RESUMO
The common octopus (Octopus vulgaris) is an excellent candidate for aquaculture diversification, due to its biological traits and high market demand. To ensure a high-quality product while maintaining welfare in captive environments, it is crucial to develop non-invasive methods for testing health biomarkers. Proteins found in skin mucus offer a non-invasive approach to monitoring octopus welfare. This study compares the protein profiles in the skin mucus of wild, aquarium-maintained, and senescent specimens to identify welfare biomarkers. A tandem mass tag (TMT) coupled with an Orbitrap Eclipse Tribrid mass spectrometer was used to create a reference dataset from octopus skin mucus, identifying 1496 non-redundant protein groups. Although similar profiles were observed, differences in relative abundances led to the identification of potential biomarkers, including caspase-3-like, protocadherin 4, deleted in malignant brain tumors, thioredoxin, papilin, annexin, cofilin and mucin-4 proteins. Some of these proteins also revealed potential as bioactive peptides. This investigation provides the most extensive analysis of the skin mucus proteome in the common octopus and is the first to explore how aquarium maintenance and senescence alter the mucus proteome. This research highlights the potential of skin mucus protein/peptides as non-invasive monitoring biomarkers in cultured animals.
Assuntos
Muco , Octopodiformes , Proteoma , Pele , Animais , Octopodiformes/metabolismo , Pele/metabolismo , Proteoma/metabolismo , Muco/metabolismo , Biomarcadores/metabolismo , Proteômica/métodos , Envelhecimento/metabolismo , AquiculturaRESUMO
Helminths are masters at manipulating host's immune response. Especially, parasitic nematodes have evolved strategies that allow them to evade, suppress, or modulate host's immune response to persist and spread in the host's organism. While the immunomodulatory effects of nematodes on their hosts are studied with a great commitment, very little is known about nematodes' own immune system, immune response to their pathogens, and interactions between parasites and bacteria in the host's organism. To illustrate the response of the parasitic nematode Anisakis simplex s.s. during simulated interaction with Escherichia coli, different concentrations of lipopolysaccharide (LPS) were used, and the proteomic analysis with isobaric mass tags for relative and absolute quantification (tandem mass tag-based LC-MS/MS) was performed. In addition, gene expression and biochemical analyses of selected markers of oxidative stress were determined. The results revealed 1148 proteins in a group of which 115 were identified as differentially regulated proteins, for example, peroxiredoxin, thioredoxin, and macrophage migration inhibitory factor. Gene Ontology annotation and Reactome pathway analysis indicated that metabolic pathways related to catalytic activity, oxidation-reduction processes, antioxidant activity, response to stress, and innate immune system were the most common, in which differentially regulated proteins were involved. Further biochemical analyses let us confirm that the LPS induced the oxidative stress response, which plays a key role in the innate immunity of parasitic nematodes. Our findings, to our knowledge, indicate for the first time, the complexity of the interaction of parasitic nematode, A. simplex s.s. with bacterial LPS, which mimics the coexistence of helminth and gut bacteria in the host. The simulation of this crosstalk led us to conclude that the obtained results could be hugely valuable in the integrated systems biology approach to describe a relationship between parasite, host, and its commensal bacteria.
Assuntos
Anisakis/efeitos dos fármacos , Proteínas de Helminto/metabolismo , Lipopolissacarídeos/farmacologia , Animais , Anisakis/genética , Anisakis/metabolismo , Anisakis/microbiologia , Escherichia coli/fisiologia , Proteínas de Helminto/genética , Interações Hospedeiro-Patógeno , Estresse Oxidativo , ProteômicaRESUMO
The common octopus (Octopus vulgaris) is nowadays the most demanded cephalopod species for human consumption. This species was also postulated for aquaculture diversification to supply its increasing demand in the market worldwide, which only relies on continuously declining field captures. In addition, they serve as model species for biomedical and behavioral studies. Body parts of marine species are usually removed before reaching the final consumer as by-products in order to improve preservation, reduce shipping weight, and increase product quality. These by-products have recently attracted increasing attention due to the discovery of several relevant bioactive compounds. Particularly, the common octopus ink has been described as having antimicrobial and antioxidant properties, among others. In this study, the advanced proteomics discipline was applied to generate a common octopus reference proteome to screen potential bioactive peptides from fishing discards and by-products such as ink. A shotgun proteomics approach by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) using an Orbitrap Elite instrument was used to create a reference dataset from octopus ink. A total of 1432 different peptides belonging to 361 non-redundant annotated proteins were identified. The final proteome compilation was investigated by integrated in silico studies, including gene ontology (GO) term enrichment, pathways, and network studies. Different immune functioning proteins involved in the innate immune system, such as ferritin, catalase, proteasome, Cu/Zn superoxide dismutase, calreticulin, disulfide isomerase, heat shock protein, etc., were found in ink protein networks. Additionally, the potential of bioactive peptides from octopus ink was addressed. These bioactive peptides can exert beneficial health properties such as antimicrobial, antioxidant, antihypertensive, and antitumoral properties and are therefore considered lead compounds for developing pharmacological, functional foods or nutraceuticals.
