RESUMO
Simple cyclic fed-batch culture (cfbc), consisting of a constant medium feed with periodic withdrawals of culture, resulted in a product yield (13.4 mg protein per gram biomass) similar to that obtained using the complex multiphase industrial production strategy (13.7 mg protein per gram biomass). In cfbc, productivity was ultimately limited by the rate at which the cells could assimilate methanol. Glycerol was inhibitory to growth at high concentrations. However, product yield continued to increase as the glycerol concentration was increased. In chemostat culture, dissolved oxygen concentration influenced product yield independently of any detectable influence on cell growth.
Assuntos
Reatores Biológicos/microbiologia , Técnicas de Cultura de Células/métodos , Glicerol/metabolismo , Periodicidade , Pichia/crescimento & desenvolvimento , Pichia/metabolismo , Albumina Sérica/biossíntese , Humanos , Metanol/farmacologia , Oxigênio/metabolismo , Pichia/classificação , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Albumina Sérica/genética , Albumina Sérica/isolamento & purificaçãoRESUMO
Mutation and selection for increased resistance to cell-wall synthesis inhibitors led to alterations in the hyphal branching rate of Saccharopolyspora erythraea NRRL 2338. Mutants with decreased branching frequency exhibited increased hyphal strength (estimated by in vitro micromanipulation). As the hyphal strength was increased, this led to a greater proportion of hyphal particles in liquid culture with a hyphal fragment diameter of greater than 88 microm. This, in turn, coincided with proportionately increased antibiotic production.
Assuntos
Eritromicina/biossíntese , Hifas/citologia , Hifas/fisiologia , Saccharopolyspora/citologia , Saccharopolyspora/fisiologia , Reatores Biológicos , Farmacorresistência Bacteriana/genética , Eritromicina/isolamento & purificação , Fermentação , Processamento de Imagem Assistida por Computador/métodos , Mutação , Reprodutibilidade dos Testes , Saccharopolyspora/genética , Sensibilidade e Especificidade , Estresse MecânicoRESUMO
To examine the relationship between maternal methadone exposure and neonatal head circumference and abstinence syndrome, we examined the records of 172 opiate-addicted gravidas enrolled in a methadone maintenance program in an urban hospital over a 2-year period. Higher doses of methadone in the third trimester were associated with increased head circumference reflecting both increased gestational duration and improved overall growth. Neonatal withdrawal was positively correlated with gestational age at delivery and race, with nonblack infants exhibiting higher neonatal abstinence scores than blacks following adjustment for maternal dose and gestational age at delivery. Selection of optimal methadone dosage is a complex problem in which the favorable neurobehavioral outcome associated with increased growth and gestational age must be weighed against the risks associated with more severe neonatal withdrawal. Our findings of improved overall fetal growth and gestational duration associated with higher methadone doses suggest that more liberal methadone dosing in pregnancy may improve long-term neonatal outcome.
Assuntos
Metadona/efeitos adversos , Transtornos Relacionados ao Uso de Opioides/tratamento farmacológico , Complicações na Gravidez , Resultado da Gravidez , Negro ou Afro-Americano , Cefalometria , Feminino , Idade Gestacional , Humanos , Recém-Nascido , Metadona/administração & dosagem , GravidezRESUMO
The murine hybridoma PQXB1/2 cannot be adapted to grow in culture media containing < 0.5 mM glutamine. Transformants selected following electroporation of PQXB1/2 cells with vectors containing a Chinese hamster glutamine synthetase (GS) cDNA under the control of the SV40 early promoter also failed to grow in the absence of glutamine in the culture medium. PQXB1/2 cells have, however, been transformed to glutamine independence following electroporation with a vector containing this glutamine synthetase cDNA under the control of the human cytomegalovirus immediate early promoter. In these cells, sufficient active glutamine synthetase was expressed from one vector per cell to enable growth in glutamine-free media. The specific activity of glutamine synthetase in two transformed cell lines producing parental levels of antibody was increased by 128 and 152%, respectively (0.57 and 0.63 mumol min-1 per 10(6) cells in transformants compared with parental levels of 0.25 mumol min-1 per 10(6) cells). This reprogramming of glutamine synthetase expression and glutamine metabolism is important for developing strategies to deal with ammonia toxicity and the production of cell lines with improved metabolic processes.
