反应详情

温度

80 °C

FaSSIF (Fasted State Simulated Intestinal Fluid) was prepared based on blank FaSSIF which is a phosphate buffer obtained by dissolving 0.696 g NaOH (BDH Laboratory Supplies, Poole, England), 7.908 g NaH2PO4.H2O (Merck, Darmstadt, Germany) and 12.37 g NaCl in 2 L of purified water (18.2 MΩ, Elga, Tex., USA). The pH was adjusted to exactly 6.5 with 1M NaOH. FaSSIF was created by adding 3.23 g sodium taurocholate (ICN Biomedicals, Eschwege, Germany) and 5.9 mL of a solution of lecithin (YDS Chemicals, Heusden, Belgium) in chloroform (100 mg·mL−1) to approximately 200 mL blank FaSSIF. This mixture was heated to 80° C. for 15 minutes to remove all chloroform, after which a clear solution was obtained. Subsequently, blank FaSSIF was added up to a volume of 11. Some experiments were performed with MES buffered FaSSIF to allow the addition of SGF (simulated gastric fluid, 0.1M HCl containing 0.2 wt. % NaCl) without a significant pH change. MES [2-(N-morpholino)-ethanesulfonic acid, Sigma-Aldrich, Steinheim, Germany] (977.3 mg), sodium taurocholate (179.1 mg) and lecithin solution (655.6 μl) were added to prepare 100 ml of MES-FaSSIF according to the same protocol as mentioned before. As receiver medium for the Caco-2 transport experiments, 0.2 wt. % TPGS (D-α-tocopherol polyethylene glycol 1000 succinate, Eastman, Anglesey, England) in transport medium (TM) [HBSS (Hanks' Balanced Salt Solution) supplemented with glucose (to obtain a final concentration of 25 mM, Sigma-Aldrich, Steinheim, Germany) and Hepes-buffer (10 mM, pH 7.4, N-2-hydroxyethyl-piperazine-N′-2-ethanesulfonic acid, Lonza, Verviers, Belgium)] was used. To maintain sink conditions, TPGS was included in the medium added to the basolateral side of the Caco-2 cell monolayer.

1. customFaSSIF (Fasted State Simulated Intestinal Fluid) was prepared
2. customobtained
3. customto remove all chloroform
4. customafter which a clear solution was obtained
5. customto prepare 100 ml of MES-FaSSIF according to the same protocol