Rhamnolipids are extensively studied biosurfactants due to their potential in many industrial applications, eco-friendly production and properties. However, their availability for broader application is severely limited by their production costs, therefore the optimization of efficacy of their cultivation gains significance as well as the information regarding the physio-chemical properties of rhamnolipids resulting from various cultivation strategies. In this work, the bioprocess design focused on optimization of the rhamnolipid yield of Pseudomonas aeruginosa DBM 3774 utilizing the response surface methodology (RSM). Six carbon sources were investigated for their effect on the rhamnolipid production. The RSM prediction improved the total rhamnolipid yield from 2.2 to 13.5 g/L and the rhamnolipid productivity from 11.6 to 45.3 mg/L/h. A significant effect of the carbon source type, concentration and the C/N ratio on the composition of the rhamnolipid congeners has been demonstrated for cultivation of P. aeruginosa DBM 3774 in batch cultivation. Especially, changes in presence of saturated fatty acid in the rhamnolipid congeners, ranging from 18.8% of unsaturated fatty acids (carbon source glycerol; 40 g/L) to 0% (sodium citrate 20 g/L) were observed. This demonstrates possibilities of model based systems as basis in cultivation of industrially important compounds like biosurfactants rhamnolipids and the importance of detailed study of interconnection between cultivation conditions and rhamnolipid mixture composition and properties.
- Publication type
- Journal Article MeSH
L-asparaginase is an essential enzyme used in cancer treatment, but its production faces challenges like low yield, high cost, and immunogenicity. Recombinant production is a promising method to overcome these limitations. In this study, response surface methodology (RSM) was used to optimize the production of L-asparaginase 1 from Saccharomyces cerevisiae in Escherichia coli K-12 BW25113. The Box-Behnken design (BBD) was utilized for the RSM modeling, and a total of 29 experiments were conducted. These experiments aimed to examine the impact of different factors, including the concentration of isopropyl-b-LD-thiogalactopyranoside (IPTG), the cell density prior to induction, the duration of induction, and the temperature, on the expression level of L-asparaginase 1. The results revealed that while the post-induction temperature, cell density at induction time, and post-induction time all had a significant influence on the response, the post-induction time exhibited the greatest effect. The optimized conditions (induction at cell density 0.8 with 0.7 mM IPTG for 4 h at 30 °C) resulted in a significant amount of L-asparaginase with a titer of 93.52 μg/mL, which was consistent with the model-based prediction. The study concluded that RSM optimization effectively increased the production of L-asparaginase 1 in E. coli, which could have the potential for large-scale fermentation. Further research can explore using other host cells, optimizing the fermentation process, and examining the effect of other variables to increase production.
- MeSH
- Asparaginase * genetics biosynthesis metabolism MeSH
- Escherichia coli K12 genetics enzymology MeSH
- Escherichia coli genetics metabolism MeSH
- Fermentation MeSH
- Isopropyl Thiogalactoside pharmacology MeSH
- Recombinant Proteins * genetics metabolism MeSH
- Saccharomyces cerevisiae * genetics metabolism MeSH
- Temperature MeSH
- Publication type
- Journal Article MeSH
Microbial transglutaminase (MTG) is an enzyme widely used in the food industry because it creates cross-links between proteins, enhancing the texture and stability of food products. Its unique properties make it a valuable tool for modifying the functional characteristics of proteins, significantly impacting the quality and innovation of food products. In this study, response surface methodology was employed to optimize the fermentation conditions for microbial transglutaminase production by the strain Streptoverticillium cinnamoneum KKP 1658. The effects of nitrogen dose, cultivation time, and initial pH on the activity of the produced transglutaminase were investigated. The significance of the examined factors was determined as follows: cultivation time > nitrogen dose > pH. The interaction between nitrogen dose and cultivation time was found to be crucial, having the second most significant impact on transglutaminase activity. Optimal conditions were identified as 48 h of cultivation with a 2% nitrogen source dose and an initial medium pH of approximately 6.0. Under these conditions, transglutaminase activity ranged from 4.5 to 5.5 U/mL. The results of this study demonstrated that response surface methodology is a promising approach for optimizing microbial transglutaminase production. Future applications of transglutaminase include the development of modern food products with improved texture and nutritional value, as well as its potential use in regenerative medicine for creating biomaterials and tissue scaffolds. This topic is particularly important and timely as it addresses the growing demand for innovative and sustainable solutions in the food and biomedical industries, contributing to an improved quality of life.
