abstract: Petroleum contamination is ubiquitous during extraction, transportation, refining, and storage. Contamination damages the soil’s ecosystem function, reduces its aesthetics, and poses a potential threat to human beings. The overall goals of this dissertation are to advance understanding of the mechanisms behind ozonation of petroleum-contaminated soil and to configure an effective integrated bioremediation + ozonation remedial strategy to remove the overall organic carbon. Using a soil column, I conducted batch ozonation experiments for different soils and at different moisture levels. I measured multiple parameters: e.g., total petroleum hydrocarbons (TPH) and dissolved organic carbon (DOC), to build a full understanding of the data that led to the solid conclusions. I first demonstrated the feasibility of using ozone to attack heavy petroleum hydrocarbons in soil settings. I identified the physical and chemical hurdles (e.g., moisture, mass transfer, pH) needed to be overcome to make the integration of chemical oxidation and biodegradation more efficient and defines the mechanisms behind the experimental observations. Next, I completed a total carbon balance, which revealed that multiple components, including soil organic matter (SOM) and non-TPH petroleum, competed for ozone, although TPH was relatively more reactive. Further experiments showed that poor soil mixing and high soil-moisture content hindered mass transfer of ozone to react with the TPH. Finally, I pursued the theme of optimizing the integration of ozonation and biodegradation through a multi-stage strategy. I conducted multi-stages of ozonation and bioremediation for two benchmark soils with distinctly different oils to test if and how much ozonation enhanced biodegradation and vice versa. With pH and moisture optimized for each step, pre-ozonation versus post-ozonation was assessed for TPH removal and mineralization. Multi-cycle treatment was able to achieve the TPH regulatory standard when biodegradation alone could not. Ozonation did not directly enhance the biodegradation rate of TPH; instead, ozone converted TPH into DOC that was biodegraded and mineralized. The major take-home lesson from my studies is that multi-stage ozonation + biodegradation is a useful remediation tool for petroleum contamination in soil. / Dissertation/Thesis / Doctoral Dissertation Civil, Environmental and Sustainable Engineering 2018

Na maior parte dos ecossistemas aquáticos a respiração excede a produção primária bruta autóctone, acarretando uma produção líquida de ecossistema negativa. Estudos recentes atribuem essa condição a processos de degradação de matéria orgânica alóctone, portanto o ciclo do carbono em corpos de água interiores deve estar pareado com o aporte de carbono orgânico originado na bacia hidrográfica. Este trabalho contribui para a melhor compreensão dos processos atuantes no ciclo de carbono em bacias hidrográficas, por meio de monitoramento de dados de campo, desenvolvimento e aplicação de modelagem matemática e simulação numérica. O estudo foi realizado utilizando dados de monitoramento na bacia hidrográfica do Rio Ijuí localizada no planalto meridional gaúcho. A exportação fluvial de carbono orgânico e inorgânico foi estimada a partir de dados de vazão e concentração. Um modelo baseado em processos para simulação da dinâmica de carbono, MGB-IPH-C, foi desenvolvido e acoplado ao modelo hidrológico MGB-IPH. O modelo hidrológico foi ajustado em diversos pontos de controle na bacia hidrográfica do Ijuí obtendo-se bons resultados, principalmente para as bacias de maior porte (>1000 km2). O modelo de carbono foi avaliado qualitativamente considerando a representação de processos conceituais e, quantitativamente, pela comparação das concentrações e fluxos simulados em relação aos obtidos pelos dados do monitoramento. Não foi possível identificar associações diretas entre as concentrações de carbono inorgânico ou orgânico e vazão, com base nos dados medidos em campo. As cargas médias de carbono nas bacias estudadas apresentaram valores abaixo da média global, na ordem de 25-40 kg.ha-1.ano-1 e 8-10 kg.ha-1.ano-1, para as frações inorgânica e orgânica, respectivamente. O MGB-IPH-C representou processos conceituais esperados, em especial, a acumulação no solo e o efeito da lavagem das águas sobre a concentração dos rios. Foi possível obter bons ajustes em escala anual para as cargas e concentrações médias de carbono na bacia de estudo. A simulação da dinâmica de carbono em ecossistemas aquáticos em pareamento com a bacia hidrográfica utilizando modelos conceituais determinísticos contribui para o entendimento dos processos operantes nesses sistemas e deve ser complementada por meio de análise de dados de monitoramento, em freqüência adequada à escala, com métodos empíricos. / In most aquatic ecosystems respiration exceeds autochtonous gross primary production leading to a