Peptone Concentration Research Articles

Effects of Hydrogen Peroxide on Different Toxigenic and Atoxigenic Isolates of Aspergillus flavus.

Drought stress in the field has been shown to exacerbate aflatoxin contamination of maize and peanut. Drought and heat stress also produce reactive oxygen species (ROS) in plant tissues. Given the potential correlation between ROS and exacerbated aflatoxin production under drought and heat stress, the objectives of this study were to examine the effects of hydrogen peroxide (H2O2)-induced oxidative stress on the growth of different toxigenic (+) and atoxigenic (−) isolates of Aspergillus flavus and to test whether aflatoxin production affects the H2O2 concentrations that the isolates could survive. Ten isolates were tested: NRRL3357 (+), A9 (+), AF13 (+), Tox4 (+), A1 (−), K49 (−), K54A (−), AF36 (−), and Aflaguard (−); and one A. parasiticus isolate, NRRL2999 (+). These isolates were cultured under a H2O2 gradient ranging from 0 to 50 mM in two different media, aflatoxin-conducive yeast extract-sucrose (YES) and non-conducive yeast extract-peptone (YEP). Fungal growth was inhibited at a high H2O2 concentration, but specific isolates grew well at different H2O2 concentrations. Generally the toxigenic isolates tolerated higher concentrations than did atoxigenic isolates. Increasing H2O2 concentrations in the media resulted in elevated aflatoxin production in toxigenic isolates. In YEP media, the higher concentration of peptone (15%) partially inactivated the H2O2 in the media. In the 1% peptone media, YEP did not affect the H2O2 concentrations that the isolates could survive in comparison with YES media, without aflatoxin production. It is interesting to note that the commercial biocontrol isolates, AF36 (−), and Aflaguard (−), survived at higher levels of stress than other atoxigenic isolates, suggesting that this testing method could potentially be of use in the selection of biocontrol isolates. Further studies will be needed to investigate the mechanisms behind the variability among isolates with regard to their degree of oxidative stress tolerance and the role of aflatoxin production.

Optimization of medium components and physicochemical parameters to simultaneously enhance microbial growth and production of lypolitic enzymes by Stenotrophomonas sp.

The optimization of lipase and esterase production (LP and EP) and bacterial growth (BG) of a Stenotrophomonas sp. strain was developed. For this purpose, the effect of five different medium components and three physicochemical parameters were evaluated using a Plackett-Burman statistical design. Among eight variables, stirring speed, pH, and peptone concentration were found to be the most effective factors on the three responses under evaluation. An optimization study applying Box-Behnken response surface methodology was used to study the interactive effects of the three selected variables on LP/EP and microorganism growth. Predicted models were found to be significant with high regression coefficients (90%-99%). By using the desirability function approach, the optimum condition applying simultaneous optimization of the three responses under study resulted to be: stirring speed of 100rpm, pH of 7.5, and a peptone concentration of 10g/L, with a desirability value of 0.977. Under these optimal conditions, it is possible to achieve in the optimized medium a 15-fold increase in esterase productivity, a 117-fold increase in lipase production, and a 9-log CFU/mL increase in BG, compared with the basal medium without agitation.

Conditions for supplemental biogenic substrates to enhance activated sludge degradation of xenobiotic.

The effects of biogenic presence on the degradation of xenobiotic organics by natural microbial populations have been reported as either advantageous or disadvantageous. The inconsistency of the reports implies there could be a turning point from disadvantageous to advantageous outcomes so that conditions may exist that could bring an optimum advantage. This study tested the supplementations of varying concentrations of sucrose and peptone, separately and combined, to acclimated activated sludge degradation of xenobiotic 2,4-D, while other operational and microbiological conditions were held constant. Our test results showed that biogenic may indeed enhance or slow down xenobiotic degradation rates. The highest enhancements exist at concentrations of 50 and 80 mg/L, respectively, for sucrose and peptone when supplemented separately, and 20 mg/L sucrose and 40 mg/L peptone combined. Conditions for advantageous biogenic supplementation were identified for activated sludge degradation of a xenobiotic; specifically, the highest degradation rate enhancements occurred when biogenic supplementation was approximately 0.5 to 0.7 the concentration of 2,4-D base on chemical oxygen demand (COD), which brought a biomass yield of approximately double that yielded by 2,4-D. Kinetics analyses provided clues for the possible causes of advantageous and disadvantageous effects due to biogenic supplementation.

