The sample was homogenized using a grinder (T 25 digital ULTRA-TURRAX®, IKA, Staufen, Germany). After weighing 10 g of the sample, 10 mL of acetonitrile was added to each weighed sample and shaken for 1 min. Thereafter, a QuEChERS extraction kit (magnesium sulfate: 98.5–101.5%; sodium chloride: ≥99.5%; sodium citrate: 99.9%; disodium citrate sesquihydrate: 99%) was added to the sample solution, followed by vigorous shaking for 1 min using a rotary mixer (DE/VIVA, Collomix GmbH, Gaimersheim, Germany). Subsequently, centrifugation was performed for 10 min at 4000 rpm using SORVALL LYNX 4000 (Thermo Scientific, Waltham, MA, USA). Then, 1 mL of the supernatant was put into the QuEChERS dispersive SPE kit (primary secondary amine, octadecyl silane end-capped, magnesium sulfate; 98.5–101.5%), mixed with Mixmate 5353 (Effendorf, Hamburg, Germany) for 1 min, and centrifuged again with Minispin plus 545 (Effendorf, Hamburg, Germany) at 10,000 rpm for 1 min. The liquid separated through this process was filtered with a 0.2 μm PTFE syringe filter (Whatman, Maidstone, UK) and was used as the final sample.
Ptfe syringe filter
Manufactured by Cytiva
Sourced in United Kingdom, United States
The PTFE syringe filter is a laboratory equipment used to filter small sample volumes. It is made of a polytetrafluoroethylene (PTFE) membrane with a pore size suitable for filtration purposes.
Automatically generated - may contain errors
Lab products found in correlation
Whatman, by Cytiva (26 mentions)
Sorvall lynx 4000, by Thermo Fisher Scientific (1 mentions)
Methanol, by Thermo Fisher Scientific (1 mentions)
Milli q gradient a10 system, by Merck Group (1 mentions)
Hplc grade formic acid, by Thermo Fisher Scientific (1 mentions)
Acetonitrile, by Merck Group (1 mentions)
Ammonium carbonate, by Merck Group (1 mentions)
Methanol meoh, by Merck Group (1 mentions)
Ods 2 hypersil, by Thermo Fisher Scientific (1 mentions)
Waters e2695 separations module, by Waters Corporation (1 mentions)
Pda 2996, by Waters Corporation (1 mentions)
Hplc grade, by Thermo Fisher Scientific (1 mentions)
Merlin compact, by Zeiss (1 mentions)
Xs204, by Mettler Toledo (1 mentions)
Gemini c18 column, by Phenomenex (1 mentions)
Cary 50 scan, by Agilent Technologies (1 mentions)
Thermomixer comfort, by Eppendorf (1 mentions)
Nanodrop one, by Thermo Fisher Scientific (1 mentions)
Uv 1700 pharmaspec, by Shimadzu (1 mentions)
26 protocols using ptfe syringe filter
1
QuEChERS Extraction of Food Samples
2
Preparative SEC Purification of PAHs
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Size-exclusion chromatography (SEC) of PAHs was conducted by injecting 5 mL of a PAH solution in CHCl3 (100 mg mL−1) to a Recycling Preparative HPLC (LC-9260 NEXT, Japan Analytical Industry) system equipped with JAIGEL-2.5 H/2 H/3 H columns and a differential refractometer. Chloroform was used as an eluent with a flow rate of 3.5 mL min−1. Before injection, the solution was filtered through a PTFE syringe filter (Whatman, 0.2 µm pore). The SEC was performed under a cycling mode until the coinciding peaks were separated. The desired fraction was collected using a fraction collector. Two prep-SEC systems ran in parallel, giving the maximum capacity of separation of 1 g.
3
Phytochemical extraction and fractionation
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Dried bark or leaves of collecting plants were crushed in a mixer separately. Two grams of each sample was extracted thrice with 75 mL of 100% methanol under sonication for 90 min, each. The extract was filtered with a filter paper (Whatman No. 2, USA). The final volume was adjusted to 75 mL with 100% methanol and filtered through a 0.45 μm PTFE syringe filter (Whatman, New York, NY, USA) before HPLC analysis. The filtrate was dried under a stream of N2 gas and then used in pancreatic lipase assay. The air-dried bark of FM (960 g) was ground and extracted in sonication with 100% methanol. The extract was filtered and concentrated in vacuo. The resulting crude extract (187 g) was suspended in water and fractionated with n-hexane, dichloromethane, ethyl acetate and n-butanol, successively to yield n-hexane fr. (6.4%, w/w), CH2Cl2 fr. (3.2%), EtOAc fr. (32.1%), n-BuOH fr. (47.1%), and aqueous fr. (7.5%) fractions, respectively [26 ].
4
Biocompatible PVA:CS Blended Solution
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A biocompatible PVA:CS blended solution was prepared by mixing a PVA solution and a chitosan solution, respectively. A PVA solution was synthesized by dissolving a mixture of 10 wt% PVA powder and 3 mL diluted deionized (DI) water. In addition, a chitosan solution was synthesized by dissolving a mixture of 2 wt% chitosan powder based on the shrimp shell, 2% acetic acid solution, and 10 mL DI water. The PVA solution and chitosan solution were completely dissolved using a magnetic stirrer with 600 rpm at 90 °C for 2 h and 800 rpm at 50 °C for 6 h, respectively. The solutions were then filtered using a 5 μm pore size polytetrafluoroethylene (PTFE) syringe filter (Whatman, International Ltd., Maidstone, UK) to remove impurities. Finally, the two solutions were completely mixed in a 1:1 ratio using a magnetic stirrer at 800 rpm at room temperature for 6 h. Figure 1 a shows the fabrication details of the PVA:CS blended solution.
