Most materials were used without further purification. Benzamide (BA) (99%), PIC (98%), NIC (98%), iNIC (99%), INH (≥99%), benzhydrazide (BHZ) (98%), isooctane (2,2,4 trimethylpentane, 99.8%), methanol (≥99.9%), activated charcoal (99.5%), chloroform (≥99.5%), deuterium oxide (99.9%), 2,2-dimethyl-2-si-lapentane-5-sulfonate sodium salt (DSS, 97%), monosodium phosphate (≥99.0%), disodium phosphate (≥99.0%), sodium hydroxide (≥98%), and hydrochloric acid (37%) were all purchased from Sigma-Aldrich. DPPC (≥99%) and DPPE (99%) were purchased from Avanti Polar Lipids. Sodium AOT (aerosol-OT, bis(2-ethylhexyl)sulfosuccinate sodium salt, ≥99.0%) was purchased from Sigma-Aldrich and was purified further as has been reported previously to remove any acidic impurities.30 (link) Briefly, 50.0 g AOT was dissolved into 150 mL of methanol to which 15 g activated charcoal was added. This suspension was stirred for 2 weeks. After mixing, the suspension was filtered to remove the activated charcoal. The filtrate was then dried under rotary evaporation at 50 °C until the water content was below 0.2 molecules of water per AOT as determined by 1H NMR spectroscopy. The pH was adjusted throughout this study using varying concentrations of NaOH or HC1 dissolved/mixed in either D2O or H2O depending on experimental requirements. NaOH or HC1 dissolved in D2O is referred to as NaOD or DC1 respectively.
Activated charcoal
Manufactured by Merck Group
Sourced in United States, Germany, Canada, United Kingdom, Poland, India, Sao Tome and Principe, Italy, Switzerland
Activated charcoal is a highly porous form of carbon that has been treated to increase its adsorptive properties. It is a versatile lab equipment used for a variety of applications, including the removal of impurities, the purification of substances, and the separation of compounds.
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Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (7 mentions)
Methanol, by Merck Group (6 mentions)
Ethanol, by Merck Group (5 mentions)
Sodium chloride (nacl), by Merck Group (5 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (4 mentions)
Sucrose, by Merck Group (4 mentions)
Glycerol, by Merck Group (4 mentions)
Ethyl acetate, by Merck Group (4 mentions)
Hydrochloric acid (hcl), by Merck Group (4 mentions)
Acetonitrile, by Merck Group (3 mentions)
Chloroform, by Merck Group (3 mentions)
Trypan blue, by Merck Group (3 mentions)
Adipic acid dihydrazide, by Merck Group (3 mentions)
Polyethyleneimine cellulose plate, by Merck Group (3 mentions)
Mgcl2, by Merck Group (3 mentions)
Kod plus mutagenesis kit, by Toyobo (3 mentions)
Methanol, by Thermo Fisher Scientific (3 mentions)
Sodium hydroxide (naoh), by Merck Group (3 mentions)
Chloroform, by Chempur (2 mentions)
Fed400, by Binder (2 mentions)
110 protocols using activated charcoal
1
Purification and Characterization of Sodium AOT
2
Embryo Development from Microspores with Antibiotics
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Five mL of a microspore suspension (microspore isolation 4.3) in the NLN-13 medium was placed in a Petri dish with a diameter of 6 mm. Before placing a microspore suspension, we added 3–4 drops of sterile agarose solution of activated charcoal (Sigma-Aldrich, St. Louis, MO, USA) melted in a microwave (1 g activated charcoal per 100 mL of 0.5% agarose solution) in each Petri dish. After solidification of agarose with activated carbon, a suspension of microspores and antibiotic effluents was added to Petri dishes until the final concentration of 50, 100, or 1000 mg/L. As a control, there were dishes to which antibiotics were not added.
The dishes were incubated in darkness for 2 days at 32 °C and then continuously at 25 °C about 21 days when the embryos formed. For each of the genotypes, at least three independent experiments were performed in four replications for each variation. To analyze the effect of antibiotics to the development of microspores in embryos, we considered experiments where the control did not show the development of bacteria and inhibition of the development of embryos.
The dishes were incubated in darkness for 2 days at 32 °C and then continuously at 25 °C about 21 days when the embryos formed. For each of the genotypes, at least three independent experiments were performed in four replications for each variation. To analyze the effect of antibiotics to the development of microspores in embryos, we considered experiments where the control did not show the development of bacteria and inhibition of the development of embryos.
