RESUMO
BACKGROUND: The presence of a wide range of bioactive organic pollutants in wastewater and municipal water sources is raising concerns about their potential effects on humans. Not surprisingly, various approaches are being explored that can efficiently degrade these persistent organic pollutants. Use of peroxidases has recently been recognized as a novel remediation approach that may have potential advantages over conventional degradation techniques. However, testing the abilities of different peroxidases to degrade diverse emerging pollutants is tedious and cumbersome. RESULTS: In the present study, we present a rapid and robust approach to easily test the degradability of 21 different emerging pollutants by five different peroxidases (soybean peroxidase, chloroperoxidase, lactoperoxidase, manganese peroxidase, and horseradish peroxidase) using an LC-MSMS approach. Furthermore, this approach was also used to examine the role of a redox mediator in these enzymatic degradation assays. Our results show that some of the organic pollutants can be easily degraded by all five of the peroxidases tested, whereas others are only degraded by a specific peroxidase (or when a redox mediator was present) and there are some that are completely resistant to degradation by any of the peroxidases tested (even in the presence of a redox mediator). The degradation of furosemide and trimethoprim by soybean peroxidase and chloroperoxidase, respectively, was investigated in detail by examining the transformation products generated during their degradation. Some of the products generated during enzymatic breakdown of these pollutants have been previously reported by others, however, we report many new transformation products. CONCLUSIONS: LC-MSMS approaches, like the one described here, can be used to rapidly evaluate the potential of different peroxidases (and redox requirements) to be used as bioremediation agents. Our preliminary result shows peroxidases hold tremendous potential for being used in a final wastewater treatment step.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Espectrometria de Massas/métodos , Peroxidases/química , Poluentes Químicos da Água/química , Biocatálise , Furosemida/química , Trimetoprima/químicaRESUMO
Water contamination by various recalcitrant organic aromatic compounds is an emerging environmental issue that is increasingly attracting the attention of environmental scientists. A great majority of these recalcitrant pollutants are industrial wastes, textile dyes, pharmaceuticals, hormones, and personal care products that are discharged into wastewater. Not surprisingly, various chemical, physical, and biological strategies have been proposed and developed to remove and/or degrade these pollutants from contaminated water bodies. Biological approaches, specifically using oxidoreductase enzymes (such as peroxidases and laccases) for pollutant degradation are a relatively new and a promising research area that has potential advantages over other methods due to their higher efficiency and the ease of handling. This review focuses on the application of different classes of oxidoreductase enzymes to degrade various classes of organic pollutants. In addition to classifying these enzymes based on structural differences, the major factors that can affect their remediation ability, such as the class of peroxidases employed, pH, molecular structure of the pollutant, temperature, and the presence of redox mediators are also examined and discussed. Interestingly, a literature survey combined with our unpublished data suggests that "peroxidases" are a very heterogeneous and diverse family of enzymes and have different pH profiles, temperature optima, thermal stabilities, requirements for redox mediators, and substrate specificities as well as varying detoxification abilities. Additionally, remediation of real-life polluted samples by oxidoreductases is also highlighted as well as a critical look at current challenges and future perspectives.
Assuntos
Biodegradação Ambiental , Peroxidases , Poluentes Químicos da Água , Proteínas de Bactérias , Corantes/isolamento & purificação , Corantes/metabolismo , Proteínas Fúngicas , Peroxidases/química , Peroxidases/metabolismo , Proteínas de Plantas , Medicamentos sob Prescrição/isolamento & purificação , Medicamentos sob Prescrição/metabolismo , Poluentes Químicos da Água/isolamento & purificação , Poluentes Químicos da Água/metabolismoRESUMO
BACKGROUND: Here, we determined in vitro antioxidant activity, total phenols and flavonoids and evaluated antiproliferative activity of three medicinal plant extracts: Trigonella foenum-graecum (Fenugreek), Cassia acutifolia (Senna) and Rhazya stricta (Harmal). METHODS: The leaves of the three medicinal plants were extracted with 70% ethanol. Antioxidant activities of the extracts were determined by using DPPH (1,1-diphenyl-2-picrylhydrazyl) assay. Total flavonoid and phenolic contents were determined using colorimetric assays. MTT assay was used to estimate the antiproliferative activities of the extracts against human hepatoma (HepG2) cancer cell line. In addition, the effects of R. stricta extract on cell cycle, colony formation, and wound healing of HepG2 cells and tube formation of HUVEC cells were assessed. RESULTS: Percentage inhibition of DPPH scavenging activity were dose-dependent and ranged between (89.9% ± 0.51) and (28.6% ± 2.07). Phenolic contents ranged between (11.5 ± 0.013) and (9.7 ± 0.008) mg GAE/g while flavonoid content ranged between (20.8 ± 0.40) and (0.12 ± 0.0.01) mg QE/g. Antiproliferative results of the extracts were found to be consistent with their antioxidant activity. Among the extracts evaluated, that of R. stricta showed the best antioxidant, antiproliferative and antimetastatic activities at low concentration. It also inhibited the colony-formation capacity of HepG2 cells and exhibited antiangiogenic activity. Cell cycle analysis showed significant arrest of cells at G2/M phase 12 and 48 h after treatment and significant arrest at G1/S phase after 24 h of treatment. Consistent data were observed in western blot analysis of protein levels of Cdc2 and its cyclin partners. CONCLUSIONS: These findings introduce R. stricta as a potentially useful anti-metastatic agent and a novel potential anti-tumour agent for hepatocellular carcinoma (HCC) treatment.
