Efficient production of ε-poly-L-lysine from cassava bagasse hydrolysate used as carbon source by Streptomyces albulus US3-18.

The production of ε-poly-L-lysine (ε-PL) from cassava bagasse hydrolysate (CBH) by Streptomyces albulus US3-18 was investigated in this study. With 30 g/L glucose from CBH, 1.30 g/L ε-PL and 10.68 g/L biomass were obtained in shake flask fermentation. Interestingly, the two values were increased by 14.0% and 21.5%, respectively, compared to the control (1.14 g/L and 8.79 g/L). Simultaneously, the activities of four key enzymes of ε-PL synthesis during CBH fermentation were enhanced to varying degrees. In batch fermentation of 5-L bioreactor, 3.39 g/L ε-PL and 10.17 g/L DCW were harvested with 40 g/L glucose from CBH. The combination of fed-batch fermentation with two-stage pH strategy significantly increased ε-PL titer and biomass to 37.41 g/L and 41.0 g/L, respectively. Moreover, eleven volatile components were detected in CBH by GC-MS, and 6-pentyl-α-pyrone (6PP) was first identified as the most abundant volatile ingredient. The results in CBH fermentation demonstrated that S. albulus US3-18 exhibited high tolerance to these volatile byproducts. Using ICP-MS, the calcium concentration in CBH was determined as 195.0 mg/(kg hydrolyzate), and cobalt, copper, lead, chromium, mercury and arsenic were not detected. By adding 0.05 g/L CaCl2 to M3G medium, ε-PL yield was improved by 28.0%, indicating calcium was one of the factors for the enhanced ε-PL production. The study provides a reference for the efficient production of ε-PL from low-cost agricultural residues.

Enhancement of erythritol production by Trichosporonoides oedocephalis ATCC 16958 through regulating key enzyme activity and the NADPH/NADP ratio with metal ion supplementation.

Erythritol, a well-known natural sweetener, is mainly produced by microbial fermentation. Various metal ions (Al3+, Cu2+, Mn2+, and Ni2+) were added to the culture medium of Trichosporonoides oedocephalis ATCC 16958 at 30 mg/L in shake flask cultures. Compared with controls, Cu2+ increased the erythritol content by 86% and decreased the glycerol by-product by 31%. After 48 hr of shake flask culture, sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that expression levels of erythrose reductase (ER) in the presence of 30 mg/L CuSO4 · 5H2O were higher than those obtained after treatment with other examined metal ions. Furthermore, after 108 hr of batch culture in a 5-L bioreactor, supplementation with 30 mg/L of CuSO4 · 5H2O increased the specific erythritol content by 27%. Further studies demonstrated that ER activity under 30 mg/L CuSO4 · 5H2O supplementation in a fermentor was overtly increased compared with the control after 60 hr, while glycerol-3-phosphate dehydrogenase activity was clearly reduced in most of the fermentation process. Furthermore, the NADPH/NADP ratio was slightly lower in T. oedocephalis cells treated with Cu2+ compared with control cells. These results provide further insights into Cu2+ effects on erythritol biosynthesis in T. oedocephalis and should improve the industrial production of erythritol by biological processes.

Transformation of the yeast Trichosporonoides oedocephalis.

The osmotolerant yeast, Trichosporonoides oedocephalis, is an excellent producer of erythritol, which has wide industrial applications. In this study, we developed an efficient transformation method for T. oedocephalis. To evaluate the T. oedocephalis transformation, we constructed a DNA fragment (loxP-Kan-loxP/Cre system) that was targeted to the mitogen-activated protein kinase HOG1 gene. Transformants were selected on plates containing G418 and response surface methodology was employed to obtain optimum transformation conditions. Optimal transformation could be achieved at an incubation time of 40 min, when the concentration of zymolyase-100T was 30 µg/mL, and when 100 mM CaCl2 was added to the mixture. The predicted optimal transformation efficiency was 133 transformants per µg of DNA. This novel method will facilitate studies in synthetic biology and metabolic engineering of T. oedocephalis.

[Study of cortex phellodendri chinensis decoction experiment based on the spectral imaging technology].

The purpose of decoction traditional Chinese medicine is to make full exhalation of medicinal materials active ingredients, thus it has the maximum effect of traditional Chinese medicine to treat disease. In order to detect the dissolution change of medicinal materials active ingredients in decoction process, this paper applys spectral imaging technology, with Chinese traditional medicine cortex phellodendri as an example, discussing its fluorescence intensity at different time in decoction process. And the analysis results reflect edgewise the dissolution rule of cortex phellodendri active ingredients.

