International Research Journal of Environment Sciences________________________________ ISSN 2319–1414Vol. 3(1), 31-37, January (2014) Int. Res. J. Environment Sci. International Science Congress Association 31 Antioxidants and Some Biochemical Parameters in Workers Exposed to Petroleum Station Pollutants in Mosul City, IraqLuay A. Al-Helaly and Tareq Y. Ahmed Department of Chemistry, College of Science, Mosul University, IRAQAvailable online at: www.isca.in, www.isca.me Received 19th December 2013, revised 29th December 2013, accepted 18th January 2014 AbstractThe present study was conducted in Mosul city to evaluted the effects of different pollution sources on antioxidants and some biochemical parameters. The parameters included: Vitamin E, Vitamin A,-carotene, Vitamin C, folic acid, ceruloplasmin (Cp), total protein, albumin, calcium, total bilirubin, uric acid, creatinine, total iron binding capacity (TIBC), iron, sulfate, glutathione (GSH), malondialdehyde (MDA), cholesterol, glucose, selenium and transferrin saturation(%). The study involved (110) persons divided into two groups, one of which was subjected to petroleum station workers included(51 nonsmokers) worked at different positions and the other considered to be control group (n = 59 nonsmokers)(Outside city center living) with similar age and sex. The results showed a significant decrease in: vitamin E, vitamin C, creatinine, GSH and cholesterol, with no significant changes in: folic acid, calcium, iron, sulfate, glucose, selenium and transferrin saturation (%), vitamin A, -carotene, total bilirubin and TIBC. On the other hand, there was a significant increase in: Cp., total protein, albumin, total bilirubin, uric acid and MDA. In addition, the increase of workers period in petroleum station cause, a decrease in: vitamin E, vitamin C, folic acid, Cp., total protein,albumin, calcium, GSH, glucose and selenium, and an increase in: vitamin A, carotene, total bilirubin, uric acid,creatinine, TIBC, iron, sulfate, MDA, cholesterol, transferrin saturation(%)were observed.It was concluded that an increase exposure of petroleum pollutants to the workers led to an increase in their oxidative stress.Keywords: Petroleum station, pollutants, workers, antioxidantsIntroductionPollution can be defined as the introduction of contaminants into a natural environment, leading to instability, disorder, harm, discomfort and stress to the ecosystem, the physical systems or living organisms. Pollutants included chemicals such as petroleum hydrocarbons, heavy metals, pesticides etc. These contaminants can lead to system stress1,2. Previous studies have demonstrated increased oxidative stress in subjects exposed to environmental air pollution through the assessment of oxidative damage to DNA, lipids, or proteins3,4. For example, free radicals may induce peroxidation of arachidonic acid generating F2-isoprostanes. Urinary or plasma F2 isoprostanes have been found to be related to smoking and exposure to environmental tobacco smoke. Occupational exposure to combustion products, including petroleum have been associated with increased oxidative stress biomarkers in some, but not all, studies8,9. Exposure to petroleum hydrocarbons could be from gasoline fumes at the pumps, spilled oils; chemicals used at home and work e.g. pesticides. Some amount may leak from underground storage tanks and enter the ground waters. Some of the petroleum fractions are released into the soil contaminating the plants grown in such soil10,11. It was stated under normal physiological condition, animals maintained a balance between generation and neutralization of reactive oxygen species (ROS)12. However, when organisms are subjected to petroleum compounds, the rate of production of ROS in cells get increased along with hydrogen peroxide, hypochlorous acid (HClO) and free radicals including hydroxyl radical (OH) and superoxide anion (O.