Inoculum Age Research Articles

Statistical optimisation of xylanase production by estuarine Streptomyces sp. and its application in clarification of fruit juice.

Recently, xylanase has become an essential option for environmental friendly industrial biotechnological applications and the rising demand for its large scale production urge to take the advantage of statistical approach of optimization to investigate the interactive effects of prominent process factors involved to enhance xylanase production. In the present study, xylanase production from Streptomyces sp. strain ER1 isolated from Cochin estuarine soil; was optimised using statistical designs- Plackett-Burman and Central composite design. Plackett-Burman design was used to identify important fermentation condition factors affecting the xylanase production using beechwood xylan as the substrate. The optimum levels of these significant factors were determined employing the Central Composite Design. Out of the thirteen factors screened, concentration of beechwood xylan and olive oil, agitation speed, and inoculum age were recognized as the most significant factors. By analyzing the response surface plots and using numerical optimization method, the optimal levels for concentration of xylan and olive oil, agitation speed and inoculum age were determined as 0.37%, 33.10 mg/L, 42.87 RPM and 21.05 h, respectively. The optimised medium resulted in a 1.56-fold increased level of the xylanase (10,220 U/mL) production compared to the initial level (3986.444 U/mL) after 120 h of fermentation. The purified enzyme could successfully clarify orange, mousambi and pineapple juice to 20.87%, 23.64% and 27.89% respectively. Thus the present study has proved that Streptomyces sp. strain ER1 (KY449279) is a potential and useful organism for xylanase production and its purified enzyme could clarify the selected fruit juices.

Production of paclitaxel with anticancer activity by two local fungal endophytes, Aspergillus fumigatus and Alternaria tenuissima.

Among 60 fungal endophytes isolated from twigs, bark, and mature leaves of different plant species, two fungal isolates named TXD105 and TER995 were capable of producing paclitaxel in amounts of up to 84.41 and 37.92μg L-1, respectively. Based on macroscopic and microscopic characteristics, ITS1-5.8S-ITS2 rDNA sequence, and phylogenetic characteristic analysis, the two respective isolates were identified as Aspergillus fumigatus and Alternaria tenuissima. In the effort to increase paclitaxel magnitude by the two fungal strains, several fermentation conditions including selection of the proper fermentation medium, agitation rate, incubation temperature, fermentation period, medium pH, medium volume, and inoculum nature (size and age of inoculum) were tried. Fermentation process carried out in M1D medium (pH6.0) and maintained at 120rpm for 10days and at 25°C using 4% (v/v) inoculum of 5-day-old culture stimulated the highest paclitaxel production to attain 307.03μg L-1 by the A. fumigatus strain. In the case of the A. tenuissima strain, fermentation conditions conducted in flask basal medium (pH6.0) and maintained at 120rpm for 14days and at 25°C using 8% (v/v) inoculum of 7-day-old culture were found the most favorable to attain the highest paclitaxel production of 124.32μg L-1. Using the MTT-based assay, fungal paclitaxel significantly inhibited the proliferation of five different cancer cell lines with 50% inhibitory concentration values varied from 3.04 to 14.8μg mL-1. Hence, these findings offer new and alternate sources with excellent biotechnological potential for paclitaxel production by fungal fermentation.

Zn2+ sequestration by Nostoc muscorum: study of thermodynamics, equilibrium isotherms, and biosorption parameters for the metal.

Microbial biosorption has evolved as an effective strategy for heavy metal removal from contaminated waters. The common cyanobacterium Nostoc muscorum isolated from the banks of a polluted river in Meghalaya, India, was tested for its potential to remove Zn2+ from aqueous solutions. Energy-dispersive X-ray (EDX) study verified Zn binding on the cyanobacterial biomass, and FTIR analysis revealed many negatively charged functional groups (hydroxyl, carbonyl, alcohol, amine, phosphoryl, sulfhydryl, and carboxyl) on the cell surface that aided in metal binding. Thermodynamic studies established the biosorption process to be energetically favorable with negative free energy change (-10.404, -10.599, and -10.796kJ/mol at 298, 303, and 308K, respectively). Sorption isotherm data fitted best in the Langmuir isotherm indicating monolayer nature of Zn sorption. The organism showed hyper-accumulation tendency towards Zn with a maximum sorption capacity as high as 2500mg of Zn taken up per gram of biomass. The separation factor R L calculated from Langmuir isotherm ranged between 0 and 1 signifying favorable interaction between the cyanobacterial biomass and the Zn ions. Various experimental parameters, viz. pH, temperature, inoculum age and size, and shaking rate, influenced Zn biosorption. Optimized experimental conditions significantly enhanced the sorption percentage. Sorption was primarily a fast surface phenomenon in the beginning with internalization of zinc ions by the live cells on prolonged exposure.

