Citric acid fermentation by the mutant strain of theAspergillus niger resistant to manganese ions inhibition

Summary A new mutant strain,Aspergillus niger GS-III, showing resistance to manganese ions inhibition of citric acid fermentation on a sugarcane molasses containing medium was induced fromAspergillus niger KCU 520, a high citric acid-yielding strain. In submerged, surface or continuous cultures in the presence of manganese ions concentration upto 1.5 ppm the mutant strain yielded citric acid about 90 KgM^–3 . The citric acid yield was comparable to that obtained with the parental strain KCU 520 in the absence of manganese ions, but it was atleast 3-fold higher than that obtained by the latter in the presence of manganese ions. The mutant strain immobilized in calcium alginate beads was used in combination with surface-stabilized cultures for about 36-days in a continuous flow horizontal fermenter without any apparent loss in citric acid productivity. These results indicate that the manganese-resistant mutant is stable and may be used in the presence of sufficient manganese ions concentration (1.5 ppm) in the fermentation medium. This capability of the mutant strainA. niger GS-III has been correlated with greatly reduced levels (about one-thirds) of the NADP⁺ -isocitric dehydrogenase, one of the control points for citric acid accumulation.     

Ammonium ion & citric acid supplementation in batch cultures ofAspergillus niger B 60

Summary The effect of a single pulse of ammonium sulphate or of citrate upon the progress and final outcome of a batch citric acid fermentation was studied. It was found that the optimum addition time for the supplemental N was in the range of 40 to 75 h. Final citric acid concentration achieved was significantly increased when the concentration of N source added was between 0.25 and 0.5 kg m^–3. The mechanism of the observed stimulation seemed to be an indirect one. Addition of exogenous citric acid to the broth, led to an increase in citrate production by the culture. The optimum time for citric acid addition was around 90 h.Nomenclature Y_p/s Yield of citric acid produced (kg) on sucrose consumed (kg) - P/t Overall citric acid productivity (kg m^–3 h^–1)     

Production of citric acid with immobilized Yarrowia lipolytica

Summary Citric acid was produced with immobilized Yarrowia lipolytica yeast in repeated batch-shake-flask and air-lift fermentations. In active and passive immobilization methods calcium alginate, -carrageenan, polyurethane gel, nylon web and polyurethane foams were tested as carriers in repeated-batch fermentations. The highest citric acid productivity of 155 mg l^–1 h^–1 was reached with alginate-bead-immobilized cells in the first batch. A decrease in bead diameter from 5–6 mm to 2–3 mm increased the volumetric citric acid productivity threefold. In an air-lift bioreactor the highest citric acid productivity of 120 mg l^–1 h^–1 with a product concentration of 16.4 g l^–1 was obtained with cells immobilized in -carrageenan beads. Offprint requests to: H. Kautola     

Lactic acid production by aggregated continuous cultures of Lactobacillus acidophilus under aerobic conditions

Summary The ability of Lactobacillus acidophilus to aggregate, to produce lactic acid for a long term continuous fermentation process and to exist as aggregate cell cultures in a gas-lift reactor under aerobic conditions was studied. The main product of fermentation was lactic acid and only the traces of other end-products were determined. The highest fermentation efficiency of lactic acid was 98.6% and the highest productivity was 9.6 g.l^–1.h^–1 of lactic acid.     

Ethanol from hydrolyzed whey permeate using Saccharomyces cerevisiae in a membrane recycle bioreactor

A diauxic fermentation was observed during batch fermentation of enzyme-hydrolyzed whey permeate to ethanol by Saccharomyces cerevisiae. Glucose was consumed before and much faster than galactose. In the continuous membrane recycle bioreactor (MRB), sugar utilization was a function of dilution rate and concentration of sugars. At a cell concentration of 160 kg/m³, optimum productivity was 31 kg/(m³ · h) at ethanol concentration of 65 kg/m³. Low levels of acetate (0.05–0.1 M) reduced cell growth during continuous fermentation, but also reduced galactose utilization.     

Ethanol production by immobilized whole cells ofZymomonas mobilis in a continuous flow expanded bed bioreactor and a continuous flow stirred tank bioreactor

Continuous ethanol fermentation by immobilized whole cells ofZymomonas mobilis was investigated in an expanded bed bioreactor and in a continuous stirred tank reactor at glucose concentrations of 100, 150 and 200 g L^–1. The effect of different dilution rates on ethanol production by immobilized whole cells ofZymomonas mobilis was studied in both reactors. The maximum ethanol productivity attained was 21 g L^–1 h^–1 at a dilution rate of 0.36 h^–1 with 150 g glucose L^–1 in the continuous expanded bed bioreactor. The conversion of glucose to ethanol was independent of the glucose concentration in both reactors.     