-). Oxidative stress have been implicated in a variety of pathological conditions such as diabetics mellitus, cancer, aging, liver damage, atherosclerosis etc.13. Recently, it was reported that airborne particles were associated with decreased in heart rate variability14. Another recent paper reported nitrogen dioxide (NO) and PM2.5(particulate matter with aerodynamic diameter less than 2.5 µm)15 were associated with defibrillator discharges due to ventricular arrhythmias in patients with implanted cardioverter defibrillators16. Antioxidants such as glutathione (GSH), uric acid, ascorbate and -tocopherol present in epithelial lining fluid (ELF) may protect the airways from oxidant injury induced by exposure to air pollutants17. The antioxidants act as sacrificial substrates scavenging oxidant pollutants from the airways and thereby preventing oxidation of macromolecules such as lipids, proteins and carbohydrates. The aim of the study was to evalute the effects of pollution for petroleum station workers in Mosul city on antioxidants and some biochemical parameters and to determine the effect of pollution period International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(1), 31-37, January (2014) Int. Res. J. Environment Sci. International Science Congress Association 32 Material and Methods The study included (110) persons represented in two groups. The first group was considered as control (59), while the second group was exposed petroleum pollutants (51) from workers in Mosul city, Iraq.Kits for determination of total protein no. (0303), albumin no. (0801), calcium no. (2403), total bilirubin no. (0401), iron no. (0502), total iron binding capacity no. (0512), glucose no. (0903), cholesterol no. (0603), were obtained from Syrbio kits, Syria.The other biochemical parameter (Vitamin A, -carotene, vitamin E, vitamin C, Folic acid, malondialdehyde, glutathion, selenium, uric acid and creatinine), were estimated using manual methods (table 1). For venipuncture, 10 ml sterile syringes equipped with (22G x 1.25) syringe needles were used and put of blood in dry and clean plain tubes. After coagulation, it was centrifuged at 4000 x g for 15 minute. Serum was transferred into plain tube equipped with tight–fitting caps by disposable tips, then stored at –20 \rC18TableMethods used to determination of biochemical parameters No. Parameters measured Method used 1 Vitamin A Needld-Pearson method 19 2 -carotene Needld-Pearson method 19 3 Vitamin E Emmerie-Engel reaction 20 4 Vitamin C 2,4-dinitrophenylhydrazine derivatization method21 5 Folic acid Microbiologyical assay 22 6 Glutathion Modified procedure utlilizing Ellman`s reagent23 7 Malondialdehyde Thiobarbituric acid method 24 8 Uric acid Phosphotungstic acid method 25 9 Total bilirubin Diazo method 26 10 Total protein Biuret methods 27 11 Albumin Bromocresol green mthod(dye binding method) 28 12 Ceruloplasmin p-Phenylenediamine oxidase method29 13 Glucose Glucose oxidase method 3 0 14 Cholesterol Cholesterol estrase methods 31 15 Creatinine Jaff method 32 16 Total Iron Binding Capacity Ramsay method33 17 Transferrin saturation Transferrin saturation(%) 34 = Serum Iron /TIBC X 100 18 Iron Bathophenanthroline method 34 19 Calcium Methylthymol blue method 35 20 Selenium Selenium-orthophenylenediamine complex36 Results and DiscussionEffects of petroleum station on the biochemical parameters of the workers: The results of different biochemical parameters in petroleum station workers and control group were listed in table (2). There is a significant decrease of biochemical parameters with the workers of petroleum station when compared with the control group in: vitamin E (P=0.008), vitamin C (P=0.0001), creatinine (P=0.0001), glutathione (P=0.0001) and cholesterol (P=0.0001). The decrease is not significant in: folic acid, calcium and iron. This finding agrees with that observation earlier37,38. In addition, the non significant decrease in the levels of sulfate, glucose, selenium and transferrin saturation (%) agree with the results obtained by other37 Table-2 The biochemical parameters of petroleum station workers and control Parameter Control group non-smokers (n=59) petroleum station non-smokers workers (n=51) mean SD mean SD Age(year) 35.7 10.9 34.62 7.86 Weight(kg) 76.31 14.5 77.46 10.09 Height(cm.) 169.0 10.57 169.6 8.5 B.M.I(k.g./m 2 ) 26.71 2.45 26.93 2.45 Vit.E(mg/dl) 1.0 0.21 0.73* 0.18 Vit.A(g/dl) 46.42 12.4 58.4 17.7 -carotene(g/dl) 80.49 21.52 162.1 35.22 Vit.C(mg/dl) 0.85 0.17 0.269* 0.02 Folic acid(ng/ml) 6.85 1.7 6.69 1.03 Cp.