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Fig. 36. EPR spectra at 77 K from liver of mice injected with bacterial lipopolysaccharide (LPS) 4 h earlier. (A) (1,2) ferrous-citrate complex and diethyldithiocarbamate (DETC); (B) (3) the ferrous-citrate complex or (4,5) the ferrous-citrate complex and thiosulfate. Spectra (2,5) were recorded in livers of mice which had been given NNLA at 2.5 h after the injection of LPS. The animals were euthanized in 15 min after injecting the ferrous-citrate complex, DETC or thiosulfate. (From Ref. [99].)

Fig. 36. EPR spectra at 77 K from liver of mice injected with bacterial lipopolysaccharide (LPS) 4 h earlier. (A) (1,2) ferrous-citrate complex and diethyldithiocarbamate (DETC); (B) (3) the ferrous-citrate complex or (4,5) the ferrous-citrate complex and thiosulfate. Spectra (2,5) were recorded in livers of mice which had been given NNLA at 2.5 h after the injection of LPS. The animals were euthanized in 15 min after injecting the ferrous-citrate complex, DETC or thiosulfate. (From Ref. [99].)

after the injection of thiosulfate ligands together with ferrous citrate (Fig. 36, curve 4). The concentration of DNIC in the tissue changed with time. The highest yields were ca 6 ^M DNIC/kg of wet tissue, and were observed 10-15 min after the injection of thiosulfate. At later times, the yields diminished. We attribute the decrease of DNIC to either oxidation of the thiosulfate ligands, or the removal of the ligands from the tissue. The inhibition by LNNA showed unequivocally that the DNIC complexes were endogenously formed from endogenous NO produced enzymatically by NOS from L-arginine. We propose that a certain quantity of DNIC is formed endogenously in any organism capable of generating NO. The basal concentration of DNIC in tissues seems quite low, and remains below the detection limit of EPR (ca 10 nM tissue concentration) as a result of destruction by superoxide radicals and peroxynitrite. However, the DNIC levels may be raised above the detection threshold by supplying thiosulfate to obtain very stable DNIC complexes, or by artificially boosting the endogenous NO production, for example by the injection of LPS.

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