Cost-effective cellulase production using Parthenium hysterophorus biomass as an unconventional lignocellulosic substrate

The potential of untreated Parthenium hysterophorus weed biomass was evaluated as a substrate for cellulase production. The cellulose in the biomass was used as the main source of carbon. Solid-state fermentation was carried out using Trichoderma reesei, and optimization of cultural conditions was done for maximization of cellulase production. The results revealed that highest cellulase production was achieved on the 8th day of incubation, at 30 °C, keeping solid-to-liquid ratio 1:2 when two discs of inoculum were used per gram of the substrate. The optimized inoculum age was 96 h for CMCase and 120 h for FPase. On studying the enhancing effect of different carbon and nitrogen sources, lactose and ammonium molybdate were found suitable, respectively. The optimized concentration of lactose for the highest CMCase and FPase activities was 1.5 and 1%, respectively. Ammonium molybdate was best at 1% concentration for both CMCase and FPase. Maximum CMCase and FPase activities obtained were 20.49 and 2.42 U/gds, respectively.

ASPERGILLUS TERREUS KMBF1501 A POTENTIAL PIGMENT PRODUCER UNDER SUBMERGED FERMENTATION

Objective: The present study was aimed to identify the fungal isolate from soil and to understand the different optimized parameters better to facilitate the pigment production that has high yield and stability.Methods: Aspergillus sp. was isolated from Western Ghats soil by the conventional serial dilution technique and assessed as a potential pigment producer. Different broth medium such as potato dextrose broth (PDB), czapek-dox broth (CDB), malt extract broth (MEB), rose bengal broth (RBB), sabouraud dextrose broth (SDB), yeast malt extract broth (YEMB), pH (3-9), temperature (24, 27, 30, 33, 37 and 40 °C), carbon (lactose,glucose,sucrose, maltose, galactose and fructose) and nitrogen source (peptone, yeast extract, urea and inorganic nitrogen sources like potassium nitrate, ammonium chloride and sodium nitrate), mineral salts such as sodium dihydrogen phosphate (Na2H2Po4), magnesium sulphate (Mg2So4), calcium chloride (CaCl2), copper sulphate (Cu2So4), potassium dihydrogen phosphate (KH2Po4) and manganese sulphate (Mn2So4) and inoculum age (2-7 d) of the medium related to high pigment production were analysed.Results: Aspergillus terreus KMBF1501 was identified by ribosomal DNA sequencing showing 99% similarity with other Aspergillus terreus and the Accession number (KX113516) was assigned. The optimum culture conditions for pigment production by Aspergillus terreus KMBF1501 was achieved at pH 5 (0.563±0.012 nm), temperature of 27 °C (0.382±0.001 nm) with glucose (0.501±0.002 nm) as carbon source, peptone (2.147±0.004 nm) as nitrogen source, Mg2SO4 (0.401±0.001 nm)as mineral salt and 4 d (0.324±0.001 nm) of inoculum age in PDB (0.761±0.006 nm).Conclusion: Aspergillus terreus KMBF1501 produced maximum pigment when cultured in modified PDB than in common PDB medium. The high concentration of the pigment can be used for various industrial purposes.

Effect of inoculum age and physical parameters on in vitro culture of the entomopathogenic nematode Steinernema feltiae.

