RESUMO
Leishmaniasis are diseases caused by parasites of the genus Leishmania and transmitted to humans by the bite of infected insects of the subfamily Phlebotominae. Current drug therapy shows high toxicity and severe adverse effects. Recently, two oligopeptidases (OPBs) were identified in Leishmania amazonensis, namely oligopeptidase B (OPB) and oligopeptidase B2 (OPB2). These OPBs could be ideal targets, since both enzymes are expressed in all parasite lifecycle and were not identified in human. This work aimed to identify possible dual inhibitors of OPB and OPB2 from L. amazonensis. The three-dimensional structures of both enzymes were built by comparative modelling and used to perform a virtual screening of ZINC database by DOCK Blaster server. It is the first time that OPB models from L. amazonensis are used to virtual screening approach. Four hundred compounds were identified as possible inhibitors to each enzyme. The top scored compounds were submitted to refinement by AutoDock program. The best results suggest that compounds interact with important residues, as Tyr490, Glu612 and Arg655 (OPB numbers). The identified compounds showed better results than antipain and drugs currently used against leishmaniasis when ADMET in silico were performed. These compounds could be explored in order to find dual inhibitors of OPB and OPB2 from L. amazonensis.
Assuntos
Antiprotozoários/química , Antiprotozoários/farmacologia , Leishmania/enzimologia , Proteínas de Protozoários/metabolismo , Serina Endopeptidases/metabolismo , Inibidores de Serina Proteinase/farmacologia , Sítios de Ligação , Simulação por Computador , Bases de Dados Factuais , Regulação Enzimológica da Expressão Gênica , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Proteínas de Protozoários/genética , Serina Endopeptidases/genética , SoftwareRESUMO
Phytase hydrolyzes phytic acid from the plant components of animal feed, releasing inorganic phosphorus. The phytase production by Aspergillus japonicus was optimized using Plackett-Burman designs (PBD), composite central rotational designs (CCRD), and response surface methodology from standard Czapek medium. The enzyme was applied in broiler chicken and laying hen foods. Analysis from PBD showed that KH2 PO2, MgSO4 · 7H2O, and yeast extract had significant influences on phytase secretion (p < 0.05). The best results from the CCRD experiments were obtained using (A) 0.040% KH2 PO4, (B) 0.050% MgSO4 · 7H2O, and (C) 0.040% yeast extract, enhancing in 49-53 U mg(-1) protein. The determination coefficient (R(2)) was 0.92 and Fcalc was 7.48 times greater than Flisted . Thus, the reduced coded model: Y (U mg-1) = 50.29 + 4.30A - 3.35(A)2 - 4.80(B)2 + 5.62C - 4.26(C)2 was considered predictive and statistically significant (p < 0.05). The optimized culture medium increased the phytase yield in 250%. A. japonicus phytase released high levels of Pi from broiler chicken and laying hen food. A. japonicus is an excellent phytase producer in a culture medium using inexpensive components and agricultural wastes. Therefore, these results provide sound arguments for the formulation of a low cost culture medium for phytase production.
Assuntos
6-Fitase/metabolismo , Ração Animal , Aspergillus/enzimologia , Animais , Aspergillus/crescimento & desenvolvimento , Galinhas , Meios de Cultura/química , Enzimas/metabolismo , Manipulação de Alimentos/métodosRESUMO
The main use of High Pressure Processing (HPP) in food processing is microorganism inactivation, and studies demonstrated that the characteristics of matrix and microorganisms can interfere on it. As the behavior of lactic acid bacteria exposed to different water activity (aw) levels in a meat product is still unclear, this study aimed to determine the effect of pressure, time, and aw to inactivate Latilactobacillus sakei, a pressure resistant lactic acid bacteria (LAB) in a meat emulsion model through a response surface methodology. The meat emulsion model was designed with adjusted aw (from 0.940 to 0.960) and was inoculated with a pressure resistant LAB and processed varying pressure (400-600 MPa) and time (180-480 s), following the Central Composite Rotational Design (CCRD). The inactivation of the microorganism ranged from 0.99 to 4.12 UFC/g depending on the applied condition. At studied conditions, according to the best fitting and most significant polynomial equation (R2 of 89.73 %), in a meat emulsion model, aw had no influenced on HPP inactivation on LAB (p > 0.05) and only pressure and holding time had significative impact on it. The results of experimental validation of the mathematical model were satisfactory, confirming the suitability of the model. The information obtained in the present study stands out the matrix, microorganism and process effects at HPP efficiency. The answers obtained support food processors in product development, process optimization and food waste reduction.
