RESUMO
SCOPE: The effect of diabetes on the pharmacokinetics, bioavailability and brain distribution of grape polyphenols and select metabolites was studied in the Zucker diabetic fatty (ZDF) rat model. METHODS AND RESULTS: (ZDF) rats and their lean controls (LN) were dosed with a Standardized Grape Polyphenol (SGP) Mixture consisting of grape seed extract, Concord grape juice and resveratrol (RES) by oral gavage for 10 days. An 8-h pharmacokinetic study was performed. After 24 h, a second dose of SGP was administered and 1 h later animals were sacrificed and brain tissue was harvested. Plasma, urine, and brain tissue were analyzed for grape polyphenols. ZDF rats exhibited significantly diminished Cmax for all catechin, epicatechin, quercetin and resveratrol conjugated metabolites. Bioavailability was significantly lower in ZDF rats for methylated flavan-3-ol, RES, and quercetin metabolites. Significantly lower levels of metabolites of RES, quercetin, and flavan-3-ols were found in brains of ZDF rats. There was no significant difference between ZDF and LN in anthocyanins in plasma and no anthocyanins were detectable in brain extracts. ZDF rats showed significantly higher urinary excretion for all polyphenols. CONCLUSION: Diabetes may alter the overall bioavailability of some polyphenols in plasma and brain in part due to higher urinary clearance.
Assuntos
Encéfalo/efeitos dos fármacos , Diabetes Mellitus Experimental/sangue , Polifenóis/sangue , Polifenóis/farmacocinética , Vitis/química , Animais , Antocianinas/sangue , Antocianinas/farmacocinética , Antocianinas/urina , Disponibilidade Biológica , Glicemia/metabolismo , Encéfalo/metabolismo , Catequina/sangue , Catequina/farmacocinética , Catequina/urina , Diabetes Mellitus Tipo 2/sangue , Flavonoides/sangue , Flavonoides/farmacocinética , Flavonoides/urina , Extrato de Sementes de Uva/sangue , Extrato de Sementes de Uva/farmacocinética , Extrato de Sementes de Uva/urina , Masculino , Polifenóis/urina , Quercetina/sangue , Quercetina/farmacocinética , Quercetina/urina , Ratos , Ratos Zucker , Resveratrol , Estilbenos/sangue , Estilbenos/farmacocinética , Estilbenos/urina , Espectrometria de Massas em TandemRESUMO
The plant secondary metabolite and common food additive dihydrocoumarin (DHC) is an inhibitor of the Sirtuin family of NAD+-dependent deacetylases. Sirtuins are key regulators of epigenetic processes that maintain silent chromatin in yeast and have been linked to gene expression, metabolism, apoptosis, tumorogenesis and age-related processes in multiple organisms, including humans. Here we report that exposure to the polyphenol DHC led to defects in several Sirtuin-regulated processes in budding yeast including the establishment and maintenance of Sir2p-dependent silencing by causing disassembly of silent chromatin, Hst1p-dependent repression of meiotic-specific genes during the mitotic cell cycle. As both transient and prolonged exposure to environmental and dietary factors have the potential to lead to heritable alterations in epigenetic states and to modulate additional Sirtuin-dependent phenotypes, we examined the bioavailability and digestive stability of DHC using an in vivo rat model and in vitro digestive simulator. Our analyses revealed that DHC was unstable during digestion and could be converted to melilotic acid (MA), which also caused epigenetic defects, albeit less efficiently. Upon ingestion, DHC was observed primarily in intestinal tissues, but did not accumulate over time and was readily cleared from the animals. MA displayed a wider tissue distribution and, in contrast to DHC, was also detected in the blood plasma, interstitial fluid, and urine, implying that the conversion of DHC to the less bioactive compound, MA, occurred efficiently in vivo.
Assuntos
Cumarínicos/farmacologia , Dieta , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Sirtuína 2/metabolismo , Animais , Biocatálise/efeitos dos fármacos , Disponibilidade Biológica , Cumarínicos/sangue , Cumarínicos/farmacocinética , Digestão , Líquido Extracelular/metabolismo , Trato Gastrointestinal/efeitos dos fármacos , Trato Gastrointestinal/metabolismo , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Genes Reporter , Loci Gênicos , Mutação/genética , Fenóis/sangue , Fenóis/urina , Fenótipo , Ratos Sprague-Dawley , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Esporos Fúngicos/efeitos dos fármacos , Esporos Fúngicos/genética , Sus scrofaRESUMO
UNLABELLED: Research suggests that subclinical hypothyroidism (SHT) influences insulin sensitivity and glucose tolerance. Reductions in thyroid stimulating hormone (TSH) concentrations are associated with exercise training (ExTr), which improves insulin sensitivity and glucose uptake. PURPOSE: A secondary analysis of previously published data was conducted to examine the relationship between SHT, TSH and glucose homeostatic control at baseline and to assess the impact of ExTr on thyroid status and how SHT affects changes in insulin sensitivity after ExTr. MATERIALS AND METHODS: Data were obtained from a 36-week ExTr and whey protein supplementation intervention trial. Subjects (n = 304, 48 ± 7 years, females = 186) were randomized to a specific whey protein group (0, 20, 40, or 60 g per day) and all subjects participated in a resistance (2 d/wk) and aerobic (1 d/wk) training program. Testing was conducted at baseline and post-intervention. RESULTS: At baseline, 36% (n = 110) and 12% (n = 35) of subjects were classified with SHT based on the TSH ≥ 3 µIU/L or TSH ≥ 4.5 µIU/L cut-offs, respectively. No association was found between baseline TSH and baseline measures of glucose homeostatic control. Whey protein supplementation did not influence intervention outcomes. Post-intervention (n = 164), no change was observed in TSH. SHT did not affect changes in insulin sensitivity following ExTr. CONCLUSION: These results support that the health benefits of ExTr for the management of insulin resistance (IR) are not blunted by SHT.
