Assaying poly(ADP-ribose) polymerase activity in plants by polarographic method

A new method has been developed to assay poly(ADP-ribose) polymerase (PARP) activity in plant tissues through determining the content of nicotinamide (NIC) produced by enzymatic reaction by linear sweeping polarographic metho d. The detection limit of NIC was 0.03 mu mol/L, the calibration graph was linear up to 5 mu mol/L (r = 0.999). The recoveries were approximately in t he range of 92% to 98% and the relative standard deviations were less than 6.6%, Moreover, NAD(+) and other interference existing in the mixture after enzymatic reaction had been removed by simple pretreatment, thus PARP assa ys were not interfered. A rapid, simple, sensitive and reliable nonisotopic method is reported to assay PARP activity in plant tissues. The results sh ow that the Km(NAD+) value of PARP in maize (Zea mays L.) seedlings is 59 a nd the optimum pH for PARP activity is 8.5. Moreover, physiological conditi ons affect PARP activity in plant tissues, which has not been reported prev iously. When tobacco (Nicotiana tobacum) suspension cells were stressed by NaCl at low concentrations (100, 200 mmol/ L), the PARP activity increased significantly; when the cells were stressed at high concentrations (400, 1 000 mmol/L). it decreased to or even below the control level. PARP activity in etiolated maize seedlings was higher than that in light-grown seedlings .