Plackett-Burman Design Research Articles

Gluconic Acid Production by Using Recombinant Escherichia coli Waksman pqq+ Cells with a Novel Approach from Biomass Sources.

Gluconic acid (GA) is widely used in the pharmaceutical, food, detergent, textile, leather, and concrete industries. However, cost-effective and high-yield production of GA remains a challenge. Due to currently high raw material inputs of GA, various alternative carbohydrate sources are being investigated. Sucrose is one of the cost-effective biomass sources that can be used as feedstock. The most common industrial production of GA is based on wild-type bacteria and fungi, but there are many problems with this production. This study aimed to optimize the production of GA from glucose produced by hydrolysis of sucrose using recombinant E. coli Waksman (W) pqq+ strain. After sucrose was enzymatically hydrolyzed, significant medium components for GA production were determined as glucose, calcium carbonate (CaCO3), peptone, and ammonium phosphate ((NH4)3PO4) using Placket-Burman Design (PBD). Detailed optimization of the medium components that are significant in GA production was carried out using central composite design (CCD), and optimum values of the independent variables examined in maximum GA production (93.5 ± 2.95g/L) were determined as glucose 95, CaCO3 25, peptone 2, and (NH4)3PO4 1.13 (g/L). Using results obtained in the Erlenmeyer experiments, GA production in the bioreactor was investigated by CCD. The maximum GA efficiency (3.20 ± 0.15g/L. h) was obtained under conditions where the air supply rate was 10.82 L/min, stirring speed was 656.87rpm, and CaCO3 concentration was 16.90g/L. In conclusion, it has been shown that GA can be produced with a high yield with this novel approach using a recombinant strain for GA production from sucrose.

Immobilization of an endophytic Bacillus sp. on Phragmites karka stem for lipase production in the presence of Cassia fistula seeds

AbstractEndophytic bacteria have not been reported widely for their lipolytic abilities, so they are not used for large‐scale lipase production. The purpose of this study was to explore an endophytic bacterium for the production of lipase implementing cost‐effective techniques, including the use of Cassia fistula seeds, as a substrate in the production medium and stem pieces of Phragmites karka as the immobilization matrix. The endophytic strain Bacillus sp. E4 was originally isolated from the halophytic plant Arthrocnemum macrostachyum. Bacillus sp. E4 produced 6.05 IU mL−1 lipase in the presence of powdered seeds of Cassia fistula (golden shower tree). Initial trial experiments using a one‐factor‐at‐a‐time approach led to an improvement in lipase titers to 10.05 IU mL−1. Consequently, investigations using the Plackett–Burman design suggested the influence of three significant factors – incubation period, inoculum size, and substrate concentration – on lipase production. They were optimized using the Box–Behnken design (BBD). In the response optimization experiment, strain E4 yielded 52.35 IU mL−1 lipase, which was in accordance with the predicted yield and indicated an overall 8.65 fold improvement in lipase production. To investigate the use of free cells, strain E4 was immobilized on the stem pieces of a halophytic plant, Phragmites karka, which was used for the first time as an immobilization matrix. The immobilized cells retained lipase production ability for up to six cycles with the highest yield of 110 IU mL−1, which corresponded to an improvement of more than eighteenfold. Scanning electron micrographs confirmed the colonization of E4 cells in the matrix and demonstrated the utilization of C. fistula seeds. Fourier transform infrared spectroscopy affirmed the utilization of components including fatty acids by the immobilized E4 cells. The study suggests that endophytic bacterial strains could be applied for the production of lipase with the utilization of nontraditional oil sources.

Agricultural Waste Management by Production of Second-Generation Bioethanol from Sugarcane Bagasse Using Indigenous Yeast Strain.

