Eucalyptus hydrolysate detoxification with activated charcoal adsorption or ion-exchange resins for xylitol production

Eucalyptus hemicellulosic hydrolysate used for xylitol production by Candida guilliermondii FTI20037 was previously treated either with ion-exchange resins or with activated charcoal adsorption combined with pH adjustment, in order that acetic acid, furfural and hydroxymethylfurfural could be removed. The best results for xylitol yield factor (0.76 g/g) and volumetric productivity (0.68 g/(l h) were attained when a three-fold concentrated hydrolysate was treated with ion-exchange resins. Using activated charcoal combined with pH adjustment for treating a three-fold concentrated hydrolysate resulted in a xylitol yield factor of 0.40 g/g and a volumetric productivity of 0.30 g/(l h). This same treatment applied to a six-fold concentrated hydrolysate resulted in a xylitol yield factor of 0.66 g/g and a volumetric productivity of 0.50 g/(l h).     

Overcoming inhibitors in a hemicellulosic hydrolysate: improving fermentability by feedstock detoxification and adaptation of <Emphasis Type="Italic">Pichia stipitis</Emphasis>

In order to improve the fermentative efficiency of sugar maple hemicellulosic hydrolysates for fuel ethanol production, various methods to mitigate the effects of inhibitory compounds were employed. These methods included detoxification treatments utilizing activated charcoal, anion exchange resin, overliming, and ethyl acetate extraction. Results demonstrated the greatest fermentative improvement of 50% wood hydrolysate (v/v) by Pichia stipitis with activated charcoal treatment. Another method employed to reduce inhibition was an adaptation procedure to produce P. stipitis stains more tolerant of inhibitory compounds. This adaptation resulted in yeast variants capable of improved fermentation of 75% untreated wood hydrolysate (v/v), one of which produced 9.8 g/l ± 0.6 ethanol, whereas the parent strain produced 0.0 g/l ± 0.0 within the first 24 h. Adapted strains RS01, RS02, and RS03 were analyzed for glucose and xylose utilization and results demonstrated increased glucose and decreased xylose utilization rates in comparison to the wild type. These changes in carbohydrate utilization may be indicative of detoxification or tolerance activities related to proteins involved in glucose and xylose metabolism.     

Detoxification of dilute acid hydrolysates of lignocellulose with lime

The hydrolysis of hemicellulose to monomeric sugars by dilute acid hydrolysis is accompanied by the production of inhibitors that retard microbial fermentation. Treatment of hot hydrolysate with Ca(OH)(2) (overliming) is an effective method for detoxification. Using ethanologenic Escherichia coli LY01 as the biocatalyst, our results indicate that the optimal lime addition for detoxification varies and depends on the concentration of mineral acids and organic acids in each hydrolysate. This optimum was shown to be readily predicted on the basis of the titration of hydrolysate with 2 N NaOH at ambient temperature to either pH 7.0 or pH 11.0. The average composition of 15 hydrolysates prior to treatment was as follows (per L): 95.24 +/- 7.29 g sugar, 5.3 +/- 2.99 g acetic acid, 1.305 +/- 0.288 g total furans (furfural and hydroxymethylfurfural), and 2.86 +/- 0.34 g phenolic compounds. Optimal overliming resulted in a 51 +/- 9% reduction of total furans, a 41 +/- 6% reduction in phenolic compounds, and a 8.7 +/- 4.5% decline in sugar. Acetic acid levels were unchanged. Considering the similarity of microorganisms, it is possible that the titration method described here may also prove useful for detoxification and fermentation processes using other microbial biocatalysts.     

