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In the last decade, the incidence of obesity has increased dramatically worldwide, reaching a dangerous pandemic spread. This condition has serious public health implications as it significantly increases the risk of chronic diseases such as type 2 diabetes, fatty liver, hypertension, heart attack, and stroke. The treatment of obesity is therefore the greatest health challenge of our time. Conventional therapeutic treatment of obesity is based on the use of various synthetic molecules belonging to the class of appetite suppressants, lipase inhibitors, hormones, metabolic regulators, and inhibitors of intestinal peptide receptors. The long-term use of these molecules is generally limited by various side effects and tolerance. For this reason, the search for natural alternatives to treat obesity is a current research goal. This review therefore examined the anti-obesity potential of natural chalcones based on available evidence from in vitro and animal studies. In particular, the results of the main in vitro studies describing the principal molecular therapeutic targets and the mechanism of action of the different chalcones investigated were described. In addition, the results of the most relevant animal studies were reported. Undoubtedly, future clinical studies are urgently needed to confirm and validate the potential of natural chalcones in the clinical prophylaxis of obesity.
Assuntos
Depressores do Apetite , Chalconas , Diabetes Mellitus Tipo 2 , Animais , Chalconas/farmacologia , Chalconas/uso terapêutico , Chalconas/química , Diabetes Mellitus Tipo 2/tratamento farmacológico , Obesidade/tratamento farmacológico , Obesidade/metabolismoRESUMO
In the context of inflammation and immunity, there are fragmented and observational studies relating to the pharmacological activity of Mangifera indica L. and its main active component, mangiferin. Therefore, we aimed to analyze the potential beneficial effects of this plant extract (MIE, 90 % in mangiferin) in a mouse model of gouty arthritis, to allow the evaluation of cellular immune phenotypes and the biochemical mechanism/s beyond MIE activity. Gouty arthritis was induced by the intra-articular administration of MSU crystals (200 µg 20 µl-1), whereas MIE (0.1-10 mg kg-1) or corresponding vehicle (DMSO/saline 1:3) were orally administrated concomitantly with MSU (time 0), 6 and 12 h after the stimulus. Thereafter, knee joint score and oedema were evaluated in addition to western blot analysis for COX-2/mPGES-1 axis. Moreover, the analysis of pro/anti-inflammatory cyto-chemokines coupled with the phenotyping of the cellular infiltrate was performed. Treatment with MIE revealed a dose-dependent reduction in joint inflammatory scores with maximal inhibition observed at 10 mg kg-1. MIE significantly reduced leukocyte infiltration and activation and the expression of different pro-inflammatory cyto-chemokines in inflamed tissues. Furthermore, biochemical analysis revealed that MIE modulated COX-2/mPGES-1 and mPGDS-1/PPARγ pathways. Flow cytometry analysis also highlighted a prominent modulation of inflammatory monocytes (CD11b+/CD115+/LY6Chi), and Treg cells (CD4+/CD25+/FOXP3+) after MIE treatment. Collectively, the results of this study demonstrate a novel function of MIE to positively affect the local and systemic inflammatory/immunological perturbance in the onset and progression of gouty arthritis.
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Artrite Gotosa , Mangifera , Extratos Vegetais , Animais , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Artrite Gotosa/tratamento farmacológico , Artrite Gotosa/metabolismo , Ciclo-Oxigenase 2/metabolismo , Mangifera/química , Camundongos , Extratos Vegetais/farmacologia , Linfócitos T Reguladores , Células Th17RESUMO
A new series of in vitro potent and highly selective histone methyl transferase enzyme G9a inhibitors was obtained. In particular, compound 2a, one the most potent G9a inhibitor identified, was endowed with >130-fold selectivity over GLP and excellent ligand efficiency. Therefore, it may represent a valuable tool compound to validate the role of highly selective G9a inhibitors in different pathological conditions. When 2a was characterized in vitro in cellular models of skeletal muscle differentiation, a relevant increase of myofibers' size and reduction of the fibroadipogenic infiltration were observed, further confirming the therapeutic potential of selective G9a inhibitors for the treatment of Duchenne muscle dystrophy.
