RESUMO
A systematic study on the various sources of ion suppression in UHPLC-MS-MS analysis was carried out for 24 phenolic antioxidants in 6 different extracts of medicinal plants from Amazonia. The contributions of matrix effects, mobile-phase additives, analyte co-elution and electric charge competition during ionization to the global ion suppression were evaluated. Herein, the influence of mobile-phase additives on the ionization efficiency was found to be very pronounced, where ion suppression of approximately 90% and ion enhancement effects greater than 400% could be observed. The negative effect caused by the wrong choice of internal standard (IS) on quantitative studies was also evaluated and discussed from the perspective of ion suppression. This work also shows the importance of performing studies with this approach even for very similar matrices, such as varieties of medicinal plants from the same species, because different effects were observed for the same analyte.
Assuntos
Antioxidantes/análise , Fenóis/análise , Extratos Vegetais/química , Brasil , Catequina/análogos & derivados , Catequina/análise , Cromatografia Líquida/métodos , Ácidos Cumáricos/análise , Cumarínicos/análise , Flavonoides/análise , Plantas Medicinais , Espectrometria de Massas por Ionização por Electrospray/métodos , Estilbenos/análise , Espectrometria de Massas em Tandem/métodosRESUMO
It is well established that the involvement of reactive species in the pathophysiology of several neurological diseases, including phenylketonuria (PKU), a metabolic genetic disorder biochemically characterized by elevated levels of phenylalanine (Phe). In previous studies, we verified that PKU patients (treated with a protein-restricted diet supplemented with a special formula not containing L-carnitine and selenium) presented high lipid and protein oxidative damage as well as a reduction of antioxidants when compared to the healthy individuals. Our goal in the present study was to evaluate the effect of Phe-restricted diet supplemented with L-carnitine and selenium, two well-known antioxidant compounds, on oxidative damage in PKU patients. We investigated various oxidative stress parameters in blood of 18 treated PKU patients before and after 6 months of supplementation with a special formula containing L-carnitine and selenium. It was verified that treatment with L-carnitine and selenium was capable of reverting the lipid peroxidation, measured by thiobarbituric acid-reactive species, and the protein oxidative damage, measured by sulfhydryl oxidation, to the levels of controls. Additionally, the reduced activity of glutathione peroxidase was normalized by the antioxidant supplementation. It was also verified a significant inverse correlation between lipid peroxidation and L-carnitine blood levels as well as a significant positive correlation between glutathione peroxidase activity and blood selenium concentration. In conclusion, our results suggest that supplementation of L-carnitine and selenium is important for PKU patients since it could help to correct the oxidative stress process which possibly contributes, at least in part, to the neurological symptoms found in phenylketonuric patients.
Assuntos
Antioxidantes/farmacologia , Carnitina/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Fenilcetonúrias/fisiopatologia , Selênio/farmacologia , Adolescente , Antioxidantes/administração & dosagem , Antioxidantes/uso terapêutico , Carnitina/administração & dosagem , Carnitina/uso terapêutico , Suplementos Nutricionais , Humanos , Fenilcetonúrias/sangue , Fenilcetonúrias/dietoterapia , Espécies Reativas de Oxigênio/metabolismo , Selênio/administração & dosagem , Selênio/uso terapêutico , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , Adulto JovemRESUMO
In this study, oil-in-water formulations were optimized to determine sodium, potassium, calcium, magnesium, zinc, and iron in emulsified egg samples by flame atomic absorption spectrometry (FAAS). This method is simpler and requires fewer reagents when compared with other sample pre-treatment procedures and allows the calibration to be carried out using aqueous standards. Different oily phases such as corn oil, decyl oleate and octyl stearate were tested, as well as Tween 80, Triton X-100 and Triton 114 were analyzed as surfactants. The optimum type and proportion of formulations were determined and their use depended on the element studied. The emulsion preparation was performed by a conventional method that involves mixing both phases at 60 degrees C by magnetic stirring and phase inversion to change the water-to-oil ratio by increasing the volume of the surfactant-water external phase and correspondingly decreasing the volume of internal phase. The accuracy of the method was further confirmed by determining the metals in a whole egg powder CRM and recoveries ranged from 97.5% for Mg to 102.2% for Na, with relative standard deviations lower than 2.3%. The precision of the procedures was determined through repeatability (intra-day precision) and intermediate precision (inter-day). The repeatability presented RSD values lower than 4.2%. The intermediate precision was evaluated using the RSD and F-test. The RSD values to intermediate precision was lower than 5.3% and the computed F-values were lower than tabulated F-values, indicating no significant difference between the results obtained on different days. The proposed method including, sample emulsification for subsequent metal determination for FAAS, has proved to be sensitive, reproducible, simple and economical.
