RESUMO
Using turbidity measurements to quantify bacterial growth is a common and well-established practice in microbiology. Automated devices offering high throughput analyses have largely contributed to the increase of its use. A main difficulty of this method is that it detects growth only at late exponential phase, making turbidity measurements limited for studies focussing on low cell numbers. This work proposes an improved estimator for the probability of growth of individual cells using turbidity-based measurements, when the initial number of cells is low and random. We modify the currently used estimator for the expected cell number per well, a Poisson-parameter, and show that an optimal scenario is when ca 20% of the wells do not become turbid, resulting in improved accuracy and precision.
Assuntos
Bactérias/crescimento & desenvolvimento , Nefelometria e Turbidimetria/métodos , Projetos de Pesquisa , ProbabilidadeRESUMO
Hereditary angioedema (HAE) is a rare autosomal dominant disease due to C1 esterase inhibitor deficiency (C1-INH). The disease is characterized by subcutaneous and submucosal edema in the absence of urticaria due to the accumulation of bradykinin. This descriptive study aimed to evaluate the clinical characteristics of patients with a confirmed diagnosis of HAE referred to our Outpatient Clinic between December 2009 and November 2017. Fifty-one patients (38 F, 13 M) with a mean age of 32 years (range: 7-70 y) were included. Family history of HAE was reported in 70% (36/51) of the cases; 33/46 patients became symptomatic by 18 years of age. The median time between onset of symptoms and diagnosis was 13 years (3 mo-50 y). The most frequent triggering factors for attacks were stress (74.4%), trauma (56.4%), and hormonal variations (56%). The main symptoms were subcutaneous edema in 93.5% (43/46) of patients, gastrointestinal symptoms in 84.8% (39/46), and obstruction in the upper airways in 34.8% (16/46). Hospitalization occurred in 65.2%, of whom 13.3% had to be transferred to the Intensive Care Unit. Prophylactic treatment was instituted in 87% (40/46) of patients, and 56.5% (26/46) required additional treatment to control attacks. Owing to our data collection over a period of 8 years, a significant number of patients were identified by this HAE reference center. Despite early recognition and prophylactic treatment, a high percentage of patients were hospitalized. HAE is still diagnosed late, reinforcing the need for more reference centers specialized in diagnosis and educational projects for health professionals.
Assuntos
Proteína Inibidora do Complemento C1/análise , Angioedema Hereditário Tipos I e II/sangue , Angioedema Hereditário Tipos I e II/etiologia , Adolescente , Adulto , Idade de Início , Idoso , Antifibrinolíticos/uso terapêutico , Criança , Antagonistas de Estrogênios/uso terapêutico , Feminino , Angioedema Hereditário Tipos I e II/tratamento farmacológico , Angioedema Hereditário Tipos I e II/prevenção & controle , Hospitalização , Humanos , Masculino , Pessoa de Meia-Idade , Nefelometria e Turbidimetria/métodos , Profilaxia Pós-Exposição/métodos , Fatores Desencadeantes , Trauma Psicológico/complicações , Fatores de Risco , Estresse Psicológico/complicações , Resultado do Tratamento , Adulto JovemRESUMO
The study of the stability of medicines is mandated by the International Conference on Harmonization and the World Health Organization. Rifaximin, an antimicrobial marketed in the form of tablets, has no record of stability studies. Thus, the objective of the present work was to investigate the behavior and stability of rifaximin tablets for 6 months under simultaneous conditions of temperature and humidity by UV, IR, HPLC, and turbidimetry techniques. After 6 months of stability study, rifaximin tablets were shown to obey zero-order kinetics when analyzed by physicochemical methods and second-order kinetics when analyzed by a microbiological method. However, the UV method was not suitable for the evaluation of rifaximin. IR, HPLC, and turbidimetry methods can already be used to evaluate the stability of rifaximin tablets. It is important to analyze products with more than one type of method before releasing results mainly in the case of antimicrobial products in which the association of physicochemical and microbiological techniques must be a rule. Rifaximin tablets can be considered stable after 6 months under conditions of 40 ± 2°C and 75 ± 5% relative humidity.