Assuntos
Octopodiformes , Proteoma , Animais , Humanos , Proteoma/metabolismo , Proteômica/métodos , Octopodiformes/química , Cromatografia Líquida , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Tinta , Espectrometria de Massas em Tandem , Peptídeos/químicaRESUMO
Food safety is a critical aspect of public health and involves the handling, preparation, and storage of food to avoid contamination and foodborne illnesses [...].
Assuntos
Microbiologia de Alimentos , Proteômica , Proteômica/métodos , Manipulação de Alimentos/métodos , Inocuidade dos Alimentos , Perfilação da Expressão Gênica , Contaminação de AlimentosRESUMO
Food allergies (FA) have dramatically increased in recent years, particularly in developed countries. It is currently well-established that food tolerance requires the strict maintenance of a specific microbial consortium in the gastrointestinal (GI) tract microbiome as alterations in the gut microbiota can lead to dysbiosis, causing inflammation and pathogenic intestinal conditions that result in the development of FA. Although there is currently not enough knowledge to fully understand how the interactions between gut microbiota, host responses and the environment cause food allergies, recent advances in '-omics' technologies (i.e., proteomics, genomics, metabolomics) and in approaches involving systems biology suggest future headways that would finally allow the scientific understanding of the relationship between gut microbiome and FA. This review summarizes the current knowledge in the field of FA and insights into the future advances that will be achieved by applying proteomic techniques to study the GI tract microbiome in the field of FA and their medical treatment. Metaproteomics, a proteomics experimental approach of great interest in the study of GI tract microbiota, aims to analyze and identify all the proteins in complex environmental microbial communities; with shotgun proteomics, which uses liquid chromatography (LC) for separation and tandem mass spectrometry (MS/MS) for analysis, as it is the most promising technique in this field.
Assuntos
Hipersensibilidade Alimentar , Microbioma Gastrointestinal , Microbiota , Humanos , Proteômica/métodos , Espectrometria de Massas em Tandem , Microbiota/fisiologiaRESUMO
The common octopus is a cephalopod species subject to active fisheries, with great potential in the aquaculture and food industry, and which serves as a model species for biomedical and behavioral studies. The analysis of the skin mucus allows us to study their health in a non-invasive way, by using a hardly exploited discard of octopus in the fishing sector. A shotgun proteomics approach combined with liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) using an Orbitrap-Elite instrument was used to create a reference dataset from octopus skin mucus. The final proteome compilation was investigated by integrated in-silico studies, including Gene Ontology (GO), the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, network studies, and prediction and characterization analysis of potential bioactive peptides. This work presents the first proteomic analysis of the common octopus skin mucus proteome. This library was created by merging 5937 identified spectra of 2038 different peptides. A total of 510 non-redundant proteins were identified. Obtained results show proteins closely related to the defense, which highlight the role of skin mucus as the first barrier of defense and the interaction with the environment. Finally, the potential of the bioactive peptides with antimicrobial properties, and their possible application in biomedicine, pharmaceutical, and nutraceutical industry was addressed.