Assuntos
Lactobacillales , Latilactobacillus sakei , Produtos da Carne , Eliminação de Resíduos , Produtos da Carne/microbiologia , Água , Emulsões , Microbiologia de Alimentos , Contagem de Colônia Microbiana , Carne/microbiologiaRESUMO
PsaA, a candidate antigen for a vaccine against pneumonia, is well-conserved in all Streptococcus pneumoniae serotypes. A sequence of two-level experimental designs was used to evaluate medium composition and seed conditions to optimize the expression of soluble mature PsaA in E. coli. A face-centered central composite design was first used to evaluate the effects of yeast extract (5 and 23.6 g/L), tryptone (0 and 10 g/L), and glucose (1 and 10 g/L), with replicate experiments at the central point (14.3 g/L yeast extract, 5 g/L tryptone, 5.5 g/L glucose). Next, a central composite design was used to analyze the influence of NaCl concentration (0, 5, and 10 g/L) compared with potassium salts (9.4 g/L K(2)HPO(4)/2.2 g/L KH(2)PO(4)), and seed growth (7 and 16 h). Tryptone had no significant effect and was removed from the medium. Yeast extract and glucose were optimized at their intermediate concentrations, resulting in an animal-derived material-free culture medium containing 15 g/L yeast extract, 8 g/L glucose, 50 µg/mL kanamycin, and 0.4% glycerol, yielding 1 g/L rPsaA after 16 h induction at 25°C in shake flasks at 200 rpm. All the seed age and salt conditions produced similar yields, indicating that no variation had a statistically significant effect on expression. Instead of growing the seed culture for 16 h (until saturation), the process can be conducted with 7 h seed growth until the exponential phase. These results enhanced the process productivity and reduced costs, with 5 g/L NaCl being used rather than potassium salts.
Assuntos
Adesinas Bacterianas/biossíntese , Escherichia coli/genética , Expressão Gênica , Lipoproteínas/biossíntese , Meios de Cultura/química , Glicerol/metabolismo , Streptococcus pneumoniae/química , Streptococcus pneumoniae/genéticaRESUMO
Carbazole 1,9a-dioxygenase (CarA), the first enzyme in the carbazole degradation pathway used by Pseudomonas sp., was expressed in E. coli under different conditions defined by experimental design. This enzyme depends on the coexistence of three components containing [2Fe-2S] clusters: CarAa, CarAc, and CarAd. The catalytic site is present in CarAa. The genes corresponding to components of carbazole 1,9a-dioxygenase from P. stutzeri were cloned and expressed by salt induction in E. coli BL21-SI (a host that allows the enhancement of overexpressed proteins in the soluble fraction), using the vector pDEST™14. The expression of these proteins was performed under different induction conditions (cell concentration, temperature, and time), with the help of two-level factorial design. Cell concentration at induction (measured by absorbance at 600 nm) was tested at 0.5 and 0.8. After salt induction, expression was performed at 30 and 37°C, for 4 h and 24 h. Protein expression was evaluated by densitometry analysis. Expression of CarAa was enhanced by induction at a lower cell concentration and temperature and over a longer time, according to the analysis of the experimental design results. The results were validated at Abs (ind) = 0.3, 25°C, and 24 h, at which CarAa expression was three times higher than under the standard condition. The behavior of CarAc and CarAd was the inverse, with the best co-expression condition tested being the standard one (Abs (ind) = 0.5, T = 37°C, and t = 4 h). The functionality of the proteins expressed in E. coli was confirmed by the degradation of 20 ppm carbazole.
