Ethanol production from sweet sorghum juice using very high gravity technology: Effects of carbon and nitrogen supplementations

Ethanol production from sweet sorghum juice by Saccharomyces cerevisiae NP01 was investigated under very high gravity (VHG) fermentation and various carbon adjuncts and nitrogen sources. When sucrose was used as an adjunct, the sweet sorghum juice containing total sugar of 280 g l⁻¹, 3 g yeast extract l⁻¹ and 5 g peptone l⁻¹ gave the maximum ethanol production efficiency with concentration, productivity and yield of 120.68 ± 0.54 g l⁻¹, 2.01 ± 0.01 g l⁻¹ h⁻¹ and 0.51 ± 0.00 g g⁻¹, respectively. When sugarcane molasses was used as an adjunct, the juice under the same conditions gave the maximum ethanol concentration, productivity and yield with the values of 109.34 ± 0.78 g l⁻¹, 1.52 ± 0.01 g l⁻¹ h⁻¹ and 0.45 ± 0.01 g g⁻¹, respectively. In addition, ammonium sulphate was not suitable for use as a nitrogen supplement in the sweet sorghum juice for ethanol production since it caused the reduction in ethanol concentration and yield for approximately 14% when compared to those of the unsupplemented juices.     

Lactic acid production by Lactobacillus sp. RKY2 in a cell-recycle continuous fermentation using lignocellulosic hydrolyzates as inexpensive raw materials

Continuous lactic acid fermentations were conducted using lignocellulosic hydrolyzates and corn steep liquor as inexpensive raw materials. Lactic acid concentrations decreased with increases in the dilution rate, whereas the residual substrate concentrations increased. However, lactic acid yields were maintained at more than 0.90 g g⁻¹ over all cases experimented. The cell-recycle cultivation system exerted positive effects on fermentation efficiency, including volumetric productivity, which is attributable to the retention of cells in the bioreactor. The cell-recycle continuous fermentation of lignocellulosic hydrolyzates yielded a lactic acid productivity of 6.7 g l⁻¹ h⁻¹ for a dilution rate of 0.16 h⁻¹ using 30 g l⁻¹ of corn steep liquor and 1.5 g l⁻¹ of yeast extract as nutrients. The productivity (6.7 g l⁻¹ h⁻¹) acquired by the cell-recycle continuous fermentation of lignocellulosic hydrolyzates was 1.6 times higher than the lactic acid productivity yielded in the continuous fermentation without cell-recycle system.     

Green and economical production of propionic acid by Propionibacterium freudenreichii CCTCC M207015 in plant fibrous-bed bioreactor

Propionic acid production by Propionibacterium freudenreichii from molasses and waste propionibacterium cells was studied in plant fibrous-bed bioreactor (PFB). With non-treated molasses as carbon source, 12.69 ± 0.40 g l^-1 of propionic acid was attained at 120 h in free-cell fermentation, whereas the PFB fermentation yielded 41.22 ± 2.06 g l^-1 at 120 h and faster cells growth was observed. In order to optimize the fermentation outcomes, fed-batch fermentation was performed with hydrolyzed molasses in PFB, giving 91.89 ± 4.59 g l^-1 of propionic acid at 254 h. Further studies were carried out using hydrolyzed waste propionibacterium cells as substitute nitrogen source, resulting in a propionic acid concentration of 79.81 ± 3.99 g l^-1 at 302 h. The present study suggests that the low-cost molasses and waste propionibacterium cells can be utilized for the green and economical production of propionic acid by P. freudenreichii.     

