Nah2po4 2h2o

A sample of 25 mg of tissue was homogenized using a solution of 1.15% KCl (Merck, Darmstadt, Germany). The homogenates were centrifuged at 4,000 rpm for 30 min at 4°C and the supernatants were separated by decantation for the measurement of GSH-Px and MDA.
Another 25 mg of tissue was washed with isotonic sodium chloride (NaCl; IE Ulagay) for tGSH analysis. First, the NaCl was removed from the samples, then the final volume was brought up to 2 ml by adding a phosphate buffer solution [0.213 g of NaH₂PO₄∙2H₂O, 1.563 g of Na₂HPO₄∙2H₂O (Merck), 0.038 g of EDTA (Sigma-Aldrich) in 100 ml of distilled water at pH 7.4].
The tissues were homogenized in an icy environment and centrifuged at 1,000 rpm for 15 min at a temperature of 4°C. The supernatants were separated and used for the measurement of protein concentration according to the method described by Bradford [13 (link)].

All
reagents were of analytical
grade and used without further purification. All of the reagents were
prepared using Millipore water (18.2 MΩ·cm). Proline (99%), l-leucine (98%), phenylalanine (98%), threonine (98%), arginine
(98%), asparagine (98%), glycine (99%), valine (98%), alanine (98%),
methionine (98%), tryptophan (98%), histidine (98%), acrylamide (99%),
NaOH, CH₃COONa·3H₂O (99.5%), Na₂HPO₄·H₂O (99%), NaH₂PO₄·2H₂O (99%), and KCl (99%) were purchased
from Merck. Methylene blue monohydrate (96%) was purchased from Acros;
HCl (33–37%) and (NH₄)₂SO₄ (99%) were purchased from Fisher Scientific.

The growth conditions of the Caco‐2 cultures and adhesion assays were carried out as described previously (Lebeer et al., 2012). Caco‐2 cells were incubated with 1 ml fresh bacterial culture or rehydrated spray‐dried powder [10⁷ CFU ml⁻¹ in Dulbecco's Modified Eagle Medium (Life Technologies, Waltham, MA, USA) without 10% foetal bovine serum (Thermo Fisher, Asse, Belgium)] for 1 h at 37°C, 5% CO₂, 100% humidity. Then, the cells were washed by adding prewarmed phosphate‐buffered saline (PBS), containing 8.2 g l⁻¹ NaCl (Fagron, Waregem, Belgium), 0.3 g l⁻¹ NaH₂PO₄.2H₂O (Merck kGaA, Darmstadt, Germany) and 1.54 g l⁻¹ Na₂HPO₄.2H₂O (Merck kGaA). To determine the number of viable adhered bacteria, appropriated serial dilutions of the remained adhered bacterial cells in PBS were plated n MRS agar in triplicate and the CFU were enumerated after 48 h. The adhesion ratio was calculated by comparing the number of cells present in the bacterial suspension which was added initially to the Caco‐2 cells, to the bacterial colonies counted that adhered to the Caco‐2 cells.