RESUMO
In the present study, microplastics (MPs) and metal concentrations were studied in the widely consumed tilapia (Oreochromis niloticus) fishes (n = 15) collected from a metropolitan reservoir of the Atoyac River basin, Mexico. Nearly 139 fibers were extracted from the gastrointestinal tracts and assessed using optical microscopy to evaluate their physical characteristics. The colour distribution of the fibers was mainly black (40%), blue (19%), red and white (14%). SEM images represented the surface morphology, while the elemental composition of the fibers was studied using EDX spectra. Polymer characterization using µFTIR aided in confirming the fibers as plastics (polyamide, polyester, and synthetic cellulose) and non-plastics (natural cellulose). Henceforth, â¼33% of the fibers, provisionally thought to be plastics, were natural fibers. The total metal concentrations were higher in the liver (259.24 mg kg-1) than the muscle (122.56 mg kg-1) due to diverse metabolic functions in the hepatic tissues. Human health risk assessment in terms of Hazard Index (HI) presented Pb and Zn values above unity in both adults and children, prompting regulatory measures. Statistical tests between MPs and fish biometry did not present any substantial correlations. The present study also affirmed that the presence of MPs and metals in fishes of a highly contaminated region is not only governed by their bioavailabilities, but also on the physiological characteristics of the individual organism.
Assuntos
Ciclídeos , Tilápia , Poluentes Químicos da Água , Animais , Criança , Monitoramento Ambiental , Água Doce , Humanos , México , Microplásticos , Plásticos , Poluentes Químicos da Água/análiseRESUMO
Reactive oxygen species are implicated as mediators of tissue damage in the acute renal failure induced by inorganic mercury. Astaxanthin (ASX), a carotenoid with potent antioxidant properties, exists naturally in various plants, algae, and seafoods. This paper evaluated the ability of ASX to prevent HgCl(2) nephrotoxicity. Rats were injected with HgCl(2) (0 or 5 mg/kg b.w., sc) 6h after ASX had been administered (0, 10, 25, or 50mg/kg, by gavage) and were killed 12h after HgCl(2) exposure. Although ASX prevented the increase of lipid and protein oxidation and attenuated histopathological changes caused by HgCl(2) in kidney, it did not prevent creatinine increase in plasma and delta-aminolevulinic acid dehydratase inhibition induced by HgCl(2). Glutathione peroxidase and catalase activities were enhanced, while superoxide dismutase activity was depressed in HgCl(2)-treated rats when compared to control and these effects were prevented by ASX. Our results indicate that ASX could have a beneficial role against HgCl(2) toxicity by preventing lipid and protein oxidation, changes in the activity of antioxidant enzymes and histopathological changes.
Assuntos
Antioxidantes/farmacologia , Nefropatias/induzido quimicamente , Nefropatias/prevenção & controle , Cloreto de Mercúrio/antagonistas & inibidores , Cloreto de Mercúrio/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Animais , Biomarcadores , Glutationa Transferase/metabolismo , Rim/efeitos dos fármacos , Rim/enzimologia , Testes de Função Renal , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Necrose/induzido quimicamente , Necrose/patologia , Sintase do Porfobilinogênio/metabolismo , Carbonilação Proteica/efeitos dos fármacos , Ratos , Ratos Wistar , Compostos de Sulfidrila/metabolismo , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , Xantofilas/farmacologiaRESUMO
BACKGROUND: Malondialdehyde (MDA) is one of the better-known secondary products of lipid peroxidation, and it is widely used as an indicator of cellular injury. The employment of the thiobarbituric acid reactive substances (TBARS) technique to measure MDA has received criticism over the years because of its lack of specificity. Thus, a specific and reliable method for MDA determination in plasma by high performance liquid chromatographic (HPLC)-VIS was validated; alkaline hydrolysis, n-butanol extraction steps and MDA stability were established. METHODS: The plasma underwent alkaline hydrolysis, acid deproteinization, derivatization with TBA and n-butanol extraction. After this, MDA was determined at 532 nm by HPLC-VIS. The method was applied to 65-year-old subjects from a retirement home. RESULTS: The assay was linear from 0.28 to 6.6 microM. The reproducibility of intra-run was obtained with CV%<4% and the inter run with CV%<11%. The accuracy (bias) ranged from 2 to -4.1%, and the recovery was greater than 95%. The limit of detection (LOD) and limit of quantification (LOQ) were 0.05 and 0.17 microM, respectively. For the stability test, every sample was stored at -20 degrees C. The plasma MDA was not stable when stored after the alkaline hydrolysis step, remained stable for 30 days after TBA derivatization storage and was stable for 3 days when stored after n-butanol extraction. The elderly subjects had MDA plasma levels of 4.45+/-0.81 microM for women and 4.60+/-0.95 microM for men. CONCLUSION: The method is reproducible, accurate, stable, sensitive, and can be used in the routines in clinical laboratories. Besides, this technique presents advantages such as the complete release of protein bound MDA with the alkaline hydrolysis step, the removal of interferents with n-butanol extraction, mobile phase without phosphate buffer and rapid analytical processes and run times.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Malondialdeído/sangue , Espectrofotometria/métodos , 1-Butanol/química , Idoso , Brasil , Feminino , Humanos , Hidrólise , Modelos Lineares , Masculino , Valores de Referência , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Hidróxido de Sódio/química , Solventes/química , Tiobarbitúricos/química , Fatores de TempoRESUMO
The 6.7 murine monoclonal antibody (mAb) recognizes the human CD18 antigen and is therefore of interest as an anti-inflammatory agent. The 6.7 heavy variable chain (VH) was humanized using the closest human germline sequence as the template on to which to graft the murine complementary determining regions (CDRs). Two versions were proposed, one in which the residue proline 45 of the murine form was maintained and another in which this framework residue was changed to the leucine found in the human sequence. These VH humanized versions were expressed in the yeast Pichia pastoris as hemi-humanized single-chain Fv (scFvs), with the VL from the murine antibody. The scFv from the murine antibody was also expressed. The binding activities of the murine and both hemi-humanized scFvs were determined by flow cytometry analysis. All the constructions were able to recognize human lymphocytes harboring CD18, indicating successful humanization with transfer of the original binding capability. Some differences between the two hemi-humanized versions were observed. The method used was simple and straightforward, with no need for refined structural analyses and could be used for the humanization of other antibodies.
Assuntos
Antígenos CD18/imunologia , Região Variável de Imunoglobulina/química , Sequência de Aminoácidos , Animais , Clonagem Molecular , Humanos , Região Variável de Imunoglobulina/genética , Região Variável de Imunoglobulina/imunologia , Camundongos , Modelos Moleculares , Dados de Sequência Molecular , Pichia/genética , Homologia de Sequência de AminoácidosRESUMO
Monoclonal antibodies are one of the most important products of biotechnology and laboratories and companies all over the world are pursuing their large-scale production. Herein we report a protocol for hybridoma cell cultivation over small glass cylinders inside a 3 liter bioreactor vessel which leads to the production and purification--in order of grams--of one MAb intended for human therapeutic use. This protocol proved to be simple, reproducible and cost effective. Three trials are reported: the first two using conventionally serum-supplemented medium culture and producing 3.15 and 2.1 g of purified MAb in 30 and 21 days respectively, and the third one using serum-free medium culture and producing 6 g of purified MAb in 36 days. We have ascertained the stability of the hybridoma by its cloning directly in serum-free medium. The downstream processing of the serum-free trial was done in a single step, concentrating large volumes of supernatant while simultaneously purifying the antibody.