Assuntos
Octopodiformes , Proteogenômica , Animais , Proteômica/métodos , Proteoma/metabolismo , Octopodiformes/química , Octopodiformes/metabolismo , Cromatografia Líquida , Espectrometria de Massas em Tandem , Peptídeos/metabolismo , Muco/metabolismoRESUMO
Biogenic amine-producing bacteria are responsible for the production of basic nitrogenous compounds (histamine, cadaverine, tyramine, and putrescine) following the spoilage of food due to microorganisms. In this study, we adopted a shotgun proteomics strategy to characterize 15 foodborne strains of biogenic-amine-producing bacteria. A total of 10,673 peptide spectrum matches belonging to 4081 peptides and corresponding to 1811 proteins were identified. Relevant functional pathways were determined, and strains were differentiated into hierarchical clusters. An expected protein-protein interaction network was created (260 nodes/1973 interactions). Most of the determined proteins were associated with networks/pathways of energy, putrescine metabolism, and host-virus interaction. Additionally, 556 peptides were identified as virulence factors. Moreover, 77 species-specific peptide biomarkers corresponding to 64 different proteins were proposed to identify 10 bacterial species. This represents a major proteomic dataset of biogenic-amine-producing strains. These results may also be suitable for new treatments for food intoxication and for tracking microbial sources in foodstuffs.
Assuntos
Proteômica , Putrescina , Putrescina/metabolismo , Aminas Biogênicas/metabolismo , Bactérias/metabolismo , Peptídeos/metabolismo , Alimentos Marinhos , Microbiologia de AlimentosRESUMO
The spectrophotometric methodology for carbonate ion determination in seawater was first published in 2008 and has been continuously evolving in terms of reagents and formulations. Although being fast, relatively simple, affordable, and potentially easy to implement in different platforms and facilities for discrete and autonomous observations, its use is not widespread in the ocean acidification community. This study uses a merged overdetermined CO2 system data set (carbonate ion, pH, and alkalinity) obtained from 2009 to 2020 to assess the differences among the five current approaches of the methodology through an internal consistency analysis and discussing the sources of uncertainty. Overall, the results show that none of the approaches meet the climate goal (± 1 % standard uncertainty) for ocean acidification studies for the whole carbonate ion content range in this study but usually fulfill the weather goal (± 10 % standard uncertainty). The inconsistencies observed among approaches compromise the consistency of data sets among regions and through time, highlighting the need for a validated standard operating procedure for spectrophotometric carbonate ion measurements as already available for the other measurable CO2 variables.
Assuntos
Dióxido de Carbono , Água do Mar , Carbonato de Cálcio , Dióxido de Carbono/análise , Carbonatos , Concentração de Íons de Hidrogênio , Oceanos e Mares , Espectrofotometria/métodosRESUMO
CONTEXT: The corpus luteum (CL) is an endocrine gland in the ovary of mature females during the oestrous cycle and pregnancy. There is evidence of a relationship between the secretory function of the CL and PPARs. AIMS: In this study, we investigated the changes in the proteome of the CL in relation to the phase of the oestrous cycle and the impact of PPARγ ligands on the proteomic profile of the CL during the mid- and late-luteal phase of the oestrous cycle. METHODS: The porcine CL explants were incubated in vitro for 6h in the presence of PPARγ ligands (agonist pioglitazone, antagonist T0070907) or without ligands. Global proteomic analysis was performed using the TMT-based LC-MS/MS method. KEY RESULTS: The obtained results showed the disparity in proteomic profile of the untreated CL - different abundance of 23 and 28 proteins for the mid- and late-luteal phase, respectively. Moreover, seven proteins were differentially regulated in the CL tissue treated with PPARγ ligands. In the mid-luteal phase, one protein, CAND1, was downregulated after treatment with T0070907. In the late-luteal phase, the proteins SPTAN1, GOLGB1, TP53BP1, MATR3, RRBP1 and SRRT were upregulated by pioglitazone. CONCLUSIONS: Comparative proteomic analysis revealed that certain proteins constitute a specific proteomic signature for each examined phase. Moreover, the study showed that the effect of PPARγ ligands on the CL proteome was rather limited. IMPLICATIONS: The results provide a broader insight into the processes that may be responsible for the structural luteolysis of the porcine CL, in addition to apoptosis and autophagy.