Optoelectronic Properties of α-MoO3 Tuned by H Dopant in Different Concentration.

The optoelectronic properties of layered α-MoO3 are greatly limited due to its wide band gap and low carrier concentration. The insertion of hydrogen (H) can effectively tune the band structure and carrier concentration of MoO3. Herein, first-principles calculations were performed to unravel the physical mechanism of a H-doped α-MoO3 system. We found that the modulation of the electronic structure of H-doped MoO3 depends on the doping concentration and position of the H atoms. It was found that the band gap decreases at 8% doping concentration due to the strong coupling between Mo-4d and O-2p orbits when H atoms are inserted into the interlayer. More interestingly, the band gap decreases to an extreme due to the Mo-4d orbit when all the H atoms are inserted into the intralayer only, which has a remarkable effect on light absorption. Our research provides a comprehensive theoretical discussion on the mechanism of H-doped α-MoO3 from the doping positions and doping concentrations, and offers useful strategies on doping modulation of the photoelectric properties of layered transition metal oxides.

Perspectives of biotechnological production of L-ribose and its purification.

L-ribose is a non-natural and expensive sugar that can be used as an important intermediate for the synthesis of L-nucleoside analogues, which are used as antiviral drugs. In contrast to chemical production, biotechnological methods can produce L-ribose from biomass under environmentally friendly conditions. In this mini-review, various strategies for synthesizing L-ribose by applying microorganisms and their enzymes are discussed, including microbial biotransformation and biocatalysis by engineering bacteria. Furthermore, subsequent isolation-and-purification techniques, as an integral step in the whole process, are accordingly described, containing the special introduction of a promising strategy of L-ribose separation. Particularly, further researches and outlook for the improvement of L-ribose preparation was solely stressed. Compared with each method, this mini-review provides a panorama of respective advantages and disadvantages existing in them.

[The research on active constituent distribution of rhizoma coptidis pieces].

In order to test the distribution of active constituent of traditional Chinese medicine and to evaluate the quality of medicinal part effectively, spectral imaging analysis technology was used, and rhizoma coptidis pieces were tested as an example. First, the fluorescence spectral cube was taken, and the spectral curve of 3 different medicinal parts of the piece was obtained; second, spectral images were reconstructed by principal components analysis method, and the differences of 3 medicinal parts on the first few principal components were focused; third, the first component image was divided by the threshold method, then the distribution and relative content of 3 medicinal parts were obtained. The results show that spectral imaging analysis technology can provide the distribution of the active constituent, which can be used as the criterion of selecting medicinal parts. The testing course is nondestructive and rapid.

Automatic spatial filtering to obtain the virtual image term in digital holographic microscopy.

The main factor that limits the quality of an image reconstructed by the process of spatial filtering in digital holographic microscopy is discussed. A spatial filter determined by the distribution of the spectrum of the virtual image is designed automatically for real time dynamic analysis of a micro-object, and an optimal reconstructed phase image can be obtained. An experiment of a holographic image with an onion specimen is presented to prove the validity of this approach. Comparing the numerical reconstruction of the hologram by employing different spatial filters with the automatic spatial filtering shows the superiority of the automatic spatial filtering method, and it is suitable for dynamic and automatic analysis.

Inactivation Mechanism of 1-Ethyl-3-Methylimidazolium-Based Ionic Liquid on ß-Glucosidase Produced by Paenibacillus sp. LLZ1 and Enhanced Activity Using a Surfactant.

ß-Glucosidase (BG) hydrolyzes cellobiose into glucose, and is a vital step in converting ionic liquids (ILs)-pretreated biomass to sustainable biofuels. The inactivation mechanism of BG from Paenibacillus sp. LLZ1 induced by microcrystalline cellulose was explored in various concentrations of ILs, composed of [Emim]+ cation and [DEP]-, [OAc]-, [Br]-, [Cl]-, and [BF4]- anions. The FTIR analysis of inactivated BG indicated that the ILs altered its ß-sheet content. Moreover, circular dichroism spectroscopy (CD) suggested that the α-helix content decreased, while the ß-sheet content increased with the presence of ILs in general. Interestingly, the secondary structure of BG had almost no change after [Emim]DEP treatment, while ionic liquid [Emim]BF4 treatment caused the irreversible denaturation of BG. Eventually, by adding 0.4 mM of Aerosol OT surfactant, the BG activity was increased by 20.1% in the presence of 25% [Emim]DEP, and the corresponding glucose yield from hydrolysis of cellobiose was increased by 23.9%.

