Lipase and esterase-catalyzed acylation of hetero-substituted nitrogen nucleophiles in water and organic solvents

The lipase- and esterase-catalyzed acylations of hydroxylamine and hydrazine derivatives with octanoic acid and ethyl octanoate are described. The influence of solvent and nucleophile on the initial reaction rate was investigated for a number of free and immobilized enzymes. Initial rates were highest in water, but the overall productivity was optimal in dioxane. Octanoic acid (250 g/L) was converted for 93% into the hydroxamic acid in 36 h with only 1% (w/w) Candida antarctica lipase B (Novozym 435) in dioxane at 40 degrees C. This translates to a catalyst productivity of 68.5 g. g(-1). day(-1) and a space time yield of 149 g. L(-1). day(-1), unprecedented figures in the direct reaction of an acid with a nitrogen nucleophile in an organic solvent.     

Continuous culture of Methanococcus jannaschii, an extremely thermophilic methanogen

Methanococcus jannaschii, an extremely thermophilic methanogen isolated from a deep-sea hydrothermal vent, was grown at 80 degrees C in continuous culture on a mineral salts medium gassed with H(2) and CO(2) at three different flow rates. The maximum specific growth rate was 0.56 h(-1), and the maximum specific methane productivity was 0.32 (mol g(-1) h(-1)). Uncoupling of growth and methane production was evidenced by an increase in teh non-growth-associated rate of methane formation, beta, with increasing gaseous input. The specific hydrogenase activity exhibited growth-assiciated behaviour at low growth rates, but showed no dependence on growth at higher growth rates. The growth dependence of hydrogenase activity is consistent with the pressure dependence of hydrogenase activity measured in previous experiments. In contrast, the specific protease activity was independent of the growth rate over the entire range of dilution rates studied. (c) 1994 John Wiley & Sons, Inc.     

Acute and delayed vasoconstriction after subarachnoid hemorrhage: local cerebral blood flow, histopathology, and morphology in the rat basilar artery.

The decreased local cerebral blood flow (LCBF) and cerebral ischemia that occur after subarachnoid hemorrhage (SAH) may be caused by acute and/or delayed vasospasm. In 36 Sprague-Dawley (350-450 g) rats SAH was induced by transclival puncture of the basilar artery. Mean arterial blood pressure (MABP), LCBF, intracranial pressure (ICP), and cerebral perfusion pressure (CPP) were measured in all rats for 30 min before and 60 min after SAH was induced. One set of control (n : 7) and experimental animals (n : 7) was sacrificed after the 60 min of initial post-hemorrhage measurements were recorded. Four days after SAH induction, LCBF and MABP were measured again for 60 min in subgroups of surviving experimental rats (n : 7) and control rats (n : 7). Histopathologic and morphologic examinations of the basilar artery were performed in each subgroup. There was a sharp drop in LCBF just after SAH was induced (55.50 +/- 11.46 mlLD/min/100 g and 16.1 +/- 3.6 mlLD/min/100 g for baseline and post-SAH, respectively; p < 0.001). The flow then gradually increased but had not returned to pre-SAH values by 60 min (p < 0.05). At 4 days after SAH induction, although LCBF was lower than that observed in the control group and pre-SAH values, it was not significantly different from either of these flow rates (p > 0.05). ICP (baseline 7.05 +/- 0.4 mmHg) increased acutely to 75.2 +/- 7.1 mmHg, but returned to normal levels by 60 min after SAH. CPP (baseline 84.5 +/- 6.3 mmHg) dropped accordingly (to 18.6 +/- 3.1 mmHg), and then increased, reaching 72.2 +/- 4.9 mmHg at 60 min after SAH (p > 0.05). Examinations of the arteries revealed decreased inner luminal diameter and distortion of the elastica layer in the early stage. LCBF in nonsurviver rats (n : 8) was lower than that in the animals that survived (p < 0.01). At 4 days post-hemorrhage, the rats' basilar arteries showed marked vasculopathy. The findings showed that acute SAH alters LCBF, ICP, and CPP, and that decreased LCBF affects mortality rate. Subsequent vasculopathy occurs in delayed fashion, and this was observed at 4 days after the hemorrhage event.     

