Biomedical subjects

Rui Oliveira

Publications and source records attributed to Rui Oliveira.

9 recordsLinked to original sources

PubMed2006

Recent advances in polyhydroxyalkanoate production by mixed aerobic cultures: from the substrate to the final product.

Numerous bacteria have been found to exhibit the capacity for intracellular polyhydroxyalkanoates (PHA) accumulation. Current methods for PHA production at the industrial scale are based on their synthesis from microbial isolates in either their wild form or by recombinant strains. High production costs are associated with these methods; thus, attempts have been made to develop more cost-effective processes. Reducing the cost of the carbon substrates (e.g., through feeding renewable wastes) and increasing the efficiency of production technologies (including both fermentation and downstream extraction and recovery) are two such examples of these attempts. PHA production processes based on mixed microbial cultures are being investigated as a possible technology to decrease production costs, since no sterilization is required and bacteria can adapt quite well to the complex substrates that may be present in waste material. PHA accumulation by mixed cultures has been found under various operational conditions and configurations at both bench-scale and full-scale production. The process known as "feast and famine" or as "aerobic dynamic feeding" seems to have a high potential for PHA production by mixed cultures. Enriched cultures submitted to a transient carbon supply can synthesize PHA at levels comparable to those of pure cultures. Indeed, the intracellular PHA content can reach around 70% of the cell dry weight, suggesting that this process could be competitive with pure culture PHA production when fully developed. Basic and applied research of the PHA production process by mixed cultures has been carried out in the past decade, focusing on areas such as microbial characterization, process configuration, reactor operational strategies, process modeling and control, and polymer characterization. This paper presents a review of the PHA production process with mixed cultures, encompassing the findings reported in the literature as well as our own experimental results in relation to each of these areas.

Bacteria, Aerobic↗

PubMed2005

Mathematical modelling of a mixed culture cultivation process for the production of polyhydroxybutyrate.

Mixed cultures submitted to acetate "feast" and "famine" cycles are able to store intracellularly high quantities of polyhydroxybutyrate (PHB). It was demonstrated in a previous study that the intracellular PHB content can be increased up to 78.5% (g HB/gVSS) of cell dry weight in a sequencing batch reactor (SBR) with optimised operating conditions. The specific PHB formation rate was also shown to be higher for mixed cultures than for pure cultures. Such high intracellular PHB contents and specific productivity open new perspectives for the industrial production of polyhydroxyalkanoates (PHA) using mixed cultures instead of pure cultures. The main goal in this work was to develop a mathematical model of mixed cultures envisaging the optimisation of PHB production. A relatively simple two-compartments cell model was developed based on experimental observations and other models proposed in the literature. A convenient experimental planing allowed to identify the kinetic parameters and yield coefficients. Experiments were performed with and without ammonia limitation enabling the analysis of PHB formation independently of the cell growth process. The experimental true yields partially confirm the theoretical values proposed in the literature. The final model exhibited high accuracy in describing the process state of most experiments performed, thus opening good perspectives for future model-based optimisation studies.

Bacteria↗

PubMed2005

A member of the sugar transporter family, Stl1p is the glycerol/H+ symporter in Saccharomyces cerevisiae.

Glycerol and other polyols are used as osmoprotectants by many organisms. Several yeasts and other fungi can take up glycerol by proton symport. To identify genes involved in active glycerol uptake in Saccharomyces cerevisiae we screened a deletion mutant collection comprising 321 genes encoding proteins with 6 or more predicted transmembrane domains for impaired growth on glycerol medium. Deletion of STL1, which encodes a member of the sugar transporter family, eliminates active glycerol transport. Stl1p is present in the plasma membrane in S. cerevisiae during conditions where glycerol symport is functional. Both the Stl1 protein and the active glycerol transport are subject to glucose-induced inactivation, following identical patterns. Furthermore, the Stl1 protein and the glycerol symporter activity are strongly but transiently induced when cells are subjected to osmotic shock. STL1 was heterologously expressed in Schizosaccharomyces pombe, a yeast that does not contain its own active glycerol transport system. In S. pombe, STL1 conferred the ability to take up glycerol against a concentration gradient in a proton motive force-dependent manner. We conclude that the glycerol proton symporter in S. cerevisiae is encoded by STL1.

