Bladder cancer biomarker array to detect aberrant levels of proteins in urine.

Bladder cancer (BCa) is a serious malignancy of the urinary tract worldwide and also prominent for its high rate of recurrence incorporating 50% of all treated patients. To reduce relapse of BCa, lifelong surveillance of patients is essential leading to high treatment costs. The gold standard for the diagnosis of bladder cancer is cystoscopy. It is very sensitive, but due to high costs and its invasive nature this method for routine diagnosis of bladder cancer remains questionable. Because of this and the required surveillance of patients suffering from bladder cancer, urine based markers represent a new potential field of investigation. Literature at the National Center of Biological Information (NCBI) was retrieved for a potential marker panel offering specific protein signatures and used to develop a sensitive and accurate chip assay to monitor BCa. Discovery of possible bladder cancer protein markers is compiled by extensive literature search including 1077 recently (15.01.2008-20.03.2014) published research articles. Validation of this literature is done by selection based on prior defined inclusion and exclusion criteria. A set of six putative biomarkers (VEGF, IL-8, MMP-9, MMP-7, survivin and Cyfra 21.1) was identified and a non-invasive microarray developed to be used for further clinical validation. Investigation regarding optimized urine preparation and assay development, to enhance assay sensitivity for the marker panel, was carried out. This protein based BCa chip enables the fast (within 5 h), simultaneous, easy to operate, cheap, early and non-invasive determination of BCa and is ready for clinical evaluation.

Benefits of a European project on diagnostics of highly pathogenic agents and assessment of potential "dual use" issues.

Quality assurance exercises and networking on the detection of highly infectious pathogens (QUANDHIP) is a joint action initiative set up in 2011 that has successfully unified the primary objectives of the European Network on Highly Pathogenic Bacteria (ENHPB) and of P4-laboratories (ENP4-Lab) both of which aimed to improve the efficiency, effectiveness, and response capabilities of laboratories directed at protecting the health of European citizens against high consequence bacteria and viruses of significant public health concern. Both networks have established a common collaborative consortium of 37 nationally and internationally recognized institutions with laboratory facilities from 22 European countries. The specific objectives and achievements include the initiation and establishment of a recognized and acceptable quality assurance scheme, including practical external quality assurance exercises, comprising living agents, that aims to improve laboratory performance, accuracy, and detection capabilities in support of patient management and public health responses; recognized training schemes for diagnostics and handling of highly pathogenic agents; international repositories comprising highly pathogenic bacteria and viruses for the development of standardized reference material; a standardized and transparent Biosafety and Biosecurity strategy protecting healthcare personnel and the community in dealing with high consequence pathogens; the design and organization of response capabilities dealing with cross-border events with highly infectious pathogens including the consideration of diagnostic capabilities of individual European laboratories. The project tackled several sensitive issues regarding Biosafety, Biosecurity and "dual use" concerns. The article will give an overview of the project outcomes and discuss the assessment of potential "dual use" issues.

Cementless acetabular reconstruction and structural bone-grafting in dysplastic hips. Surgical technique.

BACKGROUND: Studies of acetabular reconstruction with use of cement and bulk bone graft have demonstrated increasing rates of cup failure in patients with dysplastic hips seven years after total hip arthroplasty. Comparable data on the long-term results of bulk bone-grafting done in conjunction with cementless implants are limited. The aim of this study was to review the clinical and radiographic results of autologous bulk bone-grafting in conjunction with a cementless cup. METHODS: From 1987 to 1992, forty-seven patients (forty women and seven men, with an average age of 50.4 years) who had developmental dysplasia of the hip underwent fifty-six total hip arthroplasties and received a structural graft in combination with a cementless Harris-Galante type-I cup. All patients were followed prospectively. In fifty-five hips, implant migration was measured with single-image radiographic analysis. RESULTS: After an average duration (and standard deviation) of 10.2 +/- 2.9 years, three patients (four hips) had died. In the surviving patients, four implants had been revised and two had radiographic evidence of loosening. With use of revision and loosening as end points, the eleven-year survival rates were 91.6% and 88.9%, respectively. Of the fifty implants that had no loosening, fourteen had measurable cup migration, thirty-five had no migration, and one implant could not be measured. All migrations but one were progressive. With loosening used as the end point, the survival rate at eleven years was 100% for the implants with no migration; however, the survival rate for the cups that had migrated was 69.3% (p = 0.0012). CONCLUSIONS: The eleven-year survival rate for the spherical press-fit cups in combination with bulk bone-grafting is satisfactory, given the complexity of these reconstructions. However, the difference between the survival of the implants that had migrated and those that had not was significant. We expect that the thirteen implants with progressive acetabular migration at the time of the latest follow-up are at risk for loosening, which will increase the revision rate for this series in the coming years.