Fungal delignification of lignocellulosic biomass improves the saccharification of cellulosics

The biological delignification of lignocellulosic feedstocks, Prosopis juliflora and Lantana camara was carried out with Pycnoporus cinnabarinus, a white rot fungus, at different scales under solid-state fermentation (SSF) and the fungal treated substrates were evaluated for their acid and enzymatic saccharification. The fungal fermentation at 10.0 g substrate level optimally delignified the P. juliflora by 11.89% and L. camara by 8.36%, and enriched their holocellulose content by 3.32 and 4.87%, respectively, after 15 days. The fungal delignification when scaled up from 10.0 g to 75.0, 200.0 and 500.0 g substrate level, the fungus degraded about 7.69–10.08% lignin in P. juliflora and 6.89–7.31% in L. camara, and eventually enhanced the holocellulose content by 2.90–3.97 and 4.25–4.61%, respectively. Furthermore, when the fungal fermented L. camara and P. juliflora was hydrolysed with dilute sulphuric acid, the sugar release was increased by 21.4-42.4% and the phenolics content in hydrolysate was decreased by 18.46 and 19.88%, as compared to the unfermented substrate acid hydrolysis, respectively. The reduction of phenolics in acid hydrolysates of fungal treated substrates decreased the amount of detoxifying material (activated charcoal) by 25.0–33.0% as compared to the amount required to reduce almost the same level of phenolics from unfermented substrate hydrolysates. Moreover, an increment of 21.1–25.1% sugar release was obtained when fungal treated substrates were enzymatically hydrolysed as compared to the hydrolysis of unfermented substrates. This study clearly shows that fungal delignification holds potential in utilizing plant residues for the production of sugars and biofuels.     

Ethanol production from rice hull using Pichia stipitis and optimization of acid pretreatment and detoxification processes

The goal of this study was to produce ethanol from rice hull hydrolysates (RHHs) using Pichia stipitis strains and to optimize dilute acid hydrolysis and detoxification processes by response surface methodology (RSM). The optimized conditions were found as 127.14°C, solid:liquid ratio of 1:10.44 (w/v), acid ratio of 2.52% (w/v), and hydrolysis time of 22.01 min. At these conditions, the fermentable sugar concentration was 21.87 g/L. Additionally, the nondetoxified RHH at optimized conditions contained 865.2 mg/L phenolics, 24.06 g/L fermentable sugar, no hydroxymethylfurfural (HMF), 1.62 g/L acetate, 0.36 g/L lactate, 1.89 g/L glucose, and 13.49 g/L fructose + xylose. Furthermore, RHH was detoxified with various methods and the best procedures were found to be neutralization with CaO or charcoal treatment in terms of the reduction of inhibitory compounds as compared to nondetoxified RHH. After detoxification procedures, the content of hydrolysates consisted of 557.2 and 203.1 mg/L phenolics, 19.7 and 21.60 g/L fermentable sugar, no HMF, 0.98 and 1.39 g/L acetate, 0 and 0.04 g/L lactate, 1.13 and 1.03 g/L glucose, and 8.46 and 12.09 g/L fructose + xylose, respectively. Moreover, the base‐line mediums (control), and nondetoxified and detoxified hydrolysates were used to produce ethanol by using P. stipitis strains. The highest yields except that of base‐line mediums were achieved using neutralization (35.69 and 38.33% by P. stipitis ATCC 58784 and ATCC 58785, respectively) and charcoal (37.55% by P. stipitis ATCC 58785) detoxification methods. Results showed that the rice hull can be utilized as a good feedstock for ethanol production using P. stipitis. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:872–882, 2016     

Bioconversion of Saccharum spontaneum (wild sugarcane) hemicellulosic hydrolysate into ethanol by mono and co-cultures of Pichia stipitis NCIM3498 and thermotolerant Saccharomyces cerevisiae-VS₃