(mg/l) 151.1 39.4 159.7* 83.1 T.p.(gm/dl) 5.5 0.38 6.25* 0.63 Alb. (gm/dl) 4.36 0.12 4.91* 0.43 Calcium(mg/dl) 11.78 0.24 11.45 0.55 Total Bilir. (mg/dl) 0.41 0.27 0.74* 0.029 Uric acid(mg/dl) 5.05 1.58 5.68* 1.71 Creatinine(mg/dl) 0.82 0.05 0.69* 0.29 TIBC(g/dl) 186.8 32.16 219.0 39.69 Iron(mg/l) 1.6 0.19 0.661 0.289 Sulfate(mmol/l) 2.5 1.05 2.47 1.5 GSH(mol/l) 13.7 0.89 9.87* 0.86 MDA( mol/l) 6.5 2.97 15.06* 3.73 Cholesterol(mg/dl) 180.8 40.13 162.1* 7.94 Glucose(mg/dl) 68.1 35.8 55.21 11.88 Selenium(g/dl) 35.7 11.06 34.88 4.46 Tansferrin saturation% 36.65 13.5 30.18 10.21 *Different Significantly at P 0.05. Petrol chemical is a complex combination of hydrocarbons. About 95% of compounds in petrol vapors are aliphatic and alicylic compounds and less than 2% are aromatics. The volatile nature of petrol makes it readily available in the atmosphere at any time it is dispensed, especially at petrol filling stations11,39. A main environmental source of benzene exposure is vehicle exhaust emissions and evaporation losses during the handling, International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(1), 31-37, January (2014) Int. Res. J. Environment Sci. International Science Congress Association 33 distribution and storage of petrol40. The decrease of antioxidant refers to its utilization which prevents the cellular damage from free radicals, therefore a decrease in vitamin A, vitamin C, vitamin E, glutathion and selenium was observed. Vitamin C is oxidized first as ascorbyl radical and then to dehydroascobate and in the process scavenges free radicals and prevents radical-induced damage of lipoproteins and other macromolecules. Vitamin C was far more effective in inhibiting lipid peroxidation initiated by a peroxyl radical initator than other plasma components, such as urate, bilrubin, and alpha-tocopherol41, 42. Moreover, it has been observed that there was significantly increase in: ceruloplasmin (P=0.001), total protein (P=0.0001), albumin (P=0.0001), uric acid (P=0.014), Total Bilir. (P=0.029) and malondialdehyde (P=0.02), while there was no significant increase in: vitamin A, -carotene, total bilirubin and total iron binding capacity. This is in agreement with other studies which illustrated that benzene exposure has been associated with increases in the overall formation of MDA43-45. It was reported that uric acid might be a consistent and reliable biomarker of significant exposure to lead46. The pathophysiology by which lead exposure causes elevation in uric acid level is thought to be due to damage tubules which causes retention of uric acid47,48. Albumin, uric acid and bilirubin are major antioxidant components of plasma. In addition, albumin might play a major role of the total antioxidant capacity of plasma49. Albumin, one of the most important protein in human plasma, is able to bind to Cu++ tightly and with iron weakly. Copper bond to albumin is still effective in generating radicals species (Hydroxyl radicals) in the presence of hydrogen peroxide by Fenton reactions. Therefore increasing it to protect human from oxidants compounds formation50. Albumin and others, an index or marker of glomerular disease though raised in lead workers were probably a sufficiently reliable of lead nephropathy39,40. A raised total bilirubin was reported among diesel engine workers who were exposed to high-pressure resistant lubricants containing lead naphthanate51. They blamed the increase in total bilirubin to the poor compliance of the workers resulting in skin exposure, where