Assuntos
Proteínas de Bactérias/metabolismo , Dioxigenases/metabolismo , Escherichia coli/genética , Pseudomonas stutzeri/enzimologia , Proteínas de Bactérias/genética , Biotecnologia , Dioxigenases/genética , Escherichia coli/metabolismo , Fluoruracila/metabolismo , Redes e Vias Metabólicas , Pseudomonas stutzeri/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transformação BacterianaRESUMO
This study investigated the vitamin C content, total phenolic compounds (TPC), and the potential bioactivities (antioxidant, antiproliferative, antibacterial activities, and inhibition capacity against N-nitrosation) of seven neglected and underutilized species (NUS): culantro (Eryngium foetidum), false roselle (Hibiscus acetosella), roselle (Hibiscus sabdariffa), tree basil (Ocimum gratissimum), Barbados Gooseberry (Pereskia aculeata), purslane (Portulaca oleracea), and tannia (Xanthosoma sagittifolium). Phenolic-rich extracts were obtained by a sequential optimization strategy (Plackett-Burman and Central Composite Design). O. gratissimum presented the highest TPC and X. sagittifolium the greater total vitamin C content. Overall, the plant extracts presented promising bioactive capacities, as scavenging capacity against HOCl, H2O2 and ROO⢠induced oxidation. P. oleracea demonstrated the highest cytostatic effect against ovarian and kidney tumor cells. O. gratissimum effectively inhibited S. Choleraesuis growth. Maximum inhibition on n-nitrosation was showed by O. gratissimum and E. foetidum. These results highlight the studied NUS as sources of potential health-promoting compounds.
Assuntos
Hibiscus , Peróxido de Hidrogênio , Brasil , Extratos Vegetais/farmacologia , Folhas de PlantaRESUMO
This work investigated the growth of Kluyveromyces marxianus NRRL Y-7571 in solid-state fermentation in a medium composed of sugarcane bagasse, molasses, corn steep liquor and soybean meal within a packed-bed bioreactor. Seven experimental runs were carried out to evaluate the effects of flow rate and inlet air temperature on the following microbial rates: cell mass production, total reducing sugar and oxygen consumption, carbon dioxide and ethanol production, metabolic heat and water generation. A mathematical model based on an artificial neural network was developed to predict the above-mentioned microbial rates as a function of the fermentation time, initial total reducing sugar concentration, inlet and outlet air temperatures. The results showed that the microbial rates were temperature dependent for the range 27-50 degrees C. The proposed model efficiently predicted the microbial rates, indicating that the neural network approach could be used to simulate the microbial growth in SSF.
Assuntos
Reatores Biológicos/microbiologia , Kluyveromyces/crescimento & desenvolvimento , Aerobiose , Biomassa , Meios de Cultura/química , Fermentação , Modelos Teóricos , Oxigênio/metabolismo , Saccharum/metabolismo , Glycine max/metabolismo , Temperatura , Zea mays/metabolismoRESUMO
A potential application of inulinase in the food industry is the production of fructoligosaccharides (FOS) by the transfructosilation of sucrose. The FOSs present many interesting functional properties besides their ability to increase the shelf-life and flavor of many products. The use of an industrial medium represents a good alternative to producing inulinase at low cost, since the activity may improve, or at least remain the same, as that obtained using a synthetic medium. This work was an optimization study of the inulinase production by Kluyveromyces marxianus NRRL Y-7571 using industrial pre-treated culture medium in a bioreactor employing a sequential strategy of experimental design. Initially, a Plackett-Burman (Screening Design) design was used, where the studied variables were molasses, corn steep liquor, yeast extract concentration, and agitation and aeration rates. After the analysis of the effects, a central composite rotational design (CCRD) was carried out. The optimized condition for the inulinase production was: 250 g/l of molasses, 80 g/l of corn steep liquor, 6 g/l of yeast extract, 300 rpm of agitation and 1.5 vvm aeration rate, which resulted in an enzymatic activity of 1,317 +/- 65 U/ml.
