Effect of oxygen on ethanol fermentation in packed-bed tapered-column reactor

Summary In ethanol production with immobilized yeast a major problem is the provision of nutrients to these highly concentrated cells. O₂ being one of the nutrients of utmost importance to yeast cells, was fed into a column packed with beads with a cell loading of more than 40 g/l. Since addition of large volume of air or O₂ to a cylindrical column reactor would aggravate the problems of pressure build up and channelling caused by the evolving CO₂ gas, a tapered-column reactor and pulsed flow of oxygen gas was used. The supplement of O₂ gas to the tapered column increased the productivity from 21.1 g ethanol x (l gel x h)^-1 to 26.7 g x (l gel x h)^-1, when the ethanol concentration at the outlet was about 80 g/l. The yield coefficient of ethanol was also increased from 0.41 g ethanol/g glucose to 0.43 after O₂ supplement was started. The effects of frequency and duration of O₂ supplement were also determined.     

Direct Evidence for Changes in the Resistance of Legume Root Nodules to O2 Diffusion

Indirect evidence suggests that legumes can adjust rapidly theresistance of their root nodules to O₂ diffusion. Here we describeexperiments using O₂ specific micro-electrodes and dark fieldmicroscopy to study directly the operation of this diffusionbarrier. The O₂ concentration sensed by the electrode decreasedsharply in the region of the inner cortex and was less than1.0 mmol m^–3 throughout the infected tissue in nodulesof both pea (Pisum sativum) and french bean (Phaseolus vulgaris).In a number of experiments the ambient O₂ concentration wasincreased to 40% while the electrode tip was just inside theinner cortex. In 13 out of 21 cases the O₂ concentration atthis position either remained low and unchanged or increasedirreversibly to near ambient values. In the remaining casesthe O₂ concentration increased after 1 to 2.5 min and then decreasedto its former value. These results are ascribed to an increasein resistance of the barrier in response to increased O₂ fluxinto the nodule. It was shown microscopically that air spacesboth at the boundary between the infected zone and the innercortex, and within the infected zone started to disappear 3min after nodules were exposed to high ambient O₂ concentrationsand had disappeared completely after 8 min. These spaces werenot changed by exposure of the nodule for 10 min to either N₂or air. Key words: Oxygen, root nodules, air spaces     

Evolution of the 28S ribosomal RNA gene in anurans: regions of variability and their phylogenetic implications

Fifteen restriction sites were mapped to the 28S ribosomal RNA gene of individuals representing 54 species of frogs, two species of salamanders, a caecilian, and a lungfish. Eight of these sites were present in all species examined, and two were found in all but one species. Alignment of these conserved restriction sites revealed, among anuran 28S rRNA genes, five regions of major length variation that correspond to four of 12 previously identified divergent domains of this gene. One of the divergent domains (DD8) consists of two regions of length variation separated by a short segment that is conserved at least throughout tetrapods. Most of the insertions, deletions, and restriction-site variations identified in the 28S gene will require sequence-level analysis for a detailed reconstruction of their history. However, an insertion in DD9 that is coextensive with frogs in the suborder Neobatrachia, a BstEII site that is limited to representatives of two leptodactylid subfamilies, and a deletion in DD10 that is found only in three ranoid genera are probably synapomorphies.      

Evidence for inhibitory SH groups in the thiol activated K:Cl cotransporter of low K sheep red blood cells

The monofunctional thiol reagents N-ethylmaleimide (NEM) and methyl methanethiosulfonate (MMTS) stimulate ouabain resistant (OR) electroneutral K:Cl cotransport in LK sheep red blood cells at low, but not at high concentrations. Diamide (DM), on the other hand, only stimulates OR K:Cl flux (Lauf, P.K., J. Memb. Biol. 101: 179–188, 1988). The DM stimulated K:Cl cotransport was decreased toward the control value prior to DM stimulation when NEM or MMTS were added, subsequently. The inhibitory effect was dependent on the compound's concentration and exposure time and, in the case of MMTS, was reversed upon addition of dithiothreitol (DTT). The inhibition was more prominent when NEM treatment was performed at pH 8.0 and disappeared at pH 6.0. In contrast the NEM stimulatory effect was most effective when the pH of NEM treatment was 6.0 (Bauer, J. & Lauf, P.K., J. Memb. Biol. 73: 257–261, 1983). The results suggest the existence of additional, however, inhibitory thiol groups in the already thiol-activated K:Cl cotransporter, with a different pKa value and a lower affinity for NEM or MMTS as compared to the stimulatory thiol groups. Like the activating thiols, the inhibitory sulfhydryls appeared to be inaccessible to non-penetrating thiol reagents and hence, must be located deeper within the red cell membrane.     

