Effects of culture condition and nutrition on the co-production of microbial oil and exopolysaccharide by Sporidiobolus pararoseus JD-2

Microbial oil has been gaining considerable attention from researchers recently as renewable and ecofriendly oil and its potential as feedstock for food industry and biodiesel industry. In this context, we have earlier demonstrated production of microbial oil and exopolysaccharide (EPS) from the yeast Sporidiobolus pararoseus JD-2. In this study, we explored increasing its production by optimizing the culture condition and nutrition. As expected, culture temperature and dissolved oxygen (DO) are the contributing factors for co-producing microbial oil and EPS, in which 28? and lower quantum (i.e., 30 mL/500 mL) show the best conditions in shake-flasks fermentation. By contrast, the initial pH from 4 to 8 has no obvious effect on producing microbial oil and EPS. In addition, the culture nutrition (i.e., carbon/nitrogen source) were also discussed, and indicating that 20 g/L of corn steep liquor and 60 g/L of glucose are beneficial to produce microbial oil and EPS (i.e., 34.1±1.2 g/L and 11.5±0.2 g/L, respectively). Meanwhile, the residue glucose should be maintained at 20 g/L, in which the highest production of microbial oil and EPS was obtained (i.e., 34.6±1.7 g/L and 11.7±0.8 g/L, respectively). The biomass, microbial oil and EPS were further increased during optimizing the DO level, which reached to 67.8±2.1 g/L, 34.7±0.6 g/L and 11.8±0.5 g/L during maintaining DO level at 20-30%, respectively. The results suggest that appropriate culture condition and nutrition considerably improve the fermentation performance of S. pararoseus JD-2 and significantly increase co-production of microbial oil and EPS (by 11.2 and 8.3%, respectively) compared to the un-optimized fermentation.

The experimental model of intestinal epithelial cell growth established by real time cell analysis / 中国药理学通报

Aim To observe the effect of epidermal growth factor (EGF) on cell growth under different culture conditions using the real-time cell analyzer and EGF as a tool medicine to promote cell growth,and to provide reference for establishing pharmacokinetic model of IEC-6 cell growth (proliferation).Methods IEC-6 cell was inoculated on E-Plate 16 plate at a density of 1 × 104 cell/well and cultured in DMEM with 10％ serum for 24 h,then replaced by serum-free DMEM culture (serum starvation)for 20 h,then the effects of different culture conditions on cell growth as well as EGF efficacy were observed.Results ① When the serum concentration was 10％,the cell growth index of EGF group(1,10,100 μg · L-1) after drug administration 24 h,48 h and 72 h was P ＞ 0.05 compared with the blank group,suggesting that 10％ serum culture could not reflect the efficacy of EGF.② When the serum concentration was 0％,EGF (1,10 μg · L-1) improved cell growth inhibition caused by serum-free cultivation,but could not recover it to normal level (the EGF group after drug administration 24 h,48 h and 72 h was P ＜0.01 compared with 5％ serum),which suggested that serum-free culture could not reflect the EGF efficacy.③ 0％,0.5％,1％ serum had different effects on cell growth,of which 0.5％ serum could neither have obvious inhibition on cell growth,nor reflect the EGF effect due to promoting cell growth for a long time.④When the serum concentration was 0.5 ％,the cell growth index of EGF groups after drug administration 24h,48h and 72h was P ＜0.01 compared with the blank group,suggesting that 0.5％ serum culture could better reflect EGF efficacy.⑤The efficacy of EGF (10 μg· L-1) in promoting cell growth was confirmed by repeated validation of 0.5 ％ serum.Condusions A reference scheme of the IEC-6 cell growth (proliferative) pharmacological experimental model is established in the real-time cell analyzer:cells are cultured in DMEM with 10％ serum for 24h,then in serum-free DMEM (serum starvation) for 20h,then after the adding of reagent,cells are cultured in DMEM with 0.5％ serum for 48-72 h to observe its effect on cell growth (proliferation).