The lignocellulosic biomass is a low-cost renewable resource for eco-benign liquid fuel 'ethanol'. To resolve the hydrolysis of mixed sugars in lignocellulosic substrate Saccharum spontaneum, the microbial co-cultures of Pichia stipitis NCIM 3498 and thermotolerant Saccharomyces cerevisiae-VS(3) were analyzed for efficient bioconversion of mixed sugars into ethanol. Among the hydrolysis conditions, the acid hydrolysis released maximum sugars along with furans, phenolics and acetic acid. The acidic hydrolysate was detoxified and fermented by monocultures of P. stipitis NCIM3498 (P.S.) and thermotolerant S. cerevisiae VS(3) (S.C.), and co-culture of P.S. (7.5 mL) and S.C. (2.5 mL). Before the fermentation of hemicellulose acid hydrolysate, both the monocultures (P.S. and S.C.), and varying ratios of P.S. and S.C. microorganisms in co-cultures #1, #2 and #3 were grown on simulated synthetic medium. The ethanol yield from monocultures of P.S. (0.44 ± 0.021 g/g), S.C. (0.22 ± 0.01 g/g) and co-culture #3 (0.49 ± 0.02 g/g) revealed unique characteristics of each mono and co-culture technology. The fermentation of hemicellulose acid hydrolysate with monocultures of P.S., S.C. and co-culture #3 produced 12.08 ± 0.72 g/L, 1.40 ± 0.07 g/L, and 15.0 ± -0.92 g/L ethanol, respectively.     

Bioconversion of water-hyacinth (Eichhornia crassipes) hemicellulose acid hydrolysate to motor fuel ethanol by xylose-fermenting yeast

Water-hyacinth (Eichhornia crassipes) hemicellulose acid hydrolysate has been utilized as a substrate for ethanol production using Pichia stipitis NRRL Y-7124. Hydrolysate fermentability was considerable improved by boiling, and overliming up to pH 10.0 with solid Ca(OH)(2) in combination with sodium sulfite. The percent total sugar utilized and ethanol yield (Y(p/s)) for the untreated hydrolysate were 20.15+/-0.17% and 0.19+/-0.003 g(p) g(s)(-1), respectively, compared with 76.0+/-0.32% and 0.35 g(p) g(s)(-1), respectively for the treated material. The fermentation was very effective at an aeration rate of 0.02 v/v/m, temperature 30+/-0.2 degrees C and pH 6.0+/-0.2. However, the volumetric productivity (Q(p)) was still considerably less than observed in a simulated synthetic hydrolysate medium with a sugar composition similar to the hemicellulose acid hydrolysate. L-Arabinose was not fermented but assimilated. The presence of acetic acid in the hydrolysate decreased the ethanol yield and productivity considerably.     

Effects of sulfuric acid loading and residence time on the composition of sugarcane bagasse hydrolysate and its use as a source of xylose for xylitol bioproduction

A 2(2) full factorial design was employed to evaluate the effects of sulfuric acid loading and residence time on the composition of sugarcane bagasse hydrolysate obtained in a 250-L reactor. The acid loading and the residence time were varied from 70 to 130 mg acid per gram of dry bagasse and from 10 to 30 min, respectively, while the temperature (121 degrees C) and the bagasse loading (10%) were kept constant. Both the sulfuric acid loading and the residence time influenced the concentrations of xylose and inhibitors in the hydrolysate. The highest xylose concentration (22.71 g/L) was achieved when using an acid loading of 130 mg/g and a residence time of 30 min. These conditions also led to increased concentrations of inhibiting byproducts in the hydrolysate. All of the hydrolysates were vacuum-concentrated to increase the xylose concentration, detoxified by pH alteration and adsorption into activated charcoal, and used for xylitol bioproduction in a stirred tank reactor. Neither the least (70 mg/g, 10 min) nor the most severe (130 mg/g, 30 min) hydrolysis conditions led to the best xylitol production (37.5 g/L), productivity (0.85 g/L h), and yield (0.78 g/g).     

Influence of the toxic compounds present in brewer's spent grain hemicellulosic hydrolysate on xylose-to-xylitol bioconversion by Candida guilliermondii