Entomopathogenic nematodes (EPNs) of the families Steinernematidae and Heterorhabditidae have a symbiotic association with bacteria which makes them virulent against insects. EPNs have been mass produced using in vivo and in vitro methods, including both solid and liquid fermentation. This study assessed the effect of nematode inoculum age on the production of Steinernema feltiae in liquid, solid and biphasic processes. Several physical parameters were also assessed: the effect of medium viscosity, flask size and aeration speed on the recovery and yield of infective juveniles (IJs). Inoculum age treatments included inoculum liquid cultures that were 7, 14, 21 and 28 days old. Nematodes from the same inoculum were added to one liquid medium (liquid culture), one solid medium with bacteria previously grown in sponge (solid culture) and a variation of the solid medium (a biphasic culture), in which the bacteria were first grown in liquid and, then, soaked into the sponges, with the purpose of providing a more homogeneous bacterial culture before nematode inoculation. Experiments were conducted in Erlenmeyer flasks. Eight treatments were established involving combinations of three variables: two media (with and without 0.2% agar), two flask sizes (250 and 150 ml) and two agitation speeds (180 and 280 rpm). The study showed increases in nematode yield for liquid cultures, but not for solid or biphasic cultures, with the advance of the inoculum age up to 28 days of growth. Furthermore, the addition of 0.2% agar to the liquid medium and increasing the aeration rate by using larger flasks with higher agitation speed may increase nematode recovery and final yield. The experiments were conducted using shake flasks but the results may also be applicable for bioreactors.

Process development for the production of bioethanol from waste algal biomass of Gracilaria verrucosa

The algal biomass of different species of Gracilaria were collected from coasts of Orissa and Tamil Nadu, India and characterized biochemically. Among various species, G. verrucosa was found to be better in terms of total carbohydrate content (56.65%) and hence selected for further studies. The agar was extracted from algal biomass and the residual pulp was enzymatically hydrolyzed. The optimization of algal pulp hydrolysis for various parameters revealed a maximum sugar release of 75.8mg/ml with 63% saccharification yield. The fermentation of enzymatic hydrolysate of algal pulp was optimized and 8% (v/v) inoculum size, 12h inoculum age, pH 5.0 were found to be optimum parameters for maximum ethanol concentration (27.2g/L) after 12h. The process of enzymatic hydrolysis and fermentation were successfully scaled up to 2L bioreactor scale.

Screening of Important Factors for Xylanase and Cellulase Production from the Fungus C. cinerea RM-1 NFCCI-3086 through Plackett-Burman Experimental Design

A Plackett-Burman design was employed to evaluate the effect of different culture conditions associated with xylanase and cellulase production by the fungus C. cinerea RM-1 NFCCI 3086 using agro-residues as substrate. Eight variables were assessed for their significance on xylanase and cellulase production under solid state fermentation. The optimal culture conditions for xylanase production were developed by maintaining the variables of temperature, incubation period, substrate concentration, particle size, inoculum size, and inoculum age at their higher levels, while keeping pH and moisture ratio at their lower levels. For cellulase production, temperature, incubation period, substrate concentration, inoculum size, and inoculum age were fixed at their higher levels, and pH, particle size, and moisture level were kept at their lower levels. Of the eight variables, temperature, incubation period, and pH had significant influence on xylanase and cellulase production. These three variables can be further optimized for increased enzyme production.

Influence of nutritional and operational parameters on the production of butanol or 1,3-propanediol from glycerol by a mutant Clostridium pasteurianum

Butanol and 1,3-propanediol (1,3-PDO) are simultaneously produced by Clostridium pasteurianum from glycerol. In this study, random chemical mutagenesis of C. pasteurianum DSM 525 was conducted to improve its tolerance to butanol. Selected nutritional and operational parameters were evaluated to identify strategies that favour the production of each metabolite. From those experiments, it was possible to isolate cells able to produce 22% more butanol than the parent strain in serum bottles. The supplementation of the culture medium with 2mgl-1 of iron increased the production of butanol by 163%, and the optimum inoculum age was found to be 12hours. Overall, the experiments conducted in bioreactor led to lower butanol titers than in serum bottles, which is attributed to the higher pressure present in the bottles. At pH 6.0, N2 sparging notoriously favoured the production of biomass and 1,3-PDO, while a lower pH (5.0) led to a higher butanol yield, although growth was negatively affected. The results herein gathered allowed the identification of specific conditions that favour the production of either butanol or 1,3-PDO. Furthermore, it was found that N2 sparging is a suitable strategy to maximize the titer, yield and productivity of 1,3-PDO using C. pasteurianum.