Assuntos
Antineoplásicos , Antioxidantes , Fabaceae/química , Extratos Vegetais , Antineoplásicos/química , Antineoplásicos/farmacologia , Antioxidantes/química , Antioxidantes/farmacologia , Compostos de Bifenilo/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Flavonoides/análise , Células Hep G2 , Células Endoteliais da Veia Umbilical Humana , Humanos , Medicina Tradicional , Fenóis/análise , Picratos/metabolismo , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Folhas de Planta/química , Plantas Medicinais/químicaRESUMO
One of the hallmarks of malignant cell populations is the ability to undergo continuous proliferation. This property allows clonal lineages to acquire sequential aberrations that can fuel increasingly autonomous growth, invasiveness, and therapeutic resistance. Innate cellular mechanisms have evolved to regulate replicative potential as a hedge against malignant progression. When activated in the absence of normal terminal differentiation cues, these mechanisms can result in a state of persistent cytostasis. This state, termed "senescence," can be triggered by intrinsic cellular processes such as telomere dysfunction and oncogene expression, and by exogenous factors such as DNA damaging agents or oxidative environments. Despite differences in upstream signaling, senescence often involves convergent interdependent activation of tumor suppressors p53 and p16/pRB, but can be induced, albeit with reduced sensitivity, when these suppressors are compromised. Doses of conventional genotoxic drugs required to achieve cancer cell senescence are often much lower than doses required to achieve outright cell death. Additional therapies, such as those targeting cyclin dependent kinases or components of the PI3K signaling pathway, may induce senescence specifically in cancer cells by circumventing defects in tumor suppressor pathways or exploiting cancer cells' heightened requirements for telomerase. Such treatments sufficient to induce cancer cell senescence could provide increased patient survival with fewer and less severe side effects than conventional cytotoxic regimens. This positive aspect is countered by important caveats regarding senescence reversibility, genomic instability, and paracrine effects that may increase heterogeneity and adaptive resistance of surviving cancer cells. Nevertheless, agents that effectively disrupt replicative immortality will likely be valuable components of new combinatorial approaches to cancer therapy.
Assuntos
Proliferação de Células/genética , Senescência Celular/genética , Terapia de Alvo Molecular , Neoplasias/tratamento farmacológico , Neoplasias/genética , Antineoplásicos/uso terapêutico , Proliferação de Células/efeitos dos fármacos , Transformação Celular Neoplásica/genética , Instabilidade Genômica/efeitos dos fármacos , Humanos , Neoplasias/patologia , Fosfatidilinositol 3-Quinases/genética , Transdução de Sinais/genética , Telomerase/efeitos dos fármacos , Telomerase/genética , Proteína Supressora de Tumor p53/genéticaRESUMO
Deregulation of angiogenesis--the growth of new blood vessels from an existing vasculature--is a main driving force in many severe human diseases including cancer. As such, tumor angiogenesis is important for delivering oxygen and nutrients to growing tumors, and therefore considered an essential pathologic feature of cancer, while also playing a key role in enabling other aspects of tumor pathology such as metabolic deregulation and tumor dissemination/metastasis. Recently, inhibition of tumor angiogenesis has become a clinical anti-cancer strategy in line with chemotherapy, radiotherapy and surgery, which underscore the critical importance of the angiogenic switch during early tumor development. Unfortunately the clinically approved anti-angiogenic drugs in use today are only effective in a subset of the patients, and many who initially respond develop resistance over time. Also, some of the anti-angiogenic drugs are toxic and it would be of great importance to identify alternative compounds, which could overcome these drawbacks and limitations of the currently available therapy. Finding "the most important target" may, however, prove a very challenging approach as the tumor environment is highly diverse, consisting of many different cell types, all of which may contribute to tumor angiogenesis. Furthermore, the tumor cells themselves are genetically unstable, leading to a progressive increase in the number of different angiogenic factors produced as the cancer progresses to advanced stages. As an alternative approach to targeted therapy, options to broadly interfere with angiogenic signals by a mixture of non-toxic natural compound with pleiotropic actions were viewed by this team as an opportunity to develop a complementary anti-angiogenesis treatment option. As a part of the "Halifax Project" within the "Getting to know cancer" framework, we have here, based on a thorough review of the literature, identified 10 important aspects of tumor angiogenesis and the pathological tumor vasculature which would be well suited as targets for anti-angiogenic therapy: (1) endothelial cell migration/tip cell formation, (2) structural abnormalities of tumor vessels, (3) hypoxia, (4) lymphangiogenesis, (5) elevated interstitial fluid pressure, (6) poor perfusion, (7) disrupted circadian rhythms, (8) tumor promoting inflammation, (9) tumor promoting