Cementless acetabular reconstruction and structural bone-grafting in dysplastic hips.

BACKGROUND: Studies of acetabular reconstruction with use of cement and bulk bone graft have demonstrated increasing rates of cup failure in patients with dysplastic hips seven years after total hip arthroplasty. Comparable data on the long-term results of bulk bone-grafting done in conjunction with cementless implants are limited. The aim of this study was to review the clinical and radiographic results of autologous bulk bone-grafting in conjunction with a cementless cup. METHODS: From 1987 to 1992, forty-seven patients (forty women and seven men, with an average age of 50.4 years) who had developmental dysplasia of the hip underwent fifty-six total hip arthroplasties and received a structural graft in combination with a cementless Harris-Galante type-I cup. All patients were followed prospectively. In fifty-five hips, implant migration was measured with single-image radiographic analysis. RESULTS: After an average duration (and standard deviation) of 10.2 +/- 2.9 years, three patients (four hips) had died. In the surviving patients, four implants had been revised and two had radiographic evidence of loosening. With use of revision and loosening as end points, the eleven-year survival rates were 91.6% and 88.9%, respectively. Of the fifty implants that had no loosening, fourteen had measurable cup migration, thirty-five had no migration, and one implant could not be measured. All migrations but one were progressive. With loosening used as the end point, the survival rate at eleven years was 100% for the implants with no migration; however, the survival rate for the cups that had migrated was 69.3% (p = 0.0012). CONCLUSIONS: The eleven-year survival rate for the spherical press-fit cups in combination with bulk bone-grafting is satisfactory, given the complexity of these reconstructions. However, the difference between the survival of the implants that had migrated and those that had not was significant. We expect that the thirteen implants with progressive acetabular migration at the time of the latest follow-up are at risk for loosening, which will increase the revision rate for this series in the coming years.

Optimizing processing parameters for signal enhancement of oligonucleotide and protein arrays on ARChip Epoxy.

Signal enhancement of oligonucleotide and protein arrays on ARChip Epoxy was achieved by optimizing chip processing parameters. The parameters investigated were fabrication, blocking and guide dot concentration, probe concentration and modification, print buffer, humidity during arraying, slide agitation, spot volume and spotter compatibility. The optimum oligonucleotide concentration was 20 microM, while the optimum protein concentration was 0.05 mg/ml. Amino-modified oligonucleotides were best able to be bound to the resin's epoxy groups at pH 8, whereas thiol-modified oligonucleotides displayed an optimum coupling value of pH 7. So as to avoid background (BG) contamination of probes around bright guide dots, the concentration of fluorescent guide dots was set to 1 muM. The most suitable print buffers for oligonucleotide arrays using both piezo- and contact-printing systems proved to be 3 x SSC/1.5 M betaine and commercial ArrayLink. When 0.01% monochlortriazinyl-beta-cyclodextrin sodium salt (MCT) was added, the hybridization signal doubled in strength as compared to plain buffer. The optimum print buffer for proteins was 0.1 N phosphate buffer, pH 8/10% glycerine. The optimum humidity for arraying oligonucleotides was 60% and for proteins 40%. Initially agitating slides for 15 min was found just as effective as agitating slides over the total hybridization period (2.5 h), and this resulted in a three times stronger signal.