In silico optimization and low structured kinetic model of poly[(R)-3-hydroxybutyrate] synthesis by Cupriavidus necator DSM 545 by fed-batch cultivation on glycerol

Glycerol was utilized by Cupriavidus necator DSM 545 for production of poly-3-hydroxybutyrate (PHB) in fed-batch fermentation. Maximal specific growth rates (0.12 and 0.3 h⁻¹) and maximal specific non-growth PHB production rate (0.16 g g⁻¹ h⁻¹) were determined from two experiments (inocula from exponential and stationary phase). Saturation constants for nitrogen (0.107 and 0.016 g L⁻¹), glycerol (0.05 g L⁻¹), non-growth related PHB synthesis (0.011 g L⁻¹) and nitrogen/PHB related inhibition constant (0.405 g L⁻¹), were estimated. Five relations for specific growth rate were tested using mathematical models. In silico performed optimization procedures (varied glycerol/nitrogen ratio and feeding) has resulted in a PHB content of 70.9%, shorter cultivation time (23 h) and better PHB yield (0.347 g g⁻¹). Initial concentration of biomass 16.8 g L⁻¹ and glycerol concentration in broth between 3 and 5 g L⁻¹ were decisive factors for increasing of productivity.     

Enhanced 2,3-butanediol production by Klebsiella oxytoca using a two-stage agitation speed control strategy

Batch fermentative production of 2,3-butanediol by Klebsiella oxytoca was investigated using various oxygen supply methods though varying agitation speed. Based on the analysis of three kinetic parameters including specific cell growth rate (μ), specific glucose consumption rate (q_s) and specific 2,3-butanediol formation rate (q_p), a two-stage agitation speed control strategy, aimed at achieving high concentration, high yield and high productivity of 2,3-butanediol, was proposed. At the first 15 h, agitation speed was controlled at 300 rpm to obtain high μ for cell growth, subsequently agitation speed was controlled at 200 rpm to maintain high q_p for high 2,3-butanediol accumulation. Finally, the maximum concentration of 2,3-butanediol reached 95.5 g l⁻¹ with the yield of 0.478 g g⁻¹ and the productivity of 1.71 g l⁻¹ h⁻¹, which were 6.23%, 6.22% and 22.14% over the best results controlled by constant agitation speeds.     

Techno-economic implications of improved high gravity corn mash fermentation

The performance of Saccharomyces cerevisiae MBG3964, a strain able to tolerate >18% v/v ethanol, was compared to leading industrial ethanol strain, Fermentis Ethanol Red, under high gravity corn mash fermentation conditions. Compared to the industrial ethanol strain, MBG3964 gave increased alcohol yield (140 g L⁻¹ vs. 126 g L⁻¹), lower residual sugar (4 g L⁻¹ vs. 32 g L⁻¹), and lower glycerol (11 g L⁻¹ vs. 12 g L⁻¹). After 72 h fermentation, MBG3964 showed about 40% viability, whereas the control yeast was only about 3% viable. Based on modelling, the higher ethanol tolerant yeast could increase the profitability of a corn-ethanol plant and help it remain viable through higher production, lower unit heating requirements and extra throughput. A typical 50 M gal y⁻¹ dry mill ethanol plant that sells dried distiller’s grain could potentially increase its profit by nearly $US3.4 M y⁻¹ due solely to the extra yield, and potentially another $US4.1 M y⁻¹ if extra throughput is possible.     

Synergistic production of L-arabinose from arabinan by the combined use of thermostable endo- and exo-arabinanases from Caldicellulosiruptor saccharolyticus

The optimum conditions for the production of l-arabinose from debranched arabinan were determined to be pH 6.5, 75 °C, 20 g l⁻¹ debranched arabinan, 42 U ml⁻¹ endo-1,5-α-l-arabinanase, and 14 U ml⁻¹ α-l-arabinofuranosidase from Caldicellulosiruptor saccharolyticus and the conditions for sugar beet arabinan were pH 6.0, 75 °C, 20 g l⁻¹ sugar beet arabinan, 3 U ml⁻¹ endo-1,5-α-l-arabinanase, and 24 U ml⁻¹ α-l-arabinofuranosidase. Under the optimum conditions, 16 g l⁻¹l-arabinose was obtained from 20 g l⁻¹ debranched arabinan or sugar beet arabinan after 120 min, with a hydrolysis yield of 80% and a productivity of 8 g l⁻¹ h⁻¹. This is the first reported trial for the production of l-arabinose from the hemicellulose arabinan by the combined use of endo- and exo-arabinanases.     