Continuous solvent production by Clostridium beijerinckii BA101 immobilized by adsorption onto brick

The performance of a continuous bioreactor containing Clostridium beijerinckii BA101 adsorbed onto clay brick was examined for the fermentation of acetone, butanol, and ethanol (ABE). Dilution rates from 0.3 to 2.5 h^–1 were investigated with the highest solvent productivity of 15.8 g l^–1 h^–1 being obtained at 2.0 h^–1. The solvent yield at this dilution rate was found to be 0.38 g g^–1 and total solvent concentration was 7.9 g l^–1. The solvent yield was maximum at 0.45 at a dilution rate of 0.3 h^–1. The maximum solvent productivity obtained was found to be 2.5 times greater than most other immobilized continuous and cell recycle systems previously reported for ABE fermentation. A higher dilution rate (above 2.0 h^–1) resulted in acid production rather than solvent production. This reactor was found to be stable for over 550 h. Scanning electron micrographs (SEM) demonstrated that a large amount of C. beijerinckii cells were adsorbed onto the brick support.     

New cell recycle ethanol fermentation with periodic cleaning of filter with gas

High ethanol productivities were obtained by cell recycle cultures of yeast and bacterial strains at a dry cell concentration of 200 kg cells m^–3 using a new membrane bioreactor system. The filtration rates of the cultures were stabilized by removing the microbial cake on the filter with periodic back flows of the fermentation gas through the filter. For instance, the filtration flux of 0.023 m³m^–2h^–1 was maintained for 30 h with the periodic cleaning of the filter, whereas it decreased at a half time of 2 h without the cleaning. Ethanol productivity, ethanol concentration and filtration flux attained were: 68.7 kg/(m³ · h), 62.7 kg/m³ and 0.029 m³m^–2h^–1 for Saccharomyces carlsbergensis, LAM1068, the respective values for Zymomonas mobilis, ZM4, were: 93.7, 33 5 and 0.074.     

Batch and continuous ribonuclease production by immobilized Aspergillus clavatus cells in a bubble-collumn bioreactor

Summary Ribonuclease production using immobilized cells (IC) of Aspergillus clavatus has been studied under batch, repeated-batch and continuous fermentation conditions in a bubble-column bioreactor and compared with production by free cells, Immobilization was achieved by the method of cryostructurization in polyvinyl alcohol beads. The effect of various aeration rates in a column bioreactor has been investigated. Enzyme production by IC [42 000 units (U)·l^–1] during batch fermentations was comparable to that of a free-cell system. The specific productivity of IC was 8.5 times higher than that of free cells. In repeated batch fermentation at various aeration rates, successful reuse of IC was obtained, with comparable levels of enzyme production. Continuous ribonuclease production was achieved for 44 days at 1 vvm aeration and a dilution rate of 0.0 h^–1 with volumetric productivity (450 U·1^–1) and yield.     

Production of poly-γ-glutamic acid by fed-batch culture of Bacillus licheniformis

Fed-batch cultures of Bacillus licheniformis produced poly--glutamic acid (PGA), a water-soluble biodegradable polymer. PGA reached 35 g l^–1 with a productivity of 1 g l^–1 h^–1 by pulsed-feeding of citric acid (1.44 g h^–1) and l-glutamic acid (2.4 g h^–1) when citric acid was depleted from the culture medium.     

Application of a membrane recycle bioreactor for continuous ethanol production

Summary A series of continuous fermentations were carried out with a production strain of the yeast Saccharomyces cerevisiae in a membrane bioreactor. A membrane separation module composed of ultrafiltration tubular membranes retained all biomass in a fermentation zone of the bioreactor and allowed continuous removal of fermentation products into a cell-free permeate. In a system with total (100%) cell recycle the impact of fermentation conditions [dilution rate (0.03–0.3 h^–1); substrate concentration in the feed (50–300 g·1^–1); biomass concentration (depending on the experimental conditions)] was studied on the behaviour of the immobilized cell population and on ethanol formation. Maximum ethanol productivity (15 g·1^–1·h^–1) was attained at an ethanol concentration of 81 g·1^–1. The highest demands of cells for maintenance energy were found at the maximum feed substrate concentration (300 g·1^–1) and at very low concentrations of cells in the broth.     

