Influence of amino nitrogen in the culture medium enhances the production of δ-endotoxin and biomass of <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> var. <Emphasis Type="Italic">israelensis</Emphasis> for the large-scale production of the mosquito control agent

We reported here the role of amino nitrogen in the commercial production of Bacillus thuringiensis var. israelensis media design. The insect pathogen B. thuringiensis var. israelensis was cultured in different media containing varying initial levels of amino nitrogen sources obtained from three different commercial venders. The biomass, mosquito larval toxicity and spore count produced were measured during the fermentation process. The results showed that the higher level of initial amino nitrogen concentrations in the medium led to higher yield of biomass (dry weight 4.78 g l(-1)), larvicidal activity (LC(50) 18.52 ng ml(-1)) and spore count (3.24 x 10(11) CFU ml(-1)). Similarly decreasing the initial amino nitrogen concentration in the medium led to a decreased biomass (dry weight 1.64 g l(-1)), larvicidal activity (LC(50) 27.01 ng ml(-1)) and spore count (3.7 x 10(10) CFUml(-1)).     

Application of a simple yeast extract from spent brewer's yeast for growth and sporulation of Bacillus thuringiensis subsp. kurstaki: a Physiological Study

The suitability of using a simple brewer's yeast extract (BYE), prepared by autolysis of complete beer slurry, for growth and sporulation of Bacillus thuringiensis kurstaki was studied in baffled shake flasks. In a standard buffered medium with 2.5% (w/v) glucose and 1% (w/v) brewer's yeast extract, growth of B. t. kurstaki resulted in a low biomass production with considerable byproduct formation, including organic acids and a concomitant low medium pH, incomplete glucose utilization and marginal sporulation, whereas growth in the same medium with a commercial laboratory-grade yeast extract (Difco) resulted in a high biomass concentration, complete glucose utilization, relatively low levels of byproducts and complete sporulation (2.6 × 10⁹ spores/ml). When glucose was left out of the medium, however, growth parameters and sporulation were comparable for BYE and commercial yeast extract, but absolute biomass levels and spore counts were low. Iron was subsequently identified as a limiting factor in BYE. After addition of 3 mg iron sulphate/l, biomass formation in BYE-medium more than doubled, low byproduct formation was observed, and complete sporulation occurred (2.8 × 10⁹spores/ml). These data were slightly lower than those obtained in media with commercial yeast extract (3.6 × 10⁹spores/ml), which also benefited, but to a smaller extent, from addition of iron.     

Production of glomus intraradices propagules, an arbuscular mycorrhizal fungus, in an airlift bioreactor

This work addresses the symbiotic culture of the arbuscular mycorrhizal (AM) fungus Glomus intraradices with Daucus carota hairy roots transformed by Agrobacterium rhizogenes, in two submerged culture systems: Petri dish and airlift bioreactor. AM fungi play an active role in plant nutrition and protection against plant pathogens. These fungi are obligate biotrophs as they depend on a host plant for their needs in carbohydrates. The effect of the mycorrhizal roots inoculum-to-medium volume ratio on the growth of both symbionts was studied. A critical inoculating condition was observed at approximately 0.6 g dry biomass (DW). L-1 medium, above which root growth was significantly reduced when using a low-salt minimal (M) liquid medium previously developed for hairy root-AM fungi co-culture. Below critical inoculum conditions the maximum specific root growth and specific G. intraradices spore production rates of 0.021 and 0.035 d-1, respectively, were observed for Petri dish cultures. Maximum spore production in the airlift bioreactor was ten times lower than that of Petri dish cultures and obtained with the lowest inoculum assessed (0.13 g DW. L-1 medium) with 1.82 x 10(5) +/- 4.05 x 10(4) (SEM) spores (g DW inoculum)-1 (L medium)-1 in 107 d. This work proposes a second-generation bioprocess for AM fungi propagule production in bioreactors. Copyright 1999 John Wiley & Sons, Inc.     