Effect of nutrient limitation and two-stage continuous fermentor design on productivities during "Clostridium ragsdalei" syngas fermentation

The effect of three limiting nutrients, calcium pantothenate, vitamin B₁₂ and cobalt chloride (CoCl₂), on syngas fermentation using “Clostridium ragsdalei” was determined using serum bottle fermentation studies. Significant results from the bottle studies were translated into single- and two-stage continuous fermentor designs. Studies indicated that three-way interactions between the three limiting nutrients, and two-way interactions between vitamin B₁₂ and CoCl₂ had a significant positive effect on ethanol and acetic acid formation. In general, ethanol and acetic acid production ceased at the end of 9 days corresponding to the production of 2.01 and 1.95 g L⁻¹ for the above interactions. Reactor studies indicated the three-way nutrient limitation in two-stage fermentor showed improved acetic acid (17.51 g g⁻¹ cells) and ethanol (14.74 g g⁻¹ cells) yield compared to treatments in single-stage fermentors. These results further support the hypothesis that it is possible to modulate the product formation by limiting key nutrients during C. ragsdalei syngas fermentation.     

Bioethanol fermentation of concentrated rice straw hydrolysate using co-culture of Saccharomyces cerevisiae and Pichia stipitis

Rice straw is one of the abundant lignocellulosic feed stocks in the world and has been selected for producing ethanol at an economically feasible manner. It contains a mixture of sugars (hexoses and pentoses).Biphasic acid hydrolysis was carried out with sulphuric acid using rice straw. After acid hydrolysis, the sugars, furans and phenolics were estimated. The initial concentration of sugar was found to be 16.8 g L⁻¹. However to increase the ethanol yield, the initial sugar concentration of the hydrolysate was concentrated to 31 g L⁻¹ by vacuum distillation. The concentration of sugars, phenols and furans was checked and later detoxified by over liming to use for ethanol fermentation. Ethanol concentration was found to be 12 g L⁻¹, with a yield, volumetric ethanol productivity and fermentation efficiency of 0.33 g L⁻¹ h⁻¹, 0.4 g g⁻¹ and 95%, respectively by co-culture of OVB 11 (Saccharomyces cerevisiae) and Pichia stipitis NCIM 3498.     

Development of chemically defined media supporting high cell density growth of Ketogulonicigenium vulgare and Bacillus megaterium

The immediate precursor of L-ascorbic acid, or vitamin C, is 2-keto-l-gulonic acid (2-KLG). This is commonly produced commercially by Ketogulonicigenium vulgare and Bacillus megaterium, using corn steep liquor powder (CSLP) as an organic nitrogen source. In this study, the effects of the individual CSLP components (amino acids, vitamins, and metal elements) on 2-KLG production were evaluated, with the aim of developing a complete, chemically defined medium for 2-KLG production. Forty components of CSLP were analyzed, and key components were correlated to biomass, 2-KLG productivity, and consumption rate of L-sorbose. Glycine had the greatest effect, followed by serine, biotin, proline, nicotinic acid, and threonine. The combination of 0.28 g L⁻¹ serine, 0.36 g L⁻¹glycine, 0.18 g L⁻¹ threonine, 0.28 g L⁻¹proline, 0.19 g L⁻¹nicotinic acid, and 0.62 mg L⁻¹biotin in a chemically defined medium produced the highest maximum biomass concentration (4.2 × 10⁹ cfu mL⁻¹), 2-KLG concentration (58 g L⁻¹), and yield (0.76 g g⁻¹) after culturing for 28 h.     

3-Phenyllactic acid production by substrate feeding and pH-control in fed-batch fermentation of Lactobacillus sp. SK007

3-Phenyllactic acid (PLA), which is produced by some strains of lactic acid bacteria (LAB), is a known antimicrobial agent with a broad spectrum. Batch and fed-batch fermentation by the strain Lactobacillus sp. SK007 for PLA production have been reported. With batch fermentation without pH-control, PLA production yield was 2.42 g L⁻¹. When fed-batch fermentation by Lactobacillus sp. SK007 was conducted in 3 L initial volume with pH-control at 6.0 and intermittent feeding, which was developed after fermentation for 12 h and every 2 h with 120 mL 100 g L⁻¹ PPA phenylpyruvic acid (PPA) and 50 mL 500 g L⁻¹ glucose each time, PLA production yield reached 17.38 g L⁻¹. The final conversion ratio of PPA to PLA was 51.1%, and the PLA production rate was 0.241 g L⁻¹ h⁻¹. This indicated that PPA was the ideal substrate for PLA fermentation production, and fed-batch fermentation with intermittent PPA feeding and pH-control was an effective approach to improve PLA production yield.     