Assuntos
Terapia por Exercício/métodos , Hipotireoidismo/sangue , Hipotireoidismo/terapia , Avaliação de Resultados em Cuidados de Saúde , Sobrepeso/sangue , Sobrepeso/terapia , Proteínas do Soro do Leite/farmacologia , Adulto , Glicemia/metabolismo , Terapia Combinada , Suplementos Nutricionais , Feminino , Teste de Tolerância a Glucose , Humanos , Hipotireoidismo/dietoterapia , Resistência à Insulina/fisiologia , Masculino , Pessoa de Meia-Idade , Obesidade/sangue , Obesidade/dietoterapia , Obesidade/terapia , Sobrepeso/dietoterapia , Tireotropina/sangue , Proteínas do Soro do Leite/administração & dosagemRESUMO
SCOPE: The pharmacokinetics, bioavailability, and regional brain distribution of polyphenols from apple-grape seed extract (AGSE) mixture and bilberry extract were studied after 3 weeks of dosing in weanling pigs. MATERIALS AND METHODS: Weanling piglets were treated for 3 weeks with extracts of (AGSE) or bilberry extracts, using a physiological (27.5 mg/kg) or supplement (82.5 mg/kg) dose. A 24-h pharmacokinetic study was conducted and brain tissue was harvested. Major flavan-3-ol and flavonol metabolites including catechin-O-ß-glucuronide, epicatechin-O-ß-glucuronide, 3'O-methyl-catechin-O-ß-glucuronide, 3'O-methyl-epicatechin-O-ß-glucuronide, quercetin-O-ß-glucuronide, and O-methyl-quercetin-O-ß-glucuronide were analyzed in plasma, urine, and regional brain extracts from AGSE groups. Anthocyanidin-O-galactosides and O-glucosides of delphinidin (Del), cyanidin (Cyn), petunidin (Pet), peonidin (Peo), and malvidin (Mal) were analyzed in plasma, urine, and brain extracts from bilberry groups. CONCLUSION: Significant plasma dose-dependence was observed in flavan-3-ol metabolites of the AGSE group and in Mal, Del and Cyn galactosides and Pet, Peo, and Cyn glucosides of the bilberry groups. In the brain, a significant dose dependence was found in the cerebellum and frontal cortex in all major flavan-3-ol metabolites. All anthocyanidin glycosides, except for delphinidin, showed a dose-dependent increase in the cerebellum.
Assuntos
Encéfalo/metabolismo , Malus/química , Polifenóis/farmacocinética , Vaccinium myrtillus/química , Vitis/química , Animais , Antocianinas/metabolismo , Antocianinas/farmacocinética , Disponibilidade Biológica , Peso Corporal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ingestão de Alimentos/efeitos dos fármacos , Extrato de Sementes de Uva/química , Fígado/efeitos dos fármacos , Fígado/enzimologia , Extratos Vegetais/química , Polifenóis/sangue , Polifenóis/urina , Sus scrofa , Distribuição Tecidual , DesmameRESUMO
Fish has many health benefits but is also the most common source of methylmercury. The bioavailability of methylmercury in fish may be affected by other meal components. In this study, the effect of green tea on the bioavailability of methylmercury from an oral bolus of fish muscle tissue was studied in rats and compared to a water treated control group and a group treated with meso-2,3-dimercaptosuccinic acid (DMSA), a compound used medically to chelate mercury. Rats were given a single oral dose of fish tissue via gavage and one of the treatments. Rats were given access to food for 3 h at 12 h intervals. They were dosed with each of the treatments with each meal. Blood samples were collected for 95 hours. Green tea significantly increased the concentration of total mercury in blood relative to the control, whereas DMSA significantly decreased it. In addition, feeding caused a slight increase in blood mercury for several meals following the initial dose.