The effect of yeast extract supplementation on the production of lactic acid from whey permeate byLactobacillus helueticus

Summary Batch and continuous two-stage cultures have been conducted in order to determine the effect of yeast extract (YE) on the homolactic fermentation of whey permeate byLactobacillus helveticus. Supplementation with YE had a significant effet on lactic acid concentration, volumetric productivity, and substrate conversion, but not on lactic acid yield. Volumetric productivity in the first stage increased from 2 to 9 g l^–1 per hour by increasing the YE concentration from 1.5 to 25 g l^–1 At the same time conversion improved from 22% to 93% at a dilution rate of 0.2 h^–1. The second stage demonstrated the effect of YE at a lower dilution rate (0.14 h^–1. A high system conversion (97%) and a high final lactic acid concentration (40 g l^–1) were achieved with 10 g l^–1 YE.     

Continuous production of citric acid from dairy wastewater using immobilized<Emphasis Type="Italic">Aspergillus niger</Emphasis> ATCC 9142

The continuous production of citric acid from dairy wastewater was investigated using calcium-alginate immobilizedAspergillus niger ATCC 9142. The citric acid productivity and yield were strongly affected by the culture conditions. The optimal pH, temperature, and dilution rate were 3.0, 30°C, and 0.025 h⁻¹, respectively. Under optimal culture conditions, the maximum productivity, concentration, and yield of citric acid produced by the calcium-alginate immobilizedAspergillus niger were 160 mg L⁻¹ h⁻¹, 4.5 g/L, and 70.3% respectively. The culture was continuously perfored for 20 days without any apparent loss in citric acid productivity. Conversely, under the same conditions with a batch shake-flask culture, the maximum productivity, citric acid concentration, and yield were only 63.3 mg L⁻¹ h⁻¹, 4.7 g/L and 51.4%, respectively. Therefore, the results suggest that the bioreactor used in this study could be potentially used for continuous citric acid production from dairy wastewater by applying calcium-alginate immobilizedAspergillus niger.     

Variation of fatty acid profile with solar cycle in outdoor chemostat culture ofIsochrysis galbana ALII-4

Outdoor chemostat cultures of the marine microalgaIsochrysis galbana at constant dilution rate (0.034 h^–1 ) have been carried out under different weather conditions. Steady-state biomass concentrations were 1.61±0.03 kg m^–3 in May and 0.95±0.04 kg m^–3 in July, resulting in biomass output rates of 0.54 kg m^–3 d^–1 and 0.32 kg m^–3 d^–1 in May and July, respectively. Two patterns of daily variation with the solar cycle were observed in the fatty acid content. Saturated and mono-unsaturated fatty acids (16:0 and 16:1n7) show significant variation with the solar cycle, associated with short-term changes in environmental factors. Palmitic and palmitoleic acids are generated during daylight and consumed during the dark period. However, polyunsaturated fatty acids do not show a significant response to the solar cycle and their changes are associated with long-term variation in environmental factors. The maximum EPA productivity was obtained in May, 14.1 g m^–3 d^–1, which is close to that found in the literature for indoor continuous cultures. Nonetheless, the outdoor EPA content (up to 2.61 % d.wt) was lower than the indoor EPA content from a previous study (5% d.wt).     

Continuous lactic acid fermentation with concentrated product recovery by ultrafiltration and electrodialysis

Lactose of sweet whey permeate was converted into sodium lactate byLactobacillus helveticus. To increase the, productivity of the lactic acid fermentation and to reduce the amounts of effluents, the bioreactor was coupled with an ultrafiltration module and an electrodialysis unit. Without the electrodialyzer, with total cell recycling and at a dilution rate of 0.88 h^–1, a cellular concentration of 64 gl^–1 and a productivity of 22 gl^–1 h^–1 were obtained. When the electrodialysis unit is coupled, the outlet concentration of lactate was stabilized at 85±5 gl^–1.     

Propionic acid and biomass production using continuous ultrafiltration fermentation of whey

Summary This paper presents a study of propionic acid and propionibacteria production from whey by usingPropionibacterium acidi-propionici in continuous fermentation with cell recycle. The highest propionic acid volumetric productivity achieved was 5 g.l^–1.h^–1 with no biomass bleeding. A maximal biomass concentration of 130 g.l^–1 was achieved before initiating biomass bleeding to give a biomass volumetric productivity of 3.2 g.l^–1.h^–1 with a biomass of 75 g.l^–1 and a propionic acid productivity of 3.6 g.l^–1.h^–1 (for about 100 hours i.e. more than 50 residence times).     