negative net ecosystem production. Recent studies attributes this condition to allochtonous organic matter degradation processes, so the inland water carbon cycle should be linked to the input of organic carbon from the catchment. This work contributes to a better comprehension of processes operating in the carbon cycle in river basins, throughout development and application of mathematical modeling and numerical simulation. The study was conducted using data of the Ijuí river watershed, located in the Planalto Meridional Gaúcho. Fluvial exports of organic and inorganic carbon were estimated from data measured in field. A process-based model for carbon dynamics simulation, MGB-IPH-C, was developed and coupled to the hydrological model MGB-IPH. The hydrological model was calibrated to various control points in the Ijui river basin with good results, especially in larger subwatersheds (>1000 km2). The carbon model was evaluated considering the representation of carbon dynamics conceptual processes and by comparison of observed carbon concentrations and export rates obtained by simulation and monitoring data. The estimates of annual export of carbon in the study area were below the global average, in magnitude of 25-40 kg/ha.yr and 8-10 kg/ha.yr for inorganic and organic carbon, respectively. The proposed carbon model, MGB-IPH-C, was able to simulate expected conceptual processes, in particular, the build-up on soil and the effect of wash-off on carbon concentration in river reaches. It was possible to obtain good fits for annual average carbon fluvial export rates and concentrations. The simulation of carbon dynamics in inland waters coupled with the watershed using conceptual deterministic models contributes to the understanding of processes operating on these systems and may be complemented throughout analysis of field data, at frequency appropriated to the scale, with empirical methods.

A Amazônia armazena em sua floresta na ordem de 95 a 120 PgC de biomassa viva e mais 160 PgC no solo, que podem ser rapidamente liberados para a atmosfera por meio da queima de biomassa e, também, pela mudança do uso da terra. Este estudo foi desenvolvido com o objetivo de elucidar a contribuição da Bacia Amazônica nas emissões de carbono no ano de 2010. A quantificação do CO2 foi realizada por meio da coleta do ar atmosférico utilizando aviões de pequeno porte que descreveram um perfil vertical em quatro locais, estrategicamente posicionado na Bacia Amazônica, e utilizando sistemas semiautomáticos de coleta de ar em 17 ou 12 altitudes diferentes. O Fluxo de emissão/absorção foi calculado pelo método de integração de coluna, que consiste na determinação da concentração de CO2 no perfil vertical, subtraído da concentração de entrada no continente, levando-se em conta o tempo que a massa de ar despende entre a costa e o local de amostragem. Para a determinação da concentração de entrada, foram utilizadas as concentrações medidas pela NOAA nas Ilhas de Ascencion e Barbados e, como traçador de massas de ar, o SF6. Foi encontrado um caráter emissor da Amazônia em território brasileiro para o ano de 2010, em torno de 0,41 PgC, considerando a média ponderada das quatro regiões estudadas, sendo a queima de biomassa a principal responsável. Para a determinação da emissão proveniente da queima de biomassa foi utilizado o CO, como traçador, e a razão CO:CO2. Apesar de possuir um perfil emissor neste ano anormalmente seco, foi possível observar um caráter sumidor de carbono. / The Amazon forest contains on the order of 95-120 PgC in living biomass and additional 160 PgC in soils, which can be quickly released into the atmosphere through the biomass burning and also by the land use change. This study was developed with the aim to elucidate the contribution of the Amazon Basin in carbon emissions. The quantification of CO2 was performed by collecting atmospheric air using small airplanes that described a vertical profile at four locations strategically positioned in the Amazon and using semi-automatic sampling systems in 17 or 12 different heights. The emission / absorption flux was calculated by column integration method, which consists in determining the vertical profile concentration, subtracted from the background concentration, taking into account the time that the air mass spends between coast and sampling site. To determine the background concentration, were used the concentrations measured by NOAA at the Ascencion and Barbados islands and, as an air mass tracer, the SF6. Was found a emission behavior at the Amazon in Brazil for the year 2010, around 0.41 PgC considering the weighted average of the four regions studied, being the biomass burning mainly responsible. To determine the biomass burning emission, was used the CO as a tracer and, the CO:CO2 ratio. Despite having an emitter profile at this abnormally dry year, it was possible to observe it behavior of carbon sink.