Assuntos
Ciclo Estral , PPAR gama , Animais , Cromatografia Líquida , Corpo Lúteo/metabolismo , Feminino , Ligantes , PPAR gama/metabolismo , Pioglitazona/análise , Pioglitazona/metabolismo , Pioglitazona/farmacologia , Gravidez , Proteoma/metabolismo , Proteômica , Suínos , Espectrometria de Massas em TandemRESUMO
The microbiota present in the gastrointestinal tract is involved in the development or prevention of food allergies and autoimmune disorders; these bacteria can enter the gallbladder and, depending on the species involved, can either be benign or cause significant diseases. Occlusion of the gallbladder, usually due to the presence of calculi blocking the bile duct, facilitates microbial infection and inflammation, which can be serious enough to require life-saving surgery. In addition, the biliary salts are secreted into the intestine and can affect the gut microbiota. The interaction between the gut microbiota, pathogenic organisms, and the human immune system can create intestinal dysbiosis, generating a variety of syndromes including the development of food allergies and autoimmune disorders. The intestinal microbiota can aggravate certain food allergies, which become severe when the integrity of the intestinal barrier is affected, allowing bacteria, or their metabolites, to cross the intestinal barrier and invade the bloodstream, affecting distal body organs. This article deals with health conditions and severe diseases that are either influenced by the gut flora or caused by gallbladder obstruction and inflammation, as well as putative treatments for those illnesses.
Assuntos
Doenças Autoimunes , Hipersensibilidade Alimentar , Microbioma Gastrointestinal , Humanos , Vesícula Biliar , Intestinos/microbiologia , InflamaçãoRESUMO
Enterococcus species are Gram-positive bacteria that are normal gastrointestinal tract inhabitants that play a beneficial role in the dairy and meat industry. However, Enterococcus species are also the causative agents of health care-associated infections that can be found in dairy and fermented food products. Enterococcal infections are led by strains of Enterococcus faecalis and Enterococcus faecium, which are often resistant to antibiotics and biofilm formation. Enterococci virulence factors attach to host cells and are also involved in immune evasion. LC-MS/MS-based methods offer several advantages compared with other approaches because one can directly identify microbial peptides without the necessity of inferring conclusions based on other approaches such as genomics tools. The present study describes the use of liquid chromatography−electrospray ionization tandem mass spectrometry (LC−ESI−MS/MS) to perform a global shotgun proteomics characterization for opportunistic pathogenic Enterococcus from different dairy and fermented food products. This method allowed the identification of a total of 1403 nonredundant peptides, representing 1327 proteins. Furthermore, 310 of those peptides corresponded to proteins playing a direct role as virulence factors for Enterococcus pathogenicity. Virulence factors, antibiotic sensitivity, and proper identification of the enterococcal strain are required to propose an effective therapy. Data are available via ProteomeXchange with identifier PXD036435. Label-free quantification (LFQ) demonstrated that the majority of the high-abundance proteins corresponded to E. faecalis species. Therefore, the global proteomic repository obtained here can be the basis for further research into pathogenic Enterococcus species, thus facilitating the development of novel therapeutics.
Assuntos
Enterococcus , Alimentos Fermentados , Antibacterianos/farmacologia , Cromatografia Líquida , Farmacorresistência Bacteriana , Enterococcus faecalis , Microbiologia de Alimentos , Testes de Sensibilidade Microbiana , Proteômica , Espectrometria de Massas em Tandem , Fatores de VirulênciaRESUMO
Anisakis simplex s. s. is a parasitic nematode of marine mammals and causative agent of anisakiasis in humans. The cuticle and intestine of the larvae are the tissues most responsible for direct and indirect contact, respectively, of the parasite with the host. At the L4 larval stage, tissues, such as the cuticle and intestine, are fully developed and functional, in contrast to the L3 stage. As such, this work provides for the first time the tissue-specific proteome of A. simplex s. s. larvae in the L4 stage. Statistical analysis (FC ≥ 2; p-value ≤ 0.01) showed that 107 proteins were differentially regulated (DRPs) between the cuticle and the rest of the larval body. In the comparison between the intestine and the rest of the larval body at the L4 stage, 123 proteins were identified as DRPs. Comparison of the individual tissues examined revealed a total of 272 DRPs, with 133 proteins more abundant in the cuticle and 139 proteins more abundant in the intestine. Detailed functional analysis of the identified proteins was performed using bioinformatics tools. Glycolysis and the tricarboxylic acid cycle were the most enriched metabolic pathways by cuticular and intestinal proteins, respectively, in the L4 stage of A. simplex s. s. The presence of two proteins, folliculin (FLCN) and oxoglutarate dehydrogenase (OGDH), was confirmed by Western blot, and their tertiary structure was predicted and compared with other species. In addition, host-pathogen interactions were identified, and potential new allergens were predicted. The result of this manuscript shows the largest number of protein identifications to our knowledge using proteomics tools for different tissues of L4 larvae of A. simplex s. s. The identified tissue-specific proteins could serve as targets for new drugs against anisakiasis.