Clean-Up Human Embryonic Stem Cell Lines Using Humanized Culture Condition

Human embryonic stem cell (hESC) culture system has been changing culture conditions from conventional to xeno-free for therapeutic cell applications, and N-glycolylneuraminic acid (Neu5Gc) could be a useful indicator of xenogeneic contaminations in hESCs because human cells can no longer produce it genetically. We set up the humanized culture condition using commercially available humanized materials and two different adaptation methods: sequential or direct. SNUhES4 and H1 hESC lines, previously established in conventional culture conditions, were maintained using the humanized culture condition and were examined for the presence of Neu5Gc. The hESCs showed the same morphology and character as those of the conventional culture condition. Moreover, they were negative for Neu5Gc within two passages without loss of pluripotency. This study suggested that this method can effectively cleanse previously established hESC lines, bringing them one step closer to being clinical-grade hESCs.

Exploration of the Most Suitable Condition for Culturing Influenza Virus on Plateau Region / 现代检验医学杂志

Objective To explore the most suitable condition of influenza virus which was culturing on plateau region,and im-prove the effect of the separation of influenza virus.Methods The original specimens were respective inoculated in MDCK cells,by comparing CO2 percent concentration (5%,4.8%,4.6%,4.4%,4.2%),TPCK-pancreatic enzyme dosage (1,2,3, 4,5μg/m and 6,8,10μg/ml),inoculation amount (100,200,300,400,500μl),incubation time (48,72,96,120,144 h),inoc-ulation methods (adsorption,direct inoculated),and culture vessel(cells,6 orifice)that were influenced on influenza virus, the best culture conditions was determined.Hemagglutinatio (HA)method was used to detect the virus titers.Results Through the comparison,the HA titer of virus was in the highest titer with 6 orifice plate culture vessel,4.4% CO2 and 4.6% CO2 ,2μg/ml TPCK-pancreatic enzyme,300μl and 400μl inoculation amount,72h~96h of incubation time.Conclusion The optimal condition of the influenza virus cultured with the MDCK cell has been established in the laboratory on plateau region.

Microfluidic chip for tumor cell 3 D culturing establishment and its cultural conditions / 中国药理学通报

Aims To design and fabricate the 3 D cell cultural microfluidic chip for tumor cell culturing, with which to research the compatible conditions for gelatin forming and dissolving with calcium alginate as the scaffolds. To culture SMMC-7721 cells in the chip and to detect the surviving rate. Methods The microfluid-ic chip was fabricated with the software Corel Draw, the technology of soft lithography, molding, and plas-ma bonding. The applicability was tested and cells were cultured on it, on which the cell status was ob-served, their surviving rate was calculated with the help of the software IPP. Results The chip we fabri-cate was calculated is suitable for cell 3D culturing, the tumor cells showed a favorable proliferation ability in 72 h on chip, the surviving rate was ( 96. 1 ± 4. 5)%. The cells were solid and TCS appeared. Con-clusion The microfluidic chip manufactured appeared for tumor cell 3D culture, is suitable for the growth of SMMC-7721 and the cells are indubitable. They show some different status in proliferative and agminated compared with traditional 2D cell culture. With the chip and the condition found, there will be a better way to study the characteristics of tumor cells and is beneficial to the screen of anti-tumor drugs.

The Efficacy and Safety of Collagen-I and Hypoxic Conditions in Urine-Derived Stem Cell Ex Vivo Culture