In the wake of rapid industrialization and burgeoning transportation networks, the escalating demand for fossil fuels has accelerated the depletion of finite energy reservoirs, necessitating urgent exploration of sustainable alternatives. To address this, current research is focusing on renewable fuels like second-generation bioethanol from agricultural waste such as sugarcane bagasse. This approach not only circumvents the contentious issue of food-fuel conflicts associated with biofuels but also tackles agricultural waste management. In the present study indigenous yeast strain, Clavispora lusitaniae QG1 (MN592676), was isolated from rotten grapes to ferment xylose sugars present in the hemicellulose content of sugarcane bagasse. To liberate the xylose sugars, dilute acid pretreatment was performed. The highest reducing sugars yield was 1.2% obtained at a temperature of 121°C for 15min, a solid-to-liquid ratio of 1:25 (% w/v), and an acid concentration of 1% dilute acid H2SO4 that was significantly higher (P < 0.001) yield obtained under similar conditions at 100°C for 1h. The isolated strain was statistically optimized for fermentation process by Plackett-Burman design to achieve the highest ethanol yield. Liberated xylose sugars were completely utilized by Clavispora lusitaniae QG1 (MN592676) and gave 100% ethanol yield. This study optimizes both fermentation process and pretreatment of sugarcane bagasse to maximize bioethanol yield and demonstrates the ability of isolated strain to effectively utilize xylose as a carbon source. The desirable characteristics depicted by strain Clavispora lusitaniae shows its promising utilization in management of industrial waste like sugarcane bagasse by its conversion into renewable biofuels like bioethanol.

Statistical Experimental Designs for cLTB-Syn Vaccine Production Using Daucus carota Cell Suspension Cultures.

The carrot-made LTB-Syn antigen (cLTB-Syn) is a vaccine candidate against synucleinopathies based on carrot cells expressing the target antigen LTB and syn epitopes. Therefore, the development of an efficient production process is required with media culture optimization to increase the production yields as the main goal. In this study, the effect of two nitrogen sources (urea and glutamate) on callus cultures producing cLTB-Syn was studied, observing that the addition of 17 mM urea to MS medium favored the biomass yield. To optimize the MS media composition, the influence of seven medium components on biomass and cLTB-Syn production was first evaluated by a Plackett-Burman design (PBD). Then, three factors were further analyzed using a central composite design (CCD) and response surface methodology (RSM). The results showed a 1.2-fold improvement in biomass, and a 4.5-fold improvement in cLTB-Syn production was achieved at the shake-flask scale. At the bioreactor scale, there was a 1.5-fold increase in biomass and a 2.8-fold increase in cLTB-Syn yield compared with the standard MS medium. Moreover, the cLTB-Syn vaccine induced humoral responses in BALB/c mice subjected to either oral or subcutaneous immunization. Therefore, cLTB-Syn is a promising vaccine candidate that will aid in developing immunotherapeutic strategies to combat PD and other neurodegenerative diseases without the need for cold storage, making it a financially viable option for massive immunization.

Maximization of uricase production in a column bioreactor through RSM based optimization.

Uricase (EC 1.7.3.3) is an oxidoreductase enzyme which is widely exploited for diagnostic and treatment purposes in medicine. This study has been focused on producing recombinant uricase from E. coli BL21 in a bubble column bioreactor (BCB) and finding the optimal conditions for maximum uricase activity. The three most effective variables on uricase activity were selected through Plackett-Burman design from 8 different variables and were further optimized by central composite design of RSM. The selected variables included the inoculum size (%v/v), IPTG (Isopropyl β-d-1-thiogalactopyranoside) concentration (mM) and the initial pH of culture medium. The activity of uricase, final OD600 (Optical Density at 600nm wave-length) and final pH were considered as the responses of this optimization and were modeled. As a result, the activity of 5.84 U.ml-1 and final OD600 of 3.42 were obtained at optimum conditions of 3 %v/v of inoculum size, IPTG concentration of 0.54 mM and pH=6.0. By purifying the obtained enzyme using a Ni-NTA agarose affinity chromatography column, 165±1.5 mg uricase was obtained from a 600 mL cell culture. The results of this study show that bubble column bioreactors can be a highly effective option for large scale uricase production.

Ammonia nitrogen removal from coal gasification wastewater using a falling-film dielectric barrier discharge reactor: Process optimization and toxicity analysis