Assuntos
Anisaquíase , Anisakis , Animais , Anisaquíase/parasitologia , Anisakis/química , Anisakis/metabolismo , Metabolismo dos Carboidratos , Humanos , Larva/metabolismo , Mamíferos/metabolismo , Proteoma/metabolismoRESUMO
Enterococcus belongs to a group of microorganisms known as lactic acid bacteria (LAB), which constitute a broad heterogeneous group of generally food-grade microorganisms historically used in food preservation. Enterococci live as commensals of the gastrointestinal tract of warm-blooded animals, although they also are present in food of animal origin (milk, cheese, fermented sausages), vegetables, and plant materials because of their ability to survive heat treatments and adverse environmental conditions. The biotechnological traits of enterococci can be applied in the food industry; however, the emergence of enterococci as a cause of nosocomial infections makes their food status uncertain. Recent advances in high-throughput sequencing allow the subtyping of bacterial pathogens, but it cannot reflect the temporal dynamics and functional activities of microbiomes or bacterial isolates. Moreover, genetic analysis is based on sequence homologies, inferring functions from databases. Here, we used an end-to-end proteomic workflow to rapidly characterize two bacteriocin-producing Enterococcus faecium (Efm) strains. The proteome analysis was performed with liquid chromatography coupled to a trapped ion mobility spectrometry-time-of-flight mass spectrometry instrument (TimsTOF) for high-throughput and high-resolution characterization of bacterial proteins. Thus, we identified almost half of the proteins predicted in the bacterial genomes (>1100 unique proteins per isolate), including quantifying proteins conferring resistance to antibiotics, heavy metals, virulence factors, and bacteriocins. The obtained proteomes were annotated according to function, resulting in 22 complete KEGG metabolic pathway modules for both strains. The workflow used here successfully characterized these bacterial isolates and showed great promise for determining and optimizing the bioengineering and biotechnology properties of other LAB strains in the food industry.
Assuntos
Bacteriocinas , Queijo , Enterococcus faecium , Animais , Enterococcus faecium/genética , Enterococcus faecium/metabolismo , Bacteriocinas/metabolismo , Proteômica , Enterococcus , Queijo/microbiologiaRESUMO
Some Listeria species are important human and animal pathogens that can be found in contaminated food and produce a variety of virulence factors involved in their pathogenicity. Listeria strains exhibiting multidrug resistance are known to be progressively increasing and that is why continuous monitoring is needed. Effective therapy against pathogenic Listeria requires identification of the bacterial strain involved, as well as determining its virulence factors, such as antibiotic resistance and sensitivity. The present study describes the use of liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) to do a global shotgun proteomics characterization for pathogenic Listeria species. This method allowed the identification of a total of 2990 non-redundant peptides, representing 2727 proteins. Furthermore, 395 of the peptides correspond to proteins that play a direct role in Listeria pathogenicity; they were identified as virulence factors, toxins and anti-toxins, or associated with either antibiotics (involved in antibiotic-related compounds production or resistance) or resistance to toxic substances. The proteomic repository obtained here can be the base for further research into pathogenic Listeria species and facilitate the development of novel therapeutics for these pathogens.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/química , Farmacorresistência Bacteriana Múltipla , Listeria/efeitos dos fármacos , Listeria/patogenicidade , Proteoma/química , Fatores de Virulência/química , Transportadores de Cassetes de Ligação de ATP/química , Cromatografia Líquida/métodos , Genes Bacterianos , Listeria/classificação , Listeria/genética , Peptídeos/química , Proteômica/métodos , Espectrometria de Massas por Ionização por Electrospray/métodos , Espectrometria de Massas em Tandem/métodosRESUMO
BACKGROUND: This study addresses the quality loss of European hake (Merluccius merluccius) during frozen storage. Its objective was to comparatively analyse the effect of a previous high-pressure processing (HPP) (150 MPa for 2 min) when different storage temperatures (-10, -18 and -30 °C) were employed. RESULTS: Most chemical quality indices (trimethylamine, TMA; dimethylamine, DMA, formaldehyde, FA; free fatty acids, FFAs) provided a marked content increase with freezing and frozen storage time, values being higher by increasing the storage temperature. Previous HPP led to an inhibitory (P < 0.05) effect on the TMA, DMA, FA and FFA formation in frozen fish kept at -10 and -18 °C, the preservative effect being higher at the highest temperature tested; however, in agreement to the low damage development, no effect could be proved on samples stored at -30 °C. Concerning lipid oxidation, peroxides formation was found low, although a slight increasing effect (P < 0.05) was implied in fish corresponding to all temperatures as a result of the previous HPP; furthermore, an inhibitory effect (P < 0.05) on fluorescent compounds formation (tertiary lipid oxidation) was evident after freezing and at month 9 for -10 °C samples. CONCLUSION: It is concluded that a 150-MPa high-pressure treatment may inhibit the formation of degradative molecules such as DMA, FA, TMA and FFAs during the frozen storage at -18 °C (maximum recommended) and -10 °C. However, results have indicated that lowering the storage temperature showed to be more effective than the current HPP (150 MPa for 2 min). © 2020 Society of Chemical Industry.