Assuntos
Reatores Biológicos , Glicosídeo Hidrolases/biossíntese , Kluyveromyces/enzimologia , Meios de Cultura/química , Proteínas Fúngicas/biossíntese , Kluyveromyces/crescimento & desenvolvimentoRESUMO
The present work aimed to study the viability of the use of sugarcane molasses and corn steep liquor (CSL) in a sequential inulinase production performing an up-stream pretreatment of these agroindustrial residues. A sequential strategy was used applying three central composite rotatable designs (CCRDs) to optimise medium composition, followed by a down-stream step. The medium containing 150 g L(-1) molasses, 50 g L(-1) CSL and 6 g L(-1) yeast extract, yielded a maximum inulinase production of 1,294 +/- 7 U mL(-1), after 72 h of fermentation. A down-stream evaluation was carried out using an expanded bed of Streamline DAE resin (Pharmacia), with and without the up-stream treatment. The results showed that the enzyme could not be recovered from the non-pretreated medium, whereas a yield of 91% was obtained in the adsorption stage from the medium prepared with the up-stream treatment, showing the viability of producing the enzyme inulinase from agroindustrial residues using the integrated process.
Assuntos
Glicosídeo Hidrolases/biossíntese , Glicosídeo Hidrolases/isolamento & purificação , Engenharia Biomédica , Reatores Biológicos , Biotecnologia , Meios de Cultura , Estabilidade Enzimática , Fermentação , Tecnologia de Alimentos , Frutose/biossíntese , Glicosídeo Hidrolases/metabolismo , Concentração de Íons de Hidrogênio , Inulina/metabolismo , Cinética , Kluyveromyces/enzimologia , Kluyveromyces/crescimento & desenvolvimento , Melaço , Oligossacarídeos/biossíntese , Termodinâmica , Zea maysRESUMO
The production of inulinase employing agroindustrial residues as the substrate is a good alternative to reduce production costs and to minimize the environmental impact of disposing these residues in the environment. This study focused on the use of a phenomenological model and an artificial neural network (ANN) to simulate the inulinase production during the batch cultivation of the yeast Kluyveromyces marxianus NRRL Y-7571, employing a medium containing agroindustrial residues such as molasses, corn steep liquor and yeast extract. It was concluded that due to the complexity of the medium composition it was rather difficult to use a phenomenological model with sufficient accuracy. For this reason, an alternative and more cost-effective methodology based on ANN was adopted. The predictive capacity of the ANN was superior to that of the phenomenological model, indicating that the neural network approach could be used as an alternative in the predictive modeling of complex batch cultivations.