Fermentation media containing different concentrations of toxic compounds were prepared from brewer's spent grain (BSG) hemicellulosic hydrolysate, and used for xylose-to-xylitol bioconversion by Candida guilliermondii. Such fermentation media were composed of the hydrolysate in the following ways: raw (RH); concentrated four-fold (CH); concentrated and treated with activated charcoal (TCH); raw supplemented with sugars until a concentration four-fold higher (SRH); concentrated and subsequently diluted but supplemented with sugars until a concentration four-fold higher (SDCH). All media presented an initial xylose concentration of 85 g/l, except RH, which contained 23 g/l xylose. Fermentation results revealed that the sugars supplementation to raw hydrolysate favored the xylitol production. Nevertheless, xylitol production from CH was negatively affected due to the high concentration of toxic compounds present in the medium. The hydrolysate treatment with activated charcoal partially removed the toxic compounds, and the xylitol production was higher than in CH, but not so efficient as in SRH. It was thus concluded that to obtain an efficient xylose-to-xylitol bioconversion from BSG hydrolysate, the sugars concentration must be increased, but the toxic compounds concentration must be reduced to the same level present in the raw hydrolysate.     

Microbial utilization of levoglucosan in wood pyrolysate as a carbon and energy source

The Waterloo Fast Pyrolysis Process (WFPP) can produce an organic liquid high in levoglucosan (1, 6-anhydro-beta-D-glucopyranose) content from suitably pretreated lignocellulosics. A variety of fungi and yeasts were screened for their ability to utilize and ferment this organic liquid. To enhance its fermentability, the pyrolysis tar was posttreated in three different ways: (1) an aqueous extract (lignin removed); (2) activated charcoal treated (lignin and aromatics removed); and (3) acid hydrolysate (lignin and aromatics removed with the levoglucosan hydrolyzed to glucose). Four fungal strains were examined. None grew in the aqueous extract, but all grew equally well in both the activated charcoal treated and the acid hydrolysate, suggesting that the aromatic species were inhibitory to growth. Seven yeast species were examined, two of which did not grow on any of the extracts. Five of the yeast strains grew well on both the aqueous extract as well as the activated charcoal extract. The hydrolysate was optimal in terms of biomass yield and ethanol production. Ethanol yields on the hydrolysate were comparable or better than those on glucose. Ethanol was also produced in the aqueous extract and activated charcoal-treated substrate, but yields were considerably lower than on the hydrolysate or glucose. It is apparent that a wood pyrolysate maximized for levoglucosan can serve as a fermentable substrate, although postpyrolysis clean-up appears necessary. (c) 1993 John Wiley & Sons, Inc.     

活性炭对玉米芯半纤维素稀硫酸水解液的脱毒效果

用稀硫酸对玉米芯半纤维素进行水解是一种常用的方法,但是玉米芯半纤维素在水解成木糖等还原糖的同时还产生了糠醛、乙酸和酚类等抑制水解液发酵的毒物。以混合脱毒法为基础,研究活性炭在脱毒过程中的作用。结果表明,有脱毒效果的活性炭种类是GH-13和GH-15,随着活性炭添加量的增大,脱毒效果增强,但木糖损失也随之增多。其中采用5%GH-15时的脱毒效果最佳,该条件下乙酸去除率为24.60%,糠醛去除率达100%,酚类化合物去除效率R280值0.009,而木糖的损失率为23.70%。     

Fermentation of lignocellulosic sugars to acetic acid by <Emphasis Type="Italic">Moorella thermoacetica</Emphasis>

A systematic study of bioconversion of lignocellulosic sugars to acetic acid by Moorella thermoacetica (strain ATCC 39073) was conducted. Four different water-soluble fractions (hydrolysates) obtained after steam pretreatment of lignocellulosic biomass were selected and fermented to acetic acid in batch fermentations. M. thermoacetica can effectively ferment xylose and glucose in hydrolysates from wheat straw, forest residues, switchgrass, and sugarcane straw to acetic acid. Xylose and glucose were completely utilized, with xylose being consumed first. M. thermoacetica consumed up to 62 % of arabinose, 49 % galactose and 66 % of mannose within 72 h of fermentation in the mixture of lignocellulosic sugars. The highest acetic acid yield was obtained from sugarcane straw hydrolysate, with 71 % of theoretical yield based on total sugars (17 g/L acetic acid from 24 g/L total sugars). The lowest acetic acid yield was observed in forest residues hydrolysate, with 39 % of theoretical yield based on total sugars (18 g/L acetic acid from 49 g/L total sugars). Process derived compounds from steam explosion pretreatment, including 5-hydroxymethylfurfural (0.4 g/L), furfural (0.1 g/L) and total phenolics (3 g/L), did not inhibit microbial growth and acetic acid production yield. This research identified two major factors that adversely affected acetic acid yield in all hydrolysates, especially in forest residues: (i) glucose to xylose ratio and (ii) incomplete consumption of arabinose, galactose and mannose. For efficient bioconversion of lignocellulosic sugars to acetic acid, it is imperative to have an appropriate balance of sugars in a hydrolysate. Hence, the choice of lignocellulosic biomass and steam pretreatment design are fundamental steps for the industrial application of this process.     