Process optimization for hyper production of xylanase via statistical methodology from isolated Bacillus pumilus 3GAH using lignocellulosic waste

Abstract Xylanases hold immense importance in many of the industrial applications for which their economic production at large scale is desired. Isolated xylanolytic strain Bacillus pumilus 3GAH with potential of high titre productivity was selected for subsequent optimization using wheat bran as a substrate. For this, Plackett Burman design was constructed with 11 process variables including media components (wheat bran, peptone, xylose, KNO3 and yeast extract), incubation conditions (shaking conditions, incubation temperature and incubation time), pH, inoculum age and inoculum size. As soon as the most significant factors were identified, they were used to construct Central Composite Design (CCD) within the range of xylose (0.01–0.32 g/50 ml), incubation time (12–26 h), yeast extract (0.01–0.32 g/50 ml), peptone (0.01–0.32 g/50 ml) and temperature (19.7–55.3 °C). CCD predicted the maximum xylanase activity (IU/ml) at the optimum conditions of xylose (0.1 g/50 ml), incubation time (22 h), yeast extract (0.1 g/50 ml), peptone (0.1 g/50 ml) and temperature (30 °C) while the fixed variables were wheat bran (0.3 g/50 ml), KNO3 (0.1 g/50 ml), inoculum size (0.5 ml), inoculum age (16 h) and shaking at 120 rpm. These statistical designs altogether resulted in approximately 1430 IU/ml xylanase activity translating to around 200,200 IU/gds (per gram dry substrate) with 24 fold increase as compared to unoptimised 8310 IU/gds; hence proving its utility.

Optimization of fermentation condition favoring butanol production from glycerol by Clostridium pasteurianum DSM 525

Butanol is a promising biofuel and valuable platform chemical that can be produced through fermentative conversion of glycerol. The initial fermentation conditions for butanol production from pure glycerol by Clostridium pasteurianum DSM 525 were optimized via a central composite design. The effect of inoculum age, initial cell density, initial pH of medium and temperature were quantified and a quadratic model was able to predict butanol yield as a function of all four investigated factors. The model was confirmed through additional experiments and via analysis of variance (ANOVA). Subsequently, numerical optimization was used to maximize the butanol yield within the experimental range. Based on these results, batch fermentations in a 7L bioreactor were performed using pure and crude (residue from biodiesel production) glycerol as substrates at optimized conditions. A butanol yield of 0.34molebutanolmole−1glycerol was obtained indicating the suitability of this feedstock for fermentative butanol production.

Profiling and production of hemicellulases by thermophilic fungus Malbranchea flava and the role of xylanases in improved bioconversion of pretreated lignocellulosics to ethanol.

This study reports thermophilic fungus Malbranchea flava as a potent source of xylanase and xylan-debranching accessory enzymes. M. flava produced high levels of xylanase on sorghum straw containing solidified culture medium. The optimization of culture conditions for production of hemicellulases was carried out using one factor at a time approach and Box–Behnken design of experiments with casein (%), inoculum age (h) and inoculum level (ml) as process variables and xylanase, β-xylosidase, acetyl esterases and arabinofuranosidase as response variables. The results showed that casein concentration between 3.0 and 3.5 %, inoculum age (56–60 h) and inoculum level (2–2.5 ml) resulted in production of 16,978, 10.0, 67.7 and 3.8 (U/gds) of xylanase, β-xylosidase, acetyl esterase and α-l-arabinofuranosidase, respectively. Under optimized conditions M. flava produced eight functionally diverse xylanases with distinct substrate specificity against different xylan types. The peptide mass fingerprinting of 2-D gel electrophoresis resolved proteins indicated to the presence of cellobiose dehydrogenase and glycosyl hydrolases suggesting the potential of this strain in oxidative and classical cellulase-mediated hydrolysis of lignocellulosics. Addition of xylanase (300 U/g substrate) during saccharification (at 15 % substrate loading) of different pretreated (acid/alkali) substrates (cotton stalks, wheat straw, rice straw, carrot grass) by commercial cellulase (NS28066) resulted in 9–36 % increase in saccharification and subsequent fermentation to ethanol when compared to experiment with commercial enzyme only. High ethanol level 46 (g/l) was achieved with acid pretreated cotton stalk when M. flava xylanase was supplemented as compared to 39 (g/l) with xylanase without xylanase addition.Electronic supplementary materialThe online version of this article (doi:10.1007/s13205-015-0325-2) contains supplementary material, which is available to authorized users.