CARBON AND NITROGEN POOLS AND FLUXES OF TWO FOREST ECOSYSTEMS IN THE ORE MOUNTAINS With increasing atmospheric CO2 concentrations, forest ecosystems are considered for their sequestration ability. However, there are differences between coniferous and deciduous tree species in their impact on the carbon (C) and nitrogen (N) fluxes within forest ecosystems, which influence soil carbon and nitrogen pools. In natural beech and monoculture spruce stands on research sites of Czech Geological Survey in the Ore Mountains, C and N fluxes and pools were investigated. There were investigated ecosystem inputs (C, N, in throughfall, litterfall) as well as outputs (soil respiration, concentrations of C, N in seepage water discharge). Further, C and N pools of living biomass and soils were determined. Throughfall DOC was significantly higher in the spruce stand, on the other hand litterfall C flux showed the opposite relationship. At output, DOC, DON and NH4 + discharge fluxes from O horizon were significantly higher in the spruce stand whereas in the beech stand higher NO3 - flux from mineral soil was recorded. Soil respiration was quite similar in the both stands, however different parts of original sources of respiration among tree species were different. Beech stand has larger pools of carbon and nitrogen in...

How do additional data of the same and/or different type contribute to reducing model parameter and predictive uncertainties? This was the question addressed with two models – the HBV hydrological water balance model and the ICBM soil carbon balance model – that were used to investigate the usefulness of the Generalized Likelihood Uncertainty Estimation (GLUE) method for calibrations and uncertainty analyses.  The GLUE method is based on threshold screening of Monte Carlo simulations using so-called informal likelihood measures and subjective acceptance criterion. This method is highly appropriate for model calibrations when errors are dominated by epistemic rather than stochastic uncertainties.  The informative value of data for model calibrations was investigated with numerous calibrations aimed at conditioning posterior parameter distributions and boundaries on model predictions.  The key results demonstrated examples of: 1) redundant information in daily time series of hydrological data; 2) diminishing returns in the value of continued time series data collections of the same type; 3) the potential value of additional data of a different type; 4) a means to effectively incorporate fuzzy information in model calibrations; and 5) the robustness of estimated parameter uncertainty for portability of a soil carbon model between and tropical climate zones.  The key to obtaining these insights lied in the methods of uncertainty analysis used to produce them.  A paradigm for selecting between formal and informal likelihood measures in uncertainty analysis is presented and discussed for future use within a context of climate related environmental modeling. / QC 20110414

Esta pesquisa consistiu em estimar o balanço de carbono da vegetação nativa de uma paisagem antropizada, ou seja, calcular a quantidade de biomassa arbórea acima do solo (AGB) preexistente (cenário pré-colonial), o quanto foi perdido pela ação humana (desmatamento e fragmentação) e que quantidade de carbono poderia ser recuperada com a adequação ao Novo Código Florestal Brasileiro (NCFB). Os fatores desse balanço são apresentados cartograficamente, e foram obtidos baseado em dados de campo para associar valores de tonelagem aos estoques estimados. A AGB pré-colonial estimada foi de 4.62 Gigatoneladas, dos quais 87.3% (3.85 Gt) foram perdidos até 2017. Essas perdas foram