Soluble and insoluble solids contributions to high-solids enzymatic hydrolysis of lignocellulose

The rates and extents of enzymatic cellulose hydrolysis of dilute acid pretreated corn stover (PCS) decline with increasing slurry concentration. However, mass transfer limitations are not apparent until insoluble solids concentrations approach 20% w/w, indicating that inhibition of enzyme hydrolysis at lower solids concentrations is primarily due to soluble components. Consequently, the inhibitory effects of pH-adjusted pretreatment liquor on the enzymatic hydrolysis of PCS were investigated. A response surface methodology (RSM) was applied to empirically model how hydrolysis performance varied as a function of enzyme loading (12-40mg protein/g cellulose) and insoluble solids concentration (5-13%) in full-slurry hydrolyzates. Factorial design and analysis of variance (ANOVA) were also used to assess the contribution of the major classes of soluble components (acetic acid, phenolics, furans, sugars) to total inhibition. High sugar concentrations (130g/L total initial background sugars) were shown to be the primary cause of performance inhibition, with acetic acid (15g/L) only slightly inhibiting enzymatic hydrolysis and phenolic compounds (9g/L total including vanillin, syringaldehyde, and 4-hydroxycinnamic acid) and furans (8g/L total of furfural and hydroxymethylfurfural, HMF) with only a minor effect on reaction kinetics. It was also demonstrated that this enzyme inhibition in high-solids PCS slurries can be approximated using a synthetic hydrolyzate composed of pure sugars supplemented with a mixture of acetic acid, furans, and phenolic compounds, which indicates that generally all of the reaction rate-determining soluble compounds for this system can be approximated synthetically.     

Microbial oil production from corncob acid hydrolysate by <Emphasis Type="Italic">Trichosporon cutaneum</Emphasis>

Corncob was treated by dilute H₂SO₄. The hydrolysate contained 45.7 g sugar/l. Without concentration or adding other nutrients, the hydrolysate, after being detoxified by overliming and adsorption with activated charcoal, was used for oil production using Trichosporon cutaneum. After 8 days’ growth in shake-flasks, the biomass was 22.1 g/l with a lipid content of 36%. The lipid yield per mass of sugar was 17.4% (w/w). Corncob thus is a promising raw material for microbial oil production by this yeast.     

Production of polyhydroxyalkanoates by <Emphasis Type="Italic">Burkholderia cepacia</Emphasis> ATCC 17759 using a detoxified sugar maple hemicellulosic hydrolysate

Sugar maple hemicellulosic hydrolysate containing 71.9 g/l of xylose was used as an inexpensive feedstock to produce polyhydroxyalkanoates (PHAs) by Burkholderia cepacia ATCC 17759. Several inhibitory compounds present in wood hydrolysate were analyzed for effects on cell growth and PHA production with strong inhibition observed at concentrations of 1 g/l furfural, 2 g/l vanillin, 7 g/l levulinic acid, and 1 M acetic acid. Gradual catabolism of lower concentrations of these inhibitors was observed in this study. To increase the fermentability of wood hydrolysate, several detoxification methods were tested. Overliming combined with low-temperature sterilization resulted in the highest removal of total inhibitory phenolics (65%). A fed-batch fermentation exhibited maximum PHA production after 96 h (8.72 g PHA/L broth and 51.4% of dry cell weight). Compositional analysis by NMR and physical–chemical characterization showed that PHA produced from wood hydrolysate was composed of polyhydroxybutyrate (PHB) with a molecular mass (M _N) of 450.8 kDa, a melting temperature (T _m) of 174.4°C, a glass transition temperature (T _g) of 7.31°C, and a decomposition temperature (T _decomp) of 268.6°C.     