A central composite design for the optimizing lipase and protease production from Bacillus subtilis PTCC 1720

Abstract In the present study, the producing condition of lipase and protease from Bacillus subtilis PTCC 1720 was optimized by response surface methodology (RSM). A central composite design (CCD) was used to evaluate the interactive effect of the enzyme production conditions on the lipase activity (LA), protease activity (PA) and biomass of B. subtilis PTCC 1720. Independent variables including temperature, sucrose, peptone, and corn oil concentration were studied. According to the results, the optimal enzyme production condition was achieved with temperature of 43.4 °C, sucrose of 20 g/L, peptone of 20 g/L, and corn oil of 4 mL/L. In this conditions the LA, PA, and biomass were predicted 3.16 U/mL, 1744.6 U/mL, and 3.03 g/L, respectively. The experimental and predicted values of the LA, PA, and biomass show a good agreement with one another, with a high degree of accuracy of the models.

전통 장류 유래 Bacillus subtilis SCJ4의 특성확인 및 통계학적 방법을 이용한 배양조건 최적화

strains isolated from Korean traditional fermented food in Sunchang and their investigated biochemical characterization and ability of biogenic amines non-producing. We selected the SCJ4 having various activity by measurement of extracellular enzyme, antioxidant and antimicrobial activities. Selected strain SCJ4 by 16S rRNA sequencing and biochemical characterization was named Bacillus subtilis SCJ4. And then, we investigated cell growth of SCJ4, and optimized of culture medium constituents using response surface methodology as statistically method. Response surface methodology used Plackett-Burman experimental design for screening of medium constituent. Tryptone, peptone and MgSO4 as medium constituent improving cell growth selected. In order to find out optimal concentration on each constituent, we carried out central composite design. Consequently, optimized concentrations of tryptone, peptone and MgSO4 were predicted to be 15.35 g/L, 12.235 g/L, and 3.5 g/L respectively. Through the model verification, we confirmed about 1.28-fold improvement of the dried cell weight from 0.8767 g/L to 1.1222 g/L when compared to basal medium.

Response Surface Methodology for the Optimization of Kojic Acid Production by Aspergillus Flavus Using Muntingia Calabura Fruits as a Carbon Source

The current study focused on the optimization of kojic acid production by a soil isolated fungal organism Aspergillus flavus through Surface Fermentation (SF) by utilizing a novel carbon source i.e. ripened fruits of Muntingia calabura L. Initially the most significant 7 physico-chemical factors were studied for the optimization process through One-Factor-At-A-Time Method (OFAT). Among these 5 influential factors were screened and five-factor-three-level Central Composite Design (CCD) matrix and Response Surface Methodology (RSM) was performed to enhance the production rate of kojic acid. Through one-factor-at-a-time method the optimal values were Substrate concentration 100g/L, Peptone concentration 4g/L, KH 2 PO 4 concentration 2g/L, MgSO 4 concentration 0.7g/L, pH 6.0, Time 28d and Temperature 30°C and the maximum kojic acid production was 85.1g/L. The maximum production obtained at the predicted optimal conditions through response surface methodology was Substrate concentration 100g/L, Peptone concentration 4g/L, pH 6.0, Time 28d and Temperature 29°C and the maximum kojic acid production was 88.8 g/L. These results directs to the success of the model in improving a process for the production of kojic acid, a significant organic acid with vast industrial importance.

Response Surface Methodology for the Optimization of Proteases Production by a Novel Egyptian Isolate Bacillus amyloliquefaciens 35s

