Unlocking hot trub's potential: a simple method for extracting bitter acids and xanthohumol.

BACKGROUND: Hot trub is a macronutrient- and micronutrient-rich by-product generated in the brewing industry, which is still underrated as a raw material for reprocessing purposes. In this context, this study aimed to investigate the extraction of bitter acids' and xanthohumol from hot trub as well as identify the significance of parameters for the process. The research assessed various extraction parameters, such as pH, ethanol concentration, temperature, and solid-to-liquid ratio, using a Plackett-Burman design. RESULTS: Ethanol concentration and pH were the most significant parameters affecting extraction yield. ß-acids were found to be the principal components of the bitter acids, with a maximum concentration near 16 mg g-1, followed by iso-α-acids and α-acids achieving 6 and 3.6 mg g-1, respectively. The highest yields of bitter acids were observed in the highest ethanol concentration, while pH was relevant to extraction process in treatments with low ethanol ratios. Concerning the xanthohumol extraction, the approach achieved maximum concentration (239 µg g-1) in treatments with ethanol concentration above 30%. Despite their variances, the phytochemicals exhibited comparable extraction patterns, indicating similar interactions with macromolecules. Moreover, the characterization of the solid residues demonstrated that the extraction process did not bring about any alterations to the chemical and total protein profiles. CONCLUSION: Ethanol concentration was found to have the most significant impact on the extraction of bitter acids and xanthohumol, while temperature had no significant effect. The solid remains resulting from the extraction showed potential for use as a protein source. © 2024 Society of Chemical Industry.

An overview of metabolomic and proteomic profiling in bipolar disorder and its clinical value.

INTRODUCTION: Bipolar disorder (BD) is a complex psychiatric disease characterized by alternating mood episodes. As for any other psychiatric illness, currently there is no biochemical test that is able to support diagnosis or therapeutic decisions for BD. In this context, the discovery and validation of biomarkers are interesting strategies that can be achieved through proteomics and metabolomics. AREAS COVERED: In this descriptive review, a literature search including original articles and systematic reviews published in the last decade was performed with the objective to discuss the results of BD proteomic and metabolomic profiling analyses and indicate proteins and metabolites (or metabolic pathways) with potential clinical value. EXPERT OPINION: A large number of proteins and metabolites have been reported as potential BD biomarkers; however, most studies do not reach biomarker validation stages. An effort from the scientific community should be directed toward the validation of biomarkers and the development of simplified bioanalytical techniques or protocols to determine them in biological samples, in order to translate proteomic and metabolomic findings into clinical routine assays.

Application of Lipidomics in Psychiatry: Plasma-Based Potential Biomarkers in Schizophrenia and Bipolar Disorder.

In this study, we obtained a lipidomic profile of plasma samples from drug-naïve patients with schizophrenia (SZ) and bipolar disorder (BD) in comparison to healthy controls. The sample cohort consisted of 30 BD and 30 SZ patients and 30 control individuals. An untargeted lipidomics strategy using liquid chromatography coupled with high-resolution mass spectrometry was employed to obtain the lipid profiles. Data were preprocessed, then univariate (t-test) and multivariate (principal component analysis and orthogonal partial least squares discriminant analysis) statistical tools were applied to select differential lipids, which were putatively identified. Afterward, multivariate receiver operating characteristic tests were performed, and metabolic pathway networks were constructed, considering the differential lipids. Our results demonstrate alterations in distinct lipid pathways, especially in glycerophospholipids, sphingolipids and glycerolipids, between SZ and BD patients. The results obtained in this study may serve as a basis for differential diagnosis, which is crucial for effective treatment and improving the quality of life of patients with psychotic disorders.

Understanding ayahuasca effects in major depressive disorder treatment through in vitro metabolomics and bioinformatics.

Emerging insights from metabolomic-based studies of major depression disorder (MDD) are mainly related to biochemical processes such as energy or oxidative stress, in addition to neurotransmission linked to specific metabolite intermediates. Hub metabolites represent nodes in the biochemical network playing a critical role in integrating the information flow in cells between metabolism and signaling pathways. Limited technical-scientific studies have been conducted to understand the effects of ayahuasca (Aya) administration in the metabolism considering MDD molecular context. Therefore, this work aims to investigate an in vitro primary astrocyte model by untargeted metabolomics of two cellular subfractions: secretome and intracellular content after pre-defined Aya treatments, based on DMT concentration. Mass spectrometry (MS)-based metabolomics data revealed significant hub metabolites, which were used to predict biochemical pathway alterations. Branched-chain amino acid (BCAA) metabolism, and vitamin B6 and B3 metabolism were associated to Aya treatment, as "housekeeping" pathways. Dopamine synthesis was overrepresented in the network results when considering the lowest tested DMT concentration (1 µmol L-1). Building reaction networks containing significant and differential metabolites, such as nicotinamide, L-DOPA, and L-leucine, is a useful approach to guide on dose decision and pathway selection in further analytical and molecular studies.