Upper urinary tract-derived urine stem cells (USCs) are considered a valuable mesenchymal stem cell source for autologous cell therapy. However, the reported culture condition for USCs is not appropriate for large-quantity production, because cells can show limited replicativity, senescence, and undesirable differentiation during cultivation. These drawbacks led us to reconstitute a culture condition that mimics the natural stem cell niche. We selected extracellular matrix protein and oxygen tension to optimize the ex vivo expansion of USCs, and compared cell adhesion, proliferation, gene expression, chromosomal stability, differentiation capacity, immunity and safety. Culture on collagen type I (ColI) supported highly enhanced USC proliferation and retention of stem cell properties. In the oxygen tension analysis (with ColI), 5% O₂ hypoxia showed a higher cell proliferation rate, a greater proportion of cells in the S phase of the cell cycle, and normal stem cell properties compared to those observed in cells cultured under 20% O₂ normoxia. The established reconstituted condition (ColI/hypoxia, USCs(recon)) was compared to the control condition. The expanded USCs(recon) showed highly increased cell proliferation and colony forming ability, maintained transcription factors, chromosomal stability, and multi-lineage differentiation capacity (neuron, osteoblast, and adipocyte) compared to the control. In addition, USCs(recon) retained their immune-privileged potential and non-tumorigenicity with in vivo testing at week 8. Therefore, reconstituted condition allows for expanded uUSC cell preparations that are safe and useful for application in stem cell therapy.

Effect of different culture conditions on protocorm-like bodies of Dendrobium candidum in bioreactor culture / 中草药

Objective: To explore the effect of different culture conditions on protocorm-like bodies (PLBs) of Dendrobium candidum in bioreactor culture. Methods: Inoculation quantity, ventilation, sucrose concentration, and light intensity were taken to carry out the single factor experiment design. Biomass was determined by drying method, polysaccharide content by phenol-sulfuric acid method, and SPSS16.0 software was used for data analysis. Results: Under the bioreactor culture, the nutrient medium components for the PLBs proliferation of D. candidum and polysaccharide accumulation were 1/2 MS medium supplemented with 1.0 mg/L NAA, 5% coconutmilk, and 3% sucrose, pH value was 6.0. The culture conditions, such as inoculation with 40 g/L, 15 μm pore porous nozzle, aeration volumes by 1.0 and 0.5 L/min used alternately, and photon flux of 2000 lx could improve the proliferation coefficient and polysaccharide contents of PLBs. Conclusion: Different culture conditions have the significant effect on the growth of PLBs and the accumulation of polysaccharide. Suitable culture conditions have the very important practic significance to the biomass of PLBs and main medicinal ingredient production of PLBs.

Hiperidricidade: uma desordem metabólica / Hyperhydricity: a metabolic disorder

A hiperidricidade, anteriormente chamada vitrificação, é considerada uma desordem fisiológica, bioquímica e morfológica decorrente do acúmulo anormal de água no interior das células e tecidos. As plantas cultivadas in vitro estão, indubitavelmente, sob contínua condição de estresse, os quais resultam em alterações metabólicas características do estresse oxidativo. Anatomicamente, plantas ou brotos afetados frequentemente apresentam-se inchados, com coloração verde claro, folhas translúcidas e com aparência de vidro, baixa relação número de células/área celular e hipolignificação. Alterações fisiológicas que ocorrem nas principais vias metabólicas, incluindo fotossíntese, respiração e transpiração, resultam em redução de eficiência dessas vias metabólicas. Os distúrbios morfológicos, fisiológicos e bioquímicos são desencadeados por fatores físicos, relacionados ao ambiente dos recipientes de cultivo e consistência do meio de cultura ou por fatores químicos como os componentes do meio de cultura, em especial dos reguladores de crescimento em altas concentrações. A hiperidricidade ocorre em vários níveis de severidade, chegando a resultar na perda irreversível da capacidade morfogênica e o estabelecimento de um estado neoplásico das células, no entanto, na maioria dos casos, a hiperidricidade é considerada reversível. Esta revisão foca o conhecimento atual sobre o fenômeno da hiperidricidade abordando aspectos morfológicos, fisiológicos, bioquímicos e a reversibilidade do processo.

