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Applied Biochemistry and Biotechnology: Part A: Enzyme Engineering and Biotechnology (v.168, #1)

Engineering a Metabolic Pathway for Isobutanol Biosynthesis in Bacillus subtilis by Xiaoqiang Jia; Shanshan Li; Sha Xie; Jianping Wen (pp. 1-9).

Isobutanol can be biosynthesized via α-ketoisovalerate catalyzed by heterologous keto acid decarboxylase (KDC) and alcohol dehydrogenase (ADH). In this work, isobutanol biosynthesis pathway was designed in Bacillus subtilis, a notable solvent-tolerant host. In order to do that, a plasmid pPKA expressing KDC and ADH under the control of a B. subtilis strong promoter P₄₃ was constructed. Isobutanol was detected in the products of the recombinant B. subtilis harboring pPKA plasmid, whereas none was detected by the wild-type strain. Effects of the medium ingredients such as glucose concentration and valine addition, and operating parameters such as initial pH, inoculation volume, and medium work volume on isobutanol production were also investigated. Isobutanol production reached to the maximum of 0.607 g/L after 35-h cultivation under the conditions: glucose concentration of 3%, valine addition of 2%, initial pH of 7.0, inoculum of 1%, and work volume of 50 mL/250 mL. Though the isobutanol production by the recombinant was low, it was the first successful attempt to produce isobutanol in engineered B. subtilis, and the results showed its great potential as an isobutanol-producing cell factory.

Keywords: Isobutanol; Bacillus subtilis ; Genetic engineering; Biosynthesis

Effects of Preparation Method on the Performance of Ni/Al₂O₃ Catalysts for Hydrogen Production by Bio-Oil Steam Reforming by Xinbao Li; Shurong Wang; Qinjie Cai; Lingjun Zhu; Qianqian Yin; Zhongyang Luo (pp. 10-20).

Steam reforming of bio-oil derived from the fast pyrolysis of biomass is an economic and renewable process for hydrogen production. The main objective of the present work has been to investigate the effects of the preparation method of Ni/Al₂O₃ catalysts on their performance in hydrogen production by bio-oil steam reforming. The Ni/Al₂O₃ catalysts were prepared by impregnation, co-precipitation, and sol–gel methods. XRD, XPS, H₂-TPR, SEM, TEM, TG, and N₂ physisorption measurements were performed to characterize the texture and structure of the catalysts obtained after calcination and after their subsequent use. Ethanol and bio-oil model compound were selected for steam reforming to evaluate the catalyst performance. The catalyst prepared by the co-precipitation method was found to display better performance than the other two. Under the optimized reaction conditions, an ethanol conversion of 99% and a H₂ yield of 88% were obtained.

Keywords: Bio-oil; Steam reforming; Hydrogen; Ni/Al₂O₃ ; Ethanol; Model compound

Characteristics of an Immobilized Yeast Cell System Using Very High Gravity for the Fermentation of Ethanol by Hairui Ji; Jianliang Yu; Xu Zhang; Tianwei Tan (pp. 21-28).

The characteristics of ethanol production by immobilized yeast cells were investigated for both repeated batch fermentation and continuous fermentation. With an initial sugar concentration of 280 g/L during the repeated batch fermentation, more than 98% of total sugar was consumed in 65 h with an average ethanol concentration and ethanol yield of 130.12 g/L and 0.477 g ethanol/g consumed sugar, respectively. The immobilized yeast cell system was reliable for at least 10 batches and for a period of 28 days without accompanying the regeneration of Saccharomyces cerevisiae inside the carriers. The multistage continuous fermentation was carried out in a five-stage column bioreactor with a total working volume of 3.75 L. The bioreactor was operated for 26 days at a dilution rate of 0.015 h⁻¹. The ethanol concentration of the effluent reached 130.77 g/L ethanol while an average 8.18 g/L residual sugar remained. Due to the high osmotic pressure and toxic ethanol, considerable yeast cells died without regeneration, especially in the last two stages, which led to the breakdown of the whole system of multistage continuous fermentation.