Metabolomics: A Powerful Tool to Understand the Schizophrenia Biology.

ABSVTRACT: Schizophrenia, as any other psychiatric disorder, is a multifactorial and complex illness whose etiology is not completely established. Therefore, studies involving strategies that are able to describe the molecular alterations caused by the disease and, consequently, indicate the altered metabolic pathways are of increasing interest. Metabolomics is a very suitable approach that can be applied for this task, since it consists of the evaluation of the set of metabolites contained in a biological system undergoing a biological process, such as a disease or treatment. In metabolomics, state-of-the-art analytical techniques (mass spectrometry and nuclear magnetic resonance) are employed to identify and quantify the metabolites present in the studied biological samples, and chemometric and bioinformatic tools are applied to determine the specific metabolites and metabolic pathways that are relevant to the biological process under investigation. The aim of this chapter is to describe the basic principles of metabolomics, how this strategy can improve the understanding of the schizophrenia biology, and the findings obtained so far.

Comprehending Cardiac Dysfunction by Oxidative Stress: Untargeted Metabolomics of In Vitro Samples.

Cardiovascular diseases (CVDs) are noncommunicable diseases known for their complex etiology and high mortality rate. Oxidative stress (OS), a condition in which the release of free radical exceeds endogenous antioxidant capacity, is pivotal in CVC, such as myocardial infarction, ischemia/reperfusion, and heart failure. Due to the lack of information about the implications of OS on cardiovascular conditions, several methodologies have been applied to investigate the causes and consequences, and to find new ways of diagnosis and treatment as well. In the present study, cardiac dysfunction was evaluated by analyzing cells' alterations with untargeted metabolomics, after simulation of an oxidative stress condition using hydrogen peroxide (H2O2) in H9c2 myocytes. Optimizations of H2O2 concentration, cell exposure, and cell recovery times were performed through MTT assays. Intracellular metabolites were analyzed right after the oxidative stress (oxidative stress group) and after 48 h of cell recovery (recovery group) by ultra-high-performance liquid chromatography coupled to mass spectrometry (UHPLC-MS) in positive and negative ESI ionization mode. Significant alterations were found in pathways such as "alanine, aspartate and glutamate metabolism", "glycolysis", and "glutathione metabolism", mostly with increased metabolites (upregulated). Furthermore, our results indicated that the LC-MS method is effective for studying metabolism in cardiomyocytes and generated excellent fit (R2Y > 0.987) and predictability (Q2 > 0.84) values.

Follistatin-related protein 1 interacting partner of Syndecan-1 promotes an aggressive phenotype on Oral Squamous cell carcinoma (OSCC) models.

Syndecans belong to the family of transmembrane heparan sulfate proteoglycans and are associated with many physiopathological processes, including oral cancer. As previously shown soluble syndecan-1 (SDC1) fragments and synthetic SDC1 peptide were able to induce cell migration in oral cancer cell lines. In order to explore the role of SDC1 in oral cancer, we have investigated SDC1 interacting partners and its functional role in oral cancer models. Here we have shown that SDC1 interacts with follistatin-related protein 1 (FSTL1) by its ectodomain (ectoSDC1) and extracellular juxtamembrane peptide (pepSDC1) and that their transcript levels can affect tumor events. Using orthotopic mouse model we identified that the knock-down for FSTL1 (shFSTL1) or for both FSTL1 and SDC1 (sh2KD) produced less aggressive and infiltrative tumors, with lower keratinization deposition, but with increased levels of epithelial-mesenchymal transition and proliferation compared to control and SDC1 knock-down. Based on cell culture assays, we suggest that the shFSTL1 effect on tumor tissues might be from significant increase of mRNA levels of Activin A (ActA) and its resceptors. This study shows for the first time two different complexes, SDC1 and FSTL1; pepSDC1 and FSTL1, exhibiting a close relationship in cell signaling events, as FSTL1 promotes a more aggressive phenotype. SIGNIFICANCE: This work contributes to the understanding of new SDC1 functions, based on the investigation of protein-protein complex formation in Oral Squamous cell carcinoma (OSCC) models. The FSTL1 identification, as an interacting partner of SDC1 ectodomain and of its derived peptide promotes molecular events that favors cancer development and progression, as highlighted by Activin A (ActA) and Epithelial-mesenchymal transition (EMT) gene expression and by changes in the phenotype of orthotopic OSCC mouse tumor tissues when SDC1-FSTL1 expression is modulated.