High-light stress and photoprotection in Umbilicaria antarctica monitored by chlorophyll fluorescence imaging and changes in zeaxanthin and glutathione

The effect of high light on spatial distribution of chlorophyll (Chl) fluorescence parameters over a lichen thallus (Umbilicaria antarctica) was investigated by imaging of Chl fluorescence parameters before and after exposure to high light (1500 micro mol m (-2) s (-1), 30 min at 5 degrees C). False colour images of F (V)/F (M) and Phi (II) distribution, taken over thallus with 0.1 mm (2) resolution, showed that maximum F (V)/F (M) and Phi (II) values were located close to the thallus centre. Minimum values were typical for thallus margins. After exposure to high light, a differential response of F (V)/F (M) and Phi (II) was found. The marginal thallus part exhibited a loss of photosynthetic activity, manifested as a lack of Chl fluorescence signal, and close-to-centre parts showed a different extent of F (V)/F (M) and Phi (II) decrease. Subsequent recovery in the dark led to a gradual return of F (V)/F (M) and Phi (II) to their initial values. Fast (30 min) and slow (1 - 22 h) phase of recovery were distinguished, suggesting a sufficient capacity of photoprotective mechanisms in U. antarctica to cope with low-temperature photoinhibition. Glutathione and xanthophyll cycle pigments were analyzed by HPLC. High light led to an increase in oxidized glutathione (GSSG), and a conversion of violaxanthin to zeaxanthin, expressed as their de-epoxidation state (DEPS). The responses of GSSG and DEPS were reversible during subsequent recovery in the dark. GSSG and DEPS were highly correlated to non-photochemical quenching (NPQ), indicating involvement of these antioxidants in the resistance of U. antarctica to high-light stress. Heterogeneity of Chl fluorescence parameters over the thallus and differential response to high light are discussed in relation to thallus anatomy and intrathalline distribution of the symbiotic alga Trebouxia sp.     

In vitro steady-flow analysis of systemic-to-pulmonary shunt haemodynamics.

A modified Blalock-Taussig shunt is a connection created between the systemic and pulmonary arterial circulations to improve pulmonary perfusion in children with congenital heart diseases. Survival of these patients is critically dependent on blood flow distribution between the pulmonary and systemic circulations which in turn depends upon the flow resistance of the shunt. Previously, we investigated the pressure-flow relationship in rigid shunts with a computational approach. to estimate the pulmonary blood flow rate on the basis of the in vivo measured pressure drop. The present study aims at evaluating, in vitro how the anastomotic distensibility and restrictions due to suture presence affect the shunt pressure-flow relationship. Two actual Gore-Tex shunts (3 and 4 mm diameters) were sutured to compliant conduits by a surgeon and tested at different steady flow rates (0.25-11 min(-1)) and pulmonary pressures (3-34 mmHg). Corresponding computational models were also created to investigate the role of the anastomotic restrictions due to sutures. In vitro experiments showed that pulmonary artery pressure affects the pressure-flow relationship of the anastomoses. particularly at the distal site. However, this occurrence scarcely influences the total shunt pressure drop. Comparisons between in vitro and computational models without anastomotic restrictions show that the latter underestimates the in vitro pressure drops at any flow rate. The addition of the anastomotic restrictions (31 and 47% of the original area of 3 and 4 mm shunts, respectively) to the computational models reduces the gap, especially at high shunt flow rate and high pulmonary pressure.     

Effect of prosthetic mitral valve geometry and orientation on flow dynamics in a model human left ventricle

Pulsatile flow dynamics through bileaflet (St Jude and Duromedics), tilting disc (Bjork-Shiley and Omniscience), caged ball (Starr-Edwards), pericardial (Edwards) and porcine (Carpentier-Edwards) mitral valves in a model human left ventricle (LV) were studied. The model human ventricle, obtained from an in situ diastolic casting, was incorporated into a mock circulatory system. Measurements were made at various heart rates and flow rates. These included the transvalvular pressure drop and regurgitation in percent and cm3 beat-1. The effect of valve geometry and the orientation of the valve with respect to the valve annulus was analyzed using a flow visualization technique. Qualitative flow visualization study indicates certain preferred orientations for the tilting disc and bileaflet valve prostheses in order to obtain a smooth washout of flow in the LV chamber.     

