Acetobacter Sp Research Articles

Understanding Kombucha Tea Fermentation: A Review.

Kombucha is a traditional fermented tea whose consumption has increased in the recent years due to its multiple functional properties such as anti-inflammatory potential and antioxidant activity. The microbiological composition of this beverage is quite complex and still more research is needed in order to fully understand its behavior. This study comprises the chemical and microbiological composition of the tea and the main factors that may affect its production.

Enzymatic characterization of a recombinant carbonyl reductase from Acetobacter sp. CCTCC M209061

BackgroundAcetobacter sp. CCTCC M209061 could catalyze carbonyl compounds to chiral alcohols following anti-Prelog rule with excellent enantioselectivity. Therefore, the enzymatic characterization of carbonyl reductase (CR) from Acetobacter sp. CCTCC M209061 needs to be investigated.ResultsA CR from Acetobacter sp. CCTCC M209061 (AcCR) was cloned and expressed in E. coli. AcCR was purified and characterized, finding that AcCR as a dual coenzyme-dependent short-chain dehydrogenase/reductase (SDR) was more preferred to NADH for biocatalytic reactions. The AcCR was activated and stable when the temperature was under 35 °C and the pH range was from 6.0 to 8.0 for the reduction of 4′-chloroacetophenone with NADH as coenzyme, and the optimal temperature and pH were 45 °C and 8.5, respectively, for the oxidation reaction of isopropanol with NAD+. The enzyme showed moderate thermostability with half-lives of 25.75 h at 35 °C and 13.93 h at 45 °C, respectively. Moreover, the AcCR has broad substrate specificity to a range of ketones and ketoesters, and could catalyze to produce chiral alcohol with e.e. >99% for the majority of tested substrates following the anti-Prelog rule.ConclusionsThe recombinant AcCR exhibited excellent enantioselectivity, broad substrate spectrum, and highly stereoselective anti-Prelog reduction of prochiral ketones. These results suggest that AcCR is a powerful catalyst for the production of anti-Prelog alcohols.Graphical abstractThe biocatalytic reactions conducted with the recombinant AcCR

Immobilization of Acetobacter sp. CGMCC 8142 for efficient biocatalysis of 1, 3-propanediol to 3-hydroxypropionic acid

3-hydroxypropionic acid (3-HP) is an important chemical platform organic in material industry, daily chemical industry and biomedicine field due to its numerous valuable derivatives. However, no mature methods have been established in the synthesis industry for direct large scale production. The bacterium Acetobacter sp. CGMCC 8142 with high efficiency of alcohols oxidation property was immobilized for biosynthesis of 3-HP from 1, 3-propanediol (1, 3-PDO). Parameter values in mass transfer modeling indicated that mass transfer of immobilized biocatalysts affected the oxidation reaction (the internal effectiveness factor η i φ > 0.3. The optimal immobilization conditions for 3-HP biocatalysis was sodium alginate 40 g/L, gel beads diameter 1 mm, cross-linkage time 2 h and 0.1mM FeCl2. Immobilized cells showed promising substrate tolerance, pH stability, thermal stability and storability. After 5 cycles of reaction, 3-HP molar yield of immobilized beads was retained to 80.26%, and 66.95 g/L 3-HP were produced from 70 g/L 1, 3-PDO. The biocatalysis process of immobilized cells introduced in this study may provide an economical and efficiency alternative route for practical production of 3-HP.

Engineering of a novel carbonyl reductase with coenzyme regeneration in E. coli for efficient biosynthesis of enantiopure chiral alcohols