atribuídas em 90% (3.46 Gt) ao desmatamento, e 10% (0.39 Gt) à efeitos decorrentes da fragmentação. Os déficits em Reserva Legal e Áreas de Preservação Permanente estipulados pelo NCFB se traduzem em um potencial de recuperação de 0.11 Gt de AGB para São Paulo. Se reduzíssemos os efeitos da fragmentação esse potencial triplicaria (0.37 Gt). Os hotspots de biomassa atuais estão restritos às florestas costeiras (Ribeira do Iguape, Baixada Santista e Litoral Norte) e que as regiões com maior potencial de recuperação de carbono com a adequação ao NCFB são o Vale do Paraíba, Aguapeí e Pontal do Paranapanema. Finalmente, discutimos a eficácia dos mecanismos de compensação: seriam necessários 0.96 a 2.79 bilhões de US$ para restaurar o déficit legal de vegetação, possível apenas se os mercados estiverem dispostos a pagar pelo menos 17.5 US$ por tonelada de carbono fixado na vegetação / The research aimed to estimate the carbon balance of natural vegetation in a tropical anthropized landscape. That is, to calculate the amount of pre-existing tree biomass above the soil (pre-colonial scenario), how much was lost by human action (deforestation and fragmentation) and how much carbon could be recovered with the adaptation to the New Brazilian Forest Code (NCFB). This balance is represented cartographically for the state of São Paulo, in its different stages (e.g. carbon inventory map for the original vegetation cover), and is based on field data to associate tonnage values with the estimated inventories. The estimated pre-colonial AGB for São Paulo was 4.62 Gigatonnes, of which 87.3% (3.85 Gt) were lost. These losses can be attributed to 90% (3.46 Gt) to deforestation, and 10% (0.39 Gt) to the effects of fragmentation. The vegetation deficits in Legal Reserve and Areas of Permanent Preservation stipulated by the NCFB translate into a potential recovery of 0.11 Gt of AGB. If we were to reduce degradation in forest fragments this potential AGB recovery would raise to 0.37 Gt. It was observed that current biomass hotspots are restricted to coastal forests (Ribeira do Iguape, Baixada Santista and Litoral Norte) and that the regions with the greatest potential for carbon recovery with adaptation to the NCFB are the Vale do Paraíba, Aguapeí and Pontal of Paranapanema. Finally, we discussed the effectiveness of compensation mechanisms, it would require 0.96 to 2.79 billion US$ to restore the legal deficit of vegetation, feasible as long as carbon markets are willing to pay 17.5 US$ for ton of fixed Carbon

Decisões tomadas no planejamento da implantação florestal, como a densidade do plantio e o material genético, alteram a disponibilidade de recursos naturais tais como nutrientes, água e luz e consequentemente afetam a assimilação do carbono, que está diretamente relacionada ao crescimento da planta. Ainda são escassas as informações de como e quanto a densidade de plantio e o material genético afetam a fixação de carbono pelas ávores. O objetivo desse trabalho foi avaliar os padrões de fixação e alocação de carbono em plantações clonais de Eucalyptus sp. em duas densidades de plantio (3x2 m e 3x4 m), em um Latossolo Vermelho Distrófico T ípico A moderado. O estudo foi desenvolvido na Estação Experimental de Ciências Florestais de Itatinga-SP - ESALQ/USP, com três materiais clonais de Eucalyptus urophylla: AEC 0144, AEC 224 e COP 1404. O delineamento experimental utilizado foi em blocos casualizados, com esquema fatorial 3 x 2, sendo composto por três clones de eucalipto em dois espaçamentos, com 6 repetições para cada tratamento. Durante o intervalo de um ano (dos 40 aos 52 meses de idade) foram determinados a Produtividade Primária Líquida da Parte