Fig. 1.
Shikonin reactivity with NO. The reaction of shikonin with NO was tested in a competition assay with oxyhemoglobin (Oxy-Hb) and monitored by EPR. The reaction consisted of 100 µM NOC-7 in 50 mM DETC in INTRAFAT emulsion, pH 7.0, supplemented with FeSO4, to which shikonin was added (see Materials and Methods). (A) a representative EPR spectrum of the (DETC)2-Fe2+-NO complex. (B) On the left, changes of signal intensity of the (DETC)2-Fe2+-NO complex by Oxy-Hb but not by shikonin. Oxy-Hb concentrations of 0, 5, 10, 15, and 20 µg/ml are shown by black circles, white circles, black triangles, white rhombus, and white squares, respectively. The assay in the presence of shikonin (0.5 mM) is shown by the dashed line on grey circles. On the right, the NO-reacting activity of Oxy-Hb is shown as a function of concentration. Results are representative of duplicate experiments.