Cost analysis based on bioreactor cultivation conditions: Production of a soluble recombinant protein using Escherichia coli BL21(DE3).

The impact of cultivation strategy on the cost of recombinant protein production is crucial for defining cost-effective bioreactor operation conditions. This paper presents a methodology to estimate and compare cost impacts related to utilities as well as medium composition, using simple design equations and accessible data. Data from batch bioreactor cultures were used as case study involving the production of pneumococcal surface protein A, a soluble recombinant protein, employing E. coli BL21(DE3). Cultivation strategies and corresponding process costs covered a wide range of operational conditions, including different media, inducers, and temperatures. The core expenses were related to the medium and cooling. When the price of peptone was above the threshold value of US$ 30/kg, defined medium became the best choice. IPTG and temperatures around 32 °C led to shorter cultures and lower PspA4Pro production costs. The procedure offers a simple, accessible theoretical tool to identify cost-effective production strategies using bioreactors.

Cost analysis based on bioreactor cultivation conditions: production of a soluble recombinant protein using Escherichia coli BL21(DE3)

The impact of cultivation strategy on the cost of recombinant protein production is crucial for defining cost-effective bioreactor operation conditions. This paper presents a methodology to estimate and compare cost impacts related to utilities as well as medium composition, using simple design equations and accessible data. Data from batch bioreactor cultures were used as case study involving the production of pneumococcal surface protein A, a soluble recombinant protein, employing E. coli BL21(DE3). Cultivation strategies and corresponding process costs covered a wide range of operational conditions, including different media, inducers, and temperatures. The core expenses were related to the medium and cooling. When the price of peptone was above the threshold value of US$ 30/kg, defined medium became the best choice. IPTG and temperatures around 32°C led to shorter cultures and lower PspA4Pro production costs. The procedure offers a simple, accessible theoretical tool to identify cost-effective production strategies using bioreactors.

Cost analysis based on bioreactor cultivation conditions: production of a soluble recombinant protein using Escherichia coli BL21(DE3)

The impact of cultivation strategy on the cost of recombinant protein production is crucial for defining cost-effective bioreactor operation conditions. This paper presents a methodology to estimate and compare cost impacts related to utilities as well as medium composition, using simple design equations and accessible data. Data from batch bioreactor cultures were used as case study involving the production of pneumococcal surface protein A, a soluble recombinant protein, employing E. coli BL21(DE3). Cultivation strategies and corresponding process costs covered a wide range of operational conditions, including different media, inducers, and temperatures. The core expenses were related to the medium and cooling. When the price of peptone was above the threshold value of US$ 30/kg, defined medium became the best choice. IPTG and temperatures around 32°C led to shorter cultures and lower PspA4Pro production costs. The procedure offers a simple, accessible theoretical tool to identify cost-effective production strategies using bioreactors.

Production and purification of an untagged recombinant pneumococcal surface protein A (PspA4Pro) with high-purity and low endotoxin content.

Streptococcus pneumoniae is the main cause of pneumonia, meningitis, and other conditions that kill thousands of children every year worldwide. The replacement of pneumococcal serotypes among the vaccinated population has evidenced the need for new vaccines with broader coverage and driven the research for protein-based vaccines. Pneumococcal surface protein A (PspA) protects S. pneumoniae from the bactericidal effect of human apolactoferrin and prevents complement deposition. Several studies indicate that PspA is a very promising target for novel vaccine formulations. Here we describe a production and purification process for an untagged recombinant fragment of PspA from clade 4 (PspA4Pro), which has been shown to be cross-reactive with several PspA variants. PspA4Pro was obtained using lactose as inducer in Phytone auto-induction batch or glycerol limited fed-batch in 5-L bioreactor. The purification process includes two novel steps: (i) clarification using a cationic detergent to precipitate contaminant proteins, nucleic acids, and other negatively charged molecules as the lipopolysaccharide, which is the major endotoxin; and (ii) cryoprecipitation that eliminates aggregates and contaminants, which precipitate at -20 °C and pH 4.0, leaving PspA4Pro in the supernatant. The final process consisted of cell rupture in a continuous high-pressure homogenizer, clarification, anion exchange chromatography, cryoprecipitation, and cation exchange chromatography. This process avoided costly tag removal steps and recovered 35.3 ± 2.5% of PspA4Pro with 97.8 ± 0.36% purity and reduced endotoxin concentration by >99.9%. Circular dichroism and lactoferrin binding assay showed that PspA4Pro secondary structure and biological activity were preserved after purification and remained stable in a wide range of temperatures and pH values.

