Orthogonal combination design was adopted in examining the Spirulina platensis (S. platensis) yield and the influence of four factors (Se content, Se-adding method, S content and NaHCO3 content) on algae growth. The results showed that Se content, Se-adding method and NaHCO3 content were key factors in cultivation conditions of Se-enriched S. platensis with the optimal combination being Se at 300 mg/L, Se-adding amount equally divided into three times and NaHCO3 at 16.8 g/L. Algae yield had a remarkable correlation with OD560 and floating rate by linear regression analysis. There was a corresponding relationship between effects of the four factors on algae yield and on OD560, floating rate too. In conclusion, OD560 and floating rate could be served as yield-forming factors.
Bicarbonates ; analysis ; Culture Media ; Cyanobacteria ; growth & development ; Selenium ; analysis ; pharmacology

OBJECTIVETo study the cultural characteristics of Armillaria mellea (A. mellea ) on solid media.

METHODSA. mellea was cultured on semi-solid agar medium in dark conditions. Effects of different media, carbon sources, nitrogen sources, and temperature on growth and morphology of A. mellea were observed. The contents of polysaccharide, mannitol, glucose, and reducing sugars in A. mellea during different stages of development were determined.

RESULTSThe biomass and morphology of A. mellea were different in various media. Sugars were more effective carbon sources than the relevant sugar alcohols. Little molecular carbon sources such as alcohol and glycerol could be utilized by A. mellea, but starch only could be utilized slowly. Either organic or inorganic nitrogen sources could be uptaken and utilized effectively by A. mellea. No evidence was found that VitB1 affects the growth of A. mellea. The growth cycle on wort medium at 30 degrees C was shorter than that at 25 degrees C for 7 days. In logarithmic growth phase and stable phase, the polysaccharide contents of A. mellea were 9.24% and 4.70% respectively, while the mannitol contents were 10.08% and 10.58% respectively; glucose and reducing sugar contents remained low level in the whole growth stage.

CONCLUSIONSCarbon sources have a more remarked effect on the growth of A. mellea than the nitrogen sources do. Optimal temperature for the growth of A. mellea ranges 20-30 degrees C. Mannitol accumulates more than other little molecular carbohydrates in A. mellea.

Agaricales ; chemistry ; growth & development ; Culture Media ; Mannitol ; analysis

The aim of this study is to develop a synthetic medium suitable for 13C metabolic flux analysis (13C-MFA) of Streptomyces rimosus. The cell growth rate and oxytetracycline production by S. rimosus M4018 were compared when M4018 cells were growth on the optimized chemically defined media with organic nitrogen sources or inorganic nitrogen sources. First, a synthetic medium contained KNO3 as the main nitrogen source was screened, then optimized by a response surface method. Using this new medium, the oxytetracycline yield was increased from 75.2 to 145.6 mg/L. Furthermore, based on the 13C-MFA, we identified that Entner-Doudoroff pathway does not exist in S. rimosus cells cultured in a chemically defined medium with feed of 100% 1-13C labeled glucose. This study is helpful for subsequent 13C-MFA application of S. rimosus.
Carbon Isotopes ; analysis ; Culture Media ; chemistry ; Metabolic Flux Analysis ; Nitrogen ; chemistry ; Oxytetracycline ; biosynthesis ; Streptomyces rimosus ; metabolism

