Xinqing Zhao

Prospective and development of butanol as an advanced biofuel.

Butanol has been acknowledged as an advanced biofuel, but its production through acetone-butanol-ethanol (ABE) fermentation by clostridia is still not economically competitive, due to low butanol yield and titer. In this article, update progress in butanol production is reviewed. Low price and sustainable feedstocks such as lignocellulosic residues and dedicated energy crops are needed for butanol production at large scale to save feedstock cost, but processes are more complicated, compared to those established for ABE fermentation from sugar- and starch-based feedstocks. While rational designs targeting individual genes, enzymes or pathways are effective for improving butanol yield, global and systems strategies are more reasonable for engineering strains with stress tolerance controlled by multigenes. Compared to solvent-producing clostridia, engineering heterologous species such as Escherichia coli and Saccharomyces cerevisiae with butanol pathway might be a solution for eliminating the formation of major byproducts acetone and ethanol so that butanol yield can be improved significantly. Although batch fermentation has been practiced for butanol production in industry, continuous operation is more productive for large scale production of butanol as a biofuel, but a single chemostat bioreactor cannot achieve this goal for the biphasic ABE fermentation, and tanks-in-series systems should be optimized for alternative feedstocks and new strains. Moreover, energy saving is limited for the distillation system, even total solvents in the fermentation broth are increased significantly, since solvents are distilled to ~40% by the beer stripper, and more than 95% water is removed with the stillage without phase change, even with conventional distillation systems, needless to say that advanced chemical engineering technologies can distil solvents up to ~90% with the beer stripper, and the multistage pressure columns can well balance energy consumption for solvent fraction. Indeed, an increase in butanol titer with ABE fermentation can significantly save energy consumption for medium sterilization and stillage treatment, since concentrated medium can be used, and consequently total mass flow with production systems can be reduced. As for various in situ butanol removal technologies, their energy efficiency, capital investment and contamination risk to the fermentation process need to be evaluated carefully.

Yeast flocculation: New story in fuel ethanol production

Yeast flocculation has been used in the brewing industry to facilitate biomass recovery for a long time, and thus its mechanism of yeast flocculation has been intensively studied. However, the application of flocculating yeast in ethanol production garnered attention mainly in the 1980s and 1990s. In this article, updated research progress in the molecular mechanism of yeast flocculation and the impact of environmental conditions on yeast flocculation are reviewed. Construction of flocculating yeast strains by genetic approach and utilization of yeast flocculation for ethanol production from various feedstocks were presented. The concept of self-immobilized yeast cells through their flocculation is revisited through a case study of continuous ethanol fermentation with the flocculating yeast SPSC01, and their technical and economic advantages are highlighted by comparing with yeast cells immobilized with supporting materials and regular free yeast cells as well. Taking the flocculating yeast SPSC01 as an example, the ethanol tolerance of the flocculating yeast was also discussed.

Current progress and future prospect of microalgal biomass harvest using various flocculation technologies

Microalgae have been extensively studied for the production of various valuable products. Application of microalgae for the production of renewable energy has also received increasing attention in recent years. However, high cost of microalgal biomass harvesting is one of the bottlenecks for commercialization of microalgae-based industrial processes. Considering harvesting efficiency, operation economics and technological feasibility, flocculation is a superior method to harvest microalgae from mass culture. In this article, the latest progress of various microalgal cell harvesting methods via flocculation is reviewed with the emphasis on the current progress and prospect in environmentally friendly bio-based flocculation. Harvesting microalgae through bio-based flocculation is a promising component of the low-cost microalgal biomass production technology.

Bioflocculant production from Solibacillus silvestris W01 and its application in cost-effective harvest of marine microalga Nannochloropsis oceanica by flocculation

Microalgae are widely studied for biofuel production, however, current technologies to harvest microalgae for this purpose are not well developed. In this work, a bacterial strain W01 was isolated from activated sludge and identified as Solibacillus silvestris. Bioflocculant in the culture broth of W01 showed 90% flocculating efficiency on marine microalga Nannochloropsis oceanica, and no metal ion was required for the flocculation process. Chemical analysis of the purified bioflocculant indicated that it is a proteoglycan composed of 75.1% carbohydrate and 24.9% protein (w/w). The bioflocculant exhibits no effect on the growth of microalgal cells and can be reused to for economical harvesting of N. oceanica. This is the first report that strain of S. silvestris can produce bioflocculant for microalgae harvest. The novel bioflocculant produced by W01 has the potential to harvest marine microalgae for cost-effective production of microalgal bioproducts.

Impact of zinc supplementation on the improvement of ethanol tolerance and yield of self-flocculating yeast in continuous ethanol fermentation

The effects of zinc supplementation were investigated in the continuous ethanol fermentation using self-flocculating yeast. Zinc sulfate was added at the concentrations of 0.01, 0.05 and 0.1 g l(-1), respectively. Reduced average floc sizes were observed in all the zinc-supplemented cultures. Both the ethanol tolerance and thermal tolerance were significantly improved by zinc supplements, which correlated well with the increased ergosterol and trehalose contents in the yeast flocs. The highest ethanol concentration by 0.05 g l(-1) zinc sulfate supplementation attained 114.5 g l(-1), in contrast to 104.1 g l(-1) in the control culture. Glycerol production was decreased by zinc supplementations, with the lowest level 3.21 g l(-1), about 58% of the control. Zinc content in yeast cells was about 1.4 microMol g(-1) dry cell weight, about sixfold higher than that of control in all the zinc-supplemented cultures, and close correlation of zinc content in yeast cells with the cell viability against ethanol and heat shock treatment was observed. These studies suggest that exogenous zinc addition led to a reprogramming of cellular metabolic network, resulting in enhanced ethanol tolerance and ethanol production.