Assuntos
Agricultura/métodos , Reatores Biológicos , Glicosídeo Hidrolases/biossíntese , Microbiologia Industrial/instrumentação , Microbiologia Industrial/métodos , Biomassa , Meios de Cultura , Fermentação , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/metabolismo , Cinética , Kluyveromyces/metabolismo , Modelos Teóricos , Melaço , Redes Neurais de Computação , Reprodutibilidade dos Testes , Temperatura , Zea mays/metabolismoRESUMO
Objetivo: analisar as perspectivas e os desafios na concretização do dever de garantir a mais completa assistência no campo da saúde suplementar. Metodologia: pesquisa qualitativa de natureza descritiva, que analisou acervo documental composto pelos principais diplomas legais relacionados ao campo da saúde suplementar e a evolução da jurisprudência do Supremo Tribunal Federal e do Superior Tribunal de Justiça nessa área, notadamente sobre o fornecimento de medicamentos não registrados pela Anvisa ou uso off label de remédios registrados, e acerca da taxatividade mitigada do rol da ANS. Resultados: as avenças firmadas entre os planos de saúde e os beneficiários configuram relação contratual na qual o consumidor é a parte vulnerável, mas, por outro lado, detém informações que não são de conhecimento da operadora. Esta, por sua vez, detém conhecimentos técnicos sobre moléstias e tratamentos muito além da compreensão de um consumidor comum. Nesse mercado, em que predomina a assimetria informacional, avulta a importância da firme atuação da agência reguladora, entidade capaz de equilibrar o sistema, dando tratamento uniforme a questões que devem ser globalmente consideradas, tendo em vista a massa de consumidores de planos de saúde, levando em conta as especificidades de cada tipo de contrato, individual ou coletivo, anterior ou posterior à Lei nº 9.656/1998. Conclusão: a atuação do Legislativo deve levar em conta o delicado equilíbrio entre as necessidades e as possibilidades, bem como as características inerentes ao caráter privado da prestação, de forma suplementar, de serviços de natureza pública, ao passo que a intervenção do Judiciário para infirmar decisões da agência reguladora justifica-se apenas em casos de clara ilegalidade e, sobretudo, deve levar em conta a globalidade do sistema regulado, o que nem sempre é facilmente perceptível à vista de casos individualmente considerados.
Assuntos
Planos de Pré-Pagamento em Saúde , Jurisprudência , Direito à SaúdeRESUMO
Objetivo: analisar as perspectivas e os desafios na concretização do dever de garantir a mais completa assistência no campo da saúde suplementar. Metodologia: pesquisa qualitativa de natureza descritiva, que analisou acervo documental composto pelos principais diplomas legais relacionados ao campo da saúde suplementar e a evolução da jurisprudência do Supremo Tribunal Federal e do Superior Tribunal de Justiça nessa área, notadamente sobre o fornecimento de medicamentos não registrados pela Anvisa ou uso off label de remédios registrados, e acerca da taxatividade mitigada do rol da ANS. Resultados: as avenças firmadas entre os planos de saúde e os beneficiários configuram relação contratual na qual o consumidor é a parte vulnerável, mas, por outro lado, detém informações que não são de conhecimento da operadora. Esta, por sua vez, detém conhecimentos técnicos sobre moléstias e tratamentos muito além da compreensão de um consumidor comum. Nesse mercado, em que predomina a assimetria informacional, avulta a importância da firme atuação da agência reguladora, entidade capaz de equilibrar o sistema, dando tratamento uniforme a questões que devem ser globalmente consideradas, tendo em vista a massa de consumidores de planos de saúde, levando em conta as especificidades de cada tipo de contrato, individual ou coletivo, anterior ou posterior à Lei nº 9.656/1998.Conclusão: a atuação do Legislativo deve levar em conta o delicado equilíbrio entre as necessidades e as possibilidades, bem como as características inerentes ao caráter privado da prestação, de forma suplementar, de serviços de natureza pública, ao passo que a intervenção do Judiciário para infirmar decisões da agência reguladora justifica-se apenas em casos de clara ilegalidade e, sobretudo, deve levar em conta a globalidade do sistema regulado, o que nem sempre é facilmente perceptível à vista de casos individualmente considerados.