Response surface methodology (RSM) and artificial neural network-real encoded genetic algorithm (ANN-REGA) were employed to develop a process for fermentative swainsonine production from Metarhizium anisopliae (ARSEF 1724). The effect of finally screened process variables viz. inoculum size, oatmeal extract, glucose, and CaCl2 were investigated through central composite design and were further utilized for training sets in ANN with training and test R values of 0.99 and 0.94, respectively. ANN-REGA was finally employed to simulate the predictive swainsonine production with best evolved media composition. ANN-REGA predicted a more precise fermentation model with 103 % (shake flask) increase in alkaloid production compared to 75.62 % (shake flask) obtained with RSM model upon validation.
- MeSH
- Alkaloids isolation & purification metabolism MeSH
- Biotechnology methods MeSH
- Fermentation MeSH
- Culture Media chemistry MeSH
- Metarhizium genetics metabolism MeSH
- Neural Networks, Computer MeSH
- Colony Count, Microbial MeSH
- Swainsonine isolation & purification metabolism MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
β-Cyclodextrin functionalized PEGylated porous silica nanoparticles KIT-6 (denoted as [β-CD@PEGylated KIT-6] NPs) is synthesized and evaluated as an efficient and reliable pH-sensitive nano-carrier. Curcumin (CUR), an anticancer drug, has low solubility and stability and these properties diminished its bioavailability. One way to overcome this problem is employing nano-carrier for delivery of CUR. In this study, the novel [β-CD@PEGylated KIT-6] NPs nano-carrier was employed for CUR delivery successfully. The nano-DDS was characterized using different techniques such as X-ray powder diffraction (XRD), transmission and scanning electron microscopy (TEM and SEM), Fourier transform infrared (FT-IR) spectroscopy, differential scanning calorimeter (DSC), N2 adsorption–desorption measurement, and dynamic light scattering (DLS). In this study, first, the combination of central composite design (CCD) and response surface methodology (RSM) was used to achieve the optimal condition of the loading step with investigation of two important factors: the loading time and the weight ratio of drug to nano-carrier. Maximum loading efficiency 88.55% was obtained at 43 h of loading time and 1.22 of the weight ratio. Then CUR was loaded onto the nano-carrier at this optimal condition and its released was investigated by CCD-RSM. The maximum drug release was obtained at 5.16 of pH and 107 h of release time.
- MeSH
- beta-Cyclodextrins * MeSH
- Curcumin * MeSH
- Drug Delivery Systems MeSH
- Delayed-Action Preparations * chemical synthesis chemistry MeSH
- Cell Line, Tumor drug effects MeSH
- Nanoparticles MeSH
- Drug Carriers chemistry MeSH
- Silicon Dioxide MeSH
- Porosity MeSH
- Antineoplastic Agents chemical synthesis chemistry MeSH
- Drug Liberation MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
We have developed a new microextraction technique for equilibrium, non-exhaustive analyte preconcentration from aqueous solutions into organic solvents lighter than water. The key point of the method is application of specially designed and optimized bell-shaped extraction device, BSED. The technique has been tested and applied to the preconcentration of selected volatile and semi volatile compounds which were determined by gas chromatography/mass spectrometry in spiked water samples. The significant parameters of the extraction have been found using chemometric procedures and these parameters were optimized using the central composite design (CCD) for two solvents. The analyte preconcentration factors were in a range from 8.3 to 161.8 (repeatability from 7 to 14%) for heptane, and 50.0-105.0 (repeatability from 0 to 5%) for tert-butyl acetate. The reproducibility of the technique was within 1-8%. The values of limits of detection and determination were 0.1-3.3 ng mL(-1) for heptane and 0.3-10.7 ng mL(-1) for tert-butyl acetate. The new microextraction technique has been found to be a cheap, simple and flexible alternative to the common procedures, such as SPME or LLME. This BSED-LLME technique can also be combined with other separation methods, e.g., HPLC or CE.
