Fermentation of Sugarcane Bagasse Hemicellulose Hydrolysate to Xylitol by a Hydrolysate-Acclimatized Yeast

The yeast strain, Candida sp B-22, which has an increased tolerance to untreated neutralized sugarcane bagasse hemicellulose hydrolysate has been isolated by a continuous adaptation-selection technique which allows yeasts to overcome the inhibitory effect of extraneous chemicals in hydrolysate. With this yeast, xylitol is produced from untreated full-strength hydrolysate in a yield of over 85% of the theoretical value. A final xylitol concentration of 94.74 g/L was obtained from 105.35 g/L D-xylose in hemicellulose hydrolysate after 96 hr of incubation.     

THE ENHANCEMENT OF AMMONIA PRETREATMENT ON THE FERMENTATION OF RICE STRAW HYDROLYSATE TO XYLITOL

In xylitol fermentation from rice straw hydrolysate by Candida tropicalis As2.1776, ammonia steeping was employed to pretreat the rice straw. Experimental results showed that the content of toxic compounds created in the hydrolysis process, such as acetic acid and phenolic compounds, was greatly reduced and the fermentation of the hydrolysate was enhanced. Xylitol fermentation was investigated in flasks and a 2‐L bioreactor, respectively. The xylitol yield factor and volumetric productivity were 0.746 g/g and 0.686 g/(L·h) in the lab‐flask fermentation, respectively. The corresponding results conducted in bioreactor fermentation were 0.689 g/g and 0.697 g/(L·h), respectively.     

Bioethanol production from ball milled bagasse using an on-site produced fungal enzyme cocktail and xylose-fermenting Pichia stipitis

Sugarcane bagasse is one of the most promising agricultural by-products for conversion to biofuels. Here, ethanol fermentation from bagasse has been achieved using an integrated process combining mechanical pretreatment by ball milling, with enzymatic hydrolysis and fermentation. Ball milling for 2 h was sufficient for nearly complete cellulose structural transformation to an accessible amorphous form. The pretreated cellulosic residues were hydrolyzed by a crude enzyme preparation from Penicillium chrysogenum BCC4504 containing cellulase activity combined with Aspergillus flavus BCC7179 preparation containing complementary β-glucosidase activity. Saccharification yields of 84.0% and 70.4% for glucose and xylose, respectively, were obtained after hydrolysis at 45 °C, pH 5 for 72 h, which were slightly higher than those obtained with a commercial enzyme mixture containing Acremonium cellulase and Optimash BG. A high conversion yield of undetoxified pretreated bagasse (5%, w/v) hydrolysate to ethanol was attained by separate hydrolysis and fermentation processes using Pichia stipitis BCC15191, at pH 5.5, 30 °C for 24 h resulting in an ethanol concentration of 8.4 g/l, corresponding to a conversion yield of 0.29 g ethanol/g available fermentable sugars. Comparable ethanol conversion efficiency was obtained by a simultaneous saccharification and fermentation process which led to production of 8.0 g/l ethanol after 72 h fermentation under the same conditions. This study thus demonstrated the potential use of a simple integrated process with minimal environmental impact with the use of promising alternative on-site enzymes and yeast for the production of ethanol from this potent lignocellulosic biomass.     

CARBON SOURCES AND THEIR EFFECT ON GROWTH, ACETIC ACID AND ETHANOL PRODUCTION BY BRETTANOMYCES BRUXELLENSIS IN BATCH CULTURE

The influence of available low‐cost carbohydrates as carbon sources on Brettanomyces bruxellensis growth, acetic acid and ethanol production was studied in order to ascertain the viability of this yeast to eventually become an industrial acetic acid producer. Six different raw materials were included as carbon sources (glucose, sugarcane molasses, refined cane sugar, pineapple, sugarcane and beet juices). B. bruxellensis develops in a complex culture medium like plant juices and sugarcane molasses better than in a medium with a simple carbohydrate such as glucose. The maximum acid acetic yield (0.24 g/g) and productivity (0.14 g/L/h) were attained in tests carried out with sugarcane molasses containing 60 g/L sucrose. The strain produced low levels of ethanol in a refined sugarcane medium, but was able to produce a substantial quantity of acetic acid (13 g/L).     