fibroblasts and (10) tumor cell metabolism/acidosis. Following this analysis, we scrutinized the available literature on broadly acting anti-angiogenic natural products, with a focus on finding qualitative information on phytochemicals which could inhibit these targets and came up with 10 prototypical phytochemical compounds: (1) oleanolic acid, (2) tripterine, (3) silibinin, (4) curcumin, (5) epigallocatechin-gallate, (6) kaempferol, (7) melatonin, (8) enterolactone, (9) withaferin A and (10) resveratrol. We suggest that these plant-derived compounds could be combined to constitute a broader acting and more effective inhibitory cocktail at doses that would not be likely to cause excessive toxicity. All the targets and phytochemical approaches were further cross-validated against their effects on other essential tumorigenic pathways (based on the "hallmarks" of cancer) in order to discover possible synergies or potentially harmful interactions, and were found to generally also have positive involvement in/effects on these other aspects of tumor biology. The aim is that this discussion could lead to the selection of combinations of such anti-angiogenic compounds which could be used in potent anti-tumor cocktails, for enhanced therapeutic efficacy, reduced toxicity and circumvention of single-agent anti-angiogenic resistance, as well as for possible use in primary or secondary cancer prevention strategies.
Assuntos
Inibidores da Angiogênese/uso terapêutico , Antineoplásicos Fitogênicos/uso terapêutico , Neoplasias/terapia , Neovascularização Patológica/terapia , Vasos Sanguíneos/efeitos dos fármacos , Vasos Sanguíneos/crescimento & desenvolvimento , Vasos Sanguíneos/patologia , Proliferação de Células/efeitos dos fármacos , Humanos , Imunoterapia , Neoplasias/prevenção & controle , Neovascularização Patológica/prevenção & controleRESUMO
Genomic instability can initiate cancer, augment progression, and influence the overall prognosis of the affected patient. Genomic instability arises from many different pathways, such as telomere damage, centrosome amplification, epigenetic modifications, and DNA damage from endogenous and exogenous sources, and can be perpetuating, or limiting, through the induction of mutations or aneuploidy, both enabling and catastrophic. Many cancer treatments induce DNA damage to impair cell division on a global scale but it is accepted that personalized treatments, those that are tailored to the particular patient and type of cancer, must also be developed. In this review, we detail the mechanisms from which genomic instability arises and can lead to cancer, as well as treatments and measures that prevent genomic instability or take advantage of the cellular defects caused by genomic instability. In particular, we identify and discuss five priority targets against genomic instability: (1) prevention of DNA damage; (2) enhancement of DNA repair; (3) targeting deficient DNA repair; (4) impairing centrosome clustering; and, (5) inhibition of telomerase activity. Moreover, we highlight vitamin D and B, selenium, carotenoids, PARP inhibitors, resveratrol, and isothiocyanates as priority approaches against genomic instability. The prioritized target sites and approaches were cross validated to identify potential synergistic effects on a number of important areas of cancer biology.
Assuntos
Instabilidade Genômica/efeitos dos fármacos , Neoplasias/dietoterapia , Neoplasias/genética , Centrossomo/metabolismo , Dano ao DNA/genética , Reparo do DNA/genética , Dieta , Instabilidade Genômica/genética , Humanos , Neoplasias/patologia , Prognóstico , Telomerase/antagonistas & inibidores , Telomerase/genéticaRESUMO
The evasion of anti-growth signaling is an important characteristic of cancer cells. In order to continue to proliferate, cancer cells must somehow uncouple themselves from the many signals that exist to slow down cell growth. Here, we define the anti-growth signaling process, and review several important pathways involved in growth signaling: p53, phosphatase and tensin homolog (PTEN), retinoblastoma protein (Rb), Hippo, growth differentiation factor 15 (GDF15), AT-rich interactive domain 1A (ARID1A), Notch, insulin-like growth factor (IGF), and Krüppel-like factor 5 (KLF5) pathways. Aberrations in these processes in cancer cells involve mutations and thus the suppression of genes that prevent growth, as well as mutation and activation of genes involved in driving cell growth. Using these pathways as examples, we prioritize molecular targets that might be leveraged to promote anti-growth signaling in cancer cells. Interestingly, naturally occurring phytochemicals found in human diets (either singly or as mixtures) may promote anti-growth signaling, and do so without the potentially adverse effects associated with synthetic chemicals. We review examples of naturally occurring phytochemicals that may be applied to prevent cancer by antagonizing growth signaling, and propose one phytochemical for each pathway. These are: epigallocatechin-3-gallate (EGCG) for the Rb pathway, luteolin for p53, curcumin for PTEN, porphyrins for Hippo, genistein for GDF15, resveratrol for ARID1A, withaferin A for Notch and diguelin for the IGF1-receptor pathway. The coordination of anti-growth signaling and natural compound studies will provide insight into the future application of these compounds in the clinical setting.