Biological Transport↗

PubMed2005

New diagnostic criteria for acute myocardial infarction and in-hospital mortality.

BACKGROUND: The recent introduction of new diagnostic criteria for acute myocardial infarction (AMI), with troponin measurement, has increased the number of patients admitted with this diagnosis. OBJECTIVE: To evaluate the epidemiologic and prognostic implications of the new diagnostic criteria for AMI. METHODS: This was a retrospective study of 586 patients admitted for acute coronary syndrome (ACS) to the coronary care unit of our hospital, between 2002 and 2003. Data were collected from RECIMA, the Madeira Ischemic Heart Disease Registry. The population was analyzed following two different definitions of ACS: 1 - old criteria (Group I): AMI with ST elevation (typical symptoms or ECG with ST-segment elevation and raised CK-MB >2x), AMI without ST elevation (typical symptoms or ECG without ST elevation and raised CK-MB >2x) and unstable angina (UA) (symptoms or ECG indicative of ischemia, with normal CK-MB, regardless of troponin status); 2 - new criteria (Group II): AMI with ST elevation (typical symptoms or ECG with segment ST elevation and raised CK-MB >2x or troponin), AMI without ST elevation (typical symptoms or ECG without ST-segment elevation and raised CK-MB >2x or troponin) and UA (symptoms or ECG indicative of ischemia, with normal enzymes). We evaluated whether this change in criteria had any influence on in-hospital mortality. RESULTS: The new criteria significantly (by 11.9 %) increased the total number of patients admitted with AMI. This was due to an increase in AMI without ST elevation (p < 0.001) and a decrease in patients with UA (p < 0.001), with no changes in AMI with ST elevation. In-hospital mortality was lower in patients with AMI diagnosed by the new criteria and in those with UA. CONCLUSION: The overall increase in AMI resulting from the new diagnostic classification was accompanied by a decrease, although not statistically significant, of in-hospital mortality, probably due to the lower risk of the population analyzed.

Aged↗

PubMed2004

Expression studies of GUP1 and GUP2, genes involved in glycerol active transport in Saccharomyces cerevisiae, using semi-quantitative RT-PCR.

Glycerol active uptake in Saccharomyces cerevisiae, characterised physiologically as a proton symport, was previously described as repressed by glucose, induced by growth on non-fermentable carbon sources and unresponsive to growth under salt stress. GUP1 and GUP2 were identified and characterised as genes involved in glycerol active uptake. Using semi-quantitative RT-PCR, GUP1 and GUP2 transcription was measured. Unlike active transport activity determined previously, this was shown to be constitutive and not affected by either glucose repression or growth under salt stress. Furthermore, transcription of GUP1 and GUP2 was not affected in the gpd1gpd2 mutant strain grown under salt stress in the presence of small amounts of glycerol, in which case a very high Vmax of glycerol uptake was reported. Intracellular compounds were determined. Glycerol, acetate and trehalose were found to be the major compounds accumulated. Surprisingly, the gpd1gpd2 mutant was found to produce significant amounts of glycerol. Yet, the results provide no evidence for a correlation between the amount of each compound and the glycerol transport activity in any of the strains.

Acetates↗

PubMed2004

Optimization of polyhydroxybutyrate production by mixed cultures submitted to aerobic dynamic feeding conditions.