Assuntos
Conservação de Alimentos/métodos , Carne/análise , Animais , Conservação de Alimentos/instrumentação , Armazenamento de Alimentos , Congelamento , Gadiformes , Controle de QualidadeRESUMO
Jumbo squid (Dosidicus gigas) is one of the largest cephalopods, and represents an important economic fishery in several regions of the Pacific Ocean, from southern California in the United States to southern Chile. Large and considerable discards of this species, such as skin, have been reported to constitute an important source of potential by-products. In this paper, a shotgun proteomics approach was applied for the first time to the characterization of the jumbo squid (Dosidicus gigas) skin proteome. A total of 1004 different peptides belonging to 219 different proteins were identified. The final proteome compilation was investigated by integrated in-silico studies, including gene ontology (GO) term enrichment, pathways, and networks studies. Potential new valuable bioactive peptides such as antimicrobial, bioactive collagen peptides, antihypertensive and antitumoral peptides were predicted to be present in the jumbo squid skin proteome. The integration of the global proteomics results and the bioinformatics analysis of the jumbo squid skin proteome show a comprehensive knowledge of this fishery discard and provide potential bioactive peptides of this marine by-product.
Assuntos
Decapodiformes/química , Peptídeos/isolamento & purificação , Proteínas/isolamento & purificação , Pele/química , Animais , Biologia Computacional , Simulação por Computador , Peptídeos/química , Peptídeos/farmacologia , Proteínas/química , Proteínas/farmacologia , ProteômicaRESUMO
We propose a new workflow for fast phosphoproteome profiling. The workflow is based on the use of accelerated in-solution trypsin digestion under an ultrasonic field provided by high-intensity focused ultrasound (HIFU) combined with an inverse strategy based on TiO2 selective phosphopeptide enrichment, fractionation by strong cation exchange chromatography (SCX) and analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS) using a high-resolution mass spectrometer. The performance of the method was established for the global phosphoproteome analysis of unstimulated human Jurkat leukemia T cells (E6.1). Using this accelerated workflow, 15367 phosphorylation sites from 13029 different phosphopeptides belonging to 3163 different phosphoproteins were efficiently identified with high-throughput and reproducibility in less than 15 h. The functional analysis revealed significant phosphorylation-based networks that are implicated in immune function and tumor development pathways. The present strategy, HIFU-TiO2-SCX-LC-MS/MS, is the fastest analytical method reported to date for generating large-scale phosphoproteomics data sets (<15 h).
