Luteolin

DMEM/F12, DMEM high glucose, and RPMI 1640 cell culture media were purchased from Gibco (Thermo Fisher Scientific, Waltham, MA, USA). 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was obtained from Alfa Aesar (Waltham, MA, USA). Luteolin and lucidin were purchased from PhytoLab (Vestenbergsgreuth, Germany), and alizarin and taxifolin were acquired from Sigma Aldrich (St. Louis, MO, USA). Propidium iodide solution and Hoechst 33,342 were also bought from Sigma Aldrich (St. Louis, MO, USA). A Caspase 3/7 Glo Kit was obtained from Promega (Madison, WI, USA). All the other chemicals and solvents obtained commercially were of analytical grade. All solutions were freshly prepared by using ultra-pure grade water purified with a Milli-Q system from Millipore (Billerica, MA, USA).

Orientin, isoOrientin, vitexin, isovitexin, schaftoside, isoschaftoside, luteolin, apigenin, and quercetin (>95%, reference substances) were purchased from Phytolab (Vestenbergsgreuth, Germany). Caco-2 cells were obtained from the ATCC (American Type Culture Collection, Manassas, VA, USA). Acetonitrile (LiChrosolv^®), trifluoroacetic acid (TFA), dimethyl sulfoxide (DMSO), formic acid, sodium acetate, sulfatase (from Helix pomatia, type H-1, lyophilized, >10,000 U/g), β-glucuronidase (from bovine liver, type B-10, 10,100 U/g) methanol (LiChrosolv^®), 3-(4,5-dimethylthiazolyl)-2,5-diphenyl tetrazolium bromide (MTT), and sodium dodecyl sulfate (SDS) were purchased from Merck (Darmstadt, Germany). Hanks’ balanced salt solution (HBSS) was obtained from Biowest (Nuaillé, France). Dulbecco’s modified Eagle medium (DMEM) was purchased from Life Technologies (Carlsbad, CA, USA). Phosphate-buffered saline (PBS), fetal bovine serum (FBS), trypsin-ethylenediaminetetraacetic acid (trypsin-EDTA), nonessential amino acids (NEAAs), penicillin, and streptomycin were purchased from Biochrom AG (Berlin, Germany). Transwell^® plates (insert diameter 12 mm, pore size 3.0 µm, membrane growth area 1.12 cm²) were obtained from Corning Costar (Cambridge, MA, USA), and 96-well plates were purchased from TPP (Trasadingen, Switzerland).

S. bachtiarica aerial parts were used for polyphenolic compound determination based on standards including gallic acid, vanillic acid, caffeic acid, ferulic acid, p-coumaric acid, quercetin, luteolin, and apigenin (Phytolab, Germany, 98% purity). Methanolic extraction was applied, in which 2g of leaf samples was used and after grinding were extracted using 20 mL of methanol (80%), and shaken at 25 °C for 20 h. The extraction was repeated twice and the air-dried extract was dissolved in HPLC solvent A (1 mL), filtered (0.22 μm disk), and 20 μL was injected to an Agilent 1090 system with detection range of 260 and 350 nm. A 250 × 4.6 mm, 5 μm, symmetry C18 column (Waters Crop., Milford, MA, USA) was applied in this experiment. The mobile phase included formic acid (99.9:0.1) as a solution (A) and acetonitrile/formic acid (99.9:0.1) as a solution (B) with gradient elution at 25 °C and flow rate of 0.8 mL min⁻¹. The gradient program started from A: B (90:10) for 1 min, followed by 10–26% B for 40 min, 26–65% B for 30 min, and finally 65–100% B for 5 min followed by equilibration with 0–90% A for 4 min. Polyphenolic components were determined by comparing UV spectra and the retention times with pure standards, and the amount was reported in mg per 100 g of dry sample weight.