Cultivation of microalgae for oil production with a cultivation strategy of urea limitation

The microalgae, Chlorella sp., were cultivated in various culture modes to assess biomass and lipid productivity in this study. In the batch mode, the biomass concentrations and lipid content of Chlorella sp. cultivated in a medium containing 0.025–0.200 g L⁻¹ urea were 0.464–2.027 g L⁻¹ and 0.661–0.326 g g⁻¹, respectively. The maximum lipid productivity of 0.124 g d⁻¹ L⁻¹ occurred in a medium containing 0.100 g L⁻¹ urea. In the fed-batch cultivation, the highest lipid content was obtained by feeding 0.025 g L⁻¹ of urea during the stationary phase, but the lipid productivity was not significantly increased. However, a semi-continuous process was carried out by harvesting the culture and renewing urea at 0.025 g L⁻¹ each time when the cultivation achieved the early stationary phase. The maximum lipid productivity of 0.139 g d⁻¹ L⁻¹ in the semi-continuous culture was highest in comparison with those in the batch and fed-batch cultivations.     

Assessment of olive-mill wastewater as a growth medium for lipase production by Candida cylindracea in bench-top reactor

Olive-mill wastewater (OMW) was investigated for its suitability to serve as a medium for lipase production by Candida cylindracea NRRL Y-17506. The OMW that best supported enzyme production was characterized by low COD and low total sugars content. In shake flask batch cultures, OMW supplementation with 2.4 g l⁻¹ NH₄Cl and 3 g l⁻¹ olive oil led to an enzyme activity of about 10 U ml⁻¹. The addition of glucose or malt extract and supplements containing organic N (e.g., peptone, yeast extract) either depressed or did not affect the enzyme production. Further experiments were then performed in a 3-l stirred tank reactor to assess the impact of medium pH and stirring speed on the yeast enzyme activity. The lipase activity was low (1.8 U ml⁻¹) when the pH was held constant at 6.5, significantly increased (18.7 U ml⁻¹) with uncontrolled pH and was maximum (20.4 U ml⁻¹) when the pH was let free to vary below 6.5. A stirring regime, that varied depending on the dissolved oxygen concentration in the medium, both prevented the occurrence of anoxic conditions during the exponential growth phase and enabled good lipase production (i.e., 21.6 U ml⁻¹) and mean volumetric productivity (i.e., 123.5 U l⁻¹ h⁻¹).     

Anaerobic degradation of tetrachlorobisphenol-A in river sediment

This study investigated the anaerobic degradation of tetrachlorobisphenol-A (TCBPA) in sediment samples collected at three sites along the Erren River in southern Taiwan. TCBPA anaerobic degradation half-lives (t_1/2) in the sediment were 12.6, 16.9 and 21.7 d at concentrations of 50, 100, and 250 ??g g⁻¹, respectively. TCBPA (50 ??g g⁻¹) anaerobic degradation half-lives (t_1/2) in the sediment were 10.1, 11.8, 11.0, 11.6, 10.8, 9.1, 8.5, 18.2, 19.3, and 16.1 d by the addition of yeast extract (5 mg l⁻¹), cellulose (0.96 mg l⁻¹), sodium chloride (1%), brij 30 (130 mg l⁻¹), brij 35 (43 mg l⁻¹), rhamnolipid (55 ??M), surfactin (91 ??M), phthalic esters (2 mg l⁻¹), nonylphenol (2 mg l⁻¹), and heavy metals (2 mg l⁻¹), respectively. The degradation rate of TCBPA was enhanced by the addition of yeast extract, cellulose, sodium chloride, brij 30, brij 35, rhamnolipid, or surfactin. However, it was inhibited by the addition of phthalic esters, nonylphenol, or heavy metals. Also noted was the presence of dichlorobisphenol-A and bisphenol-A, two intermediate products resulting from the anaerobic degradation of TCBPA accumulated in the sediments.     