The novel anti-Prelog stereospecific carbonyl reductase from Acetobacter sp. CCTCC M209061 was successfully expressed in E. coli combined with glucose dehydrogenase (GDH) to construct an efficient whole-cell biocatalyst with coenzyme NADH regeneration. The enzymatic activity of GAcCR (AcCR with a GST tag) reached 304.9U/g-dcw, even 9 folds higher than that of wild strain, and the activity of GDH for NADH regeneration recorded 46.0U/mg-protein in the recombinant E. coli. As a whole-cell biocatalyst, the recombinant E. coli BL21(DE3)pLysS (pETDuet-gaccr-gdh) possessed a broad substrate spectrum for kinds of carbonyl compounds with encouraging yield and stereoselectivity. Besides, the asymmetric reduction of ethyl 4-chloroacetoacetate (COBE) to optically pure ethyl 4-chloro-3-hydroxybutyrate (CHBE) catalyzed by the whole-cell biocatalyst was systematically investigated. Under the optimal reaction conditions, the optical purity of CHBE was over 99% e.e. for (S)-enantiomer, and the initial rate and product yield reached 8.04μmol/min and 99.4%, respectively. Moreover, the space-time yield was almost 20 folds higher than that catalyzed by the wild strain. Therefore, a new, high efficiency biocatalyst for asymmetric reductions was constructed successfully, and the enantioselective reduction of prochiral compounds using the biocatalyst was a promising approach for obtaining enantiopure chiral alcohols.

Markedly improving asymmetric oxidation of 1-(4-methoxyphenyl) ethanol with Acetobacter sp. CCTCC M209061 cells by adding deep eutectic solvent in a two-phase system.

BackgroundEnantiopure (S)-1-(4-methoxyphenyl) ethanol {(S)-MOPE} can be employed as an important synthon for the synthesis of cycloalkyl [b] indoles with the treatment function for general allergic response. To date, the biocatalytic resolution of racemic MOPE through asymmetric oxidation in the biphasic system has remained largely unexplored. Additionally, deep eutectic solvents (DESs), as a new class of promising green solvents, have recently gained increasing attention in biocatalysis for their excellent properties and many successful examples in biocatalytic processes. In this study, the biocatalytic asymmetric oxidation of MOPE to get (S)-MOPE using Acetobacter sp. CCTCC M209061 cells was investigated in different two-phase systems, and adding DES in a biphasic system was also explored to further improve the reaction efficiency of the biocatalytic oxidation.ResultsOf all the examined water-immiscible organic solvents and ionic liquids (ILs), 1-butyl-3-methylimidazolium hexafluorophoshpate ([C4MIM][PF6]) afforded the best results, and consequently was selected as the second phase of a two-phase system for the asymmetric oxidation of MOPE with immobilized Acetobacter sp. CCTCC M209061 cells. For the reaction performed in the [C4MIM][PF6]/buffer biphasic system, under the optimized conditions, the initial reaction rate, the maximum conversion and the residual substrate e.e. recorded 97.8 μmol/min, 50.5 and >99.9 % after 10 h reaction. Furthermore, adding the DES [ChCl][Gly] (10 %, v/v) to the aqueous phase, the efficiency of the biocatalytic oxidation was rose markedly. The optimal substrate concentration and the initial reaction rate were significantly increased to 80 mmol/L and 124.0 μmol/min, respectively, and the reaction time was shortened to 7 h with 51.3 % conversion. The immobilized cell still retained over 72 % of its initial activity after 9 batches of successive reuse in the [C4MIM][PF6]/[ChCl][Gly]-containing buffer system. Additionally, the efficient biocatalytic process was feasible up to a 500-mL preparative scale.ConclusionThe biocatalytic asymmetric oxidation of MOPE with Acetobacter sp. CCTCC M209061 cells was successfully conducted in the [C4MIM][PF6]-containing biphasic system with high conversion and enantioselectivity, and the reaction efficiency was further enhanced by adding [ChCl][Gly] to the reaction system. The efficient biocatalytic process was promising for the preparation of enantiopure (S)-MOPE.