Pressure drop increase by biofilm accumulation in spiral wound RO and NF membrane systems: role of substrate concentration,flow velocity,substrate load and flow direction

In an earlier study, it was shown that biofouling predominantly is a feed spacer channel problem. In this article, pressure drop development and biofilm accumulation in membrane fouling simulators have been studied without permeate production as a function of the process parameters substrate concentration, linear flow velocity, substrate load and flow direction. At the applied substrate concentration range, 100–400 μg l^?1 as acetate carbon, a higher concentration caused a faster and greater pressure drop increase and a greater accumulation of biomass. Within the range of linear flow velocities as applied in practice, a higher linear flow velocity resulted in a higher initial pressure drop in addition to a more rapid and greater pressure drop increase and biomass accumulation. Reduction of the linear flow velocity resulted in an instantaneous reduction of the pressure drop caused by the accumulated biomass, without changing the biofilm concentration. A higher substrate load (product of substrate concentration and flow velocity) was related to biomass accumulation. The effect of the same amount of accumulated biomass on the pressure drop increase was related to the linear flow velocity. A decrease of substrate load caused a gradual decline in time of both biomass concentration and pressure drop increase. It was concluded that the pressure drop increase over spiral wound reverse osmosis (RO) and nanofiltration (NF) membrane systems can be reduced by lowering both substrate load and linear flow velocity. There is a need for RO and NF systems with a low pressure drop increase irrespective of the biomass formation. Current efforts to control biofouling of spiral wound membranes focus in addition to pretreatment on membrane improvement. According to these authors, adaptation of the hydrodynamics, spacers and pressure vessel configuration offer promising alternatives. Additional approaches may be replacing heavily biofouled elements and flow direction reversal.     

An immobilized cell reactor with simultaneous product separation. II. Experimental reactor performance

The simultaneous separation of volatile fermentation products from product-inhibited fermentations can greatly increase the productivity of a bioreactor by reducing the product concentration in the bioreactor, as well as concentrating the product in an output stream free of cells, substrate, or other feed impurities. The Immobilized Cell Reactor-Separator (ICRS) consists of two column reactors: a cocurrent gas-liquid "enricher" followed by a countercurrent "stripper" The columns are four-phase tubular reactors consisting of (1) an inert gas phase, (2) the liquid fermentation broth, (3) the solid column internal packing, and (4) the immobilized biological catalyst or cells. The application of the ICRS to the ethanol-from-whey-lactose fermentation system has been investigated. Operation in the liquid continuous or bubble flow regime allows a high liquid holdup in the reactor and consequent long and controllable liquid residence time but results in a high gas phase pressure drop over the length of the reactor and low gas flow rates. Operation in the gas continuous regime gives high gas flow rates and low pressure drop but also results in short liquid residence time and incomplete column wetting at low liquid loading rates using conventional gas-liquid column packings. Using cells absorbed to conventional ceramic column packing (0.25-in. Intalox saddles), it was found that a good reaction could be obtained in the liquid continuous mode, but little separation, while in the gas continuous mode there was little reaction but good separation. Using cells sorbed to an absorbant matrix allowed operation in the gas continuous regime with a liquid holdup of up to 30% of the total reactor volume. Good reaction rates and product separation were obtained using this matrix. High reaction rates were obtained due to high density cell loading in the reactor. A dry cell density of up to 92 g/L reactor was obtained in the enricher. The enricher ethanol productivity ranged from 50 to 160 g/L h while the stripper productivity varied from 0 to 32 g/L h at different feed rates and concentrations. A separation efficiency of as high as 98% was obtained from the system.     

