The effect of electric field stimulation on the biosorption of uranium by non-living biomass derived from Kluyveromyces marxianus IMB3

Summary Non-living biomass from the thermotolerant, ethanol-producing yeast strain Kluyveromyces marxianus IMB3 is capable of uranium biosorption. The biomass has an observed biosorption capacity of 115mg uranium/g dry weight of biomass with a calculated value of 127mg uranium/g dry weight. Following exposure of the biomass to electric fields of 2,500 V/cm for 20msec. the maximum biosorption capacity (observed or calculated) for uranium did not differ significantly for the untreated biomass. However, at lower residual concentrations of uranium (<10mg/L) the capacity of the treated biomass for uranium was significantly increased above values obtained with untreated material.     

Iminodiacetate and nitrilotriacetate degradation by Kluyveromyces marxianus IMB3.

The thermotolerant yeast Kluyveromyces marxianus IMB3 was capable of utilising either iminodiacetate or nitrilotriacetate as a sole source of nitrogen for growth. Cell extracts contained iminodiacetate dehydrogenase and nitrilotriacetate monooxygenase activities, suggesting the presence in the yeast of orthologues of these bacterial enzymes. The activities were not detectable in complete medium-growth cells, nor in nitrogen-starved cells, suggesting an inducible biodedgradation pathway for biodegradation of these xenobiotics, which has not been previously reported in a eukaryotic cell system. This observation emphasises the hitherto unrealised importance of yeast strains in the biodegradation of xenobiotics in the environment.     

Biosorption of textile dyes by biomass derived from Kluyveromyces marxianus IMB3

Since it had previously been found that biomass derived from the thermotolerant ethanol-producing yeast strain Kluyveromyces marxianus IMB3 exhibited a relatively high affinity for heavy metals it was decided to determine whether or not it might be capable of textile dye biosorption. To this end, biosorption isotherm analysis was carried out using the biomass together with commonly-used textile dyes including Remazol Black B, Remazol Turquoise Blue, Remazol Red, Remazol Golden Yellow and Cibacron Orange. Although the dyes Remazol Black B, Remazol Turquoise Blue and Remazol Red adhered to the Langmuir model, the remaining dyes failed to do so. The observed biosorption capacities at equilibrium dye concentrations of 100?mg/l were compared and it was found that the biomass exhibited a significant affinity for each dye. The potential use of this biosorptive material in the bioremediation of textile processing effluents is discussed.     

Studies on the biosorption of uranium by a thermotolerant, ethanol-producing strain of Kluyveromyces marxianus

The ability of residual biomass from the thermotolerant ethanol-producing yeast strain Kluyveromyces marxianus IMB3 to function as a biosorbent for uranium has been examined. It was found that the biomass had an observed maximum biosorption capacity of 120?mg U/g dry weight of biomass. The calculated value for the biosorption maximum, obtained by fitting the data to the Langmuir model was found to be 130?mg U/g dry weight biomass. Maximum biosorption capacities were examined at a number of temperatures and both the observed and calculated values obtained for those capacities increased with increasing temperature. Decreasing the pH of the biosorbate solution resulted in a decrease in uptake capacity. When biosorption reactions were carried out using sea-water as the diluent it was found that the maximum biosorption capacity of the biomass increased significantly. Using transmission electron microscopy, uranium crystals were shown to be concentrated on the outer surface of the cell wall, although uranium deposition was also observed in the interior of the cell.     