A phase I clinical and pharmacokinetic study of the camptothecin glycoconjugate, BAY 38-3441, as a daily infusion in patients with advanced solid tumors.

BACKGROUND: The aim of this study was to define the maximum tolerated dose (MTD), dose-limiting toxicity (DLT) and pharmacokinetics of the camptothecin glycoconjugate BAY 38-3441, administered as an infusion for 30 min on two separate schedules every 3 weeks. PATIENTS AND METHODS: A total of 81 patients with advanced solid tumors were treated with BAY 38-3441 either at doses of 20, 40, 67, 100, 140, 210, 315, 470 and 600 mg/m2/day for 1 day every 3 weeks (single-dose schedule), or at doses of 126, 189, 246, 320 and 416 mg/m2/day once daily for three consecutive days every 3 weeks (3-day schedule). Plasma sampling was performed to characterize the pharmacokinetics of BAY 38-3441 and camptothecin with these schedules. RESULTS: DLTs included renal toxicity, granulocytopenia and thrombocytopenia on the single-day schedule at doses > or = 470 mg/m2/day, and diarrhea and thrombocytopenia on the 3-day schedule at doses > or = 320 mg/m2/day. Other non-DLTs were gastrointestinal, dermatological and hematological. Pharmacokinetics of BAY 38-3441 and camptothecin appear to be dose-dependent, but not linear. CONCLUSIONS: Renal toxicity was dose-limiting for BAY 38-3441 using 30-min infusions on the single-dose schedule. Dose escalation to 470 mg/m2/day is feasible using a 2-h infusion. However, because of the superior safety profile, we recommend the 3-day schedule for BAY 38-3441 at a dose of 320 mg/m2/day as 30-min infusions for further phase II studies.

[Long-term results of uncemented Parhofer-Mönch screw rings]. / Langzeitergebnisse der zementfreien Parhofer-Mönch Schraubpfanne.

AIM: The aim of this study was an analysis of the long-term behaviour and implant migration of the Parhofer-Mönch-screw cup seen in patients between 1982 and 1991. METHOD: 92 cups (m : f = 53 : 39, mean age 53 +/- 7 years) were included mainly prospectively. After 118 +/- 45 months all patients were examined clinically and radiologically. Digital migration analysis was performed using the single-film X-ray analysis (Einbildröntgenanalyse, EBRA). RESULTS: 5 patients had died. 32 cups were revised, in 7 patients a loosening of the cup was suspected. The 10-year-survival was 71.4 %. In 53 of 81 analysed cups a migration of more than 1 mm was shown, 28 cups did not migrate. In comparison to these stable implants the survival of migrated cups was significantly inferior. CONCLUSION: The 10-year-survival and the high rate of implant migration document the poor results of the PM cup. In spite of an extraordinary primary stability, the failure of secondary osseointegration represents the main cause of loosening in this type of cup.

Overexpression of the ATP-dependent helicase RecG improves resistance to weak organic acids in Escherichia coli.

Increased resistance to several weak organic acids was conferred on Escherichia coli by overexpression of the ATP-dependent helicase RecG and, to a lesser extent, by overexpressing the helicase RuvAB. This property of helicases was identified by reproducible selection of recG-bearing clones from genomic libraries of the acetate-resistant species Acetobacter aceti and Staphylococcus capitis. We show that overexpression of RecG from both species, but also from E. coli, increased the maximum biomass concentration attained by E. coli cultures that were grown in the presence of various weak organic acids and uncouplers. Furthermore, overexpression of RecG from A. aceti significantly improved the maximum growth rates of E. coli under weak organic acid challenge. Based on the known role of RecG in DNA replication/repair, our data provide a first indication that weak organic acids negatively affect DNA replication and/or repair, and that these negative effects may be counteracted by helicase activity.

