To reduce MitoTEMPOL to the hydroxylamine, MitoTEMPOL-H, MitoTEMPOL (10 μL of a 0.5 M solution in ethanol) was mixed with 10 μL of 0.5 M sodium ascorbate (pH∼6), reacted for ∼ 1 min at room temperature, during which time the colour changed from orange to pale yellow [ 29].
Stock solutions (30 mg/ml in ethanol with 250 μg/ml BHT) of 7-KC, 7βOHC and 7αOHC (an optical isomer of 7βOHC) were freshly prepared as required.
Briefly, DPPH solution was prepared by diluting 1.2 ml of DPPH stock solution (0.2 M DPPH in ethanol) with 3 ml ethanol and 0.5 ml DMSO.
For tissue amino-acid analysis, liver and tumour samples were homogenised in 6.6 m M solution of acetic acid in ethanol with a homogeniser (Ingenieurbüro CAT, M Zipperer, Staufen im Breisgau, Germany) for 30 s on ice.
After washed twice in PBS, labelled cells were trypsinized, washed in PBS, and fixed in a 70 : 30 vol/vol 50 mM glycine water solution in ethanol (pH adjusted at 2.0 with HCl) for 30 min at 4°C.
The TLC plate was monitored by spraying with 10%H2SO44 solution in ethanol followed by heating.
Free bases were then converted into the hydrochloride salts by treatment of their solution in anhydrous ethanol with 1 M HCl in dioxane.
For photoluminescence experiments, we immersed the specimens in a common laser dye solution (i.e. Rhodamine 800 in ethanol with a weight ratio of 0.3%).
For the aluminum doping, the aluminum acetate solution prepared in ethanol was mixed with 0.1 M zinc acetate with different molar concentrations.
Sections were immersed for 4 min in Schiff's reagent and washed in water for 5 min. Sections were counterstained with Hematoxylin solution, dehydrated in ethanol and HemoD solution, mounted, and imaged.
