A simple chemiluminescence assay for detecting oxidative stress in ischemic limb injury.

This study was designed to evaluate a simple, sensitive, nonstimulated chemiluminescence assay to measure the oxidative stress production in the whole blood of rabbits after an ischemic insult. By using an ultra-sensitive chemiluminescence (CL) analyzer and lucigenin amplification, the assay system can be performed without leukocyte isolation and stimulant administration. The blood CL levels of healthy rabbits were 122 +/- 18 counts/10 s. After 8 h of ischemia, the CL levels of whole blood immediately after reperfusion, 1 h, 2 h, 24 h, 48 h, 7 days, and 14 days of reperfusion were 409 +/- 78 counts/10 s, 283 +/- 55 counts/10 s, 256 +/- 43 counts/10 s, 228 +/- 33 counts/10 s, 185 +/- 32 counts/10 s, 160 +/- 16 counts/10 s, and 119 +/- 15 counts/10 s. The differences were statistically significant between the control and samples obtained up to 24 h after reperfusion. The corresponding creatine phosphate kinase (CPK) levels of the rabbits in this study were 938 +/- 43 U/l for the control blood sample and 5921 +/- 498 U/l, 6948 +/- 427 U/l, 6860 +/- 1115 U/l, 5763 +/- 516 U/l, 1545 +/- 291 U/l, 478 +/- 60 U/l, and 458 +/- 48 U/l for the experimental blood samples. The CPK levels of the blood samples before 24 h and after 7 days of reperfusion were significantly different to that of the control blood sample. The changes in the CL and CPK levels were quite similar before 48 h reperfusion. This assay has proved to be valuable in the quantitative measurement of ischemic insults of skeletal muscles.

Mutagenicity of dipyridyls and their methylated derivatives in Salmonella typhimurium/rat liver microsome system.

The Salmonella/microsome assay with strains TA97, TA98, TA100, TA1535, TA1537 and TA1538 was used to examine the potential mutagenicity of 5 dipyridyls, 1 tripyridyl, 3 dipyridinium diiodides and 2 pyridinium monoiodides. The widely used herbicide paraquat (1,1-dimethyl-4,4'-dipyridinium diiodide) and its precursor 4,4'-dipyridyl gave weak and marginal mutagenic activity to Salmonella typhimurium TA1535 and TA1538 in the presence of S9-mix. Significantly high mutagenicity was obtained with 2,2'-, 3,3'-, 2,3'-, and 2,4'-dipyridyls, 2,2',2"-tripyridyl, and 5 pyridinium salts under the same conditions. The positive mutagenic response of 2,2',2"-tripyridyl suggests that higher polymers of pyridine contaminating paraquat preparations might be mutagenic. The dose-response curves of 1,1-dimethyl-3,3'-dipyridinium diiodide and 1,1'-dimethyl-2,2'-dipyridinium diiodide revealed an exponential relationship between the number of induced revertants and the compound concentrations. The results suggested that the mechanism of mutation induced by these two compounds might be attributed to the chain reactions of their free-radicals with molecular oxygen.