Optimization of Alkaline Protease Production by Alkaliphilic Bacillus sp. KW2 in Low Cost Medium using Statistical Approaches

Objective: Isolation of alkaline protease producing alkaliphilic bacteria and optimization of protease production in low cost medium. Methods: Optimization of protease production was performed by one-variable-at-a-time (OVAAT) approach and statistical approaches using Design Expert 6 software. Results: Nine (9) morphologically distinct alkaliphilic bacterial strains were isolated. Of these isolates 6 strains were found to be positive for protease production. Among these proteolytic strains, Bacillus sp. KW2 was selected for protease production optimization studies. The optimal protease production was observed at 30°C and pH 10.7. Among the various carbon and nitrogen sources studied rice bran and soybean meal were found to be the best Carbon and Nitrogen sources respectively. Plackett-Burman design (PBD) was used to screen signal factors that influenced the protease production. Rice bran, soybean meal, inoculum age, CaCl2 and inoculum size gave positive effects whereas KH2PO4 and MgSO4 gave negative effects on protease production. The four most critical factors viz. rice bran, soybean meal, calcium chloride and potassium dihydrogen phosphate were selected for optimization studies by Response Surface Methodology (RSM). The response surface graphs showed significant interactions among the factors and a final optimized medium (FOM) was obtained after PBD and RSM experiments. Overall, a 4.8 fold increase in protease production was observed after optimization. Studies of the time course of protease production by Bacillus sp. KW2 revealed that maximal protease production occurred at the stationary phase (84 h) of growth. Conclusions: The present study shows that protease production is greatly influenced by cultural conditions and media constituents. This study also established that the strain KW2 could produce alkaline protease in a low cost medium containing rice bran and soybean meal as major carbon and nitrogen sources. KW2 and its protease may be promising agents for biotechnological applications.

Extracellular Biofabrication, Characterization, and Antimicrobial Efficacy of Silver Nanoparticles Loaded on Cotton Fabrics Using Newly IsolatedStreptomycessp. SSHH-1E

Biological method for silver nanoparticles synthesis has been developed to obtain cost effective, clean, nontoxic, and ecofriendly size-controlled nanoparticles. The objective of this study is extracellular biosynthesis of antimicrobial AgNPs using cell-free supernatant of a localStreptomycessp. strain SSHH-1E. Different medium composition and fermentation conditions were screened for maximal AgNPs biosynthesis using Plackett-Burman experimental design and the variables with statistically significant effects were selected to study their combined effects and to find out the optimum values using a Box-Behnken design. The synthesized AgNPs were characterized using UV-visible spectroscopy, transmission electron microscopy, Fourier transform infrared spectroscopy, and energy dispersive X-ray spectroscopy. Rapid biosynthesis of AgNPs was achieved by addition of 1 mM AgNO3solution to the cell-free supernatant. The produced particles showed a single surface plasmon resonance peak at 400 nm by UV-Vis spectroscopy which confirmed the presence of AgNPs.Streptomycessp. SSHH-1E was identified asStreptomyces narbonensisSSHH-1E. Transmission electron microscopy study indicated that the shape of AgNPs is spherical and the size is ranging from 20 to 40 nm. Fourier transform infrared spectroscopy analysis provides evidence for proteins as possible reducing and capping agents. Furthermore, the biosynthesized AgNPs significantly inhibited the growth of medically important pathogenic Gram-positive and Gram-negative bacteria and yeast. The maximum biosynthesis of AgNPs was achieved at initial pH of 8, peptone of 0.5 g, and inoculum age of 48 h. The statistical optimization resulted in a 4.5-fold increase in the production of AgNPs byStreptomyces narbonensisSSHH-1E.