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Histona-Lisina N-Metiltransferase , Histonas , Inibidores Enzimáticos/farmacologiaRESUMO
Chronic hepatitis B (CHB) is a major worldwide public health problem and novel anti-HBV therapies preventing liver disease progression to cirrhosis and hepatocellular carcinoma are urgently needed. Over the last several years, capsid assembly modulators (CAM) have emerged as clinically effective anti-HBV agents which can inhibit HBV replication in CHB patients. As part of a drug discovery program aimed at obtaining novel CAM endowed with high in vitro and in vivo antiviral activity, we identified a novel series of sulfamoylbenzamide (SBA) derivatives. Compound 10, one of the most in vitro potent SBA-derived CAM discovered to date, showed excellent pharmacokinetics in mice suitable for oral dosing. When studied in a transgenic mouse model of hepatic HBV replication, it was considerably more potent than NVR 3-778, the first sulfamoylbenzamide (SBA) CAM that entered clinical trials for CHB, at reducing viral replication in a dose-dependent fashion. We present herein the discovery process, the SAR analysis and the pre-clinical profile of this novel SBA CAM.
Assuntos
Antivirais , Capsídeo , Animais , Antivirais/farmacocinética , Proteínas do Capsídeo , Vírus da Hepatite B , Camundongos , Montagem de Vírus , Replicação ViralRESUMO
Proanthocyanidins (PACs) are a group of bioactive molecules found in a variety of plants and foods. Their bioavailability depends on their molecular size, with monomers and dimers being more bioavailable than those that have a higher polymerization degree. This study aimed to develop a method to convert high-molecular-weight PACs to low-molecular-weight ones in a grape seed extract (GSE) from Vitis vinifera L. Therefore, GSE was subjected to alkaline treatment (ATGSE), and its difference in chemical composition, compared to GSE, was evaluated using a molecular networking (MN) approach based on results obtained from HPLC-ESI HRMS/MS characterization analysis. The network analysis mainly noted the PAC cluster with about 142 PAC compounds identified. In particular, the obtained results showed a higher content of monomeric and dimeric PACs in ATGSE compared to GSE, with 58% and 49% monomers and 31% and 24% dimers, respectively. Conversely, trimeric (9%), polymeric (4%), and galloylated PACs (14%) were more abundant in GSE than in ATGSE (6%, 1%, and 4%, respectively). Moreover, in vitro antioxidant and anti-inflammatory activities were investigated, showing the high beneficial potential of both extracts. In conclusion, ATGSE could represent an innovative natural matrix rich in bioavailable and bioaccessible PACs for nutraceutical applications with potential beneficial properties.
Assuntos
Extrato de Sementes de Uva , Proantocianidinas , Vitis , Proantocianidinas/química , Disponibilidade Biológica , Peso Molecular , Extrato de Sementes de Uva/farmacologia , Extrato de Sementes de Uva/química , Vitis/química , Sementes/química , Extratos Vegetais/químicaRESUMO
A new strategy that takes advantage of the synergism between NMR and UHPLC-HRMS yields accurate concentrations of a high number of compounds in biofluids to delineate a personalized metabolic profile (SYNHMET). Metabolite identification and quantification by this method result in a higher accuracy compared to the use of the two techniques separately, even in urine, one of the most challenging biofluids to characterize due to its complexity and variability. We quantified a total of 165 metabolites in the urine of healthy subjects, patients with chronic cystitis, and patients with bladder cancer, with a minimum number of missing values. This result was achieved without the use of analytical standards and calibration curves. A patient's personalized profile can be mapped out from the final dataset's concentrations by comparing them with known normal ranges. This detailed picture has potential applications in clinical practice to monitor a patient's health status and disease progression.
Assuntos
Metabolômica/métodos , Medicina de Precisão/métodos , Urina/química , Adulto , Idoso , Idoso de 80 Anos ou mais , Cromatografia Líquida de Alta Pressão/métodos , Cistite/metabolismo , Cistite/urina , Feminino , Humanos , Imageamento por Ressonância Magnética/métodos , Espectroscopia de Ressonância Magnética/métodos , Masculino , Metaboloma/fisiologia , Pessoa de Meia-Idade , Espectrometria de Massas em Tandem/métodos , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/urinaRESUMO
A new visible light-induced photocatalytic protocol enabling the formation of secondary amides from electron-poor organic bromides and isocyanides was developed. In addition, the in situ interception of ketenimine intermediates with nitrogen nucleophiles such as amines, hydrazines, and TMSN3 afforded, in a one-pot two-step procedure, valuable scaffolds such as ketene aminals, pyrazolones, and tetrazoles. Mechanistic evidence confirmed a radical pathway where isocyanides acted as radical geminal acceptors generating key imidoyl radical species.