Ethanol adaptation in a thermotolerant yeast strain Kluyveromyces marxianus IMB3

The maximum ethanol concentration produced from glucose in defined media at 45°C by the thermotolerant yeast Kluyveromyces marxianus IMB3 was 44 g L⁻¹. Acclimatisation of the strain through continuous culture at ethanol concentrations up to 80 g L⁻¹, shifted the maximum ethanol concentration at which growth was observed from 40 g L⁻¹ to 70 g L⁻¹. Four isolates were selected from the continuous culture, only one of which produced a significant increase in final ethanol concentration (50 ± 0.4 g L⁻¹), however in subsequent fermentations, following storage on nutrient agar plates, the maximum ethanol concentration was comparable with the original isolate. The maximum specific ethanol production rates (approximately 1.5 g (gh)⁻¹) were also comparable with the original strain except for one isolate (0.7 g (gh)⁻¹). The specific ethanol productivity decreased with ethanol concentration; this decrease correlated linearly (rval 0.92) with cell viability. Due to the transience of induced ethanol tolerance in the strain it was concluded that this was not a valid method for improving final ethanol concentrations or production rates. Received 18 July 1997/ Accepted in revised form 19 February 1998     

Ethanol production at 45°C by alginate-immobilized Kluyveromyces marxianus IMB3 during growth on lactose-containing media

The thermotolerant yeast strain Kluyveromyces marxianus IMB3 was immobilized in calcium alginate and this was used in batch-fed reactor systems to convert lactose (4?g/l) to ethanol. Production of ethanol by the free and immobilized biocatalyst in the presence and absence of Mn²⁺ was compared. In systems containing the free microorganism in the presence and absence of Mn²⁺, ethanol increased to a maximum of 8?g/l within 40 hours with no significant difference in production by both systems. Ethanol production by the immobilized system in the absence of Mn²⁺ increased to a maximum of 13?g/l within 40 hours and then decreased to 9?g/l within 80 hours. Ethanol production by the immobilized system in the presence of Mn²⁺ increased to 14?g/l within 60 hours and this decreased to 13?g/l at 80 hours. When all systems were re-fed at 80 hours, ethanol production by systems containing the free biocatalyst increased to a maximum of 3?g/l while the immobilized system in the presence of Mn²⁺ increased to a maximum of 12?g/l. Subsequent experiments involving re-feeding the system at shorter time intervals demonstrated that ethanol production by the immobilized system on lactose-containing media at 45?°C was far superior to ethanol production by the free biocatalyst.     

The effect of Mn2+ on ethanol production from lactose using Kluyveromyces marxianus IMB3 immobilized in magnetically responsive matrices

The thermotolerant, ethanol producing yeast strain, K. marxianus IMB3 was immobilized in calcium alginate containing magnetically responsive Fe₃O₄ particles. In these studies the β-galactosidase derived from K. marxianus IMB3 was immobilized onto the Fe₃O₄ particles prior to inclusion into the alginate matrix. Ethanol production by the immobilized microorganism in the presence of Fe₃O₄ reached a maximum of 16?g/L on 40?g/L lactose whereas prior immobilization of the enzyme to the particles and inclusion into the alginate matrix increased ethanol production to a maximum concentration of 18 g/L. When Mn²⁺ was incorporated into fermentations containing the immobilized enzyme in the alginate matrix, ethanol production increased further to a maximum concentration of 20?g/L. In addition, the behaviour of the magnetically responsive biocatalyst containing the co-immobilized enzyme was examined in a batch-fed system in the presence and absence of Mn²⁺.     

High-temperature alcoholic fermentation of whey using Kluyveromyces marxianus IMB3 yeast immobilized on delignified cellulosic material

A novel system for high-temperature alcoholic fermentation of whey is described. This system consists of Kluyveromyces marxianus yeast immobilized on delignified cellulosic material (DCM). The effect of pH, initial lactose concentration and temperature on the fermentation of a synthetic medium containing lactose was studied. Batch fermentations of whey were also carried out and the formation of volatile by-products was examined. The concentrations of higher alcohols were found to be in very low levels leading to a product of improved quality. The fermented whey had an improved characteristic aroma compared to unfermented whey. The possibility to use fermented whey as raw material for the production of a novel, low alcohol content drink was also investigated.     