Assuntos
Carcinogênese/genética , Proliferação de Células/genética , Neoplasias/genética , Neoplasias/terapia , Transdução de Sinais , Proteínas de Ligação a DNA , Fator 15 de Diferenciação de Crescimento/genética , Via de Sinalização Hippo , Humanos , Fatores de Transcrição Kruppel-Like/genética , Terapia de Alvo Molecular , Proteínas Nucleares/genética , PTEN Fosfo-Hidrolase/genética , Proteínas Serina-Treonina Quinases/genética , Proteína do Retinoblastoma/genética , Somatomedinas/genética , Fatores de Transcrição/genética , Proteína Supressora de Tumor p53/genéticaRESUMO
Cancers harbor significant genetic heterogeneity and patterns of relapse following many therapies are due to evolved resistance to treatment. While efforts have been made to combine targeted therapies, significant levels of toxicity have stymied efforts to effectively treat cancer with multi-drug combinations using currently approved therapeutics. We discuss the relationship between tumor-promoting inflammation and cancer as part of a larger effort to develop a broad-spectrum therapeutic approach aimed at a wide range of targets to address this heterogeneity. Specifically, macrophage migration inhibitory factor, cyclooxygenase-2, transcription factor nuclear factor-κB, tumor necrosis factor alpha, inducible nitric oxide synthase, protein kinase B, and CXC chemokines are reviewed as important antiinflammatory targets while curcumin, resveratrol, epigallocatechin gallate, genistein, lycopene, and anthocyanins are reviewed as low-cost, low toxicity means by which these targets might all be reached simultaneously. Future translational work will need to assess the resulting synergies of rationally designed antiinflammatory mixtures (employing low-toxicity constituents), and then combine this with similar approaches targeting the most important pathways across the range of cancer hallmark phenotypes.
Assuntos
Antineoplásicos/uso terapêutico , Inflamação/tratamento farmacológico , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Transformação Celular Neoplásica/efeitos dos fármacos , Heterogeneidade Genética/efeitos dos fármacos , Humanos , Inflamação/genética , Inflamação/patologia , Terapia de Alvo Molecular , Neoplasias/genética , Neoplasias/patologia , Transdução de Sinais/efeitos dos fármacosRESUMO
Cancer immune evasion is a major stumbling block in designing effective anticancer therapeutic strategies. Although considerable progress has been made in understanding how cancers evade destructive immunity, measures to counteract tumor escape have not kept pace. There are a number of factors that contribute to tumor persistence despite having a normal host immune system. Immune editing is one of the key aspects why tumors evade surveillance causing the tumors to lie dormant in patients for years through "equilibrium" and "senescence" before re-emerging. In addition, tumors exploit several immunological processes such as targeting the regulatory T cell function or their secretions, antigen presentation, modifying the production of immune suppressive mediators, tolerance and immune deviation. Besides these, tumor heterogeneity and metastasis also play a critical role in tumor growth. A number of potential targets like promoting Th1, NK cell, γδ T cell responses, inhibiting Treg functionality, induction of IL-12, use of drugs including phytochemicals have been designed to counter tumor progression with much success. Some natural agents and phytochemicals merit further study. For example, use of certain key polysaccharide components from mushrooms and plants have shown to possess therapeutic impact on tumor-imposed genetic instability, anti-growth signaling, replicative immortality, dysregulated metabolism etc. In this review, we will discuss the advances made toward understanding the basis of cancer immune evasion and summarize the efficacy of various therapeutic measures and targets that have been developed or are being investigated to enhance tumor rejection.