Aims: The present work aimed to optimize proteases production by Bacillus amyloliquefaciens 35s using the response surface methodology (RSM). Study Design: Variables affecting proteases production were screened using a Plackett–Burman design. Face Centered Central Composite Design (FCCCD) of RSM was adopted for the augmentation of total proteases production assessed at three coded levels (–1, 0, +1). All obtained data were analyzed by ANOVA with post hoc multiple comparison analysis performed using Tukey’s HSD. Place and Duration of Study: Department of Agricultural Microbiology, Faculty of Agriculture, Ain Shams University, between March 2014 and September 2014. Methodology: Bacillus amyloliquefaciens 35s was used for proteases production. Modified TGY (Tryptone gluscose yeast extarct) medium was the basal medium. Impacts of nutritional factors (carbon and nitrogen and mineral salts) were studied using Plackett-Burman design with fold over augmenting method. “Design Expert ® 8.0.7.1” Stat-Ease was used to analyze the experimental Original Research Article Nassar et al.; BMRJ, 6(5):255-269, 2015; Article no.BMRJ.2015.078 256 Plackett–Burman design. Temperature, pH and agitation rate (using shake flask) were optimized statistically by the factorial FCCCD of the RSM. Validation of statistical model of physical factors was done by carrying out the experiment at optimum conditions of the process parameters as determined from the model. Optimum conditions obtained through RSM in terms of FCCCD were examined and verified in a 5 L bench top continuous stirred tank bioreactor and production process was scaled-up in a batch process with controlled and non-controlled pH. Fermented medium was centrifuged to collect cells and determination of biomass and protease concentration. Results: Among the significant media components, peptone and starch showed to have significant effects on the response as for protease production, with confidence level > 98% and were further optimized using FCCCD. Conditions promoted proteases production were different from those enhanced cell growth. Physical parameters indicated that production of proteases by Bacillus amyloliquefaciens is non-growth dependent. Maximum proteases production predicted (992.12 u/ml) was observed near the mid-point (0) values (concentrations) of both peptone (10 g/l) and starch (10 g/l) and the experimental value 935 u/ml was very close to the predicted value validating the model. The final proteases production in the bioreactor reached 1530 u/ml obtained within 12-14 h at 0.6 vvm aeration and 120 rpm of agitation speed. Conclusion: Instead of conventional method of one variable at time approach, Response Surface Methodology, as statistical approach, showed to be adequate and efficient to optimize protease production by Bacillus amyloliquefaciens.

Optimization of Cultural Parameters for Cost Effective Production of Kojic Acid by Fungal Species Isolated from Soil

A novel isolate capable of producing opulent yields of kojic acid with surface fermentation was screened from ten different isolated soil fungi. The organism was confirmed as Aspergillus flavus FJ537130 strain using 18SrRNA based molecular analysis and it was identified as a negative producer of aflatoxin. Though twelve different carbon sources were used, significant yields of kojic acid crystals was noticed with Sago starch. The optimized conditions established were substrate concentration 1000 ml (100 g of starch powder in 1000 ml of H2O), pH 6.0, Time 28 d, Temperature 28°C, Peptone concentration 4 g/l, KH2PO4 concentration 1 g/l, MgSO4 concentration 0.5 g/l and the yield achieved was 28.5 g/l. The resulted fermentation broth was subjected to solvent extraction followed by gel filtration for the separation of kojic acid and crystallization. The structural characterization of purified kojic acid was confirmed by Proton NMR, FTIR and XRD. The molecular weight and purity of kojic acid was confirmed by LC/MS and HPLC. The isolated kojic acid crystals shows high antimicrobial activity against pathogenic bacteria Staphylococcus aureus and Escherichia coli and maximum zone of inhibition was 9 mm. The inhibitory effect of kojic acid was more on the cell line K562 (Leukemia) when compared to the MDAMB435S (Breast cancer) cell line.

Adaptation of the tween 80 assay with a resolution V fractional factorial design and its application to rank Ophiostoma fungi with wood extractive degrading capabilities

Wood extractives in radiata pine are the source of the observed pitch problems in the pulp and paper industry. Various methodologies have been studied and used to reduce or eliminate their negative effects. A biological treatment with albino fungi of the Ophiostoma genus, fungi that degrade the lipidic components of extractives, has been proposed as a more environmentally friendly alternative for pitch degradation. The current methods used to search for Ophiostoma albino strains with the highest degradation rates of wood extractives are labor intensive and require a large amount of resources. The Tween 80® Opacity test, an assay designed to measure lipolytic enzyme activity in filamentous fungi, was studied to verify the feasibility of its application as a methodology to rank Ophiostoma albino strains with deresination capabilities. The Tween 80® Opacity Test was first characterized by implementing a non-replicated 2 k-p fractional factorial design of resolution V for a 5 factors with 16 treatments and then to study the effect of the fungus species on the lipolytic enzyme activity; a randomized one factor general factorial design was conducted. The incubation temperature; antibiotics presence; and Tween 80® , CaCl 2 , and peptone concentrations were investigated in the first experiment. The Ophiostoma species effect was studied in the second experiment. In both experiments, the halo area size, which was formed by the fatty acid-calcium complex precipitate, was the response variable. The results demonstrated the effectiveness of the Tween 80® opacity test to measure the lipolytic enzyme activity of Ophiostoma albino strains. Similarly, the incubation temperature and the concentrations of Tween 80® , CaCl 2 , and bacteriological peptone had the highest statistically significant effect on the response variable. In addition, our data demonstrated that the albino strains from the specie O. floccosum exhibited the highest rate of lipolytic enzyme production.