Ethanol production by Kluyveromyces marxianus IMB3 during growth on straw-supplemented whiskey distillery spent wash at 45 °C

The thermotolerant, ethanol-producing yeast strain, Kluyveromyces marxianus IMB3 was grown on media consisting of straw-supplemented distillery spent wash from The Old Bushmill's Distillery Co. Ltd., Bushmills, Co Antrim, Northern Ireland. Media were supplemented with cellulase activity and fermentations were carried out at 45?°C. When pulverized straw was used as substrate in this system at concentrations of 2, 4 and 6% (w/v), ethanol concentrations increased to maxima of 1.45, 2.2 and 3?g/l, respectively. Based on straw containing a maximum of 40% cellulose, these ethanol concentrations accounted for 36, 27 and 24% of the maximum theoretical yield, respectively. When the straw was pre-treated with NaOH and used in the spent wash containing system at concentrations of 2, 4 and 6% (w/v) ethanol, concentrations increased to maxima of 3, 6.2 and 10.5?g/l, respectively and these accounted for 75, 76 and 86% of the maximum theoretical yield. When these results are compared with previously published data relating to the use of straw in laboratory-based media, they suggest that whiskey distillery spent wash may provide an adequate medium for supplementation with complex carbohydrate and subsequent ethanol production in simultaneous saccharification and fermentation processes.     

Simultaneous saccharification and fermentation of Kanlow switchgrass pretreated by hydrothermolysis using Kluyveromyces marxianus IMB4

A thermotolerant yeast strain named Kluyveromyces marxianus IMB4 was used in a simultaneous saccharification and fermentation (SSF) process using Kanlow switchgrass as a feedstock. Switchgrass was pretreated using hydrothermolysis at 200 degrees C for 10 min. After pretreatment, insoluble solids were separated from the liquid prehydrolyzate by filtration and washed with deionized water to remove soluble sugars and inhibitors. Insoluble solids were then hydrolyzed using a commercial cellulase preparation and the released glucose was fermented to ethanol by K. marxianus IMB4 in an SSF process. SSF temperature was 37, 41, or 45 degrees C and pH was 4.8 or 5.5. SSF was conducted for 7 days. Results were compared with a control of Saccharomyces cerevisiae D(5)A at 37 degrees C and pH 4.8. Fermentation by IMB4 at 45 and 41 degrees C ceased after 3 and 4 days, respectively, when a pH 4.8 citrate buffer was used. Fermentation continued for all 7 days using IMB4 at 37 degrees C and the control. When pH 5.5 citrate buffer was used, fermentation ceased after 96 h using IMB4 at 45 degrees C, and ethanol yield was greater than when pH 4.8 citrate buffer was used (78% theoretical). Ethanol yield using IMB4 at 45 degrees C, pH 5.5 was greater than the control after 48, 72, and 96 h (P < 0.05).     

Industrial scale ethanol production using the thermotolerant yeast Kluyveromyces marxianus IMB3 in an Indian distillery

The thermotolerant ethanol producing Kluyveromyces marxianus IMB3 yeast was used in eight 60m3 fermenters for industrial ethanol production in India using sugarcane molasses. Ethanol ranged between 6.0–7.2% (w/v) with added advantages of elimination of cooling during fermentation and shorter fermentation periods of 20h. © Rapid Science Ltd. 1998     

Studies on the growth of a thermotolerant yeast strain,Kluyveromyces marxianus IMB3, on sucrose containing media

A thermotolerant alcohol-producing yeast strain, Kluyveromyces marxianus IMB3 was shown to grow on sucrose (10% [w/v]) containing media at 45 °C. Under such conditions the organism reached stationary phase within 20 hours and yielded ethanol concentrations in the region of 33g/L. During growth on sucrose containing media the organism was found to produce a cell- associated activity capable of hydrolysing sucrose. This activity was shown to have a Km of 5.0mM when sucrose was used as the substrate. In addition the enzyme was shown to have a pH optimum of 5.0 and a temperature optimum of 50–55 °C and under those conditions the enzyme was shown to be relatively thermostable.     