The continuous use of coal gasification technology for green energy generation to supplement the dwindling energy reserves generates significant amounts of wastewater containing ammonia nitrogen, posing a significant risk to the finite drinking water sources and aquatic lives. Herein, two falling-film dielectric barrier discharge (DBD) reactors with copper and titanium high-voltage electrodes, respectively, were designed to remove ammonia nitrogen (NH3−N) and chemical oxygen demand (COD) simultaneously from coal gasification wastewater. The operational parameters were screened using the Plackett-Burman design, and the significant parameters were further optimized via the response surface methodology. The results showed that the power parameter significantly influenced NH3-N removal, and COD removal was influenced considerably by power and treatment time. Under the optimum conditions (treatment time = 20.32 min, pH = 3, airflow rate = 1 L/min, power = 116 W, and recirculating flow rate = 50.02 mL/min), a removal ratio of 62.33 % and 37.35 % was achieved for NH3-N and COD for the reactor with the copper electrode, and 67.66 % and 44.90 % for NH3-N, and COD for the reactor with the titanium electrode. Additionally, the concentrations of O3, ·OH, and H2O2 produced by the reactors were quantified, and their crucial roles in the removal process of NH3-N were evaluated. Although smaller concentrations of NO2− and NO3− were detected in the treated wastewater, over 97 % of N2 selectivity encompassed NH3-N removal. Finally, acute phytotoxicity tests were performed, and the results indicated that at 100 mL/L dosage, the treated wastewater from both reactors did not restrict Barbarea verna growth.

Design and validation of a sampling module for a cyclonic wet-wall bioaerosol sampler using a computational fluid dynamics model

Pollen in the atmosphere is one of the common bioaerosol materials that can be harmful to human health. To collect and predict the pollen in the atmosphere, a sampling module of a cyclone wet-wall bioaerosol sampler was designed using the computational fluid dynamics (CFD) method and response surface method (RSM). The parabolic shape was chosen for the sampling module based on the axial and tangential velocities of flow in the module. Then, the main diameter of the sampling module was determined. The significance analysis of the main parameters of the sampling module on the capture rate was conducted using the Plackett-Burman (PB) design method. In descending order, the items are inlet width > outlet diameter > inlet height > base diameter > cone height > cylinder height > outlet pipe insertion depth. The test center point of Box-Behnken design (BBD) for the four significant parameters, namely inlet width, outlet diameter, inlet height, and cone height, were determined using the steepest climb test. Then, the optimal parameters of the sampling module were obtained using RSM and the optimal parameters were an inlet height of 15.4 mm, an inlet width of 5.4 mm, an outlet diameter of 14.7 mm, and a base diameter of 13.3 mm. Finally, the simulated capture rate of the sampling module was validated by the experimental results. The capture rate of the optimal sampling module for 2.5 μm PSL particles was 82.93 ± 4.25%. The results of this study can provide a practical method for designing the sampling module using the CFD model and RSM. Additionally, it can provide a high collection rate sampling module for pollen and bioaerosol in the environment.

Two-pot optimization of nutrient sources to enhance antioxidants in Citrus reticulata peel powder through solid-state fermentation with Aspergillus niger CGMCC 3.6189

This study aimed to optimize nutrient sources for enhancing the accumulation of antioxidants in Citrus reticulata peel powder (CRPP) through solid-state fermentation (SSF) using Aspergillus niger CGMCC 3.6189. A Plackett-Burman design revealed that four medium compositions, including yeast extract, NH4Cl, CaCl2, and MgCl2, significantly affected antioxidant accumulation in fermented CRPP. The optimal concentrations of these four compositions were predicted as 13.86, 0.40, 0.06, and 0.04 mg/g of CRPP, respectively, using a Box-Behnken design (BBD) when simultaneous optimization of total phenolic content (TPC), total flavonoid content (TFC), 2,2′-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging capacities using Derringer's desirability function. Under the optimal medium compositions, TPC, TFC, ABTS, and DPPH scavenging capacities of fermented CRPP (FO-CRPP) increased by 25.2%, 26.9%, 45.8%, and 57.3%, respectively, in comparison with the unfermented CRPP (UF-CRPP). The HPLC analysis exhibited notable decreases in the levels of hesperidin, narirutin, nobiletin, and tangeretin, while enhancements in the contents of p-coumaric acid, ferulic acid, narirutin, and hesperetin in FO-CRPP compared to UF-CRPP were observed. Scanning electron microscopy and Fourier transform infrared spectroscopy analyses demonstrated changes in the surface microstructure and functional groups in FO-CRPP. Our findings suggested that yeast extract and NH4Cl significantly improve the accumulation of antioxidants in CRPP during SSF by A. niger CGMCC 3.6189.

