N 1 naphthyl ethylenediamine dihydrochloride

The chemicals gallic acid, Folin ciocalteu reagent, trichloroacetic acid, sodium salicylate and ethylenediamine tetra acetic acid (EDTA) were purchased from Sigma Chemicals Co. (P.O. Box 14508, St. Louis, MO 63178 USA). 1,1-Diphenyl-2-picrylhydrazyl (DPPH) free radical, (−)-epigallocatechin gallate, aluminium chloride and sulfanilamide were purchased by Fluka (Flukachemie GmbH, CH-9471 Buchs). L-ascorbic acid, hydrogen peroxide, N-(1-naphthyl)-ethylenediamine dihydrochloride and ethanol were purchased from BDH Chemicals (BDH Chemicals Ltd, Poole, England). Ferric chloride, potassium ferricyanide and sodium nitrite were purchased from Riedel De Haen Ag, Wunstorfer Strasse 40, SEELZE1, D3016, Germany.
Lactate dehydrogenase (LDH) enzyme assay kit was purchased from DiaSys (Alte Strasse 9, 65558, Holzheim, Germany). Alanine transaminase (ALT) and Aspartate transaminase (AST) enzyme assay kits were purchased from POINTE, SCIENTIFIC, INC (5449 Research Drive, Canton MI 48188, USA).

The chemicals gallic acid, Folin Ciocalteu reagent, trichloroacetic acid, 2-deoxy-D-ribose and ethylenediamine tetra acetic acid (EDTA) were purchased from Sigma Chemicals Co. (P.O. Box 14508, St. Louis, MO 63178 USA). 1,1-Diphenyl-2-picrylhydrazyl (DPPH) free radical, (-)-epigallocatechin gallate, aluminium chloride and sulfanilamide were purchased by Fluka (Fluka chemie GmbH, CH-9471 Buchs). L-ascorbic acid, hydrogen peroxide, N-(1-naphthyl)-ethylene diamine dihydrochloride and ethanol were purchased from BDH Chemicals (BDH Chemicals Ltd, Poole, England). Sodium nitroprusside was purchased from Qualigens (A division of GlaxoSmith Kline Pharmaceuticals Ltd). Ferric chloride, potassium ferricyanide and sodium nitrite were purchased from Riedel De Haen Ag, Wunstorfer Strasse 40, SEELZE1, D3016, Germany.
Lactate dehydrogenase (LDH) enzyme assay kit was purchased from DiaSys (Alte Strasse 9, 65558, Holzheim, Germany). Alanine transaminase (ALT) and Aspartate transaminase (AST) enzyme assay kits were purchased from POINTE, SCIENTIFIC, INC (5449 Research Drive, Canton MI 48188, USA).
SHIMADZU UV 1601 UV Visible spectrophotometer (Shimadzu Corporation, Kyoto, Japan) was used to read the absorbance. Deionized water was obtained from LABCONCO WATER PRO-PS UV ultra filtered water system (LABCONCO Corporation, Kansas city, Missouri).

HD11 cells were stimulated with ODN-2006 as described above. Supernatant was collected to measure NO production. Briefly, 30 μL of sample were added to the well in a 96-well flat bottom plate. An equal volume of 1% sulfanilamide (Merck, Darmstadt, Germany) was added in each well, followed by 30 μL 0.1% N-(1-naphthyl) ethylenediamine dihydrochloride (VWR) at room temperature for 5 min. The nitrite concentration was determined by measuring optical density at 550 nm. Sodium nitrite (Merck, Darmstadt, Germany,) was used as a standard to accurately determine the nitrite concentration in the cell supernatant.