RESUMO
In the present paper it was investigated the role of sauroxine, an alkaloid of Phlegmariurus saururus, as a modulator of some types of learning and memory, considering the potential nootropic properties previously reported for the alkaloid extract and the main alkaloid sauroine. Sauroxine was isolated by means of an alkaline extraction, purified by several chromatographic techniques, and assayed in electrophysiological experiments on rat hippocampus slices, tending towards the elicitation of the long-term potentiation (LTP) phenomena. It was also studied the effects of intrahippocampal administration of sauroxine on memory retention in vivo using a Step-down test. Being the bio distribution of a drug an important parameter to be considered, the concentration of sauroxine in rat brain was determined by GLC-MS. Sauroxine blocked LTP generation at both doses used, 3.65 and 3.610-2µM. In the behavioral test, the animals injected with this alkaloid (3.6510-3nmol) exhibited a significant decrease on memory retention compared with control animals. It was also showed that sauroxine reached the brain (3.435µg/g tissue), after an intraperitoneal injection, displaying its ability to cross the blood-brain barrier. Thus, sauroxine demonstrated to exert an inhibition on these mnemonic phenomena. The effect here established for 1 is defeated by other constituents according to the excellent results obtained for P. saururus alkaloid extract as well as for the isolated alkaloid sauroine.
Assuntos
Alcaloides/farmacologia , Hipocampo/fisiologia , Potenciação de Longa Duração/efeitos dos fármacos , Memória/efeitos dos fármacos , Alcaloides/química , Animais , Cromatografia Gasosa-Espectrometria de Massas , Masculino , Ratos WistarRESUMO
Uno de los principales mecanismos para la resistencia de Candida albicans a azoles es la sobreexpresión de transportadores que extruyen los fármacos antimicóticos y disminuyen su concentración intracelular. Anteriormente nuestros laboratorios habían informado que el flavonoide prenilado 2', 4'-dihidroxi - 5'-(1'''-dimetilalil)-6-prenil pinocembrina (6PP) inhibe en forma competitiva los transportadores cdr, dependientes del ATP, además de ser antimicrobiano per se. En este trabajo realizamos estudios bioinformáticos para analizar los posibles sitios de interacción molecular estereoespecífica del 6PP, fluconazol y adenosín trifosfato (ATP) con los transportadores cdr1 y cdr2. Se confeccionaron los modelos tridimensionales de los ligandos y de los transportadores a través de estudios de homología de datos respecto de otras macromoléculas. Se calculó la variación de energía libre de Gibbs ( G) asociada a 45 modelos de interacción molecular y se estudiaron cdr1 y cdr2 en formacomparativa, dada su similitud estructural. Se seleccionaron los modelos más probables desde el punto de vista termodinámico. El 6PP y el fluconazol podrían competir de diversas maneras por un sitio estereoespecífico que atraviesa la membrana de la levadura. Este sitio, numerado como 1, fue estudiado exhaustivamente, es común a los transportadores cdr1 y cdr2 y aporta selectivamente distintosaminoácidos. Entre estos últimos, los de mayor participación en los modelos de interacción fueron, en cdr1 y cdr2, respectivamente: PHE 754,756 y TYR 747,749 . Se seleccionaron los 9 mejores complejos formados por cada ligando, de acuerdo a su energía de afinidad decreciente (en KJ/mol en mejor modelo de cdr2: 6PP = -9,3; fluconazol = -6,8). Además el flavonoide prenilado competiría con el ATP en el dominio citosólico y con menor probabilidad lo imitaría el fluconazol ( G en KJ/mol en mejor modelo de cdr2: 6P P= -8,2, ATP = -7,6; fluconazol = -6,7)...