固定化热带假丝酵母发酵氨浸稻秸水解液生产木糖醇

采用海藻酸钙固定化热带假丝酵母细胞发酵氨水浸泡稻秸半纤维素水解液生产木糖醇。为了提高木糖醇的转化率,对发酵条件进行了研究。发酵在250 mL锥形瓶中进行。向水解液中补充适量氮源和营养盐等营养物质提高了木糖醇的生产速率,但木糖醇转化率没有因此而提高。适宜的初始pH和细胞干浓度分别为4-5和1.22 g/L。在这些条件下,进行了固定化细胞重复法较高浓缩度水解液的试验。结果发现,固定化细胞能在初始木糖浓度为104.2 g/L的水解液中重复批式发酵5次,木糖醇平均得率和生产速率分别为0.737 g/g和0.533 g/(L.h)。     

利用纤维床反应器驯化原壳小球藻提高对蔗渣水解液耐受性研究

原壳小球藻可快速利用蔗渣水解液中的可发酵糖,但水解液中副产物对细胞生长有抑制作用。为了提高其在高浓度水解液中的异养生长能力,本研究利用纤维床反应器(FBB)驯化细胞,系统研究了蔗渣水解液的制备及其组成、分批补料培养种子液、FBB中的细胞固定化,并在FBB中利用水解液为培养基进行细胞驯化。结果表明,蔗渣经酸解酶解后,其水解液的主要成分为葡萄糖、木糖、乙酸、纤维二糖和阿拉伯糖,浓度分别为18.40g/L、16.17g/L、6.13g/L、5.10g/L和2.29g/L;在发酵罐中采用Basal培养基补料分批培养细胞,117h后细胞密度可达到12.37g/L;将发酵罐与FBB连接并循环培养基33h后形成了固定化细胞床;随后以水解液培养基代替Basal培养基,通过逐级提高水解液培养基浓度来驯化培养固定化细胞,最终从纤维床上分离获得了能在含有35g/L葡萄糖的水解液中异养生长的高耐受性藻株,而野生型藻株不能生长。     

Ethanol production by a new pentose-fermenting yeast strain, Scheffersomyces stipitis UFMG-IMH 43.2, isolated from the Brazilian forest

The ability of a recently isolated Scheffersomyces stipitis strain (UFMG-IMH 43.2) to produce ethanol from xylose was evaluated. For the assays, a hemicellulosic hydrolysate produced by dilute acid hydrolysis of sugarcane bagasse was used as the fermentation medium. Initially, the necessity of adding nutrients (MgSO(4)·7H(2)O, yeast extract and/or urea) to this medium was verified, and the yeast extract supplementation favoured ethanol production by the yeast. Then, in a second stage, assays under different initial xylose and cell concentrations, supplemented or not with yeast extract, were performed. All these three variables showed significant (p < 0.05) influence on ethanol production. The best results (ethanol yield and productivity of 0.19 g/g and 0.13 g/l/h, respectively) were obtained using the hydrolysate containing an initial xylose concentration of 30 g/l, supplemented with 5.0 g/l yeast extract and inoculated with an initial cell concentration of 2.0 g/l. S. stipitis UFMG-IMH 43.2 was demonstrated to be a yeast strain with potential for use in xylose conversion to ethanol. The establishment of the best fermentation conditions was also proved to be of great importance to increasing the product formation by this yeast strain. These findings open up new perspectives for the establishment of a feasible technology for ethanol production from hemicellulosic hydrolysates.     