Myocardial perfusion and coronary flow reserve assessed by positron emission tomography in patients after Fontan-like operations.

Ventricular dysfunction in patients after Fontan-like operations (FLOs) is a serious complication that might contribute to poor long-term results. Ischemic heart disease will have debilitating consequences on a Fontan heart. Ten patients (15.8 +/- 5.01 years) after FLO had transesophageal echocardiography and cardiac catheterization 9.3 +/- 4.2 years after surgery. Myocardial perfusion was assessed by NH3-positron emission tomography (rest/adenosine) and compared with that of 10 healthy adults (26.1 +/- 6.3 years). Ventricular function was normal in 4 and reduced in 6 patients; end systolic and end diastolic meridional wall stress was significantly elevated in the FLO group. Coronary angiography revealed no stenosis of the coronaries. Compared to normals, myocardial blood flow (MBF) at rest was higher in the FLO group (0.99 +/- 0.25 vs 0.77 +/- 0.17 ml/g/min, p <0.05), whereas MBF after vasodilatation (2.12 +/- 0.78 vs 3.10 +/- 0.85 ml/g/min, p <0.05) and coronary flow reserve (CFR) was reduced (2.5 +/- 0.88 vs 4.1 +/- 1.01, p <0.05), especially in those with impaired ventricular function. Coronary vascular resistance after vasodilatation was elevated in the FLO group (38.2 +/- 17.4 vs 24.5 +/- 8.3 mmHg/ml/g/min, p <0.05). Altered MBF, increased meridional wall stress, and impaired CFR are common findings in FLO. Attenuated CFR and reduced ventricular function are significantly correlated and may be risk factors for the long-term outcome.

Metabolic flux response to phosphoglucose isomerase knock-out in Escherichia coli and impact of overexpression of the soluble transhydrogenase UdhA.

Blocking glycolytic breakdown of glucose by inactivation of phosphoglucose isomerase (Pgi) in Escherichia coli led to a greatly reduced maximum specific growth rate. Examination of the operational catabolic pathways and their flux ratios using [U-(13)C(6)]glucose-labeling experiments and metabolic flux ratio analysis provide evidence for the pentose phosphate (PP) pathway as the primary route of glucose catabolism in the knock-out mutant. The resulting extensive flux through the PP pathway disturbs apparently the reducing power balance, since overexpression of the recently identified soluble transhydrogenase UdhA improves significantly the growth rate of the Pgi mutant. The presented results provide first evidence that UdhA restores the cellular redox balance by catalyzing electron transfer from NADPH to NADH.

White paper on the future chemical policy in the European Union - implications for animal welfare.

In February 2001, the European Commission adopted a White Paper on a Future Chemicals Policy. Its main goals are better to protect humans and the environment from unknown risks through chemicals. The "promotion of non-animal testing" is one of the key elements of the proposed strategy. For low production volume chemicals, only data from in vitro-tests are to be requested. The data requirements for higher production volume chemicals shall be designed flexibly so that only data relevant for the respective chemical are collected. From the point of view of animal welfare concrete risk management strategies should be defined before test batteries are put together. The test catalogues currently listed in the Classification Directive 67/548/EEC are to be replaced by tiered testing strategies, and concrete waiving strategies are to be designed so that the requested tailor-made testing can actually be realized. Another essential prerequisite for the promotion of non-animal testing is that the funding of alternative method research is formulated as a key action with a concrete budget in the Sixth Research Framework Program of the European Union.

Proteins induced during adaptation of Acetobacter aceti to high acetate concentrations.