Assuntos
Humanos , Trifosfato de Adenosina/química , Antifúngicos/farmacologia , Candida albicans , Farmacorresistência Fúngica , Flavonoides/fisiologia , Prenilação de Proteína , Fluconazol/farmacocinética , Fluconazol/farmacologia , Imageamento Tridimensional , Ligantes , Substâncias Macromoleculares , Moléculas de Adesão Celular/fisiologia , EstereoisomerismoRESUMO
Huperzine A, a Lycopodium alkaloid produced by Chinese folk herb Huperzia serrata (Lycopodiaceae), has been shown to be a promising agent for the treatment of Alzheimer's disease due to its potent acetylcholinesterase (AChE) activity, as well its efficacy in the treatment of memory of aged patients. Thus, the effects of two Huperzia species of habitats in Brazil (H. quadrifariata and H. reflexa) with described in vitro AChE inhibition activities were studied and their effects on mice brain AChE inhibition were determined after a single intraperitoneal (i.p.) injection. The alkaloid extracts were administered to mice in various doses (10, 1 and 0.5mg/kg) and acetylcholinesterase activity was measured post mortem in two brain areas using the Ellman's colorimetric method. The AChE activity was found to be significantly reduced in both the cortex and hippocampus, although this activity was less potent than that of reference inhibitor huperzine A (0.5mg/kg). Thus, it appears that H. quadrifariata and H. reflexa alkaloid extracts, shown to inhibit acetylcholinesterase in vitro, also have very potent in vivo effects, suggesting that the Huperzia species may still constitute a promising source of compounds with pharmaceutical interest for Alzheimer's disease.
Assuntos
Acetilcolinesterase/metabolismo , Alcaloides/farmacologia , Doença de Alzheimer/metabolismo , Encéfalo/efeitos dos fármacos , Inibidores da Colinesterase/farmacologia , Huperzia/química , Extratos Vegetais/farmacologia , Alcaloides/uso terapêutico , Doença de Alzheimer/tratamento farmacológico , Animais , Encéfalo/metabolismo , Córtex Cerebral/efeitos dos fármacos , Inibidores da Colinesterase/uso terapêutico , Hipocampo/efeitos dos fármacos , Injeções Intraperitoneais , Masculino , Camundongos , Camundongos Endogâmicos , Fitoterapia , Extratos Vegetais/uso terapêutico , Sesquiterpenos/farmacologia , Sesquiterpenos/uso terapêuticoRESUMO
The quantitative determination of boldine alkaloid in boldo leaf extracts by employing cyclic voltammetry, at a liquid/liquid interface as well as the validation of this methodology against the reference method, high performance liquid chromatography (HPLC), are reported in the present paper. The voltammetric analysis was performed successfully and economically using two kinds of liquid/liquid interfaces: water/1,2-dicholoroethane and water/PVC (polyvinyl chloride)-gelled 1,2-dichloroethane. Linear calibration curves in the concentration range of 1.04 × 10(-5)mol L(-1) to 5.19 × 10(-4)mol L(-1) were obtained with a detection limit equal to (6.1 ± 0.7) × 10(-5)mol L(-1) and the quantitative determination of this alkaloid, in complex matrixes such as boldo leaf extracts, by the electrochemical technique proposed was found to be equal to the values obtained using the standard HPLC method. The validation analysis of this methodology against HPLC demonstrated that accuracy, linearity, limit of detection (LOD), limit of quantification (LOQ), specificity and precision are acceptable. The electroanalytical technique proposed is economical and selective, involves simple equipment and can be applied for the quantitative determination of boldine alkaloid in complex matrixes such as leaf extracts without special drug separation. Moreover, cyclic voltammetry (CV) experiments applied at the liquid/liquid interface under different experimental conditions allowed us to study the transfer mechanism of boldine, and determine a value of pK(a)(w)=6.90 for protonated boldine, from the variation of voltammetric peak current with pH.