3
Reactivation of Virus Inhibitor CVI
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Immediately before use, guanidine hydrochloride (6 mol/l) was cleared over night by activated charcoal (Merck, Darmstadt, Germany) and adjusted to pH 8.0 by adding 50 mmol/l tris(hydroxymethyl)aminomethane (Tris). Freeze dried CVI (10 mg/ml) was mixed with 1 ml Tris-buffered guanidine-hydrochloride, incubated at room temperature for 1 h with frequent thorough vortex followed by 1 h at 37°C. The buffer from the clear supernatant after centrifugation (10.000 x g, 10 min, 4°C) was replaced for 150 mmol/l HAc by gelfiltration (Sephadex-G25; GE Healthcare, Munich, Germany). This procedure regains the biological activity of CVI on CFSC (S1 Fig ).
4
Bacterial Production and Purification of P(3HB)
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Polyhydroxybutyrate (P(3HB)) was produced by bacterial fermentation from glycerol (Orlen Południe S.A., Trzebinia, Poland) in the presence of NaCl (Chempur, Piekary Śląskie, Poland) at 45 °C using bacterial strain Zobellella denitrificans (Wilhelms-Universität Münster, Germany) as described in [28 (link)]. The biomass after fermentation was lyophilized and then extracted with chloroform (Chempur, Piekary Śląskie, Poland). The resulting solution was passed through activated charcoal (Merck, Warsaw, Poland) and 0.2 µm polytetrafluoroethylene (PTFE) filter (Avantor, Gdańsk, Poland). Next, the P(3HB) solution was concentrated on a rotatory evaporator (Heidolph Hei-VAP Industrial B, Heidolph Instruments GmbH & Co. KG, Schwabach, Germany), precipitated in an ice-cold methanol solution (Chempur, Piekary Śląskie, Poland) and dried in an oven (Binder FED400, Binder GmbH, Tuttlingen, Germany). The cleaned-up polymer was reconstituted in chloroform (5% w/v) for further experiments.
5
Quantitative Analysis of Food Compounds
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Acrylamide (purum, ≥98%, GC), methAcrylamide (purum, >98%, GC), acetonitrile (for HPLC), as well as standards for HPLC (glucose, lactose, lactic acid and acetic acid) were provided by Sigma-Aldrich (Sp. z o.o, Poznan, Poland). Activated charcoal (for analysis) and silica gel 60 (0.040–0.063 mm, for column chromatography) were obtained from Merck Sp. z o.o. (Warsaw, Poland), while methanol (HPLC-suitable), sodium chloride, ethyl acetate, sulphuric acid, and sodium sulphate anhydrous (all pure p.a.) from Avantor Performance Materials Poland S.A. lactose, as well as Carrez’s Reagent I (1 mol/L of zinc acetate in 0.5 mol/L acetic acid solution) and Carrez’s Reagent II (aqueous 0.25 mol/L solution of potassium hexacyanoferrate(II)), used for precipitation and clarification, were provided by Chempur (Piekary Śląskie, Poland). Casein was obtained from Pol-Aura, Chemicals (Olsztyn, Poland).
6
Biphenyl Synthesis and Ionic Liquid Preparation
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For the CTF-biphenyl synthesis, anhydrous aluminum chloride (AlCl3; 98.5%) was received from Arcos Organics, cyanuric chloride (99%) was obtained from Sigma-Aldrich, and biphenyl (99%) from Alfa Aesar. biphenyl was recrystallized from ethanol before usage. For the synthesis of the IL, [BMIm][NTf2] 1-chlorobutane (99%) was purchased from Merck, 1-methylimidazole (99%) and lithium bis(trifluoromethanesulfonyl)imide (Li[NTf2]; 99%) were obtained from Fluorochem, activated charcoal (extra pure) from Merck, and acidic aluminum oxide (150 m2/g; Brockmann grade 1) from Alfa Aesar. The solvents acetone (≥99.8%), dichloromethane (DCM; 99.99%), and ethyl acetate (≥99%) were received from Fisher Scientific; and methanol (MeOH; ≥99.8%) and tetrahydrofuran (THF; ≥99.9%) from Sigma-Aldrich. The polymer Matrimid® 5218 (BTDA/DAPI) was provided by Huntsman Advanced Materials. The gases CO2 (grade 4.5), CH4 (grade 4.5), and He (grade 5.0) were received from Air Liquide.
7
Loperamide-Based Antidiarrheal Protocol
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Loperamide (Lopax, Axar Pharmaceuticals, Baroda, India) was used as a standard antidiarrheal drug. Castor oil (Fine, Mumbai, India) and prostaglandin E2 (PGE2) (Sigma-Aldrich Chemical Pvt. Ltd., USA) as diarrhea-inducing agents, activated charcoal (Merck, India) as an intestinal transit marker, and Gum Acacia (Fine Chem, Boisar, India) and Tragacanth powder (Central Drug House Pvt. Ltd., Bombay, India) as suspension agents were used in this study.