lead absorption might have occurred through damaged skin, in addition to that it can be inhaled through the lungs. Many countries have reduced or eliminated the use of lead additives in motor gasoline due to the health consequences of lead exposure as well as to the introduction of catalytic converter, but in other countries leaded gasoline remains the norm, and still acts as a major source of human lead exposure. Although the exposure to lead has declined over the last several decades, lead toxicity remains a public health problem in most industrialized countries52,53. Effects of working period in petroleum station on the biochemical parameters: Depending on the period workers in petroleum station, the group was subdivided into three groups as listed in table 3. The results were indicated that vitamin E was decreased significantly with increasing of the period of petroleum station workers while for vitamin C and folic acid non significant difference was observed. Organic components carried on the particle surface play an important role in mediating the toxic effect. For example, polycyclic aromatic hydrocarbons, toluene and benzene can induce oxidative stress indirectly by changing the activities of antioxidant enzymes needed to composite for defense54, 55. -Carotene increased significantly with increasing of the period of petroleum station workers, but no significant difference was shown in vitamin A Ceruloplasmin was not affected with increasing the period of petroleum station workers and no significant difference was observed. The total protein wasn’t affected, but significant differences in groups was observed, similar results were published39. Albumin was decreased with increasing the period of petroleum station workers and there was a significant difference among the groups.Calcium was decreased significantly with increasing the period of petroleum station workers and a significant difference was observed among all groups. The increase of subjecting period leads that calcium is readily mobilized in bone deposits39,56causing its level to be decreased.The total bilirubin was increased with increasing the period of petroleum station workers and significant differences among the groups. This finding was in a good agreement with investigated research57, 58 .Uric acid was increased significantly with the increase of the period of petroleum station workers except last group and a significant difference was observed among all groups. This finding agrees with the other observations found earlier40, 59 . It was reported that uric acid is an endoantioxidant, thus raising urate level might in part be an antioxidant responses to protect against the prooxidant effect of lead40. Creatinine was increased significantly with increasing the period of petroleum station workers .The increase level of heavy metals from pollution as the period of pollution increased might increase creatinine60. It was found that chronic leads exposure decreases the glomerular filtration rate (GFR) with subsequent renal tubular fibrosis, renal atrophy and disturbances of the renal functions61,62 . Iron and sulfate were increased significantly with increasing the period of petroleum station workers, While TIBC was not affected. This result is in accords with other37. Glutathion was significantly decreased with the increase of the period of petroleum station workers. GSH is an important antioxidant produced by liver cells and the major soluble antioxidant in cell compartment. The significant decrease in GSH concentration might be attributed to its use in various protective roles against oxidants63. International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(1), 31-37, January (2014) Int. Res. J. Environment Sci. International Science Congress Association 34 Table-3 The biochemical parameters of petroleum station workers at different periods of working, and control using Duncan test Parameter Control group n=59 (0-5 year) n=22 (6-15 year) n=17 (16year and