- MeSH
- Water Pollutants, Chemical analysis isolation & purification MeSH
- Equipment Design MeSH
- Liquid Phase Microextraction instrumentation methods MeSH
- Mineral Waters analysis MeSH
- Drinking Water analysis MeSH
- Gas Chromatography-Mass Spectrometry methods MeSH
- Reproducibility of Results MeSH
- Water analysis MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
- Research Support, Non-U.S. Gov't MeSH
Response surface methodology was used to evaluate the effect of main variables such as concentration of galactose, yeast extract and wheat bran on alpha-galactosidase production from Aspergillus parasiticus MTCC-2796 under submerged fermentation conditions. A full factorial Central Composite Design was applied to study these main factors that affected alpha-galactosidase production. The experimental results showed that the optimum concentration of galactose, yeast extract and wheat bran were 1.5 %, 0.06 % and 1.5 %, respectively. This method was efficient as only 20 experiments were necessary to asses these conditions, and model adequacy was very satisfactory as the coefficient of determination was 0.9921.
Production of amylases by fungi under solid-state fermentation is considered the best methodology for commercial scaling that addresses the ever-escalating needs of the worldwide enzyme market. Here response surface methodology (RSM) was used for the optimization of process variables for α-amylase enzyme production from Trichoderma virens using watermelon rinds (WMR) under solid-state fermentation (SSF). The statistical model included four variables, each detected at two levels, followed by model development with partial purification and characterization of α-amylase. The partially purified α-amylase was characterized with regard to optimum pH, temperature, kinetic constant, and substrate specificity. The results indicated that both pH and moisture content had a significant effect (P < 0.05) on α-amylase production (880 U/g) under optimized process conditions at a 3-day incubation time, moisture content of 50%, 30 °C, and pH 6.98. Statistical optimization using RSM showed R2 values of 0.9934, demonstrating the validity of the model. Five α-amylases were separated by using DEAE-Sepharose and characterized with a wide range of optimized pH values (pH 4.5-9.0), temperature optima (40-60 °C), low Km values (2.27-3.3 mg/mL), and high substrate specificity toward large substrates. In conclusion, this study presents an efficient and green approach for utilization of agro-waste for production of the valuable α-amylase enzyme using RSM under SSF. RSM was particularly beneficial for the optimization and analysis of the effective process parameters.
Lipases are industrially important enzymes having vast applications in various fields. Cloning and expression of lipase enzyme-encoding genes in suitable host lead to their widespread use in different fields. The present study represents the first attempt towards the expression of the synthetic lipase gene in Pseudomonas aeruginosa. An alkalophilic lipase gene (GenBank accession number: NP_388152) from Bacillus subtilis was synthetically designed and introduced in the pJN105 vector and subsequently cloned in Pseudomonas aeruginosa SDK-6. Agarose gel electrophoresis confirmed the transformation of SDK-6, exhibiting a band difference of ~ 700 bp between native and recombinant pJN105. Further amplification of cloned lipase gene was confirmed using PCR amplification with Lip 1 and Lip 2 primers respectively, followed by restriction analysis. Approximately 15-fold increase in lipase production was observed in recombinant Pseudomonas as compared to the native strain. One factor at a time (OFAT) analysis revealed L-arabinose, inoculum size (0.5%; v/v), and agitation (120 rpm) as significant factors affecting the over-expression of lipase enzyme. Optimization of enzyme induction conditions by central composite design (CCD) led to 1.60-fold increase in the production of lipase at 0.65% (w/v) inducer concentration, OD600-1.075 before induction and 35 °C post induction temperature with overall lipase production of 50.50 IU/mL. Statistical validation of observed value via ANOVA showed an F-value of 138.70 at p < 0.01 with R2 of 0.9921.
- MeSH
- Arabinose metabolism MeSH
- Bacillus subtilis * genetics enzymology MeSH
- Bacterial Proteins genetics metabolism MeSH
- Gene Expression MeSH
- Genetic Vectors genetics MeSH
- Cloning, Molecular MeSH
- Lipase * genetics metabolism MeSH
- Pseudomonas aeruginosa * genetics enzymology MeSH
- Recombinant Proteins * genetics metabolism MeSH
- Publication type
- Journal Article MeSH
- MeSH
- Electromyography methods utilization MeSH
- Methods MeSH
- Muscles physiology MeSH
- Publication type
- Review MeSH
