Cultivation of Aspergillus niger in a pilot plant external-loop air-lift bioreactor

Abstract: Changes of the main hydrodynamic and oxygen transfer parameters during Aspergillus niger cultivation in an external-loop air-lift bioreactor of 200 dm³ operating capacity were investigated. The final average concentrations of biomass and citric acid obtained in batch fermentations were about 17 g 1^-1 and 90 g 1^-1, respectively. Significant influence of the increasing biomass concentration on the rheological properties of the broth and operating parameters was found. Volumetric oxygen transfer coefficient. k ^L a , was found to be dependent on the apparent viscosity of the broth with an exponent of -0.984.     

Physiological effects of five months exposure to low concentrations of O3 and NH3 on Douglas fir (Pseudotsuga menziesii)

Three years old seedlings of Douglas fir (Pseudotsuga menziesii) were exposed lo filtered air, O₃ (day and night concentrations of 78 and 30 μgm^?3: respectively). NH₃ (54 μg m^?3) and to a mixture of NH₃+O₃ (day and night concentrations of 49 + 83 and 49 + 44 μg m^?3 respectively), for 5 months in fumigation chambers. Both gas exchange and chlorophyll fluorescence were measured on shoots which had sprouted at the beginning of the exposure period. After 4. 8, 10 and 20 weeks of exposure, light response curves of electron transport rate (J) were determined, in which J was deduced from chlorophyll fluorescence. Net CO₂ assimiialion was measured at maximum light intensity of 560) μmol m^?2 S^?1 (P_n.560). After 8 and 10 weeks of exposure also light response curves of CO₂ assimilation were assessed. Shoots exposed to O₃ showed a reduction in net CO₂ assimilation as compared to the control shoots during the entire exposure period. The reduction was related lo a lower chlorophyll content and a lower electron transport rate, whereas no effect on quantum yield efficiency (qy) was observed. In contrast, shoots exposed to NH₃ showed a positive effect on photosynthesis. Shoots exposed to NH₃. + O₃ showed a rapid increase in P_n.560, in the period between 4 and 8 weeks to a level equal of that of the NH₃-treatment. After this period a decline in P_n.560 was observed. After 10 weeks of exposure shoots exposed to O₃ showed an increased transpiration rate in the dark as compared to the control shoots. In addition, water use efficiency (WUE) declined as a result of an increase in leaf conductance. Both observations indicate that the stomatal apparatus was affected by O₃. A high transpiration rate in the dark was also found for shoots esposed to NHX. However, shoots exposed to NH₃+ O₃ showed neither an effect on WUE, nor an effect on transpiration rate in the dark. The possibility that NH₃ delayed the O₃ induced effects on photosynthesis and stomatal conductance is discussed.     

Analysis of DNA polymerase activity in Petunia protoplasts treated with clastogenic agents

Clastogenic agents, i.e. agents that can induce chromosome or DNA breakage, have been shown to enhance the rale of direct gene transfer to protoplasts. The effect was analysed at the enzymatic level using protoplast homogenates as well as intact protoplasts. For that purpose existing procedures were modified to enable measurement of DNA polymerase in vivo. In the system used, external DNA was able to enter the cells without the addition of membrane-permeabilizing compounds. When comparing total DNA polymerase activity of protoplasts irradiated with X-rays or UV-light with that of untreated cells we did not observe significant differences. Incubation of protoplasts with high doses of bleomycin affected total DNA polymerase activity negatively. but dideoxythymidine triphosphate-sensitive activity was not influenced. We conclude that the DNA strand-breaks induced by low doses of X-rays. UV-light or bleomycin do not increase the total or the repair-DNA polymerase activity and. therefore. that the increase in the transformation rates after DNA strand-breaking is not preceded by enhanced DNA polymerase activity.     

Enzyme IIBcellobiose of the phosphoenol-pyruvate-dependent phosphotransferase system of Escherichia coli: backbone assignment and secondary structure determined by three-dimensional NMR spectroscopy.