Assuntos
Peixes , Rim/efeitos dos fármacos , Mercúrio/toxicidade , Chá/efeitos adversos , Animais , Quelantes/química , Rim/patologia , Mercúrio/sangue , Ratos , Succímero/administração & dosagemRESUMO
The development of an on-animal separation-based sensor that can be employed for monitoring drug metabolism in a freely roaming sheep is described. The system consists of microdialysis sampling coupled to microchip electrophoresis with electrochemical detection (MD-ME-EC). Separations were accomplished using an all-glass chip with integrated platinum working and reference electrodes. Discrete samples from the microdialysis flow were introduced into the electrophoresis chip using a flow-gated injection approach. Electrochemical detection was accomplished in-channel using a two-electrode isolated potentiostat. Nitrite was separated by microchip electrophoresis using reverse polarity and a run buffer consisting of 50 mM phosphate at pH 7.4. The entire system was under telemetry control. The system was first tested with rats to monitor the production of nitrite following perfusion of nitroglycerin into the subdermal tissue using a linear probe. The data acquired using the on-line MD-ME-EC system were compared to those obtained by off-line analysis using liquid chromatography with electrochemical detection (LC-EC), using a second microdialysis probe implanted parallel to the first probe in the same animal. The MD-ME-EC device was then used on-animal to monitor the subdermal metabolism of nitroglycerin in sheep. The ultimate goal is to use this device to simultaneously monitor drug metabolism and behavior in a freely roaming animal.
Assuntos
Microtecnologia/instrumentação , Movimento , Preparações Farmacêuticas/isolamento & purificação , Preparações Farmacêuticas/metabolismo , Ovinos , Animais , Eletroquímica , Eletrodos , Eletroforese em Microchip , Desenho de Equipamento , Masculino , Microdiálise , RatosRESUMO
SCOPE: Grape seed polyphenol extract (GSPE) is receiving increasing attention for its potential preventative and therapeutic roles in Alzheimer's disease (AD) and other age-related neurodegenerative disorders. The intestinal microbiota is known to actively convert many dietary polyphenols, including GSPE, to phenolic acids. There is limited information on the bioavailability and bioactivity of GSPE-derived phenolic acid in the brain. METHODS AND RESULTS: We orally administered GSPE to rats and investigated the bioavailability of 12 phenolic acids known to be generated by microbiota metabolism of anthocyanidins. GSPE treatment significantly increased the content of two of the phenolic acids in the brain: 3-hydroxybenzoic acid and 3-(3´-hydroxyphenyl)propionic acid, resulting in the brain accumulations of the two phenolic acids at micromolar concentrations. We also provided evidence that 3-hydroxybenzoic acid and 3-(3´-hydroxyphenyl)propionic acid potently interfere with the assembly of ß-amyloid peptides into neurotoxic ß-amyloid aggregates that play key roles in AD pathogenesis. CONCLUSION: Our observation suggests important contribution of the intestinal microbiota to the protective activities of GSPE (as well as other polyphenol preparations) in AD. Outcomes from our studies support future preclinical and clinical investigations exploring the potential contributions of the intestinal microbiota in protecting against the onset/progression of AD and other neurodegenerative conditions.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Peptídeos beta-Amiloides/metabolismo , Microbioma Gastrointestinal , Fragmentos de Peptídeos/metabolismo , Polifenóis/farmacocinética , Peptídeos beta-Amiloides/antagonistas & inibidores , Animais , Antocianinas/administração & dosagem , Antocianinas/sangue , Antocianinas/farmacocinética , Disponibilidade Biológica , Encéfalo/metabolismo , Fermentação , Extrato de Sementes de Uva/administração & dosagem , Extrato de Sementes de Uva/sangue , Extrato de Sementes de Uva/farmacocinética , Hidroxibenzoatos/sangue , Hidroxibenzoatos/metabolismo , Intestinos/efeitos dos fármacos , Intestinos/microbiologia , Masculino , Fragmentos de Peptídeos/antagonistas & inibidores , Fenóis/metabolismo , Polifenóis/administração & dosagem , Polifenóis/sangue , Propionatos/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
Alzheimer's disease (AD) is the most prevalent neurodegenerative disease of aging and currently has no cure. Its onset and progression are influenced by multiple factors. There is growing consensus that successful treatment will rely on simultaneously targeting multiple pathological features of AD. Polyphenol compounds have many proven health benefits. In this study, we tested the hypothesis that combining three polyphenolic preparations (grape seed extract, resveratrol, and Concord grape juice extract), with different polyphenolic compositions and partially redundant bioactivities, may simultaneously and synergistically mitigate amyloid-ß (Aß) mediated neuropathology and cognitive impairments in a mouse model of AD. We found that administration of the polyphenols in combination did not alter the profile of bioactive polyphenol metabolites in the brain. We also found that combination treatment resulted in better protection against cognitive impairments compared to individual treatments, in J20 AD mice. Electrophysiological examination showed that acute treatment with select brain penetrating polyphenol metabolites, derived from these polyphenols, improved oligomeric Aß (oAß)-induced long term potentiation (LTP) deficits in hippocampal slices. Moreover, we found greatly reduced total amyloid content in the brain following combination treatment. Our studies provided experimental evidence that application of polyphenols targeting multiple disease-mechanisms may yield a greater likelihood of therapeutic efficacy.