Effect of Additives on the Production, Viability and Virulence of Blastospores of Metarhizium anisopliae

Studies on the blastospore production of Metarhizium anisopliae var. anisopliae were conducted in Adamek's medium used as a standard, enriched with lecithin, collagen, lactic acid or polyethylene glycol 200 (PEG 200) to increase spore yield and suppress mycelial pellet formation. The addition of 5% lecithin resulted in a significant 10-fold increase in spore yield up to 1.9 108 blastospores/ml compared with 1.9 107 spores/ml in the standard medium. Collagen (3%) increased the number of blastospores 3.7-fold, and lactic acid (1.5%) two-fold. A reduction of mycelial pellet formation in favour of spore production was noted with each additive. The viability of blastospores at 40IC from media with lecithin, collagen and lactic acid suspended in 25% Ringer's solution was comparable to that of spores produced in the standard medium. Striking differences were noticed in the viability of spores produced with 5% PEG 200 in standard medium. The half-life of blastospores produced in standard medium suspended in sunflower oil was 33.6 h and that of 5% PEG 200 spores only 25.2 h. In bioassays, the virulence of spores produced in standard medium to which 3% lecithin, 3% collagen, 1.5% lactic acid or 5% PEG 200 had been added was tested against third-instar nymphs of Locusta migratoria migratorioides (R. & F.). The median lethal time and the mortality of L. migratoria achieved with blastospores produced with 3% lecithin (5.7 days, 99%) was comparable to that of blastospores from standard medium (5.1 days, 98%). The virulence of blastospores from all other media with additives was significantly reduced.     

Influence of growth conditions on the production of a nisin-like bacteriocin by Lactococcus lactis subsp. lactis A164 isolated from kimchi

The influence of growth parameters on the fermentative production of a nisin-like bacteriocin by Lactococcus lactis subsp. lactis A164 isolated from kimchi was studied. The bacteriocin production was greatly affected by carbon and nitrogen sources. Strain A164 produced at least 4-fold greater bacteriocin in M17 broth supplemented with lactose than other carbon sources. The amount of 3% yeast extract was found to be the optimal organic nitrogen source. While the maximum biomass was obtained at 37 degrees C, the optimal temperature for the bacteriocin production was 30 degrees C. The bacteriocin production was also affected by pH of the culture broth. The optimal pH for growth and bacteriocin production was 6.0. Although the cell growth at pH 6.0 was nearly the same level at pH 5.5 and 6.5, the greater bacteriocin activity was observed at pH 6.0. Exponential growth took place only during an initial period of the cultivation, and then linear growth was observed. Linear growth rates increased from 0.160 g(DCW) x l(-1) x h(-1) to 0.245 g(DCW) x l(-1) x h(-1) with increases in lactose concentrations from 0.5 to 3.0%. Maximum biomass was also increased from 1.88 g(DCW) x l(-1) to 4.29 g(DCW) x l(-1). However, increase in lactose concentration did not prolong the active growth phase. After 20 h cultivation, cell growth stopped regardless of lactose concentration. Production of the bacteriocin showed primary metabolic kinetics. However, bacteriocin yield based on cell mass increased greatly during the late growth phase. A maximum activity of 131x10(3) AU x ml(-1) was obtained at early stationary growth phase (20 h) during the batch fermentation in M17L broth (3.0% lactose) at 30 degrees C and pH 6.0.     

Spore Yield and Microcycle Conidiation of Colletotrichum gloeosporioides in Liquid Culture