Increased efficiency of substrate utilization by exposure of the thermotolerant yeast strain,Kluyveromyces marxianus IMB3 to electric-field stimulation

Summary In order to enhance cellobiose utilization and conversion of substrate to ethanol by the thermotolerant yeast strain, Kluyveromyces marxianus IMB3, the organism was exposed to short, intense electric pulses. When cells were treated with pulses measuring 0.25kV for 10mS, in the presence of cellobiose, ethanol production was found to increase by almost 40% above that found in fermentations containing non-treated cells. When the extracellular culture filtrate was assayed for -glucosidase activity no significant difference in levels was detected between treated and control systems. Increasing the voltage of the pulses resulted in a decrease in ethanol production.     

Fermentation of molasses using a thermotolerant yeast,Kluyveromyces marxianus IMB3: simplex optimisation of media supplements

 The use of molasses as a substrate for ethanol production by the thermotolerant yeast Kluyveromyces marxianus var. marxianus was investigated at 45°C. A maximum ethanol concentration of 7.4% (v/v) was produced from unsupplemented molasses at a concentration of 23% (v/v). The effect on ethanol production of increasing the sucrose concentration in 23% (v/v) molasses was determined. Increased sucrose concentration had a similar detrimental effect on the final ethanol produced as the increase in molasses concentration. This indicated that the effect may be due to increased osmotic activity as opposed to other components in the molasses. The optimum concentration of the supplements nitrogen, magnesium, potassium and fatty acid for maximum ethanol production rate was determined using the Nelder and Mead (Computer J 7:308–313, 1965) simplex optimisation method. The optimum concentrations of the supplements were 0.576 g l^-1 magnesium sulphate, 0.288 g l^-1 potassium dihydrogen phosphate and 0.36% (v/v) linseed oil. Added nitrogen in the form of ammonium sulphate did not affect the ethanol production rate. Received: 29 January 1996/Received revision: 23 April 1996/Accepted: 29 April 1996     

Simultaneous saccharification and fermentation of Kanlow switchgrass by thermotolerant Kluyveromyces marxianus IMB3: the effect of enzyme loading, temperature and higher solid loadings

Switchgrass (Panicum virgatum) was subjected to hydrothermolysis pretreatment and then used to study the effect of enzyme loading and temperature in a simultaneous saccharification and fermentation (SSF) with the thermotolerant yeast strain Kluyveromyces marxianus IMB3 at 8% solid loading. Various loadings of Accellerase 1500 between 0.1 and 1.1 mL g(-1) glucan were tested in SSF at 45 °C (activity of enzyme was 82.2 FPU mL(-1)). The optimum enzyme loading was 0.7 mL g(-1) glucan based on the six different enzyme loadings tested. SSFs were performed at 37, 41 and 45 °C with an enzyme loading of 0.7 mL g(-1) glucan. The highest ethanol concentration of 22.5 g L(-1) was obtained after 168 h with SSF at 45 °C, which was equivalent to 86% yield. Four different batch and fed-batch strategies were evaluated using a total solid loading of 12% (dry basis). About 32 g L(-1) ethanol was produced with the four strategies, which was equivalent to 82% yield.     

Effect of hydrothermolysis process conditions on pretreated switchgrass composition and ethanol yield by SSF with Kluyveromyces marxianus IMB4

Hot compressed liquid water was used to treat switchgrass in a method called hydrothermolysis to disrupt lignin, dissolve hemicellulose, and increase accessibility of cellulose to cellulase. Three temperatures (190, 200, and 210 °C) and hold times (10, 15, and 20 min) were tested. Switchgrass treated at 190 °C for 10 min had the greatest xylan recovery in the prehydrolyzate. Less than 0.65 g/L glucose were released into the prehydrolyzate for all pretreatment conditions, indicating most glucose was retained as cellulose in the solid substrate. 5-Hydroxymethylfurfural (HMF) and furfural formation in the prehydrolyzate were found to be less than 1 g/L for all treatments. The highest concentration of ethanol, 16.8 g/L (72% of theoretical), was produced from switchgrass pretreated at 210 °C and 15 min using simultaneous saccharification and fermentation (SSF) at 45 °C with the thermotolerant yeast Kluyveromyces marxianus IMB4 and 15 FPU cellulase/g glucan.     