Optimization of Streptomyces sp.A11 Medium Cultivation on Cyclo(Tyrosyl-Prolyl) Production Using the Response Surface Methodology

The Response Surface Methodology is a three factorial design which provides the relationship between one or more measured dependent response and a number of input (independent) factors. Response surface methodology was used for optimization of fermentation medium for antibacterial agent cyclo(tyr-pro) production by Streptomyces sp. A11. The optimal response region of the significant factor was predicted by using a second order polynomial model fitted to the results obtained by applying the Central Composite Design (CCD) statistical design. Determination of the best production medium of cyclo(tyr-pro) showed among 6 carbon sources used, dextrin produced the highest cyclo(tyr-pro). Among the five nitrogen sources used, peptone produced the highest cyclo(tyr-pro), and the mineral salts group I produced the highest cyclo(tyr-pro) compared with others. The results of optimization showed that three variables (dextrin as a carbon source, peptone as nitrogen source, and mixture of mineral salts) used showed significantly affected to the activity of cyclo(tyr-pro). The mathematical model used, obtained by the composition of the three variables most optimum were concentration of dextrin was 32.55 g.L-1 , peptone concentration was 11.22 g L-1 , and 8.65 mL of mineral salts. By using the composition of mathematic model, cyclo(tyr-pro) was obtained 51.54 mg L-1 (model expectations) and 50.04 mg L-1 from experiments. The differences of experiment with expectation response value was 2.91%. Using the optimized parameters, the studies demonstrated an increase in the cyclo(tyr-pro) was 2.5 fold from 20 mg L-1 to 50 mg L-1

Statistical optimization of culture medium for the overproduction of chicken anemia virus immunogen‐ VP1 protein in a recombinant E. coli for vaccine application

AbstractThe capsid protein VP1 of chicken anemia virus (CAV) is an immunogen usually to be used when developing the diagnostic kits and subunit vaccines. In this study, a codon‐optimized VP1 gene of CAV was used to express VP1 in Escherichia coli. To establish the best culture medium for VP1 production, the modified super optimal broth with catabolic repressor (mSOC) medium was selected, out of eight commonly used media, as the basal medium for optimization. The 24 factorial experimental results demonstrated that only two constituents of mSOC, peptone and mixed ionic solution, had a significant effect on VP1 production. After a steepest ascent experiment, a rotatable central composite design (CCD) approach was applied to optimize these variables with respect to VP1 production. When the concentrations of peptone (1.79%) and mixed ionic solution (0.27%) predicted by the CCD approach were used with mSOC, a maximum VP1 yield of 138.68 µg mL−1 was achieved. The use of this optimized mSOC medium resulted in a 1.84‐fold increase in VP1 protein production compared to LB medium. The optimized mSOC identified herein has potential to be used in the future for the industrial large‐scaled VP1 production to diagnostic or a subunit vaccine application. © 2014 Curtin University of Technology and John Wiley & Sons, Ltd.

Optimization Statistical Condition of Incubation for the Growth State of a Recombinant Strain Produced Alkaline Protease

Recombinant E. coli strain was used to yield alkaline protease. The effects of components of medium and condition of incubation for the growth state of a recombinant strain including glucose concentration, peptone concentration, volume of salt solution, incubation time and agitation frequency on density of recombinant strain were optimized using one-factor-at-a-time experiment. The optimal components of medium were glucose concentration of 8 g/L, peptone concentration of 6 g/L and volume of salt solution 10 mL. And condition of incubation was obtained at agitation frequency of 210 r/min, keep it at 37°C for 32 h.