The assignment of backbone resonances and the secondary structure determination of the Cys 10 Ser mutant of enzyme IIBcellobiose of the Escherichia coli cellobiose-specific phosphoenol-pyruvate-dependent phosphotransferase system are presented. The backbone resonances were assigned using 4 triple resonance experiments, the HNCA and HN(CO)CA experiments, correlating backbone 1H, 15N, and 13C alpha resonances, and the HN(CA)CO and HNCO experiments, correlating backbone 1H,15N and 13CO resonances. Heteronuclear 1H-NOE 1H-15N single quantum coherence (15N-NOESY-HSQC) spectroscopy and heteronuclear 1H total correlation 1H-15N single quantum coherence (15N-TOCSY-HSQC) spectroscopy were used to resolve ambiguities arising from overlapping 13C alpha and 13CO frequencies and to check the assignments from the triple resonance experiments. This procedure, together with a 3-dimensional 1H alpha-13C alpha-13CO experiment (COCAH), yielded the assignment for all observed backbone resonances. The secondary structure was determined using information both from the deviation of observed 1H alpha and 13C alpha chemical shifts from their random coil values and 1H-NOE information from the 15N-NOESY-HSQC. These data show that enzyme IIBcellobiose consists of a 4-stranded parallel beta-sheet and 5 alpha-helices. In the wild-type enzyme IIBcellobiose, the catalytic residue appears to be located at the end of a beta-strand.     

Is there an energy conservation “system” in brain that protects against the consequences of energy depletion?

A poorly understood marked decrease (circa 50% of control) in local cerebral glucose utilization is caused by sublethal doses of NaCN. The decrease is global, occurring in essentially all brain regions and is entirely reversible within hours, leaving no obvious pathology. This event is not unique to NaCN in so far as a strikingly similar pattern of decreased glucose utilization occus with some other toxins. Nor can it be attributed to a direct action of NaCN since local application by microdialysis to the striatum produces a global depression. These results imply that some widely distributed system or substance is involved. We speculate the existence of a system possibly related to the reticular activating system that senses a fall in energy production and acts globally to make cells quiescent and thus would give some protection from excitotoxic driven damage.Special issue dedicated to Dr. Sidney Ochs.     

Chiral copper(I)—thiolate clusters in metallothionein and glutathione

Metallothionein (MT) is a ubiquitous mammalian protein comprising 61 or 62 nonaromatic amino acids of which 20 are cysteine residues. The high sulfhydryl content imparts to this protein a unique and remarkable ability to bind multiple metal ions in structurally significant metal–thiolate clusters. MT can bind seven divalent metal ions per protein molecule in two domains with exclusive tetrahedral metal coordination. The domain stoichiometries for the M₇S₂₀ structure are M₄(S_cys)₁₁ (α domain) and M₃(S_cys)₉ (β domain). Up to 12 Cu(I) ions can displace the 7 Zn²⁺ ions bound per molecule in Zn₇–MT. The incoming Cu(I) ions adopt a trigonal planar geometry with domain stoichiometries for the Cu₁₂S₂₀ structure of Cu₆(S_cys)₁₁ and Cu₆(S_cys)₉ for the α and β domains, respectively. The circular dichroism (CD) spectra recorded as Cu⁺ is added to Zn₇–MT to form Cu₁₂–MT directly report structural changes that take place in the metal binding region. The spectrum arises under charge transfer transitions between the cysteine S and the Cu(I); because the Cu(I)–thiolate cluster units are located within the chiral binding site, intensities in the CD spectrum are directly related to changes in the binding site. The CD technique clearly indicates stoichiometries of several Cu(I)–MT species. Model Cu(I)–thiolate complexes, using the tripeptide glutathione as the sulfhydryl source, were examined by CD spectroscopy to obtain transition energies and the Cu(I)–thiolate coordination geometries which correspond to these bands. Possible structures for the Cu(I)–thiolate clusters in the α and β domains of Cu₁₂–MT are proposed. © 1994 Wiley-Liss, Inc.     