The hyperhydricity, formerly called vitrification, is considered a physiological, biochemistry and morfologic disorder due to abnormal accumulation of water inside the cells and tissues. Plants grown in vitro are undoubtedly under continuous stress condition which results in metabolic changes characteristic of oxidative stress. Anatomically plants or shoots affected often become swollen, with pale green, translucent sheets, glass-like, low relative number of cells / cell area and hipolignification. Physiological changes occur in major metabolic pathways including photosynthesis, respiration and transpiration resulting in reduced efficiency of these metabolic pathways. Morphological, physiological and biochemical disorders are triggered by physical factors related to the environment of cultivation vessels and consistency of the culture medium or by chemical factors such as culture medium components, especially the growth regulators in high concentrations. The hyperhydricity occurs at various levels of severity, reaching result in irreversible loss of morphogenic capacity and the establishment of a state of neoplastic cells, however, in most cases hyperhydricity is considered reversible. This review focuses on the current knowledge about the phenomenon of hyperhydricity addressing morphological, physiological, biochemical, and reversibility of the process.

Rapid propagation of Schisandra chinensis / 中草药

Objective: To develop a rapid propagation of Sehisandra chinensis. Methods: The tissue culture of S. chinensis was studied by orthogonal test and the media for the rapid propagation of S. chinensis were optimized. Results: When NAA 0.3, ZT 0.1, and 6-BA 1.5 mg/L were added in the MS, the best rate of bud proliferation was obtained, while the NAA 0.2 mg/L was added, the best rooting was got. The optimal cultural contion is mat temperature is 25 °C, light intensity 2 000 lx, and photoperiod 12 h. Conclusion: The optimal media and cultural condition in the test could be used to improve the yield and output, as well as quality of S. chinensis.

Culture Conditions for Mycelial Growth of Coriolus versicolor

Coriolus versicolor, is one of the most popular medicinal mushrooms due its various biologically active components. This study was conducted to obtain basic information regarding the mycelial culture conditions of C. versicolor. Based on the culture, and MCM media were suitable for the mycelial growth of the mushroom. The optimum carbon and nitrogen sources were dextrin and yeast extract, respectively, and the optimum C/N ratio was 10 to 2 when 2% glucose was used. Other minor components required for optimal growth included thiamine-HCl and biotin as vitamins, succinic acid, lactic acid and citric acid as organic acids, as well as MgSO4.7H2O as mineral salts.

Culture conditions for the production of a-galactosidase by Aspergillus parasiticus MTCC-2796: a novel source / Condiciones del cultivo para la producción de a-galactosidasa por Aspergillus parasiticus MTCC-2796: una nueva fuente

Aspergillus parasiticus microbial type culture collection (MTCC)-2796, a new source of a-galactosidase is an efficient producer of enzyme in basic medium under submerged fermentation conditions. Maximum a-galactosidase production (156.25 Uml-1) was obtained when the basic medium is supplemented with galactose (0.5 percent w/v) and raffinose (0.5 percent w/v) as carbon source and yeast extract as nitrogen source. Enzyme production was also enhanced considerably in the presence of wheat bran (1.0 percent w/v). Enzyme secretion was strongly inhibited by the presence of Hg2+, Cu2+, and Co2+ in the medium and to some extent by Zn2+ and Ni2+, while marginal increase in the enzyme production was observed when Mg2+ and Mn2+ were added in the medium. Among amino acids checked (aparagine, cysteine, glutamine, leucine and proline), glutamine (1 mM) was found to be an enhancer for the enzyme production. The temperature and pH range for the production of enzyme were 25ºC to 35ºC and 6.5 to 7.5, respectively with maximum activity (50 Uml-1) at 30ºC and pH 6.5 under static fermentation condition.

Culture Conditions for the Mycelial Growth of Ganoderma applanatum

Ganoderma applanatum is one of the most popular medicinal mushrooms due to the various biologically active components it produces. This study was conducted to obtain basic information regarding the mycelial culture conditions of Ganoderma applanatum. Based on the colony diameter and mycelial density, PDA, YMA and MCM media were suitable for the mycelial growth of the mushroom. The optimum temperature for mycelial growth was found to be 25~30degrees C. The optimum carbon and nitrogen sources were mannose and dextrin, respectively, and the optimum C/N ratio was 2 to 10 when 2% glucose was used. Other minor components required for the optimal growth included thiamine-HCl and biotin as vitamins, succinic acid and lactic acid as organic acids, and MgSO4.7H2O, KH2PO4 and NaCl as mineral salts.