Keywords: Ethanol; Cell immobilization; Sweet sorghum bagasse; Repeated batch fermentation; Multistage continuous fermentation

Effect of Agitation Rate on Ethanol Production from Sugar Maple Hemicellulosic Hydrolysate by Pichia stipitis by Alan M. Shupe; Shijie Liu (pp. 29-36).

Concentrated dilute acid hydrolysate was obtained from hot water extracts of Acer saccharum (sugar maple) and was fermented to ethanol by Pichia stipitis in a 1.3-L-benchtop bioreactor. The conditions under which the highest ethanol yield was achieved were when the air flow rate was set to 100 cm³ and the agitation rate was set to 150 rpm resulting in an overall mass transfer coefficient (K _L a) of 0.108 min⁻¹. A maximum ethanol concentration of 29.7 g/L was achieved after 120 h of fermentation; however, after 90 h of fermentation, the ethanol concentration was only slightly lower at 29.1 g/L with a yield of 0.39 g ethanol per gram of sugar consumed. Using the same air flow rate and adjusting the agitation rate resulted in lower ethanol yields of 0.25 g/g at 50 rpm and 0.30 g/g at 300 rpm. The time it takes to reach the maximum ethanol concentration was also affected by the agitation rate. The ethanol concentration continued to increase even after 130 h of fermentation when the agitation rate was set at 50 rpm, whereas the maximum ethanol concentration was reached after only 68.5 h at 300 rpm.

Keywords: Pichia stipitis ; Hemicellulosic hydrolysate; Ethanol fermentation; Agitation rate

Structure and Pyrolysis Characteristics of Lignin Derived from Wood Powder Hydrolysis Residues by Bin Zhang; Xiuli Yin; Chuangzhi Wu; Zejing Qiu; Congwei Wang; Yanqin Huang; Longlong Ma; Shubin Wu (pp. 37-46).

Physicochemical characteristics of wood powder acid hydrolysis residue (WAHR) were studied firstly in this study, and WAHL (lignin derived from WAHR) was separated successfully from WAHR based on an improved isolating method. The content of functional group such as phenolic hydroxyl group of guaiacyl, syringyl, and hydroxyl–phenyl units in WAHL were identified by ³¹P-NMR and DFRC (derivatization followed by reductive cleavage) method. Thermal degradation experiments were carried out on a thermogravimetric (TG) analyzer to show pyrolysis characteristics of WAHL. The compositions of pyrolysis products of WAHL were also studied throughout a pyrolysis–gas chromatography–mass spectrometry (Py–GC–MS) analyzer. It was shown that the pyrolysis of WAHL took place in a wide temperature range and there were two obvious peaks in the differential thermogravimetric diagram. Results of Py–GC–MS analysis indicated that pyrolysis products were mainly formed through cleavage of the β-O-4 connection and multiple pyrolysis.

Keywords: WAHL; Pyrolysis; NMR; FTIR; Py–GC–MS

Understanding the Influence of Phosphatidylcholine on the Molecular Weight of Hyaluronic Acid Synthesized by Streptococcus zooepidemicus by Jingfeng Sun; Miaomiao Wang; Yanrong Chen; Fei Shang; Hua Ye; Tianwei Tan (pp. 47-57).

The effect of phosphatidylcholine on the molecular weight properties of hyaluronic acid (HA) was studied in batch culture of Streptococcus zooepidemicus by adding phosphatidylcholine at the early stage of exponential phase. With the addition of 80 mg/L of phosphatidylcholine, maximum HA yield (2.47 g/L) and weight-average molecular weight (902.60 KDa) were achieved, increased by 17.4% and 67.1%, respectively, as compared to the control. Metabolic flux analysis was employed to study the mechanism of phosphatidylcholine on the molecular weight of HA. The normalized flux distribution maps based on fermentation data at phosphatidylcholine addition indicated that phosphatidylcholine resulted in higher flux flowing to the HA pathway and lower flux flowing to the glycolysis and biomass synthesis pathway, coupling with higher level of UDPNAG generation and extra regeneration of ATP. The GC-MS analysis of fatty acids in the plasma membrane showed that the addition of phosphatidylcholine could promote the mobility and permeability of the cell membrane, making the HA chain pass through the membrane more easily, thus decreasing the energy consumption. All these results led to higher molecular weight of hyaluronic acid.