Production of detoxified sorghum straw hydrolysates for fermentative purposes

Sorghum straw can be hydrolysed to obtain monosaccharide solutions, mainly containing xylose. The usual biotechnological application of xylose is its bioconversion to xylitol. The global process from straw to xylitol can give an added value to the sorghum straw. The process has the following sequential steps: reduction of size, acid hydrolysis, neutralization, detoxification, fermentation, recovery and purification. This work deals with the optimization of the detoxification process of sorghum straw hydrolysates with activated charcoal. The variables evaluated were pH (1–5), contact time (20–60 min) and activated charcoal charge (20–33 g kg^?1). Mathematical models were obtained through a factorial experimental design. The models suggest that optimal conditions for detoxification are pH 1, contact time of 29 min and a charcoal charge of 33 g kg^?1. These conditions allowed hydrolysates with 54.2 g xylose L^?1, 13.5 g glucose L^?1, 12 g arabinose L^?1, 0.2 g furfural L^?1 and 0.0 g acetic acid L^?1 to be obtained. The results suggest that performing the detoxification step before the neutralization step gave the best outcome. Fermentations by Candida parapsilosis NRRL Y‐2315 were performed and it was confirmed that the treated hydrolysate is suitable for xylitol production, yielding up to 17 g L^?1 of this polyol. Copyright © 2006 Society of Chemical Industry     

驯化和胶囊包裹的假丝酵母LF04发酵玉米芯水解液生产木糖醇(英文)

玉米芯的酸水解液是木糖醇生产的重要原料,但是该水解液中含有糠醛、酚类等对后续微生物发酵有毒害作用的化合物。本研究从土壤中分离了一株似假丝酵母LF01,通过驯化和微胶囊包裹来提高其对水解液的抗性。结果表明通过多次驯化并进行包裹的假丝酵母LF04能在玉米芯水解液中不经任何脱毒处理发酵木糖生产木糖醇。在pH5.5 溶氧为 0.15vvm 的条件下发酵 88h,木糖转化率为 76%,木糖醇浓度达 61.768g/L。远高于其出发菌株。该结果表明采用该方法有望用于木糖醇的工业化生产。     

Use of spent brewer's yeast in L‐(+) lactic acid fermentation

The application of by‐products from the brewing industry in lactic acid (LA) production was investigated in order to replace expensive nitrogen sources (such as yeast extract) with cheaper and renewable nitrogenous materials such as brewer's yeast (BY). In this study, brewer's spent grain (BSG) hydrolysate was used for L‐(+)‐LA fermentation by Lactobacillus rhamnosus ATCC 7469. The effect of pH control during the fermentation and the addition of various BY contents (5–50 g/L) in BSG hydrolysate on fermentation parameters was evaluated. BY addition significantly increased free amino nitrogen (FAN) concentration (by 25.2% at 5 g/L to 616% at 50 g/L). A strong positive correlation between FAN concentration in the hydrolysate and concentration of L‐(+)‐LA produced was observed (correlation coefficient of 0.913). A high cell viability of L. rhamnosus ATCC 7469 (1.95–3.32 × 10⁹ CFU/mL at the end of fermentation) was achieved in all fermentations with the addition of brewer's yeast. The addition of BY increased L‐(+)‐lactic acid yield and volumetric productivity up to 8.4% (5 g/L) and 48.3% (50 g/L). The highest L‐(+)‐LA yield (89%) and volumetric productivity (0.89 g/L h^?1) were achieved in fermentation of BSG hydrolysate with 50 g/L of BY. © 2019 The Institute of Brewing & Distilling     

Xylitol bioproduction from wheat straw: hemicellulose hydrolysis and hydrolyzate fermentation