RESUMO
SCOPE: Metabolic syndrome has become an epidemic and poses tremendous burden on the health system. People with metabolic syndrome are more likely to experience cognitive decline. As obesity and sedentary lifestyles become more common, the development of early prevention strategies is critical. In this study, we explore the potential beneficial effects of a combinatory polyphenol preparation composed of grape seed extract, Concord purple grape juice extract, and resveratrol, referred to as standardized grape polyphenol preparation (SGP), on peripheral as well as brain dysfunction induced by metabolic syndrome. METHODS AND RESULTS: We found dietary fat content had minimal effect on absorption of metabolites of major polyphenols derived from SGP. Using a diet-induced animal model of metabolic syndrome (DIM), we found that brain functional connectivity and synaptic plasticity are compromised in the DIM mice. Treatment with SGP not only prevented peripheral metabolic abnormality but also improved brain synaptic plasticity. CONCLUSION: Our study demonstrated that SGP, comprised of multiple bioavailable and bioactive components targeting a wide range of metabolic syndrome related pathological features, provides greater global protection against peripheral and central nervous system dysfunctions and can be potentially developed as a novel prevention/treatment for improving brain connectivity and synaptic plasticity important for learning and memory.
Assuntos
Síndrome Metabólica/tratamento farmacológico , Plasticidade Neuronal/efeitos dos fármacos , Polifenóis/farmacologia , Sinapses/efeitos dos fármacos , Vitis/química , Animais , Disponibilidade Biológica , Encéfalo/efeitos dos fármacos , Encéfalo/fisiopatologia , Gorduras na Dieta/farmacologia , Modelos Animais de Doenças , Feminino , Extrato de Sementes de Uva/farmacologia , Masculino , Síndrome Metabólica/dietoterapia , Camundongos , Camundongos Endogâmicos C57BL , Polifenóis/farmacocinética , Ratos , Ratos Sprague-Dawley , Resveratrol , Estilbenos/farmacologiaRESUMO
Epidemiological and preclinical studies indicate that polyphenol intake from moderate consumption of red wines may lower the relative risk for developing Alzheimer's disease (AD) dementia. There is limited information regarding the specific biological activities and cellular and molecular mechanisms by which wine polyphenolic components might modulate AD. We assessed accumulations of polyphenols in the rat brain following oral dosage with a Cabernet Sauvignon red wine and tested brain-targeted polyphenols for potential beneficial AD disease-modifying activities. We identified accumulations of select polyphenolic metabolites in the brain. We demonstrated that, in comparison to vehicle-control treatment, one of the brain-targeted polyphenol metabolites, quercetin-3-O-glucuronide, significantly reduced the generation of ß-amyloid (Aß) peptides by primary neuron cultures generated from the Tg2576 AD mouse model. Another brain-targeted metabolite, malvidin-3-O-glucoside, had no detectable effect on Aß generation. Moreover, in an in vitro analysis using the photo-induced cross-linking of unmodified proteins (PICUP) technique, we found that quercetin-3-O-glucuronide is also capable of interfering with the initial protein-protein interaction of Aß(1-40) and Aß(1-42) that is necessary for the formation of neurotoxic oligomeric Aß species. Lastly, we found that quercetin-3-O-glucuronide treatment, compared to vehicle-control treatment, significantly improved AD-type deficits in hippocampal formation basal synaptic transmission and long-term potentiation, possibly through mechanisms involving the activation of the c-Jun N-terminal kinases and the mitogen-activated protein kinase signaling pathways. Brain-targeted quercetin-3-O-glucuronide may simultaneously modulate multiple independent AD disease-modifying mechanisms and, as such, may contribute to the benefits of dietary supplementation with red wines as an effective intervention for AD.
Assuntos
Doença de Alzheimer/dietoterapia , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Quercetina/análogos & derivados , Administração Oral , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Antocianinas/administração & dosagem , Antocianinas/farmacocinética , Disponibilidade Biológica , Células Cultivadas , Suplementos Nutricionais , Modelos Animais de Doenças , Glucosídeos , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Plasticidade Neuronal/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fármacos Neuroprotetores/administração & dosagem , Fármacos Neuroprotetores/farmacocinética , Polifenóis/administração & dosagem , Polifenóis/metabolismo , Polifenóis/farmacocinética , Multimerização Proteica/efeitos dos fármacos , Quercetina/administração & dosagem , Quercetina/farmacocinética , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Vinho/análiseRESUMO
Little is known about the effects of different quantities of whey protein on exercise training-induced changes in body composition and indices of metabolic syndrome in middle-aged overweight and obese adults. Therefore, we examined the effects of consuming 0.8-MJ supplements with 0 (n = 126), 10 (n = 112), 20 (n = 44), or 30 (n = 45) g whey protein twice daily in conjunction with resistance (2 d/wk) and aerobic (1 d/wk) exercise training in a double-blind, randomized, placebo-controlled, community-based 9-mo study in men (n = 117) and women (n = 210); (age: 48 ± 7.9 y; BMI: 30.0 ± 2.8 kg/m(2)). Whey protein supplementation did not influence any of the following outcomes, some of which were affected by training. Among all participants, strength increased by 15 ± 12% (P < 0.001) and maximal oxygen uptake capacity (VO(2)max) increased by 9 ± 15% (P < 0.001). Body weight was unchanged (0.1 ± 3.7 kg, P = 0.80), lean body mass increased by 1.9 ± 2.8% (0.95 ± 1.3 kg, P < 0.001), and fat mass decreased by 2.6 ± 9.4% (-0.86 ± 3.1 kg, P = 0.001). Oral-glucose-tolerance testing showed that plasma glucose AUC was unchanged (-18.0 ± 170 mmol/L· 3 h, P = 0.16), insulin AUC decreased by 2.6 ± 32% (-7.5 ± 29 nmol/L· 3 h, P = 0.01), and HOMA-IR (0.2 ± 2.0, P = 0.81) and the insulin sensitivity index (0.3 ± 3.0, P = 0.63) were unchanged. Plasma concentrations of TG; total, LDL, and HDL cholesterol; C-reactive protein; plasminogen activator inhibitor-1; blood pressure; and waist circumference were unchanged. Whey protein supplementation did not affect exercise training-induced responses in body composition and indices of metabolic syndrome in middle-aged overweight and obese adults who maintained body weight.