Assuntos
Alcaloides/química , Aporfinas/análise , Cromatografia Líquida de Alta Pressão/métodos , Aporfinas/química , Calibragem , Calorimetria/métodos , Eletroquímica/métodos , Eletrodos , Modelos Químicos , Análise de Regressão , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Água/químicaRESUMO
In inflammation, nitric oxide (NO) acts as a pro-inflammatory mediator, which is synthesized by inducible nitric oxide synthase (iNOS) in response to pro-inflammatory agents such as lipopolysaccharide (LPS). Quercetin (Qt) has anti-inflammatory properties through its ability to inhibits nitric oxide production and iNOS expression in different cellular types. In the present study, we evaluated the effect of a semi-synthetic acetyl (quercetin-3,5,7,3'-tetraacetyl: TAQt) Qt derivative and two natural sulphated (quercetin-3-acetyl-7,3',4'-trisulphate: ATS and quercetin-3,7,3',4'-tetrasulphate: QTS) Qt derivatives on the LPS-induced NO production and iNOS expression in J774A.1 cells. Our results demonstrate that only TAQt inhibited the NO production by decreasing the iNOS mRNA and protein levels. In addition, we showed that TAQt blocked the LPS-induced nuclear NF-kappaB translocation by inhibiting the IkappaB-alpha degradation. Hence, as TAQt inhibited the LPS-induced iNOS expression and NO production, it could therefore be considered as a potential therapeutic agent for the treatment of inflammatory diseases related with the NO system.
Assuntos
Anti-Inflamatórios/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Lipopolissacarídeos/farmacologia , Óxido Nítrico Sintase Tipo II/biossíntese , Quercetina/farmacologia , Animais , Antioxidantes/farmacologia , Linhagem Celular , Proteínas I-kappa B/metabolismo , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Mediadores da Inflamação/metabolismo , Macrófagos , Camundongos , Inibidor de NF-kappaB alfa , NF-kappa B/metabolismo , Óxido Nítrico/biossíntese , Quercetina/análogos & derivados , RNA Mensageiro/biossínteseRESUMO
Huperzia saururus is reported in Argentinean popular medicine as a memory improver. Chemical studies have shown that the main constituents of the species are Lycopodium alkaloids. On the basis of this information, a purified alkaloid extract was obtained by alkaline extraction of the aerial parts. The aim of this work was to investigate the effects of intra-hippocampal administration of the purified alkaloid extract (AE) on memory retention in vivo, using a step down test, in order to correlate with previous results obtained in vitro in an electrophysiological model. The AE administration significantly increased the latency time in comparison to control animals. For treated animals the latency time was 37.61 +/- 2.84, 80.94 +/- 2.37, and 180.00 +/- 5.74 s for 1, 5, and 10 ng/rat, respectively versus 14.89 +/- 2.38 s for controls. According to these results there is a good relationship between the ethnopharmacological use and the effects hereby showed.
Assuntos
Alcaloides/farmacologia , Huperzia/química , Memória/efeitos dos fármacos , Extratos Vegetais/farmacologia , Alcaloides/administração & dosagem , Alcaloides/isolamento & purificação , Animais , Aprendizagem da Esquiva/efeitos dos fármacos , Comportamento Animal , Relação Dose-Resposta a Droga , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Masculino , Medicina Tradicional , Componentes Aéreos da Planta , Extratos Vegetais/administração & dosagem , Extratos Vegetais/isolamento & purificação , Ratos , Ratos WistarRESUMO
Huperzia saururus (Lam.) Trevis. (Lycopodiaceae) known as cola de quirquincho is used in folk medicine to improve memory. The cholinergic neurons of the basal forebrain, including those in the medial septum, and in the vertical limbs of the diagonal band of Broca and the nucleus basalis of Meynert, provide a major source of cholinergic enervation of the cortex and hippocampus. These neurons have also been shown to play an important role in learning and memory processes. Thus, the effects of this traditional Argentinean species were studied in relation to its activity on synaptic transmission in the hippocampus. The alkaloid extract obtained first by decoction of the aerial parts and by subsequent alkaline extraction, was purified by using a Sephadex LH 20 packed column. Electrophysiological experiments were developed with the purified extract (E(2)) on rat hippocampus slices, thus eliciting long-term potentiation (LTP). Results show a marked increase in the hippocampal synaptic plasticity. The threshold value for generation of LTP was 22 +/- 1.01 Hz on average for E(2), while for controls it was 86 +/- 0.92 Hz. All of these factors could explain the use of Huperzia saururus as a memory improver as is reported in the ethnomedicine.
