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After centrifugation, supernatants from both extractions were combined and washed with 2 mL of saturated NaCl.
Briefly, dry extracts were resuspended in 0.5 ml of methanolic base, vortexed, and incubated at 100°C for 3 min, followed by addition of 14% BF3 in methanol (0.5 ml), vortexing, incubation at 100°C for 1 min, addition of hexane (0.5 ml), vortexing, incubation at 100°C for 1 min, and addition of 6.5 ml of saturated NaCl.
Large scale cultures (1 L volume in 2 L baffled shake flasks) were inoculated with 10 mL of saturated overnight cultures per liter.
The reaction mixture was stirred at room temperature for 10 min. After completion of the reaction (monitored by TLC), the reaction mixture was diluted with CH2Cl2 (20 mL), and the organic layer was then washed 2 times with 5 mL of saturated aqueous NaHCO3.
Briefly, 20 30 mg of gastric tissue sample were saponified with 1.0 ml NaOH in methanol and the FAME were prepared with 2 mL 14% boron trifluoride in methanol following extraction with hexane after washing with 3.0 ml of saturated NaCl.
35 g of AMPS was dissolved in 300 ml of saturated solution of Ca(OH 2 with stirring at 25 ± 2 °C.
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A 100 ml stock solution of saturated sodium chloride (salt solution) was prepared with distilled water and 38 g of NaCl; a 100 ml stock solution of 5M sodium hydroxide (base solution) was prepared with distilled water and 20 g of NaOH; and a 100 ml stock solution of 5M hydrochloric acid (acid solution) was prepared by diluting concentrated hydrochloric acid with distilled water.
The supernatant was added to 12 ml of 40% saturated sucrose solution and centrifuged at 95,000 × g at 4 °C for 1 h.
The optimum conditions were: 20 min of extraction at 70 °C using 60 mg of the cork sample and 10 mL of water saturated with sodium chloride in a 20 mL amber vial with constant magnetic stirring.
The reaction mixture was extracted twice with 2.0 ml of water saturated n-butanol.
Crystals were removed by filtering through a Buchner funnel to remove the urea complexes, which were washed twice with 25 ml of methanol saturated with urea.
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