Assuntos
Antibacterianos/química , Rifamicinas/química , Cromatografia Líquida de Alta Pressão/métodos , Estabilidade de Medicamentos , Umidade , Cinética , Nefelometria e Turbidimetria/métodos , Rifaximina , Espectrofotometria Infravermelho/métodos , Espectrofotometria Ultravioleta/métodos , Comprimidos , TemperaturaRESUMO
Hereditary angioedema (HAE) is a rare autosomal dominant disease due to C1 esterase inhibitor deficiency (C1-INH). The disease is characterized by subcutaneous and submucosal edema in the absence of urticaria due to the accumulation of bradykinin. This descriptive study aimed to evaluate the clinical characteristics of patients with a confirmed diagnosis of HAE referred to our Outpatient Clinic between December 2009 and November 2017. Fifty-one patients (38 F, 13 M) with a mean age of 32 years (range: 7-70 y) were included. Family history of HAE was reported in 70% (36/51) of the cases; 33/46 patients became symptomatic by 18 years of age. The median time between onset of symptoms and diagnosis was 13 years (3 mo-50 y). The most frequent triggering factors for attacks were stress (74.4%), trauma (56.4%), and hormonal variations (56%). The main symptoms were subcutaneous edema in 93.5% (43/46) of patients, gastrointestinal symptoms in 84.8% (39/46), and obstruction in the upper airways in 34.8% (16/46). Hospitalization occurred in 65.2%, of whom 13.3% had to be transferred to the Intensive Care Unit. Prophylactic treatment was instituted in 87% (40/46) of patients, and 56.5% (26/46) required additional treatment to control attacks. Owing to our data collection over a period of 8 years, a significant number of patients were identified by this HAE reference center. Despite early recognition and prophylactic treatment, a high percentage of patients were hospitalized. HAE is still diagnosed late, reinforcing the need for more reference centers specialized in diagnosis and educational projects for health professionals.
Assuntos
Humanos , Masculino , Feminino , Criança , Adolescente , Adulto , Pessoa de Meia-Idade , Idoso , Adulto Jovem , Proteína Inibidora do Complemento C1/análise , Angioedema Hereditário Tipos I e II/etiologia , Angioedema Hereditário Tipos I e II/sangue , Estresse Psicológico/complicações , Fatores Desencadeantes , Fatores de Risco , Resultado do Tratamento , Idade de Início , Antagonistas de Estrogênios/uso terapêutico , Angioedema Hereditário Tipos I e II/prevenção & controle , Angioedema Hereditário Tipos I e II/tratamento farmacológico , Profilaxia Pós-Exposição/métodos , Trauma Psicológico/complicações , Hospitalização , Antifibrinolíticos/uso terapêutico , Nefelometria e Turbidimetria/métodosRESUMO
PURPOSE:: Avastin® (bevacizumab) is an anti-vascular endothelial growth factor (VEGF) monoclonal antibody given as an off-label drug by intravitreal administration for treatment of ocular diseases. The drug's clinical application and its cost-benefit profile has generated demand for its division into single-use vials to meet the low volume and low-cost doses necessary for intraocular administration. However, the safety of compounding the drug in single-use vials is still under discussion. In this study, the stability and efficacy of Avastin® repacked in individual single-use glass vials and glass ampoules by external compounding pharmacies were evaluated. METHODS:: Polyacrylamide gel electrophoresis (PAGE), size-exclusion chromatography (SEC), dynamic light scattering (DLS), and turbidimetry were selected to detect the formation of aggregates of various sizes. Changes in bevacizumab biological efficacy were investigated by using an enzyme-linked immunosorbent assay (ELISA). RESULTS:: Repacked and reference bevacizumab showed similar results when analyzed by PAGE. By SEC, a slight increase in high molecular weight aggregates and a reduction in bevacizumab monomers were observed in the products of the three compounding pharmacies relative to those in the reference bevacizumab. A comparison of repacked and reference SEC chromatograms showed that the mean monomer loss was ≤1% for all compounding pharmacies. Protein aggregates in the nanometer- and micrometer-size ranges were not detected by DLS and turbidimetry. In the efficacy assay, the biological function of repacked bevacizumab was preserved, with <3% loss of VEGF binding capacity relative to that of the reference. CONCLUSION:: The results showed that bevacizumab remained stable after compounding in ampoules and single-use glass vials; no significant aggregation, fragmentation, or loss of biological activity was observed.