Thin-film coupled fluid-solid analysis of flow through the Ahmed glaucoma drainage device

The Ahmed glaucoma valve (AGV) is a popular glaucoma drainage device, allowing maintenance of normal intraocular pressure in patients with reduced trabecular outflow facility. The uniquely attractive feature of the AGV, in contrast to other available drainage devices, is its variable resistance in response to changes in flow rate. As a result of this variable resistance, the AGV maintains a pressure drop between 7 and 12 mm Hg for a wide range of aqueous humor flow rates. In this paper, we demonstrate that the nonlinear behavior of the AGV is a direct result of the flexibility of the valve material. Due to the thin geometry of the system, the leaflets of the AGV were modeled using the von Kármán plate theory coupled to a Reynolds lubrication theory model of the aqueous humor flow through the valve. The resulting two-dimensional coupled steady-state partial differential equation system was solved by the finite element method. The Poisson's ratio of the valve was set to 0.45, and the modulus was regressed to experimental data, giving a best-fit value 4.2 MPa. Simulation results compared favorably with previous experimental studies and our own pressure-drop/flow-rate data. For an in vitro flow of 1.6 microL/min, we calculated a pressure drop of 5.8 mm Hg and measured a pressure drop of 5.2 +/- 0.4 mm Hg. As flow rate was increased, pressure drop rose in a strongly sublinear fashion, with a flow rate of 20 microL/min giving a predicted pressure drop of only 10.9 mm Hg and a measured pressure drop of 10.5 +/- 1.1 mm Hg. The AGV model was then applied to simulate in vivo conditions. For an aqueous humor flow rate of 1.5-3.0 microL/min, the calculated pressure drops were 5.3 and 6.3 mm Hg.     

Upper airway collapsibility and patterns of flow limitation at constant end-expiratory lung volume

The passive pharyngeal critical closing pressure (Pcrit) is measured using a series of pressure drops. However, pressure drops also lower end-expiratory lung volume (EELV), which independently affects Pcrit. We describe a technique to measure Pcrit at a constant EELV. Continuous positive airway pressure (CPAP)-treated obstructive sleep apnea (OSA) patients and controls were instrumented with an epiglottic catheter, magnetometers (to measure change in EELV), and nasal mask/pneumotachograph and slept supine on nasal CPAP. Pcrit was measured in standard fashion and using our novel "biphasic technique" in which expiratory pressure only was lowered for 1 min before the inspiratory pressure was dropped; this allowed EELV to decrease to the drop level before performing the pressure drop. Seven OSA and three controls were studied. The biphasic technique successfully lowered EELV before the inspiratory pressure drop. Pcrit was similar between the standard and biphasic techniques (-0.4 ± 2.6 vs. -0.6 ± 2.3 cmH(2)O, respectively, P = 0.84). Interestingly, we noted three different patterns of flow limitation: 1) classic Starling resistor type: flow fixed and independent of downstream pressure; 2) negative effort dependence within breaths: substantial decrease in flow, sometimes with complete collapse, as downstream pressure decreased; and 3) and negative effort dependence across breaths: progressive reductions in peak flow as respiratory effort on successive breaths increased. Overall, EELV changes do not influence standard passive Pcrit measurements if breaths 3-5 of pressure drops are used. These results also highlight the importance of inspiratory collapse in OSA pathogenesis. The cause of negative effort dependence within and across breaths is not known and requires further study.     

Estimation of inspiratory pressure drop in neonatal and pediatric endotracheal tubes.

