptídeos/análise , Oligopeptídeos/farmacologia , Peptídeos/farmacologia , Espectrometria de Massas em Tandem
11.
Analyst ; 143(22): 5487-5496, 2018 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-30289422

RESUMO

Antibody-drug conjugates (ADCs) are an emerging type of biotherapeutics that utilize multiple tissue-specific antibodies combined with a range of linker designs to enable the transportation and selective release of cytotoxic drugs in close proximity to tumours. Consisting of antibodies conjugated to small drug molecules through a variety of linkers, ADCs are chemically complex analytes. Here we present a unique experimental comparison of four techniques for ADC analysis: hydrophobic interaction chromatography (HIC-UV/Vis), reversed phase liquid chromatography mass spectrometry (RPLC-MS), using either a QToF or an Orbitrap analyser, and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Four different ADCs consisting of Trastuzumab, monomethyl auristatin E (MMAE) and a peptidic linker moiety differing in their respective stoichiometric ratios in regard to drug-to-antibody ratio (DAR) were used for the comparison. We found that the determined DAR from all techniques was comparable, while the accuracy of the molecular weights for the conjugated light and heavy chain differed more extensively. This indicates that the choice of a mass analyser is more crucial for determining the accurate weights of the light and heavy chains than to evaluate the DAR of a given batch. However, ambiguous DAR assignment in HIC-UV/Vis or bias for either the light or heavy chain fragments in the mass spectrometry-based techniques can influence the obtained average DAR value and the use of complementary techniques is advisable. Out of the four techniques evaluated, HIC-UV/Vis and MALDI required less time to obtain an average DAR value and would therefore be good for initial screenings in the early stages of the discovery phase of new ADCs.


Assuntos
Cromatografia de Fase Reversa/métodos , Imunoconjugados/análise , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Imunoconjugados/química , Peso Molecular , Oligopeptídeos/análise , Trastuzumab/análise
12.
Food Res Int ; 113: 407-413, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30195535

RESUMO

The purpose of this preliminary study was to discriminate the chemical fingerprints of Protected Designation of Origin (PDO) Grana Padano cheeses from non-PDO "Grana-type" cheeses by means of an untargeted metabolomic approach based on ultra-high-pressure liquid chromatography coupled to quadrupole time-of-flight mass spectrometer (UHPLC/QTOF-MS). Hierarchical cluster analysis and Orthogonal Projections to Latent Structures Discriminant Analysis (OPLS-DA) allowed discriminating PDO vs. non-PDO cheeses. Lipids (fatty acids and their derivatives, phospholipids and monoacylglycerols), amino acids and oligopeptides, together with plant-derived compounds were the markers having the highest discrimination potential. It can be postulated that Grana Padano value chain, as strictly defined in the PDO production specification rules, can drive the biochemical processes involved in cheese making and ripening in a distinct manner, thus leaving a defined chemical signature on the final product. These preliminary findings provide the basis for further authenticity studies, aiming to protect the designation of origin of PDO Grana Padano cheese by applying a comprehensive foodomics-based approach.


Assuntos
Queijo/análise , Queijo/classificação , Metabolômica/métodos , Aminoácidos/análise , Aminoácidos/química , Cromatografia Líquida de Alta Pressão , Análise por Conglomerados , Análise Discriminante , Análise de Alimentos , Lipídeos/análise , Lipídeos/química , Espectrometria de Massas , Análise Multivariada , Oligopeptídeos/análise , Oligopeptídeos/química
13.
Drug Test Anal ; 10(11-12): 1755-1760, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30239151

RESUMO

In this work, a novel initial testing assay based on liquid chromatography-mass spectrometry is presented, enabling the detection of peptidic drugs and drug candidates (< 2 kDa) prohibited in sports. The assay covers representatives and metabolites of gonadotropin releasing hormone and its analogs (GnRHs), growth hormone secretagogues (GHS), growth hormone releasing peptides (GHRPs), and the Vasopressin-analog Desmopressin. The general objective of this work was to reduce sample preparation efforts to a minimum while preserving highest possible sensitivity and specificity of the assay, demonstrating limits of detection between 50 and 200 pg/mL. Here, a "dilute-and-inject" strategy provides the simplest conceivable sample preparation procedure. Furthermore, the combination of well-established strategies for the determination of peptides, such as two-dimensional liquid chromatography, dimethyl sulfoxide (DMSO)-assisted electrospray ionization, high resolution mass spectrometric detection and a tailored reporter template, which facilitates data review enormously, provides a high-throughput initial testing assay for lower molecular mass peptidic and peptide-related analytes.