Process optimization for microfluidic preparation of liposomes using food-grade components

This study explores the potential for optimizing a sustainable manufacturing process that maintains the essential characteristics of conventional liposomes using food-grade solvents and components. The focus was comparing the physicochemical, morphological, and interfacial properties of liposomes produced with these food-grade ingredients to those made by conventional methods. It was found that there was no significant difference in particle size (195.87 ± 1.40 nm) and ζ-potential (−45.13 ± 0.65 mV) between liposomes made from food-grade and conventional materials. The manufacturing process for liposomes, utilizing food-grade solvents and components, was optimized through the application of Plackett-Burman design and response surface methodology. This approach helped identify key parameters (soy lecithin, β-sitosterol, W/O ratio) and their optimal values (3.17 g, 0.25 g, 1:2.59). These findings suggest that it is possible to enhance the use of liposomes as an effective and safe delivery system in the food industry, adhering to the strict guidelines set by regulatory agencies.

Agile quality-by-design development of alginate microparticles for encapsulation of hydrophilic drug

The development of alginate microparticles for encapsulation of hydrophilic drug is addressed. The alginate microparticles are produced by the emulsification/gelation process. The objective of this study is to optimize the encapsulation of sunset yellow, a model of small hydrophilic drug, by the means of an agile quality by design (QbD) approach. Five input factors are considered: the alginate concentration (2–7% w/V), the drug/polymer ratio (0.1/1–0.5/1), the cross-linker addition flow rate (1–1.6 mL/min), the cross-linker volume (5–10 mL), and the crosslinking time (15–60 min). Critical quality attributes (and their associated specifications) are the particle size (∈30;150µm) and polydispersity, the encapsulation efficiency (>50%), and the drug loading (>10%). The implemented agile method follows three successive QbD sprints, each based on a specific design of experiments (DoE). First, a screening of the process parameters is performed using a Plackett-Burman design, followed in a second sprint by the implementation of a central composite Hartley’s design to identify the design space and to extract four eligible control operating regions where the probability to meet the CQA specifications is above 95%. In the last sprint, one of these optimal operating conditions has been qualified by testing eight end points of the region through the application of a full factorial design. This operating region corresponds to a combination of three factors: alginate concentration in [6.7;7]%, drug/polymer ratio in [0.29;0.34]w/w and a curing time in [40;60]min.

Optimization of the fermentation media and growth conditions of Bacillus velezensis BHZ-29 using a Plackett-Burman design experiment combined with response surface methodology.

Bacillus velezensis occurs extensively in the soil environment. It produces a range of antimicrobial compounds that play an important role in the field of biological control. However, during the actual application process it is often affected by factors such as the medium formulation and fermentation conditions, and therefore biocontrol measures often do not achieve their expected outcomes. In this study, the B. velezensis BHZ-29 strain was used as the research object. The carbon and nitrogen sources, and inorganic salts that affect the number of viable bacteria and antibacterial potency of B. velezensis BHZ-29, were screened by a single factor test. A Plackett-Burman design experiment was conducted to determine the significant factors affecting the number of viable bacteria and antibacterial potency, and a Box-Behnken design experiment was used to obtain the optimal growth of B. velezensis BHZ-29. The medium formula that produced the highest number of viable bacteria and most antibacterial substances was determined. The initial pH, temperature, amount of inoculant, liquid volume, shaking speed, and culture time were determined by a single factor test. The factors that had a significant influence on the number of viable bacteria of B. velezensis BHZ-29 were selected by an orthogonal test. A Box-Behnken design experiment was conducted to obtain the optimal fermentation conditions, and highest number of viable bacteria and antibacterial titer. Molasses, peptone, and magnesium sulfate had significant effects on the viable count and antibacterial titer of B. velezensis BHZ-29. The viable count of B. velezensis BHZ-29 increased from 7.83 × 109 to 2.17 × 1010 CFU/mL, and the antibacterial titer increased from 111.67 to 153.13 mm/mL when the optimal media were used. The optimal fermentation conditions for B. velezensis BHZ-29 were as follows: temperature 25.57°C, pH 7.23, culture time 95.90 h, rotation speed 160 rpm, amount of inoculant 2%, and liquid volume 100 ml. After the optimization of fermentation conditions, the number of viable bacteria increased to 3.39 × 1010 CFU/mL, and the bacteriostatic titer increased to 158.85 mm/ml.The plant height and leaf number of cotton plants treated with BHZ-29 fermentation broth were higher than those of cotton inoculated with Verticillium dahliae. The number of bacteria was 1.15 × 107 CFU/g, and the number of fungi was 1.60 × 105 spores/g. The disease index of the cotton seedlings treated with the optimized fermentation broth was 2.2, and a control effect of 93.8% was achieved. B. velezensis BHZ-29 could reduce the disease index of cotton Verticillium wilt and had a controlling effect on the disease. The best effect was achieved in the treatment group with an inoculation concentration of 2 × 108 CFU/ml, the disease index was 14.50, and a control effect of 84.18% was achieved. The fermentation process parameters of the number of viable bacteria and antibacterial titer by strain B. velezensis BHZ-29 were optimized to lay a foundation for the practical production and application of strain B. velezensis BHZ-29 in agriculture.

