Butylhydroxytoluene

All-trans-β-carotene (Type II synthetic, purity >95%), lutein from marigold, β-cryptoxanthin (purity >97%), zeaxanthin (purity >95%), lycopene from tomato (purity >90%), butylhydroxytoluene (BHT, purity ≥99%) and triethylamine (TEA, purity ≥99%) were obtained from Sigma-Aldrich (Shnelldorf, Germany). High-performance liquid chromatography (HPLC) organic solvents were of analytical grade: methanol (MeOH), absolute ethanol (EtOH), chloroform and hexane were from Carlo Erba (Val-de-Reuil, France), methyl-tert-butyl-ether (MTBE) was from Fisher Scientific (Loughborough, UK), petroleum ether (PE) was from VWR Prolabo (Fontenay-sous-Bois, France). Ultrapure water was obtained from a purified water system Arium^® 611UV from Sartorius (Göttingen, Germany) with a resistivity of 18.2 MΩ*cm. Sodium chloride (NaCl, purity >99%) and potassium hydroxide (KOH, purity >99%) were obtained from VWR Prolabo (Fontenay-sous-Bois, France), and activated magnesium silicate Florisil^® at 100–200 mesh (Sigma-Aldrich, Shnelldorf, Germany). Liquid Nitrogen was from Air Liquide (Nancy, France). The synthesis of 5-(4-carboxyphenyl)-10,15,20-triphenyl porphyrin (P1COOH) has been described elsewhere [25 (link)].

Homogenates of the brain (10%; weight/volume) were prepared by blending the sections of this organ (of known weight) with an ice-cold potassium phosphate buffer (50 mM, pH = 7.4; made up with the use of dipotassium hydrogen phosphate, potassium dihydrogen phosphate [POCh], and distilled water) and butyl-hydroxytoluene (Sigma-Aldrich GmbH, Steinheim, Germany) in a glass/teflon homogenizer (Schütt homogen^plus, Schutt Labortechnik GmbH, Göttingen, Germany). Homogenates prepared for the assay of SOD, GPx, and GR were subjected to centrifugation (MPW-350R centrifugator, Medical Instruments, Warsaw, Poland) at 20000× g for 30 min (4 °C), whereas other measurements were performed in the aliquots of the homogenates centrifuged for 20 min at 700× g (4 °C) [49 (link)]. The received aliquots of the brain tissue were stored frozen (−70 °C) until the assay.

Chitosan, medium molecular weight, deacetylation value of 75–85%, and viscosity of 200–800 cP, was purchased from Aldrich Chemical Co.
Bovine serum albumin, acrylamide, N,N′-methylenebisacrylamide, trichloroacetic acid (TCA), FeSO₄, sulfuric acid, cumene hydroperoxide (CHP), butylhydroxytoluene, methanol, xylenol orange, di-nitrophenylhydrazine (DNPH), guanidine, ethanol, ethyl acetate, hydrochloric acid, Coomassie® Brilliant Blue R-250, thioglycolic acid, NBD-Cl, and o-phthalaldehyde (OPA) were purchased from Sigma-Aldrich (St. Louis, Missouri, USA).
Thiobarbituric acid was purchased (TBA) from Fluka (Sigma-Aldrich, Toluca, MX); sodium chloride, EDTA disodium salt, N,N′,N′- tetramethylethylenediamine (TEMED), Tris (base), urea, β-mercaptoethanol, glycine, acetic acid glacial, sodium phosphate monobasic, sodium phosphate dibasic, and copper sulfate pentahydrate were purchased from J.T. Baker (Pennsylvania, USA); sodium carbonate and lactic acid were purchased from Fermont (Monterrey, MX); sodium dodecyl sulfate (SDS) and bromophenol blue were obtained from Hycel (Mexico, MX); and plate count agar was purchased from Bioxon, Becton and Dickinson (Mexico, MX). All reagents used were of analytical grade.

Folin-Ciocalteu reagent, sodium carbonate, bovine serum albumin (BSA), acetate buffer, sodium dodecyl sulphate (SDS), triethanolamine, ferric chloride, sodium acetate trihydrate, 2,4, 6-tripyridyl-s-triazine (TPTZ), 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, β-carotene, linoleic acid, Tween 20, sodium phosphate, sodium chloride, potassium sodium tartrate tetrahydrate, 3,5-dinitrosalicylic acid, sodium hydroxide, α-amylase, starch, α-glucosidase, potassium phosphate monobasic, 4-nitrophenyl α-D-glucopyranoside, and butyl hydroxytoluene (BHT) were acquired from Sigma-Aldrich, Milan (Italy). Standards as tannic acid, gallic acid, 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), and acarbose were purchased from Sigma-Aldrich, Milan (Italy). Solvents as n-hexane, ethyl acetate, acetonitrile, methanol, hydrochloric acid, chloroform, and glacial acetic acid were purchased from Carlo Erba (Milan, Italy). HPLC grade solvents, as methanol and Trifluoroacetic acid (TFA), were acquired from Romil (Cambridge, UK). All spectrophotometric measurements were done on a UV/Vis spectrophotometer (SPECTROstar^Nano, BMG Labtech) and each reaction was performed in triplicate.

Kraft lignin (KL) was purchased from Sigma Aldrich (St. Louis, MO, USA) and used after purification by standard procedures including alkali–acid treatment and continuous washing with deionized water. Dimethyl isosorbide (DMI), ethanol, 2,2-diphenyl-1-picrylhydrazyl (DPPH), L-ascorbic acid AA, 3-O-ethylascorbate 1, ferulic acid 2, palmitic acid 3, 6-O-Palmitoyl-L-ascorbic acid 6, diisopropyl azodicarboxylate (DIAD), triphenylphosphine (TPP), butylhydroxytoluene (BHT), para-4-nitrophenyl laurate (para-NPL), chitosan (non-animal derived, average Mw 50 kDa, CH), concanavalin A (Con A, from Canavalia ensiformis, Jack bean type VI), and Lipase B from Candida antarctica were purchased from Sigma Aldrich and used without further purification. Analytical grade tetrahydrofuran THF and 2-methyl tetrahydrofuran (2-MeTHF) were refluxed over Na–benzophenone under nitrogen, followed by distillation and storage over 4 Å molecular sieves. Deuterated and HPLC purity-grade solvents were purchased from VWR. 1H and 13C NMR spectra were obtained in MeOD₃ using a Bruker 400 MHZ spectrometer (Bruker, Billerica, MA, USA).