Characterization of flocculating agent from the self-flocculating microalga Scenedesmus obliquus AS-6-1 for efficient biomass harvest

In the present work, the extracellular biopolymers from the self-flocculating microalga Scenedesmus obliquus AS-6-1 were studied. It was revealed that the self-flocculation of the microalgal cells was mediated by cell wall-associated polysaccharides with a molecular weight of 127.9 kDa. Sugar compositions analysis indicated that the monomers consist of glucose, mannose, galatose, rhamnose and fructose with the molar ratio of 8:5:3:2:1. Addition of 0.6 mg/L purified flocculating agent resulted in the fast flocculation of freely suspended cells of S. obliquus and Chlorella vulgaris. The flocculating activity is stable between pH 6 and 8 and at 20-60°C.

Establishment of an efficient genetic transformation system in Scenedesmus obliquus

Scenedesmus obliquus belongs to green microalgae, which is attracting attention as a feedstock for biofuels production and biorefinery as well as in bioremediation of environmental pollutants, making its genetic modifications for more efficient growth and accumulation of aimed metabolites significant. However, the genetic transformation system of S. obliquus is still not well established. In the current work, S. obliquus was transformed via electroporation using a plasmid containing chloramphenicol resistance gene (CAT) as a selectable marker and the green fluorescent protein gene (gfp) as a reporter. Using the optimized transformation conditions, the transformation efficiency was 494±48 positive transgenic clones per 10(6) recipient cells, which is more efficient comparing with those reported in other microalgal transformation studies. Green fluorescence was observed after six months of cultivation, and CAT-specific products were also detected in the transformants by PCR, Southern blot and RT-PCR analysis. This is the first report on establishing such an efficient and stable transformation system for S. obliquus, a prerequisite for both functional genomic studies and strain improvement for other biotechnology applications of this important microalgal species.

Ethanol fermentation with Kluyveromyces marxianus from Jerusalem artichoke grown in salina and irrigated with a mixture of seawater and freshwater

Aims:  To study fuel ethanol fermentation with Kluyveromyces marxianus ATCC8554 from Jerusalem artichoke (Helianthus tuberosus) grown in salina and irrigated with a mixture of seawater and freshwater.

Streptomyces xinghaiensis sp. nov., isolated from marine sediment

A novel actinomycete, strain S187(T), was isolated from a marine sediment sample collected from Xinghai Bay, Dalian, China. Growth occurred on ISP medium 2 containing 0-9 % NaCl and at pH 6.0-9.0 and 10-45 degrees C. The cell wall of strain S187(T) contained the isomer ll-diaminopimelic acid as the diagnostic diamino acid. The predominant menaquinones were MK-9(H(6)) (40.8 %), MK-9(H(8)) (38.2 %) and MK-9(H(2)) (8.8 %). The major fatty acids were iso-C(16 : 0) (29.6 %), anteiso-C(15 : 0) (14.0 %) and anteiso-C(17 : 0) (11.6 %). Cells contained phosphatidylethanolamine, phosphatidylinositol, phosphatidylglycerol, phosphatidylinositol mannosides and one unknown phospholipid. The G+C content of the genomic DNA was 72.01 mol%. The 16S rRNA gene sequence of the isolate had similarities of 98.1 and 97.5 % with those of Streptomyces flavofuscus NRRL B-8036(T) (=DSM 41426(T)) and Streptomyces albiaxialis DSM 41799(T), respectively, showing that the novel strain should be assigned to the genus Streptomyces. DNA-DNA hybridizations with the two above-mentioned Streptomyces species showed 31.4 and 46.9 % relatedness, respectively. Moreover, the three strains differed in some physiological and biochemical properties. Thus, on the basis of phenotypic and genotypic analyses, it is proposed that strain S187(T) represents a novel species of the genus Streptomyces, for which the name Streptomyces xinghaiensis sp. nov. is proposed; the type strain is S187(T) (=NRRL B-24674(T)=CCTCC AA 208049(T)=KCTC 19546(T)).

Zinc and yeast stress tolerance: Micronutrient plays a big role

Zinc is an essential element for the normal growth, metabolism and physiology of yeast. Besides acting as a cofactor for many enzymes, zinc is also required for the structural stability of zinc finger proteins, many of which exert important controls on cellular metabolic processes. Although the effect of zinc on ethanol fermentation has previously been reported, the role of zinc on cellular stress tolerance is not well studied. In this review, the latest progress regarding the important roles of zinc and zinc containing proteins in yeast metabolism and cellular function, especially the recent findings demonstrating the involvement of zinc finger proteins in cellular stress responses, are summarized, with the focus on stress tolerance of yeast in the context of fuel ethanol production. An in-depth understanding of the status of zinc in the reprogramming of yeast metabolic network is vital for the breeding of robust yeast strains for ethanol fermentation, as well as for improved production efficiency of yeast-based bioindustry.