Assuntos
Desamino Arginina Vasopressina/análise , Hormônio Liberador de Gonadotropina/análise , Oligopeptídeos/análise , Detecção do Abuso de Substâncias/métodos , Espectrometria de Massas em Tandem/métodos , Cromatografia Líquida/métodos , Doping nos Esportes , Hormônio Liberador de Gonadotropina/análogos & derivados , Ensaios de Triagem em Larga Escala/métodos , Humanos , Limite de Detecção , Manejo de Espécimes
14.
Anal Chem ; 90(16): 9682-9686, 2018 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-30063332

RESUMO

Tandem mass spectrometry (MS/MS) is the primary method for discovering, identifying, and localizing post-translational modifications (PTMs) in proteins. However, conventional positive ion mode collision induced dissociation (CID)-based MS/MS often fails to yield site-specific information for labile and acidic modifications due to low ionization efficiency in positive ion mode and/or preferential PTM loss. While a number of alternative methods have been developed to address this issue, most require specialized instrumentation or indirect detection. In this work, we present an amine-reactive TEMPO-based free radical initiated peptide sequencing (FRIPS) approach for negative ion mode analysis of phosphorylated and sulfated peptides. FRIPS-based fragmentation generates sequence informative ions for both phosphorylated and sulfated peptides with no significant PTM loss. Furthermore, FRIPS is compared to positive ion mode CID, electron transfer dissociation (ETD), as well as negative ion mode electron capture dissociation (niECD) and CID, both in terms of sequence coverage and fragmentation efficiency for phospho- and sulfo-peptides. Because FRIPS-based fragmentation has no particular instrumentation requirements and shows limited PTM loss, we propose this approach as a promising alternative to current techniques for analysis of labile and acidic PTMs.


Assuntos
Radicais Livres/química , Oligopeptídeos/análise , Fosfopeptídeos/análise , Colecistocinina/análise , Colecistocinina/química , Hirudinas/análise , Hirudinas/química , Oligopeptídeos/química , Fosfopeptídeos/química , Fosforilação , Processamento de Proteína Pós-Traducional , Análise de Sequência de Proteína , Espectrometria de Massas em Tandem/métodos
15.
Astrobiology ; 18(11): 1403-1413, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30160529

RESUMO

In a previous study, we have showed that the elongation of an alanine oligopeptide [L-alanyl-L-alanyl-L-alanyl-L-alanine ((Ala)4)] to higher oligopeptides is enhanced by calcite and dolomite at 275°C, using a mineral-mediated hydrothermal flow reactor system. However, a problem during the use of hydrothermal flow reactor system was that some of the minerals, such as clay, could not be tested due to their clogging in the reactor. In this article, we attempted to analyze the scope of enhancement for the formation of L-alanyl-L-alanyl-L-alanyl-L-alanyl-L-alanine ((Ala)5) and higher oligopeptides with different minerals including clay minerals for the elongation of alanine oligopeptide at 175°C. First, carbonate minerals and some clay minerals showed an enhancement of the formation of (Ala)5 from (Ala)4. On the contrary, volcanic products showed strong inhibitory activities. According to the pH dependence on the (Ala)4 elongations, we confirmed that most enhancement and inhibitory activities are due to the pH influence on the elongation of (Ala)4. However, the enhancement of montmorillonite (Tsukinuno), sphalerite, apatite, tourmaline, calcite (Nitto Funka), and the inhibitory activities by volcanic ash (Shinmoedake), volcanic ash (Sakurajima), dickite, and pyrophillite are not simply due to the pH change in the presence of these minerals. The difference found between the previous and present studies suggests that the interaction kinetics of the aqueous phase with the mineral phase is also an important factor for the elongation of (Ala)4. These data imply that the environments with pH near neutral to weak alkaline and with minerals might have been useful for the accumulation of oligopeptides in hydrothermal conditions.