Assuntos
Inibidores da Angiogênese/química , Bevacizumab/química , Embalagem de Medicamentos , Controle de Qualidade , Inibidores da Angiogênese/análise , Bevacizumab/análise , Cromatografia em Gel/métodos , Estabilidade de Medicamentos , Difusão Dinâmica da Luz/métodos , Eletroforese em Gel de Poliacrilamida/métodos , Ensaio de Imunoadsorção Enzimática/métodos , Injeções Intravítreas , Peso Molecular , Nefelometria e Turbidimetria/métodos , Fator A de Crescimento do Endotélio Vascular/análiseRESUMO
ABSTRACT Purpose: Avastin® (bevacizumab) is an anti-vascular endothelial growth factor (VEGF) monoclonal antibody given as an off-label drug by intravitreal administration for treatment of ocular diseases. The drug's clinical application and its cost-benefit profile has generated demand for its division into single-use vials to meet the low volume and low-cost doses necessary for intraocular administration. However, the safety of compounding the drug in single-use vials is still under discussion. In this study, the stability and efficacy of Avastin® repacked in individual single-use glass vials and glass ampoules by external compounding pharmacies were evaluated. Methods: Polyacrylamide gel electrophoresis (PAGE), size-exclusion chromatography (SEC), dynamic light scattering (DLS), and turbidimetry were selected to detect the formation of aggregates of various sizes. Changes in bevacizumab biological efficacy were investigated by using an enzyme-linked immunosorbent assay (ELISA). Results: Repacked and reference bevacizumab showed similar results when analyzed by PAGE. By SEC, a slight increase in high molecular weight aggregates and a reduction in bevacizumab monomers were observed in the products of the three compounding pharmacies relative to those in the reference bevacizumab. A comparison of repacked and reference SEC chromatograms showed that the mean monomer loss was ≤1% for all compounding pharmacies. Protein aggregates in the nanometer- and micrometer-size ranges were not detected by DLS and turbidimetry. In the efficacy assay, the biological function of repacked bevacizumab was preserved, with <3% loss of VEGF binding capacity relative to that of the reference. Conclusion: The results showed that bevacizumab remained stable after compounding in ampoules and single-use glass vials; no significant aggregation, fragmentation, or loss of biological activity was observed.
RESUMO Objetivos: Avastin® (bevacizumabe) é um anticorpo monoclonal inibidor do fator de crescimento endotelial de vasos (VEGF) utilizado "off-label" por meio de administração intravítrea para o tratamento de doenças oculares. A sua aplicação clínica associada ao custo-benefício do medicamento gerou uma demanda para seu fracionamento em frascos de dose única para utilização pela via intraocular. No entanto, a segurança do fracionamento do anticorpo em frascos de dose única ainda é alvo de discussão. Neste trabalho, a estabilidade e a eficácia do Avastin® fracionado em frascos ou ampolas de vidro de dose unitária por farmácias de manipulação do mercado foram avaliadas. Métodos: As técnicas de eletroforese em gel de poliacrilamida (PAGE), cromatografia por exclusão de tamanho (SEC), espalhamento dinâmico da luz (DLS) e turbidimetria foram empregadas para avaliar a formação de agregados de diferentes tamanhos. Alterações na atividade biológica do bevacizumabe foram estudadas utilizando ELISA. Resultados: Amostras referência e do bevacizumabe fracionado apresentaram resultados semelhantes quando analisado por gel de poliacrilamida. Por cromatografia por exclusão de tamanho, um pequeno aumento na quantidade de agregados de alta massa molar seguido de uma redução nos monômeros do bevacizumabe foram observados para as amostras das três farmácias de manipulação quando comparado ao referência. A comparação dos cromatogramas mostrou uma quantidade de redução do monômero inferior a 1% para todas as amostras fracionadas. Por espalhamento dinâmico da luz e turbidimetria, não foram detectados agregados de proteína na faixa de tamanho de micrômetro e nanômetro. No ensaio de eficácia, o bevacizumabe fracionado preservou sua função biológica pois apresentou menos de 3% de perda na capacidade de ligação ao VEGF quando comparado ao referência. Conclusão: Este estudo sugere que o bevacizumabe se mantem estável após fracionamento em ampolas e frascos de vidro de dose unitária pois não foram observadas agregação e/ou fragmentação de proteínas e perda de atividade biológica em quan tidades significativas.