Endotracheal tubes (ETTs) constitute a resistive extra load for intubated patients. The ETT pressure drop (DeltaP(ETT)) is usually described by empirical equations that are specific to one ETT only. Our laboratory previously showed that, in adult ETTs, DeltaP(ETT) is given by the Blasius formula (F. Lofaso, B. Louis, L. Brochard, A. Harf, and D. Isabey. Am. Rev. Respir. Dis. 146: 974-979, 1992). Here, we also propose a general formulation for neonatal and pediatric ETTs on the basis of adimensional analysis of the pressure-flow relationship. Pressure and flow were directly measured in seven ETTs (internal diameter: 2.5-7.0 mm). The measured pressure drop was compared with the predicted drop given by general laws for a curved tube. In neonatal ETTs (2.5-3.5 mm) the flow regime is laminar. The DeltaP(ETT) can be estimated by the Ito formula, which replaces Poiseuille's law for curved tubes. For pediatric ETTs (4.0-7.0 mm), DeltaP(ETT) depends on the following flow regime: for laminar flow, it must be calculated by the Ito formula, and for turbulent flow, by the Blasius formula. Both formulas allow for ETT geometry and gas properties.     

Numerical modeling of simulated blood flow in idealized composite arterial coronary grafts: steady state simulations

This paper presents a comparative study of simulated blood flow in different configurations of simplified composite arterial coronary grafts (CACGs). Even though the composite arterial grafting is increasingly used in cardiac surgery, it is still questionable whether or not the blood flow in such grafts can adequately meet the demands of the native myocardial circulation. A computational fluid dynamics (CFD) model was developed to conduct computer-based studies of simulated blood flow in four different geometric configurations of CACGs, corresponding to routinely used networks in cardiac surgery coronary grafts (T, Y, Pi and sequential). The flow was assumed three-dimensional, laminar and steady and the fluid as Newtonian, while the vessel walls were considered as inelastic and impermeable. It was concluded that local haemodynamics, practically described by velocity, pressure drop, wall shear stress (WSS) and flow rates, may be strongly influenced by the local geometry, especially at the anastomotic sites. The computations were made at mean flow rates of 37.5, 75 and 150ml/min. The side-branch outflow rates, computed for each bypass graft, showed noticeable differences. The results, which were found both qualitatively and quantitatively consistent with other studies, indicate that the Pi-graft exhibits significantly less uniform distribution of outflow rates than the other geometric configurations. Moreover, prominent variations in WSS and velocity distribution among the assessed CACGs were predicted, showing remarkable flow interactions among the arterial branches. The lowest shear stress regions were found on the lateral walls of bifurcations, which are predominantly susceptible to the occurrence of coronary artery disease (CAD). In contrast, the highest WSS were observed at the turn of the arterial branches.     

Pressure drop,gas hold-up,and oxygen transfer in tower systems

Pressure drop, gas hold-up, and oxygen transfer were investigated in a sieve tray column, a column with Koch motionless mixers, and a bubble column. The oxygen transfer experiments were conducted using cocurrent flow of gas and liquid under steady-state conditions with oxygen transfer from the gas to the liquid phase. The oxygen transfer rates and efficiencies of the sieve tray column and the column with Koch mixers were found to be superior to those of the conventional bubble column. Gas hold-up was also greater when sieve trays or Koch mixers were inserted in the tower. The pressure drop was found to be primarily due to the liquid head in all three columns.     

Physiological flow simulation in residual human stenoses after coronary angioplasty

To evaluate the local hemodynamic implications of coronary artery balloon angioplasty, computational fluid dynamics (CFD) was applied in a group of patients previously reported by [Wilson et al. (1988), 77, pp. 873-885] with representative stenosis geometry post-angioplasty and with measured values of coronary flow reserve returning to a normal range (3.6 +/- 0.3). During undisturbed flow in the absence of diagnostic catheter sensors within the lesions, the computed mean pressure drop delta p was only about 1 mmHg at basal flow, and increased moderately to about 8 mmHg for hyperemic flow. Corresponding elevated levels of mean wall shear stress in the midthroat region of the residual stenoses, which are common after angioplasty procedures, increased from about 60 to 290 dynes/cm2 during hyperemia. The computations (Ree approximately equal to 100-400; alpha e = 2.25) indicated that the pulsatile flow field was principally quasi-steady during the cardiac cycle, but there was phase lag in the pressure drop-mean velocity (delta p - u) relation. Time-averaged pressure drop values, delta p, were about 20 percent higher than calculated pressure drop values, delta ps, for steady flow, similar to previous in vitro measurements by Cho et al. (1983). In the throat region, viscous effects were confined to the near-wall region, and entrance effects were evident during the cardiac cycle. Proximal to the lesion, velocity profiles deviated from parabolic shape at lower velocities during the cardiac cycle. The flow field was very complex in the oscillatory separated flow reattachment region in the distal vessel where pressure recovery occurred. These results may also serve as a useful reference against catheter-measured pressure drops and velocity ratios (hemodynamic endpoints) and arteriographic (anatomic) endpoints post-angioplasty. Some comparisons to previous studies of flow through stenoses models are also shown for perspective purposes.     