Monoclonal antibodies to three phospholipase C isozymes from bovine brain

Murine hybridoma cell lines secreting antibodies against the three bovine isozymes of phosphoinositide-specific phospholipase C (PLC) were established: 6, 23, and 12 lines were obtained for PLC-I (150 kDa), PLC-II (145 kDa), and PLC-III (85 kDa), respectively. The antibodies were purified from ascites fluid, and their properties were studied in detail. All the antibodies cross-reacted with their corresponding PLC enzymes, but not with the other two isozymes, suggesting that the three enzymes contain very different antigenic determinants. The six antibodies elicited by bovine PLC-I also cross-reacted with human and rat enzyme, whereas three each from anti-PLC-II antibodies and anti-PLC-III antibodies did not react with the enzymes from different species. Each antibody exerts different effects on the phosphatidylinositol-hydrolyzing activity of PLC. The most inhibitory antibody for either isozyme PLC-I or PLC-II exhibits 80% inhibition, whereas no more than 20% inhibition was observed for the anti-PLC-III antibodies. Purified PLC-I frequently contains catalytically active 140- and 100-kDa forms and an inactive 41-kDa protein in addition to the intact 150-kDa form, probably due to its high sensitivity to an unidentified endogenous protease. The five anti-PLC-I antibodies which bind to the denatured 150-kDa polypeptide also recognized the 140-kDa form, whereas only three cross-reacted with the 100-kDa form, and the remaining two bound to the 41-kDa protein. Competitive binding studies with intact PLC enzymes and Western blot experiments with proteolytic digests revealed that the 6 anti-PLC-I, 23 anti-PLC-II, and 12 anti-PLC-III antibodies bind at least five, six, and seven different epitopes on PLC-I, PLC-II, and PLC-III, respectively. The fact that these monoclonal antibodies bind to different epitopes on the same enzyme allowed one to develop a highly specific and sensitive tandem radioimmunoassay for quantitating PLC-I, PLC-II, and PLC-III. The principle of the assay is that binding of an 125I-labeled antibody to the antigen immobilized by another antibody at a distinctive binding site is proportional to the amount of antigen present. By using this method, PLC-I, PLC-II, and PLC-III could be measured quantitatively in the presence of other proteins, detergents, lipids, polyanions, and metal ions, all of which greatly affect the activity of PLC enzymes.     

Serum IgE levels in rats infected with Paragonimus westermani]

Paragonimus westermani is a common fluke in Korea. The present study aimed to determine serum total IgE and specific IgG levels in experimental paragonimiasis of rats. Each Wistar rat was inoculated orally with 20-25 metacercariae of P. westermani from Cambaroides similis. Before and after infection (1, 2, 3, 4, 6, 8 weeks) of P. westermani, the blood was collected from the retro-orbital venous plexus of rats and kept serum at -70 degrees C. Serum total IgE and specific IgG levels were determined by the capture and conventional enzyme-linked immunosorbent assay, respectively. The results were as follows; 1. Serum IgE values were increased to 0.18 +/- 0.042 at 2 weeks, 0.28 +/- 0.151 at 4 weeks and 0.43 +/- 0.055 at 8 weeks after infection. The absorbances of non-infected rats ranged 0.07 +/- 0.021-0.12 +/- 0.025. 2. Specific IgG values were slightly increased at 3 weeks (0.20 +/- 0.032) and gradually increased up to 8 weeks (0.31 +/- 0.067) after infection. The absorbances of non-infected rats ranged 0.11 +/- 0.035-0.18 +/- 0.019. The present results suggested that P. westermani could elevate serum IgE and specific IgG antibodies in Wistar rats which were not a good definitive host.     

Application of an enzyme-linked immunosorbent assay for screening of T-2 toxin-producing Fusarium spp.

Culture filtrates of Fusarium species were subjected without clean-up procedures to an indirect competitive enzyme-linked immunosorbent assay with anti-T-2 toxin monoclonal antibody. Fusarium sporotrichioides, F. poae, F. tricinctum, and F. culmorum strains were positive for T-2 toxin, with a minimum detection limit of 5 pg per assay (100 pg/ml of culture filtrate), and the assay data correlated well with the gas-liquid chromatographic data.     

Recirculating bioreactor-separator system for simultaneous biotransformation and recovery of product: Immobilized L-aspartate beta-decarboxylase reactor system

A new combined bioreactor-separator system was designed and its operational feasibility demonstrated in order to develop a bioprocess that enables us to handle simultaneous biotransformation and recovery of product by crystallization. Enzymatic conversion of L-aspartate to L-alanine by L-aspartate beta-decarboxylase from Pseudomonas dacunhae (ATCC 21192) was used as a model system for this study to demonstrate the principles involved in the bioprocess design. Immobilized cells of P. dacunhae containing the enzyme were fluidized in a tapered column type of bioreactor and a filter-crystallizer combination was used as a separator unit in our experimental system.It was found that almost a theoretical yield was achieved, and the process control for both the bioreactor operation and separation was relatively easy. The Production systems, namely, the recirculating bioreactor separator combination system and the conventional batch reactor system, were analyzed and compared based on the results obtained form this study, and it was found that a significant cost reduction, by about 20%, can be achieved when the recirculating bioreactor-separator combination system was employed. Based on these findings, it is anticipated that the conceptual design of the bioreactor-separator combination system evaluated in this study has some potential for industrial application.