Activation of the poly(ADP-ribose) polymerase after oxidative damage is implicated in different responses of the cells, for example, cell recovery after sublethal damage or cell death after lethal damage. However, the extent and mechanism of involvement of the enzyme in these two processes appear to be different. Inhibitors of this polymerase, such as benzamides, which do not completely inhibit PARP have been shown to protect the cells from killing by massive oxidant damage, could neither reduce the cellular recovery after mild oxidant damage nor completely inhibit DNA repair in vitro. We report here that 1,5-dihydroxyisoquinoline, which was earlier shown to be a strong inhibitor of this polymerase in vitro, is also its potent inhibitor in vivo. Using sensitive techniques for measuring low levels of cellular poly(ADP-ribose) polymer, we show that this inhibitor can completely abolish oxidant-induced activation of the polymerase in C3H10T1/2 cells. We show that only a minor fraction of the poly(ADP-ribose) polymerase activity is sufficient in cellular recovery after sublethal oxidant damage. We also demonstrate that cells are unable to recover from oxidant damage in the complete absence of polymerase activity.
We have studied the effect of H2O2 and O2- produced by xanthine and xanthine oxidase on NAD catabolism, poly(ADP-ribose) synthesis, and production of DNA single-strand breaks in C3H10T1/2 cells. The results show a correlation between the induction of DNA single-strand breaks, the decrease of NAD pool, and the accumulation of polymer. New techniques, based on affinity chromatography and reversed-phase high pressure liquid chromatography, have allowed an accurate determination of polymer contents and showed a 20-fold stimulation of polymer biosynthesis induced by active oxygen species. Inhibition experiments performed with 3-aminobenzamide have shown that the decrease in NAD levels after exposure of cells to active oxygen species was caused by stimulation of poly(ADP-ribosyl)ation and of another cellular process.
Studies were conducted to evaluate the effectiveness of industrial peanut dry roasting parameters in Salmonella reduction using a Salmonella surrogate, Enterococcus faecium, which is slightly more heat tolerant than Salmonella. Runner-type peanuts were inoculated with E. faecium and roasted in a laboratory scale roaster simulator in which temperature, airflow, airflow direction and bed depth were highly controlled, allowing for conditions that duplicate industrial dry roasting. Temperature data were collected at the top, middle and bottom of the roasting bed in addition to internal peanut temperature via thermocouples in the bed of peanuts and embedded in a peanut. Regardless of roast conditions, peanuts in the middle of the roasting bed received the least amount of heat and hence, represent the worst case scenario for microbial reduction. E. faecium reductions, reported as the logarithm of colony forming units/g (log CFU/g), followed a linear trend with increasing roasting time when peanuts were roasted at 149, 163, and 177 C, with > 5-log CFU/g reductions occurring at the middle of the peanut bed after 21, 15 and 11 min, respectively, at a bed depth of 75 mm and an air flow of 1.3 m/s. Increased air flow increased E. faecium reduction. At 16 min roast time and a 75 mm bed depth, reduction at the middle of the bed was ≤ 3-log CFU/g at 1 m/s and > 5-log CFU/g at 1.3 m/s. When all other roast parameters were held constant, decreasing bed depth also increased reduction of E. faecium in the middle of the bed. Comparing various samples roasted at 149, 163 and 177 C over a range of times, roast color (Hunter L-value) was positively correlated (R2 = 0.73) with the log reduction of E. faecium. Most peanuts with an L-value darker than 53, a common threshold for light roast had ≥ 5-log CFU/g reductions; however, further study is required, including roasting peanuts from different origins and maturity, to fully understand the implications of roast color development and microbial reduction. This work provides valuable practical information for manufacturers of roasted peanuts when validating Salmonella reductions under a particular set of roasting parameters.
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