Polygalacturonase production by Kluyveromyces marxianus: effect of medium composition

A strain of Kluyveromyces marxianus (CCT 3172), isolated from a cocoa fermentation in Brazil, secreted an endopolygalacturonase (PG) when grown under self-induced anaerobic conditions; neither polymethylesterase nor pectate lyase appeared in culture filtrates. Replacing glucose in the medium with sucrose had no effect on PG secretion or ethanol production. Growth in fructose-containing medium retarded secretion of PG and ethanol, but had no effect on growth. Growth and ethanol production in media containing galactose resembled those in fructose-containing medium, although PG secretion was lowered. Growth and PG secretion were considerably retarded in xylose-containing medium, and were similarly affected in media containing different concentrations of glucose. Varying the concentration of ammonium sulphate in media had no effect on growth or PG secretion.     

The effect of pulse voltage and capacitance on biosorption of uranium by biomass derived from whiskey distillery spent wash

Biosorption of uranium by residual biomass from The Old Bushmill's Distillery Co. Ltd., Bushmills, Co. Antrim, Northern Ireland, following exposure to short and intense electric pulses has been examined. The biomass was prepared from the distillery spent wash and consisted of non-viable yeast and bacterial cells. As shown previously, untreated biomass had a maximum biosorption capacity of 170?mg uranium/g dry weight biomass. When biosorption reactions were placed between two electrodes and exposed to electric pulses with field strengths ranging from 1.25–3.25?kV/cm at a capacitance of 25?μF, biosorption increased from 170?mg of uranium to 275?mg uranium/g dry weight biomass. The data were obtained from biosorption isotherm analyses and taken as the degree of biosorption at residual uranium concentrations of 3?mM. In addition, when the capacitance of the electric pulses increased from 0.25?μF to 25?μF at a fixed pulse field strength the degree of biosorption increased from 210?mg uranium to 240?mg uranium/g dry weight biomass. The results suggest that application of short and intense electric pulses to biosorption reactions may play an important role in enhancing microbial biosorption of toxic metals/radionuclides from waste water streams.     

Short communication: Ethanol production from cellulose at 45°C using a batch-fed system containing alginate-immobilized Kluyveromyces marxianus IMB3

The thermotolerant yeast, Kluyveromyces marxianus IMB3, was grown in batch culture at 45°C on cellulose-containing media, supplemented with exogenous cellulase activity. At various stages during fermentation, both substrate and enzyme were added in batch mode and fermentation was continued for 220 h. Ethanol production increased to 20 g/l at 200 h, representing 45% of the maximum theoretical yield. In subsequent experiments, the organism was immobilized in calcium alginate beads and these were used in a similar, batch-fed system at 45°C. Again, fermentation was continued for 220 h and ethanol production increased to its maximum, of 28 g/l, within 100 h and this represented in excess of 60% of the maximum theoretical yield.     

Production of ethanol by the thermotolerant yeastKluyveromyces marxianus IMB3 during growth on lactose-containing media

Summary The thermotolerant yeast strain,Kluyveromyces marxianus IMB3 was shown to be capable of growth and ethanol production on lactose containing media at 45°C. On media containing 4% (w/v) lactose, ethanol production increased to 6.0g/l within 50h and this represented 29% of theoretical yield. During growth on lactose containing media the organism was shown to produce a cell-associated β-galactosidase and no significant enzyme could be detected in the extracellular culture filtrate. Addition of β-galactosidase, released fromKluyveromyces marxianus IMB3 cells, to active fermentations, resulted in increasing ethanol production to 53% of theoretical yield at 45°C.