5
Enzymatic Hydrolysis and Lactic Acid Fermentation
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The enzymatic hydrolysis of KCW was conducted in a 100-mL total volume containing 1.5–6.0% (w/v) dry matter, cellulase (5.6 mg/g KCW), pectinase (4.2 mg/g KCW), 2.0% yeast extract, 0.5% sodium acetate, 0.2% dipotassium phosphate, 0.01% magnesium sulfate, and 0.005% manganese sulfate at E°C for 24 h. The fermentation of 40 mL hydrolysates (1.5–6.0%) with 0.4 mL LAB—W. cibaria WiKim28, L. sakei WiKim31, and L. curvatus WiKim38, respectively—was performed at 30°C for 6 days. Culture broths of LAB were centrifuged at 8000 rpm (5810R, fixed angle type; Eppendorf, USA), serially diluted, and filtered through a 0.45-μm PTFE syringe filter (Whatman, USA). The content of organic acid was measured by HPLC (Waters Alliance e2695 system, USA) at 30°C using an Aminex HPX-87H column (300 mm × 7.8 mm, Bio-Rad, Hercules, CA, USA). Elution was carried out isocratically using 5 mmol/L sulfuric acid. The flow rate and detection wavelength were 0.6 mL/min and 210 nm, respectively. Quantitative analysis of organic acid was performed using standard curves.
6
Buccal Film Drug Content Analysis
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The drug content in the buccal film was determined as follows: three films, each measuring 1.5 cm2, were dissolved in their respective mobile phases and sonicated for 10 min to ensure complete dissolution. After that, the solutions were filtered by a 0.45 μm PTFE syringe filter (Whatman, Maidstone, UK) for HPLC analysis.
7
Extraction of Sulfate Metabolites
8
Synthesis of LEU-Oleic Acid Conjugate
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LEU acetate (127 mg) was dissolved in 4 mL of deionized water (DW) and stirred at 700 rpm until completely dissolved. Then, 14 µL of triethylamine was dissolved in LEU acetate solution to desorb the acetate anions. C18 fatty acids (25 mM) were dissolved in 4 mL of THF and stirred at 700 rpm until completely dissolved. To deprotonate the carboxylic group of fatty acids for the reaction, triethylamine (14 µL) was added into fatty acids/THF solution and followed by the adding of benzoyl chloride (12 µL) and DMAP (1.22 mg). Herein, the deprotonated fatty acids reacted with benzoyl chloride to form the acid anhydride. Then, the obtained acid anhydride was attacked with DMAP, forming another leaving group (fatty acid-DMAP reactive) and then it was replaced by the hydroxyl group of LEU. Yamaguchi chloride (2,4,6-tribenzoyl chloride) used in the existing mechanism causes steric hindrance, therefore benzoyl chloride was used instead.25 (link)
Figure 1 shows a schematic diagram for the hydroxyl group targeted LEU-oleic acid conjugate (LOC). After stirring for 1 h, the mixture was injected into the DW through a 0.45 µm PTFE syringe filter (Whatman, England) and stirred at 700 rpm for 3 h. Finally, the solution was dialyzed against absolute ethanol to purify the unreacted substances.![]()
Schematic diagram for the hydroxyl group targeted LEU-oleic acid conjugate (LOC).
9
Antioxidant Capacity Determination Protocol
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Methanol, sodium carbonate, gallic acid, Folin-Ciocalteu′s phenol reagent, DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS (2,2′ azino-bis (3-ethylbenzothiazoline-6-sulfonic-acid), potassium persulphate, sodium hydroxide (NaOH) and Trolox (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid) were obtained from Fluka (Buchs, Switzerland). HPLC-grade water was obtained by purifying double distilled water in a Milli-Q Gradient A10 system (Millipore, Bedford, MA, USA), 0.45 µm PTFE syringe filter (Whatman, Milan, Italy).
10
HPLC-MS Sample Preparation Protocol
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Methanol (MeOH) and acetonitrile were HPLC grade and were both purchased from Merck (Darmstadt, Germany). Ammonium carbonate used in the sample extraction was purchased from Merck (Darmstadt, Germany). HPLC-grade formic acid was purchased from Fisher Chemical (Toronto, ON, Canada). Ammonium formate for LC/MS was obtained from Honeywell (Charlotte, NC, USA). Magnesium sulfate (MgSO4) and sodium chloride (NaCl) were used to extract salt, and primary and secondary amine (PSA), C18 and MgSO4, which were used for clean-up, were purchased from Bekolut GmbH & Co (Hauptstuhl, Germany). Direct-Q®3 UV with pump devices from Millipore (Bedford, MA, USA) was used to purify the demineralized water. A polytetrafluoroethylene (PTFE) syringe filter from Whatman (Maidstone, UK) was used for filtration.
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