Design of a bubble-swawm bioreactor for animal cell culture

A stationary bubble-swarm has been used to aerate a mammalian cell culture bioreactor with an extremely low gas flow rate. Prolonging the residence time of the gas bubbles within the medium improved the efficiency of the gas transfer into the liquid phase and suppressed foam formation. An appropriate field of speed gradients prevented the bubbles from rising to the surface. This aeration method achieves an almost 90% transfer of oxygen supplied by the bubbles. Consequently, it is able to supply cells with oxygen even at high cell densities, while sparging with a gas flow of only 0.22·10^–3–1.45·10^–3 vvm (30–200 ml/h).The reactor design, the oxygen transfer rates and the high efficiency of the system are presented. Two repeated batch cultures of a rat-mouse hybridoma cell line are compared with a surface-aerated spinner culture. The used cell culture medium was serum-free, either with or without BSA and did not contain surfactants or other cell protecting agents. One batch is discussed in detail for oxygen supply, amino acid consumption and specific antibody production.     

Nonlinear effect of dynamic self-organization in macromolecular systems caused by photocontrolled electron flux

We use the electron-conformational interaction approach to develop a physical model which self-consistently describes the photomobilized electron transfer kinetics and structure conformational transitions in reaction centers (RCs) of purple bacteria. We consider the kinetics of electron transition from pigment onto primary acceptor and the subsequent charge recombination accounting for the change of distance between the above-mentioned cofactors. It is shown that, given natural values of RC parameters, the kinetic constant's dependence on the acting light intensity is monotone. As opposed to the previous case, similar dependencies for the chain of electron transfer between primary and secondary quinone acceptors revealed anS-like relationship. This can lead to bistability of the RC optical transmission coefficient and a fundamental dependence of charge recombination kinetics upon the prehistory of the RC's interaction with exciting radiation.     

The Response to Exercise with Constant Energy Intake in Identical Twins

Seven pairs of young adult male identical twins completed a negative energy balance protocol during which they exercised on cycle ergometers twice a day, 9 out of 10 days, over a period of 93 days while being kept on a constant daily energy and nutrient intake. The total energy deficit caused by exercise above the estimated energy cost of body weight maintenance reached 244 ± 9.8 MJ (Mean ± SEM). Baseline energy intake was estimated over a period of 17 days preceding the negative energy balance protocol. Mean body weight loss was 5.0 kg (SEM = 0.6) (p <0.001) and it was entirely accounted for by the loss of fat mass (p <0.001). Fat-free mass was unchanged. Body energy losses reached 191 MJ (SEM = 24) (p <0.001) which represented about 78% of the estimated energy deficit. Subcutaneous fat loss was slightly more pronounced on the trunk than on the limbs as estimated from skinfolds, circumferences, and computed tomography (CT). The reduction in CT-assessed abdominal visceral fat was quite striking, from 81 cm² (SEM = 5) to 52 cm² (SEM = 6) (p <0.001). At the same submaximal power output level, subjects oxidized more lipids than carbohydrates after the program as indicated by the changes in the respiratory exchange ratio (p <0.05). Intrapair resemblance was observed for the changes in body weight (p <0.05), fat mass (P <0.01), percent fat (p <0.01), body energy content (p <0.01), sum of 10 skinfolds (p <0.01), abdominal visceral fat (p <0.01), fasting plasma triglycerides (p <0.05) and cholesterol (p <0.05), maximal oxygen uptake (p <0.05), and respiratory exchange ratio during submaximal work (p <0.01). We conclude that even though there were large individual differences in response to the negative energy balance and exercise protocol, subjects with the same genotype were more alike in responses than subjects with different genotypes particularly for body fat, body energy, and abdominal visceral fat changes. High lipid oxidizers and low lipid oxidizers during sub-maximal exercise were also seen despite the fact that all subjects had experienced the same exercise and nutritional conditions for about three months.     

Transfer of a marine DNA virus fromEctocarpus toFeldmannia (Ectocarpales,Phaeophyceae): aberrant symptoms and restitution of the host

Summary The marine brown algaEctocarpus siliculosus is invaded by a polyhedric virus, whose genome consists of circular, double-stranded DNA. In laboratory experiments this virus can infect a different host species,Feldmannia simplex. InfectedFeldmannia plants show severe somatic malformations. However, no functional virus particles are formed. SuchFeldmannia plants recover to resume a normal, symptom-free appearance. This result raises the possibility of intergeneric gene transfer in the natural habitat.