Facile synthesis of magnetic layered double hydroxide/metal–organic framework MIL-100(Fe) for simultaneous extraction and enantiomeric determination of three chiral triazole fungicides in water and juice samples

For the first time, magnetic layered double hydroxide/metal–organic framework (Fe3O4@CuZnAl-LDH@MIL-100(Fe)) was successfully fabricated and applied for enrichment of triazole fungicides from water and juice samples prior to chiral HPLC-UV analysis. Several key parameters influencing the extraction efficiency including extraction time, desorption time, sample solution pH, the ionic strength of sample solution, adsorbent amount, desorption volume, desorption solvent and sample volume were investigated by Plackett-Burman design and Box-Behnken design. Under the optimum conditions, the limit of detections (LODs) and limit of quantifications (LOQs) were in the ranges of 0.35 to 1.20 μg L−1 and from 1.00 to 3.80 μg L−1, respectively. The average spiked recoveries and enrichment factors ranged from 86.28-98.22 % and 21.6–23.8, respectively. The linearity was in the range of (5–1000 μg L−1) with satisfactory correlation coefficients (R2) values (0.9987–0.9999) and a good precision (RSDs ≤ 5.74 %). Moreover, the computational simulation was first employed to analyze adsorption properties of MIL-100(Fe) against triazole fungicides. This work provides a promising method for trace chiral pesticide analysis in aqueous samples.

Combining EDTA-intercalated (Zn-Al) layered double hydroxides/graphite and two-phase hollow fiber for liquid phase microextraction of nonsteroidal anti-inflammatory drugs

This study synthesized and characterized layered double hydroxides (LDHs) intercalated with ethylenediaminetetraacetic acid(EDTA) to increase the basal spacing of electrosynthesized Zn-Al-LDHs on pencil graphite (PG). The characteristic verification of the Zn-Al-EDTA-LDH nanostructures was conducted utilizing Fourier-transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM), energy dispersive and X-ray analysis (EDS), and X-ray diffraction (XRD). To improve the extraction efficiency, Zn-Al-EDTA-LDHs as adsorbent in solid phase microextraction (SPME) fiber was combined with hollow fiber liquid–liquid phase microextraction (HF-LLPME) to investigate the extraction efficiency of HF-LLME@PG-SPME technique the chosen non-steroidal anti-inflammatory drugs (NSAIDs) from aqueous solution Several variables, comprising extraction time, desorption time, stirring rate, desorption volume, pH of donor phase, and salt effects were screened and optimized by performing placket-Burman design (PBD). After evaluating the extraction and desorption procedures under optimal conditions, NSAIDs analysis was performed employing high-performance liquid chromatography-ultraviolet detection (HPLC-UV). Compared to the most common clean-up and sample preparation methods, the current study focused on developing a greener analytical chemistry process due to reduced solvent consumption, cost-effectiveness, miniaturization and simplified design. The proposed method for NSAIDs exhibited a good linear dynamic range (LDRs) (1.00–200.00 µg/L) with a coefficient of determination of > 0.9956, low limits of detection (LODs) (0.26–0.29 µg/L) and acceptable limits of quantifications (LOQs) (0.86–0.96 µg/L). Relative standard deviations of 5.31 % could be determined. The absolute recoveries of the urine sample analyses could be obtained in a 89.91–97.61 % range and enrichment factors (EFs) (178.92 to 183.88). Investigations of the developed method were also conducted in various urine samples containing the selected drugs. As a result of the obtained recoveries, the method proved to be valuable and suitable to complicated biological samples.