Production and Optimization of Alkaline Pectin Lyase fromBacillus cereusUnder Submerged Fermentation

Bacillus cereus was isolated from soil sample and selected as maximum pectin lyase producing bacterium under alkaline conditions. The morphological, cultural and biochemical characteristics of bacterium were studied and it was identified on the basis of nucleotide homology and phylogenetic analysis by 16S rDNA analysis. Pectin lyase gave considerable activity with pectin (3% w/v) as a sole carbon source, there was a significant increase in enzyme activity (p<0.001). A combination of both yeast extract and peptone (0.5% w/v each) as nitrogen source also showed significant increase in enzyme activity (p<0.001). Enzyme gave maximum activity with inoculum age of 18 h, inoculum size of 2% (v/v), at temperature of 40°C and pH 10.0 for 24 h incubation time. The glycine-NaOH buffer (0.25M) of pH 10.0 was found to be the most appropriate buffer and optimum reaction temperature and incubation time observed were 30°C and 55 min respectively. Enzyme showed greater affinity towards pectin (0.5% w/v) as substrate. Among all the metal ions studied only Mn2+ ion was effective to stimulate the activity whereas other metal ions inhibited the enzyme activity. SDS was found to be the strongest inhibitor and among thiols, L-cysteine and ascorbic acid stimulated enzyme activity. All the phenolics studied inhibited the enzyme activity. The enzyme gave half-life of 150 min at optimized pH 10.0. Optimal enzyme concentration observed was 0.01mg/ml. This enzyme has tremendous potential in textile industry and paper and pulp industry.

Single-cell protein production through microbial conversion of lignocellulosic residue (wheat bran) for animal feed

Agricultural residue (wheat bran) rich in carbohydrates was utilized in the fermentation process to produce microbial biomass. Single-cell biomass consists of the dried cells of microorganisms, which are used as protein supplements in human food and in animal feed. In the present study, two different microorganisms (Candida utilis and Rhizopus oligosporus) were studied for biomass production. To enhance the nutritional contents of wheat bran, a number of different fermentation parameters (effect of inoculum size, age of inoculum, incubation period, moisture to substrate ratio and incubation temperature) were optimized. Maximum yield was obtained at an inoculum size of 10% (v/w), with the age of the inoculum being a 48 h old culture. A fermentation period of 48 h was found to give the maximum protein yield and viable counts of yeast cells and mould hyphae. The microorganisms showed good growth at 30 °C. After complete optimization of the fermentation parameters, a batch of wheat bran was fermented with C. utilis and R. oligosporus under the optimized conditions, resulting in a maximum crude protein yield of 41.02% compared with the 4.21% crude protein of the non-fermented wheat bran. Copyright © 2015 The Institute of Brewing & Distilling

Inoculum optimization of Clostridium beijerinckii for reproducible growth.

Spore-forming solventogenic Clostridium spp. are receiving renewed attention due to their butanol production abilities. However, there is an absence of literature describing the preparation of dense, vigorous and homogeneous seed cultures of Clostridium spp., guaranteeing reproducibility during fermentation. Therefore, we performed a series of growth experiments of Clostridium beijerinckii NCIMB 8052 and its offspring SA-1 to evaluate the influence of inoculum age (harvest time) on the subsequent population's maximum specific growth rate, as a signal of population homogeneity. The organisms were cultivated in Reinforced Clostridial Medium and supplemented sweet sorghum juice. The best inoculum ages coincided with the late-exponential growth phase: between 9 and 11 h in the conditions tested. Additionally, the harvest time was delayed up to 4 h by pre-adapting the seed culture with 0.75 g L(-1) butyric acid. These findings were validated by performing a series of bench-top batch fermentations showcasing reproducibility in growth kinetics with 95% confidence limits. Overall, these experiments allowed us to understand the transient nature of seed cultures of C. beijerinckii NCIMB 8052 and SA-1, while enabling reproducibility and consistent culture performance.