Characterization of Acetobacter sp. Strain CV1 Isolated from a Fermented Vinegar

Characterization of Acetobacter sp. Strain CV1 Isolated from a Fermented Vinegar

흑찰미 식초 제조를 위한 초산균주 동정 및 품질특성 비교

This study was carried out to investigate the quality characteristics and identification of acetic acid bacteria for black waxy rice vinegar. Eight (8) strains of acetic acid bacteria were isolated for the production of acetic acid and their acidities were then compared with commercial acetic acid bacteria. Among them, F1, H4, and two types of commercial bacteria (four best strains by vinegar zymogen) were selected. After analyzing the 16S rRNA sequence, both F1 and H4 strains were identified as Acetobacter genus. Therefore, the F-1 and H-4 strains were named as Acetobacter sp. F-1 and Acetobacter sp. H-4, respectively. Acidity of black waxy rice vinegar during fermentation was steadily increased up to 16 days and the acidity was then constant. Total acidity content was higher when used FV-1 strain. In the results of Hunter's color value of black waxy vinegar, L value was at 75.01 to 80.11, while (＋a) value was at 3.34 to 3.92, and (＋b) value was at 12.84 to 18.09. The major organic acid of the black waxy vinegar was acetic acid. The total organic acid content was high when used H-4, F-1, C-2 and C-1 strains. The total free amino acid content of the black waxy vinegar by strain was the highest (351.43 mg%) of F-1 vinegar strain, and the lowest (247.74 mg%) of C-2 vinegar strain. A sensory evaluation of black waxy vinegar indicated that F-1 vinegar strain was better than the other samples in aspect of flavor, color, and overall preference.

Asymmetric reduction of ethyl acetoacetate catalyzed by immobilized Acetobacter sp. CCTCC M209061 cells in hydrophilic ionic liquid hybrid system

The asymmetric reduction of ethyl acetoacetate (EAA) to ethyl (R)-3-hydroxybutyrate [(R)-EHB] using immobilized Acetobacter sp. CCTCC M209061 cells was successfully conducted in a hydrophilic ionic liquid (IL)-containing system. The best one of all the tested watermiscible ILs was 1-(hydroxyethyl)-3-methylimidazolium hydrochloride (C2OHMIM·Cl). In C2OHMIM·Cl-aqueous buffer hybrid system, it was found that the optimal IL concentration, substrate and co-substrate concentration, reaction temperature, buffer pH and shaking rate were 0.5mol/L, 45 mmol/L, 80 mmol/L, 35°C, pH 5.5 and 200 rpm, respectively. Under the optimized reaction conditions, the initial reaction rate, the yield and the product e.e. reached 4.90 µmol/min, 95.3 and > 99.0%, respectively, which were much higher than the corresponding values reported previously. The efficient biocatalytic process mediated by the immobilized cells was feasible on 500 mL preparative scale, and the biocatalysts showed good operational stability and could be recycled for at least 10 batches.

Enhancing Asymmetric Reduction of 3-Chloropropiophenone with Immobilized Acetobacter sp. CCTCC M209061 Cells by Using Deep Eutectic Solvents as Cosolvents

Use of deep eutectic solvents (DESs) to improve biocatalytic asymmetric reduction of 3-chloropropiophenone to (S)-3-chloro-1-phenylpropanol catalyzed by whole-cell of Acetobacter sp. CCTCC M209061 was successfully performed. The cells immobilized on PVA-sodium sulfate exhibited markedly enhanced stability. Diverse DESs, as cosolvents, manifested significantly different influences on the reaction. Among them, the DES choline chloride/urea ([ChCl][U]) showed the best biocompatibility and moderately increased the cell member permeability, as demonstrated by MAR and flow cytometry assays, and consequently gave the best results. For the bioreduction conducted in the [ChCl][U]-containing system, the optimum [ChCl][U] content, substrate concentration, glucose concentration, pH and temperature were 5% (v/v), 10.0 mmol/L, 60 mmol/L, 5.5 and 30 °C, respectively. Under the optimized conditions, the obtained yield and product e.e. were 82.3% and above 99.0% at a reaction time of 6 h, respectively, and the productivit...

Using deep eutectic solvents to improve the resolution of racemic 1-(4-methoxyphenyl)ethanol through Acetobacter sp. CCTCC M209061 cell-mediated asymmetric oxidation

The DES [ChCl][Gly] could significantly enhance the efficiency of resolution of racemic MOPE to obtain (S)-MOPE using immobilized Acetobacter sp. CCTCC M209061 cells.

Bacterial cellulose membrane produced by Acetobacter sp. A10 for burn wound dressing applications.

