Mass transfer and biochemical reaction in enzyme membrane reactor systems—I. Single enzyme reactions

The effective use of an enzyme as a biological catalyst for process scale conversion is greatly enhanced by immobilizing the enzyme within the matrix of a membrane whose walls are permeable to the substrate and product. The enzyme is thus stabilized for reuse, but the apparent activity is reduced compared to that obtained with the free enzyme because of transport resistances in the bulk fluid near the membrane and within the membrane itself. To account for this reduction, solutions to the differential equations describing the mass transport-kinetic models can be obtained for certain important cases. The reaction is considered to be first-order and isothermal, and to occur in one of two modes of operation: fixed-bed or batch (limited volume). Irreversible and reversible reactions and plane sheet and spherical geometries of the membrane are considered. An example of the application of these solutions to actual data obtained with a laboratory scale immobilized enzyme reactor is presented.     

A/B-Ratio and Transport Properties of (La0.85Sr0.15)sCoO3−δ Perovskites

The perovskites (La_0.85Sr_0.15)_0.98CoO_{3 –} and (La_0.85Sr_0.15)_1.00CoO_{3 –} have been investigated using x-ray diffraction (XRD), scanning electron microscopy (SEM) and electrical conductivity relaxation (ECR). This system was chosen in order to investigate the influence of cation vacancies on the transport properties in the materials. From ECR-measurements it is concluded that no difference in the chemical diffusion coefficient for oxide ions between the two samples can be found. The activation energy for the chemical diffusion coefficient has been found to be 107 ± 5 kJ mol^{– 1}. However, the surface exchange coefficient differs between the two samples. The Co-rich sample has a significantly higher surface exchange coefficient than the A/B-stoichiometric sample. For both samples the surface exchange coefficient was almost independent of the temperature. At all temperatures the Co-rich sample had a significantly higher electrical conductivity. The difference in electrical conductivity between the two samples diminished when going to higher temperatures. Both materials can be assigned to a single phase hexagonal perovskite. An annealed sample of (La_0.85Sr_0.15)_0.98CoO_{3 –} did however contain an unidentified secondary phase on the surface.     

Phase-only optical decryption of a fixed mask

We present a system for the implementation of phase-only optical decryption of an encrypted fixed phase mask. We achieve decryption by superimposing a decrypting phase key, implemented on a phase-only spatial light modulator with an encrypted phase mask in two equivalent image planes in an optical system. The decrypted phase information is retrieved by the generalized phase-contrast technique. We have constructed a compact experimental system that uses a 635-nm diode laser in which a fixed encrypted 0/pi binary phase mask is decrypted by a binary phase key produced electronically on the spatial light modulator. The key is aligned by electronic scrolling of its position with respect to the mask.     

The mismatch negativity to frequency deviant stimuli during natural sleep

Eight subjects spent a single night in the sleep laboratory. Event-related potentials (ERPs) were recorded during the presentation of two auditory "oddball' stimulus conditions in which tonal frequency was manipulated. In the first condition, 1000 Hz "standard' and 2000 Hz "deviant' tones were presented. In the second condition, the deviant tone was reduced to 1050 Hz. In both conditions, deviant probability was 0.2. Stimuli were presented every 600 ms during wakefulness and stages 2, 4, and REM of sleep. A distinctive N1 wave was visible in both stimulus conditions when the subject was awake. The deviant stimuli elicited a "mismatch negativity' (MMN) that inverted in polarity at the mastoid. In REM sleep, an N1 and a MMN were also elicited in both conditions. In the large deviance condition, the MMN had a slightly attenuated amplitude and was shorter in duration while in the small deviant condition, its peak latency was unusually early. Neither the N1 nor the MMN could be recorded in non-REM sleep.     

Transmembrane passage of cytokine-inducing bacterial products across new and reprocessed polysulfone dialyzers

The widespread use of bicarbonate dialysate and reprocessed high-efficiency and "high-flux" dialyzers has raised concerns about the increased risk of reverse-transfer of dialysate contaminants into the blood compartment. To evaluate this concern, the reverse-transfer of bacterial products from contaminated bicarbonate dialysate into the blood compartment was compared during in vitro dialysis with new or reprocessed high-flux polysulfone dialyzers. In vitro dialysis was carried out at 37 degrees C by use of a counter-current recirculating loop dialysis circuit with either new high-flux polysulfone dialyzers or dialyzers reprocessed once or 20 times with formaldehyde (0.75%) and bleach (< 1%) with an automated system. Heparinized whole blood from healthy volunteers was circulated through the blood compartment, and bicarbonate dialysate was circulated in the dialysate compartment. The dialysate was challenged sequentially by 1:1000 and 1:100 dilutions of a sterile Pseudomonas aeruginosa culture supernatant (bacterial challenge). Samples were drawn from the blood and dialysate compartments 1 h after each challenge. Peripheral blood mononuclear cells (PBMC) were harvested by Ficoll-Hypaque separation from whole blood in the blood compartment and a 5 x 10(6) PBMC/mL cell suspension was prepared. Likewise, dialysate samples (0.5 mL) were added to 0.5 mL suspension of 5 x 10(6) PBMC/mL drawn at baseline. All PBMC suspensions were incubated upright in a humidified atmosphere at 37 degrees C with 5% CO2 for 24 h, and total interleukin-1 alpha (IL-1 alpha) and tumor necrosis factor-alpha (TNF alpha) cytokine production (cell-associated and secreted) was measured by radioimmunoassay. Eight experiments were performed for each arm of the study with the same donor for each arm. One hour after contaminating the dialysate with a 1:1000 dilution of the bacterial challenge, IL-1 alpha production by PBMC harvested from the blood compartment was 160 +/- 0, 171 +/- 11, and 270 +/- 35 pg, respectively, for new dialyzers, dialyzers reprocessed once, and dialyzers reprocessed 20 times (P = 0.004). One hour after challenging the dialysate with 1:100 dilution, IL-1 alpha production by PBMC harvested from the blood compartment was 188 +/- 20, 228 +/- 35, and 427 +/- 67 pg, respectively, for new dialyzers, dialyzers reprocessed once, and dialyzers reprocessed 20 times (P = 0.006). IL-1 alpha production by PBMC from dialyzers reprocessed 20 times was significantly greater than both new and dialyzers reprocessed once. However, there were no significant differences between new dialyzers and dialyzers reprocessed once. Similarly, after the 1:1000 challenge, TNF alpha production by PBMC harvested from the blood compartment was 160 +/- 0, 160 +/- 0, and 213 +/- 22 pg, respectively, for new dialyzers, dialyzers reprocessed once, and dialyzers reprocessed 20 times (P = 0.008). After the 1:100 challenge, TNF alpha production was 168 +/- 8, 188 +/- 20, and 225 +/- 32 pg, respectively, for new dialyzers, dialyzers reprocessed once, and dialyzers reprocessed 20 times (P = 0.20). These results demonstrate that reprocessing of high-flux polysulfone dialyzers with bleach increases the risk of reverse-transfer of bacterial products from contaminated dialysate, and this risk appears to increase with the number of reuses. Consequently, units that reprocess membranes with bleach and have suboptimal water quality might subject their patients to a higher risk of cytokine-related morbidity.     