Phase reconstruction of digital holography with the peak of the two-dimensional Gabor wavelet transform.

We describe a numerical reconstruction technique for digital holography by means of the two-dimensional Gabor wavelet transform (2D-GWT). Applying the 2D-GWT to digital holography, the object wave can be reconstructed by calculating the wavelet coefficients of the hologram at the peak of the 2D-GWT automatically. At the same time the effect of the zero-order diffraction image and the twin image are eliminated without spatial filtering. Comparing the numerical reconstruction of a holographic image by the analysis of the one-dimensional Gabor wavelet transform (1D-GWT) with the 2D-GWT, we show that the 2D-GWT method is superior to the 1D-GWT method, especially when the fringes of the hologram are not just along the y direction. The theory and the results of a simulation and experiments are shown.

Digital reconstruction based on angular spectrum diffraction with the ridge of wavelet transform in holographic phase-contrast microscopy.

A numerical reconstruction technique of digital holography based on angular spectrum diffraction by means of the ridge of Gabor wavelet transform (GWT) is presented. Appling the GWT, the object wave can be reconstructed by calculating the wavelet coefficients of the hologram at the ridge of the GWT automatically even if the spectrum of the virtual image is disturbed by the other spectrum. It provides a way to eliminate the effect of the zero-order and the twin-image terms without the spatial filtering. In particular, based on the angular spectrum theory, GWT is applied to the digital holographic phase-contrast microscopy on biological specimens. The theory, the results of a simulation and an experiment of an onion specimen are shown.

A solid ionic Lactate biosensor using doped graphene-like membrane of Au-EVIMC-titania nanotubes-polyaniline.

Herein, a doped graphene-like membrane was designed, which was copolymerized to be a solid ionic biosensor by using titania nanotubes (TiNTs), polyaniline (PANI), EvimCl (1-ethyl-3-vinylimidazolium chloride, EVIMC) and chloroauric acid (HAuCl4). The structure of graphene-like arrangement and the copolymerization mechanism of the film were discussed in detail. Because of the high catalytic property, Lactate could be determined on the membrane catalyzed by lactate dehydrogenase (LDH) containing isocitrate dehydrogenase (NAD+), which was immobilized onto the film by electrostatic attraction. The electrochemical response on the LDH/Au-EVIMC-TiNTs-PANI/ITO was increased by twice than the LDH/TiNTs/PANI/ITO, and exhibited two linear responses within the concentration range from 5.5 × 10-7 M to 5.55 × 10-6 M, and 5.55 × 10-6 M to 3.33 × 10-3 M, with a detection limit of 1.65 × 10-7 M (S / N = 3). The interference study for other common coexistence such as uric acid and hemoglobin revealed that there was no overlapping signal for the detection of Lactate on the biosensor. The developed method proved the most stability in the determination of real blood samples, and the recoveries ranged from 96.7% to 105.8% with a satisfactory result.

Characterization of Ribose-5-Phosphate Isomerase B from Newly Isolated Strain Ochrobactrum sp. CSL1 Producing L-Rhamnulose from L-Rhamnose.

In this study, we attempted to find new and efficient microbial enzymes for producing rare sugars. A ribose-5-phosphate isomerase B (OsRpiB) was cloned, overexpressed, and preliminarily purified successfully from a newly screened Ochrobactrum sp. CSL1, which could catalyze the isomerization reaction of rare sugars. A study of its substrate specificity showed that the cloned isomerase (OsRpiB) could effectively catalyze the conversion of L-rhamnose to L-rhamnulose, which was unconventional for RpiB. The optimal reaction conditions (50°C, pH 8.0, and 1 mM Ca2+) were obtained to maximize the potential of OsRpiB in preparing L-rhamnulose. The catalytic properties of OsRpiB, including Km, kcat, and catalytic efficiency (kcat/Km), were determined as 43.47 mM, 129.4 sec-1, and 2.98 mM/sec. The highest conversion rate of L-rhamnose under the optimized conditions by OsRpiB could reach 26% after 4.5 h. To the best of our knowledge, this is the first successful attempt of the novel biotransformation of L-rhamnose to L-rhamnulose by OsRpiB biocatalysis.