Bacteria cellulose membranes (BCM) are used for wound dressings, bone grafts, tissue engineering, artificial vessels, and dental implants because of their high tensile strength, crystallinity and water holding ability. In this study, the effects of BCM application for 15 days on healing of burn wounds were investigated based on evaluation of skin regeneration and angiogenesis in burn injury skin of Sprague-Dawley (SD) rats. BCM showed a randomly organized fibrils network, 12.13 MPa tensile strength, 12.53% strain, 17.63% crystallinity, 90.2% gel fraction and 112.14 g × m(2)/h highest water vapor transmission rate (WVTR) although their swelling ratio was enhanced to 350% within 24h. In SD rats with burned skin, the skin severity score was lower in the BCM treated group than the gauze (GZ) group at all time points, while the epidermis and dermis thickness and number of blood vessels was greater in the BCM treated group. Furthermore, a significant decrease in the number of infiltrated mast cells and in vascular endothelial growth factor (VEGF) and angiopoietin-1 (Ang-1) expression was observed in the BCM treated group at day 10 and 15. Moreover, a significant high level in collagen expression was observed in the BCM treated group at day 5 compared with GZ treated group, while low level was detected in the same group at day 10 and 15. However, the level of metabolic enzymes representing liver and kidney toxicity in the serum of BCM treated rats was maintained at levels consistent with GZ treated rats. Overall, BCM may accelerate the process of wound healing in burn injury skin of SD rats through regulation of angiogenesis and connective tissue formation as well as not induce any specific toxicity against the liver and kidney.

유기염소계 난분해성 산업폐수의 처리를 위한 미생물제제의 개발

유기염소계 난분해성 산업폐수처리에 효과적인 미생물제제 개발을 위하여 PCP (pentachlorophenol)와 TCE (trichloroethylene) 등과 같은 유기계 염소화합물이 오염되어있는 토양 및 산업폐수로부터 PCP 분해활성이 높은 GP5, GP19와 TCE 분해활성이 높은 GA6, GA15를 분리하였다. 이들 분리균주 GP5, GP19, GA6, GA15등은 Acetobactor sp., Pseudomonas sp., Arthrobacer sp., Xanthomonas sp.과 유사한 것으로 나타나 최종적으로 Acetobacter sp. GP5, Pseudomonas sp. GP19, Arthrobacer sp. GA6, Xanthomoas sp. GA15로 명명하였다. 유기염소계 산업폐수의 처리를 위한 복합미생물제제 OC17은 PCP와 TCE를 분해하는 4개의 분리 분리균주와 방향족화합물 분해균주인 Acinetobacter sp. KN11, Neisseria sp. GN13의 배양액을 혼합하여 제조하였다. 복합미생물제제 OC17은 <TEX>$2.8{\times}10^9CFU/g$</TEX>의 균체수를 갖고 있으며, 밀도는 <TEX>$0.299g/cm^3$</TEX>, 수분함량은 26.8%를 나타내었다. 복합미생물 제제 OC17은 PCP 500 mg/l가 포함되어있는 인공폐수를 이용한 실험에서 배양 65시간에 87%의 분해효율을 나타내었고, TCE (300 uM)의 분해효율은 배양 50시간에 90%의 분해효율을 나타내었다. 복합미생물제제 OC17을 이용한 유기 염소계 산업폐수의 처리효율 시험을 위한 연속배양 실험 에서 10일간 처리 하였을 때 91%의 COD 제거효율을 나타내었다. The screening of the microorganisms degrading chlorinated organic compounds such as PCP (pentachlorophenol) and TCE (trichloroethylene) was conducted with soil and industrial wastewater contaminated with various chlorinated organic compounds. Isolates (GP5, GP19) capable of degrading PCP and isolates (GA6, GA15) capable of degrading TCE were identified as Acetobactor sp., Pseudomonas sp., Arthrobacer sp., Xanthomonas sp. and named Acetobacter sp. GP5, Pseudomonas sp. GP19, Arthrobacer sp. GA6 and Xanthomoas sp. GA15, respectively. The microbial augmentation, OC17 formulated with the mixture of bacteria including isolates (4 strains) degrading chlorinated organic compounds and isolates (Acinetobacter sp. KN11, Neisseria sp. GN13) degrading aromatic hydrocarbons. Characteristics of microbial augmentation OC-17 showed cell mass of <TEX>$2.8{\times}10^9CFU/g$</TEX>, bulk density of <TEX>$0.299g/cm^3$</TEX> and water content of 26.8%. In the experiment with an artificial wastewater containing PCP (500 mg/l), degradation efficiency of the microbial augmentation OC17 was 87% during incubation of 65 hours. The degradation efficiency of TCE (300 uM) by microbial augmentation OC17 was 90% during incubation of 50 hours. In a continuous culture experiment, analysis of the biodegradation of organic compounds by microbial augmentation OC17 in industry wastewater containing chlorinated hydrocarbons showed that the removal rate of COD was 91% during incubation of 10 days. These results indicate that it is possible to apply the microbial augmentation OC17 to industrial wastewaters containing chlorinated organic compounds.