Assuntos
Composição Corporal/fisiologia , Exercício Físico/fisiologia , Síndrome Metabólica/metabolismo , Proteínas do Leite/farmacologia , Sobrepeso/tratamento farmacológico , Adulto , Apetite , Proteínas Alimentares , Suplementos Nutricionais , Método Duplo-Cego , Metabolismo Energético , Feminino , Humanos , Rim/efeitos dos fármacos , Rim/fisiologia , Masculino , Pessoa de Meia-Idade , Fenômenos Fisiológicos da Nutrição , Sobrepeso/metabolismo , Proteínas do Soro do LeiteRESUMO
While polyphenolic compounds have many health benefits, the potential development of polyphenols for the prevention/treatment of neurological disorders is largely hindered by their complexity as well as by limited knowledge regarding their bioavailability, metabolism, and bioactivity, especially in the brain. We recently demonstrated that dietary supplementation with a specific grape-derived polyphenolic preparation (GP) significantly improves cognitive function in a mouse model of Alzheimer's disease (AD). GP is comprised of the proanthocyanidin (PAC) catechin and epicatechin in monomeric (Mo), oligomeric, and polymeric forms. In this study, we report that following oral administration of the independent GP forms, only Mo is able to improve cognitive function and only Mo metabolites can selectively reach and accumulate in the brain at a concentration of â¼400 nM. Most importantly, we report for the first time that a biosynthetic epicatechin metabolite, 3'-O-methyl-epicatechin-5-O-ß-glucuronide (3'-O-Me-EC-Gluc), one of the PAC metabolites identified in the brain following Mo treatment, promotes basal synaptic transmission and long-term potentiation at physiologically relevant concentrations in hippocampus slices through mechanisms associated with cAMP response element binding protein (CREB) signaling. Our studies suggest that select brain-targeted PAC metabolites benefit cognition by improving synaptic plasticity in the brain, and provide impetus to develop 3'-O-Me-EC-Gluc and other brain-targeted PAC metabolites to promote learning and memory in AD and other forms of dementia.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Antioxidantes/uso terapêutico , Proantocianidinas/uso terapêutico , Peptídeos beta-Amiloides/antagonistas & inibidores , Peptídeos beta-Amiloides/toxicidade , Animais , Antioxidantes/metabolismo , Disponibilidade Biológica , Biotransformação , Western Blotting , Cromatografia Líquida de Alta Pressão , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Dieta , Sistemas de Liberação de Medicamentos , Feminino , Potenciação de Longa Duração/efeitos dos fármacos , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Camundongos , Camundongos Transgênicos , Polifenóis/química , Polifenóis/farmacologia , Proantocianidinas/farmacocinética , Proantocianidinas/farmacologia , Ratos , Ratos Sprague-Dawley , Transmissão Sináptica/efeitos dos fármacos , Vitis/químicaRESUMO
Two different strategies for investigating the likely fate, after ingestion, of natural, bioactive berry constituents (anthocyanins and other non-nutritive flavonoids) are compared. A model of the human gastrointestinal tract (TIM-1) that mimicked the biological environment from the point of swallowing and ingestion through the duodenum, jejunum, and ileum (but not the colon) was used to monitor the stability and bioaccessibility of anthocyanins from both maqui berry and wild blueberry. TIM-1 revealed that most anthocyanins were bioaccessible between the second and third hours after intake. Alternatively, biolabeled anthocyanins and other flavonoids generated in vitro from berry and grape cell cultures were administered to in vivo (rodent) models, allowing measurement and tracking of the absorption and transport of berry constituents and clearance through the urinary tract and colon. The advantages and limitations of the alternative strategies are considered.