Assuntos
Glutamina/metabolismo , Hibridomas/metabolismo , Animais , Sequência de Bases , Biotecnologia , DNA Complementar/genética , Eletroporação , Engenharia Genética , Vetores Genéticos , Glutamato-Amônia Ligase/genética , Humanos , Hibridomas/imunologia , Camundongos , Dados de Sequência Molecular , TransfecçãoRESUMO
Hybridoma batch cultures were extended using feed formulations based on nutrient consumption measured during different batch culture phases when (a) growth but negligible antibody production was taking place; (b) maximum antibody production rate and declining viable cell growth rate were observed. Strategy (a) was the more successful (2.8-fold compared with 1.8-fold antibody titer increase) and maintained cell viability for longer. Analysis of the effects of omitting individual amino acids yielded results which were consistent with those from the feeding experiment.
RESUMO
The use of partial cubic spline data interpolation for the calculation of volumetric metabolite exchange rates suggested the existence of three distinct metabolic phases during bioreactor culture of a hybridoma cell line. During phase 1, a rapid amino acid uptake rate and ammonia release rate were observed. The growth rate was low and glutamine synthetase activity fell. In phase 2, maximum growth rate and minimum glutamine assimilation and ammonium production rates were observed. Attempts to corroborate the apparent ammonia assimilation in this phase using (15)NH(4)Cl resulted in low incorporation rates into alanine and glutamine. Maximum glutamine synthetase activity took place during this period. Maximum antibody production rate was observed during phase 3 during which peaks in glutamine assimilation, ammonia release, and glutamine synthetase activity were observed. The apparent existence of the three phases prompted us to carry out Northern blot analysis of glutamine synthetase RNA at appropriate times during the process. This revealed a pattern of appearance and dis-appearance of mRNA consistent with the three phases indicated by the fermentation parameters.
RESUMO
Microsomes were prepared from the term placentas of 5 drug-dependent and 4 normal (control) mothers and from the livers of 3 normal fetuses (gestational age = 22-24 weeks) to determine if chronic maternal drug addiction can induce metabolic pathways in the placenta which are utilized in the biotransformation of drugs of abuse. Using as model substrates aminopyrine for demethylation, aniline for hydroxylation, and bilirubin for conjugation, we observed little to none of substrate biotransformation in both drug-dependent and control placentas. Similarly, such enzymatic activity in the fetal liver was also low. We conclude that chronic maternal addiction does not induce metabolic pathways in the placenta for the biotransformation of drugs of abuse.
Assuntos
Drogas Ilícitas/metabolismo , Placenta/metabolismo , Complicações na Gravidez/metabolismo , Transtornos Relacionados ao Uso de Substâncias , Biotransformação , Feminino , Feto/metabolismo , Humanos , Drogas Ilícitas/farmacocinética , Fígado/metabolismo , GravidezRESUMO
In natural ecosystems, microbial activity is often associated with the presence of a surface, particularly in low-nutrient environments. The chemostat allows the study of such low-nutrient environments together with the precise control of other growth parameters. By using this system, enrichment cultures with inocula from two different river sources have been made. A more diverse community attached itself to surfaces placed in the chemostat when the cultures were carbon-limited than when the limiting nutrient was nitrogen. Further studies on a pseudomonad isolated from the carbon-limited enrichment cultures have shown that surface-associated organisms grow at approximately twice the rate of the same organism in the free surrounding medium. A hypothesis to explain this phenomenon based on the chemiosmotic theory is discussed.
Assuntos
Fenômenos Fisiológicos Bacterianos , Bactérias/crescimento & desenvolvimento , Membrana Celular/fisiologia , Metabolismo Energético , Fímbrias Bacterianas/fisiologia , Dinâmica Populacional , Especificidade da Espécie , Propriedades de SuperfícieRESUMO
Neonatal withdrawal is described for a sample of 70 infants born to addicted women treated with methadone and comprehensive prenatal care. Although symptoms were manifested by over 90% of the infants, those born to women receiving near-term average doses of less than or equal to 20 mg/day had significantly less symptomatology, weight loss, and need for pharmacologic treatment than those of mothers still on higher doses. Reduction of methadone dose levels during the last 6 weeks of pregnancy to less than or equal to 20 mg/day appears to reduce the severity of neonatal withdrawal.