Activated sludge submitted to aerobic dynamic feeding conditions showed a good and stable capacity to store polyhydroxybutyrate (PHB). The system, working for 2 years, selected a microbial population with a high PHB storage capacity. The influence of carbon and nitrogen concentrations on the PHB accumulation yield was studied in a range of 15-180 Cmmol/l for acetate and 0-2.8 Nmmol/l for ammonia. Low ammonia concentrations favored PHB accumulation. The maximum PHB content, 67.5%, was obtained for 180 Cmmol/l of acetate supplied in one pulse. However, such high substrate concentration proved to be inhibitory for the storage mechanism, causing a slowdown of the specific PHB storage rate. In order to avoid substrate inhibition, 180 Cmmol/l of acetate was supplied in different ways: continuously fed and in three pulses of 60 Cmmol/l each. In both cases the specific PHB storage rate increased and the PHB content obtained were 56.2% and 78.5%, respectively. The latter value of PHB content is similar to that obtained by pure cultures and was never reported for mixed cultures. Addition of acetate by pulses controlled by the oxygen concentration was kept for 16 days, the PHB content being always above 70% of cell dry weight.

Acetates↗

PubMed2004

Yeast orthologues associated with glycerol transport and metabolism.

Glycerol is a key compound in the regulation of several metabolic pathways in Saccharomyces cerevisiae. From this yeast most of the genes involved in glycerol consumption, production and transport are now available. Some of the mechanisms involving glycerol metabolism and transport are common to other yeasts. This work presents a search for GPD1/2, GUT1, GUP1/2 and FPS1 orthologues in a series of hemiascomycetous yeasts. All the genes cloned were able to complement S. cerevisiae mutant phenotypes and presented a high degree of similarity to the corresponding genes in this yeast. A phylogenetic analysis is presented. The allocation of GUP genes in the membrane bound O-acyl transferases (MBOAT) family is suggested as more consistent than their inclusion in the TC-DB/glycerol uptake family.

Biological Transport↗

PubMed2003

Fps1p channel is the mediator of the major part of glycerol passive diffusion in Saccharomyces cerevisiae: artefacts and re-definitions.

Glycerol has been shown to cross the plasma membrane of Saccharomyces cerevisiae through (1) a H(+)/symport detected in cells grown on non-fermentable carbon sources, (2) the constitutively expressed Fps1p channel and (3) by passive diffusion. The Fps1p channel has been named a facilitator for mediating glycerol low affinity transport of the facilitated diffusion type. We present experimental evidence that this kinetic is an artefact created by glycerol kinase activity. Instead, the channel is shown to mediate the major part of glycerol's passive diffusion. This is not incompatible with Fps1p's major role in vivo, which has been previously shown to be the control of glycerol export under osmotic stress or in reaction to turgor changes. We also verified that FPS1 overexpression caused an increase in H(+)/symport V(max). Furthermore, yfl054c and fps1 mutants were equally affected by exogenously added ethanol, being the correspondent passive diffusion stimulated. For the first time, to our knowledge, a phenotype attributed to the functioning of YFL054c gene is presented. Glycerol passive diffusion is thus apparently channel-mediated. This is discussed according to glycerol's chemical properties, which contradict the widely spread concept of glycerol's liposoluble nature. The discussion considers the multiple roles that the intracellular levels of glycerol and its pathway regulation might play as a central key to metabolism control.

Artifacts↗

PubMedDate not supplied

Bioprocess iterative batch-to-batch optimization based on hybrid parametric/nonparametric models.

This paper presents a novel method for iterative batch-to-batch dynamic optimization of bioprocesses. The relationship between process performance and control inputs is established by means of hybrid grey-box models combining parametric and nonparametric structures. The bioreactor dynamics are defined by material balance equations, whereas the cell population subsystem is represented by an adjustable mixture of nonparametric and parametric models. Thus optimizations are possible without detailed mechanistic knowledge concerning the biological system. A clustering technique is used to supervise the reliability of the nonparametric subsystem during the optimization. Whenever the nonparametric outputs are unreliable, the objective function is penalized. The technique was evaluated with three simulation case studies. The overall results suggest that the convergence to the optimal process performance may be achieved after a small number of batches. The model unreliability risk constraint along with sampling scheduling are crucial to minimize the experimental effort required to attain a given process performance. In general terms, it may be concluded that the proposed method broadens the application of the hybrid parametric/nonparametric modeling technique to "newer" processes with higher potential for optimization.

Bioreactors↗