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The identification of a new series of growth inhibitors of Trypanosoma cruzi, the causative agent of Chagas' disease, is described. In vitro screening of a subset of compounds from our in-house compound collection against the parasite led to the identification of hit compound 1 with low micromolar inhibition of T. cruzi growth. SAR exploration on the hit compound led to the identification of compounds that show nanomolar parasite growth inhibition (T. cruzi EC50 ≤ 100 nM) and no cytotoxicity in human cells (HeLa CC50 > 50 µM). Further investigation identified CYP51 inhibition (compound 11 CYP51 IC50 52 nM) as a possible mechanism of action of this new class of anti-parasitic agents.
Assuntos
Descoberta de Drogas , Inibidores do Crescimento/farmacologia , Piridinas/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Relação Dose-Resposta a Droga , Inibidores do Crescimento/síntese química , Inibidores do Crescimento/química , Humanos , Estrutura Molecular , Testes de Sensibilidade Parasitária , Piridinas/síntese química , Piridinas/química , Relação Estrutura-Atividade , Tripanossomicidas/síntese química , Tripanossomicidas/químicaRESUMO
A previous publication from our laboratory reported the identification of a new class of 2-(1H-imidazo-2-yl)piperazines as potent T. brucei growth inhibitors as potential treatment for Human African Trypanosomiasis (HAT). This work describes the structure-activity relationship (SAR) around the hit compound 1, which led to the identification of the optimized compound 18, a single digit nanomolar inhibitor (EC50 7 nM), not cytotoxic and with optimal in vivo profile that made it a suitable candidate for efficacy studies in a mouse model mimicking the second stage of disease.
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Inibidores do Crescimento/química , Piperazinas/química , Tripanossomicidas/química , Trypanosoma brucei brucei/efeitos dos fármacos , Tripanossomíase Africana/tratamento farmacológico , Sobrevivência Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Inibidores do Crescimento/farmacologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Isomerismo , Morfolinas/química , Piperazinas/farmacologia , Quinolinas/química , Relação Estrutura-Atividade , Tripanossomicidas/farmacologiaRESUMO
Inhibition of KEAP1-NRF2 protein-protein interaction is considered a promising strategy to selectively and effectively activate NRF2, a transcription factor which is involved in several pathologies such as Huntington's disease (HD). A library of linear peptides based on the NRF2-binding motifs was generated on the nonapeptide lead Ac-LDEETGEFL-NH2 spanning residues 76-84 of the Neh2 domain of NRF2 with the aim to replace E78, E79 and E82 with non-acidic amino acids. A deeper understanding of the features and accessibility of the T80 subpocket was also targeted by structure-based design. Approaches to improve cell permeability were investigated using both different classes of cyclic peptides and conjugation to cell-penetrating peptides. This insight will guide future design of macrocycles, peptido-mimetics and, most importantly, small neutral brain-penetrating molecules to evaluate whether NRF2 activators have utility in HD.