The effect of V8 juice concentration (5 to 40%, vol/vol), spore inoculum density (10⁵ and 10⁷ spores per ml), and liquid batch or fed-batch culture condition on mycelium and spore production by Colletotrichum gloeosporioides was evaluated. The amount of mycelium produced, the time required for initiation of sporulation following attainment of maximum mycelium, and the time for attainment of maximum spore concentration increased with increasing V8 juice concentration in batch culture. Cultures containing V8 juice at >10% achieved a similar spore density (apparent spore-carrying capacity) of about 0.8 mg of spores per ml (1 × 10⁷ to 2 × 10⁷ spores per ml) independent of inoculum density and V8 juice concentration. The relative spore yield decreased from a high of 64% of the total biomass for the low-inoculum 5% V8 culture, through 13% for the analogous 40% V8 culture, to a low of 2% for the high-inoculum 27% V8 culture. Fed-batch cultures were used to establish conditions of high spore density and low substrate availability but high substrate flux. The rate of addition of V8 juice was adjusted to approximate the rate of substrate utilization by the (increasing) biomass. The final spore concentration was about four times higher (3.0 mg of spores per ml) than the apparent spore-carrying capacity in batch culture. This high spore yield was obtained at the expense of greatly reduced mycelium, resulting in a high relative spore yield (62% of the total biomass). Microcycle conidiation occurred in the fed-batch but not batch systems. These data indicate that substrate-limited, fed-batch culture can be used to increase the amount and efficiency of spore production by C. gloeosporioides by maintaining microcycle conidiation conditions favoring allocation of nutrients to spore rather than mycelium production.     

Optimization of the spore production of Penicillium roqueforti in solid substrate fermentation on buckwheat seeds

Summary The effect of substrate (buckwheat seeds) pretreatment on the growth and the sporulation behaviour of Penicillium roqueforti is presented. When a saccharifying enzyme (-amylase) is added to a medium which exhibits a low water content (0.46 g water/g initial dry matter, IDM), a more rapid internal colonization of the seeds occurs, but the final spore production does not increase and remains close to 8.10⁹ spores/g dry matter (DM) at 500 h. No carbon source limitation is then observed. The addition of casein hydrolysate to this medium gives rise to a great increase of the sporulation, since 14.5 10⁹ spores/g DM are obtained after 600 h. This result is attained by a better spore yield from the mycelium, the substrate colonization being unchanged. High water content (0.60 g water/g IDM) of buckwheat seeds induces a shorter cultivation time along with a higher biomass production. However, the spore content of the medium remains close to the low water content one, but 60% total spores are external against 30% to 35% in the other media.     

A kinetic model describing Shewanella oneidensis MR-1 growth, substrate consumption, and product secretion

Aerobic growth of Shewanella oneidensis MR-1 in minimal lactate medium was studied in batch cultivation. Acetate production was observed in the middle of the exponential growth phase and was enhanced when the dissolved oxygen (DO) concentration was low. Once the lactate was nearly exhausted, S. oneidensis MR-1 used the acetate produced during growth on lactate with a similar biomass yield as lactate. A two-substrate Monod model, with competitive and uncompetitive substrate inhibition, was devised to describe the dependence of biomass growth on lactate, acetate, and oxygen and the acetate growth inhibition across a broad range of concentrations. The parameters estimated for this model indicate interesting growth kinetics: lactate is converted to acetate stoichiometrically regardless of the DO concentration; cells grow well even at low DO levels, presumably due to a very low K(m) for oxygen; cells metabolize acetate (maximum specific growth rate, micro(max,A) of 0.28 h(-1)) as a single carbon source slower than they metabolize lactate (micro(max,L) of 0.47 h(-1)); and growth on acetate is self-inhibiting at a concentration greater than 10 mM. After estimating model parameters to describe growth and metabolism under six different nutrient conditions, the model was able to successfully estimate growth, oxygen and lactate consumption, and acetate production and consumption under entirely different growth conditions.     

A procedure for high-yield spore production by Bacillus subtilis

Bacillus subtilis spores have a number of potential applications, which include their use as probiotics and competitive exclusion agents to control zoonotic pathogens in animal production. The effect of cultivation conditions on Bacillus subtilis growth and sporulation was investigated in batch bioreactions performed at a 2-L scale. Studies of the cultivation conditions (pH, dissolved oxygen concentration, and media composition) led to an increase of the maximum concentration of vegetative cell from 2.6 x 10(9) to 2.2 x 10(10) cells mL(-)(1) and the spore concentration from 4.2 x 10(8) to 5.6 x 10(9) spores mL(-)(1). A fed-batch bioprocess was developed with the addition of a nutrient feeding solution using an exponential feeding profile obtained from the mass balance equations. Using the developed feeding profile, starting at the middle of the exponential growth phase and finishing in the late exponential phase, an increase of the maximum vegetative cell concentration and spore concentration up to 3.6 x 10(10) cells mL(-)(1) and 7.4 x 10(9) spores mL(-)(1), respectively, was obtained. Using the developed fed-batch bioreaction a 14-fold increase in the concentration of the vegetative cells was achieved. Moreover, the efficiency of sporulation under fed-batch bioreaction was 21%, which permitted a 19-fold increase in the final spore concentration, to a final value of 7.4 x 10(9) spores mL(-)(1). This represents a 3-fold increase relative to the highest reported value for Bacillus subtilis spore production.     