The New Omics Era into Systems Approaches: What Is the Importance of Separation Techniques?

Omics sciences have been facing challenges in different fields, especially in life sciences. One of these challenges involves assessing biology into systems interpretation. With the advance of genomics, molecular biology has been projected into the realm of systems biology. In a different direction, systems approaches are making definitive strides toward scientific understanding and biotechnological applications. Separation techniques provided meaningful progress in the omics era, conducting the classical molecular biology to contemporary systems biology. In this introductory chapter, the relevance of these techniques to the development of different omics sciences, within the systems biology context, will be discussed.

A pilot study indicating the dysregulation of the complement and coagulation cascades in treated schizophrenia and bipolar disorder patients.

A better understanding of the proteome profile after bipolar disorder (BD) and schizophrenia (SCZ) treatment, besides monitoring disease progression, may assist on the development of novel therapeutic strategies with the ability to reduce or control possible side effects. In this pilot study, proteomics analysis employing nano liquid chromatography coupled to mass spectrometry (nLC-MS) and bioinformatic tools were applied to identify differentially abundant proteins in serum of treated BD and SCZ patients. In total, 10 BD patients, 10 SCZ patients, and 14 healthy controls (HC) were included in this study. 24 serum proteins were significantly altered (p < 0.05) in BD and SCZ treated patients and, considering log2FC > 0.58, 8 proteins presented lower abundance in the BD group, while 7 proteins presented higher abundance and 2 lower abundance in SCZ group when compared against HC. Bioinformatics analysis based on these 24 proteins indicated two main altered pathways previously described in the literature; furthermore, it revealed that opposite abundances of the complement and coagulation cascades were the most significant biological processes involved in these pathologies. Moreover, we describe disease-related proteins and pathways associations suggesting the necessity of clinical follow-up improvement besides treatment, as a precaution or safety measure, along with the disease progression. Further biological validation and investigations are required to define whether there is a correlation between complement and coagulation cascade expression for BD and SCZ and cardiovascular diseases.

Chemical Composition of Traditional and Analog Ayahuasca.

Traditional ayahuasca can be defined as a brew made from Amazonian vine Banisteriopsis caapi and Amazonian admixture plants. Ayahuasca is used by indigenous groups in Amazonia, as a sacrament in syncretic Brazilian religions, and in healing and spiritual ceremonies internationally. The study aimed to determine concentrations of the main bio- and psychoactive components of ayahuasca used in different locations and traditions. We collected 102 samples of brews from ayahuasca-using communities. Concentrations of N,N-dimethyltryptamine (DMT), tetrahydroharmine, harmine, and harmaline were determined by ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS). Qualitative analyses for non-traditional additives (moclobemide, psilocin, yuremamine) were performed by high resolution mass spectrometry. Higher and more variable concentrations of DMT in neoshamanic ayahuasca samples compared to indigenous samples may indicate use of higher and more variable proportions of DMT-containing admixture plants. From European samples, we found two related samples of analog ayahuasca containing moclobemide, psilocin, DMT, yuremamine, and very low concentrations of B. caapi alkaloids. Some analogs of ayahuasca (Peganum harmala, Mimosa tenuiflora) were used in Europe. No analogs were found from Brazil or Santo Daime ceremonies in Europe. We recommend awareness about the constituents of the brew and ethical self-regulation among practitioners of ayahuasca ceremonies.

Periplasm-enriched fractions from Xanthomonas citri subsp. citri type A and X. fuscans subsp. aurantifolii type B present distinct proteomic profiles under in vitro pathogenicity induction.