over) n=12 mean SD Mean SD mean SD mean SD Age(year) 35.7 a 10.9 31.3 a 10.17 37.66 a 7.58 45.0 a 0.81 Weight(kg) 76.31 a 14.5 74.2 a 11.64 74.33 a 9.33 86.25 a 2.98 Height(cm.) 169.0 a 10.57 167.8 a 6.61 171.0 a 7.53 170.1 a 13.54 B.M.I(k.g./m 2 ) 26.71 a 2.45 26.35 a 2.31 25.41 a 2.46 29.8 a 1.35 Vit.E(mg/dl) 1.0 a 0.21 0.82 b 0.5 0.68 c 0.27 0.66 d 0.41 Vit.A(g/dl) 46.42 a 12.4 52.71 a 18.1 63.13 a 19.6 62.18 a 16.28 -carotene(g/dl) 80.49 d 21.52 97.7c 45.4 179.7 b 66.1 261.3 a 79.6 Vit.C(mg/dl) 0.85 a 0.17 0.66 b 0.11 0.65 b 0.13 0.57 b 0.03 Folic acid(ng/ml) 6.85 a 1.7 6.8 a 1.64 5.72 a 0.88 5.93 a 89.12 Cp.(mg/l) 151.1 b 39.4 271.9 a 93.7 246.5 a 85.3 219.8 a 77.9 T.p.(gm/dl) 5.5 a 0.38 5.47 b 0.28 5.28 b 0.77 5.89 b 0.5 Alb. (gm/dl) 4.36 a 0.12 4.94 a 1.1 4.05 b 0.39 3.65 b 0.94 Calcium(mg/dl) 11.78 a 0.24 11.64 b 0.3 11.62 c 0.19 10.93 d 0.4 Total Bilir. (mg/dl) 0.41 a 0.27 0.34 b 0.1 0.54 a 0.17 0.46 b 0.29 Uric acid(mg/dl) 5.05 d 1.58 5.62 b 1.9 5.96 a 1.52 5.41 c 1.8 Creatinine(mg/dl) 0.82 d 0.05 0.95 c 0.11 1.21 b 0.07 1.41 a 0.08 TIBC(g/dl) 186.8 a 32.16 202.8 a 16.2 237.48 a 57.7 208.32 a 13.25 Iron(mg/l) 1.6 a 0.19 0.52 d 0.19 0.69 c 0.34 0.81 b 0.28 Sulfate(mmol/l) 2.5 a 1.05 2.423 a 0.14 2.89 a 0.43 2.69 b 0.83 GSH(mol/l) 13.7 a 0.89 8.46 b 0.72 7.0 c 0.69 6.31 d 0.56 MDA( mol/l) 6.5 b 2.97 11.34 b 3.20 15.93 a 4.88 18.41 a 5.01 Cholesterol(mg/dl) 170.8 d 40.13 177.6 c 37.0 182.2 b 10.4 190.26 a 44.04 Glucose(mg/dl) 68.1 a 15.8 58.83 a 14.08 54.71 a 10.0 65.54 a 12.32 Selenium(g/dl) 35.7 a 11.06 35.42 a 7.21 34.15 b 1.25 32.45 b 5.1 Tansferrin saturation% 36.65 b 13.5 25.64 d 9.21 29.05 c 12.2 38.88 a 13.5 Different letters horizontally a, b, c, d indicate that the means are different significantly at P 0.05. Malondialdehyde was increased with increasing the period of petroleum station workers and a significant difference was observed among each other. Malondialdehyde concentrations was increased significantly, indicating the amount of cellular damage. This increase could be significantly damage, from lipid peroxidation. Lipid peroxidation initiated by free radicals generated as a result of diesel intoxication is usually deleterious to cell membranes and is implicated in a number of pathological conditions64Cholesterol and transferrin saturation (%) were increased with increasing the period of petroleum station workers, and significant difference among each other. This finding was in a good agreement with others59. Moreover an increase in transferrin saturation(%) resulting from pollution was also observed . Aging might increase it with increasing mortality65. An increase in serum cholesterol emphasizes liver dysfunction, as cholesterol is synthesized in the liver and also excreted by the liver. The increase in the concentration of LDL-cholesterol is probably because of the increase concentration of cholesterol. LDL-cholesterol could be deposited on the walls of the artery leading to arthrosclerosis and other cardiovascular diseases66. High concentration of cholesterol in the serum could lead to diseases like arthrosclerosis and stroke67. Also high serum cholesterol has been reported to decrease concentration of circulating insulin and serum glutathione68. Glucose was not affected significantly with increasing the period of petroleum station workers and non significant difference among each other in all groups. Selenium was not affected significantly with increasing the period of petroleum station workers.ConclusionConclusion was drawn from the study that, an increase exposure of petroleum pollutants led to an increase in the oxidative stress for station workers. This is due to decrease in the antioxidant levels and an increase in the oxidants especially in the last period of work. This behaviors might suggest that workers subject to petroleum pollutants must supply with antioxidants dose especially with increasing period of exposure. International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(1), 31-37, January (2014) Int. Res. J. Environment Sci. International Science Congress Association 35 References 1.Prasad T.I., Suresh K.M. and Prasad D.A. Charecterization of Diffuse Chemical Pollution in Satna District of Vindhya Region, IndiaInt. Res. J. Environment Sci2(11), 46-60 (2013)2.Chatterjee P. and Das P. Changes in Lung Function status of Adult Female over last one decade: A Cross-Sectional study in Kolkata, IndiaInt. Res. J. Environment Sci., 2(7), 30-34 (2013)3.Singh V.K., Patel D.K., Jyoti Ram S., Mathur N. and Siddiqui M.K. Blood Levels of Polycyclic Aromatic Hydrocarbons in Children and their Association with Oxidative Stress Indices: An Indian PerspectiveClin. Biochem.41,152–161(2008)4.Svecova V., Rossner P., Jr., Dostal M., Topinka J., Solansky I. and Sram R.J. Urinary 8-oxodeoxyguanosine levels in children exposed to air pollutantsMutat. Res66237-43(2009)5.Cracowski J.L., Durand T. and Bessard G. Isoprostanes as a Biomarker of Lipid Peroxidation in Humans: Physiology, Pharmacology and Clinical ImplicationsTrends Pharmacol. Sci23, 360–366(2002)6.Dietrich M., Block, G., Hudes, M., Morrow, J. D., Norkus, E.P., Traber, M.G., Cross, C.E. and Packer, L. Antioxidant supplementation decreases lipid peroxidation biomarker F(2)-isoprostanes in plasma of smokers, Cancer Epidemiol. Biomarkers Prev.11, 7–13(2002)7.Dietrich M., Block G., Benowitz N.L., Morrow J.D., Hudes M., Jacob P., 3rd, Norkus, E.P. and Packer, L. Vitamin C supplementation decreases oxidative stress biomarker f2-isoprostanes in plasma of nonsmokers exposed to environmental tobacco smoke, Nutr. Cancer,45, 176–184(2003). 8.Rossner P., Jr., Svecova V., Milcova A., Lnenickova Z., Solansky I. and Sram R.J., Seasonal variability of oxidative stress markers in city bus drivers, Part II. Oxidative damage to lipids and proteins, Mutat. Res.,642, 21–27 (2008) 9.Marie C., Ravanat J.L., Badouard C., Marques M., Balducci F. and Maitre A., Urinary levels of oxidative DNA and RNA damage among workers exposed to polycyclic aromatic hydrocarbons in silicon production: Comparison with 1-hydroxypyrene, Environ. Mol. Mutagen.,50, 88–95 (2009) 10.Ujowundu C.O., Kalu F.N., Nwaoguikpe R.N., Kalu O.I., Ihejirika C.E., Nwosunjoku E.C., Okechukwu R.I. Biochemical and physical characterization of diesel oil contaminated soil in southeastern Nigeria, Res. J. Chem. Sci., 1(8), 57-62 (2011)11.Saini B., Verma R., Himanshu S.K. and Gupta S., Analysis of exhaust emissions from gasoline powered vehicles in a Sub-urban Indian town, Int. Res. J. Environment Sci.,2(1), 37-42(2013)12.Valko M.D., Leibfritz J., Moncol M.T. C. and Mazur M., Free radicals and antioxidants in normal physiological functions and human disease, Int. J. Biochem. Cell Biol.,39, 44-84 (2007)13.Nakagawa, H., Miyata, N. 2, 2, 6, 6-tetramethylpiperdim-1 oxyl process for evaluating oxidative stress on the cell membrane and mitochondria, Pub Med indexed for med.Line (2008)14.Gold, D.R., Litonjua, A., Schwartz, J., Verrier, M., Milstein, R., Larson, A., Lovett, E., Verrier, R. Ambient pollution and heart rate variability, Circulation. 101,1267–1273(2000). 15.Sadhana, Ch., Ashwani, K. and Anand Dev G. Air pollution and air quality index of Kodinar Gujrat, India, Int. Res. J. Environment Sci.,2(5), 62-67(2013). 16.Peters, A., Liu, E., Verrier, R.L., Schwartz, J., Gold, D.R., Mittleman, M., Baliff, J., Oh, A., Allen, G., Monahan, K. Air pollution and incidences of cardiac arrhythmia,Epidemiology,11,11–17 (2000) 17.Kelly, F.J., Blomberg, A. Frew A., Holgate ST, Sandström T. Antioxidant kinetics in lung lavage fluid following exposure of humans to nitrogen dioxide, Am. J. Respir. Crit. Care. Med.,154, 1700-1705 (1996)18.Liang L., D'Haese, P., Lamberts, L.V., DeBroe, M.E. Direct determination of iron in urine and serum using graphite furnace atomic absorption spectrometry. Analysis., 114,143-147(1989). 