Effects of pellet characteristics on L-lactic acid fermentation by R. oryzae: pellet morphology, diameter, density, and interior structure.

The effects of pellet morphology, diameter, density, and interior structure on L-lactic acid fermentation by Rhizopus oryzae were characterized for different inoculum sizes and concentrations of peptone and CaCO3. Inoculum size was the most important factor determining pellet formation and diameter. The diameter decreased with increasing inoculum size, and larger pellets were observed for lower inoculum sizes. Peptone concentration had the greatest effect on pellet density, which increased with increasing peptone concentration. L-lactic acid production depended heavily on pellet density but not on pellet diameter. Low-density pellets formed easily under conditions of low peptone concentration and often had a relatively hollow structure, with a thin condensed layer surrounding the pellet and an extraordinarily loose biomass or hollow center. As expected, this structure greatly decreased production. The production of L-lactic acid increased until the density reached a certain level (50-60 kg/m(3)), in which the compact part was distributed homogeneously in the thick outer layer of the pellet and loose in the central layer. Homogeneously structured, denser pellets had limited mass transfer, causing a lower overall turnover rate. However, the interior structure remained nearly constant throughout all fermentation phases for pellets with the same density. CaCO3 concentration only had a slight influence on pellet diameter and density, probably because it increases spore germination and filamentous hypha extension. This work also provides a new analysis method to quantify the interior structure of pellets, thus giving insight into pellet structure and its relationship with productivity.

A study on biological characteristics of hybrid of Lentinula edodes from a wildstrain (Langbiang mountain, Lam Dong) and a commercial one (Japan)

Lentinula edodes is a favorable edible mushroom in the world. Its successful cultivation requires not only good and high yield strains but also the appropriate nutrient sources and environmental conditions. A hybrid strain (Len 003-009) is the result of the mating between a commercial strain (Japan) (Len 009) and a wild one (Langbiang mountain, Lam Dong) (Len 003). Studying some biological characteristics of the hybrid showed that its growth rate was similar to or quicker than that of the commercial strain and about twice as rapid as that of the wild one. The hybrid also represented some heredity characteristics from its parents. In addition, the optimal growth of the hybrid and the commercial strain were similar with the temperature from 24 to 27 degree Celsius, the concentration of glucose and peptone being 20 g/l and 1 g/l, respectively. Moreover, its parents could only optimally grow at the different levels of pH (pH=5 in Len 009 and pH=6 in Len 003), while the hybrid could do that in both mentioned levels. Again, this showed good characteristics of hybrid got from its parents. All strains did not have any demand for yeast extract during their growth and development. Thus, it could not evaluate whether there was any inheritance from its parents or not from this experiment.

English

Trichoderma viride Sn-9106 with high cellulase activity was used to produce enzyme on residues of Chinese herbs as substrate in solid state fermentation. Residues of Chinese herbs and peptone were found to be the best combination of carbon and nitrogen source for the production of cellulase. The nutrient composition of medium was optimized using response surface methodology. A fractional factorial design (33) was applied to elucidate the nutrient medium components that significantly affect cellulase production. The concentration of peptone and wheat bran in the medium was a significant factor. The composition of nutrient fermentation medium optimized with response surface methodology was in g/L: wheat bran, 19.8, peptone, 2.06 and KH2PO4, 2.9. Compared to the original medium, the cellulase activity increased from 3.8 to 7.5 IU/mL. Key words: cellulase, Trichoderma viride Sn-9106, response surface methodology (RSM), solid state fermentation (SSF), residues of Chinese herbs (RCH).

96-well microtiter plates for biofouling simulation in biomedical settings

Microtiter plates with 96 wells are routinely used in biofilm research mainly because they enable high-throughput assays. These platforms are used in a variety of conditions ranging from static to dynamic operation using different shaking frequencies and orbital diameters. The main goals of this work were to assess the influence of nutrient concentration and flow conditions on biofilm formation by Escherichia coli in microtiter plates and to define the operational conditions to be used in order to simulate relevant biomedical scenarios. Assays were performed in static mode and in incubators with distinct orbital diameters using different concentrations of glucose, peptone and yeast extract. Computational fluid dynamics (CFD) was used to simulate the flow inside the wells for shaking frequencies ranging from 50 to 200 rpm and orbital diameters from 25 to 100 mm. Higher glucose concentrations enhanced adhesion of E. coli in the first 24 h, but variation in peptone and yeast extract concentration had no significant impact on biofilm formation. Numerical simulations indicate that 96-well microtiter plates can be used to simulate a variety of biomedical scenarios if the operating conditions are carefully set.