As a typical product of microbial metabolism, the weak acid acetate is well known for its cytotoxic effects. In contrast to most other microbes, the so-called acetic acid bacteria can acquire significant resistance to high acetate concentrations when properly adapted to such hostile conditions. To characterize the molecular events that are associated with this adaptation, we analyzed global protein expression levels during adaptation of Acetobacter aceti by two-dimensional gel electrophoresis. Adaptation was achieved by using serial batch and continuous cultivations with increasing acetate supplementation. Computer-aided analysis revealed a complex proteome response with at least 50 proteins that are specifically induced by adaptation to acetate but not by other stress conditions, such as heat or oxidative or osmotic stress. Of these proteins, 19 were significantly induced in serial batch and continuous cultures and were thus noted as acetate adaptation proteins (Aaps). Here we present first microsequence information on such Aaps from A. aceti. Membrane-associated processes appear to be of major importance for adaptation, because some of the Aap bear N-terminal sequence homology to membrane proteins and 11 of about 40 resolved proteins from membrane protein-enriched fractions are significantly induced.

Immunohistochemical analysis of major TGF-beta isoforms and their receptors in laryngeal carcinomas.

We analyzed immunohistochemically the tissue distribution of the three major transforming growth factors-beta isoforms (TGF-beta1, -2, -3) and their receptors (TBR-I, -II and -III) in tissue samples from 38 patients with laryngeal squamous cell carcinomas. Besides a qualitative evaluation, the number of the respectively labeled cells was determined by morphometric analysis. In all tumor samples a significant staining of most tumor cells was seen both for the TGF-beta isoforms and the TBRs. Similarly, the majority of stromal cells were labeled. On semiserial sections, there were only minor differences in the distribution pattern and in the number of labeled cells between the three TGF-beta isoforms and the TBRs, suggesting that most tumor cells are actively involved in the neosynthesis of TGF-betas and TBRs; accordingly, at least most tumor cells seem to be capable of producing more than one TGF-beta form and in parallel several TBRs. With decreasing tumor cell differentiation the number of TGF-beta- and TBR-positive tumor cells decreased slightly (but not to a statistically significant degree). Interestingly, the stromal cells were labeled for TGF-betas and TBRs to a lower extent than the epithelial cells, and there was no significant difference between non-tumor-associated control stroma and the immediate peritumoral stroma. Our observations suggest an even, enhanced level of TGF-beta production in laryngeal squamous cell carcinomas, which may explain some well-known side-effects of tumor growth, such as stromal desmoplasia. In addition, the presence of immunoreactive TBR-proteins in the vast majority of tumor cells excludes the mere absence of TBRs in those carcinomas as the cause for inappropriate TGF-beta function in the tumor cells. This in turn suggests that molecular alterations either of the TBR-proteins non-affecting the synthesis and turnover or downstream alterations of the TGF-beta signaling pathway may be main reasons for the loss of response of the tumor cells to the enhanced amounts of TGF-betas.

Bacillus subtilis metabolism and energetics in carbon-limited and excess-carbon chemostat culture.

The energetic efficiency of microbial growth is significantly reduced in cultures growing under glucose excess compared to cultures growing under glucose limitation, but the magnitude to which different energy-dissipating processes contribute to the reduced efficiency is currently not well understood. We introduce here a new concept for balancing the total cellular energy flux that is based on the conversion of energy and carbon fluxes into energy equivalents, and we apply this concept to glucose-, ammonia-, and phosphate-limited chemostat cultures of riboflavin-producing Bacillus subtilis. Based on [U-(13)C(6)]glucose-labeling experiments and metabolic flux analysis, the total energy flux in slow-growing, glucose-limited B. subtilis is almost exclusively partitioned in maintenance metabolism and biomass formation. In excess-glucose cultures, in contrast, uncoupling of anabolism and catabolism is primarily achieved by overflow metabolism, while two quantified futile enzyme cycles and metabolic shifts to energetically less efficient pathways are negligible. In most cultures, about 20% of the total energy flux could not be assigned to a particular energy-consuming process and thus are probably dissipated by processes such as ion leakage that are not being considered at present. In contrast to glucose- or ammonia-limited cultures, metabolic flux analysis revealed low tricarboxylic acid (TCA) cycle fluxes in phosphate-limited B. subtilis, which is consistent with CcpA-dependent catabolite repression of the cycle and/or transcriptional activation of genes involved in overflow metabolism in the presence of excess glucose. ATP-dependent control of in vivo enzyme activity appears to be irrelevant for the observed differences in TCA cycle fluxes.