Assuntos
Hipocampo/efeitos dos fármacos , Huperzia , Transmissão Sináptica/efeitos dos fármacos , Animais , Eletrofisiologia , Hipocampo/fisiologia , Técnicas In Vitro , Potenciação de Longa Duração/efeitos dos fármacos , Masculino , Extratos Vegetais/farmacologia , Ratos , Ratos WistarRESUMO
Huperzia saururus (Lam.) Trevis. (Lycopodiaceae) is used widely in Argentinian traditional medicine as an aphrodisiac and for memory improvement. An aqueous extract from the aerial parts was obtained by decoction, revealing the presence of alkaloids, among other constituents. By partition with organic solvent in alkaline media, alkaloids were extracted and then purified by gel permeation. We studied the anticholinesterase activity in vitro of the alkaloid extract using erythrocyte membranes and human serum as sources of acetylcholinesterase and pseudocholinesterase, respectively. The results show a marked inhibition of true acetylcholinesterase with an IC50 value of 0.58 microg/ml. Low inhibition of pseudocholinesterase was observed (IC50 value = 191 microg/ml). This shows a selectivity of the extract for the true acetylcholinesterase. Furthermore, chemical study of the bioactive extract was performed by GC-MS, revealing the presence of seven Lycopodium alkaloids, including some not identified previously: sauroxine, 6-hydroxylycopodine, N-acetyllycodine, lycopodine, lycodine, N-methyllycodine, and clavolonine. Further investigations will be undertaken in order to discover which compound/s are responsible for the aqueous extract's acetylcholinesterase activity.
Assuntos
Inibidores da Colinesterase/farmacologia , Huperzia , Fitoterapia , Extratos Vegetais/farmacologia , Acetilcolinesterase/efeitos dos fármacos , Acetilcolinesterase/metabolismo , Inibidores da Colinesterase/administração & dosagem , Inibidores da Colinesterase/uso terapêutico , Relação Dose-Resposta a Droga , Membrana Eritrocítica/enzimologia , Humanos , Concentração Inibidora 50 , Componentes Aéreos da Planta , Extratos Vegetais/administração & dosagem , Extratos Vegetais/uso terapêuticoRESUMO
The number, phenotype, localisation and development of intraepithelial lymphocytes (IEL) from duodenum (Du) and ileum (Il) were studied by immunohistochemistry (IHC) and light and electron microscopy in unweaned (0-7 weeks old) and six months-old pigs. Developmental changes at birth showed that 38% of the total lymphocytes in the villi were IEL, mainly of the CD2+CD4-CD8- double negative (DN) phenotype. That proportion rose to over 50% at week 5 after birth, resembling adult proportion, although still with fewer cells than in adult pigs. CD4+ cells appeared relatively early in life although they were confined to the lamina propria (LP) and CD8+ cells were found only in low numbers. In the villi of adult animals, almost half of the total number of lymphocytes were IEL (49% Du, 52% Il). Over half of these IEL (52% Du, 53% Il) showed the CD2+CD4-CD8+ phenotype and were localized at the epithelium's basement membrane. Numerous (43% Du, 42% Il) DN IEL were found grouped at the enterocyte nucleus level and relatively few (5% in Du and Il) granular IEL were found apically in the epithelium. These proportions were homogeneously maintained along the villi's tip, middle and bottom, suggesting that the IEL may have their origin in the LP. Therefore, the IEL compartment in the porcine intestine develops slowly with age and is actually composed by a heterogeneous population of cells (null, DN and CD8+). These results may explain the increased susceptibility of young animals to disease during the lactation period and should be taken into account when functional studies are carried out with IEL. The quantitative results of this paper established a model for studies on the effect of age, diet, normal flora, infection and oral immunization on the IEL of the gut.