Aérea (ANPP: incremento da biomassa aérea somada ao folhedo), a Respiração Autotrófica da Parte Aérea (Rp: respiração das folhas e do tronco com base com base na ANPP, assumindo um valor constante de eficiência de uso do carbono (CUE) de 0,53 (GIARDINA et al., 2003)), o Fluxo de Carbono Abaixo do Solo (TBCF: produção e respiração das raízes grossas e finas, exsudatos das raízes e produção de substratos usados por micorrizas) e a Produtividade Primária Bruta (GPP: somatório dos fluxos de carbono) para os seis tratamentos avaliados. A maior produtividade encontrada para o clone AEC 0144 no espaçamento 3x2 m foi resultado de uma maior GPP (5997,45 g C m-2 ano-1), maior partição de carbono para incremento de tronco (30%) e menor partição da GPP para TBCF (34%). A ANPP variou de 1453,99 g C m-2 ano-1 (Clone COP 1404 no espaçamento 3x2 m) a 2288,78 g C m-2 ano-1 (Clone AEC 0144 no espaçamento 3x2 m), sendo os maiores fluxos encontrados para os clones AEC 0144 em ambos espaçamentos e para o clone COP 1404 no espaçamento 3x4 m. A variação encontrada nos valores de respiração da parte aérea seguiram o mesmo padrão dos resultados da ANPP. O TBCF foi significadamente superior para os clones AEC 0144 e AEC 224 no espaçamento 3x2 m com 2056,36 g C m-2 ano-1 e 1903,83 g C m-2 ano-1, respectivamente; e para o clone COP 1404 no espaçamento 3x4 m (1927,43 g C m-2 ano-1). Houve correlação positiva do TBCF com a GPP, mas não com a ANPP. / Decisions not planned for forest deployment, such as planting density and genetic material, alter the availability of natural resources such as nutrients, water and light and consequently affect the assimilation of carbon, which is directed to plant growth. They are still scarce as information on how and how much planting density and genetic material for a fixation of carbon by trees. The objective of this work was to evaluate the carbon allocation patterns in clonal plantations of Eucalyptus sp. in two planting densities (3x2 m and 3x4 m), in a typical Typic A moderate Dystrophic Red Latosol. The study was developed at the Experimental Station of Forest Sciences of Itatinga-SP - ESALQ / USP, with three clonal materials of Eucalyptus urophylla: AEC 0144, AEC 224 and COP 1404. The experimental design was a randomized complete block design with a factorial scheme 3 x 2, being composed of three clones of eucalyptus in two spacings, with 6 replicates for each treatment. During the one-year interval (from 40 to 52 months of age) was determined Aboveground Net Primary Productivity (ANPP: increase of the aerial biomass added to the litterfall), Aboveground Autotrophic Respiration (Rp: leaf respiration and (CUE) of 0.53 (GIARDINA et al., 2003)), Total Belowground Carbon Flux (TBCF: production and respiration of the roots) and Gross Primary Productivity (GPP: sum of the carbon fluxes) for the six treatments evaluated. The higher productivity found for the clone AEC 0144 in the 3x2 m spacing resulted from a higher GPP (5997.45 g C m-2 year-1), larger carbon partition for trunk increment (30%) and smaller partition from GPP for TBCF (34%). The ANPP ranged from 1453.99 g C m-2 year-1 (Clone COP 1404 in spacing 3x2 m) to 2288.78 g C m-2 year-1 (Clone AEC 0144 in 3x2 m spacing) Found for clones AEC 0144 in both spacings and COP 1404 in 3x4 m spacing. The variation found in aboveground autotrophic respiration values followed the same pattern of ANPP results. The TBCF was significantly higher for clones AEC 0144 and AEC 224 at 3x2 m spacing with 2056.36 m-2 year-1 and 1903.83 m-2 year-1, respectively; and for clone COP 1404 in 3x4 m spacing (1927.43 m-2 year-1). There was a positive correlation between TBCF and GPP, but not with ANPP.