The causative agent of Asiatic citrus canker, the Gram-negative bacterium Xanthomonas citri subsp. citri (XAC), produces more severe symptoms and attacks a larger number of citric hosts than Xanthomonas fuscans subsp. aurantifolii XauB and XauC, the causative agents of cancrosis, a milder form of the disease. Here we report a comparative proteomic analysis of periplasmic-enriched fractions of XAC and XauB in XAM-M, a pathogenicity- inducing culture medium, for identification of differential proteins. Proteins were resolved by two-dimensional electrophoresis combined with liquid chromatography-mass spectrometry. Among the 12 proteins identified from the 4 unique spots from XAC in XAM-M (p<0.05) were phosphoglucomutase (PGM), enolase, xylose isomerase (XI), transglycosylase, NAD(P)H-dependent glycerol 3-phosphate dehydrogenase, succinyl-CoA synthetase ß subunit, 6-phosphogluconate dehydrogenase, and conserved hypothetical proteins XAC0901 and XAC0223; most of them were not detected as differential for XAC when both bacteria were grown in NB medium, a pathogenicity non-inducing medium. XauB showed a very different profile from XAC in XAM-M, presenting 29 unique spots containing proteins related to a great diversity of metabolic pathways. Preponderant expression of PGM and XI in XAC was validated by Western Blot analysis in the periplasmic-enriched fractions of both bacteria. This work shows remarkable differences between the periplasmic-enriched proteomes of XAC and XauB, bacteria that cause symptoms with distinct degrees of severity during citrus infection. The results suggest that some proteins identified in XAC can have an important role in XAC pathogenicity.

New SDC function prediction based on protein-protein interaction using bioinformatics tools.

The precise roles for SDC have been complex to specify. Assigning and reanalyzing protein and peptide identification to novel protein functions is one of the most important challenges in postgenomic era. Here, we provide SDC molecular description to support, contextualize and reanalyze the corresponding protein-protein interaction (PPI). From SDC-1 data mining, we discuss the potential of bioinformatics tools to predict new biological rules of SDC. Using these methods, we have assembled new possibilities for SDC biology from PPI data, once, the understanding of biology complexity cannot be capture from one simple question.

Validation of an analytical method for the determination of the main ayahuasca active compounds and application to real ayahuasca samples from Brazil.

Ayahuasca is a brew prepared from the water decoction of two Amazonian plants, which is legally used for religious, cultural or therapeutic activities. The potential use of ayahuasca as a natural or phytotherapeutic drug is directly linked to the action of its active compounds and their connection with the therapeutic efficacy of the beverage. In this context, the aim of the present study was to establish a selective, sensitive and reproducible analytical method for the quantification of the main active ayahuasca compounds. Thirty-eight samples from the state of São Paulo, Brazil, were analyzed and the simultaneous quantifications of N,N-dimethyltryptamine (DMT), tetrahydroharmine (THH), harmine (HME) and harmaline (HML) were performed. This study enabled the development of a fast validated analytical method with minimal matrix interference and high reproducibility for the tracing of active ayahuasca compound concentrations for the first time. This method is important as an auxiliary tool for the study of active compound effects in biological responses using different multi-omic platforms.

Agrin has a pathological role in the progression of oral cancer.

BACKGROUND: The extracellular matrix modulates the hallmarks of cancer. Here we examined the role of agrin-a member of this matrix-in progression of oral squamous cell carcinoma (OSCC). METHODS: We evaluated the immunohistochemical expression of agrin in OSCC and dysplasias. Benign lesions were used as control. In subsequent experiments, we investigated whether the silencing of agrin interferes with tumour expansion both in vitro as well as in vivo. To gain insights into the role of agrin, we identified its protein network (interactome) using mass spectrometry-based proteomics and bioinformatics. Finally, we evaluated the clinical relevance of agrin interactome. RESULTS: Agrin was elevated in malignant and premalignant lesions. Further, we show that agrin silencing interferes with cancer cell motility, proliferation, invasion, colony and tumour spheroid formation, and it also reduces the phosphorylation of FAK, ERK and cyclin D1 proteins in OSCC cells. In orthotopic model, agrin silencing reduces tumour aggressiveness, like vascular and neural invasion. From a clinical perspective, agrin contextual hubs predict a poor clinical prognosis related with overall survival. CONCLUSIONS: Altogether, our results demonstrate that agrin is a histological marker for the progression of oral cancer and is a strong therapeutic target candidate for both premalignant and OSCC lesions.

Comparative proteomic analysis of Xanthomonas citri ssp. citri periplasmic proteins reveals changes in cellular envelope metabolism during in vitro pathogenicity induction.