Application of Plackett-Burman Design in the Screening of Factors Acting on the Sulfuric Leaching Process of Rare Earths from Moroccan Phosphogypsum

As the main waste of the phosphates industry (200–300 million tons per annum produced globally), phosphogypsum (PG) is a major environmental concern (85% of PG is disposed of on land or at sea). The valorization of PG as a secondary source of rare earth elements (REE) is one of the most recent valorization options under investigation. This study investigated the sulfuric acid leaching process of REE from a Moroccan PG. Plackett -Burman design was used to statistically screen significant factors affecting the leaching efficiency of REE named the studied response (Y). Seven factors considered potentially influential were studied as the model variables : Sulfuric acid concentration (X1); Solid to liquid ratio (X2); Drying temperature (X3); Leaching time (X4); Stirring speed (X5); Temperature (X6), Particle size (X7). According to the results, the predicted values of the fitted model were in good agreement with the experimental values by a coefficient of determination (R2) equal to (99 %). drying temperature has a statistically insignificant effect with p=0.336 &gt; 0.05. All the other six variables have significant effects (p &lt; 0.05), but 97,91 % of Y was affected only by the following four factors: X1 (63.71%), X4 (17.94%), X2 (9.98%) , X5 (6.24% ).

Green ecofriendly enhancement of cellulase productivity using agricultural wastes by Aspergillus terreus MN901491: statistical designs and detergent ability on cotton fabrics

BackgroundCellulase is considered a group member of the hydrolytic enzymes, responsible for catalyzing the hydrolysis of cellulose and has various industrial applications. Agricultural wastes are used as an inexpensive source for several utilizable products throughout the world. So, searching for cellulase enzymes from fungal strains capable of utilizing agricultural wastes to increase productivity, reduce costs and overcome waste accumulation in the environment is very important to evaluate its potency as a bio-additive to detergent agents.ResultsIn the current study, the previously identified fungal strain Aspergillus terreus MN901491 was screened and selected for cellulase production. Medium parameters were optimized using one-factor-at-a-time (OFAT) and multi-factorial (Plackett-Burman and Box-Behnken) design methods. OFAT showed the ability of the fungal strain to utilize agricultural wastes (corn cob and rice straw) as a substrate. Also, yeast extract was the best nitrogen source for enhancing cellulase productivity. The most significant variables were determined by Plackett-Burman Design (PBD) and their concentrations were optimized by Response Surface Methodology (RSM) using Box-Behnken Design (BBD). Among eleven independent variables screened by PBD, malt extract, (NH4)2SO4, and KCl were the most significant ones followed by rice straw which affected cellulase production positively. The ANOVA results particularly the R2-value of PBD (0.9879) and BBD (0.9883) confirmed the model efficiency and provided a good interpretation of the experiments. PBD and BBD improved cellulase productivity by 6.1-fold greater than that obtained from OFAT. Medium optimization using OFAT and statistical models increased cellulase production from A. terreus MN901491 by 9.3-fold compared to the non-optimized medium. Moreover, the efficiency of cellulase activity on cotton fabrics as a bio-additive detergent was evaluated and estimated using whiteness and scanning electron microscope (SEM) that affirmed its potential effect and remarkable detergent ability to improve whiteness by 200% in comparison with non-washed fabric and by 190% in comparison with fabric washed by water.ConclusionThe presented work was stabilized as a multi-efficiency in which wastes were used to produce cellulase enzyme from the fungal strain, Aspergillus terreus MN901491 as a bio-additive to detergent applications that involved ecofriendly and green processes.

Eggshell waste bioprocessing for sustainable acid phosphatase production and minimizing environmental hazards