Bicarbonate use in three aquatic plants

Our study aimed to test the ability of aquatic plants to use bicarbonate when acclimated to three different bicarbonate concentrations. To this end, we performed experiments with the three species Ceratophyllum demersum, Egeria densa, Lagarosiphon major to determine photosynthetic rates under varying bicarbonate concentrations. We measured bicarbonate use efficiency, photosynthetic performance and respiration. For all species, our results revealed that photosynthetic rates were highest in replicates grown at low alkalinity. Thus, E. densa had approx. five times higher rates at low (264 ± 15 μmol O₂ g⁻¹ DW h⁻¹) than at high alkalinity (50 ± 27 μmol O₂ g⁻¹ DW h⁻¹), C. demersum had three times higher rates (336 ± 95 and 120 ± 31 μmol O₂ g⁻¹ DW h⁻¹), and L. major doubled its rates at low alkalinity (634 ± 114 and 322 ± 119 μmol O₂ g⁻¹ DW h⁻¹). Similar results were obtained for bicarbonate use efficiency by E. densa (136 ± 44 and 43 ± 10 μmol O₂ mequiv. L⁻¹ g⁻¹ DW h⁻¹) and L. major (244 ± 29 and 82 ± 24 μmol O₂ mequiv. L⁻¹ g⁻¹ DW h⁻¹). As to C. demersum, efficiency was high but unaffected by alkalinity, indicating high adaptation ability to varied alkalinities. A pH drift experiment supported these results. Overall, our results suggest that the three globally widespread worldwide species of our study adapt to low inorganic carbon availability by increasing their efficiency of bicarbonate use.     

Harvesting biohydrogen from cellobiose from sulfide or nitrite-containing wastewaters using Clostridium sp. R1

Harvesting biohydrogen from inhibiting wastewaters is of practical interest since the toxicity of compounds in a wastewater stream commonly prevents the bioenergy content being recovered. The isolated Clostridium sp. R1 is utilized to degrade cellobiose in sulfide or nitrite-containing medium for biohydrogen production. The strain can effectively degrade cellobiose free of severe inhibitory effects at up to 200 mg l⁻¹ sulfide or to 5 mg l⁻¹ nitrite, yielding hydrogen at >2.0 mol H₂ mol⁻¹ cellobiose. Principal metabolites of cellobiose fermentation are acetate and butyrate, with the concentration of the former increases with increasing sulfide and nitrite concentrations. The isolated strain can yield hydrogen from cellobiose in sulfide-laden wastewaters. However, the present of nitrite significantly limit the efficiency of the biohydrogen harvesting process.     

Enhanced ethanol production by fermentation of rice straw hydrolysate without detoxification using a newly adapted strain of Pichia stipitis

An enhanced inhibitor-tolerant strain of Pichia stipitis was successfully developed through adaptation to acid-treated rice straw hydrolysate. The ethanol production obtained by fermentation of NaOH-neutralized hydrolysate without detoxification using the adapted P. stipitis was comparable to fermentation of overliming-detoxified hydrolysate. The ethanol yield using the adapted P. stipitis with both types of hydrolysate at pH 5.0 achieved 0.45 g_p g_s⁻¹, which is equivalent to 87% of the maximum possible ethanol conversion. Furthermore, the newly adapted P. stipitis demonstrated significantly enhanced tolerance to sulfate and furfural despite the fact that both inhibitors had not been removed from the hydrolysate by NaOH neutralization. Finally, the ethanol conversion could be maintained at 60% and above when the neutralized hydrolysate contained 3.0% sulfate and 1.3 g L⁻¹ furfural.     