Assuntos
Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Peptídeos Cíclicos/química , Peptídeos/química , Sequência de Aminoácidos , Sítios de Ligação , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular/efeitos dos fármacos , Desenho de Fármacos , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/antagonistas & inibidores , Simulação de Dinâmica Molecular , Fator 2 Relacionado a NF-E2/antagonistas & inibidores , Peptídeos/metabolismo , Peptídeos/farmacologia , Peptídeos Cíclicos/metabolismo , Peptídeos Cíclicos/farmacologia , Ligação Proteica , Relação Estrutura-AtividadeRESUMO
The pandemic evolution of SARS-CoV-2 infection is forcing the scientific community to unprecedented efforts to explore all possible approaches against COVID-19. In this context, targeting virus entry is a promising antiviral strategy for controlling viral infections. The main strategies pursued to inhibit the viral entry are considering both the virus and the host factors involved in the process. Primarily, direct-acting antivirals rely on inhibition of the interaction between ACE2 and the receptor binding domain (RBD) of the Spike (S) protein or targeting the more conserved heptad repeats (HRs), involved in the membrane fusion process. The inhibition of host TMPRSS2 and cathepsins B/L may represent a complementary strategy to be investigated. In this review, we discuss the development entry inhibitors targeting the S protein, as well as the most promising host targeting strategies involving TMPRSS2 and CatB/L, which have been exploited so far against CoVs and other related viruses.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Inibidores de Serina Proteinase/farmacologia , Internalização do Vírus/efeitos dos fármacos , Animais , Betacoronavirus/metabolismo , Betacoronavirus/fisiologia , Humanos , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/metabolismoRESUMO
Huntington's disease (HD) is a neurodegenerative disease caused by polyglutamine expansion in the huntingtin protein. For drug candidates targeting HD, the ability to cross the blood-brain barrier (BBB) and reach the site of action in the central nervous system (CNS) is crucial for achieving pharmacological activity. To assess the permeability of selected compounds across the BBB, we utilized a two-dimensional model composed of primary porcine brain endothelial cells and rat astrocytes. Our objective was to use this in vitro model to rank and prioritize compounds for in vivo pharmacokinetic and brain penetration studies. The model was first characterized using a set of validation markers chosen based on their functional importance at the BBB. It was shown to fulfill the major BBB characteristics, including functional tight junctions, high transendothelial electrical resistance, expression, and activity of influx and efflux transporters. The in vitro permeability of 54 structurally diverse known compounds was determined and shown to have a good correlation with the in situ brain perfusion data in rodents. We used this model to investigate the BBB permeability of a series of new HD compounds from different chemical classes, and we found a good correlation with in vivo brain permeation, demonstrating the usefulness of the in vitro model for optimizing CNS drug properties and for guiding the selection of lead compounds in a drug discovery setting.
Assuntos
Barreira Hematoencefálica/metabolismo , Fármacos do Sistema Nervoso Central/uso terapêutico , Descoberta de Drogas/métodos , Doença de Huntington/tratamento farmacológico , Modelos Biológicos , Transportadores de Cassetes de Ligação de ATP/metabolismo , Animais , Astrócitos/metabolismo , Permeabilidade Capilar/fisiologia , Células Cultivadas , Córtex Cerebral/citologia , Técnicas de Cocultura , Impedância Elétrica , Células Endoteliais/metabolismo , Permeabilidade , Ratos , Ratos Sprague-Dawley , Proteínas Carreadoras de Solutos/metabolismo , Suínos , Junções Íntimas/metabolismoRESUMO
The identification of a new series of growth inhibitors of Trypanosoma brucei rhodesiense, causative agent of Human African Trypanosomiasis (HAT), is described. A selection of compounds from our in-house compound collection was screened in vitro against the parasite leading to the identification of compounds with nanomolar inhibition of T. brucei growth. Preliminary SAR on the hit compound led to the identification of compound 34 that shows low nanomolar parasite growth inhibition (T. brucei EC50 5â¯nM), is not cytotoxic (HeLa CC50â¯>â¯25,000â¯nM) and is selective over other parasites, such as Trypanosoma cruzi and Plasmodium falciparum (T. cruzi EC50 8120â¯nM, P. falciparum EC50 3624â¯nM).