Optimal Culture Condition for Antifungal Susceptibility Tests of Malassezia globosa / 대한의진균학회지

BACKGROUND: Although numerous culture conditions for Malassezia species were suggested, there were not so many objective evaluation articles in the literature. OBJECTIVES: We examined the various culture conditions for Malassezia globosa. METHODS: Malassezia globosa culture conditions were assessed by Dixon's agar, modified Leeming-Notman medium in diverse oil content and temperature conditions. RESULTS: Maximum growth rate of Malassezia globosa was achieved at 3% olive oil. The optimal temperatures for the maximal growth of M. globosa were observed at 32~34degrees C. CONCLUSION: In this study, we established the optimal culture condition for M. globosa, and confirmed its excellent utility for the antifungal susceptibility tests for M. globosa and M. restricta. Our results can help the investigators plan to do the prospective researches involving Malassezia species, such as the susceptibility test for newly developed antifungal agents.

Favorable Culture Conditions for Mycelial Growth of Korean Wild Strains in Ganoderma lucidum

Ganoderma lucidum (Fr.) Karst (Polyporaceae), belonging to basidiomycota, is one of the most famous medicinal mushrooms. This study was carried out to investigate favorable mycelial growth conditions, such as pH, temperature, growth media, carbon sources and nitrogen sources of Korean strains in G. lucidum. The most suitable temperature for the mycelial growth was obtained at 30degrees C. In general, optimal temperature range for the mycelial growth was found at 25~30degrees C. This Mushroom has a broad pH range (5~9) for its mycelial growth and mostly favorable growth was found at pH 5. Generally, Hamada, Glucose peptone, YM, Mushroom complete and Lilly media were the most suitable for the mycelial growth of G. lucidum. Among 10 different carbon sources, dextrin, galactose and fructose were best but the rest of other carbon sources also facilitated the growth of mycelia. The most suitable nitrogen sources were ammonium acetate, glycine, arginine and calcium nitrate, but to a certain extent, all of the supplemented nitrogen sources also stimulated the mycelial growth.

Effect of Culture Conditions on the Protein Fibril Expression of Candida albicans

Candida albicans is an important human pathogen that causes systemic infections, predominantly among population with weakened immune system. Cell wall structures of C. albicans are important to adhere to host tissue and evade to host immune system. Among cell wall structure, the outermost fibrillar layer of C. albicans is of interest since it may play important roles in antigenicity, phagocytosis, and adherence. The expression of virulent factors could be affected by the growth conditions. The dynamic nature of the cell surface alters the physical properties of the fungal interface with host cells and thereby influences adhesion to the host and recognition by components of the host immune system. In this study, we investigated the effects of culture conditions on cell surface fibril expression of C. albicans by a transmitting electron microscopy and SDS-PAGE. The protein fibril of C. albicans was expressed in the presence of whole serum, however, the fibril expression was decreased in 25% serum and serum containing 1% glucose. Also, germ tube can be induced by serum, RMPI medium, N-acetyl glucosamine, and 39 degrees C culture condition, hence, the fibrillar structure of C. albicans was detected only in serum-induced germ tube. The expression of fibril layer and the major fibril proteins of 66, 47, 30 kDa were reduced as increasing cell concentration of intial inoculum from 2x10(7) cells/ml to 8x10(7) cells/ml. The fibrillar layer of C. albicans was expressed in serum early within 10 min, and the thickness of fibril layer was increased according to the increase of culture time. When the fibrillar proteins were analysed by SDS-PAGE, major protein of 30 kDa was maintained continuously during over night culture although expression of the other proteins were various. These results suggest that expression of serum induced protein fibril is influenced by culture conditions and is not related to hyphal transition of C. albicans.