Fed-batch fermentation of Lactobacillus lactis for hyper-production of l-lactic acid

A fed-batch fermentation of Lactobacillus lactis to produce l-lactic acid was developed in which the residual glucose concentration in the culture was used to control a continuous feeding strategy. Up to 210 g l-lactic acid l^–1 (97% yield) was obtained. The maximal dry cell was 2.7 g l^–1 and the average l-lactic acid productivity was 2.2 g l^–1 h^–1.     

In-situ recovery of butanol during fermentation

End-product inhibition in the acetone-butanol fermentation was reduced by using extractive fermentation to continuously remove acetone and butanol from the fermentation broth. In situ removal of inhibitory products from Clostridium acetobutylicum resulted in increased reactor productivity; volumetric butanol productivity increased from 0.58 kg/(m³h) in batch fermentation to 1.5 kg/(m³h) in fed-batch extractive fermentation using oleyl alcohol as the extraction solvent. The use of fed-batch operation allowed glucose solutions of up to 500 kg/m³ to be fermented, resulting in a 3.5- to 5-fold decrease in waste water volume. Butanol reached a concentration of 30–35 kg/m³ in the oleyl alcohol extractant at the end of fermentation, a concentration that is 2–3 times higher than is possible in regular batch or fed-batch fermentation. Butanol productivities and glucose conversions in fed-batch extractive fermentation compare favorable with continuous fermentation and in situ product removal fermentations.List of Symbols C _g kg/m³ concentration of glucose in the feed - C _w dm³/m³ concentration of water in the feed - F(t) cm³/h flowrate of feed to the fermentor at time t - V(t) dm³ broth volume at time t - V _i dm³ initial broth volume - V _si dm³ volume of the i-th aqueous phase sample - effective fraction of water in the feed Part 1. Bioprocess Engineering 2 (1987) 1–12     

Optimization of a packed bed bioreactor with immobilized cells using experimental design

A central composite design was employed for the optimization of heterogeneous enzymatic hydrolysis of sucrose. The reaction was catalyzed by whole yeast cells of Saccharomyces cerevisiae immobilized in Ca-pectate gel. Bioreactor volumetric productivity was chosen as an optimization criterion, while temperature and gel biomass concentration were optimization parameters. Sucrose inlet concentration of 700 kg m^–3 and outlet conversion of 65% were constant in all experiments. In the temperature range 51–73 °C and biomass concentration range 11–39 kg m^–3 (dry mass of cells), the dependence of bioreactor productivity on the two factors was described by a second order polynom regression equation. No simple optimum was revealed by the experimental design. The bioreactor productivity increased within the whole experimental range of biomass concentration, whereas a temperature optimum was found to be between 60 and 65 °C.List of Symbols b _j jth regression coefficient - c _Si kg m^–3 inlet sucrose concentration - F m³ min^–1 flow rate - F F distribution - f _LF degrees of freedom of lack of fit variance - f _P degrees of freedom of pure error variance - N total number of runs - n ₀ number of runs in the centre of design - P kg m^–3 min^–1 productivity - s _LF ² lack of fit variance - SS _LF lack of fit sum of squares - S _p ² pure error variance - SS _P pure error sum of squares - SS _R total residual sum of squares - V _b m³ bioreactor bed volume - X _O outlet conversion - x ₁ 1st factor - coded temperature - x ₂ 2nd factor - coded biomass concentration - y kgm^–3min^–1 measured response (productivity) - kg m^–3 min^–1 estimated response (productivity) - y _Oi kg m^–3 min^–1 measured response in the centre of design - ¯y ₀ kg m^–3 min^–1 average of response in the centre of design     

Use of cell recycle in the aerobic fermentative production of citric acid by yeast

Summary An aerobic continuous stirred tank bioreactor with cell recycle was used to produce citric acid from glucose with a yeastSaccharomycopsis lipolytica NRRL Y7576. Specific rate of total acid production was 0.045h^–1, yield on glucose was 0.86 g/g and volumetric productivity was 1.16 g acid/Lh; all higher than or similar to batch values. Effluent acid concentration was 75g/L. In batch, under nitrogen limited. conditions, stability of citric acid synthesis and excretion was constant over a period of 700 hours. Under conditions of cell recycle, cell concentration and rate of acid production were constant over 200 hours of operation.