Keywords: Hyaluronic acid; Phosphatidylcholine; Molecular weight; Metabolic flux analysis; Streptococcus zooepidemicus

High Production of β-Glucosidase by Aspergillus niger on Corncob by Chunli Wang; Gaihong Wu; Chang Chen; Shulin Chen (pp. 58-67).

Using low-cost raw material is an effective approach for reducing the cost of cellulolytic enzymes. The farmland waste corncob was found in this study to be the best carbon source for the production of β-glucosidase by Aspergillus niger. The maximum yield of β-glucosidase activity was 48.7 IU ml⁻¹ by using 50 g l⁻¹ of corncob powder as the substrate. It was found that the water-soluble components of the corncob could increase β-glucosidase production significantly only when mixed with Avicel or wheat bran. The soluble components could not enhance the biomass and β-glucosidase production when used alone. On the other hand, the water-insoluble components of the corncob still produced high level of β-glucosidase (30 IU ml⁻¹) although lower than that of using whole corncob. The results suggested that the water-insoluble components of corncob were beneficial for β-glucosidase production. It was further demonstrated that the xylan in the water-insoluble parts of corncob was the important factor in producing β-glucosidase by A. niger.

Keywords: Corncob; β-Glucosidase; Aspergillus niger ; Xylan; The water-soluble components; Water-insoluble components

Enzymatic Synthesizing of Phytosterol Oleic Esters by Xinxin Pan; Biqiang Chen; Juan Wang; Xinzhi Zhang; Biyun Zhul; Tianwei Tan (pp. 68-77).

A method of synthesizing the phytosterol esters from oleic acid and sterols was studied, using immobilized lipase Candida sp. 99–125 as catalyst. Molar ratio (oleic acid/phytosterols), temperature, reaction period, organic solvents, catalyst, and silica-gel drier were optimized, and the result showed that 93.4% of the sterols had been esterified under the optimal synthetic condition: the molar ratio of oleic acid/phytosterol is 1:1 in 10 mL iso-octane, immobilized lipase (w, 140% of the sterols), incubated in an orbital shaker (200 rpm) at a temperature of 45 °C for 24 h. The immobilized lipase could be reused for at least 13 times with limited loss of esterification activity. The conversion still maintained up to 86.6%. Hence, this developed process for synthesizing phytosterol esters could be considered as simple and low-energy consumption compared to existing chemical processes.

Keywords: Phytosterol oleic esters; Phytosterols; Lipase synthesis; Immobilized enzyme

Biogasification of Green and Food Wastes Using Anaerobic-Phased Solids Digester System by Guangqing Liu; Ruihong Zhang; Hamed M. El-Mashad; Renjie Dong; Xiaoying Liu (pp. 78-90).

The performance of a laboratory-scale anaerobic-phased solid (APS) digester system treating food and green wastes was evaluated at thermophilic condition. The APS system comprised of four hydrolysis digesters and one biogasification reactor. The hydrolysis reactors were operated batchwised at a 12-day retention time, while the biogasification reactor was continuously operated at different hydraulic retention times (HRT). The biogas and methane yields from green waste were determined to be 0.438 and 0.252 L/g volatile solid (VS), respectively, with VS removal of 78%. The biogas and methane yields from food waste were 0.596 and 0.379 L/g VS, respectively, with VS removal of 85%. Hydrogen was produced from hydrolysis reactors during the digestion of food waste. Its content was 30.1% and 8.5% of the biogas produced on the first and second day of digestion, respectively. Hydrogen yield from the whole system was determined to be 0.029 L/g VS representing about 4.9% of the total biogas production from the system. The ratio between the volumes of biogasification and hydrolysis reactors (BR/HR) was found to be a factor that affects the process performance and stability.

Keywords: Biogas; Hydrogen; Anaerobic-phased solid-bed (APS); Biogasification; Food waste; Green waste

Fermentative Hydrogen Production from Soybean Protein Processing Wastewater in an Anaerobic Baffled Reactor (ABR) Using Anaerobic Mixed Consortia by Ge-fu Zhu; Jian-zheng Li; Chao-xiang Liu (pp. 91-105).