Physiology of the yeast Kluyveromyces marxianus during batch and chemostat cultures with glucose as the sole carbon source

Growth, substrate consumption, metabolite formation, biomass composition and respiratory parameters of Kluyveromyces marxianus ATCC 26548 were determined during aerobic batch and chemostat cultivations, using mineral medium with glucose as the sole carbon source, at 30 degrees C and pH 5.0. Carbon balances closed within 95-101% in all experiments. A maximum specific growth rate of 0.56 h(-1), a biomass yield on glucose of 0.51 g g(-1), and a maximum specific consumption of oxygen of 11.1 mmol g(-1) h(-1) were obtained during batch cultures. The concentration of excreted metabolites was very low at the culture conditions applied, representing 6% of the consumed carbon at most. Acetate and pyruvate were excreted to a larger extent than ethanol under the batch conditions, and the protein content accounted for 54.6% of the biomass dry weight. Steady states were obtained during chemostats at dilution rates of 0.1, 0.25 and 0.5 h(-1). At the two former dilution rates, cells grew at carbon limitation and the biomass yield on glucose was similar to that obtained under the batch conditions. Metabolite formation was rather low, accounting for a total of 0.005 C-mol C-mol(-1) substrate. At 0.5 h(-1), although the biomass yield on glucose was similar to the value obtained under the above-mentioned conditions, the cultivation was not under carbon limitation. Under this condition, 2-oxoglutarate, acetate, pyruvate and ethanol were the prevalent metabolites excreted. Total metabolite formation only accounted to 0.056 C-mol C-mol(-1) of substrate. A very high protein and a low carbohydrate content (71.9% and 9.6% of biomass dry weight, respectively) were measured in cells under this condition. It is concluded that K. marxianus aligns with the so-called aerobic-respiring or Crabtree-negative yeasts. Furthermore, it has one of the highest growth rates among yeasts, and a high capacity of converting sugar into biomass, even when carbon is not the limiting nutrient. These results provide useful data regarding the future application of K. marxianus in processes aimed at the production of biomass-linked compounds, with high yields and productivities.     

Issues in the culture of the extremely thermophilic methanogen, Methanothermus fervidus

Basic issues in the culture of the extremely thermophilic archaeon, Methanothermus fervidus, have been investigated, including culture medium formulation, substrate yield and product yield coefficient, growth rate and stoichiometry, and H(2) uptake kinetics. The pH optimum for growth of this organism was estimated at 6.9. Growth medium buffered with PIPES instead of bicarbonate supported both increased growth rate and maximum biomass concentration. Substitution of titanium(III) citrate for the reducing agent sodium sulfide improved culture performance as well. However, independent adjustment of iron and nickel concentrations from 11 to 111 muM, respectively, and carbon dioxide partial pressure from 5 to 20 psia did not impact the culture of M. fervidus significantly. An elemental balance approach was utilized to aid in design of a defined medium to support growth to a target maximum biomass concentration of at least 1.0 g dry wt/L. The growth of this organism was limited by H(2) availability in this reformulated culture medium. The maximum growth rate and biomass concentration achieved in anaerobic vials with the defined medium was 0.16 h(-1) and 0.74 g dry wt/L, respectively. This maximum biomass concentration was a 72% improvement over that obtained with a literature-based defined medium. The Monod parameter, K(s), with H(2) as limiting substrate, was estimated at 1.1 +/- 0.4 psia (55 +/- 20 muM in the broth), based on a H(2) consumption study. Representative values for the substrate yield, Y(X/CO(2) ), and product yield coefficient, Y(CH(4)/) (X), were determined experimentally to be 1.78 +/- 0.04 g dry wt/mol CO(2), and 0.52 +/- 0.01 mol CH(4)/g dry wt, respectively. A bench-scale fermentation system suitable for the culture of extremely thermophilic anaerobes was designed and constructed and proved effective for the culture of M. fervidus. (c) 1993 Wiley & Sons, Inc.     