Assuntos
Carcinogênese/imunologia , Evasão da Resposta Imune , Neoplasias/imunologia , Neoplasias/terapia , Apresentação de Antígeno/imunologia , Carcinogênese/efeitos dos fármacos , Humanos , Tolerância Imunológica/efeitos dos fármacos , Tolerância Imunológica/imunologia , Neoplasias/patologia , Compostos Fitoquímicos/uso terapêutico , Linfócitos T Reguladores/imunologia , Evasão Tumoral/efeitos dos fármacos , Evasão Tumoral/imunologiaRESUMO
Proliferation is an important part of cancer development and progression. This is manifest by altered expression and/or activity of cell cycle related proteins. Constitutive activation of many signal transduction pathways also stimulates cell growth. Early steps in tumor development are associated with a fibrogenic response and the development of a hypoxic environment which favors the survival and proliferation of cancer stem cells. Part of the survival strategy of cancer stem cells may manifested by alterations in cell metabolism. Once tumors appear, growth and metastasis may be supported by overproduction of appropriate hormones (in hormonally dependent cancers), by promoting angiogenesis, by undergoing epithelial to mesenchymal transition, by triggering autophagy, and by taking cues from surrounding stromal cells. A number of natural compounds (e.g., curcumin, resveratrol, indole-3-carbinol, brassinin, sulforaphane, epigallocatechin-3-gallate, genistein, ellagitannins, lycopene and quercetin) have been found to inhibit one or more pathways that contribute to proliferation (e.g., hypoxia inducible factor 1, nuclear factor kappa B, phosphoinositide 3 kinase/Akt, insulin-like growth factor receptor 1, Wnt, cell cycle associated proteins, as well as androgen and estrogen receptor signaling). These data, in combination with bioinformatics analyses, will be very important for identifying signaling pathways and molecular targets that may provide early diagnostic markers and/or critical targets for the development of new drugs or drug combinations that block tumor formation and progression.
Assuntos
Proteínas de Ciclo Celular/genética , Proliferação de Células/efeitos dos fármacos , Neoplasias/patologia , Neoplasias/terapia , Antineoplásicos/uso terapêutico , Proteínas de Ciclo Celular/biossíntese , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Humanos , Terapia de Alvo Molecular , Neoplasias/genética , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Transdução de Sinais/efeitos dos fármacosRESUMO
Apoptosis or programmed cell death is natural way of removing aged cells from the body. Most of the anti-cancer therapies trigger apoptosis induction and related cell death networks to eliminate malignant cells. However, in cancer, de-regulated apoptotic signaling, particularly the activation of an anti-apoptotic systems, allows cancer cells to escape this program leading to uncontrolled proliferation resulting in tumor survival, therapeutic resistance and recurrence of cancer. This resistance is a complicated phenomenon that emanates from the interactions of various molecules and signaling pathways. In this comprehensive review we discuss the various factors contributing to apoptosis resistance in cancers. The key resistance targets that are discussed include (1) Bcl-2 and Mcl-1 proteins; (2) autophagy processes; (3) necrosis and necroptosis; (4) heat shock protein signaling; (5) the proteasome pathway; (6) epigenetic mechanisms; and (7) aberrant nuclear export signaling. The shortcomings of current therapeutic modalities are highlighted and a broad spectrum strategy using approaches including (a) gossypol; (b) epigallocatechin-3-gallate; (c) UMI-77 (d) triptolide and (e) selinexor that can be used to overcome cell death resistance is presented. This review provides a roadmap for the design of successful anti-cancer strategies that overcome resistance to apoptosis for better therapeutic outcome in patients with cancer.
Assuntos
Apoptose/genética , Terapia de Alvo Molecular , Neoplasias/tratamento farmacológico , Neoplasias/genética , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Autofagia/genética , Proliferação de Células/genética , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Neoplasias/patologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
Targeted therapies and the consequent adoption of "personalized" oncology have achieved notable successes in some cancers; however, significant problems remain with this approach. Many targeted therapies are highly toxic, costs are extremely high, and most patients experience relapse after a few disease-free months. Relapses arise from genetic heterogeneity in tumors, which harbor therapy-resistant immortalized cells that have adopted alternate and compensatory pathways (i.e., pathways that are not reliant upon the same mechanisms as those which have been targeted). To address these limitations, an international task force of 180 scientists was assembled to explore the concept of a low-toxicity "broad-spectrum" therapeutic approach that could simultaneously target many key pathways and mechanisms. Using cancer hallmark phenotypes and the tumor microenvironment to account for the various aspects of relevant cancer biology, interdisciplinary teams reviewed each hallmark area and nominated a wide range of high-priority targets (74 in total) that could be modified to improve patient outcomes. For these targets, corresponding low-toxicity therapeutic approaches were then suggested, many of which were phytochemicals. Proposed actions on each target and all of the approaches were further reviewed for known effects on other hallmark areas and the tumor microenvironment. Potential contrary or procarcinogenic effects were found for 3.9% of the relationships between targets and hallmarks, and mixed evidence of complementary and contrary relationships was found for 7.1%. Approximately 67% of the relationships revealed potentially complementary effects, and the remainder had no known relationship. Among the approaches, 1.1% had contrary, 2.8% had mixed and 62.1% had complementary relationships. These results suggest that a broad-spectrum approach should be feasible from a safety standpoint. This novel approach has potential to be relatively inexpensive, it should help us address stages and types of cancer that lack conventional treatment, and it may reduce relapse risks. A proposed agenda for future research is offered.