초산균에 의한 환경친화적 기능성소재인 세균 셀룰로오스 생산을 위한 정치배양조건 최적화

Bacterial cellulose (BC) has played important role as new functional material for food industry and industrial products based on its unique properties. The interest in BC from static cultures has increased steadily in recent years because of its potential for use in medicine and cosmetics. In this study, we investigated culture condition for BC production by Acetobacter sp. F15 in static culture. The strain F15, which was isolated from decayed fruit, was selected on the basis of BC thickness. The optimal medium compositions for BC production were glucose 7%, soytone 12%, <TEX>$K_2HPO_4$</TEX> 0.2%, <TEX>$NaH_2PO_4{\cdot}_2H_2O$</TEX> 0.2%, lactic acid 0.05% and ethanol 0.3%, respectively. The strain F15 was able to produce BC at <TEX>$26^{\circ}C-36^{\circ}C$</TEX> with a maximum at <TEX>$32^{\circ}C$</TEX>. BC production occurred at pH 4.5-8 with a maximum at pH 6.5. Under these conditions, a maximum BC thickness of 12.15 mm was achieved after 9 days of cultivation; this value was about 2.3-fold higher than the thickness in basic medium. Scanning electron micrographs showed that BC from the optimal medium was more compact than plant cellulose and was reticulated structure consisting of ultrafine cellulose fibrils. BC from the optimal medium was found to be of cellulose type I, the same as typical native cellulose.

A novel carbonyl reductase with anti-Prelog stereospecificity from Acetobacter sp. CCTCC M209061: purification and characterization.

A novel carbonyl reductase (AcCR) catalyzing the asymmetric reduction of ketones to enantiopure alcohols with anti-Prelog stereoselectivity was found in Acetobacter sp. CCTCC M209061 and enriched 27.5-fold with an overall yield of 0.4% by purification. The enzyme showed a homotetrameric structure with an apparent molecular mass of 104 kDa and each subunit of 27 kDa. The gene sequence of AcCR was cloned and sequenced, and a 762 bp gene fragment was obtained. Either NAD(H) or NADP(H) can be used as coenzyme. For the reduction of 4′-chloroacetophenone, the Km value for NADH was around 25-fold greater than that for NADPH (0.66 mM vs 0.026 mM), showing that AcCR preferred NADPH over NADH. However, when NADH was used as cofactor, the response of AcCR activity to increasing concentration of 4′-chloroacetophenone was clearly sigmoidal with a Hill coefficient of 3.1, suggesting that the enzyme might possess four substrate-binding sites cooperating with each other The Vmax value for NADH-linked reduction was higher than that for NADPH-linked reduction (0.21 mM/min vs 0.17 mM/min). For the oxidation of isopropanol, the similar enzymological properties of AcCR were found using NAD+ or NADP+ as cofactor. Furthermore, a broad range of ketones such as aryl ketones, α-ketoesters and aliphatic ketones could be enantioselectively reduced into the corresponding chiral alcohols by this enzyme with high activity.