Assuntos
Ovos/análise , Análise de Alimentos/métodos , Metais/análise , Metais/química , Métodos Analíticos de Preparação de Amostras , Animais , Cálcio/análise , Cálcio/química , Calibragem , Galinhas , Emulsões , Análise de Alimentos/normas , Ferro/análise , Ferro/química , Magnésio/análise , Magnésio/química , Potássio/análise , Potássio/química , Padrões de Referência , Reprodutibilidade dos Testes , Sódio/análise , Sódio/química , Espectrofotometria Atômica , Zinco/análise , Zinco/químicaRESUMO
Aluminum (Al), a neurotoxic agent, has been associated with Alzheimer's disease (AD), which is characterized by cholinergic dysfunction in the central nervous system. In this study, we evaluated the effect of long-term exposure to aluminum on acetylcholinesterase (AChE) activity in the central nervous system in different brain regions, in synaptosomes of the cerebral cortex and in erythrocytes. The animals were loaded by gavage with AlCl(3) 50 mg/kg/day, 5 days per week, totalizing 60 administrations. Rats were divided into four groups: (1) control (C); (2) 50 mg/kg of citrate solution (Ci); (3) 50 mg/kg of Al plus citrate (Al + Ci), and (4) 50 mg/kg of Al (Al). AChE activity in striatum was increased by 15% for Ci, 19% for Al + Ci and 30% for Al, when compared to control (P < 0.05). The activity in hypothalamus increased 23% for Ci, 26% for Al + Ci and 28% for Al, when compared to control (P < 0.05). AChE activity in cerebellum, hippocampus and cerebral cortex was decreased by 11%, 23% and 21% respectively, for Al, when compared to the respective controls (P < 0.05). AChE activity in synaptosomes was increased by 14% for Al, when compared to control (P < 0.05). Erythrocyte AChE activity was increased by 17% for Al + Ci and 11% for Al, when compared to control (P < 0.05). These results indicate that Al affects at the same way AChE activity in the central nervous system and erythrocyte. AChE activity in erythrocytes may be considered a marker of easy access of the central cholinergic status.
Assuntos
Acetilcolinesterase/metabolismo , Alumínio/toxicidade , Sistema Nervoso Central/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Animais , Sistema Nervoso Central/enzimologia , Eritrócitos/enzimologia , Masculino , Ratos , Ratos WistarRESUMO
Acute effects of mercury on mouse blood, kidneys, and liver were evaluated. Mice received a single dose of mercuric chloride (HgCl2, 4.6 mg/kg, subcutaneously) for three consecutive days. We investigated the possible beneficial effects of antioxidant therapy (N-acetylcysteine (NAC) and diphenyl diselenide (PhSe)2) compared with the sodium salt of 2,3-dimercapto-1-propanesulfonic acid (DMPS), an effective chelating agent in HgCl2 exposure in mice. We also verified whether metallothionein (MT) induction might be involved in a possible mechanism of protection against HgCl2 poisoning and whether different treatments would modify MT levels and other toxicological parameters. The results demonstrated that HgCl2 exposure significantly inhibited delta-aminolevulinate dehydratase (delta-ALA-D) activity in liver and only DMPS treatment prevented the inhibitory effect. Mercuric chloride caused an increase in renal non-protein thiol groups (NPSH) and none of the treatments modified renal NPSH levels. Urea concentration was increased after HgCl2 exposure. NAC plus (PhSe)2 was partially effective in protecting against the effects of mercury. DMPS and (PhSe)2 were effective in restoring the increment in urea concentration caused by mercury. Thiobarbituric acid-reactive substances (TBARS), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) activities and ascorbic acid levels were not modified after mercury exposure. Mercuric chloride poisoning caused an increase in hepatic and renal MT levels and antioxidant treatments did not modify this parameter. Our data indicated a lack of therapeutic effect of the antioxidants tested.