Assuntos
Piperazinas/química , Piperazinas/farmacologia , Tripanossomicidas/química , Tripanossomicidas/farmacologia , Trypanosoma brucei brucei/efeitos dos fármacos , Tripanossomíase Africana/tratamento farmacológico , Células HeLa , Humanos , Imidazóis/química , Imidazóis/farmacologia , Malária Falciparum/tratamento farmacológico , Plasmodium falciparum/efeitos dos fármacos , Relação Estrutura-Atividade , Trypanosoma brucei brucei/crescimento & desenvolvimento , Tripanossomíase Africana/parasitologiaRESUMO
Key epitranscriptomic players have been increasingly characterized for their structural features and their involvement in several diseases. Accordingly, the design and synthesis of novel epitranscriptomic modulators have started opening a glimmer for drug discovery. m6A is a reversible modification occurring on a specific site and is catalyzed by three sets of proteins responsible for opposite functions. Writers (e.g., methyl-transferase-like protein (METTL) 3/METTL14 complex and METTL16) introduce the methyl group on adenosine N-6, by transferring the methyl group from the methyl donor S-adenosyl-methionine (SAM) to the substrate. Despite the rapidly advancing drug discovery progress on METTL3/METTL14, the METTL16 m6A writer has been marginally explored so far. We herein provide the first comprehensive overview of structural and biological features of METTL16, highlighting the state of the art in the field of its biological and structural characterization. We also showcase initial efforts in the identification of structural templates and preliminary structure-activity relationships for METTL16 modulators.
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Several medicinal herbal plants are extensively used as sources of bioactive compounds with beneficial effects on human health. This study assessed the procyanidin and polyphenol profiles together with the antioxidant potential of seven herbal medical matrices. To achieve this aim, procyanidin extraction from grape pomace was optimized and validated by monitoring monomeric-trimeric procyanidins. The proposed quantification method was applied to the seven medical herbs, and it proved to be a very efficient protocol for procyanidin-rich extracts analysis. In addition, the Paullinia cupana Kunth. seed was identified as a very rich source of procyanidins (about 5 mg/g dry matrix of each dimeric and about 3 mg/g dry matrix trimeric) with high antioxidant properties. The polyphenolic profile was assessed by HPLC-HESI-MS/MS analysis. The in vitro antioxidant activity was evaluated by DPPH assay to explore the antioxidant properties of the extracts, which were substantially higher in Peumus boldus Molina leaves extracts (935.23 ± 169 µmol of Trolox equivalent/g of dry weight) concerning the other matrices. Moreover, a high Pearson coefficient value was observed between the total flavonoid content (TFC) and DPPH in comparison with the total polyphenol content (TPC) and DPPH, indicating flavonoids as the principal bioactive with antioxidant activity in the extracts.
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Environmental contamination by heavy metals (HMs) has emerged as a significant global issue in recent decades. Among natural substances, food-deriving polyphenols have found a valuable application in chelating therapy, partially limited by their low water solubility. Thus, three different hydroalcoholic extracts titrated in quercetin (QE), ellagic acid (EA), and curcumin (CUR) were formulated using maltodextrins as carriers, achieving a powder with a valuable water solubility (MQE 91.3 ± 1.2%, MEA 93.4 ± 2.1, and MCUR 89.3 ± 2%). Overcoming the problem of water solubility, such formulations were tested in an in vitro simulated gastrointestinal digestion experiment conducted on a water sample with standardized concentrations of the principal HMs. Our results indicate that regarding the nonessential HMs investigated (Pb, Cd, As, Sb, and Hg), MQE has been shown to be the most effective in increasing the HMs' non-bioaccessible concentration, resulting in concentration increases in Cd of 68.3%, in As of 51.9%, in Hg of 58.9%, in Pb of 271.4, and in Sb of 111.2% (vs control, p < 0.001) in non-bioaccessible fractions. Regarding the essential HMs, MEA has shown the greatest capability to increase their intestinal bioaccessibility, resulting in +68.5%, +61.1, and +22.3% (vs control, p < 0.001) increases in Cu, Zn, and Fe, respectively. Finally, considering the strong relation between the antiradical and chelating activities, the radical scavenging potentials of the formulations was assayed in DPPH and ABTS assays.
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Unripe tomatoes represent an agri-food waste resulting from industrial by-processing products of tomatoes, yielding products with a high content of bioactive compounds with potential nutraceutical properties. The food-matrix biological properties are attributed to the high steroidal glycoalkaloid (SGA) content. Among them, α-tomatine is the main SGA reported in unripe green tomatoes. This review provides an overview of the main chemical and pharmacological features of α-tomatine and green tomato extracts. The extraction processes and methods employed in SGA identification and the quantification are discussed. Special attention was given to the methods used in α-tomatine qualitative and quantitative analyses, including the extraction procedures and the clean-up methods applied in the analysis of Solanum lycopersicum L. extracts. Finally, the health-beneficial properties and the pharmacokinetics and toxicological aspects of SGAs and α-tomatine-containing extracts are considered in depth. In particular, the relevant results of the main in vivo and in vitro studies reporting the therapeutic properties and the mechanisms of action were described in detail.