Use of Ionic Liquid To Significantly Improve Asymmetric Reduction of Ethyl Acetoacetate Catalyzed by Acetobacter sp. CCTCC M209061 Cells

Biocatalytic reduction of ethyl acetoacetate (EAA) to ethyl (R)-3-hydroxybutyrate [(R)-EHB] with Acetobacter sp. CCTCC M209061 cells was successfully conducted in ionic liquid (IL)-based biphasic systems. Several water-immiscible ILs were used to construct biphasic systems. The best IL investigated was 1-butyl-3-methylimidazolium hexafluorophosphate (C4mim·PF6), which also had good biocompatibility. Several influential variables were examined. The optimum parameters were as follows: volume ratio of buffer to C4mim·PF6, 1/2 (v/v); substrate concentration, 55 mmol/L; buffer pH, 5.5; cosubstrate concentration, 80 mmol/L; reaction temperature, 35 °C; and shaking speed, 220 rpm. Under these optimal conditions, the initial reaction rate, the yield, and the product e.e. were 0.39 mmol/(L min), 90.8%, and >99%, respectively, which were much better than results reported previously. This efficient whole-cell biocatalytic process was feasible on a 450-mL preparative scale, and the immobilized cells showed excellent ...

Acetic acid bacteria in grape must

Acetic acid bacteria (AAB) have undergone continuous taxonomic revision, resulting in an increased number of genera and species ascribed to this group. Thus, the description of the most common AAB in grapes, must and wine has changed dramatically since the initial assessments, which were primarily based on physiological methods. In the present study, we identified the AAB isolated from different grape musts by sequencing the 16S rRNA gene and 16S-23S rRNA gene internal transcribed spacer region. We had previously performed studies from the same winery. However, in this study we now identified AAB species that have been recently described in wines, as well as others that were identified for first time in this niche. &lt;em&gt;Gluconobacter cerinus&lt;/em&gt;, &lt;em&gt;G. japonicus&lt;/em&gt;, &lt;em&gt;G. thailandicus &lt;/em&gt;and &lt;em&gt;G. oxydans&lt;/em&gt; were previously identified as &lt;em&gt;G. oxydans&lt;/em&gt;. &lt;em&gt;Kozakia baliensis&lt;/em&gt; was also within this group. &lt;em&gt;Acetobacter pasteurianus&lt;/em&gt;, &lt;em&gt;A. cerevisiae&lt;/em&gt; and &lt;em&gt;A. malorum&lt;/em&gt; were formerly grouped as &lt;em&gt;Acetobacter sp&lt;/em&gt;. Many isolates previously described as &lt;em&gt;G. oxydans&lt;/em&gt; or &lt;em&gt;A. aceti&lt;/em&gt; likely correspond to other, newly described species of the same genera.

Biocatalytic anti-Prelog stereoselective reduction of ethyl acetoacetate catalyzed by whole cells of Acetobacter sp. CCTCC M209061

The biocatalytic anti-Prelog stereoselective reduction of ethyl acetoacetate (EAA) to ethyl (R)-3-hydroxybutyrate {(R)-EHB} was successfully conducted in the aqueous system using immobilized Acetobacter sp. CCTCC M209061 cells. Among various microorganisms tested, Acetobacter sp. CCTCC M209061 gave the best performance and showed great potential for the bioreduction of EAA to (R)-EHB. Acetobacter sp. CCTCC M209061 cells were immobilized on calcium alginate and then coated with chitosan, and the immobilized cells clearly surpassed the free cells in terms of better thermal stability, storability and recyclability. The optimal co-substrate for the reaction was found to be glucose. The optimal substrate concentration, buffer pH, glucose concentration, reaction temperature, biocatalyst concentration and shaking speed were 35.0mmol/L, 5.5, 80.0mmol/L, 35°C, 0.45g/mL and 200rpm, respectively. Under the optimized conditions, the initial reaction rate, the yield and the enantiomeric excess (e.e.) of the product were 1.28μmol/min, 82.6% and above 99.0%, respectively, which were much higher than those reported previously. The efficient whole-cell biocatalytic process was feasible on a 500-mL preparative scale, and the immobilized cells showed excellent operational stability and could be re-used for at least 10 batches. Additionally, the product e.e. constantly remained above 99.0% regardless of re-use cycles of the biocatalyst.