19.Neeld J.B., Person W.N., Macro- and micromethods for the determination of serum vitamin A using trifluoroacetic acid, J.Nutr., 79, 454-462 (1963) 20.Emmerie A. and Engel C., Serum total tocopherol estimation by colorimetric method, Nature, 142, 873 (1938)21.Roe J.H. and Kuther C.H., The determination of dehydroascorbic acid and ascorbic acid in plant tissues by the 2,4-dinitophenylhydrazine method, J.Biol.Chem., 147, 399 (1943) 22.Sedlak J. and Lindsay R.H., Estimation of total, protein-bound, and nonprotein sulfhydryl groups in tissue with Ellman's reagent, Anal. Biochem., 25(1), 192–205 (1968) 23.Lunec J., Free radicals: their involvement in disease processes, Review Articale, Ann. Clin. Biochem., 27, 173– 182 (1990)24.Varley H., Practical clinical biochemistry, 4th ed. The White Friars Press Ltd. UK, 82(1967)25.Toro, G., Ackermann P. G.Practical clinical chemistry, Little, Brown and Company(Inc.), USA, 497-506 (1975) International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(1), 31-37, January (2014) Int. Res. J. Environment Sci. International Science Congress Association 36 26.Kingsley G.R., The direct biuret method for the determinations of serum proteins as applied to photoelectric and visual colorimetry, J Clin. Lab. 27, 840-847 (1942)27.Doumas, B.T., Waston, W.A., Bigg, H.G. Albumin standards and the measurement of serum albumin with BCG, Clin Chim Acta, 31,87-96 (1971)28.Sunderman, F.W., Nomato, S. Measurement of human serum ceruloplasmin by its para phenylenediamine oxidase activity.Clin. Chem. 16(11), 903-910 (1970)29.Trinder P., Determination of glucose in blood using glucose oxidase with an alternative oxygen acceptor, Ann. Clin. Biochem. , 24-27 (1969)30.Richmond, W. Preparation and properties of a cholesterol oxidase from Nocardia Sp. And its application to the enzymatic assay of total cholesterol, Clin. Chem., 19(12), 1350-1356 (1973)31.Jaffe M., Uber, denNiederschlag, Wwlchen Pikrinsaure in normalem harn erzeugt and uber eine neue reaktion des creatinine, Hoppe- Seylers Z. Physiol. Chem., 10, 391-400 (1886) 32.William L., White J.M., Flashka C.M., The measurment of iron and total iron binding capacityby using calorimetric test (ferrozine), Clin. Chem.23, 237-239 (1977) 33.Burtis C.A. and Ashwood E.R., Tietz textbook of clinical chemistry, 3 rd ed. W.B. Saunders Company, USA, 490, 482, 1048, 500, 1241(1999)34.Rbertson W.G. and Marshall R.W., Calcium measurments in serum and plasma : total and ionized, Crit. Rev. Clin. Lab. Sci., 11, 271-304 (1979)35.Snell F.D., Photometric and fluorometric methods of analysis non metals, John Wiley and Sons, Newyork, 482-517 (1981)36.Sellappa S., Sadhanandhan B. and Francis A., Evaluation of genotoxicity in petrol station workers in south India using micronucleus assay, Industrial Health,48, 852–856 (2010)37.Anetor J.I. Serum uric acid and urinary protein: reliable bioindirectors of lead nephropathy in Nigerian lead workers, African J. Biomed. Res., 5(1-2), 19-24 (2002)38. Sies H., Stahl W. and Sundquist A.R., Antioxidant functions of vitamins. vitamins E and C, beta-carotene and other carotenoids, Ann. N.Y. Acad. Sci., 669, 7–20 (1992)39.Hong Y, Park E, Park M., Community level exposure to chemicals and oxidative stress in adult population,Toxicol Lett., 184(2), 139–144 (2009) 40.Al-Fhady N.H.A.M., Comparison of carbon monoxide, lead and cadmium effect upon blood of workers in contact, Ph. D. Thesis, College of Science, University of Mosul, Mosul, Iraq, 54 (2002)41.Chavez J., Cano C., Souki A., Bermudez V., Medina M., Ciszek A. and Amell A., Hernandez-Hernandez, R. and Valasco, M. Effect of cigarette smoking on the oxidant/antioxidant balance in healthy subjects, Am. J. Ther., 14,189–193 (2007)42.Chen Y., Effects of benzene on lipid peroxidation and the activity of relevant enzymes in humans, Chin J Prevent Med.,26, 336–338 (1992)43.Georgieva T., Michailova A. and Panev T., Possibilities to control the health risk of petrochemical workers, Int Arch Occup Environ Health75, 21–26 (2002) 44.Lai Ch. H., Saou-Hsing Liou, Jouni J.K. Jaakkola, Han-Bin Huang4, Ting-Yao Su, Paul T. Strickland. Exposure to polycyclic aromatic