Establishment Of New Methods To Evaluate Derivatives Of Reactive Oxygen Metabolites (DROM) And Biological Antioxidant Potential Tests (BAP) And Their Roles In Hyperthyroidism Disease Assessment

ObjectiveTo establish the simple, low-cost methods to measure derivatives of reactive oxygen metabolites (DROM) and biological antioxidant potential test (BAP) based on automated biochemical analyzer and to evaluate the analytical performance of these new methods. To evaluate the oxidative stress levels in the serum of patients with hyperthyroidism. To explore the role of DROM and BAP as biomarkers of oxidative stress in evaluating disease severity in hyperthyroidism.MethodsBuilt on measuring principles, at the very start, we established new methods to measure derivatives of reactive oxygen metabolites (DROM) and biological antioxidant potential test (BAP) using automatic biochemical analyzer and spectrophotometer and optimized measuring parameters. According to CLSI documents, we evaluated the analytical performance of these new methods including precision experiment, sensitivity experiment, linearity experiment and interference test.287 healthy individuals were recruited to measure DROM and BAP before we established the normal reference interval.73 hyperthyroidism patients and matched 100 healthy individuals were recruited in Beijing Hospital from November 2014 to February 2015. All subjects were evaluated with TSH, FT3, FT4 and markers of oxidative stress. The biological markers of oxidative stress include DROM, superoxide dismutase (SOD) and BAP test.Results1. The target low and high concentration TBHP solution test results show that. The total imprecision is< 6.67%; the fresh normal serum specimen test results show that the total imprecision is< 4.69%. The blank limit is 2.85 mmol/L TBHP; detection limit for fresh serum specimen is 11.43 mmol/L TBHP; quantitative limit for serum specimen is 31.77 mmol/L TBHP. TBHP concentration in the 31.77-400 mmol/L range is liner. The test results are linear (R2= 0.999). The normal reference interval was 107≥mmol/L TBHP. When hemoglobin≤ 175g/L, triglyceride≤27.18mmol/L and Vic≤60 μmol/L, the deviation was 47.3%,5.62% and 14.89% respectively.2. The target low and high concentration vitamin C solution test results show that, the total imprecision is< 5.87%; the low and high fresh serum specimen test results show that the total imprecision is< 14.14%. The blank limit is 0.068 mmol/L; detection limit for fresh serum specimen is 0.154 mmol/L; quantitative limit for serum specimen is 0.224 mmol/L. Vitamin C concentration in the 0-4 mmol/L range, the test results are linear (R2=0.9996). The normal reference interval was≥2.89 mmol/L VitC. When hemoglobin≤ 170g/L, triglyceride≤7.385mmol/L and bilirubin≤73.8 μmol/L, the deviation was -2.58%,-13.11% and 83.1% respectively.3. Serum levels of SOD are positively associated with TSH (r=0.264, P=0.024),meanwhile the levels of SOD are negatively associated with FT3 (r=-0.250, P=0.035).There is no significant correlation between the levels of DROM with TSH (r=-0.087,P=0.464),FT3 (r=0.201,P=0.092),FT4 (r=0.072,P=0.548).There is no significant correlation between the levels of BAP with TSH (r=0.193, P=0.101),FT3 (r=-0.101,P=0.403),FT4 (r=-0.095, P=0.425)ConclusionResults of DROM and BAP study show that the precision, the detection limit and the linear range of DROM and BAP using automatic biochemical analyzer can comply with the requirements of the clinical laboratory.Levels of DROM are increased in the serum of patients with hyperthyroidism compared with control. Serum levels of BAP and SOD are decreased. The level of SOD is positively correlated with the level of TSH and negatively correlated with the level of FT3.