Production and purification of an untagged recombinant pneumococcal surface protein A (PspA4Pro) with high-purity and low endotoxin content

Streptococcus pneumoniae is the main cause of pneumonia, meningitis, and other conditions that kill thousands of children every year worldwide. The replacement of pneumococcal serotypes among the vaccinated population has evidenced the need for new vaccines with broader coverage and driven the research for protein-based vaccines. Pneumococcal surface protein A (PspA) protects S. pneumoniae from the bactericidal effect of human apolactoferrin and prevents complement deposition. Several studies indicate that PspA is a very promising target for novel vaccine formulations. Here we describe a production and purification process for an untagged recombinant fragment of PspA from clade 4 (PspA4Pro), which has been shown to be cross-reactive with several PspA variants. PspA4Pro was obtained using lactose as inducer in Phytone auto-induction batch or glycerol limited fed-batch in 5-L bioreactor. The purification process includes two novel steps: (i) clarification using a cationic detergent to precipitate contaminant proteins, nucleic acids, and other negatively charged molecules as the lipopolysaccharide, which is the major endotoxin; and (ii) cryoprecipitation that eliminates aggregates and contaminants, which precipitate at -20 A degrees C and pH 4.0, leaving PspA4Pro in the supernatant. The final process consisted of cell rupture in a continuous high-pressure homogenizer, clarification, anion exchange chromatography, cryoprecipitation, and cation exchange chromatography. This process avoided costly tag removal steps and recovered 35.3 +/- 2.5% of PspA4Pro with 97.8 +/- 0.36% purity and reduced endotoxin concentration by > 99.9%. Circular dichroism and lactoferrin binding assay showed that PspA4Pro secondary structure and biological activity were preserved after purification and remained stable in a wide range of temperatures and pH values.

Análise dos aspectos antropométricos em jovens atletas de alto rendimento praticantes da modalidade futebol / Analysis of the anthropometric features in young athletes of high yield practitioners of the soccer modality

As medidas antropométricas estão sendo amplamente utilizadas, na verificação das adaptações em resposta ao treinamento, para acompanhamento e desenvolvimento na seleção de atletas. Este estudo teve como objetivo analisar e caracterizar a composição corporal de atletas da categoria de base de um clube de alto rendimento do Estado do Rio de Janeiro, na modalidade futebol participantes do período competitivo do ano de 2008. Todos os dados da amostra foram coletados no término da temporada em dezembro. A amostra desse estudo foi caracterizada por um grupo de 198 futebolistas do sexo masculino divididos em 5 grupos: Sub19 n=40, Sub17 n=42, Sub15 n=41, Sub13 n=35 e Sub11 n=40. Foi utilizada estatística descritiva (média ± desvio padrão) através do programa BioEstat 4.0 e utlilizou-se da “ANOVA one way” complementando-se com o teste “Bonferroni” com nível de significância para as amostras de P<0,01. As variáveis antropométricas ligadas à composição corporal: peso corporal, estatura, percentual de gordura, SDC (somatório de dobras cutâneas), peso gordo e peso magro mostraram perfeita evolução dentre as categorias analisadas, o que significa ter uma harmonia no processo de evolução entre os grupos. O fator que caracterizou o desenvolvimento desses atletas foi à diminuição da massa óssea (Relativo) provavelmente por causa do fim do desenvolvimento longitudinal e o aumento da massa muscular (Relativo e Absoluto) proveniente da necessidade imposta pela dinâmica do jogo ser mais competitiva na transição entre as categorias.

The anthropometric measures are being used, very broadly, in order to verify the adjustments in connection to the response of training, monitoring and development for the chose of athletes. The scope of this study was to analyze and characterize the body composition of athletes of a base category of a high yield Sport Club in the State of Rio de Janeiro, in the form of soccer player’s competitors regarding the period of the year 2008. All the taster data were collected at the end of the season in December. The test of this study was developed based on a group of 198 male soccer players divided into 5 groups: Under19 n=40, Under17 n=42, Under15 n=41, Under13 n=35 and Under11 n=40. It was used a descriptive statistic ( average ± standard ) by means of the BioEstat 4.0 Program and it was also used the “ANOVA one way” complementing the test “Bonferroni” with a significant level to the tests of P<0,01. The anthropometric variables related to the composition: body weight, height, body fat percentage, SDC (sum of skin folds), fat weight and thin body weight perfectly shows the evolution among the analyzed categories, which means having a line in the process of evolution of the groups. The factor that has characterized the development of these athletes was the reduction of the bone mass (Relative) probably caused by the end of the longitudinal development and the increased of muscle mass (Relative and Absolute) raised by the imposed dynamic of the game be more competitive in the transition of the categories.