A 2² central composite design with five center points was performed to estimate the effects of temperature (120, 130 and 140 °C) and acid loading (100, 150 and 200 mg g^?1) on the yield of monomeric xylose recovery from wheat straw hemicellulose (Y_S/RM). Under the best hydrolysis condition (140 °C and 200 mg g^?1), a Y_S/RM of 0.26 g g^?1 was achieved. After vacuum concentration and detoxification by pH alteration and active charcoal adsorption, the hydrolyzate was used as source of xylose for xylitol bioproduction in a stirred tank reactor. A xylitol production of 30.8 g L^?1 was achieved after 54 h^?1 of fermentation, resulting in a productivity (Q_P) of 0.57 g L^?1 h^?1 and bioconversion yield (Y_P/S) of 0.88 g g^?1. The maximum specific rates of xylose consumption and xylitol production were 0.19 and 0.15 g g^?1 h^?1, respectively. Copyright © 2006 Society of Chemical Industry     

稳定高效表达木糖还原酶基因工业酿酒酵母的构建及木糖醇发酵的初步研究

将树干毕赤氏酵母(Pichia stipitis)木糖还原酶基因XYL1连接到适用于酿酒酵母工业菌株的多拷贝整合载体pYMIKP中,构建得到表达质粒pYMIKP-XYL1,转化酿酒酵母工业菌株Saccharomyces cerevisiae6508。在G418平板上筛选转化子,得到含高拷贝木糖还原酶基因的酿酒酵母重组菌株XGH2,,该菌株的木糖还原酶比活力为0.8 U/mg(蛋白),比出发菌株提高了80倍以上,表明外源基因在工业菌株中实现了高效表达。摇瓶发酵结果显示,重组菌株XGH2木糖消耗为27.9 g/L,木糖消耗率为51%;木糖醇产量为30.2 g/L,木糖醇的转化率大于1.0 g/g木糖。     

Xylitol production from rice straw hemicellulose hydrolyzate by polyacrylic hydrogel thin films with immobilized Candida subtropicalis WF79

Xylose from rice straw hemicellulose hydrolysate was fermented for xylitol production using Candida subtropicalis WF79 cells immobilized in polyacrylic hydrogel thin films of 200 mum thickness. Cell immobilization was conducted by first suspending the yeast cells in a mixture of 2-hydroxyethyl methacrylate (HEMA, hydrophilic monomer), polyethylene glycol diacrylate (PEG-DA, crosslinking agent), and benzoin isopropyl ether (photoinitiator). The mixture was then allowed to form polyacrylic hydrogel thin films, between two pieces of glass sheets, by UV-initiated photopolymerization. The hemicellulose of rice straw was hydrolyzed using dilute sulfuric acid at 126 degrees C. The hydrolysate was neutralized with calcium hydroxide. After separating the solid residues and calcium sulfate precipitates by filtration, the hydrolysate was treated with charcoal to partially remove potential inhibitory substances, followed by vacuum concentration to obtain solutions of desired xylose concentrations for yeast fermentation. The thin films with immobilized yeast cells were submerged in the xylose solution from rice straw hydrolysate for fermentation in an Erlenmeyer flask. The maximum yield was 0.73 g of xylitol per gram of xylose consumed. In the 52.5-day long durability test, after 40 d of repeated batchwise operation, the fermentation activities of the cell immobilized in thin films began to decline to a yield of 0.57 g/g at the end.     

Effects of culture substrates on taste component content and taste quality of Lentinula edodes

In this research, the effects of culture substrates on taste component content of Lentinula edodes were studied, and the resulting taste quality of L. edodes was evaluated. The results revealed that single‐carbon and single‐nitrogen sources were beneficial to production of soluble sugars and polyols, organic acids and sweet amino acids, whereas a single‐carbon source was beneficial to essential amino acid production, and a mixed‐carbon source was beneficial to umami 5′‐nucleotides and high mushroom production yield. High C/N values were beneficial to trehalose, arabitol, malic acid, and succinic acid production, while low C/N values were beneficial to mannitol and citric acid production. L. edodes fruiting bodies, harvested from a culture substrate containing single carbon, high proportion of cereal bran, had a more palatable taste quality, while a substrate containing bagasse and low C/N values was not suitable for L. edodes cultivation due to unfavourable taste quality. These results provide cultivation information how to obtain fruiting bodies L. edodes with more taste components.     