Stoichiometric growth model for riboflavin-producing Bacillus subtilis.

Rate equations for measured extracellular rates and macromolecular composition data were combined with a stoichiometric model to describe riboflavin production with an industrial Bacillus subtilis strain using errors in variables regression analysis. On the basis of this combined stoichiometric growth model, we explored the topological features of the B. subtilis metabolic reaction network that was assembled from a large amount of literature. More specifically, we simulated maximum theoretical yields of biomass and riboflavin, including the associated flux regimes. Based on the developed model, the importance of experimental data on building block requirements for maximum yield and flux calculations were investigated. These analyses clearly show that verification of macromolecular composition data is important for optimum flux calculations.

Metabolic flux analysis with a comprehensive isotopomer model in Bacillus subtilis.

Fluxes in central carbon metabolism of a genetically engineered, riboflavin-producing Bacillus subtilis strain were investigated in glucose-limited chemostat cultures at low (0.11 h(-1)) and high (0.44 h(-1)) dilution rates. Using a mixture of 10% [U-(13)C] and 90% glucose labeled at natural abundance, (13)C-labeling experiments were carried out to provide additional information for metabolic flux balancing. The resulting labeling pattern in the proteinogenic amino acids were analyzed by two-dimensional [(13)C, (1)H] nuclear magnetic resonance (NMR) spectroscopy. To account rigorously for all available data from these experiments, we developed a comprehensive isotopomer model of B. subtilis central metabolism. Using this model, intracellular carbon net and exchange fluxes were estimated on the basis of validated physiological data and biomass composition in combination with 2D NMR data from 45 individual carbon atom spectra in the amino acids. Glucose catabolism proceeded primarily via glycolysis but pentose phosphate pathway fluxes increased with increasing growth rate. Moreover, significant back fluxes from the TCA cycle to the lower part of glycolysis via the gluconeogenic PEP carboxykinase were detected. The malic enzyme reaction, in contrast, was found to be inactive. A thorough statistical analysis was performed to prove the reliability of the isotopomer balance model and the obtained results. Specifically, a chi(2) test was applied to validate the model and the chi-square criterion was used to explore the sensitivity of model predictions to the experimental data.

Quantitation of TGF-beta1 mRNA in porcine mesangial cells by comparative kinetic RT/PCR: comparison with ribonuclease protection assay and in situ hybridization.

Gene expression can be examined with different techniques including ribonuclease protection assay (RPA), in situ hybridisation (ISH), and quantitative reverse transcription-polymerase chain reaction (RT/PCR). These methods differ considerably in their sensitivity and precision in detecting and quantifying low abundance mRNA. Although there is evidence that RT/PCR can be performed in a quantitative manner, the quantitative capacity of this method is generally underestimated. To demonstrate that the comparative kinetic RT/PCR strategy-which uses a housekeeping gene as internal standard-is a quantitative method to detect significant differences in mRNA levels between different samples, the inhibitory effect of heparin on phorbol 12-myristate 13-acetate (PMA)-induced-TGF-beta1 mRNA expression was evaluated by RT/PCR and RPA, the standard method of mRNA quantification, and the results were compared. The reproducibility of RT/PCR amplification was calculated by comparing the quantity of G3PDH and TGF-beta1 PCR products, generated during the exponential phases, estimated from two different RT/PCR (G3PDH, r = 0.968, P = 0.0000; TGF-beta1, r = 0.966, P = 0.0000). The quantitative capacity of comparative kinetic RT/PCR was demonstrated by comparing the results obtained from RPA and RT/PCR using linear regression analysis. Starting from the same RNA extraction, but using only 1% of the RNA for the RT/PCR compared to RPA, significant correlation was observed (r = 0.984, P = 0.0004). Moreover the morphometric analysis of ISH signal was applied for the semi-quantitative evaluation of the expression and localisation of TGF-beta1 mRNA in the entire cell population. Our results demonstrate the close similarity of the RT/PCR and RPA methods in giving quantitative information on mRNA expression and indicate the possibility to adopt the comparative kinetic RT/PCR as reliable quantitative method of mRNA analysis.