Assuntos
Antocianinas/farmacocinética , Mirtilos Azuis (Planta)/química , Frutas/química , Polifenóis/farmacocinética , Absorção , Animais , Antocianinas/análise , Disponibilidade Biológica , Isótopos de Carbono/análise , Isótopos de Carbono/farmacocinética , Linhagem Celular , Simulação por Computador , Trato Gastrointestinal/efeitos dos fármacos , Trato Gastrointestinal/metabolismo , Humanos , Modelos Biológicos , Compostos Fitoquímicos/análise , Compostos Fitoquímicos/farmacocinética , Polifenóis/análise , Ratos , Vitis/químicaRESUMO
There is potential that the pathological effects of oxidative stress (OS) associated diseases such as diabetes could be ameliorated with antioxidants, but this will require a clearer understanding of the pathway(s) by which proteins are damaged by OS. This study reports the development and use of methods that assess the efficacy of dietary antioxidant supplementation at a mechanistic level. Data reported here evaluate the impact of green tea supplementation on oxidative stress induced post-translational modifications (OSi-PTMs) in plasma proteins of Zucker diabetic fatty (ZDF) rats. The mechanism of antioxidant protection was examined through both the type and amount of OSi-PTMs using mass spectrometry based identification and quantification. Carbonylated proteins in freshly drawn blood samples were derivatized with biotin hydrazide. Proteins thus biotinylated were selected from plasma samples of green tea fed diabetic rats and control animals by avidin affinity chromatography, further fractionated by reversed phase chromatography (RPC); fractions from the RPC column were tryptic digested, and the tryptic digest was fractionated by RPC before being identified by tandem mass spectrometry (MS/MS). Relative quantification of peptides bearing carbonylation sites was achieved for the first time by RPC-MS/MS using selective reaction monitoring (SRM). Seventeen carbonylated peptides were detected and quantified in both control and treated plasma. The relative concentration of eight was dramatically different between control and green tea treated animals. Seven of the OSi-PTM bearing peptides had dropped dramatically in concentration with treatment while one increased, indicating differential regulation of carbonylation by antioxidants. Green tea antioxidants were found to reduce carbonylation of proteins by lipid peroxidation end products most, followed by advanced glycation end products to a slightly lower extent. Direct oxidation of proteins by reactive oxygen species (ROS) was protected the least by green tea.
Assuntos
Antioxidantes/farmacologia , Hemoglobinas/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Carbonilação Proteica , Espectrometria de Massas em Tandem , Animais , Antioxidantes/química , Biotina/análogos & derivados , Biotina/química , Peptídeos/análise , Ratos , Ratos Zucker , Espécies Reativas de Oxigênio/metabolismo , Chá/química , Tripsina/metabolismoRESUMO
This study reports for the first time qualitative and quantitative differences in carbonylated proteins shed into blood as a function of increasing levels of OS. Carbonylated proteins in freshly drawn blood from pairs of diabetic and lean rats were derivatized with biotin hydrazide, dialyzed, and enriched with avidin affinity chromatography. Proteins thus selected were used in several ways. Differences between control and diabetic subjects in relative concentration of proteins was achieved by differential labeling of tryptic digests with iTRAQ reagents followed by reversed phase chromatography (RPC) and tandem mass spectrometry (MS/MS). Identification and characterization of OS induced post-translational modification sites in contrast was achieved by fractionation of affinity selected proteins before proteolysis and RPC-MS/MS. Relative quantification of peptides bearing oxidative modifications was achieved for the first time by selective reaction monitoring (SRM). Approximately 1.7% of the proteins in Zucker diabetic rat plasma were selected by the avidin affinity column as compared to 0.98% in lean animal plasma. Among the 35 proteins identified and quantified, Apo AII, clusterin, hemopexin precursor, and potassium voltage-gated channel subfamily H member 7 showed the most dramatic changes in concentration. Seventeen carbonylation sites were identified and quantified, 11 of which changed more than 2-fold in oxidation state. Three types of carbonylation were identified at these sites: direct oxidative cleavage from reactive oxygen species, glycation and addition of advanced glycation end products, and addition of lipid peroxidation products. Direct oxidation was the dominant form of carbonylation observed while hemoglobin and murinoglobulin 1 homologue were the most heavily oxidized proteins.