Fermentative H₂ production from soybean protein processing wastewater (SPPW) was investigated in a four-compartment anaerobic baffled reactor (ABR) using anaerobic mixed cultures under continuous flow condition in the present study. After being inoculated with aerobic activated sludge and operated at the inoculants of 5.98 gVSS L⁻¹, COD of 5000 mg L⁻¹, HRT of 16 h and temperature of (35 ± 1) °C for 22 days, the ABR achieved stable ethanol-type fermentation. The specific hydrogen production rate of anaerobic activated sludge was 165 LH₂ kg MLVSS⁻¹ day⁻¹, the substrate conversion rate was 600.83 LH₂ kg COD⁻¹and the COD removal efficiency was 44.73% at the stable operation status. The ABR system exhibited a better stability and higher hydrogen yields than continuous stirring tank reactor under the same operational condition. The experimental data documented the feasibility of substrate degradation along with molecular H₂ generation utilizing SPPW as primary carbon source in the ABR system.

Keywords: Anaerobic baffled reactor (ABR); Fermentative hydrogen production; Wastewater treatment; Hydrogen-producing acetogenic bacteria; Hydrogen yields

Production of Tung Oil Biodiesel and Variation of Fuel Properties During Storage by Qiong Shang; Jiao Lei; Wei Jiang; Houfang Lu; Bin Liang (pp. 106-115).

The crude Tung oil with 4.72 mg KOH/g of acid value (AV) was converted by direct transesterification, and the reaction mixture was quantified. The phase distribution data showed that 38.24% of excess methanol, 11.76% of KOH, 10.13% of soap and 4.36% of glycerol were in the biodiesel phase; 0.35% of biodiesel dissolved in the glycerol phase. Tung oil biodiesel as well as its blends with 0^# diesel was investigated under different storage conditions. The results indicated that higher temperature greatly influenced the storage stability, especially when the volume fraction of Tung oil biodiesel is increased in the blends.

Keywords: Tung oil; Biodiesel; 0^# diesel; Blends; Storage stability

Influence of Blocking of 2,3-Butanediol Pathway on Glycerol Metabolism for 1,3-Propanediol Production by Klebsiella oxytoca by Gang Zhang; Guang Yang; Xu Wang; Qingjuan Guo; Ying Li; Jilun Li (pp. 116-128).

Glycerol metabolism is a typical biological oxidoreductive reaction. 1,3-Propanediol (1,3-PD) is the final product of the reductive branch, while acetate, succinate, lactate, 2,3-butanediol (2,3-BD), and ethanol were produced in the oxidative branch. 2,3-BD, which has similar properties of high boiling point and water solubility with 1,3-PD, not only contests the carbon flow and NADH with 1,3-PD but also serves as an obstacle for obtaining high purity 1,3-PD in downstream processes. In this study, a 2,3-BD pathway-deficient mutant of Klebsiella oxytoca ZG36 was constructed by knocking out the budA gene of the wild-type strain M5al. The results of fed-batch fermentation by ZG36 indicated that the glycerol flux and the distribution of metabolites were altered in the K. oxytoca when the 2,3-BD pathway was blocked. No 2,3-BD was produced, and the activity of α-acetolactate decarboxylase (α-ALDC) can not be detected in the fermentation processes. The indexes of the 1,3-PD titer, the conversion from glycerol to 1,3-PD, and the productivity per cell dry weight (CDW) increased by 42%, 62%, and 46%, respectively, compared with the M5al, and the yield of the byproducts also increased obviously. The assay of the enzyme activities in the oxidative branch and the reductive branch of the glycerol metabolism, as well as the intracellular redox state, exposited the results logically.

Keywords: 1,3-Propanediol; 2,3-Butanediol; α-Acetolactate decarboxylase; Reducing equivalent; Klebsiella oxytoca

New Feruloyl Esterases to Access Phenolic Acids from Grass Biomass by Meiqun Wu; Kofi Abokitse; Stephan Grosse; Hannes Leisch; Peter C. K. Lau (pp. 129-143).