The level of glucose-6-phosphate dehydrogenase activity strongly influences xylose fermentation and inhibitor sensitivity in recombinant Saccharomyces cerevisiae strains

Disruption of the ZWF1 gene encoding glucose-6-phosphate dehydrogenase (G6PDH) has been shown to reduce the xylitol yield and the xylose consumption in the xylose-utilizing recombinant Saccharomyces cerevisiae strain TMB3255. In the present investigation we have studied the influence of different production levels of G6PDH on xylose fermentation. We used a synthetic promoter library and the copper-regulated CUP1 promoter to generate G6PDH-activities between 0% and 179% of the wild-type level. G6PDH-activities of 1% and 6% of the wild-type level resulted in 2.8- and 5.1-fold increase in specific xylose consumption, respectively, compared with the ZWF1-disrupted strain. Both strains exhibited decreased xylitol yields (0.13 and 0.19 g/g xylose) and enhanced ethanol yields (0.36 and 0.34 g/g xylose) compared with the control strain TMB3001 (0.29 g xylitol/g xylose, 0.31 g ethanol/g xylose). Cytoplasmic transhydrogenase (TH) from Azotobacter vinelandii has previously been shown to transfer NADPH and NAD(+) into NADP(+) and NADH, and TH-overproduction resulted in lower xylitol yield and enhanced glycerol yield during xylose utilization. Strains with low G6PDH-activity grew slower in a lignocellulose hydrolysate than the strain with wild-type G6PDH-activity, which suggested that the availability of intracellular NADPH correlated with tolerance towards lignocellulose-derived inhibitors. Low G6PDH-activity strains were also more sensitive to H(2)O(2) than the control strain TMB3001.     

Steam pretreatment and fermentation of the straw material "Paja Brava" using simultaneous saccharification and co-fermentation

Pretreatment, enzymatic hydrolysis and simultaneous saccharification and fermentation (SSF) of the South American straw material Paja Brava were investigated. Suitable process conditions for an SO₂-catalyzed steam pretreatment of the material were determined and assessed by enzymatic digestibility of obtained fiber slurries for 72 h at a water insoluble solids (WIS) content of 2%. The best pretreatment conditions obtained (200°C, 5 min holding time and 2.5% SO₂) gave an overall glucose yield following enzymatic hydrolysis of more than 90%, and a xylose yield of about 70%. Simultaneous saccharification and co-fermentation of glucose and xylose (SSCF) of the pretreated material using the xylose-fermenting strain Saccharomyces cerevisiae TMB3400 was examined at WIS contents between 5% and 10%. In agreement with previous studies on other materials, the overall ethanol yield and also the xylose conversion decreased somewhat with increasing WIS content in the SSCF. In batch SSCF, the xylose conversion obtained was almost 100% at 5% WIS content, but decreased to 69% at 10% WIS. The highest ethanol concentration obtained for a WIS content of 10% was about 40 g/L, corresponding to a yield of 0.41 g/g in a fed-batch SSCF. The Paja Brava material has previously been found difficult to hydrolyze in a dilute-acid process. However, the SSCF results obtained here show that similar sugar yields and fermentation performance can be expected from Paja Brava as from materials such as wheat straw, corn stover or sugarcane bagasse.     