Using water-miscible ionic liquids to improve the biocatalytic anti-Prelog asymmetric reduction of prochiral ketones with whole cells of Acetobacter sp. CCTCC M209061

The utilization of hydrophilic ionic liquids to improve the synthesis of enantiopure alcohols was successfully performed, via the anti-Prelog asymmetric reduction of ketones with whole cells of Acetobacter sp. CCTCC M209061 newly isolated from Chinese kefir. The best results were obtained with C2OHMIM·NO3, which showed good biocompatibility and also increased moderately cell membrane permeability, thus improving the reaction efficiency. Additionally, the optimal C2OHMIM·NO3 content, buffer pH, reaction temperature and substrate concentration for 4-(trimethylsilyl)-3-butyn-2-one reduction to (R)-4-(trimethylsilyl)-3-butyn-2-ol were 10.0% (v/v), 5.0, 30 °C and 12 mM, respectively. Under the optimized conditions, the initial reaction rate, the maximum yield and the product e.e. were 14.0 μmol/min·gcell, 91%, and >99%, respectively, which were much better than the results reported previously. The efficient whole-cell biocatalytic process proved to be feasible on a 400-mL preparative scale, and the immobilized cells still retained above 88.0% of their original activity after successive re-use for 10 batches, showing the good operational stability in the presence of C2OHMIM·NO3. Furthermore, the established biocatalytic system with Acetobacter sp. CCTCC M209061 and C2OHMIM·NO3 was shown to be highly effective for the anti-Prelog asymmetric reduction of other aryl ketones to the corresponding (R)-alcohols.

Cellulose synthesized by Enterobacter sp. FY-07 under aerobic and anaerobic conditions

Enterobacter sp. FY-07 can produce bacterial cellulose (BC) under aerobic and anaerobic conditions. In static cultivation at 30°C for 72h under anoxic, oxygen-limited and aerated conditions, cellulose production exceeded 5g/l, which indicated that oxygen was not essential for production of BC by Enterobacter sp. FY-07. X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR) and scanning electron microscopy (SEM) analysis showed that the microstructure of the BC was similar to that produced by aerobic bacteria such as Gluconacetobacter xylinum BCRC12335 and Acetobacter sp. V6. The crystallinity index of the BC was 63.3%. Water-holding capacity (approximately 11000%) and rehydration ratio (24.4%) were superior to those reported for BC produced by the aerobic bacteria G. xylinum BCRC12335 and Acetobacter sp. V6. These results will facilitate static submerged fermentation for the production of BC.

Immobilization of Acetobacter sp. CCTCC M209061 for efficient asymmetric reduction of ketones and biocatalyst recycling

BackgroundThe bacterium Acetobacter sp. CCTCC M209061 is a promising whole-cell biocatalyst with exclusive anti-Prelog stereoselectivity for the reduction of prochiral ketones that can be used to make valuable chiral alcohols such as (R)-4-(trimethylsilyl)-3-butyn-2-ol. Although it has promising catalytic properties, its stability and reusability are relatively poor compared to other biocatalysts. Hence, we explored various materials for immobilizing the active cells, in order to improve the operational stability of biocatalyst.ResultsIt was found that Ca-alginate give the best immobilized biocatalyst, which was then coated with chitosan to further improve its mechanical strength and swelling-resistance properties. Conditions were optimized for formation of reusable immobilized beads which can be used for repeated batch asymmetric reduction of 4′-chloroacetophenone. The optimized immobilized biocatalyst was very promising, with a specific activity of 85% that of the free-cell biocatalyst (34.66 μmol/min/g dw of cells for immobilized catalyst vs 40.54 μmol/min/g for free cells in the asymmetric reduction of 4′-chloroacetophenone). The immobilized cells showed better thermal stability, pH stability, solvent tolerance and storability compared with free cells. After 25 cycles reaction, the immobilized beads still retained >50% catalytic activity, which was 3.5 times higher than degree of retention of activity by free cells reused in a similar way. The cells could be recultured in the beads to regain full activity and perform a further 25 cycles of the reduction reaction. The external mass transfer resistances were negligible as deduced from Damkohler modulus Da < <1, and internal mass transfer restriction affected the reduction action but was not the principal rate-controlling step according to effectiveness factors η < 1 and Thiele modulus 0.3<∅ <1.ConclusionsCa-alginate coated with chitosan is a highly effective material for immobilization of Acetobacter sp. CCTCC M209061 cells for repeated use in the asymmetric reduction of ketones. Only a small cost in terms of the slightly lower catalytic activity compared to free cells could give highly practicable immobilized biocatalyst.