Separate hydrolysis and fermentation (SHF) of Prosopis juliflora, a woody substrate, for the production of cellulosic ethanol by Saccharomyces cerevisiae and Pichia stipitis-NCIM 3498

Prosopis juliflora (Mesquite) is a raw material for long-term sustainable production of cellulosics ethanol. In this study, we used acid pretreatment, delignification and enzymatic hydrolysis to evaluate the pretreatment to produce more sugar, to be fermented to ethanol. Dilute H(2)SO(4) (3.0%,v/v) treatment resulted in hydrolysis of hemicelluloses from lignocellulosic complex to pentose sugars along with other byproducts such as furfural, hydroxymethyl furfural (HMF), phenolics and acetic acid. The acid pretreated substrate was delignified to the extent of 93.2% by the combined action of sodium sulphite (5.0%,w/v) and sodium chlorite (3.0%,w/v). The remaining cellulosic residue was enzymatically hydrolyzed in 0.05 M citrate phosphate buffer (pH 5.0) using 3.0 U of filter paper cellulase (FPase) and 9.0 U of beta-glucosidase per mL of citrate phosphate buffer. The maximum enzymatic saccharification of cellulosic material (82.8%) was achieved after 28 h incubation at 50 degrees C. The fermentation of both acid and enzymatic hydrolysates, containing 18.24 g/L and 37.47 g/L sugars, with Pichia stipitis and Saccharomyces cerevisiae produced 7.13 g/L and 18.52 g/L of ethanol with corresponding yield of 0.39 g/g and 0.49 g/g, respectively.     

Xylitol production from corn fiber and sugarcane bagasse hydrolysates by Candida tropicalis

A natural isolate, Candida tropicalis was tested for xylitol production from corn fiber and sugarcane bagasse hydrolysates. Fermentation of corn fiber and sugarcane bagasse hydrolysate showed xylose uptake and xylitol production, though these were very low, even after hydrolysate neutralization and treatments with activated charcoal and ion exchange resins. Initial xylitol production was found to be 0.43 g/g and 0.45 g/g of xylose utilised with corn fiber and sugarcane bagasse hydrolysate respectively. One of the critical factors for low xylitol production was the presence of inhibitors in these hydrolysates. To simulate influence of hemicellulosic sugar composition on xylitol yield, three different combinations of mixed sugar control experiments, without the presence of any inhibitors, have been performed and the strain produced 0.63 g/g, 0.68 g/g and 0.72 g/g of xylose respectively. To improve yeast growth and xylitol production with these hydrolysates, which contain inhibitors, the cells were adapted by sub culturing in the hydrolysate containing medium for 25 cycles. After adaptation the organism produced more xylitol 0.58 g/g and 0.65 g/g of xylose with corn fiber hydrolysate and sugarcane bagasse hydrolysate respectively.     

ABE production from yellow poplar through alkaline pre-hydrolysis, enzymatic saccharification, and fermentation