Myocardial blood flow and flow reserve after coronary reimplantation in patients after arterial switch and ross operation.

BACKGROUND: Coronary reimplantation is used in therapy for congenital heart disease, such as in the arterial switch (ASO) and Ross operations. The adequacy of myocardial perfusion may remain a matter of concern. The aim of the present study was to stratify the effect of coronary reimplantation on myocardial perfusion and to highlight the clinical relevance of any attenuation in myocardial perfusion. METHODS AND RESULTS: A total of 21 children with transposition of the great arteries at a mean interval of 11.2+/-2.9 years after ASO and 9 adolescents at a mean interval of 4.2+/-2.1 years after the Ross procedure were investigated. All patients were asymptomatic and had a normal exercise capacity. On stress echocardiography, 2 of the ASO patients had dyskinetic areas within the left ventricular myocardium, and 5 had adenosine-induced perfusion defects on positron emission tomography. No coronary obstruction was detected on coronary angiography in any patient, but a common finding was right coronary dominance and a small caliber of the distal part of the left anterior descending artery. Coronary flow reserve (CFR) was significantly reduced in all patients after ASO when compared with 10 normal healthy volunteers (age, 25.6+/-5.3 years). CFR was normal in the 9 patients who had the Ross operation (age, 19.2+/-7.6 years); exercise-induced perfusion defects were not detected in the Ross patients. CONCLUSIONS: Children after ASO are asymptomatic, without clinical signs of coronary dysfunction. In contrast to patients who had the Ross operation, stress-induced perfusion defects and an attenuated CFR were documented. The prognostic implications of these findings and the clinical consequences are unclear; nevertheless, close clinical follow-up of ASO patients is mandatory.

Central carbon metabolism of Saccharomyces cerevisiae explored by biosynthetic fractional (13)C labeling of common amino acids.

Aerobic and anaerobic central metabolism of Saccharomyces cerevisiae cells was explored in batch cultures on a minimal medium containing glucose as the sole carbon source, using biosynthetic fractional (13)C labeling of proteinogenic amino acids. This allowed, firstly, unravelling of the network of active central pathways in cytosol and mitochondria, secondly, determination of flux ratios characterizing glycolysis, pentose phosphate cycle, tricarboxylic acid cycle and C1-metabolism, and thirdly, assessment of intercompartmental transport fluxes of pyruvate, acetyl-CoA, oxaloacetate and glycine. The data also revealed that alanine aminotransferase is located in the mitochondria, and that amino acids are synthesized according to documented pathways. In both the aerobic and the anaerobic regime: (a) the mitochondrial glycine cleavage pathway is active, and efflux of glycine into the cytosol is observed; (b) the pentose phosphate pathways serve for biosynthesis only, i.e. phosphoenolpyruvate is entirely generated via glycolysis; (c) the majority of the cytosolic oxaloacetate is synthesized via anaplerotic carboxylation of pyruvate; (d) the malic enzyme plays a key role for mitochondrial pyruvate metabolism; (e) the transfer of oxaloacetate from the cytosol to the mitochondria is largely unidirectional, and the activity of the malate-aspartate shuttle and the succinate-fumarate carrier is low; (e) a large fraction of the mitochondrial pyruvate is imported from the cytosol; and (f) the glyoxylate cycle is inactive. In the aerobic regime, 75% of mitochondrial oxaloacetate arises from anaplerotic carboxylation of pyruvate, while in the anaerobic regime, the tricarboxylic acid cycle is operating in a branched fashion to fulfill biosynthetic demands only. The present study shows that fractional (13)C labeling of amino acids represents a powerful approach to study compartmented eukaryotic systems.