Assuntos
Acetilcisteína/uso terapêutico , Antioxidantes/uso terapêutico , Sequestradores de Radicais Livres/uso terapêutico , Cloreto de Mercúrio/toxicidade , Intoxicação por Mercúrio/metabolismo , Metalotioneína/biossíntese , Animais , Derivados de Benzeno/uso terapêutico , Peso Corporal/efeitos dos fármacos , Quelantes/uso terapêutico , Quimioterapia Combinada , Injeções Subcutâneas , Rim/efeitos dos fármacos , Rim/metabolismo , Fígado/efeitos dos fármacos , Fígado/enzimologia , Masculino , Intoxicação por Mercúrio/etiologia , Intoxicação por Mercúrio/prevenção & controle , Camundongos , Compostos Organosselênicos/uso terapêutico , Sintase do Porfobilinogênio/antagonistas & inibidores , Sintase do Porfobilinogênio/metabolismo , Compostos de Sulfidrila/metabolismo , Unitiol/uso terapêutico , Ureia/sangueRESUMO
Long time ago aluminum (Al) was considered as a non-toxic element and its use had no restrictions. However, over the last two decades, scientific publications have indicated that Al is a toxic element. In line with this, aluminum accumulation in the organism is associated with a variety of human pathologies. Efficient therapeutics approach to treat Al intoxication are still not available, but there is a consensus that chelation therapy is the procedure to be used. However, the development of new chelating agents are highly desirable to improve the efficacy of the treatment of Al intoxication. The present study evaluates the chelating effect of two novel pyrimidines: 4-tricloromethyl-1-H-pyrimidin-2-one (THP) and (4-methyl-6-trifluoromethyl-6-pyrimidin-2-il)-hydrazine (MTPH) in a mice model of aluminum intoxication and compares their efficacy with those of desferrioxamine (DFO), a classical agent used for treat Al accumulation. The animals were exposed to aluminum by gavage (0.1 mmol aluminum/kg/day) 5 days/week for 4 weeks. At the end of this period, DFO was injected i.p. and the novel pyrimidines were given by gavage at 0.2 mmol/kg/day for five consecutive days. Aluminum concentration in tissues (brain, liver, kidney and blood) was determined by graphite furnace atomic absorption spectroscopy (GFAAS). The results showed that when administered by gavage, aluminum accumulated in the brain, kidney and liver of mice. MTPH was able to decrease aluminum levels in aluminum plus citrate animal groups, whereas THP was inefficient for this purpose. However, the novel pyrimidines used in this study were unable to surpass the aluminum chelating property of DFO. Thus, new studies must be performed utilizing other chelating agents which can decrease aluminum toxicity.
Assuntos
Alumínio/toxicidade , Quelantes/química , Modelos Químicos , Pirimidinas/química , Alumínio/química , Animais , Masculino , CamundongosRESUMO
The interaction of aluminium with some amino acids present in human blood was studied combining ion-chromatography (IC), atomic absorption spectrometry (AAS) and ultrafiltration (UF) techniques. An IC system for simultaneous determination of ornithine, lysine, glutamic acid, aspartic acid and tyrosine was developed. By adding aluminium to standard solutions of the amino acids and keeping the pH at 6 and 7 it was possible to verify that aluminium caused a reduction on the amino acid chromatographic signals. Similar experiment, carried out for copper showed the same behaviour (with different percentage of signal reductions) and validated the results for aluminium, considering that the interaction Cu-amino acid is well-established. The AAS analysis of sample fractions (500 microl) after the IC separation showed that aluminium (as copper as well) is not present in the fractions in which the amino acid peaks appear in the chromatogram. These approaches carried out with serum samples after UF showed that part of the "free" fraction of serum aluminium is distributed, besides other ligands, among these amino acids. It was found that in serum the affinity for aluminium followed the sequence Lys>Orn>Tyr>Glu approximately Asp.