Assuntos
Heterogeneidade Genética , Terapia de Alvo Molecular , Neoplasias/terapia , Medicina de Precisão , Antineoplásicos Fitogênicos/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Neoplasias/genética , Neoplasias/patologia , Neoplasias/prevenção & controle , Transdução de Sinais , Microambiente Tumoral/genéticaRESUMO
BACKGROUND: Peroxidases are emerging as an important class of enzymes that can be used for the efficient degradation of organic pollutants. However, detailed studies identifying the various intermediates produced and the mechanisms involved in the enzyme-mediated pollutant degradation are not widely published. RESULTS: In the present study, the enzymatic degradation of an azo dye (Crystal Ponceau 6R, CP6R) was studied using commercially available soybean peroxidase (SBP) enzyme. Several operational parameters affecting the enzymatic degradation of dye were evaluated and optimized, such as initial dye concentration, H2O2 dosage, mediator amount and pH of the solution. Under optimized conditions, 40 ppm dye solution could be completely degraded in under one minute by SBP in the presence of H2O2 and a redox mediator. Dye degradation was also confirmed using HPLC and TOC analyses, which showed that most of the dye was being mineralized to CO2 in the process. CONCLUSIONS: Detailed analysis of metabolites, based on LC/MS results, showed that the enzyme-based degradation of the CP6R dye proceeded in two different reaction pathways- via symmetric azo bond cleavage as well as asymmetric azo bond breakage in the dye molecule. In addition, various critical transformative and oxidative steps such as deamination, desulfonation, keto-oxidation are explained on an electronic level. Furthermore, LC/MS/MS analyses confirmed that the end products in both pathways were small chain aliphatic carboxylic acids.
Assuntos
Compostos Azo/metabolismo , Corantes/metabolismo , Glycine max/enzimologia , Peroxidase/metabolismo , Compostos Azo/análise , Compostos Azo/química , Dióxido de Carbono/química , Dióxido de Carbono/metabolismo , Cromatografia Líquida de Alta Pressão , Corantes/análise , Corantes/química , Peróxido de Hidrogênio/química , Concentração de Íons de Hidrogênio , Espectrometria de Massas , OxirreduçãoRESUMO
UNLABELLED: Saffron has been proposed as a promising candidate for cancer chemoprevention. The purpose of this investigation was to investigate the chemopreventive action and the possible mechanisms of saffron against diethylnitrosamine (DEN)-induced liver cancer in rats. Administration of saffron at doses of 75, 150, and 300 mg/kg/day was started 2 weeks prior to the DEN injection and was continued for 22 weeks. Saffron significantly reduced the DEN-induced increase in the number and the incidence of hepatic dyschromatic nodules. Saffron also decreased the number and the area of placental glutathione S-transferase-positive foci in livers of DEN-treated rats. Furthermore, saffron counteracted DEN-induced oxidative stress in rats as assessed by restoration of superoxide dismutase, catalase, and glutathione-S-transferase levels and diminishing of myeloperoxidase activity, malondialdehyde and protein carbonyl formation in liver. The results of immunohistochemical staining of rat liver showed that saffron inhibited the DEN-mediated elevations in numbers of cells positive for Ki-67, cyclooxygenase 2, inducible nitric oxide synthase, nuclear factor-kappa B p-65, and phosphorylated tumor necrosis factor receptor. Saffron also blocked the depletion in the number of cells positive for TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling) and M30 CytoDeath in liver tissues of DEN-treated rats. In vitro experiments carried out using HepG2 cells also confirmed these findings and showed inhibition of nuclear factor-kappa B activation, increased cleavage of caspase-3, as well as DNA damage and cell cycle arrest upon saffron treatment. CONCLUSION: This study provides evidence that saffron exerts a significant chemopreventive effect against liver cancer through inhibition of cell proliferation and induction of apoptosis. This report also shows some evidence that saffron protects rat liver from cancer via modulating oxidative damage and suppressing inflammatory response.