Objective: to analyze the perspectives and challenges in the realization of the duty to guarantee the most complete assistance in the field of supplementary health. Methods: qualitative research of descriptive nature that analyzed documentary collection composed of the main legal diplomas related to the field of supplementary health and the evolution of jurisprudence of the Supreme Court and the Superior Court of Justice in this area, especially about the supply of medicines not registered by Anvisa or off label use of registered medicines, and about the mitigated taxation of the ANS list. Results: the agreements signed between the health plans and the beneficiaries configure contractual relationship in which the consumer is the vulnerable party, but, on the other hand, holds information that is not known to the operator. This, in turn, has technical knowledge about diseases and treatments far beyond the understanding of a common consumer. In this market, in which information asymmetry predominates, the importance of the firm action of the regulatory agency, an entity capable of balancing the system, giving uniform treatment to issues that should be considered globally, in view of the mass of consumers of health plans, taking into account the specificities of each type of contract, individual or collective, before or after Bill9, 656/1998. Conclusion: the Legislative action must take into account the delicate balance between needs and possibilities, as well as the characteristics inherent in the private character of the provision, in a supplementary nature, of services of a public nature, whereas intervention by the judiciary to rule against decisions of the regulatory agency is justified only in cases of clear illegality and, above all, must take into account the whole of the regulated system, which is not always easily perceived in view of individual cases.
Objetivo: analizar las perspectivas y los desafíos en la realización del deber de garantizar la más completa asistencia en el campo de la salud complementaria. Metodología: investigación cualitativa de naturaleza descriptiva que analizó acervo documental compuesto por los principales diplomas legales relacionados al campo de la salud complementaria y la evolución de la jurisprudencia del Supremo Tribunal Federal y del Superior Tribunal de Justicia en esa área, especialmente sobre el suministro de medicamentos no registrados por la Anvisa o uso fuera de etiqueta de remedios registrados, y acerca de la taxactividad mitigada del rol de la ANS. Resultados: las avencias firmadas entre los planes de salud y los beneficiarios configuran relación contractual en la cual el consumidor es la parte vulnerable, pero, por otro lado, posee informaciones que no son de conocimiento del operador. Ésta, a su vez, posee conocimientos técnicos sobre enfermedades y tratamientos mucho más allá de la comprensión de un consumidor común. En este mercado, en el que predomina la asimetría informacional, la importancia de la firme actuación de la agencia reguladora, capaz de equilibrar el sistema, dando tratamiento uniforme a cuestiones que deben ser consideradas globalmente, teniendo en cuenta las especificidades de cada tipo de contrato, individual o colectivo, anterior o posterior a la Ley 9.656/1998. Conclusión: la actuación del Legislativo debe tener en cuenta el delicado equilibrio entre las necesidades y las posibilidades, así como las características inherentes al carácter privado de la prestación, en carácter suplementario, de servicios de carácter público, mientras que la intervención del poder judicial para invalidar las decisiones de la agencia reguladora sólo se justifica en casos de clara ilegalidad y, sobre todo, debe tener en cuenta la totalidad del sistema regulado, lo que no siempre es fácilmente perceptible a la vista de casos individuales considerados.
Assuntos
Direito SanitárioRESUMO
A simple, rapid and cost-effective sample preparation method for the determination of fumonisin B1 in rice was optimised using a strategy of sequential experimental designs. Initially, a Plackett-Burman design was applied to select the statistically significant variables for the determination of fumonisin B1, and then, a central composite rotatable design was used to define the optimal conditions of these variables. The method involves extraction with a 50% acetonitrile aqueous solution and glacial acetic acid, liquid-liquid partitioning with addition of anhydrous sodium sulphate and sodium chloride, followed by dispersive SPE clean-up with diatomaceous earth. The final extract was analysed by HPLC-FLD after precolumn derivatisation with ortho-phthaldialdehyde. The optimised method was validated for selectivity, linearity, matrix effect, limits of detection and quantification, trueness, and precision, and then applied to commercial samples of polished rice. This is the first report of the occurrence of fumonisin B1 in commercial samples of polished rice from the Southeast region of Brazil.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Fumonisinas/química , Oryza/químicaRESUMO
Background: Fuels and chemicals from renewable feedstocks have a growing demand, and acetone, butanol and ethanol (ABE) are some relevant examples. These molecules can be produced by the bacterial fermentation process using hydrolysates generated from lignocellulosic biomass as sugarcane bagasse, one of the most abundant sources of lignocellulosic biomass in Brazil. It originates as a residue in mills and distilleries in the production of sugar and ethanol. Results: In the present work, two strategies to generate hydrolysates of sugarcane bagasse were adopted. The fermentation of the first hydrolysate by Clostridium acetobutylicum DSM 6228 resulted in final concentrations of butanol, acetone and ethanol of 6.4, 4.5 and 0.6 g/L, respectively. On the other hand, the second hydrolysate presented better results (averages of 9.1, 5.5 and 0.8 g/L, respectively), even without the need for nutrient supplementation, since key elements were already present in the medium. The productivity (QP) and yield (YP/S) of the solvents with second hydrolysate were 0.5 g/Lâ¢h-1 and 0.4 g/g, respectively. Conclusions: The results described herein open new perspectives for the production of important molecules from residual lignocellulosic biomass for the fuel and chemical industries within the context of second-generation biorefinery.
