Batch phases

Tests were carried out at 25°C and at initial pH 6.9. Cultures in the liquid medium were incubated in 50 mL Falcon tubes, continuously shaked at 220 rpm. Each culture contained a fresh Pseudomonas sp. 0X1 colony in 10 mL of medium. The airlift with 10 g of pumice was sterilized at 121°C for 30 min and then housed in a sterile room. One-day culture was transferred to the reactor and, after a batch phase, liquid medium with phenol as the only carbon source was continuously fed. The reactor volume V was fixed at 0.13 L. Aerobic conditions were established sparging technical air. Under these conditions microorganism started to grow immobilized on the solid s support. When immobilized biomass approached steady state, cyclic operation of the airlift was started by alternating aerobic/anaerobic conditions. 

A semicontinuous reactor is a reactor for a multiphase reaction in which one phase flows continuously through a vessel containing a batch of another phase. The operation is thus unsteady-state with respect to the batch phase, and may be steady-state or unsteady-state with respect to the flowing phase, as in a fixed-bed catalytic reactor (Chapter 21) or a fixed-bed gas-solid reactor (Chapter 22), respectively. 

In many enzyme fermentations, the limiting component, usually the C-source, has to be added semi-continuously to keep its concentration at a predetermined, usually low, value. This measure makes it possible either to influence selectivity between different pathways or to uncouple predominantly cell growth during the first phase of the fermentation from predominantly product (i.e., enzyme) formation in the later stages of the fermentation cycle. Often, protein formation is induced by adding an inducer (see Chapter 4). During the fed-batch phase, the broth volume increases. Either the broth is harvested when the maximum volume is reached, or broth is withdrawn from time to time. The product is present in high concentrations. 

The pH was maintained at 5.0 by the addition of 2 M NaOH (Micro DCU-300 B. Braun Biotech). The fermentation temperature was 30°C and the stirrer speed was 500 rpm (MCU-200 B. Braun Biotech). Cell mass was produced in an initial batch phase using a glucose concentration of 64 g/L. The concentrations of mineral salts, trace metals, vitamins, and Ergosterol/ Tween-80 were the same as in the fed-batch experiments. When the glucose in the batch medium was completely consumed, 1.9 L of dilute-acid hydrolysate was pumped into the reactor at maximum pump speed, by using a peristaltic pump (Ul-M Alitea AB). The ratio between batch volume and the final volume (i.e., after all the hydrolysate had been added) was similar to the ratio in the PDU fed-batch experiments, approx 1 4. The reactor medium was sparged with nitrogen (600 mL/min). The C02 content in the exhaust gas was measured with a gas analyzer (TanDem Adaptive Biosystems, Luton, UK). 

After a 500-mL batch phase with an initial glucose concentration of 64 g/L, 1.9 L of dilute-acid hydrolysate was added using the maximum pump speed. Batch experiments with both TMB 3000 and CBS 8066 were concluded (Figs. 2 and 3). In addition, TMB 3000 was tested with two different batches of dilute-acid hydrolysate (Table 1). 

The productivity of the strain TMB 3000 was clearly superior to that of CBS 8066, and therefore strain TMB 3000 was analyzed further. Two fed-batch fermentations with dilute-acid hydrolysate A (Table 1) were run using two different glucose concentrations in the batch phase. 

Fig. 5. Concentrations of glycerol (— - -), glucose (— —), and HMF (— —) during two fed-batch fermentations with TMB 3000. Dilute-acid hydrolysate A was used. (A) 64 g/L of glucose in the batch phase (B) 45 g/L of glucose in the batch phase.

Yield coefficients Explanation Batch phase (C-mol/C-mol) (g/g) Fed-batch phase (C-mol/C-mol) (g/g) ... 

The initial glucose concentration in the batch phase was 64 g/L. Ysc could not be calculated during the batch phase because the gas bulk in the fermentor initially consisted of air and not C02, as when the fed-batch was started. 

The visualization method also worked with a 500-L perfusion reactor system for production of recombinant human coagulation factor VIII (hFVIII) in Chinese hamster ovary (CHO) cells [36,37]. Despite the diluted concentration of CHO cells and low titer of hFVIII in the medium, the nose could differentiate between the batch phase, medium replacement phase, and the high and low productivity phases during the five-week long cultivations (Fig. 10). The low concentration of hFVIII makes it credible to believe that there are other components associated with the product formation that the electronic nose responds to. 

Fig. 5 Effects of reducing the pH fix)m 6 to 4.5 at the beginning of the fed-batch phase. Solid line rMnP activity (U/1), dashed line cell density (dry weight g/1). Symbols are as follows triangle pH 6.0 for batch/pH 4.5 for fed batch, square pH 4.5 during the whole cultivation...

In bioreactors maintained at pH 4.5 throughout the batch and fed-batch phases, the rMnP enzyme activity was very low (54 U/1) compared with activities greater than 2,000 U/1 in pH 6.0 fermentations at the end of the fed-batch phase (Fig. 4). Decreasing the culture pH from 6.0 to 4.5 at the start of the fed-batch phase also resulted in little rMnP accumulation in the medium (Fig. 5). Decreasing the culture pH from 6.0 to 4.5 after 20 h in the fed-batch phase resulted in a rapid loss of more than 1,000 U/1 rMnP activity from the rMnP produced at pH 6 (Fig. 6). The failure of P. pastoris aMnPl-1 to produce appreciable amounts of rMnP at low pH and the decline in rMnP activity upon lowering the cultivation pH are somewhat unexpected. P. pastoris grows equally well at pH 6 and 4.5, and wtMnP and rMnP are both stable and active at pH 4.5 [3]. Furthermore, the wtMnP host (P. chrysosporium) is known to produce active wtMnP at pH 4.5 [34]. Because of the rapid inactivation of rMnP in cultures at pH 4.5, a pH of 6.0 was chosen for the routine cultivation of P. pastoris aMnPl-1, and further experiments were undertaken to investigate the cause of low rMnP production at low pH. 

The initial concentrations, which are required for the calculation of the initial reaction rate and the Damkoehler number, are preferably referenced to the final volume. This, on the one hand, allows for a straightforward calculation of the post-feed reaction phase as the final SBR concentrations directly correspond to the input concentrations of the concluding batch phase. On the other hand, this allows for a simple comparison to other ideal reactors without additional recalculations. 

When setting up the mass balance the Damkoehler number was referenced to the final reaction volume, this way enabling an easier comparison with other ideal reactors and a convenient calculation of SBR phase and post reaction batch phase. This shall be adopted for the definition of the Stanton number. 

Separation of variables and integration subject to the initial condition that at the time the fed-batch phase of the operation begins, X = Xgg gives... 

Readers should note that for purposes of process control the flow rate of the feed stream can be determined at any time using equation (M) and the value of the total weight of biomass in the bioreactor at that time. When the fed batch phase of operation is initiated, the requisite feed rate is 14.4 mL/h, and at the end of this phase the necessary feed rate is 35.5 mL/h. 

The use of powders is generally in a batch phase. The solution to be treated is mixed with an appropriate quantity of the activated-carbon powder, agitated for less than I h. and then separated by settling or filtration. The used carbon may be discard or "eluted (regenerated) for further use. The use of particulate forms in liquid treating employs fixed beds through which the solution to be treated is passed. When the beds become saturated, they can be revivified by gas oxidation or solvent extraction. 

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