hydrocarbons associated with traffic exhaust: The increase of lipid peroxidation and reduction of antioxidant capacity, Aerosol and Air Quality Research, 12, 941–950 (2012)45.Gittleman et al, 1994. Cited by Anetor, J. I. Serum uric acid and urinary protein: reliable bioindirectors of lead nephropathy in nigerian lead workers, African J. Biomed. Res., 5(1-2), 19-24 (2002)46.Ball G.V. and Sorensen L.B., Pathogenesis of hyperuricaemia in Saturnine gout, N. EngI. J. Med.,280, 1199-203 (1969)47.Farkas W.R., Skai J. and Schneider M., Saturnine gout: Lead-induced formation of guanine crystals, Science, 199, 786-797 (1978)48.Aycicek A., Erel O. and Kocyigit A., Decreased total antioxidant capacity and increased oxidative stress in passive smoker infants and their mothers, Pediatr. Int.,47, 635–639 (2005)49.López-Tinoco C., Roca M., García-Valero, A., Murri M., Tinahones F.J, Segundo C., Bartha JL, Aguilar-Diosdado M. Oxidative stress and antioxidant status in patients with late-onset gestational diabetes mellitus, Acta. Diabetol., (2011)50.Clausen J. and Rastogi C., Heavy metal pollution among autoworkers lead, Br. J. Ind. Med., 34, 208-215 (1977)51.Seema T., Tiwari H.L. and Tripathi I.P., Review paper lead effects on health, Int. Res. J. Environment Sci., 2(8), 83-87 (2013)52.Seema T. and Tripathi I.P., Lead pollution – An Overview, Int. Res. J. Environment Sci.,1(5), 84-86 (2012)53.Dündarz M.R., Türkbay T., Cemal Akay S., Antioxidant enzymes and lipid peroxidation in adolescents with inhalant abuse, Turk. J. Pediat.,45, 43-45 (2003)54.Kelly F.J., Oxidative stress: its role in air pollution and adverse health effects, Occup. Enviro. Med., 60, 612-616 (2003) International Research Journal of Environment Sciences______________________________________________ ISSN 2319–1414 Vol. 3(1), 31-37, January (2014) Int. Res. J. Environment Sci. International Science Congress Association 37 55.Zielhuis R.L., Lead alloys and compounds. I: Encyclopedia of occupational health and safety, International Labour Office (Geneva), 2(2), 767-771 (1972)56.Birgitta A., Haeger J. and Aronsen, L. An assessment of the laboratory tests used to monitor the exposure of lead workers, Br. J. Ind. Med.,28, 52-58 (1971)57.Al-Nori M.K.J.N., Levels of some trace metals and related biochemicals in different laboratories, M.Sc. Thesis, College of Medicine, University of Mosul. Mosul, Iraq, 76 (2002)58.Kelle M., Diken H., Sermet A., Atmaca M., Mer C.T.,Effect of exercise on blood antioxidant status and erythrocyte lipid peroxidation: role of dietary supplementation of vitamin E., Tr. J. of Med. Sci.,29, 95-100 (1999)59.Welinder H., Skerfring S., Henriken O., Cadmium metabolism in man, Br. J. Indus. Med.,34, 221-228(1977)60.Sushil K.T., Madhur B.S., Toxicity of lead, Indian J. Physiol. Pharmacol., 37(1), 3-7 (1993)61.Rokho K.M., Andea R., David S., Weiss T., Cane W., Howird H., A longitudinal study of low level lead exposure and impairment of renal function, JAMA, 275(15), 117-1181 (1996)62.Pompella A., De Yeta V., Casini A.F., The changing faces of glutathione, a cellular protagonist, Biochem. Pharmacol., 66(8), 499-503 (2003)63.Srilaxmi P., Sareddy G.R., Kavi kishor P.B., Setty O.H, Babu P.P., Protective efficacy of natansnin, a dibenzoyl glycoside from Salvinia natans against CCl4 induced oxidative stress and cellular degeneration in rat liver, BMC Pharmacology, 10, 13 (2010)64.Mainous A.G., Gill J.M., Carek P.J., Elevated serum transferrin saturation and mortality, Ann. Fam. Med., 133, 138-142 (2004)65.Mayers G.D., Karmanna V.S., Kashyap M.L., Naicin therapy atherosclerosis, Current Opinion in Lipidology, 15(6), 659-665 (2004)66.Brown W.V., High density lipoprotein and transport of cholesterol and triglycerides in blood, J. Clin. Lipidol., , 7-19 (2007)67.Scholtz S.C., Pieters M., Oosthuizen W., Jerling J.C., Bosman M.J., Vorster H.H., The effect of red palm olein and refined palm olein on lipids and haemostatic factors in hyperfibrinogenaemic subjects, Throm. Res., 113, 13-25 (2004)