Assuntos
Acetona/metabolismo , Celulose/metabolismo , Saccharum/metabolismo , Etanol/metabolismo , Butanóis/metabolismo , Brasil , Celulose/química , Saccharum/química , Clostridium acetobutylicum/metabolismo , Biocombustíveis , FermentaçãoRESUMO
Polyamines play an important role as growth promoters, in the maturation of the intestinal tract of infants and in the modulation of the immune response; consequently, the importance of polyamines in the diet of infants and adults is well established. However, information on the occurrence and levels of polyamines in cow's milk and milk from other species (including human milk) is contradictory. Furthermore, the methods used for the extraction of amines from these samples vary widely. Therefore, a method for the accurate analysis of amines in milk from different species and in milk products is needed. A sequential strategy of experimental designs was used to optimize the analytical extraction of polyamines from milk. The dependent variables that significantly affected the recoveries were screened through a Plackett-Burman design. Sulfosalicylic acid (SSA) provided better recoveries compared to trichloroacetic acid. Centrifugation time and speed during extraction were independent variables. The Central Composite Rotational Design used to optimize the dependent variables indicated that the optimal conditions for the extraction of polyamines were 40s vortexing, four successive extractions, and an SSA concentration of 1.5%. These conditions provided recoveries ≥ 92.8% and CV ≤ 5.8%. The experiments confirmed the predicted results, indicating that the optimized conditions and models used were effective in the determination of amines from milk. Samples of raw milk and milk beverages were observed to be poor sources of spermine and spermidine, whereas human milk contained both amines.
Assuntos
Leite Humano/química , Leite/química , Poliaminas/análise , Extração em Fase Sólida/métodos , Animais , Bovinos , Cromatografia Líquida de Alta Pressão/métodos , Cromatografia Líquida de Alta Pressão/normas , Feminino , Humanos , Extração em Fase Sólida/normasRESUMO
Inulinase belongs to an important class of enzymes as it can be used to produce high-fructose syrups by enzymatic hydrolysis of inulin and fructooligosaccharides, which has been used as functional food. This work aimed to carry out a partial characterization of the crude enzymatic extract of two different inulinases, obtained by solid-state fermentation (SSF) and submerged fermentation (SmF), using agroindustrial residues as substrates. The crude enzymatic extract obtained by SmF showed an optimal pH and temperature for hydrolytic activity of 4.5 and 55 degrees Celsius, respectively; and that obtained by SSF conducted to optimal pH and temperature of 5.0 and 55 degrees Celsius, respectively. Both enzymes presented high thermostability, with a D value of 230.4 h and 123.1 h for SmF and SSF, respectively. The inulinase produced by SmF showed highest stability at pH 4.4, while inulinase obtained by SSF was more stable at pH 4.8. The results showed that inulinase obtained by SmF is less susceptible to pH effect and the inulinase obtained by SSF is more resistant to higher temperatures.