In the Sorangium cellulosum strain So ce56 genome, two putative esterase-encoding genes (loci sce1896 and sce8927) were cloned, expressed in Escherichia coli, and the resulting enzymes (designated ScFAE1 and ScFAE2) were used to assess the possible release of ferulic acid (FA) from triticale and wheat brans, and an aqueous fraction of steam-exploded wheat straw. The two polypeptides, sharing only 30% sequence identity, exhibit a typical catalytic Ser-Asp-His triad, a characteristic of α/β-hydrolase fold proteins. Both ScFAE1 (35 kDa) and ScFAE2 (34 kDa) were purified to apparent homogeneity and comparison of their kinetic parameters indicated an apparent higher affinity of ScFAE2 than ScFAE1 towards the various feruloyl substrates. This property was reflected by the observation that ScFAE2 was capable of yielding up to 85% of FA from destarched triticale bran. In the steam-exploded wheat sample, more than 85% yield of FA or p-coumaric acid was also effected by ScFAE2 without the decomposition of valuable chemical such as furfural. The two cloned FAEs represent the first of myxobacterial origin to be characterized and they are classified as new members of the type D family of FAEs.

Keywords: Biorefinery; Platform chemicals; Antioxidant; Phenolic acid; Genome mining; α/ß-Hydrolase; Triticale; Lignocellulose; Myxobacteria; Biocatalyst

A Fractal-Like Kinetic Equation to Investigate Temperature Effect on Cellulose Hydrolysis by Free and Immobilized Cellulase by Yu Zhang; Jing-Liang Xu; Wei Qi; Zhen-Hong Yuan; Xin-Shu Zhuang; Yun Liu; Min-Chao He (pp. 144-153).

According to fractal-like theory in the heterogeneous system, a cellulase-catalyzed kinetic equation that contained two parameters (rate constant k and fractal dimension h) was deduced. The equation described directly the mathematical relationship between reducing sugar concentration and hydrolytic time, and accurately fitted the experimental data of free/immobilized cellulase at 37, 40, 44, 47, and 50 °C (R ² > 0.99). The fitted h value is estimated as a constant (0.6148) in these tested temperatures. The fitted k value increased with temperature increase, and the relationship agreed with Arrhenius equation (R ² > 0.98). The fractal-like equation could predict accurately the experimental data at low temperature 34 °C for free/immobilized cellulase and high temperature 53 °C for immobilized cellulase, but the prediction at 53 °C for free cellulase was not accurate enough due to its lower stability than immobilized cellulase. The application of fractal-like theory in cellulase kinetics is successful.

Keywords: Biomass; Cellulase; Fractal-like kinetics; Rate constant; Arrhenius equation; Simulation and prediction

Production of Energy and Activated Carbon from Agri-Residue: Sunflower Seed Example by Adam A. Donaldson; Parag Kadakia; Murlidhar Gupta; Zisheng Zhang (pp. 154-162).

In this work, a biomass processing facility is designed and simulated for the annual conversion of 77 ktons of sunflower residue into electricity and activated carbon. The residue is initially pyrolized to produce low hydrocarbon gases (35 wt%), bio-oils (30 wt%), and char (35 wt%). The gases and bio-oils are separated and combusted to generate high pressure steam, electricity, and steam for conversion of char into activated carbon. Assuming 35% of the char’s mass is lost during activation, the proposed process produces 15.6 ktons activated carbon and 5.5 ktons ash annually, while generating 10.2 MW of electricity. Economic analysis of the proposed facility yielded capital costs of $31.64 million, annual operating costs of $31.58 million, and a yearly gross revenue of $38.9 million. A discounted payback period of 6.1 years was determined for the current design, extending to 10 years if the facility were operated at 75% capacity. While the proposed process appears to be economically viable, profitability is highly sensitive to the selling price of electricity and activated carbon, highlighting the need for additional research into the pyrolysis reactor design, char/ash separation techniques, and the quality of activated carbon obtained using char from sunflower residue pyrolysis.