Assuntos
Neoplasias Hepáticas Experimentais/prevenção & controle , Fitoterapia , Extratos Vegetais/uso terapêutico , Animais , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Crocus , Ciclo-Oxigenase 2/análise , Dietilnitrosamina , Glutationa Transferase/análise , Células Hep G2 , Humanos , Fígado/efeitos dos fármacos , Neoplasias Hepáticas Experimentais/patologia , Óxido Nítrico Sintase Tipo II/análise , Extratos Vegetais/farmacologia , Ratos , Fator de Necrose Tumoral alfa/antagonistas & inibidoresRESUMO
In the recent past, numerous new types of nanostructured carriers, as support matrices, have been engineered to advance the traditional enzyme immobilization strategies. The current research aimed to develop a robust enzyme-based biocatalytic platform and its effective deployment in the industrial biotechnology sectors at large and catalysis area, in particular, as low-cost biocatalytic systems. Suitable coordination between the target enzyme molecules and surface pendent multifunctional entities of nanostructured carriers has led an effective and significant contribution in myriad novel industrial, biotechnological, and biomedical applications. As compared to the immobilization on planar two-dimensional (2-D) surface, the unique physicochemical, structural and functional attributes of nano-engineered matrices, such as high surface-to-volume ratio, surface area, robust chemical and mechanical stability, surface pendant functional groups, outstanding optical, thermal, and electrical characteristics, resulted in the concentration of the immobilized entity being substantially higher, which is highly requisite from applied bio-catalysis perspective. Besides inherited features, nanostructured materials-based enzyme immobilization aided additional features, such as (1) ease in the preparation or green synthesis route, (2) no or minimal use of surfactants and harsh reagents, (3) homogeneous and well-defined core-shell nanostructures with thick enzyme shell, and (4) nano-size can be conveniently tailored within utility limits, as compared to the conventional enzyme immobilization. Moreover, the growing catalytic needs can be fulfilled by multi-enzymes co-immobilization on these nanostructured materials-based support matrices. This review spotlights the unique structural and functional attributes of several nanostructured materials, including carbon nanotubes, graphene, and its derivate constructs, nanoparticles, nanoflowers, and metal-organic frameworks as robust matrices for laccase immobilization. The later half of the review focuses on the applied perspective of immobilized laccases for the degradation of emergent contaminants, biosensing cues, and lignin deconstruction and high-value products.
Assuntos
Biocatálise , Enzimas Imobilizadas/química , Lacase/química , Nanopartículas/química , Nanotecnologia/métodos , Enzimas Imobilizadas/metabolismo , Lacase/metabolismo , Estruturas Metalorgânicas/químicaRESUMO
Various organic pollutants so-called emerging pollutants (EPs), including active residues from pharmaceuticals, pesticides, surfactants, hormones, and personal care products, are increasingly being detected in numerous environmental matrices including water. The persistence of these EPs can cause adverse ecological and human health effects even at very small concentrations in the range of micrograms per liter or lower, hence called micropollutants (MPs). The existence of EPs/MPs tends to be challenging to mitigate from the environment effectively. Unfortunately, most of them are not removed during the present-day treatment plants. So far, a range of treatment processes and degradation methods have been introduced and deployed against various EPs and/or MPs, such as ultrafiltration, nanofiltration, advanced oxidation processes (AOPs) and enzyme-based treatments coupled with membrane filtrations. To further strengthen the treatment processes and to overcome the EPs/MPs effective removal dilemma, numerous studies have revealed the applicability and notable biocatalytic potentialities of laccases and peroxidases to degrade different classes of organic pollutants. Exquisite selectivity and unique catalytic properties make these enzymes powerful biocatalytic candidates for bio-transforming an array of toxic contaminants to harmless entities. This review focuses on the use of laccases and peroxidases, such as soybean peroxidase (SBP), horseradish peroxidase (HRP), lignin peroxidase (LiP), manganese peroxidase (MnP), and chloroperoxidase (CPO) as a greener oxidation route towards efficient and effective removal or degradation of EPs/MPs.
Assuntos
Poluentes Ambientais/análise , Biocatálise , Biodegradação Ambiental , Humanos , Lacase , Peroxidases , PraguicidasRESUMO
Accurate, sensitive, and rapid screening of performance-enhancing drugs, including glucocorticoids, is critical to combat doping in animal racing sports. Samples of urine, saliva, and serum are usually used for random screening of controlled substances; however, they tend to provide only acute usage and doping information. Hair testing has the potential to detect long-term drug use in racing animals. In the present study, commercially available ELISA-kits were used to rapidly screen and analyze various natural and synthetic glucocorticoids in the hair of camels. The four glucocorticoids that were screened using the competitive ELISA were hydrocortisone, dexamethasone, flumethasone, and methylprednisolone. However, comparison of the results obtained using the ELISA method with our previously published LC-MS/MS assay did not show a good correlation. The results obtained with the ELISA analysis of hair samples of 27 different camels showed that this technique consistently yielded significantly higher levels of glucocorticoids compared with the LC-MS/MS assay. This is an interesting finding and suggests that commercially available ELISA tests may overestimate the amount of glucocorticoids present in camel hair, perhaps due to specificity and cross-reactivity issues.