Assuntos
Alanina/análise , Fontes Hidrotermais , Minerais/análise , Oligopeptídeos/análise , Carbonato de Cálcio/química , Concentração de Íons de Hidrogênio
16.
Int J Mol Sci ; 19(7)2018 07 04.
Artigo em Inglês | MEDLINE | ID: mdl-29973491

RESUMO

Redox signaling regulates different gastrointestinal (G.I.) epithelium functions. At the intestinal level, the loss of redox homeostasis in intestinal epithelial cells (IECs) is responsible for the pathogenesis and development of a wide diversity of G.I. disorders. Thus, the manipulation of oxidative stress in IECs could represent an important pharmacological target for different diseases. In this study, peptides released from in vitro gastro intestinal digestion of different buffalo-milk commercial dairy products were identified and evaluated for their bioactive properties. In particular, six G.I. digests of dairy products were tested in a model of oxidative stress for IECs. Among them, buffalo ricotta cheese was the most active and the presence of an abundant ß-lactoglobulin peptide (YVEELKPTPEGDL, f:60-72) was also revealed. The antioxidant potential of the identified peptide was also evaluated in a model of hydrogen peroxide (H2O2)-induced oxidative stress in the IEC-6 cell line. The peptide was able to reduce ROS release, while, on the other hand, it increased nuclear factor (erythroid-derived 2)-like 2 (Nrf2) activation and the expression of antioxidant cytoprotective factors, such as heme oxygenase 1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO1), and superoxide dismutase (SOD). These results indicate that buffalo ricotta cheese-isolated peptide could have potential in the treatment of some gastrointestinal disorders.


Assuntos
Antioxidantes/farmacologia , Queijo/análise , Laticínios/análise , Lactoglobulinas/química , Leite/química , Oligopeptídeos/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Animais , Antioxidantes/análise , Búfalos , Linhagem Celular , Heme Oxigenase (Desciclizante)/metabolismo , Humanos , Mucosa Intestinal/metabolismo , NAD(P)H Desidrogenase (Quinona)/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Oligopeptídeos/análise , Oligopeptídeos/isolamento & purificação , Ratos , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo
17.
Ticks Tick Borne Dis ; 9(6): 1391-1394, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29960872

RESUMO

Tick-borne encephalitis virus (TBEV) is a zoonotic agent causing severe encephalitis in humans. IgM antibody detection is useful for the serological diagnosis of TBEV infection, because IgM has high specificity for each flavivirus and indicates a recent infection. Commercial IgM-ELISA kits are somewhat expensive and difficulties in their sensitivity have been suggested due to their format and formalin-inactivated antigens. Therefore, the development of an inexpensive IgM-ELISA with high specificity and sensitivity is needed. In this study, a µ-capture ELISA was developed to detect TBEV-specific IgM antibodies using subviral particles (SPs) with strep-tag (strep-SP-IgM-ELISA). The results of our strep-SP-IgM-ELISA were highly correlated with diagnoses made by the neutralization test (sensitivity: 94.1%), and our strep-SP-IgM-ELISA could detect anti-TBEV IgM antibodies in patients who could not be diagnosed with the neutralization test. Besides, 51 of 52 positive samples by a commercial IgM-ELISA were also diagnosed as positive by our strep-SP-IgM-ELISA (98.1%), and our strep-SP-IgM-ELISA could detect anti-TBEV IgM antibodies in all samples that were inconclusive based on the commercial IgM-ELISA. Our strep-SP-IgM-ELISA will be useful for diagnoses in TBE-endemic areas.