8
Immobilization of Bacterial Cells on Carrier Materials
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Activated charcoal (Merck), saw dust (of Sesamum indicum obtained from local saw mill) and fly ash (collected from thermal power plant) were procured to use as carrier material for immobilization of bacterial cells.
To determine pH, 10 g solid carrier was mixed with 90 mL of distilled water with thorough stirring and pH of mixture was determined.27 The moisture content of each carrier was determined on a wet and dry weight basis. Water-holding capacity of each carrier was determined by adding 100 g oven-dried carrier material in a 500 mL size beaker containing appropriate amount of water for dipping the material. Saturated material was decanted on gauze to remove extra water and weighed.27
The bacterial suspension used for immobilization contained 12 h grown cells of the screened bacteria incubated at 28 °C in the nutrient broth. Broth cultures were added separately to each carrier in the ratio 1:3 and thoroughly mixed to obtain a homogeneous carrier-inoculum mixture. This mixture was spread in the trays and kept for 48 h curing at room temperature.27 Carrier with un-inoculated broth served as control. Each inoculated and uninoculated carrier was filled up to 1/3 in 60 cm glass columns (Borosil). The column diameter used was 7 cm so as to allow the effluent run at an optimum flow rate of 5 mL min−1.28
To determine pH, 10 g solid carrier was mixed with 90 mL of distilled water with thorough stirring and pH of mixture was determined.27 The moisture content of each carrier was determined on a wet and dry weight basis. Water-holding capacity of each carrier was determined by adding 100 g oven-dried carrier material in a 500 mL size beaker containing appropriate amount of water for dipping the material. Saturated material was decanted on gauze to remove extra water and weighed.27
The bacterial suspension used for immobilization contained 12 h grown cells of the screened bacteria incubated at 28 °C in the nutrient broth. Broth cultures were added separately to each carrier in the ratio 1:3 and thoroughly mixed to obtain a homogeneous carrier-inoculum mixture. This mixture was spread in the trays and kept for 48 h curing at room temperature.27 Carrier with un-inoculated broth served as control. Each inoculated and uninoculated carrier was filled up to 1/3 in 60 cm glass columns (Borosil). The column diameter used was 7 cm so as to allow the effluent run at an optimum flow rate of 5 mL min−1.28
9
Bacterial Extraction and Purification of P(3HB)
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Poly(3-hydroxybutyrate) (P(3HB)) was produced by bacterial fermentation from glycerol (Orlen Południe S.A., Trzebinia, Poland) in the presence of NaCl (Chempur, Piekary Śląskie, Poland) at 45 °C using bacterial strain Zobellella denitrificans (Wilhelms-Universität Münster, Münster, Germany), as described previously [59 (link)]. The biomass after fermentation was lyophilized and then extracted with chloroform (Chempur, Piekary Śląskie, Poland). The resulting solution was filtered through activated charcoal (Merck, Warsaw, Poland) and 0.2 µm polytetrafluoroethylene (PTFE) filter (Avantor, Gdańsk, Poland). Next, the P(3HB) solution was concentrated on a rotatory evaporator (Heidolph Hei-VAP Industrial B, Heidolph Instruments GmbH & Co. KG, Schwabach, Germany), precipitated in an ice-cold methanol solution (Chempur, Piekary Śląskie, Poland), and dried in an oven (Binder FED400, Binder GmbH, Tuttlingen, Germany). The purified polymer was reconstituted in chloroform (5% w/v) for further experiments.
10
Listeria monocytogenes Cultivation and Infection Preparation
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Bacterial strains used in the work are listed in Table 1 . L. monocytogenes were routinely cultivated in the Brain Heart Infusion (BHI, BD, USA) medium. To get a virulence regulon induction, L. monocytogenes was grown in BHI supplemented with 0,2% activated charcoal (Merck) [26 (link)]. E. coli used in cloning and expression procedures were cultivated in the LB medium (Sigma-Aldrich). All bacteria were grown at 37°C with agitation at 180 rpm. Antibiotics were added up to the following concentrations: erythromycin up to 10 and 300 μg ml−1 for L. monocytogenes and E. coli, respectively; ampicillin up to 100 μg ml−1; kanamycin up to 200 μg ml−1. All antibiotics were purchased from Sigma-Aldrich. To prepare a culture for infection, bacteria were grown up to mid-exponential phase, washed with PBS, aliquoted, and frozen in the presence of 10% glycerin. The concentration was determined by plating serial dilutions from the frozen culture the day before the experiment. Bacteria were thawed immediately before the experiment and resuspended in PBS up to the required concentration.
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