Elucidation of <Emphasis Type="Italic">Bacillus subtilis</Emphasis> KATMIRA 1933 Potential for Spore Production in Submerged Fermentation of Plant Raw Materials

In this study, the effects of several key factors to increase spore production by Bacillus subtilis subsp. KATMIRA 1933 were evaluated in shake flask experiments. In a synthetic medium, glucose concentration played a crucial role in the expression of bacilli sporulation capacity. In particular, maximum spore yield (2.3 × 10⁹ spores/mL) was achieved at low glucose concentration (2 g/L), and further gradual increase of the carbon source content in the medium caused a decrease in sporulation capacity. Substitution of glucose with several inexpensive lignocellulosic materials was found to be a reasonable way to achieve high cell density and sporulation. Of the materials tested, milled mandarin peels at a concentration of 40 g/L served as the best growth substrate. In these conditions, bacilli secreted sufficient levels of glycosyl hydrolases, providing slow hydrolysis of the mandarin peel’s polysaccharides to metabolizable sugars, providing the bacterial culture with an adequate carbon and energy source. Among nitrogen sources tested, peptone was found to favor spore production. Moreover, it was shown that cheese and cottage cheese whey usage, instead of distilled water, significantly increases spore formation. After optimization of the nutrient medium in the shake flask experiments, the technical feasibility of large-scale spore production by B. subtilis KATMIRA 1933 was confirmed in a laboratory fermenter. The spore yield (7 × 10¹⁰ spores/mL) obtained using a bioreactor was higher than those previously reported.     

<Emphasis Type="Italic">Bacillus amyloliquefaciens</Emphasis> Spore Production Under Solid-State Fermentation of Lignocellulosic Residues

This study was conducted to elucidate cultivation conditions determining Bacillus amyloliquefaciens B-1895 growth and enhanced spore formation during the solid-state fermentation (SSF) of agro-industrial lignocellulosic biomasses. Among the tested growth substrates, corncobs provided the highest yield of spores (47?×?10¹⁰ spores g^?1 biomass) while the mushroom spent substrate and sunflower oil mill appeared to be poor growth substrates for spore formation. Maximum spore yield (82?×?10¹⁰ spores g^?1 biomass) was achieved when 15 g corncobs were moistened with 60 ml of the optimized nutrient medium containing 10 g peptone, 2 g KH₂PO₄, 1 g MgSO₄·7H₂O, and 1 g NaCl per 1 l of distilled water. The cheese whey usage for wetting of lignocellulosic substrate instead water promoted spore formation and increased the spore number to 105?×?10¹⁰ spores g^?1. Addition to the cheese whey of optimized medium components favored sporulation process. The feasibility of developed medium and strategy was shown in scaled up SSF of corncobs in polypropylene bags since yield of 10?×?10¹¹ spores per gram of dry biomass was achieved. In the SSF of lignocellulose, B. amyloliquefaciens B-1895 secreted comparatively high cellulase and xylanase activities to ensure good growth of the bacterial culture.     

Factors affecting the production of a single-chain antibody fragment by Aspergillus awamori in a stirred tank reactor

A recombinant strain of Aspergillus awamori expressing anti-lysozyme single chain antibody fragments (scFv), under the control of a xylanase promoter, was studied in order to investigate the impact of medium, induction regime and protease production on the expression of the product. Experiments with the time of induction showed that the optimum results are achieved when induction is started in the late exponential phase (21 h after inoculation) improving the titer of the product from 14.5 mg L(-1), obtained in the early exponential phase (7 h after inoculation), to 16.2 mg L(-1). A 100% increase of the carbon (fructose) and nitrogen (ammonium sulfate) sources in the growth medium resulted in an increase in product concentration from 16.2 to 108.9 mg L(-1) and an increase in maximum dry cell weight from 7.5 to 11.5 g L(-1). A 50% reduction in the concentration of the inducer resulted in an increase in the product yield from 10 mg g(-1) dry cell weight to 12 mg g(-1). Proteolytic enzymes were produced during the fermentation up to concentrations equivalent to 1.4 g L(-1) trypsin, but they had no detrimental effect on the concentration of the antibody fragment.     