BackgroundThe Environmental Protection Agency has listed eggshell waste as the 15th most significant food industry pollution hazard. Using eggshell waste as a renewable energy source has been a hot topic recently. Therefore, finding a sustainable solution for the recycling and valorization of eggshell waste by investigating its potential to produce acid phosphatase (ACP) and organic acids by the newly-discovered B. sonorensis was the target of the current investigation.ResultsDrawing on both molecular and morphological characterizations, the most potent ACP-producing B. sonorensis strain ACP2, was identified as a local bacterial strain obtained from the effluent of the paper and pulp industries. The use of consecutive statistical experimental approaches of Plackett–Burman Design (PBD) and Orthogonal Central Composite Design (OCCD), followed by pH-uncontrolled cultivation conditions in a 7 L bench-top bioreactor, revealed an innovative medium formulation that substantially improved ACP production, reaching 216 U L−1 with an ACP yield coefficient Yp/x of 18.2 and a specific growth rate (µ) of 0.1 h−1. The metals Ag+, Sn+, and Cr+ were the most efficiently released from eggshells during the solubilization process by B. sonorensis. The uncontrolled pH culture condition is the most suitable and favoured setting for improving ACP and organic acids production. Quantitative and qualitative analyses of the produced organic acids were carried out using liquid chromatography-tandem mass spectrometry (LC–MS/MS). Lactic acid, citric acid, and hydroxybenzoic acid isomer were the most common organic acids produced throughout the cultivation process. The findings of TGA, DSC, SEM, EDS, FTIR, and XRD analysis emphasize the significant influence of organic acids and ACP activity on the solubilization of eggshell particles.ConclusionsThis study emphasized robust microbial engineering approaches for the large-scale production of a newly discovered acid phosphatase, accompanied by organic acids production from B. sonorensis. The biovalorization of the eggshell waste and the production of cost-effective ACP and organic acids were integrated into the current study, and this was done through the implementation of a unique and innovative medium formulation design for eggshell waste management, as well as scaling up ACP production on a bench-top scale.

Plackett–Burman screening of physico-chemical variables affecting Citrus peel-mediated synthesis of silver nanoparticles and their antimicrobial activity

With the growing resistance of pathogenic microbes to traditional drugs, biogenic silver nanoparticles (SNPs) have recently drawn attention as potent antimicrobial agents. In the present study, SNPs synthesized with the aid of orange (Citrus sinensis) peel were engineered by screening variables affecting their properties via Plackett–Burman design. Among the variables screened (temperature, pH, shaking speed, incubation time, peel extract concentration, AgNO3 concentration and extract/AgNO3 volume ratio), pH was the only variable with significant effect on SNPs synthesis. Therefore, SNPs properties could be enhanced to possess highly regular shape with zeta size of 11.44 nm and zeta potential of − 23.7 mV. SNPs purified, capped and stabilized by cloud point extraction technique were then checked for their antimicrobial activity against Bacillus cereus, Listeria innocua, Listeria monocytogenes, Staphylococcus aureus, Enterobacter cloacae, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella typhimurium and Candida albicans. The maximum antimicrobial activity of SNPs was recorded against E. coli, L. monocytogenes and C. albicans with clear zone diameter of 33.2, 31.8 and 31.7 mm, respectively. Based on minimum inhibition concentration and minimum bactericidal concentration of SNPs (300 mg/l) as well as their effect on respiratory chain dehydrogenases, cellular sugar leakage, protein leakage and lipid peroxidation of microbial cells, E. coli was the most affected. Scanning electron microscopy, protein banding and DNA fragmentation proved obvious ultrastructural and molecular alterations of E. coli treated with SNPs. Thus, biogenic SNPs with enhanced properties can be synthesized with the aid of Citrus peel; and such engineered nanoparticles can be used as potent antimicrobial drug against E. coli.

Plackett-Burman and Box-Wilson designs for the removal of mercury (II) by cypress pines and leaves, optimizing the surrounding conditions

In this study, the removal of Hg (II) from cypress leaves and cypress pines was examined. Factors like time of contact (1–60 min), amount of biosorbent used (0.02–0.06 min), pH levels (1.2–6.52 min), initial metal concentration (100–250 ppm) temperature (20–40 min), strength of ions (0.5–1.5 M), and speed of stirring (0–250 min) were studied. Kinetic data were described using both pseudo-first-order and pseudo-second-order models. Equilibrium data were analyzed using Langmuir and Freundlich models, with the Langmuir model fitting the data best. The kinetic data were best described by the pseudo-second-order model. Thermodynamic studies showed that the removal of Hg(II) by cypress leaves was exothermic. The Plackett–Burman design (PBD) and Box–Wilson designs were used for optimization, recommending values of 250 mg/L, 5.98, 7.98 min, and 0.5 g for the variables Co (initial concentration of Hg(II) ions), pH (acidity level), contact time, and mass of KNO3 (potassium nitrate) for the removal of mercury (II) by cypress pines.