Assuntos
Estresse Oxidativo/fisiologia , Animais , Biotina/análogos & derivados , Proteínas Sanguíneas/análise , Proteínas Sanguíneas/química , Cromatografia de Fase Reversa , Bases de Dados de Proteínas , Diabetes Mellitus Experimental , Isoprostanos/urina , Oxirredução , Fragmentos de Peptídeos/análise , Fragmentos de Peptídeos/química , Carbonilação Proteica , Proteômica/métodos , Ratos , Ratos Zucker , Espectrometria de Massas em TandemRESUMO
Polyphenols in dietary and botanical matrices are usually present as simple and complex O-glycosides. In fermented dietary materials, the glycosidic moiety is removed and accompanied in some cases by more complex changes to the polyphenol. As for most xenobiotics, polyphenols undergo phase II conjugation in the intestinal wall during their absorption from the gut. In contrast, a few polyphenols, such as puerarin in the kudzu vine, are C-glycosides and are stable in the gut and during absorption, distribution and excretion. Large bowel bacteria reduce polyphenol aglycones, causing opening of the heterocyclic B-ring and ring cleavage. The products are mostly absorbed and enter the bloodstream. Phase I and II metabolism events occur in the intestine and the liver - most polyphenols predominantly circulate as ß-glucuronides and sulfate esters with very little as the aglycones, the presumed active forms. In addition, metabolism can occur in non-hepatic tissues and cells including breast tumor cells that have variable amounts of cytochrome P450s, sulfatase and sulfotransferase activities. Inflammatory cells produce chemical oxidants (HOCl, HOBr, ONO(2)(-)) that will react with polyphenols. The isoflavones daidzein and genistein and the flavonol quercetin form mono- and dichlorinated products in reaction with HOCl. Genistein is converted to 3'-nitrogenistein in the lung tissue of lipopolysaccharide-treated rats. Whereas polyphenols that can be converted to quinones or epoxides react with glutathione (GSH) to form adducts, chlorinated isoflavones do not react with GSH; instead, they are converted to ß-glucuronides and are excreted in bile. Analysis of polyphenols and their metabolites is routinely carried out with great sensitivity, specificity and quantification by LC-tandem mass spectrometry. Critical questions about the absorption and tissue uptake of complex polyphenols such as the proanthocyanins can be answered by labeling these polyphenols with (14)C-sucrose in plant cell culture and then purifying them for use in animal experiments. The (14)C signature is quantified using accelerator mass spectrometry, a technique capable of detecting one (14)C atom in 10(15) carbon atoms. This permits the study of the penetration of the polyphenols into the interstitial fluid, the fluid that is actually in contact with non-vascular cells.
Assuntos
Alimentos , Genisteína/análise , Isoflavonas/análise , Isoflavonas/metabolismo , Polifenóis/análise , Polifenóis/metabolismo , Animais , Disponibilidade Biológica , Genisteína/metabolismo , Glucuronídeos/análise , Glucuronídeos/metabolismo , Glicosídeos/análise , Glicosídeos/metabolismo , Humanos , Mucosa Intestinal/metabolismo , Fígado/metabolismo , Espectrometria de MassasRESUMO
Hairy roots were induced in four genotypes from three kudzu species (Pueraria montana var. lobata, P. lobata and P. phaseoloides) in vitro using Agrobacterium rhizogenes to stimulate rapid secondary metabolite synthesis. Hairy roots from P. montana var. lobata (United States Department of Agriculture no. PI 434246) yielded the highest puerarin and total isoflavone content and the greatest new biomass per growth cycle among the genotypes evaluated. Hairy roots from this genotype were selected for radiolabeling using (14)C-sucrose as a carbon source. Isoflavones from radiolabeled kudzu hairy root cultures were extracted with 80% methanol, partitioned by solvent extraction, and then subfractionated by Sephadex LH-20 gel filtration. Radiolabeled isoflavones were isolated in a highly enriched fraction, which contained predominantly puerarin, daidzin and malonyl-daidzin and had an average radioactivity of 8.614 MBq/g (232.8 µCi/g) dry fraction. The (14)C-radiolabeled, isoflavone-rich fraction was orally administered at a dose of 60 mg/kg body weight to male Sprague-Dawley rats implanted with a jugular catheter, a subcutaneous ultrafiltrate probe and a brain microdialysate probe. Serum, interstitial fluid, brain microdialysate, urine and feces were collected using a Culex(®) Automated Blood Collection System for 24 h. At the end of this period, rats were sacrificed and major tissues were collected. Analysis by a scintillation counter confirmed that a bolus dose of (14)C-radiolabeled, isoflavone-rich kudzu fraction reached bone tissues, which accumulated 0.011%, 0.09% and 0.003% of the administered dose in femur, tibia and vertebrae, respectively. Femurs extracted with 80% methanol were analyzed by high-performance liquid chromatography with electrospray ionization-mass spectrometry and were found to contain trace quantities of puerarin, daidzein and puerarin glucuronide. This study demonstrates that kudzu isoflavones and metabolites are capable of reaching bone tissues, where they may contribute to the prevention of osteoporosis and the promotion of bone health.
Assuntos
Osso e Ossos/metabolismo , Isoflavonas/farmacocinética , Pueraria/química , Animais , Osso e Ossos/efeitos dos fármacos , Radioisótopos de Carbono , Cromatografia Líquida de Alta Pressão , Isoflavonas/administração & dosagem , Isoflavonas/isolamento & purificação , Masculino , Osteoporose/prevenção & controle , Extratos Vegetais/administração & dosagem , Extratos Vegetais/farmacocinética , Raízes de Plantas/química , Ratos , Ratos Sprague-Dawley , Espectrometria de Massas por Ionização por Electrospray , Distribuição TecidualRESUMO
Grape polyphenols confer potential health benefits, including prevention of neurodegenerative diseases. To determine the absorption and tissue distribution of the complex grape polyphenol mixture, (14)C-labeled polyphenols were biosynthesized by grape cell suspension cultures, during co-incubation with radioisotopically labeled sucrose, and fractionated into polyphenolic subfractions. The pharmacokinetics and distribution of grape polyphenols into blood, brain, and peripheral interstitial fluid were determined by tracking the (14)C label. The blood peak (14)C concentration of the fractions ranged from 15 minutes to 4 hours. Absorption and tissue distribution varied greatly between fractions. Concentrations in interstitial fluid were lower than in blood. The amount of residual label in the brain at 24 hours ranged from 0.1% to 1.7% of the dose, depending on the fraction. (14)C label found in the brain tissue and brain microdialysate indicated that grape polyphenols or their metabolites are able to cross the blood-brain barrier. Using (14)C-labeled plant polyphenols it is possible to track the compounds or their metabolic products into any tissue and determine distribution patterns in spite of low concentrations. A central question regarding the potential role of dietary polyphenolics in neurodegenerative research is whether they are bioavailable in the brain. Our observations indicate that some grape-derived polyphenolics do reach the brain, which suggests their potential value for applications in neurodegenerative disorders.