Assuntos
Pelo Animal/química , Camelus , Glucocorticoides/análise , Cabelo/química , Drogas Ilícitas/análise , Detecção do Abuso de Substâncias/métodos , Animais , Camelus/metabolismo , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Espectrometria de Massas em Tandem/métodosRESUMO
Nanostructured materials constitute an interesting and novel class of support matrices for the immobilization of peroxidase enzymes. Owing to the high surface area, robust mechanical stability, outstanding optical, thermal, and electrical properties, nanomaterials have been rightly perceived as immobilization matrices for enzyme immobilization with applications in diverse areas such as nano-biocatalysis, biosensing, drug delivery, antimicrobial activities, solar cells, and environmental protection. Many nano-scale materials have been employed as support matrices for the immobilization of different classes of enzymes. Nanobiocatalysts, enzymes immobilized on nano-size materials, are more stable, catalytically robust, and could be reused and recycled in multiple reaction cycles. In this review, we illustrate the unique structural/functional features and potentialities of nanomaterials-immobilized peroxidase enzymes in different biotechnological applications. After a comprehensive introduction to the immobilized enzymes and nanocarriers, the first section reviewed carbonaceous nanomaterials (carbon nanotube, graphene, and its derivatives) as a host matrix to constitute robust peroxidases-based nanobiocatalytic systems. The second half covers metallic nanomaterials (metals, and metal oxides) and some other novel materials as host carriers for peroxidases immobilization. The next section vetted the potential biotechnological applications of the resulted nanomaterials-immobilized robust peroxidases-based nanobiocatalytic systems. Concluding remarks, trends, and future recommendations for nanomaterial immobilized enzymes are also given.
Assuntos
Biocatálise , Enzimas Imobilizadas , Grafite/química , Nanoestruturas , Peroxidases/química , Antibacterianos/química , Técnicas Biossensoriais , Monitoramento AmbientalRESUMO
Small amounts of highly reactive oxygen species (oxyradicals) are normal by-products of cellular metabolism. However, under certain conditions large amounts of oxyradicals are generated inside cells which may cause extensive cellular damage. Not surprisingly, a large number of disease states have been linked to oxidative stress, including cancer, diabetes, Parkinson's disease, Alzheimer's disease, and heart disease. Previously, we had shown that fluorescence spectroscopy could be used to study the pH-dependence of GFP denaturation with various agents. In this report, we show that GFP readily loses its auto-fluorescence upon exposure to oxyradicals as measured by fluorescence spectroscopy. We further show that oxyradical scavengers can prevent this loss of GFP fluorescence, thus oxyradical-induced loss of GFP fluorescence could be used to screen for antioxidants. We have evaluated various parameters which could affect the sensitivity of this GFP-based oxyradical scavenging assay, such as concentration H(2)O(2) used to produce oxyradicals, pH of the buffer, as well as UV intensity. Surprisingly we found that pH had a very dramatic effect on oxyradical-induced GFP damage. GFP was found to be most susceptible to oxyradical-induced damage at pH 6.5, and least susceptible at pH 8.5. This is the first demonstration that GFP loses its fluorescence upon exposure to oxyradicals. Furthermore, the data presented here suggest that GFP could be used to develop assays to screen for antioxidants or radical scavengers.
Assuntos
Proteínas de Fluorescência Verde/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Sequestradores de Radicais Livres/metabolismo , Concentração de Íons de Hidrogênio , Desnaturação Proteica , Espectrometria de FluorescênciaRESUMO
Vitamin D deficiency has been implicated in a plethora of diseases including rheumatoid arthritis, Parkinson's disease, Alzheimer's disease, and osteoporosis. Deficiency of this vitamin is a global epidemic affecting both developing and developed nations. Within a clinical context, the qualitative and quantitative analysis of vitamin D is therefore vital. The main metabolic markers for assessing vitamin D status in humans are the hydroxylated forms of vitamin D, 25OHD3 and 25OHD2 on account of their long half-lives within the body and excellent stability. An adequate level for healthy individuals of these hydroxylated forms is estimated to be around 20-40ng/ml of blood. There are three main analytical techniques for determining the levels of 25OHD3 and 25OHD2. The first technique is immunoassay-based and can be performed in a rapid, high throughput, automated manner, allowing as many as 240 tests per hour with the duration of each assay as little as 18min. Furthermore, it offers excellent sensitivity with a detection range of 3.4-156ng/ml. A major downside of immunoassays is that they are unable to distinguish between the various forms of vitamin D. While HPLC is a highthroughput low cost instrument it is not a very sensitive technique and cannot quantify the down stream metabolites of vitamin D. The third technique, namely liquid chromatography-mass spectrometry (LC-MS/), provides excellent sensitivity with a wide dynamic range from 0.068pg/ml to 100ng/ml. Additionally, it offers a high level of separation and permits identification of vitamin D-related metabolites. However, a huge limitation with LC/MS/MS is their poor throughput for sample analyses. As yet, there is no analytical technique which combines the fine detection capabilities of LC/MS/MS and the rapid, automated format of immunoassay, for vitamin D analyses. Future attention therefore needs to be given to this area if the current clinical diagnostic tools for vitamin D analysis are to be further improved.