Assuntos
Controle de Qualidade , Inibidores da Angiogênese/química , Embalagem de Medicamentos , Bevacizumab/química , Ensaio de Imunoadsorção Enzimática/métodos , Cromatografia em Gel/métodos , Inibidores da Angiogênese/análise , Fator A de Crescimento do Endotélio Vascular/análise , Estabilidade de Medicamentos , Eletroforese em Gel de Poliacrilamida/métodos , Injeções Intravítreas , Bevacizumab/análise , Difusão Dinâmica da Luz/métodos , Peso Molecular , Nefelometria e Turbidimetria/métodosRESUMO
Objective To evaluate under laboratory conditions, the removal efficiency of turbidity and E. coli of two household water filters: LifeStraw® family (MF) and ceramic pot filter (CPF). Methods The two systems were operated over 6 months using two identical control units per system, treating 7.5 L/d of a synthetic substrate used as raw water. The turbidity of the substrate was adjusted with Kaolinite and the E. coli concentration, with a replica of the ATCC 95922 strain. The differences of effluent quality of the systems, in terms of turbidity and E. coli, were evaluated with Analysis of Variance (ANOVA). Operative and maintenance aspects, that could limit or enhance the use of the systems, were also considered in the evaluation. Results The water synthetic substrate quality had an average of 32.2 ± 2.8 NTU for turbidity and 3,9x105 UFC/100 mL for E. coli. Both systems reduce the turbidity to values below 2 NTU with an inactivation of 100 % of E. coli. Statistical differences were found between the systems in terms of turbidity removal, MF being more efficient than the CPF (99,2 ± 0.4 % and 97.6 % ± 1.14, respectively). Conclusions Both systems are suitable for household water supply treatment, acheiving the water quality standards established by Colombian regulations. The MF was more efficient for suspended solids removal and filtration rate, but when economic, operative, and maintenance aspects along with social acceptability and lifespan are considered, the CPF seems more suitable, especially in rural areas.
Assuntos
Cerâmica , Filtração/instrumentação , Purificação da Água/instrumentação , Abastecimento de Água , Análise de Variância , Escherichia coli/isolamento & purificação , Nefelometria e Turbidimetria/métodos , Microbiologia da ÁguaRESUMO
The present study aimed to determine the feasibility of using bovine teeth as a suitable alternative for human teeth, in experiments involving in vitro endotoxin contamination. Twenty bovine central incisors and 20 human single-root premolars had their dental crowns removed and root lengths set at 16 mm. Root canals were prepared up to #60 K-file size and sterilized with cobalt-60 gamma irradiation (20 kGy, 6 h). The teeth were randomly divided into four groups: G1-bovine teeth (bovine negative control, n = 10), G2-human teeth (human negative control, n = 10), G3-bovine teeth, inoculated with Escherichia coli (055:B55) LPS, and G4-human teeth inoculated with E. coli LPS. The G1 and G2 groups were exposed to apyrogenic water. After the teeth had been incubated at 37 °C and atmospheric humidity for 24 h, the samples of solutions in the main canals were collected with apyrogenic absorbent paper tips. LPS levels were quantified using Limulus Amebocyte Lysate assay. The data obtained were statistically analyzed using one-way ANOVA, with a significance level of 5%. A high amount of endotoxin was detected in the inoculated human teeth (G4) when compared to the sterilized teeth (G2), as well as in the inoculated bovine teeth (G3) when compared to the inoculated human teeth (G4). However, there was no statistical difference between bovine teeth before and after the E. coli endotoxin inoculation. Therefore, under the mentioned experimental conditions, the use of bovine teeth should not be a choice for laboratory research on endotoxin contamination.
Assuntos
Cavidade Pulpar/microbiologia , Lipopolissacarídeos/análise , Análise de Variância , Animais , Bovinos , Cobalto/química , Escherichia coli , Estudos de Viabilidade , Humanos , Técnicas In Vitro , Teste do Limulus , Nefelometria e Turbidimetria/métodos , Distribuição Aleatória , Reprodutibilidade dos Testes , Fatores de TempoRESUMO
Sulfated and pyruvylated galactans are the major soluble polysaccharides produced by seaweeds of the Bryopsidales. Their backbones have a complex and variable pattern of substitution which, until now, has only been elucidated for a few species. Methods for determination of sulfate and pyruvic acid content, and chemical strategies to determine their position in the galactan chain are outlined here. These methods can also be applied to other sulfated and/or pyruvylated polysaccharides.