Assuntos
Anticorpos Antivirais/sangue , Vírus da Encefalite Transmitidos por Carrapatos/isolamento & purificação , Encefalite Transmitida por Carrapatos/diagnóstico , Ensaio de Imunoadsorção Enzimática/métodos , Imunoglobulina M/sangue , Oligopeptídeos , Testes Sorológicos/métodos , Humanos , Oligopeptídeos/análise , Vírion/metabolismo
18.
Biotechniques ; 64(6): 245-253, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29939094

RESUMO

The goal in the presented study was to develop a simple, fast and accurate method for measuring the surface density of a short peptide sequence bound to a polymeric substrate. We analyzed polyurethane samples chemically modified with acrylic acid and polyurethane-grafted peptide (GSGREDVGSG) and investigated the possibility of using the bicinchoninic acid (BCA) assay to determine surface density of the solid-supported peptide. We set the conditions (temperature, time) under which the test should be conducted. We also studied the interaction of the BCA reagent with polyurethane substrate and the effect of drying conditions as well as material type and form on the test response. We have proposed potential factors that might interfere with the BCA assay and chosen the proper control materials.


Assuntos
Colorimetria/métodos , Oligopeptídeos/análise , Poliuretanos/análise , Quinolinas/química , Indicadores e Reagentes , Modelos Lineares
19.
Anal Chem ; 90(12): 7761-7768, 2018 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-29851349

RESUMO

Early screening of clinically relevant pathogens in the environment is a highly desirable goal in clinical care, providing precious information that will improve patient-care outcomes. In this work, a glove-based electrochemical sensor has been designed for point-of-use screening of Pseudomonas aeruginosa's virulence factors. The methodology used for the elaboration of the fabric platform relied on printing the conductive inks on the index and middle fingers of the glove, with the goal of screening pyocyanin and pyoverdine targets. The analytical signatures of the analytes were recorded in about 4 min, via the rapid and selective square-wave-voltammetry technique. Finger-based sensors display good performance and discrimination between the targets and potential interferences, along with good reproducibility. The sensors featured linearity over the 0.01-0.1 µM range for pyocyanin and 5-50 µM range for pyoverdine, with sensitivities of 2.51 µA/µM for pyocyanin and 1.09 nA/µM for pyoverdine ( R2 = 0.990 and 0.995, respectively) and detection limits of 3.33 nM for pyocyanin and 1.66 µM for pyoverdine. Moreover, the sensors were tested in binary mixtures of analytes, with successful outcomes. In order to gain information from the surrounding environment, the active electronic areas of the printed fingers were coated with a conductive hydrogel matrix, and relevant target surfaces were "swiped for notification" of contaminants. The simple fabrication, low-cost, and reusability of the proposed glove are likely to underpin the progressive drive of wearable sensors toward decentralized environmental and healthcare applications.


Assuntos
Técnicas Eletroquímicas , Oligopeptídeos/análise , Impressão , Pseudomonas aeruginosa/química , Piocianina/análise , Fatores de Virulência/análise , Técnicas Eletroquímicas/instrumentação , Elétrons , Humanos , Impressão/instrumentação , Soluções
20.
Growth Horm IGF Res ; 41: 1-6, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29864719

RESUMO

Detecting agents allegedly or evidently promoting growth such as human growth hormone (GH) or growth hormone releasing peptides (GHRP) in doping controls has represented a pressing issue for sports drug testing laboratories. While GH is a recombinant protein with a molecular weight of 22 kDa, the GHRPs are short (3-6 amino acids long) peptides with GH releasing properties. The endogenously produced GH (22 kDa isoform) consists of 191 amino acids and has a monoisotopic molecular mass of 22,124 Da. Within this study, a slightly modified form of GH was discovered consisting of 192 amino acids carrying an additional alanine at the N-terminus, leading to a monoisotopic mass of 22,195 Da. This was confirmed by top-down and bottom-up experiments using liquid chromatography coupled to high resolution/high accuracy mass spectrometry. Additionally, three analogues of GHRPs were identified as Gly-GHRP-6, Gly-GHRP-2 and Gly-Ipamorelin, representing the corresponding GHRP extended by a N-terminal glycine residue. The structure of these peptides was characterised by means of high resolution (tandem) mass spectrometry, and for Gly-Ipamorelin and Gly-GHRP-2 their identity was additionally confirmed by custom synthesis. Further, established in-vitro experiments provided preliminary information considering the potential metabolism after administration.


Assuntos
Doping nos Esportes , Hormônio do Crescimento Humano/análise , Oligopeptídeos/análise , Detecção do Abuso de Substâncias/métodos , Cromatografia Líquida , Humanos , Espectrometria de Massas em Tandem
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA