Scale-down and optimization studies of the gluconic acid fermentation by Gluconobacter oxydans

To simulate production-scale conditions of gluconic acid fermentation by Gluconobacter oxydans, different experimental setups are presented in this study. From the determination of the time constants of a production-scale reactor, it can be concluded that mixing and oxygen transfer are the rate-limiting mechanisms. This results in oxygen concentration gradients which were simulated in a one-compartment reactor in which the oxygen concentration was fluctuated by a fluctuated gassing with air and nitrogen. It could be concluded that only very long periods of absence of oxygen (ca. 180 s) results in lower specific oxygen uptake rates by Gluconobacter oxydans. From scale-down studies carried out in a two-compartment system to simulate a production-scale reactor more accurately, it could be concluded that not only the residence time in the aerated part of the system is important, but the liquid flow in between the different parts of the reactor is also an essential parameter. It could also be concluded that the microorganisms are not influenced negatively by the fluctuated oxygen concentrations with respect to their maximal oxidation capacity. The two-compartment system can also be used for optimization experiments in which the "aerated" compartment was gassed with pure oxygen. From these experiments it was concluded that also a short residence of the cells at high oxygen concentrations diminished the growth and product formation rates. These experiments show the necessity of the scale-down experiments if optimization is carried out. The two-compartment system presented in this study is a very attractive tool for reliable scale-down experiments.     

Genetic programming assisted stochastic optimization strategies for optimization of glucose to gluconic acid fermentation

This article presents two hybrid strategies for the modeling and optimization of the glucose to gluconic acid batch bioprocess. In the hybrid approaches, first a novel artificial intelligence formalism, namely, genetic programming (GP), is used to develop a process model solely from the historic process input-output data. In the next step, the input space of the GP-based model, representing process operating conditions, is optimized using two stochastic optimization (SO) formalisms, viz., genetic algorithms (GAs) and simultaneous perturbation stochastic approximation (SPSA). These SO formalisms possess certain unique advantages over the commonly used gradient-based optimization techniques. The principal advantage of the GP-GA and GP-SPSA hybrid techniques is that process modeling and optimization can be performed exclusively from the process input-output data without invoking the detailed knowledge of the process phenomenology. The GP-GA and GP-SPSA techniques have been employed for modeling and optimization of the glucose to gluconic acid bioprocess, and the optimized process operating conditions obtained thereby have been compared with those obtained using two other hybrid modeling-optimization paradigms integrating artificial neural networks (ANNs) and GA/SPSA formalisms. Finally, the overall optimized operating conditions given by the GP-GA method, when verified experimentally resulted in a significant improvement in the gluconic acid yield. The hybrid strategies presented here are generic in nature and can be employed for modeling and optimization of a wide variety of batch and continuous bioprocesses.     

Process optimization of continuous gluconic acid fermentation by isolated yeast-like strains of Aureobasidium pullulans

This study was focused on the optimization of a new fermentation process for continuous gluconic acid production by the isolated yeast-like strain Aureobasidium pullulans DSM 7085 (isolate 70). Operational fermentation parameters were optimized in chemostat cultures, using a defined glucose medium. Different optima were found for growth and gluconic acid production for each set of operation parameters. Highest productivity was recorded at pH values between 6.5 and 7.0 and temperatures between 29 and 31 degrees C. A gluconic acid concentration higher than 230 g/L was continuously produced at residence times of 12 h. A steady state extracellular gluconic acid concentration of 234 g/L was measured at pH 6.5. 122% air saturation yielded the highest volumetric productivity and product concentration. The biomass-specific productivity increased steadily upon raising air saturation. An intracellular gluconic acid concentration of about 159 g/L (0.83 mol) was determined at 31 degrees C. This is to be compared with an extracellular concentration of 223 g/L (1.16 mol), which indicates the possible existence of an active transport system for gluconic acid secretion, or the presence of extracellular glucose oxidizing enzymes. The new process provides significant advantages over the traditional discontinuous fungi operations. The process control becomes easier, thus offering stable product quality and quantity.     

Multicriteria optimization of gluconic acid production using net flow

The biochemical process industry is often confronted with the challenge of making decisions in an atmosphere of multiple and conflicting objectives. Recent innovations in the field of operations research and systems science have yielded rigorous multicriteria optimization techniques that can be successfully applied to the field of biochemical engineering. These techniques incorporate the expert's experience into the optimization routine and provide valuable information about the zone of possible solutions. This paper presents a multicriteria optimization strategy that generates a Pareto domain, given a set of conflicting objective criteria, and determines the optimal operating region for the production of gluconic acid using the net flow method (NFM). The objective criteria include maximizing the productivity and concentration of gluconic acid, while minimizing the residual substrate. Three optimization strategies are considered. The first two strategies identify the optimal operating region for the process inputs. The results yielded an acceptable compromise between productivity, gluconic acid concentration and residual substrate concentration. Fixing the process inputs representing the batch time, initial substrate concentration and initial biomass equal to their optimal values, the remaining simulations were used to study the sensitivity of the optimum operating region to changes in the oxygen mass transfer coefficient, K(L) a, by utilizing a multi-level K(L) a strategy. The results show that controlling K(L) a during the reaction reduced the production of biomass, which in turn resulted in increased productivity and concentration of gluconic acid above that of a fixed K(L) a.     

Citric and gluconic acid production from fig by Aspergillus niger using solid-state fermentation

The production of citric and gluconic acids from fig by Aspergillus niger ATCC 10577 in solid-state fermentation was investigated. The maximal citric and gluconic acids concentration (64 and 490 g/kg dry figs, respectively), citric acid yield (8%), and gluconic acid yield (63%) were obtained at a moisture level of 75%, initial pH 7.0, temperature 30°C, and fermentation time in 15 days. However, the highest biomass dry weight (40 g/kg wet substrate) and sugar utilization (90%) were obtained in cultures grown at 35°C. The addition of 6% (w/w) methanol into substrate increased the concentration of citric and gluconic acid from 64 and 490 to 96 and 685 g/kg dry fig, respectively. Journal of Industrial Microbiology & Biotechnology (2000) 25, 298–304. Received 15 April 2000/ Accepted in revised form 11 August 2000     

Modelling of batch fermentation and estimation of state variables using self-organizing feature maps

Summary A self-organizing feature map was used for modelling of batch yeast cultures. The model was constructed by training the neural network with experimental data of the specific rates. Estimates of state variables were obtained from the neural network model and differential mass balance equations via integration. They were compared with the experimental data. The neural network model showed a good modelling accuracy and interpolation capability.     

Cell reuse in submerged lactic acid batch fermentation

The reusability of biomass in lactic acid batch fermentation with free cells of Lactobacillus paracasei was studied in a 2–1 fermenter and in a 50-1 fermenter. In lab-scale fermentation experiments, 33 to 100% of the cell mass formed was reused in the subsequent batch in each case. In a series of seven consecutive batches, maximum values of lactate formation productivity of 6.32 to 11.54 g/l × h were observed at initial cell concentrations of 2.1 to 24.6 g/l. In all of the experiments, the initial cell viability was 78% or greater than 78%, and the final cell viability did not fall below 70%. At cell concentrations above 20 g/l, the productivity of lactic acid formation did not increase further, but remained constant. Because its level could be influenced by varying the proportions between the content of yeast extract, peptone and initial cell mass (1:1:2, 1:1:1 and 3.3.1) in the medium and no inhibitory effects were observed, this finding can be attributed to nutrient limitation. A low degree of cell reuse was reached in an analogous series of experiments carried out in a 50-1 fermenter. In this case, the initial cell concentration varied between 0.5 and 1.1 g/l, and therefore cell growth was not limited by nutrients in the first period of fermentation. Lactate production was still stable after six cell-reuse operations. The lactic acid yield did not fall below 90%. Temporary storage of the biomass in a refrigerator for a time interval of one to two weeks caused no significant impairment of overall lactate production, but a proportional prolongation of the lag phase occurred with increasing duration of storage.     

Modelling of temperature effects on batch microbial transglutaminase fermentation with Streptoverticillium mobaraense

Batch transglutaminase (MTG) fermentations by Streptovertivillium mobaraense WSH-Z2 at various temperatures ranging from 25 to 35 °C were studied. Dry cell weight and MTG activity could reach their maximal values of 25.1 g/l and 2.94 U/ml, respectively at 30 °C. One typical equation was used to describe the relationship between specific growth rate and culture temperature by comparing several typical equations. Different lag time was observed under various culture temperature. The low lag time was observed under high culture temperature. X = –a ₀(T – T ₀)² + X ₁ + a ₁ (1 – exp(a ₂ (T – T ₁))) and U = –a ₀ (T – T ₀ )² + U ₀ + a ₁ (1 – exp(a ₂ (T – T ₁ ))) could be used to describe the relationship between temperature and the maximal dry cell weight as well as the maximal MTG activity at each temperature.     

Optimisation of fermentation conditions for gluconic acid production by a mutant of Aspergillus niger

Aspergillus niger ORS-4, isolated from the sugarcane industry waste materials was found to produce notable level of gluconic acid. From this strain, a mutant Aspergillus niger ORS-4.410 having remarkable increase in gluconic acid production was isolated and compared for fermentation properties. Among the various substrates used, glucose resulted into maximum production of gluconic acid (78.04 g/L). 12% concentration led to maximum production. Effect of spore age and inoculum level on fermentation indicated an inoculum level of 2% of the 4-7 days old spores were best suited for gluconic acid production. Maximum gluconate production could be achieved after 10-12 days of the fermentation at 30 degrees C and at a pH of 5.5. Kinetic analysis of production indicated that growth of the mutant was favoured during initial stages of the fermentation (4-8 days) and production increased during the subsequent 8-12 days of the fermentation. CaCO3 and varying concentrations of different nutrients affected the production of gluconic acid. Analysis of variance for the factors evaluated the significant difference in the production levels.     