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Bacterial infections represent a key public health issue due to the occurrence of multidrug-resistant bacteria. Recently, the amount of data supporting the dynamic control of epigenetic pathways by environmental cues has triggered research efforts toward the clarification of their role in microbial infections. Among protein post-translational modifications, reversible acetylation is the most implicated in the feedback to environmental stimuli and in cellular homeostasis. Accordingly, the latest studies identified the histone deacetylase 6 (HDAC6) enzyme as a crucial player in the complex molecular machinery underlying bacterial clearance or killing. A very important milestone for the elucidation of the consequence of HDAC6 activity in bacterial infections is herein described, unveiling for the first time the role of a potent HDAC6 inhibitor in interfering with biofilm formation and modulating virulence factors of P. aeruginosa. We demonstrated that compound F2F-2020202 affected the production of some important virulence factors in P. aeruginosa, namely pyocyanin and rhamnolipids, clearly impairing its ability to form biofilm. Furthermore, evidence of possible QS involvement is supported by differential regulation of specific genes, namely RhlI, phAz1, and qsrO. The data herein obtained also complement and in part explain our previous results with selective HDAC6 inhibitors able to reduce inflammation and bacterial load in chronic infection models recapitulating the cystic fibrosis (CF) phenotype. This study fosters future in-depth investigation to allow the complete elucidation of the molecular mechanisms underlying HDAC6's role in bacterial infections.
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Unripe tomatoes are among the main waste produced during tomato cultivation and processing. In this study, unripe tomatoes from seven different Italian cultivars have been investigated to evaluate their nutraceutical potential. Phytochemical investigation allowed shedding light on the identification of seventy-five bioactive compounds. The highest amount of polyphenolic and glycoalkaloids along with the high level of antioxidant activities was found in the Datterini tomatoes variety. The peculiarity of this variety is the high chlorogenic acid content, being ten times higher compared to the other cultivars examined. Moreover, the total α-tomatine amount has been found substantially higher (34.699 ± 1.101 mg/g dry weight) with respect to the other tomato varieties analyzed. Furthermore, the cultivars metabolomic profiles were investigated with the PCA approach. Based on Datterini cultivar's metabolomic profile, its waste-recovery could represent a good option for further added value products in pharmaceutical and nutraceutical areas with a high α-tomatine content.
Assuntos
Antioxidantes , Solanum lycopersicum , Antioxidantes/química , Ácido Clorogênico , Compostos Fitoquímicos , Extratos Vegetais/químicaRESUMO
Gut microbiota plays a crucial role in human health homeostasis, and the result of its alteration, known as dysbiosis, leads to several pathologies (e.g., inflammatory bowel disease, metabolic syndrome, and Crohn's disease). Traditional methods used to assess dysbiosis include the dual sugar absorption test and the urinary lactulose/mannitol ratio (LMR) measurement using mass spectrometry. Despite its precision, this approach is costly and requires specialized equipment. Hence, we developed a rapid and reliable spectrofluorimetric method for measuring LMR in urine, offering a more accessible alternative. This spectrofluorimetric assay quantifies the fluorescence of nicotinamide adenine dinucleotide (NADH) and nicotinamide adenine dinucleotide phosphate (NADPH) produced during the enzymatic oxidation of mannitol and lactulose, respectively. The assay requires 100 µL of urine samples and detects LMR values lower (eubiosis) and higher (dysbiosis) than 0.05, ultimately being amenable to high-throughput screening and automatization, making it practical for clinical and research settings. A validation of the method demonstrated its high precision, accuracy, and robustness. Additionally, this study confirmed analyte stability under various storage conditions, ensuring reliable results even with delayed analysis. Overall, this spectrofluorimetric technique reduces costs, time, and the environmental impact associated with traditional mass spectrometry methods, making it a viable option for widespread use in the assessment of dysbiosis.