Assuntos
Alumínio/química , Aminoácidos/sangue , Aminoácidos/química , Ácido Aspártico/química , Cromatografia , Cromatografia Líquida de Alta Pressão , Cromatografia por Troca Iônica , Cobre/química , Filtração , Ácido Glutâmico/química , Humanos , Concentração de Íons de Hidrogênio , Íons , Ligantes , Lisina/química , Metais/química , Ornitina/química , Espectrofotometria Atômica , Tirosina/química , o-Ftalaldeído/químicaRESUMO
The interference of the saline concentration of fluids for peritoneal dialysis and concentrates for hemodialysis on the Limulus amebocyte lysate (LAL) assay for endotoxins was investigated. The experiments were carried out individually with each substance that compose fluids for hemodialysis, to determine the possible inhibition or enhancement effects that they could cause on the LAL assay. The compositions were also assayed to investigate the possibility of synergistic effect. They were assayed by the gel-clot method from two different suppliers, and the samples that showed inhibition effect were also assayed by the chromogenic method. The samples were analysed at successive dilutions, with different LAL sensitivities, to satisfy the endotoxin limits of 5 EU/ml for the concentrate and 0.25 EU/ml for the fluid for dialysis peritoneal. The results showed that the major interference on the gel-clot assay occurs in presence of acetic acid and in concentrates containing acid acetic, even the pH being adjusted between 6.5 and 7.5. However, the test, after an adequate dilution, could be validating for all samples. Chromogenic test can be used for peritoneal dialysis fluids considering a limit of 0.25 EU/ml and sample dilution of eight times, but it cannot be used for concentrates for hemodialysis without further dilution. Considering the results and that the chromogenic is a more time-consuming method, endotoxins in fluids for hemodialysis can be satisfactorily assayed by the gel-clot method.
Assuntos
Soluções para Diálise/análise , Endotoxinas/análise , Teste do Limulus , Diálise Renal , Géis/análise , Concentração de Íons de Hidrogênio , Indicadores e Reagentes , Diálise Peritoneal , Padrões de Referência , Sais/análiseRESUMO
A method has been developed for the chromatographic determination of methanethiol (MT) and ethanethiol (ET) as contaminants in amino acid parenteral nutrition (PN) solutions. The clean-up of the samples before chromatographic analysis was investigated by solid-phase extraction (SPE) on pre-columns filled with polyethylene powder (PE), aluminium oxide (AlOx), silica (SiOx), or polyurethane foam (PUF) as adsorbents. The thiols were more efficiently separated from the matrices by SPE on PUF pre-columns. Simultaneous derivatization and elution with DTNB (5,5'-dithiobis(2-nitrobenzoic acid)) enabled further discrimination between MT and ET by reversed-phase HPLC with spectrophotometric detection. The retention times for the derivatized MT and ET species were 12.5 and 23.0 min, respectively. Recoveries from spiked PN samples were calculated to be approximately 90%, and the MT and ET content of commercial PN solutions was determined using the methodology described. Detection limits of 15 and 10 microg L(-1) were calculated for MT and ET, respectively.
Assuntos
Aminoácidos/química , Cromatografia Líquida de Alta Pressão , Nutrição Parenteral/normas , Compostos de Sulfidrila/análise , Óxido de Alumínio/química , Ácido Ditionitrobenzoico/química , Óxidos/química , Polietileno/química , Poliuretanos/química , Compostos de Silício/químicaRESUMO
Deproteinization of human serum was carried out on-line using a polyethylene powder cartridge as pre-column. The serum sample, after dilution, was injected into the chromatograph, and when passing through the cartridge the proteins were adsorbed by the polyethylene. A protein-free eluate was carried to the analytical column while the pre-column was washed with methanol and water to elute the adsorbed proteins by changing the pump channels. After washing, the pre-column was conditioned with the eluent to receive the next sample. Deproteinization was evaluated off- and on-line using pooled serum, and testing the protein residue after passing through a polyethylene cartridge with the Comassie Brilliant Blue reaction. Parameters such as serum volume, eluent and washing solution were investigated. Sodium, potassium, magnesium and calcium determinations were performed by high-performance liquid chromatography with conductimetric detection. The proposed method is suitable for the determination of these cations in serum samples without further treatments. The total analysis time was about 20 min and a linear range from 0.3 to 1.2 mg/l for sodium, potassium, magnesium and calcium was observed. The method was characterized by a precision of about 95% and recoveries from spiked samples were between 96 and 102%. The results for serum samples analysed by the proposed method were compared with photometry and atomic absorption spectrometry.