2nd IeCAB symposium June

2 Mohamed M. Ibrahim 1* and Sameera O. Bafeel 2 1 Science Department ( Biology section), Teacher's College, King Saud University, Riyadh, Saudi Arabia 2 Biology Department, College of Science, King Abdul Aziz University, Jeddah, Saudi Arabia E. mail.

3 What is an Oxidant and Oxidative Stress? Oxidant : Refers to all “Reactive oxygen/Nitrogen species” including O 2, H 2 O 2 and OH and NO. Oxidative stress : Is an imbalance between pro- oxidants and antioxidants in favor of the pro-oxidant leading to cell injury or death.

2nd IeCAB symposium June Environmental stresses such as heavy metals that disturb the normal cellular metabolism can upset the balance of production and quenching of reactive oxygen species (ROS) like O, HO and OH and NO Environmental stresses such as heavy metals that disturb the normal cellular metabolism can upset the balance of production and quenching of reactive oxygen species (ROS) like O 2, H 2 O 2 and OH and NO..

2nd IeCAB symposium June Inevitable biological process associated with general decline in physiological function. Increased probability of death with accumulation of diverse, adverse changes as an organism gets older.

6 - A rapid change of gene expression in plants in response to heavy metal stress is important for environmental adaptation. - mRNA Differential Display technique was used in this study to analyze alterations in gene expression in garden cress (Lepidium sativum) in response to different concentrations of lead.

7 - Some Molecular, physiological as well as the response of oxidative stress indices to Pb 2+ toxicity were investigated in an effort to identify the possible involvement of Pb 2+ in the generation of oxidative stress. Also we investigated the molecular and physiological mechanisms by which Lepidium sativum cope with Pb 2+ toxicity to overcome the drastic effect of ROS.

2nd IeCAB symposium June Estimation of lipid peroxides Determination of superoxide anion radical (O2_– ) production rate Determination of lead content Isoenzyme profile of some antioxidant enzymes Semi-quantitative RT-PCR

2nd IeCAB symposium June

Table 1: Effect of increasing concentration of Pb 2+ in the growth medium on Lead accumulation in shoot and root and total lipid peroxide of Lepidium sativum roots after 10 days treatment. Each value is the mean of three replicates (  SD). Values carrying asterisk are significantly different at P  Total lipid peroxide TBARS (nmol g -1 F.W.) Pb 2+ (µg g -1 D.W) Treatments Pb 2+ (ppm) RootShoot 80.24± C 91.56±   ±5.23 ٭ 0.90  0.17 ٭ 0.24  0.03 ٭ ± 6.23 ٭ 1.94  ٭ 0.38  0.08 ٭ ± 7.23 ٭ 3.18  0.91 ٭ 0.85  0.10 ٭ 600

Fig. (1): Changes in the rate of superoxide production rate in roots of control and stressed plants of Lepidium sativum subjected to various concentrations of Pb 2+ for 10 days. The control for these measurements are non treated roots. Each value represents the mean ±SE of five replicates.

2nd IeCAB symposium June Fig. 2. Effect of increasing concentration of Pb 2+ in the growth medium on SOD, CAT and APX activities in roots of Lepidium sativum after 10 days treatment. Values are mean±/S.D. Based on three replicates, bars carrying asterisk are significantly different at P  0.05.

2nd IeCAB symposium June Fig.3: Isoenzyme profile of superoxide dismutase (SOD), A; catalase(CAT), B; ascorbic peroxidase (APX), C in enzyme preparations from leaves of 25 days grown seedlings of Lepidum sativum. Seedlings were raised for 10 days under different concentrations of Pb 2+ in the growth medium. (C) Control; 100 = 100 ppm Pb 2+ ; 200 = 200 ppm Pb 2+ ; 400 = 400 ppm Pb 2+ ; 600 = 600 ppm Pb 2+. For details see text.

2nd IeCAB symposium June

Deleterious effects resulted from the oxidative stress caused by lower and moderate concentrations of lead can be prevented by the defense mechanism in plants via scavenging of the ROS.

16 The antioxidative system, is composed not only of the low-molecular components but also of enzymatic components such as SOD, CAT, and APX. Can serve in convalescing from deleterious effect of ROS.

2nd IeCAB symposium June

18 When Pb 2+ concentration exceeded 200 ppm, the emergence rate of roots of Lepidium sativum was inhibited, this effect became more marked when Pb 2+ concentration was above 400 ppm.

2nd IeCAB symposium June Both the plant’s physiological oxidative stress indicator (lipid peroxide) and the indicator of unbalanced metabolism of free radicals (superoxide anion radical (O2_–) production rate) varied with Pb 2+ accumulation especially in roots.

20 A close relationship was found between the activities of enzymes (SOD, CAT and APX) and Pb 2+ accumulation in roots. Also the differential gene expression of different isozymes was associated with varying Pb 2+ treatment.

2nd IeCAB symposium June Genetic, Physiological and Biochemical responses of some plants to interactive biotic and abiotic stress. Effect of heavy metals on ultra structure and antioxidant isozymes of some economically important plants. 21

2nd IeCAB symposium June Deanship of Scientific Research at King Abdul-Aziz University, for supporting this research project no. 428/524; Professor Nigel, Professor of Environmental Pollution for his suggestions; Mubarak City for Scientific Research for excellent technical assistance in RT-PCR.

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24 Thank You