Citrus canker is a plant disease caused by Gram-negative bacteria from the genus Xanthomonas. The most virulent species is Xanthomonas citri ssp. citri (XAC), which attacks a wide range of citrus hosts. Differential proteomic analysis of the periplasm-enriched fraction was performed for XAC cells grown in pathogenicity-inducing (XAM-M) and pathogenicity-non-inducing (nutrient broth) media using two-dimensional electrophoresis combined with liquid chromatography-tandem mass spectrometry. Amongst the 40 proteins identified, transglycosylase was detected in a highly abundant spot in XAC cells grown under inducing condition. Additional up-regulated proteins related to cellular envelope metabolism included glucose-1-phosphate thymidylyltransferase, dTDP-4-dehydrorhamnose-3,5-epimerase and peptidyl-prolyl cis-trans-isomerase. Phosphoglucomutase and superoxide dismutase proteins, known to be involved in pathogenicity in other Xanthomonas species or organisms, were also detected. Western blot and quantitative real-time polymerase chain reaction analyses for transglycosylase and superoxide dismutase confirmed that these proteins were up-regulated under inducing condition, consistent with the proteomic results. Multiple spots for the 60-kDa chaperonin and glyceraldehyde-3-phosphate dehydrogenase were identified, suggesting the presence of post-translational modifications. We propose that substantial alterations in cellular envelope metabolism occur during the XAC infectious process, which are related to several aspects, from defence against reactive oxygen species to exopolysaccharide synthesis. Our results provide new candidates for virulence-related proteins, whose abundance correlates with the induction of pathogenicity and virulence genes, such as hrpD6, hrpG, hrpB7, hpa1 and hrpX. The results present new potential targets against XAC to be investigated in further functional studies.

MicroRNA and protein profiles in invasive versus non-invasive oral tongue squamous cell carcinoma cells in vitro.

Complex molecular pathways regulate cancer invasion. This study overviewed proteins and microRNAs (miRNAs) involved in oral tongue squamous cell carcinoma (OTSCC) invasion. The human highly aggressive OTSCC cell line HSC-3 was examined in a 3D organotypic human leiomyoma model. Non-invasive and invasive cells were laser-captured and protein expression was analyzed using mass spectrometry-based proteomics and miRNA expression by microarray. In functional studies the 3D invasion assay was replicated after silencing candidate miRNAs, miR-498 and miR-940, in invasive OTSCC cell lines (HSC-3 and SCC-15). Cell migration, proliferation and viability were also studied in the silenced cells. In HSC-3 cells, 67 proteins and 53 miRNAs showed significant fold-changes between non-invasive vs. invasive cells. Pathway enrichment analyses allocated "Focal adhesion" and "ECM-receptor interaction" as most important for invasion. Significantly, in HSC-3 cells, miR-498 silencing decreased the invasion area and miR-940 silencing reduced invasion area and depth. Viability, proliferation and migration weren't significantly affected. In SCC-15 cells, down-regulation of miR-498 significantly reduced invasion and migration. This study shows HSC-3 specific miRNA and protein expression in invasion, and suggests that miR-498 and miR-940 affect invasion in vitro, the process being more influenced by mir-940 silencing in aggressive HSC-3 cells than in the less invasive SCC-15.

Neoplastic extracellular matrix environment promotes cancer invasion in vitro.

The invasion of carcinoma cells is a crucial feature in carcinogenesis. The penetration efficiency not only depends on the cancer cells, but also on the composition of the tumor microenvironment. Our group has developed a 3D invasion assay based on human uterine leiomyoma tissue. Here we tested whether human, porcine, mouse or rat hearts as well as porcine tongue tissues could be similarly used to study carcinoma cell invasion in vitro. Three invasive human oral tongue squamous cell carcinoma (HSC-3, SCC-25 and SCC-15), melanoma (G-361) and ductal breast adenocarcinoma (MDA-MB-231) cell lines, and co-cultures of HSC-3 and carcinoma-associated or normal oral fibroblasts were assayed. Myoma tissue, both native and lyophilized, promoted invasion and growth of the cancer cells. However, the healthy heart or tongue matrices were unable to induce the invasion of any type of cancer cells tested. Moreover, when studied in more detail, small molecular weight fragments derived from heart tissue rinsing media inhibited HSC-3 horizontal migration. Proteome analysis of myoma rinsing media, on the other hand, revealed migration enhancing factors. These results highlight the important role of matrix composition for cancer invasion studies in vitro and further demonstrate the unique properties of human myoma organotypic model.