Ginseng is a valuable medicinal plant with ginsenosides as its mian effective components. Because ginseng is a perennial plant and has a very strict demand for soil conditions, the way of cultivating ginseng by cutting woods is still used in China at present and thus forest resources has been extremely destroyed. Increasing attention has been paid to the hairy roots induced by the infection of Agrobacterium rhizogenes in the production of plant secondary metabolic products for the hairy roots are characterized by rapid growth and stable hereditary and biochemical traits. That has opened a new way for the industrial production of ginseosides. However, there is little report for such studies from China. In this paper, hairy roots of ginseng were induced from the root explants of two-year-old ginseng by Agrobacterium rhizogenes A4 with directly inoculating. The transformed hairy roots could grow rapidly on MS medium and 1/2 MS medium without hormones. The cultured clones of the hairy roots were established on a solid 1/2 MS medium. After 4 - 5 subcultures the hairy roots still maintained a vigorous growth. A pair of primers were designed and synthesized according to the analytical results of RiA4TL-DNA sequence by Slightom et al . 0.8kb rolC was obtained by PCR using the genome DNA of hairy root of ginseng. Transformation was confirmed by PCR amplification of rolC genes from the hairy roots of P. ginseng. Growth rate of hairy roots on liquid medium increased by 2 times then that of the solid medium. The growth of the hairy roots can be divided into three stages: high speed in the first two weeks, middle speed in the 3 - 4 weeks and low speed hereafter. Changing the culture solution at 2 weeks regular intervals is conductive to maintaining the rapid growth of the hairy roots. By means of determination for specific growth rate and ginsenosides content, the high-yield hairy root clone R9923 was selected. The content of monomer gisenoside of Rg1, Re, Rf, Rbl, Rc, Rb2 and Rd in hairy root clone R9923 was determined by the HPLC. The total ginsenosides content in the hairy toot clone R9923 came up to 15.2 mg/g. The suitable culture conditions for ginseng hairy roots growing were 1/2 MS liquid medium (30 g/L glucose), in a shaker at 110 r/min, changing the culture solution at 2 weeks and subculture time 4 weeks. In the liquid fermented culture of 2L medium, the yield of the hairy roots could amount to 270.10 g in 4 weeks. The industrial production of ginsenosides has been preliminarily realized. Effect factors on biomass and ginsenosides content such as culture volume, inoculation, in steps cultural technology at the scale-up process of hairy roots culture were also explorated. Our results have laid a foundation for defining optimum culture manner for large-scale cultivation and large-scale production of ginsenosides.
Culture Media ; metabolism ; Culture Techniques ; methods ; Glucosides ; analysis ; Panax ; growth & development ; Plant Roots ; growth & development ; Rhizobium ; physiology

OBJECTIVETo conduct the biotransformation of ursolic acid by alternaria longipes AS3. 2875, and optimize the culture medium and biotransformation conditions.

METHODWith the consumption rate of ursolic acid and the generation rate of 28-O-beta-D-glucopyranose ursolic acid as indicators, the impact of different biotransformation conditions such as pH, phosphate, different kinds of metal ions, spore concentration, substrate quantity, temperature, shaking speed and cultivation time on the transformation of ursolic acid in alternaria longipes culture solution were detected to obtain the optimal biotransformation conditions of ursolic acid by alternaria longipes.

RESULTThe optimized biotransformation conditions were as follows: initial pH value was 5.0, MgSO4 was 0.25 g x L(-1) K2HPO4 was 1.0 g x L(-1), FeSO4 was 0.083 g x L(-1), spore concentration was 4%, substrate quantity was 0.3 g x L(-1), shaking speed was 140 r x min(-1), cultivation temperature was at 28 degrees C and cultivating time was 3 days.

CONCLUSIONThe generation rate of 28-O-beta-D-glucopyranose ursolic acid by alternaria longipes stabilizes at around 5%.

Alternaria ; chemistry ; metabolism ; Biotransformation ; Culture Media ; metabolism ; Culture Techniques ; methods ; Hydrogen-Ion Concentration ; Temperature ; Triterpenes ; analysis ; metabolism

Using 50% biogas slurry as basic medium, we investigated the effect of pH on the growth and lipid accumulation of Chlorella vulgaris. Setting two-group experiments, one was only control the initial medium pH, the initial pH was set at 6.0, 6.5, 7.0, 7.5, 8.0, and 8.5, respectively. One was control the medium pH constant, set constant pH at 6.0, 6.5, 7.0, 7.5, 8.0, and 8.5, respectively. Using HCl and NaOH regulated the pH. Results showed that algae Chlorella vulgaris grows better at pH 6.5 and 7.0, accumulate the lipid at pH 7.0-8.5, so the optimal pH for the growth and the lipid accumulation of Chlorella vulgaris was 7.0. The average removal rate of nitrate from biogas slurry was 95%, phosphate was 97%.
Biofuels ; Chlorella vulgaris ; growth & development ; metabolism ; Culture Media ; Culture Techniques ; methods ; Fermentation ; Hydrogen-Ion Concentration ; Lipids ; analysis ; biosynthesis

The novel antimicrobial peptide in submerged fermentation by Bacillus sp. fmbJ224 is strongly influenced by many internal and external factors, namely medium constituents and fermentation conditions. In this study, Plackett-Burman design was undertaken to evaluate the effects of the seventeen factors. By the statistical regression analysis, the significant factors affecting the novel antimicrobial peptide in submerged fermentation by Bacillus sp. fmbJ224 were determined as follows: glucose, NH4NO3, glutamic acid, CaCl2, MnSO4. In the second phase of the optimization process, a response surface methodology (RSM) was used to optimize the above critical internal factors, and to find out the optimization concentraction levels and the relationships between these factors. By solving the quadratic regression model equation using appropriate statistic methods, the optimal concentration of the variables were determined as: 8.13 g/L glucose, 6.14 g/L NH4NO3, 4.2 g/L glutamic acid, 3.98 mg/L CaCl2, 4.87 mg/L MnSO4. The content of the novel antimicrobial peptide was increased from 1304.21 microg/mL to 1487.58 microg/mL. The experimental data under various conditions have validated the theoretical values.
Anti-Infective Agents ; metabolism ; Bacillus ; growth & development ; metabolism ; Cell Culture Techniques ; methods ; Culture Media ; Fermentation ; Peptides ; metabolism ; Regression Analysis