Influence of water activity on the enantioselective esterification of (R,S)-ibuprofen by crosslinked crystals of Candida antarctica lipase B in organic solvent media.

The lipase B from Candida antarctica was purified from a commercial source and crystallized. The microcrystals were crosslinked using glutaraldehyde. The crosslinked crystals were then used to catalyze the esterification of (R,S)-ibuprofen with dodecanol in octane at various water activities. As for the commercial preparation immobilized on acrylic resin, Novozym 435, low water activities foster better enantioselectivity, and the maximum reaction rates are obtained for a(W) = 0.1.     

Toluene removal from waste air using a flat composite membrane bioreactor

In this report, gaseous toluene biodegradation results in a flat composite membrane reactor inoculated with Pseudomonas putida TVA8 are presented. Preliminary abiotic experiments showed that transport of toluene through the membrane was linearly and negatively correlated with the gas residence time (tau). During a 339-day biofiltration experiment, the influence of gas residence time (2-24 sec) and mass loading rate (B(v); 10-483 g x m(-3) h(-1)) on the toluene elimination capacity was investigated. A maximum elimination capacity (EC(max)) of 397 g x m(-3) h(-1) was achieved at tau = 24 sec and B(v) = 473 g x m(-3) h(-1). Expressed per unit membrane area, the EC(m,max) was 0.793 g x m(-2) h(-1), which is five times higher than results obtained with other membrane bioreactor experiments in the same range of loading rates. At low gas residence times, reactor performance was limited by mass transfer. Toluene concentration profiles along the membrane were measured for several biotic and abiotic conditions. For inlet concentrations (C(in)) up to 1 g x m(-3), more than 90% was eliminated at 15 cm from the reactor inlet. For C(in) > 1.65 g x m(-3), longer membranes are necessary to obtain these high removal efficiencies.     

Effects of long-term losartan and L-arginine treatment on haemodynamics, glomerular filtration, and SOD activity in spontaneously hypertensive rats

Recently, it has been reported that losartan, an angiotensin II receptor (ATR) antagonist, depresses the angiotensin II-induced production of superoxide radicals. Also, in spontaneously hypertensive rats (SHR) endothelial dysfunction is associated with decreased nitric oxide (NO) synthesis. In this study, we examined the effects of long-term ATR blockade and L-arginine supplementation on the haemodynamic parameters, glomerular filtration, and oxidative status in SHR. Adult male SHR were treated with losartan (10 mg/kg) and with the NO donor L-arginine (2 g/kg) for 4 weeks. The animals were divided into the following experimental groups: control (n = 7), L-arginine (n = 7), losartan (n = 7), and L-arginine + losartan (n = 7). Mean arterial pressure (MAP), regional blood flow, urea clearance, and activity of superoxide dismutase (SOD) were measured at the end of treatment. MAP was significantly reduced in the losartan group compared with the control group (133.3 +/- 7.3 vs. 161.5 +/- 14.5 mm Hg). Aortic blood flow was significantly higher and aortic vascular resistance was significantly lower in all treated groups than in the control. Urea clearance rose significantly in the L-arginine + losartan group compared with control (393.27 +/- 37.58 vs. 218.68 +/- 42.03 microL x min(-1) x 100 g(-1)) as did the activity of SOD (1668.97 +/- 244.57 vs. 1083.18 +/- 169.96 U/g Hb). Our results suggest that the antihypertensive effect of losartan and L-arginine in SHR is not primarily mediated by increased SOD activity. Also, combined treatment with ATR blockade and L-arginine supplementation has a beneficial effect on renal function that is, at least in part, mediated by increased SOD activity in SHR.     