The objective of this work is to better understand themetabolism and physiology ofEscherichiacoli(W3110) in defined medium cultures with thelong-term goal of improving cell yield and recombinant proteinproductivity. The order of amino acid utilization inE. colibatch cultures was investigated in a medium with16 amino acids and glucose. Ser, Pro, Asp, Gly, Thr, Glu andAla were rapidly consumed and depleted at the end of theexponential phase, while His, Arg, Val, Met, Ile, Leu, Phe, Lysand Tyr were consumed slowly during the following linear growthphase. The uptake order correlated to the maximum specificconsumption rate. Of the rapidly consumed amino acids onlyglyine and threonine improved growth when added individually.Serine was the first amino acid to be consumed, but inhibitedglucose uptake initially, which presumably is related to thefunction of PTS. Valine inhibited cell growth could be releasedby isoleucine. The critical medium concentration of valinetoxicity was 1.5 - 3 µmol L-1. Valine uptake was associated with exchange ofisoleucine out of the cells. Glycine significantly increased the cell yield,Yx/s,and growth rate ofE. coliin batch cultures in a glucose-mineral medium.Maximum effect occurred at pH 6.8, at 6 - 12 mmol L-1glycine, and below 1.15 g dw L-1.13C NMR technique was employed to identify [1-13C], [2-13C]and [1,2-13C]acetate in the cultures supplied with [2-13C]glycine. The NMR data revealed that littledegradation of added glycine occurred, and that serine/glycinebiosynthesis was repressed below 1.15 g dw L-1, implicating that glycine was a source ofglycine, serine, one-carbon units, and threonine. Above 1.15 gdw L-1, 53% of the consumed glycine carbon was excretedas acetate. Degradation of glycine was associated with anincreased uptake rate, cleavage by GCV, and degradation of bothglycine- and glucose-derived serine to pyruvate. This switch inmetabolism appears to be regulated by quorum sensing. A cell density-dependent metabolic switch occurred also inthe central metabolism. A 2 - 3 fold decrease in mostglycolytic and TCA cycle metabolites, but an increase inacetyl-CoA, occurred after the switch. The acetate productionrate decreased throughout the culture with a temporary increaseat the switch point, but the intracellular acetate poolremained relatively constant. Two mixtures of amino acids were fed together with glucosein fed-batch cultures ofE. coliW3110 pRIT44T2, expressing the recombinantprotein ZZT2. One mixture contained 20 amino acids and theother 5 so-called 'protein amino acids': Ala, Arg, Met, His andPhe. Although the amino aids increased the cell yield anddecreased the proteolysis rate in both cases, ZZT2 productionwas decreased. A decrease of ZZT2 synthesis rate is consideredto be the reason. Further studies of the 5 amino acidsindicated that a few amino acids disturb metabolism. Carbon mass balances were calculated in glucose limitedfed-batch cultures ofE. coli. In the end, the carbon recovery was ~90% basedon biomass, CO2and acetate, but ~100% if the all carbon in themedium was included. Outer membrane (OM) constituents,lipopolysaccharide, phospholipids, and carbohydratescontributed to 63% of the extracellular carbon. Little celllysis occurred and the unidentified (~30%) carbon was assumedto constitute complex carbohydrates. A novel cultivationtechnique Temperature-Limited Fed-Batch (TLFB) is developed toprevent OM shedding in high-cell density cultures. <b>Keywords</b>: Escherichia coli, amino acids, glycine, quorumsensing, metabolic switch, metabolite pools, carbon balance,outer membrane, lipopolysaccharide, batch culture, fed-batchculture

Eddy-covariance (EC) CO₂ flux data were analysed and annual carbon (C) balances estimated for a four-year period (2004-2007) following clearcut harvesting of a boreal jack pine stand in northern Saskatchewan. The site was a source of C to the atmosphere for all years, with annual net ecosystem productivity (NEP) increasing from -153 g C m⁻² yr⁻¹ in 2004 to -63 g C m⁻² yr⁻¹ in 2007. This increase was mainly due to gross primary productivity (GPP) increasing significantly from 78 to 200 g C m⁻² yr⁻¹ , while ecosystem respiration (R) increased only slightly from 231 to 263 g C m⁻² yr⁻¹ over the same period. In the 2006 growing season (GS), a field campaign was conducted to investigate the relationships between monthly destructive measurements of leaf area index (LAI) and daily measurements of the normalized difference vegetation index (NDVI) and photosynthetic capacity (Amax). The latter was derived from 5-day, 16-day, 30-day and annual Michaelis-Menten light response analyses using daytime measurements of NEP and incident photosynthetically active radiation. Digital-camera data were used to evaluate the potential of using the rectilinear-lens vegetation index (RLVI) as a surrogate for NDVI of a young forest stand. Results showed that LAI was linearly related to NDVI and RLVI, which was largely the result of changes in the deciduous vegetation component across the GS. These results indicate that RLVI could be used as a surrogate for NDVI up to a GS maximum LAI of 0.91 m2 m⁻² observed in 2006. Measured mean (± 1 S.D.) GS LAI was 0.67 (± 0.24) m² m⁻² in 2006. LAI accounted for the majority of the variability in Amax at the 30-day time scale, while at shorter time scales air temperature was the dominant control. For 2004 to 2007, mean spring estimates of LAI were 0.25, 0.29, 0.38 (compared to 0.40 m² m⁻² from measurements) and 0.41 m² m⁻², respectively. Results suggest that a steady increase in the jack pine LAI component accounted for the annual increases in GPP and hence NEP over the four years.