Assuntos
Cromatografia por Troca Iônica/métodos , Fosfoproteínas/metabolismo , Espectrometria de Massas em Tandem/métodos , Titânio/química , Cromatografia Líquida de Alta Pressão , Humanos , Células Jurkat , Fosfopeptídeos/análise , Fosfoproteínas/análise , Reprodutibilidade dos Testes , Linfócitos T/citologia , Linfócitos T/metabolismoRESUMO
The aim of this work was the purification and identification of the major angiotensin converting enzyme (ACE) inhibitory peptides produced by enzymatic hydrolysis of a protein concentrate recovered from a cuttlefish industrial manufacturing effluent. This process consisted on the ultrafiltration of cuttlefish softening wastewater, with a 10 kDa cut-off membrane, followed by the hydrolysis with alcalase of the retained fraction. Alcalase produced ACE inhibitors reaching the highest activity (IC50 = 76.8 ± 15.2 µg mL⻹) after 8 h of proteolysis. Sequential ultrafiltration of the 8 h hydrolysate with molecular weight cut-off (MWCO) membranes of 10 and 1 kDa resulted in the increased activity of each permeate, with a final IC50 value of 58.4 ± 4.6 µg mL⻹. Permeate containing peptides lower than 1 kDa was separated by reversed-phase high performance liquid chromatography (RP-HPLC). Four fractions (A-D) with potent ACE inhibitory activity were isolated and their main peptides identified using high performance liquid chromatography coupled to an electrospray ion trap Fourier transform ion cyclotron resonance-mass spectrometer (HPLC-ESI-IT-FTICR) followed by comparison with databases and de novo sequencing. The amino acid sequences of the identified peptides contained at least one hydrophobic and/or a proline together with positively charged residues in at least one of the three C-terminal positions. The IC50 values of the fractions ranged from 1.92 to 8.83 µg mL⻹, however this study fails to identify which of these peptides are ultimately responsible for the potent antihypertensive activity of these fractions.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Decapodiformes/metabolismo , Resíduos Industriais/análise , Peptídeos/farmacologia , Águas Residuárias/análise , Algoritmos , Inibidores da Enzima Conversora de Angiotensina/isolamento & purificação , Animais , Cromatografia Líquida de Alta Pressão , Decapodiformes/química , Relação Dose-Resposta a Droga , Indústria Alimentícia , Hidrólise , Peso Molecular , Dinâmica não Linear , Peptídeos/isolamento & purificação , Espectrometria de Massas por Ionização por Electrospray , UltrafiltraçãoRESUMO
Food allergy, referred to as the atypical physiological overreaction of the immune system after exposure to specific food components, is considered one of the major concerns in food safety. The prevalence of this emerging worldwide problem has been increasing during the last decades, especially in industrialized countries, being estimated to affect 6-8% of young children and about 2-4% of adults. Marine organisms are an important source of bioactive substances with the potential to functionally improve the immune system, reduce food allergy sensitization and development, and even have an anti-allergic action in food allergy. The present investigation aims to be a comprehensive report of marine bioactive compounds with verified actions to improve food allergy and identified mechanisms of actions rather than be an exhaustive compilation of all investigations searching beneficial effects of marine compounds in FA. Particularly, this research highlights the capacity of bioactive components extracted from marine microbial, animal, algae, and microalgae sources, such as n-3 long-chain polyunsaturated fatty acids (LC-PUFA), polysaccharide, oligosaccharide, chondroitin, vitamin D, peptides, pigments, and polyphenols, to regulate the immune system, epigenetic regulation, inflammation, and gut dysbiosis that are essential factors in the sensitization and effector phases of food allergy. In conclusion, the marine ecosystem is an excellent source to provide foods with the capacity to improve the hypersensitivity induced against specific food allergens and also bioactive compounds with a potential pharmacological aptitude to be applied as anti-allergenic in food allergy.
Assuntos
Organismos Aquáticos , Hipersensibilidade Alimentar , Hipersensibilidade Alimentar/imunologia , Humanos , AnimaisRESUMO
Fishery products are one of the main human nutritional sources, and due to the consumption increase, the quality of the derived products may be modified, during catching, technological processing, and storage. Detection and identification of pathogenic and spoilage microorganisms in fishery products is needed because the first may be involved in human diseases, while the second is responsible of significant economic losses. In this sense, liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method and computational analysis of MS data are useful tools for characterizing and identifying different microorganisms and to develop promising strategies for food science investigations. Moreover, in the past decade, metaproteomic methodologies have progressed for the study of microorganisms isolated from their natural samples and independently of the culture restrictions. Metaproteomics enables assessment of proteins and pathways from individual members of the consortium. Metaproteomics can provide a detailed understanding of which organisms occupy specific metabolic niches, how they interact, and how they utilize nutrients, and these insights can be obtained directly from environmental samples.According to that, the sample preparation of the fishery product, the LC-ESI-MS/MS dedicated method, and the MS data analysis were described in the present chapter to obtain the metaproteomic analysis of the respective microbiomes or microbial communities.