Production of Bacillus thuringiensis spores in total cell retention culture and two-stage continuous culture using an internal ceramic filter system

The production of Bacillus thuringiensis spores was investigated in a bioreactor incorporating a ceramic membrane filter to improve spore concentration and volumetric productivity. Two cultivation methods were used in this study: a total cell retention culture (TCRC), and a two-stage continuous culture with partial cell bleeding. In the TCRC, fed by 50 g/L of glucose, a spore concentration of 1.6 x 10(10) CFU/mL was obtained with a spore percentage of greater than 95% and a maximum cell mass of 82.2 g/L. The volumetric productivity was four times higher than that obtained from batch cultivation. In the two-stage continuous culture with partial cell bleeding spore concentration was strongly dependent on the bleed ratio. The spore concentration of 1.8 x 10(9) CFU/mL and the spore percentage of 70% were obtained at the second stage when a bleed ratio of 0.33 and a dilution rate of 0.23 h(-1) were used. (c) 1993 John Wiley & Sons, Inc.     

Optimization of process parameters by response surface methodology and kinetic modeling for batch production of canthaxanthin by Dietzia maris NIT-D (accession number: HM151403)

Dietzia maris NIT-D, a canthaxanthin producer, was isolated during routine screening of pigment-producing bacteria. Response surface methodology was applied for statistical designing of process parameters for biomass and canthaxanthin production. The effects of four process parameters (considered as independent variables), namely temperature (10-30?°C), pH (4.75-5.75), shaker speed (75-135?rpm) and percentage inoculum (0.5-2.5?%) on the biomass and canthaxanthin yield (considered as dependent variables) were studied. As much as 122?mg?L(-1) of canthaxanthin was obtained when Dietzia maris NIT-D was incubated for 120?h at 25?°C and 120?rpm, initial pH and percentage inoculum being 5.5 and 2?% respectively. The pigment yield is the highest reported till date, with Dietzia maris as the test organism. The maximum biomass yield was 7.39?g?L(-1) under optimized process parameters. The predicted values were also verified by validation experiments in 5-day fermentation. Different mathematical models were used to describe growth and production, considering the effect of glucose in batch mode. The kinetic constants were calculated by fitting the experimental data to the models. Cell growth was inhibited beyond a glucose concentration of 15?g?L(-1). Andrews' model gave the best fit with a R (2) value of 0.9993. During the exponential growth phase, the specific growth rate was found to remain fairly constant with respect to time. There was no inhibitory effect due to intracellular product accumulation for all concentrations of glucose. This observation is the first of its kind, as previous studies have reported that increasing accumulation of intracellular carotenoid exerts greater degree of inhibition on growth.     

Fermentative capacity in high-cell-density fed-batch cultures of baker's yeast

High-cell-density fed-batch processes for bakers' yeast production will involve a low-average-specific growth rate due to the limited oxygen-transfer capacity of industrial bioreactors. The relationship between specific growth rate and fermentative capacity was investigated in aerobic, sucrose-limited fed-batch cultures of an industrial bakers' yeast strain. Using a defined mineral medium, biomass concentrations of 130 g dry weight/L were reproducibly attained. After an initial exponential-feed phase (mu = 0.18 h(-1)), oxygen-transfer limitation necessitated a gradual decrease of the specific growth rate to ca. 0.01 h(-1). Throughout fed-batch cultivation, sugar metabolism was fully respiratory, with a biomass yield of 0.5 g biomass/g sucrose(-1). Fermentative capacity (assayed off-line as ethanol production rate under anaerobic conditions with excess glucose) showed a strong positive correlation with specific growth rate. The fermentative capacity observed at the end of the process (mu = 0.01 h(-1)) was only half that observed during the exponential-feed phase (mu = 0.18 h(-1)). During fed-batch cultivation, activities of glycolytic enzymes, pyruvate decarboxylase and alcohol dehydrogenase in cell extracts did not exhibit marked changes. This suggests that changes of fermentative capacity during fed-batch cultivation were not primarily caused by regulation of the synthesis of glycolytic enzymes.     