Comparison of llama VH sequences from conventional and heavy chain antibodies

Forty different PCR clones encoding a llama variable heavy chain domain were analysed. The majority of these clones are derived from heavy-chain antibody cDNA in which the entire CH1 exon is absent. It appears from the amino acid within the VHH framework 1 and 3 that all the llama clones belong to the VH III family. However, the individual llama VHH sequences differ more substantially from each other than expected for members of the same family. Several remarkable amino acid substitutions in the framework 2 hinder the proper association of the VL. However, they lay the foundation for the secretion from the endoplasmic reticulum and good solubility behaviour of llama H2 antibodies. The repertoire of the llama VHHs may be extensive due to the presence of a long CDR3-loop, often constrained by a disulfide bridge and the occurrence of H1 and H2 loop conformations not yet encountered in mice or human VHs. The variability plot of the amino acids in the VHH shows that the first hypervariable region coincides with the structural H1 loop in contrast to the situation found in mice and man where the CDR1 and H1 are slightly offset. We propose that the amino acids of the llama H1 loop participate actively in the antigen binding. All these observations are characteristic for the llama VHHs of the homodimeric heavy-chain H2 antibodies, but are not maintained in the llama clones from conventional heterotetrameric H2L2 immunoglobulins.     

Physiological phosphorylation of protein kinase A at Thr-197 is by a protein kinase A kinase

Phosphorylation of the catalytic subunit of cyclic AMP-dependent protein kinase, or protein kinase A, on Thr-197 is required for optimal enzyme activity, and enzyme isolated from either animal sources or bacterial expression strains is found phosphorylated at this site. Autophosphorylation of Thr-197 occurs in Escherichia coli and in vitro but is an inefficient intermolecular reaction catalyzed primarily by active, previously phosphorylated molecules. In contrast, the Thr-197 phosphorylation of newly synthesized protein kinase A in intact S49 mouse lymphoma cells is both efficient and insensitive to activators or inhibitors of intracellular protein kinase A. Using [35S]methionine-labeled, nonphosphorylated, recombinant catalytic subunit as the substrate in a gel mobility shift assay, we have identified an activity in extracts of protein kinase A-deficient S49 cells that phosphorylates catalytic subunit on Thr-197. The protein kinase A kinase activity partially purified by anion-exchange and hydroxylapatite chromatography is an efficient catalyst of protein kinase A phosphorylation in terms of both a low Km for ATP and a rapid time course. Phosphorylation of wild-type catalytic subunit by the kinase kinase activates the subunit for binding to a pseudosubstrate peptide inhibitor of protein kinase A. By both the gel shift assay and a [gamma-32P]ATP incorporation assay, the enzyme is active on wild-type catalytic subunit and on an inactive mutant with Met substituted for Lys-72 but inactive on a mutant with Ala substituted for Thr-197. Combined with the results from mutant subunits, phosphoamino acid analysis suggests that the enzyme is specific for phosphorylation of Thr-197.     

卟啉类L—B膜分子间相互作用的光谱研究

以无取代的meso-四-(4-N)-吡啶基卟啉及其过渡族金属(主要Cu~(2+)、Zn~(2+))络合物制备L-B膜,以近紫外-可见吸收光谱和荧光光谱为手段,研究叶啉类分子在氯仿溶液中,L-B膜状态下以及固态状态下的相互作用。探讨分子聚集体的存在对光谱性质的影响。 为了研究叶咻类分子间的相互作用及其对光谱性质的影响,我们首先分析了叶啉在CHCl_3溶液中及固态状态下的近紫外-可见吸收光谱和荧光光谱。并将其与叶啉类分子的L-B膜作比较。结果表明,卟啉类的Soret吸收带带宽及峰位置在三种状态下均不相同,L-B膜的情况介于溶液中的和固体下的情况之间,说明了在L-B膜中,卟啉分子存在着某种形式的聚集体,且在这种聚集体中分子间的相互作用程度比固体弱,可以认为L-B膜上的分子呈准晶体状态。