Keywords: Biomass conversion; Pyrolysis; Combined heat and power; Agri-residue; Sunflower

Cadmium Tolerance and Bioaccumulation of 18 Hemp Accessions by Gangrong Shi; Caifeng Liu; Meicheng Cui; Yuhua Ma; Qingsheng Cai (pp. 163-173).

Hemp (Cannabis sativa L.) is a fast-growing and high biomass producing plant species, which has been traditionally grown as multiple-use crop and recently considered as an energy crop. In order to screen accessions that can be cultivated in cadmium (Cd)-contaminated soils for biodiesel production, the ability of Cd tolerance and bioaccumulation of 18 hemp cultivars or ecotypes were evaluated in pot experiment under 25 mg Cd kg⁻¹ (dry weight, DW) soil condition, in terms of plant growth, pigment contents, chlorophyll fluorescence, and Cd accumulation at 45 days after seedling emergence. Results showed that seedlings of all cultivars, except USO-31, Shenyang and Shengmu, could grow quite well under 25 mg Cd kg⁻¹ (DW) soil condition. Among them, Yunma 1, Yunma 2, Yunma 3, Yunma 4, Qujing, Longxi, Lu'an, Xingtai, and Shuyang showed great biomass (>0.5 g plant⁻¹), high tolerance factors (68.6–92.3%), and little reduction of pigment content and chlorophyll fluorescence under 25 mg Cd kg⁻¹ (DW) soil stress, indicating these cultivars had a strong tolerance to Cd stress and could be cultivated in Cd-contaminated soils. Cultivars Longxi, Lu'an, Xingtai, Yunma 2, Yunma 3, Yunma 4, and Qujing exhibited higher Cd concentrations and total Cd in shoots. These cultivars, therefore, are good candidates for the implementation of the new strategy of cultivating biodiesel crops for phytoremediation of Cd-contaminated soils.

Keywords: Cadmium; Hemp; Tolerance; Accumulation

Study on the Pyrolysis of Cellulose for Bio-Oil with Mesoporous Molecular Sieve Catalysts by Feng-wen Yu; Deng-xiang Ji; Yong Nie; Yao Luo; Cheng-jie Huang; Jian-bing Ji (pp. 174-182).

Mesoporous materials possess a hexagonal array of uniform mesopores, high surface areas, and moderate acidity. They are one of the important catalysts in the field of catalytic pyrolysis. In this paper, mesoporous materials of Al-MCM-41, La-Al-MCM-41, and Ce-Al-MCM-41 were synthesized, characterized, and tested as catalysts in the cellulose catalytic pyrolysis process using a fixed bed pyrolysis reactor. The results showed that mesoporous materials exhibited a strong influence on the pyrolytic behavior of cellulose. The presence of these mesoporous molecular sieve catalysts could vary the yield of products, which was that they could decrease the yield of liquid and char and increase the yield of gas product, and could promote high-carbon chain compounds to break into low-carbon chain compounds. Mesoporous molecular sieve catalysts were benefit to the reaction of dehydrogenation and deoxidation and the breakdown of carbon chain. Further, La-Al-MCM-41 and Ce-Al-MCM-41 catalysts can produce more toluene and 2-methoxy-phenol, as compared to the non-catalytic runs.

Keywords: Catalytic pyrolysis; Mesoporous molecular sieve catalysts; Cellulose; Bio-oil

The Effect of Ni²⁺ and Cu²⁺ on the Photocatalytic Degradation of Dyes by the Chitosan–TiO₂ Complex by Xiaolei Zhao; Gang Xiao; Xin Zhang; Haijia Su; Tianwei Tan (pp. 183-197).