Response of river metabolism to restoration of flow in the Kissimmee River, Florida, U.S.A.

1. The single station diel oxygen curve method was used to determine the response of system metabolism to backfilling of a flood control canal and restoration of flow through the historic river channel of the Kissimmee River, a sub‐tropical, low gradient, blackwater river in central Florida, U.S.A. Gross primary productivity (GPP), community respiration (CR), the ratio of GPP/CR (P/R) and net daily metabolism (NDM) were estimated before and after canal backfilling and restoration of continuous flow through the river channel. 2. Restoration of flow through the river channel significantly increased reaeration rates and mean dissolved oxygen (DO) concentrations from <2 mg L⁻¹ before restoration of flow to 4.70 mg L⁻¹ after flow was restored. 3. Annual GPP and CR rates were 0.43 g O₂ m⁻² day⁻¹ and 1.61 g O₂ m⁻² day⁻¹ respectively, before restoration of flow. After restoration of flow, annual GPP and CR rates increased to 3.95 O₂ m⁻² day⁻¹ and 9.44 g O₂ m⁻² day⁻¹ respectively. 4. The ratio of P/R (mean of monthly values) increased from 0.29 during the prerestoration period to 0.51 after flow was restored, indicating an increase in autotrophic processes in the restored river channel. NDM values became more negative after flow was restored. 5. After flow was restored, metabolism parameters were generally similar to those reported for other blackwater river systems in the southeast U.S.A. Postrestoration DO concentrations met target values derived from free flowing, minimally impacted reference streams.     

Synthesis, protonation behavior, conformational analysis, and regioselective enzymatic acylation of the novel diamino analogue of (E)-5-(2-bromovinyl)-2'-deoxyuridine (BVDU)

(E)-3',5'-Diamino-5-(2-bromovinyl)-2',3',5'-trideoxyuridine (5), the diamino analogue of BVDU (1), was synthesized from BVDU. The protonation behavior of 5 has been studied by means of pH-metric measurements and NMR spectroscopy. This study allows the determination of the basicity constants and the stepwise protonation sites. Thus, the main species at physiological pH is the monoprotonated form. The conformational analysis of this nucleoside analogue was also carried out through 1H NMR spectroscopy. In addition, a convenient synthesis of N-3' and N-5' acylated derivatives was developed by regioselective enzymatic acylation. Thus, Candida antarctica lipase B (CAL-B) selectively acylated the 5'-amino group, thus furnishing nucleosides 8. On the other hand, immobilized Pseudomonas cepacia lipase (PSL-C) exhibited the opposite selectivity, conferring acylation at the 3'-amino group, thus affording derivatives 9.     

Microfiltration cell-recycle pilot system for continuous thermoanaerobic production of exo-beta-amylase

A microfiltration cell-recycle pilot-scale system was developed comprised of a conventional continuous-flow fermentor connected to an in situ steam-sterilizable cross-flow ceramic filter with a backflushing device. A microcomputer was used to control filtration pressure, tangential flow velocity, and backflushing. Performance of the system was tested with the anaerobic production of thermostable extracellular beta-amylase at 60 degrees C by Clostridium thermosulfurogenes on maltose or malto-dextrin media. Filtration rates during continuous cultivation were between 20 and 60 L/m(2)/h. The maltodextrin and cell debris occurring at high retentate flow rates or filtration pressures impaired the performance of the filter. Backflushing initially improved the permeate flux to 42% in a maltose medium and to 10% in a maltodextrin medium, but the effect diminished with time. The productivity of beta-amylase (as much as 48 U/mL/h) and concentration of biomass (as much as 14 g/L) were increased 11- and 12-fold, respectively, if compared to values obtained in a chemostat. The concentration of beta-amylase rose to 220 U/mL in the reactor, which was 5.5-fold more than under comparable conditions in a chemostat.