Through scale-up cultivation of Eriobotrya japonica suspension cells using WAVE bioreactor, the cell growth and ursolic acid (UA) accumulation were studied. The comparison test was carried out in the flask and the reactor with cell dry weight (DW) and UA content as evaluation indexes. The culture medium, DW and UA content were compared in 1 L and 5 L working volumes of bioreactor. The orthogonal test with main actors of inoculation amount, speed and angle of rotation was developed to find the optimal combination, in 1 L working volume of bioreactor. DW of the cell growth and the UA content in bioreactor were higher than those of the shaker by 105.5% and 27.65% respectively. In bioreactor, the dynamic changes of elements in the fluid culture, the dry weight of the cell growth and the UA content in 1 L and 5 L working volumes were similar. Inoculation of 80 g, rotational speed of 26 r · min(-1), and angle of 6 ° was the optimal combination, and the cell biomass of 19.01 g · L(-1) and the UA content of 27.750 mg · g(-1) were achieved after 100 h cultivation in 1 L working volume of bioreactor. WAVE Bioreactor is more suitable than flasks for the E. japonica cell suspension culture, and culture parameters can be achieved from 1 L to 5 L amplification.
Biomass ; Bioreactors ; Cell Culture Techniques ; instrumentation ; methods ; Culture Media ; chemistry ; metabolism ; Eriobotrya ; chemistry ; growth & development ; metabolism ; Triterpenes ; analysis ; metabolism

Analysis is carried out on millimeter-wave (MMW) dosimetry in culture dishes used in experiments on MMW biological effects at the cellular level. Finite-difference time-domain (FDTD) technique is employed to compute the 6 mm MMW power density (PD) irradiated into cells in a typical culture dish and the MMW power absorption density (PAD) of cells, followed by the qualitative and quantitative analyses on the outcomes. Indicated in the results, distributions of MMW PD irradiated into cells and the PAD of cells are complicated with evident inhomogeneity so that MMW dosimetry varies a lot in different position whose influence on the experimental outcomes is not neglectable. Consequently, the precise determination of dosimetry is of great importance to be conducted in related experiments.
Cell Culture Techniques ; methods ; Computer Simulation ; Culture Media ; Electromagnetic Fields ; Finite Element Analysis ; Humans ; Radiation Dosage ; Radiometry

Objective: To investigate whether in vitro culture medium (IVCM) for sparse spermatozoa can improve human sperm motility for the purpose of helping clinicians, laboratorians and patients choose a better strategy of assisted reproduction. METHODS: Semen samples were obtained from 178 males for routine semen examination from March to August 2016, including 151 cases of asthenozoospermia and 27 cases of normal sperm motility. A total of 200 μl was collected from each sample and divided into two equal portions and equal volumes of IVCM (experimental group) and F10 (1×) (control group) were added to the two portions, respectively, followed by 30-minute incubation at 37℃ in an incubator with 5% CO2. Sperm concentration, motility and viability and the percentages of progressively motile, non-progressively motile and immotile sperm were recorded before and after incubation. RESULTS: After activated with IVCM, neither the samples with asthenozoospermia nor those with normal sperm motility showed any statistically significant difference in sperm viability from the baseline or the control group (P>0.05). The rates of progressively and non-progressively motile sperm from the asthenozoospermia males were increased by 14.02% and 4.86% respectively, while that of immotile sperm decreased by 19.01% in the experimental group (P >0.01), and similar results were observed in the semen samples from the men with normal sperm motility. The percentage of reduced immotile viable sperm was positively correlated with that of immotile viable sperm in both the asthenozoospermia patients (r = 0.260, P <0.01) and the men with normal sperm motility (r = 0.679, P <0.01). CONCLUSIONS IVCM can increase sperm motility without affecting sperm viability in men with either asthenozoospermia or normal sperm motility. The larger the proportion of immotile viable sperm, the higher the percentages of progressively and non-progressively motile sperm in the semen after IVCM activation, and this correlation is more significant in men with normal sperm motility than in asthenozoospermia patients.
Asthenozoospermia ; physiopathology ; therapy ; Culture Media ; Culture Techniques ; Humans ; Male ; Semen ; Semen Analysis ; methods ; Sperm Count ; Sperm Motility ; Spermatozoa ; physiology