The present research combines biosorption and photocatalysis in a functional TiO₂-immobilized chitosan adsorbent (CTA). CTA can degrade organic pollutants and adsorb metal ions simultaneously. Target pollutants were dyes of cationic (rhodamine B, Rh.B) and anionic (methyl orange, MO) nature, with Ni²⁺ and Cu²⁺ selected as heavy metals. The presence of Ni²⁺ or Cu²⁺ improved the degradation ability of CTA for MO, but inhibited the degradation of Rh.B, with Cu²⁺ exhibiting stronger effects than Ni²⁺. There was no significant difference in CTA activity when the metal ions were pre-adsorbed or when they coexisted in the solution with the organic dyes. Protons in the reaction system affected the degradation performance in a similar way for Ni²⁺ and Cu²⁺ leading to a different effect on the degradation for MO and Rh.B. An X-ray photoelectron spectroscopy analysis of the binding energies of the metal ions on the surface in the presence of the cationic or anionic dyes explained the different behaviors. Since anionic and cationic dyes possess chromogenic groups of different charges, they adversely affect the production of OH• radicals when coexisting with Cu²⁺ or Ni²⁺.

Keywords: TiO₂ ; Chitosan; Organic dyes; Metal ions; Adsorption; Photocatalytic degradation

The Effect of Intracellular Amino Acids on GSH Production by High-cell-density Cultivation of Saccharomyces cerevisiae by Miaomiao Wang; Jingfeng Sun; Feiyan Xue; Fei Shang; Zheng Wang; Tianwei Tan (pp. 198-205).

The present paper studies the effects of precursor amino acids, i.e., l-glutamic acid (Glu), l-glycine (Gly), and l-cysteine (Cys), on the glutathione (GSH) production. The three amino acids were added during the fermentations. The GSH production was analyzed by gas chromatography–mass spectrometry (GC–MS). It was observed that the cell content of Cys reduced continually, Gly maintained a fairly constant concentration, while Glu remained at a high concentration compared with Cys and Gly. The synthesis of GSH was found to significantly increase after 28 h of fermentation upon addition of 6 mmol l⁻¹ of each of the three amino acids. Under these conditions, the GSH yields reached 2,250 ± 50 mg l⁻¹ at 34 h from 1,050 ± 50 mg l⁻¹ at 28 h. The GC–MS analyses on the effect caused by the addition of amino acids indicated that the addition of Glu was not necessary to improve the GSH production by high-cell-density cultivation of Saccharomyces cerevisiae. The addition of Cys or Gly individually enhances the production of GSH.

Keywords: Cysteine (Cys); Gas chromatography–mass spectrometry (GC–MS); Glutathione (GSH); Glutamic acid (Glu); Metabolic profiling; Saccharomyces cerevisiae

Decomposition Behavior of Hemicellulose and Lignin in the Step-Change Flow Rate Liquid Hot Water by Xinshu Zhuang; Qiang Yu; Wen Wang; Wei Qi; Qiong Wang; Xuesong Tan; Zhenhong Yuan (pp. 206-218).

Hemicellulose and lignin are the main factors limiting accessibility of hydrolytic enzymes besides the crystallinity of cellulose. The decomposition behavior of hemicellulose and lignin in the step-change flow rate hot water system was investigated. Xylan removal increased from 64.53% for batch system (solid concentration 4.25% w/v, 18 min, 184°C) to 83.78% at high flow rates of 30 ml/min for 8 min, and then 10 ml/min for 10 min. Most of them (80–90%) were recovered as oligosaccharide. It was hypothesized that the flowing water could enhance the mass transfer to improve the sugars recovery. In addition, the solubilization mechanism of lignin in the liquid hot water was proposed according to the results of Fourier transform-infrared spectroscopy and scanning electron microscopy of the water-insoluble fraction and gas chromatography–mass spectrometry of the water-soluble fraction. It was proposed that lignin in the liquid hot water first migrated out of the cell wall in the form of molten bodies, and then flushed out of the reactor. A small quantity of them was further degraded into monomeric products such as vanillin, syringe aldehyde, coniferyl aldehyde, ferulic acid, and p-hydroxy-cinnamic acid. All of these observations would provide important information for the downstream processing, such as purification and concentration of sugars and the enzymatic digestion of residual solid.

Keywords: Decomposition; Hemicellulose; Lignin; Step-change flow rate liquid hot water (SCFLHW); Sweet sorghum bagasse (SSB)

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Applied Biochemistry and Biotechnology: Part A: Enzyme Engineering and Biotechnology (v.168, #1)

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Applied Biochemistry and Biotechnology: Part A: Enzyme Engineering and Biotechnology (v.168, #1)