Ethanol production from syngas by Clostridium strain P11 using corn steep liquor as a nutrient replacement to yeast extract

The feasibility of replacing yeast extract (YE) by corn steep liquor (CSL), a low cost nutrient source, for syngas fermentation to produce ethanol using Clostridium strain P11 was investigated. About 32% more ethanol (1.7 g L⁻¹) was produced with 20 g L⁻¹ CSL media in 250-mL bottle fermentations compared to media with 1 g L⁻¹ YE after 360 h. Maximum ethanol concentrations after 360 h of fermentation in a 7.5-L fermentor with 10 and 20 g L⁻¹ CSL media were 8.6 and 9.6 g L⁻¹, respectively, which represent 57% and 60% of the theoretical ethanol yields from CO. Only about 6.1 g L⁻¹ of ethanol was obtained in the medium with 1 g L⁻¹ YE after 360 h, which represents 53% of the theoretical ethanol yield from CO. The use of CSL also enhanced butanol production by sevenfold compared to YE in bottle fermentations. These results demonstrate that CSL can replace YE as the primary medium component and significantly enhance ethanol production by Clostridium strain P11.     

Effect of lignocellulosic inhibitory compounds on growth and ethanol fermentation of newly-isolated thermotolerant Issatchenkia orientalis

A newly isolated thermotolerant ethanologenic yeast strain, Issatchenkia orientalis IPE 100, was able to produce ethanol with a theoretical yield of 85% per g of glucose at 42 °C. Ethanol production was inhibited by furfural, hydroxymethylfurfural and vanillin concentrations above 5.56 g L⁻¹, 7.81 g L⁻¹, and 3.17 g L⁻¹, respectively, but the strain was able to produce ethanol from enzymatically hydrolyzed steam-exploded cornstalk with 93.8% of theoretical yield and 0.91 g L⁻¹ h⁻¹ of productivity at 42 °C. Therefore, I. orientalis IPE 100 is a potential candidate for commercial lignocelluloses-to-ethanol production.     

Inter-population variability of leaf morpho-anatomical and terpenoid patterns of Pistacia atlantica Desf. ssp. atlantica growing along an aridity gradient in Algeria

Three Algerian populations of female Pistacia atlantica shrubs were investigated in order to check whether their terpenoid contents and morpho-anatomical parameters may characterize the infraspecific variability. The populations were sampled along a gradient of increasing aridity from the Atlas mountains into the northwestern Central Sahara.As evidenced by Scanning Electron Microscopy, tufted hairs could be found only on seedling leaves from the low aridity site as a population-specific trait preserved also in culture. Under common garden cultivation seedlings of the high aridity site showed a three times higher density of glandular trichomes compared to the low aridity site. Increased aridity resulted also in reduction of leaf sizes while their thickness increased. Palisade parenchyma thickness also increases with aridity, being the best variable that discriminates the three populations of P. atlantica.Analysis of terpenoids from the leaves carried out by GC-MS reveals the presence of 65 compounds. The major compounds identified were spathulenol (23 μg g⁻¹ dw), α-pinene (10 μg g⁻¹ dw), verbenone (7 μg g⁻¹ dw) and β-pinene (6 μg g⁻¹ dw) in leaves from the low aridity site; spathulenol (73 μg g⁻¹ dw), α-pinene (25 μg g⁻¹ dw), β-pinene (18 μg g⁻¹ dw) and γ-amorphene (16 μg g⁻¹ dw) in those from medium aridity and spathulenol (114 μg g⁻¹ dw), α-pinene (49 μg g⁻¹ dw), germacrene D (29 μg g⁻¹ dw) and camphene (23 μg g⁻¹ dw) in leaves from the high aridity site. Terpene concentrations increased with the degree of aridity: the highest mean concentration of monoterpenes (136 μg g⁻¹ dw), sesquiterpenes (290 μg g⁻¹ dw) and total terpenes (427 μg g⁻¹ dw) were observed in the highest arid site and are, respectively, 3-, 5- and 4-fold higher compared to the lower arid site. Spathulenol and α-pinene can be taken as chemical markers of aridity. Drought discriminating compounds in low, but detectable concentrations are δ-cadinene and β-copaene. The functional roles of the terpenoids found in P. atlantica leaves and principles of their biosynthesis are discussed with emphasis on the mechanisms of plant resistance to drought conditions.