Aspergillus niger as an eco-friendly agent for potassium release from K- bearing minerals: Isolation, screening and culture medium optimization using Plackett-Burman design and response surface methodology

The potential of Aspergillus niger, to enhance non-exchangeable potassium (K+) release from mineral structures were investigated as a cost-effective and environmentally friendly alternative to traditional chemical fertilizers. Optimizing the culture medium for maximum K+ release, alongside identifying potential mechanisms of action of the A. niger including the production of various organic acids and pH reduction in the minerals feldspar and phlogopite, were among the primary objectives of the present study. K+ dissolution from feldspar and phlogopite in the presence of Aspergillus niger were examined through a two-step experiment; impact of different carbon sources (glucose, sucrose, and fructose) on K+ release using the Plackett-Burman design (PBD) with 12 experimental runs and effect of other independent variables including pH (ranging from 5 to 10), carbon concentration (3–12.3 g l−1), and incubation time (5–18 days) on K+ release using the central composite design (CCD). Our results indicated that the PBD demonstrated a strong predictive capacity (RMSE = 0.012–0.018 g l−1 and R2 = 0.85–0.89) for K+ release. According to the CCD model, pH exerted a significant positive influence on increasing soluble K+ release (P < 0.001). The highest levels of K+ release (157.8 and 175.3 mg l−1 in feldspar and phlogopite, respectively) were observed at the central levels (0) of time and carbon source, and at the +α level (+1.68) of pH. Furthermore, based on the CCD model, the optimal conditions for achieving high K+ release from feldspar and phlogopite in a medium were pHs of 10.36 and 10.31, sucrose concentrations of 11.23 and 11.32 g l−1, and incubation times of 15 and 18 days, respectively. The determination coefficients of the CCD model indicated that 89.5% and 92.6% of the changes in soluble K+ for feldspar and phlogopite, could be explained by this model, respectively. In the current study, the production of organic acids and the resulting pH reduction, along with the reduction in mineral particle size in feldspar and phlogopite, were identified as potential mechanisms influencing the enhancement of potassium solubility. The predominant acids in both feldspar and phlogopite were lactic acid (70.9 and 69.15 mg l−1) and citric acid (40.48 and 22.93 mg l−1), although the production levels of organic acids differed in the two minerals. Overall, our findings highlight the potential of A. niger to proficiently release non-exchangeable potassium from mineral matrices, indicating its promising potential in agricultural applications.

Optimizing solid phase extraction and HPLC-MS/MS parameters for reliable quantification of COVID-19 pharmaceuticals in Mexico City’s wastewater: A design of experiments approach

Through the COVID-19 health crisis, a plethora of new medications, along with some discontinued pharmaceuticals, were directly tested on population to assess their efficacy in symptom management. Consequently, residues from these treatments were systematically disposed into sewage systems, presenting a novel contamination challenge in urban centers. To effectively address this issue by environmental authorities, the presence of these pollutants in wastewater must be accurately quantified. Herein, we conducted a meticulous study to validate an analytical method based on solid-phase extraction (SPE) and high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS) to detect six pharmaceuticals (metformin, famotidine, indomethacin, dexamethasone, azithromycin, and ivermectin) in wastewater. Using a Plackett-Burman design (PBD), the eight parameters significantly impacting on the HPLC-MS/MS technique were established based on the analytical signal of the six analytes. Subsequently, in conjunction with a Box-Behnken design (BBD), optimal analysis conditions were determined as follows: injection volume: 10 µL, column temperature: 20 °C, capillary voltage: 4100 V, drying gas temperature and flow rate of 350 °C and 7 L min−1 respectively, nebulizer pressure: 15 psi, concentration of formic acid and ammonium acetate in the mobile phase of 0.1 % and 10 mmol/L, respectively. SPE was optimized through a univariable experimental design. OASIS HLB cartridges exhibited the highest extraction efficiency compared to highly polar and non-polar adsorbents. The optimal sample loading conditions were set as 250.0 mL of water sample at pH = 9.0. The washing conditions were 5.0 mL of a 95:5 water:methanol solution, while elution conditions were optimal using 6.0 mL of a 98:2 methanol:formic acid solution. After evaporating the eluate and reconstituting in 1.0 mL of a 98:2 methanol:formic acid solution, a concentration factor of 250 was achieved. The analytical method was validated using for matrices, namely, raw and treated wastewater, wastewater percolated through soil and groundwater. Acceptable levels of precision and accuracy were achieved for all analytes in the tested matrices. Using the validated method, the six pharmaceuticals were quantified in environmental samples, revealing high levels of metformin, dexamethasone, and azithromycin in Mexico City’s sewage. Highly polar compounds displayed mobility as they were present in effluent samples, percolated wastewater, and groundwater.