Assuntos
Fermentação , Indústria Alimentícia , Imersão , Insulisina/metabolismo , Ativação Enzimática , Estabilidade Enzimática , Concentração de Íons de Hidrogênio , Insulisina/química , Insulisina/isolamento & purificação , Cinética , TemperaturaRESUMO
A mathematical model for an expanded bed column was developed to predict breakthrough curves for inulinase adsorption on Streamline SP ion-exchange adsorbent, using a crude fermentative broth with cells as the feedstock. The kinetics and mass transfer parameters were estimated using the PSO (particle swarm optimization) heuristic algorithm. The parameters were estimated for each expansion degree (ED) using three breakthrough curves at initial inulinase concentrations of 65.6UmL(-1). In sequence, the model parameters for an ED of 2.5 were validated using the breakthrough curve at an initial concentration of 114.4UmL(-1). The applicability of the validated model in process optimization was investigated, using the model as a process simulator and experimental design methodology to optimize the column and process efficiencies. The results demonstrated the usefulness of this methodology for expanded bed adsorption processes.
Assuntos
Cromatografia por Troca Iônica/métodos , Proteínas Fúngicas/química , Glicosídeo Hidrolases/química , Adsorção , Proteínas Fúngicas/isolamento & purificação , Glicosídeo Hidrolases/isolamento & purificação , Cinética , Kluyveromyces/enzimologia , Modelos TeóricosRESUMO
The production of a lipase by a wild-type Brazilian strain of Penicillium simplicissimum in solid-state fermentation of babassu cake, an abundant residue of the oil industry, was studied. The enzyme production reached about 90 U/g in 72 h, with a specific activity of 4.5 U/mg of total proteins. The crude lipase showed high activities at 35-60 degrees C and pH 4.0-6.0, with a maximum activity at 50 degrees C and pH 4.0-5.0. Enzyme stability was enhanced at pH 5.0 and 6.0, with a maximum half-life of 5.02 h at 50 degrees C and pH 5.0. Thus, this lipase shows a thermophilic and thermostable behavior, what is not common among lipases from mesophilic filamentous fungi. The crude enzyme catalysed the hydrolysis of triglycerides and p-nitrophenyl esters (C4:0-C18:0), preferably acting on substrates with medium-chain fatty acids. This non-purified lipase in addition to interesting properties showed a reduced production cost making feasible its applicability in many fields.
Assuntos
Ácidos/metabolismo , Fermentação , Lipase/biossíntese , Penicillium/enzimologia , Análise de Variância , Proteínas de Bactérias , Concentração de Íons de Hidrogênio , Penicillium/crescimento & desenvolvimento , Padrões de Referência , Análise de Regressão , Especificidade por Substrato , Propriedades de Superfície , Temperatura , Fatores de TempoRESUMO
Foi otimizada a metodologia analítica para determinar quatro flavonóis: miricetina (M), quercetina (Q), kanferol (K) e isoramnetina (I); e duas flavonas: luteolina (L) e apigenina (A) em amostras de pólen apícola desidratado produzidas em três estados brasileiros: Bahia (BA), São Paulo (SP) e Santa Catarina (SC). Delineamento Composto Central Rotacional (DCCR) foi utilizado para investigar os efeitos da concentração de HCl e do tempo de hidrólise sobre a concentração de cada flavonoide. A condição ótima encontrada para extração/hidrólise dos flavonoides estudados foi: 1,0M HCl/30 minutos. A melhor separação dos flavonoides foi conseguida com a coluna de fase reversa Symmetry C18 e fase móvel de metanol: tetrahidrofurano: água(26:57:17), acidificados com 0,3 de ácido fórmico em corrida isocrática (CLAE). As curvas-padrão apresentaram coeficientes de correlação superiores a 0,99. Os limites de detecção foram de 1,04, 0,88, 0,89,1,64, 0,82 e 1,19 μg/mL respectivamente para M, L, Q, A, I e K.