利用皮状丝孢酵母对甘蔗渣酶解液产油脂的研究

本试验旨在研究碱和高温液态水2种预处理方式对发酵的影响。甘蔗渣分别利用碱和高温液态水(LHW)预处理,然后使用纤维素酶进行酶解。在LHW和碱预处理后的酶解液中,分别含22.06和39.28 g/L总糖。不经过浓缩,也不需要添加任何营养物质,酶解液被用于皮状丝孢酵母的油脂发酵试验。经过3 d的培养,在LHW预处理的酶解液中,皮状丝孢酵母的生物量达到了8.08 g/L,油脂质量分数为52.00%,油脂系数高达19.03%。在碱预处理的酶解液中,经过8 d的培养,皮状丝孢酵母的生物量为13.67 g/L,油脂质量分数为43.20%,油脂系数为15.03%。脂肪酸组成分析表明所产油脂与棕榈油的组成相似。试验结果表明,甘蔗渣是一种有前景的产油脂原料,LHW是一种良好的预处理方式。该研究为利用甘蔗渣发酵产油脂提供了新思路。     

Lactic acid fermentation of brewer's spent grain hydrolysate by Lactobacillus rhamnosus with yeast extract addition and pH control

Lactic acid (LA) is a versatile chemical with a wide range of applications in food, pharmaceutical, cosmetic, textile and polymer industries. Brewer's spent grain (BSG) is the most abundant brewing by‐product. In this study BSG hydrolysates were used for LA fermentation by Lactobacillus rhamnosus ATCC 7469. The aim of this study was to evaluate the effects of pH control during fermentation, reducing sugar content and yeast extract content in BSG hydrolysate on LA fermentation parameters. The pH control greatly increased reducing sugar utilization, l ‐(+)‐LA content, yield and volumetric productivity. The highest l ‐(+)‐LA yield and volumetric productivity were achieved with the reducing sugar content of 54 g/L. Yeast extract addition significantly increased reducing sugar utilization, l ‐(+)‐LA content, L. rhamnosus cell viability, l ‐(+)‐LA yield and volumetric productivity. The highest l ‐(+)‐LA content (39.38 g/L), L. rhamnosus cell viability (9.67 log CFU/mL), l ‐(+)‐LA yield (91.29%) and volumetric productivity (1.69 g/L/h) were achieved with the reducing sugar content of 54 g/L and yeast extract content of 50 g/L. Copyright © 2017 The Institute of Brewing & Distilling     

Production of coumaric acid from sugarcane bagasse

Phenolic acids were released from sugarcane bagasse by alkaline hydrolysis at 30 °C for 4 h; The alkaline hydrolysates were ultrafiltrated, the permeates purified with anion exchange resin. The phenolic acids bound by the resin were desorbed by a mixture of water–ethanol–HCl solution (36: 60: 4) after washing the resin with water, ethanol and dilute HCl respectively. The combined eluents were concentrated for crystalization, and the crystals filtered and washed using 1% (v/v) HCl. After this purification process, the purity of products reached 89.7% based on coumaric acid. Results of HPLC/MS, HPLC using standard coumaric acid and ferulic acid showed that the main component of the purified bagasse hydrolysate was p-coumaric acid rather than ferulic acid. The purified products showed the same antioxidant activity, reducing power and free radical scavenging capacity as the standard p-coumaric acid.The technology could be applied on industrial scale.Industrial relevanceThis research presents a technology to produce coumaric acids from sugarcane bagasse. The first step is to release coumaric acid by alkaline hydrolysis. The second step is to remove the viscous polysaccharides and protein by ultrafiltration. The third step is to purify coumaric acid from the permeate of ultrafiltration by anion chromatography, and the alkaline could be reused to hydrolyze the bagasse. The technology showed potential application on industrial scale.     

Favorable effect of very low initial KLa value on xylitol production from xylose by a self-isolated strain of Pichia guilliermondii

Xylitol production from xylose by a self-isolated furfural and 5-hydroxymethyl furfural assimilating Pichia guilliermondii was studied under oxygen limitation. An extremely low initial volumetric oxygen transfer coefficient (0.075 h^− 1) was found most favorable to the xylitol production with yield of 0.61 g g^− 1. Related enzymes activities were also investigated and discussed.