Assuntos
Sistema Nervoso Central/metabolismo , Líquido Extracelular/metabolismo , Flavonoides/farmacocinética , Fenóis/farmacocinética , Extratos Vegetais/farmacocinética , Vitis/química , Administração Oral , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Animais , Radioisótopos de Carbono/análise , Radioisótopos de Carbono/farmacocinética , Sistema Nervoso Central/química , Sistema Nervoso Central/efeitos dos fármacos , Modelos Animais de Doenças , Líquido Extracelular/química , Líquido Extracelular/efeitos dos fármacos , Flavonoides/administração & dosagem , Flavonoides/química , Humanos , Masculino , Fenóis/administração & dosagem , Fenóis/química , Extratos Vegetais/administração & dosagem , Extratos Vegetais/química , Polifenóis , Ratos , Ratos Sprague-Dawley , Distribuição TecidualRESUMO
The impact of carbohydrates and milk on the bioavailability of catechin (C) and epicatechin (EC) from chocolate has been previously studied. However, little data exist regarding potential modulation of the phase II metabolism by these chocolate matrix factors. The objectives of this study were to assess the impact of matrix composition on qualitative and quantitative profiles of circulating catechins and their metabolites following administration of commercially relevant chocolate confections. Sprague-Dawley rats were administered 1.5 g of a confection (reference dark, high sucrose, or milk chocolate) by intragastric gavage, and plasma samples were collected over 8 h. High-performance liquid chromatography-mass spectrometry analysis was performed to quantify C, EC, and their metabolites. The predominant metabolites were O-glucuronides (two metabolites) and O-Me-O-glucuronides (three metabolites). Plasma concentrations of metabolites were generally the highest for high sucrose treatment and lowest for milk treatment, while the reference dark treatment generally resulted in intermediate concentrations. The O-Me-(+/-)-C/EC-O-beta-glucuronide (peak 4) was significantly higher for the high sucrose treatment (2325 nM h) versus the milk treatment (1300 nM h). Additionally, C(MAX) values for (+/-)-C/EC-O-beta-glucuronide (peak 3) and two O-Me-(+/-)-C/EC-O-beta-glucuronides (peaks 4 and 6) were significantly higher for the high sucrose treatment (4012, 518, and 2518 nM, respectively) versus the milk treatment (2590, 240, and 1670 nM, respectively). Milk and sucrose appear to modulate both metabolism and plasma pharmacokinetics and, to a lesser extent, the overall bioavailability of catechins from chocolate confections.
Assuntos
Cocos/química , Flavonoides/farmacocinética , Extratos Vegetais/farmacocinética , Administração Oral , Animais , Flavonoides/administração & dosagem , Flavonoides/metabolismo , Masculino , Extratos Vegetais/administração & dosagem , Extratos Vegetais/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
Bioactive compounds in botanicals may be beneficial in preventing age-related neurodegenerative diseases, but for many compounds conventional methods may be inadequate to detect if these compounds cross the blood brain barrier or to track the pharmacokinetics in the brain. By combining a number of unique technologies it has been possible to utilize the power of AMS to study the pharmacokinetics of bioactive compounds in the brain at very low concentrations. (14)C-labeled compounds can be biosynthesized by plant cell suspension cultures co-incubated with radioisotopically-labeled sucrose and isolated and separated into a series of bioactive fractions.To study the pharmacokinetics and tissue distribution of (14)C labeled plant polyphenols, rats were implanted with jugular catheters, subcutaneous ultrafiltration probes and brain microdialysis probes. Labeled fractions were dosed orally. Interstitial fluid (ISF) and brain microdialysate samples were taken in tandem with blood samples. It was often possible to determine (14)C in blood and ISF with a ß-counter. However, brain microdialysate samples (14)C levels on the order of 10(7) atoms/sample required AMS technology. The Brain Microdialysate(AUC)/Serum(AUC) ranged from .021- to .029, with the higher values for the glycoside fractions. By using AMS in combination with traditional methods, it is possible to study uptake by blood, distribution to ISF and determine the amount of a dose which can reach the brain and follow the pharmacokinetics in the brain.