Integration of Artificial Neural Network Modeling and Genetic Algorithm Approach for Enrichment of Laccase Production in Solid State Fermentation by Pleurotus ostreatus

Black gram husk was used as a solid substrate for laccase production by Pleurotus ostreatus, and various fermentation conditions were optimized based on an artificial intelligence method. A total of six parameters, i.e., temperature, inoculum concentration, moisture content, CuSO4, glucose, and peptone concentrations, were optimized. A total of 50 experiments were conducted, and the obtained data were modeled by a hybrid of artificial neural network (ANN) and genetic algorithm (GA) approaches. ANN was employed to model the experimental data, and the predicted values were further optimized by GA. Employment of ANN–GA hybrid methodology resulted in a significant improvement, as approximately two-fold laccase production (4244 U/gds) was achieved.

English

The remarkable properties of humic acids have generated a broad spectrum of applications in pharmaceutical, cosmetic and agricultural fields, and encouraged fermentation studies focusing on humic acids production. This work compares the humic acids production of Trichoderma (viride and reesei) species using empty fruit bunch as the substrate during submerged fermentation. The performance of each species was compared by examining spore production in oat medium, and the significant medium components and fermentation conditions were identified using Plackett and Burman statistical design. For both Trichoderma species, the results indicate that humic acids production can be enhanced by increasing the temperature, empty fruit bunch and peptone concentrations and by decreasing the (NH4)2SO4 concentration. T. reesei performed better than T. viride, generating 3-fold more of humic acids.   Key words: Humic acids, Trichoderma reesei, Trichoderm aviride, submerged fermentation, empty fruit bunch.

The Effect of Different Factors on Microbial Oil Production by Trichosporon Fermentans on Rice Straw Acid Hydrolysate

Microbial oil production by Trichosporon fermentans was carried out on the rice straw acid hydrolysate in this work. In order to reduce the detoxification cost, activated carbon was used as absorbent. The effect of peptone concentration, inoculum size, cultural temperature, and initial pH on the lipid fermentation of T. fermentans was evaluated that initial pH showed more significant influence than other three factors. The optimal peptone concentration, inoculum size, cultural temperature, and initial pH for the lipid production were 0.6 g/L, 10%, 25°C, and 7.0, respectively. Under the optimal condition, the highest lipid yield of T. fermentans was 12.1 g/L (corresponding to 31.0 g/L biomass and 39.0% lipid content) after 10 days’ fermentation. This work showed that low-cost activated carbon could be utilized for detoxification of rice straw acid hydrolysate, and T. fermentans is a promising strain for microbial oil and thus biodiesel production from lignocellulosic biomass.

Statistical approach for production of PUFA from Kocuria sp. BRI 35 isolated from marine water sample.

In this study, Plackett-Burman design was used to identify the most influential parameters affecting PUFA production by Kocuria sp. BRI 35 isolated from Antarctic water sample. Amongst 10 variables evaluated, magnesium chloride, protease peptone, glucose, and temperature were significant. Response surface methodology consisting of a central composite design was developed to study the interactions between the variables and to determine optimal values of significant variables. A quadratic model (R = 0.9652, F = 14.64, P < 0.0001) was built. The contour plots indicated that the isolate produced maximum PUFA at lower concentrations of magnesium sulfate (0.9 g/L) and higher concentrations of protease peptone (5 g/L) and glucose (10 g/L) at 15°C. MgSO4 and glucose exhibited quadratic as well as interactive effect on PUFA production whereas protease peptone and temperature showed interactive effects only. After optimization, PUFA production per unit biomass increased from 0.94 mg/g to 11.12 mg/g. This represented an increase from 3% to 58.62% of the total fatty acids. Among PUFAs, the yield of ω-6 fatty acids increased from 9.66 mg/L to 107.71 mg/L with significant increase in linoleic acid (20.36 mg/L) whereas ω-3 fatty acids increased up to 12.37 mg/L with DHA being the major ω-3 fatty acid produced.