<p>The objective of this work is to better understand themetabolism and physiology of<i>Escherichiacoli</i>(W3110) in defined medium cultures with thelong-term goal of improving cell yield and recombinant proteinproductivity.</p><p>The order of amino acid utilization in<i>E. coli</i>batch cultures was investigated in a medium with16 amino acids and glucose. Ser, Pro, Asp, Gly, Thr, Glu andAla were rapidly consumed and depleted at the end of theexponential phase, while His, Arg, Val, Met, Ile, Leu, Phe, Lysand Tyr were consumed slowly during the following linear growthphase. The uptake order correlated to the maximum specificconsumption rate. Of the rapidly consumed amino acids onlyglyine and threonine improved growth when added individually.Serine was the first amino acid to be consumed, but inhibitedglucose uptake initially, which presumably is related to thefunction of PTS. Valine inhibited cell growth could be releasedby isoleucine. The critical medium concentration of valinetoxicity was 1.5 - 3 µmol L^-1. Valine uptake was associated with exchange ofisoleucine out of the cells.</p><p>Glycine significantly increased the cell yield,<i>Y</i>_x/s,and growth rate of<i>E. coli</i>in batch cultures in a glucose-mineral medium.Maximum effect occurred at pH 6.8, at 6 - 12 mmol L^-1glycine, and below 1.15 g dw L^-1.¹³C NMR technique was employed to identify [1-¹³C], [2-¹³C]and [1,2-¹³C]acetate in the cultures supplied with [2-¹³C]glycine. The NMR data revealed that littledegradation of added glycine occurred, and that serine/glycinebiosynthesis was repressed below 1.15 g dw L^-1, implicating that glycine was a source ofglycine, serine, one-carbon units, and threonine. Above 1.15 gdw L^-1, 53% of the consumed glycine carbon was excretedas acetate. Degradation of glycine was associated with anincreased uptake rate, cleavage by GCV, and degradation of bothglycine- and glucose-derived serine to pyruvate. This switch inmetabolism appears to be regulated by quorum sensing.</p><p>A cell density-dependent metabolic switch occurred also inthe central metabolism. A 2 - 3 fold decrease in mostglycolytic and TCA cycle metabolites, but an increase inacetyl-CoA, occurred after the switch. The acetate productionrate decreased throughout the culture with a temporary increaseat the switch point, but the intracellular acetate poolremained relatively constant.</p><p>Two mixtures of amino acids were fed together with glucosein fed-batch cultures of<i>E. coli</i>W3110 pRIT44T2, expressing the recombinantprotein ZZT2. One mixture contained 20 amino acids and theother 5 so-called 'protein amino acids': Ala, Arg, Met, His andPhe. Although the amino aids increased the cell yield anddecreased the proteolysis rate in both cases, ZZT2 productionwas decreased. A decrease of ZZT2 synthesis rate is consideredto be the reason. Further studies of the 5 amino acidsindicated that a few amino acids disturb metabolism.</p><p>Carbon mass balances were calculated in glucose limitedfed-batch cultures of<i>E. coli</i>. In the end, the carbon recovery was ~90% basedon biomass, CO₂and acetate, but ~100% if the all carbon in themedium was included. Outer membrane (OM) constituents,lipopolysaccharide, phospholipids, and carbohydratescontributed to 63% of the extracellular carbon. Little celllysis occurred and the unidentified (~30%) carbon was assumedto constitute complex carbohydrates. A novel cultivationtechnique Temperature-Limited Fed-Batch (TLFB) is developed toprevent OM shedding in high-cell density cultures.</p><p><b>Keywords</b>: Escherichia coli, amino acids, glycine, quorumsensing, metabolic switch, metabolite pools, carbon balance,outer membrane, lipopolysaccharide, batch culture, fed-batchculture</p>