Growth kinetics of vitis vinifera cell suspension cultures: I. Shake flask cultures

Vitis vinifera cell suspension cultures carried out in shake flasks were closely examined for biomass growth and cell division in relation to carbohydrate, NH(4), NO(3)PO(4), and dissolved oxygen (DO)consumption. After inoculation, the oxygen uptake rate of the cultures measured on-tine was observed to increase continuously to a maximum value of 3.8 mmol O(2)L(-1)h(-1) at day 7 when cell division ceased and dissolved oxygen reached its lowest level of 17% air saturation. During this first phase of growth, the specific oxygen uptake rate remained constant at approximately 0.6 mmol 02 O(2) g(-1) dw h(-1)or approximately 2.2 mumol O(2), (10(6) cells)(-1) h(-1) whereas dry biomass concentration increased exponentially from 1.5 to 6.0 g dw L(-1). Thereafter, dry biomass concentration increased linearly to approximately 14 g dw L(-1) at day 14 following nitrate and carbohydrate uptake. During this second phase of growth, the biomass wet-to-dry weight ratio was found to increase in an inverse relationship with the estimated osmotic pressure of the culture medium. This corresponded to inflection points in the dry and wet biomass concentration and packed cell volume curves. Furthermore, growth and nutrient uptake results suggest that extracellular ammonium or phosphate ion availability may limit cell division. These findings indicate that cell division and biomass production of plant cell cultures may not always be completely associated, which suggests important new avenues to improve their productivity. (c) 1995 John Wiley & Sons, Inc.     

Improved production of the anticancer compound 1403C by glucose pulse feeding of marine Halorosellinia sp. (No. 1403) in submerged culture

Effects of different pulse fed-batch methods on production of the anti-cancer compound 1403C by marine mangrove endophytic fungus Halorosellinia sp. (No. 1403) in a 5-L bioreactor were investigated. Since high glucose concentrations improved mycelial growth but inhibited 1403C production, the cultures were pulse fed with glucose solutions to keep the residual glucose lower than 4 g/L but higher than 0.5 g/L during rapid growth phase (0-50 h). In this way, a maximum dry biomass, 1403C production and yield coefficient (Y1403C/X) of up to 4.5 g/L, 2.64 g/L and 0.59 g/g dry cell weight, respectively were achieved. These values are 22.7%, 98.0% and 61.4%, respectively higher than those obtained with batch cultures. This strategy is valuable for fermentation scale-up of Halorosellinia sp. (No. 1403) for 1403C production, and might also be applicable to other marine fungi cultures.     

Ethanol production from concentrated whey permeate using a fed-batch culture ofKluyveromyces fragilis

Summary Fed-batch fermentation of non-supplemented concentrated whey permeate resulted in high ethanol productivity for feeds of lactose for which batch fermentation had a poor performance. At an initial lactose concentration of 100 g/L and a constant lactose feeding rate of 18 g/h we have obtained: ethanol concentration 64 g/L, ethanol productivity 3.3 g/Lh, lactose consumption 100%, ethanol yield 0.47 g/g, and biomass yield 0.058 g/g.Nomenclature St total lactose fed per medium volume in the bioreactor, g/L - Si initial lactose concentration, g/L - F lactpse feeding rate, g/h - P final ethanol concentration, g/L - Y_p/s ethanol yield, g ethanol/g lactose - Y_x/s biomass yield, g biomass/g lactose - X_S lactose consumption, % - Qp overall ethanol volumetric productivity, g/Lh - m maximum specific growth rate, h - qsm maximum specific lactose consumption rate, g/gh - qpm maximum specific ethanol production rate, g/gh