ABE (acetone-butanol-ethanol) was produced through alkaline pre-hydrolysis, enzymatic saccharification, and fermentation using yellow poplar as a raw material. In alkaline pre-hydrolysis, 51.1% of the biomass remained as a residue. In the main woody components, the degrees of lignin and xylan removal were 94.3 and 62.0%, respectively. A yield of 80.9% for cellulose-to-glucose and 81.2% for xylan-to-xylose were obtained by enzymatic hydrolysis. The sugar composition of enzymatic hydrolysate was 95.1 g/L of glucose and 21.4 g/L of xylose. The enzymatic hydrolysate also contained 0.5 g/L of acetic acid and 0.5 g/L of total phenolics. Furfural and 5-hydroxymethylfurfural (5-HMF) were not detected in this hydrolysate. The yellow poplar hydrolysate (YPH) from enzymatic saccharification was used for the production of ABE using Clostridium acetobutylicum and C. beijerinckii. In YPH fermentation, C. acetobutylicum produced 18.1 g/L total ABE (productivity 0.38 g/L h, and yield 0.42), and C. beijerinckii produced 12.1 g/L (productivity 0.25 g/L h, and yield 0.37). Although the ABE productivity by C. beijerinckii was slightly low, the general performance of ABE fermentation in YPH was similar to or higher than those reported previously. Therefore, alkaline pre-hydrolysis could be a very effective pretreatment step prior to enzymatic hydrolysis.     

Biohydrogen production from wheat straw hydrolysate by dark fermentation using extreme thermophilic mixed culture

Hydrolysate was tested as substrate for hydrogen production by extreme thermophilic mixed culture (70°C) in both batch and continuously fed reactors. Hydrogen was produced at hydrolysate concentrations up to 25% (v/v), while no hydrogen was produced at hydrolysate concentration of 30% (v/v), indicating that hydrolysate at high concentrations was inhibiting the hydrogen fermentation process. In addition, the lag phase for hydrogen production was strongly influenced by the hydrolysate concentration, and was prolonged from approximately 11 h at the hydrolysate concentrations below 20% (v/v) to 38 h at the hydrolysate concentration of 25% (v/v). The maximum hydrogen yield as determined in batch assays was 318.4 ± 5.2 mL‐H₂/g‐sugars (14.2 ± 0.2 mmol‐H₂/g‐sugars) at the hydrolysate concentration of 5% (v/v). Continuously fed, and the continuously stirred tank reactor (CSTR), operating at 3 day hydraulic retention time (HRT) and fed with 20% (v/v) hydrolysate could successfully produce hydrogen. The hydrogen yield and production rate were 178.0 ± 10.1 mL‐H₂/g‐sugars (7.9 ± 0.4 mmol H₂/g‐sugars) and 184.0 ± 10.7 mL‐H₂/day L_reactor (8.2 ± 0.5 mmol‐H₂/day L_reactor), respectively, corresponding to 12% of the chemical oxygen demand (COD) from sugars. Additionally, it was found that toxic compounds, furfural and hydroxymethylfurfural (HMF), contained in the hydrolysate were effectively degraded in the CSTR, and their concentrations were reduced from 50 and 28 mg/L, respectively, to undetectable concentrations in the effluent. Phylogenetic analysis of the mixed culture revealed that members involved hydrogen producers in both batch and CSTR reactors were phylogenetically related to the Caldanaerobacter subteraneus, Thermoanaerobacter subteraneus, and Thermoanaerobacterium thermosaccharolyticum. Biotechnol. Bioeng. 2010;105: 899–908. © 2009 Wiley Periodicals, Inc.     

Detoxification of acidic catalyzed hydrolysate of Kappaphycus alvarezii (cottonii)

Red seaweed, Kappaphycus alvarezii, holds great promise for use in biofuel production due to its high carbohydrate content. In this study, we investigated the effect of fermentation inhibitors to the K. alvarezii hydrolysate on cell growth and ethanol fermentation. In addition, detoxification of fermentation inhibitors was performed to decrease the fermentation inhibitory effect. 5-Hydroxymethylfurfural and levulinic acid, which are liberated from acidic hydrolysis, was also observed in the hydrolysate of K. alvarezii. These compounds inhibited ethanol fermentation. In order to remove these inhibitors, activated charcoal and calcium hydroxide were introduced. The efficiency of activated charcoals was examined and over-liming was used to remove the inhibitors. Activated charcoal was found to be more effective than calcium hydroxide to remove the inhibitors. Detoxification by activated charcoal strongly improved the fermentability of dilute acid hydrolysate in the production of bioethanol from K. alvarezii with Saccharomyces cerevisiae. The optimal detoxifying conditions were found to be below an activated charcoal concentration of 5%.