The Culture Conditions for the Mycelial Growth of Phellinus spp

Phellinus genus belonged to Hymenochaetaceae of Basidiomycetes and has been well known as one of the most popular medicinal mushrooms due to high antitumor activity. This study was carried out to obtain the basic information for mycelial culture conditions of Phellinus linteus, P. baumii, and P. gilvus. According to colony diameter and mycelial density, the media for suitable mycelial growth of them were shown in MEA, glucose peptone, and MCM. The optimum temperature for mycelial growth was 30degrees C. Carbon and nitrogen sources were mannose and malt extract, respectively. The optimum C/N ratio was 10 : 1 to 5 : 1 with 2% glucose concentration, vitamin was thiamine-HCl, organic acid was succinic acid, and mineral salt was MgSO4.7H2O.

Cell suspension culture of Gentiana macrophylla(Ⅰ) / 中草药

Objective To establish cell suspension culture system of Gentiana macrophylla.Methods Good cell line was selected by the methods of small cell aggregate and the effects of different media,inoculation age of cells,initial pH value,culture temperature,and rotation speed on cell growth and gentiopicroside accumulation were investigated.Results The eligible cell suspension culture conditions were:MS media as the basic medium,cells with inoculation age of 15 d,initial pH 7,and rotation speed of 110 r/min at 25 ℃,culturing in Erlenmeyer flasks.Conclusion Optimal cell suspension culture system of G.macrophylla has been established by shake flasks.The different effects of media on cell growth and gentiopicroside accumulation indicate that some inorganic ions have significant effects on regulation.

Optimum culture media and liquid conditions in shaking flasks for laccase production by Ganoderma lucidum / 中草药

Objective To study the optimum culture media and liquid conditions in shaking flasks for laccase production by Ganoderma lucidum. Methods Taking S_3 stain of G. lucidum as test materials and laccase activity of G. lucidum as measurement index to optimize the culture media and liquid conditions through orthogonal test. Results The optimum culture components of media were: Glucose 30 g/L, cotton 0.2%, (NH_4)_2HPO_4 0.66 g/L, casein 0.5%, Tween-80 0.15 mL; the optimum conditions were: initial pH value of medium was 5.5, 75 mL medium was in 250 mL-flask, inoculation was 12.5%, mycelium age was 5?24 h for 9?24 h culture. Conclusion Laccase activity of G. lucidum has been improved remarkably in the optimum culture media and liquid conditions.

Conditions for Pseudomonas sp.W2 Culture and Bisphenol A Degradation / 环境与健康杂志

Objective To study the condition for Pseudomonas sp. W2 culture and degradation of bisphenol A(BPA). Methods The growth curve of Pseudomonas sp. W2 and BPA degradation curve were made concerned pH, temperature, aeration, carbon sources and nitrogen sources affecting Pseudomonas sp. W2 culture and BPA degradation. Results Under the conditions of pH=7, 25-35 ℃, certain carbon sources and nitrogen sources, Pseudomonas sp. W2 grew well and showed a good efficiency of BPA degradation. Conclusion Some wide conditions can meet the requirement of Pseudomonas sp. W2 growth and BPA degradation.

Preliminary Research on Conditions of Toxins Produced by Pestalotiopsis photiniae / 微生物学通报

Pestalotiopsis photiniae is one of the predominant pathogens of strawberry root rot disease. Based on preliminary research, it was proved that crude toxins were main pathogenic substances of the pathogen. For further investigation and utilization of toxins produced by this fungus, conditions of producing toxins were analyzed with the leaf disk method in this experiment. The result showed that pH values, cultural time, vibration, and tested temperatures obviously affected the production of toxins, except for light treatment. The most suitable culture conditions for the toxin production were pH 6.2, 25?C, darkness and stillness, for 5 d～7 d. Besides, it was discovered that crude toxins could significantly inhibit seed germination and elongation growth of roots or shoots for maize, rye and mung bean.