The optimization of Marasmius androsaceus submerged fermentation conditions in five-liter fermentor

Using desirability function, four indexes including mycelium dry weight, intracellular polysaccharide, adenosine and mannitol yield were uniformed into one expected value (Da) which further served as the assessment criteria. In our present study, Plackett–Burman design was applied to evaluate the effects of eight variables including initial pH, rotating speed, culture temperature, inoculum size, ventilation volume, culture time, inoculum age and loading volume on Da value during Marasmius androsaceus submerged fermentation via a five-liter fermentor. Culture time, initial pH and rotating speed were found to influence Da value significantly and were further optimized by Box–Behnken design. Results obtained from Box–Behnken design were analyzed by both response surface regression (Design-Expert.V8.0.6.1 software) and artificial neural network combining the genetic algorithm method (Matlab2012a software). After comparison, the optimum M. androsaceus submerged fermentation conditions via a five-liter fermentor were obtained as follows: initial pH of 6.14, rotating speed of 289.3rpm, culture time of 6.285days, culture temperature of 26°C, inoculum size of 5%, ventilation volume of 200L/h, inoculum age of 4days, and loading volume of 3.5L/5L. The predicted Da value of the optimum model was 0.4884 and the average experimental Da value was 0.4760. The model possesses well fitness and predictive ability.

Fermentation optimization of maltose-binding protein fused to neutrophil-activating protein from Escherichia coli TB1

Background: The fermentation conditions of recombinant maltose-binding protein fused to neutrophil-activating protein (rMBP-NAP) of Helicobacter pylori were optimized from Escherichia Coli TB1 with varying medium, inoculum age and size, time, inducer, pH and temperature in batch fermentation. Results: It was revealed that the optimal conditions for the production of rMBP-NAP in shake flask were as follows: M9 medium (with 3% yeast extract powder added), the inoculum age of 19 h, the inoculum size of 6%, the initial pH of 6.6, temperature at 37oC and 0.7 mmoL/L IPTG inducted 21 h in a 50 mL/250 mL shake flask. The recombinant protein yield was increased from 59 to 592 mg/L after optimization. Fermentation process conducted in a 10 L fermenter with similar conditions could get 30 g/L wet cell and 1.738 g/L soluble protein with rMBP-NAP expression level of 11.9%. Conclusion: The results improve the expression level of rMBP-NAP, and it is expected that these optimized conditions can be well applied for large scale production of rMBP-NAP. Normal 0 21 false false false ES X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:Tabla normal; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:10.0pt; font-family:Calibri,sans-serif;}

Optimization of Culture Conditions for Production of the Anti-Leukemic Glutaminase Free L-Asparaginase by Newly Isolated Streptomyces olivaceus NEAE-119 Using Response Surface Methodology.

Among the antitumor drugs, bacterial enzyme L-asparaginase has been employed as the most effective chemotherapeutic agent in pediatric oncotherapy especially for acute lymphoblastic leukemia. Glutaminase free L-asparaginase producing actinomycetes were isolated from soil samples collected from Egypt. Among them, a potential culture, strain NEAE-119, was selected and identified on the basis of morphological, cultural, physiological, and biochemical properties together with 16S rRNA sequence as Streptomyces olivaceus NEAE-119 and sequencing product (1509 bp) was deposited in the GenBank database under accession number KJ200342. The optimization of different process parameters for L-asparaginase production by Streptomyces olivaceus NEAE-119 using Plackett-Burman experimental design and response surface methodology was carried out. Fifteen variables (temperature, pH, incubation time, inoculum size, inoculum age, agitation speed, dextrose, starch, L-asparagine, KNO3, yeast extract, K2HPO4, MgSO4·7H2O, NaCl, and FeSO4·7H2O) were screened using Plackett-Burman experimental design. The most positive significant independent variables affecting enzyme production (temperature, inoculum age, and agitation speed) were further optimized by the face-centered central composite design-response surface methodology.