Formation of gluconic acid at low pH-values by free and immobilized Aspergillus niger cells during citric acid fermentation

Summary A recently developed immobilization method, characterized by the adsorption of the mycelia onto a glass-carrier in a fixed-bed reactor, was applied for citric acid production by Aspergillus niger ATCC 9142, and compared with conventional culture techniques.In a fixed-bed reactor and in a stirred fermenter a rapid gluconic acid production started immediately after nitrate exhaustion, though the pH was below 2.5 During a second production phase a comparatively small amount of citric acid was formed.In surface and shaken-flask cultures nearly no gluconic acid could be found, whereas citric acid yields were significantly higher than in the fixed-bed reactor and in the stirred fermenter.Manganese (0.8×10^–7 Mol×dm^–3 after 6 days incubation) from the stainless steel parts of the vessel seemed to be responsible for both gluconic acid production and small citric acid yields in the stirred fermenter and in the fixed-bed reactor.     

Influence of carbon sources on the viability and resuscitation of Acetobacter senegalensis during high-temperature gluconic acid fermentation

Much research has been conducted about different types of fermentation at high temperature, but only a few of them have studied cell viability changes during high-temperature fermentation. In this study, Acetobacter senegalensis, a thermo-tolerant strain, was used for gluconic acid production at 38 °C. The influences of different carbon sources and physicochemical conditions on cell viability and the resuscitation of viable but nonculturable (VBNC) cells formed during fermentation were studied. Based on the obtained results, A. senegalensis could oxidize 95 g l^− 1 glucose to gluconate at 38 °C (pH 5.5, yield 83%). However, despite the availability of carbon and nitrogen sources, the specific rates of glucose consumption (q_s) and gluconate production (q_p) reduced progressively. Interestingly, gradual q_s and q_p reduction coincided with gradual decrease in cellular dehydrogenase activity, cell envelope integrity, and cell culturability as well as with the formation of VBNC cells. Entry of cells into VBNC state during stationary phase partly stemmed from high fermentation temperature and long-term oxidation of glucose, because just about 48% of VBNC cells formed during stationary phase were resuscitated by supplementing the culture medium with an alternative favorite carbon source (low concentration of ethanol) and/or reducing incubation temperature to 30 °C. This indicates that ethanol, as a favorable carbon source, supports the repair of stressed cells. Since formation of VBNC cells is often inevitable during high-temperature fermentation, using an alternative carbon source together with changing physicochemical conditions may enable the resuscitation of VBNC cells and their use for several production cycles.

     

Using banana to generate lactic acid through batch process fermentation

We evaluated the usefulness of waste banana for generating lactic acid through batch fermentation, using Lactobacillus casei under three treatments. Two treatments consisted of substrates of diluted banana purée, one of which was enriched with salts and amino acids. The control treatment comprised a substrate suitable for L. casei growth. When fermentation was evaluated over time, significant differences (P<0.05) were found in the three treatments for each of five variables analyzed (generation and productivity of lactic acid, and consumption of glucose, fructose, and sucrose). Maximum productivity was (in g l^–1 h^–1) 0.13 for the regular banana treatment, 1.49 for the enriched banana, and 1.48 for the control, with no significant differences found between the latter two treatments. Glucose consumption curves showed that L. casei made greater use of the substrate in the enriched banana and control treatments than in the regular banana treatment. For fructose intake, the enriched banana treatment showed significantly better (P<0.05) results than the regular one. Sucrose consumption was insignificant (P<0.05), probably because fermentation time was too short. Even when enriched, diluted banana purée is an ineffective substrate for L. casei, probably because it lacks nutrients.     

Kinetic studies of the lactic acid fermentation in batch and continuous cultures

The fermentation kinetics of the homofermentative organism Lactobacillus delbrueckii in a glucose-yeast extract medium is studied in both batch and continuous culture under conditions of controlled pH. From a graphical analysis of the batch data, a mathematical model of the process is derived which relates bacterial growth, glucose utilization, and lactic acid formation. The parameters in the model represent the activity of the organism and are a function of pH, having a maximum value at about 5.90. In a continuous stirred tank fermentor (CSTF), the effect of pH, feed concentration, and residence time is observed. The feed medium is a constant ratio of two parts glucose to one part yeast extract plus added mineral salts. An approximate prediction of the steady-state behavior of the CSTF can be made using a method based on the kinetic model derived for the batch case. In making step changes from one steady state to another, the transient response is observed. Using the kinetic model to simulate the transient period, the calculated behavior qualitatively predicts the observed response.     

Optimization of fermentation conditions for gluconic acid production using Aspergillus niger immobilized on cellulose microfibrils

The optimisation of gluconic acid fermentation using immobilized Aspergillus niger on a highly porous cellulose support is described. Experimental results showing the effects of variations in oxygen partial pressure, glucose concentration and biomass concentration have been obtained with a continuous recirculation reactor. Levels of dissolved oxygen and glucose concentrations during fermentation significantly affect the production and fermentation time. The optimum biomass requirement on a porous cellulose support has been estimated to be 0.234 mg cm⁻² for efficient bioconversion. Increasing the quantum of biomass beyond this value resulted in an overgrown biofilm, which affected productivity adversely. Morphological characteristics of immobilized A. niger have also been investigated.