Improved in situ saccharification of cellulose pretreated by dimethyl sulfoxide/ionic liquid using cellulase from a newly isolated Paenibacillus sp. LLZ1.

A cellulase producing strain was newly isolated from soil samples and identified as Paenibacillus sp. LLZ1. A novel aqueous-dimethyl sulfoxide (DMSO)/1-ethyl-3-methylimidazolium diethyl phosphate ([Emin]DEP)-cellulase system was designed and optimized. In the pretreatment, DMSO was found to be a low-cost substitute of up to 70% ionic liquid to enhance the cellulose dissolution. In the enzymatic saccharification, the optimum pH and temperature of the Paenibacillus sp. LLZ1 cellulase were identified as 6.0 and 40°C, respectively. Under the optimized reaction condition, the conversion of microcrystalline cellulose and bagasse cellulose increased by 39.3% and 37.6%, compared with unpretreated cellulose. Compared to current methods of saccharification, this new approach has several advantages including lower operating temperature, milder pH, and less usage of ionic liquid, indicating a marked progress in environmental friendly hydrolysis of biomass-based materials.

Construction of an Efficient Mutant Strain of Trichosporonoides oedocephalis with HOG1 Gene Deletion for Production of Erythritol.

The mitogen-activated protein kinase HOG1 (high-osmolarity glycerol response pathway) plays a crucial role in the response of yeast to hyperosmotic shock. Trichosporonoides oedocephalis produces large amounts of polyols (,e.g., erythritol and glycerol) in a culture medium. However, the effects of HOG1 gene knockout and environmental stress on the production of these polyols have not yet been studied. In this study, a To-HOG1 null mutation was constructed in T. oedocephalis using the loxP-Kan-loxP/Cre system as replacement of the targeted genes, and the resultant mutants showed much smaller colonies than the wild-type controls. Interestingly, compared with the wild-type strains, the results of shake-flask culture showed that To-HOG1 null mutation increased erythritol production by 1.44-fold while decreasing glycerol production by 71.23%. In addition, this study investigated the effects of citric acid stress on the T. oedocephalis HOG1 null mutants and the wild-type strain. When the supplementation of citric acid in the fermentation medium was controlled at 0.3% (w/v), the concentration of erythritol produced from the wild-type and To-HOG1 knockout mutant strains improved by 18.21% and 21.65%, respectively.

Anti-proliferative effects of Atractylis lancea (Thunb.) DC. via down-regulation of the c-myc/hTERT/telomerase pathway in Hep-G2 cells.

Atractylis lancea (Thunb.) DC. (AL), an important medicinal herb in Asia, has been shown to have anti-tumor effects on cancer cells, but the involved mechanisms are poorly understood. This study focused on potential effects and molecular mechanisms of AL on the proliferation of the Hep-G2 liver cancer cell line in vitro. Cell viability was assessed by MTT test in Hep-G2 cells incubated with an ethanol extract of AL. Then, the effects of AL on apoptosis and cell cycle progression were determined by flow cytometry. Telomeric repeat amplification protocol (TRAP) assays was performed to investigate telomerase activity. The mRNA and protein expression of human telomerase reverse transcriptase (hTERT) and c-myc were determined by real-time RT-PCR and Western blotting. Our results show that AL effectively inhibits proliferation in Hep-G2 cells in a concentration- and time-dependent manner. When Hep-G2 cells were treated with AL after 48h,the IC50 was about 72.1 µg/ mL. Apoptosis was induced by AL via arresting the cells in the G1 phase. Furthermore, AL effectively reduced telomerase activity through inhibition of mRNA and protein expression of hTERT and c-myc. Hence, these data demonstrate that AL exerts anti-proliferative effects in Hep-G2 cells via down-regulation of the c-myc/hTERT/ telomerase pathway.

Digital holographic microscopy by use of surface plasmon resonance for imaging of cell membranes.

A technique called surface plasmon resonance digital holographic microscopy (SPRDHM) for optical imaging of cell membranes is proposed. The intensity and phase distributions of the reflected light that is modulated by the cell membrane in surface plasmon resonance can be simultaneously obtained. The imaging principle and capability are theoretically analyzed and demonstrated by experiments. In addition, the technique is compared with total internal reflection digital holographic microscopy (TIRDHM) in theory and experiment, respectively. The results show that the SPRDHM technique is better in spatial resolution and phase sensitivity than the TIRDHM technique for imaging of cell membranes.