Nwose EU, Bwititi PT, Chalada MJ (2018) Influence of Anticoagulants on Determination of H2O2 Levels in Blood: Comparison of Citrate and EDTA. Int J Pathol Clin Res 4:083.


© 2018 Nwose EU, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

RESEARCH ARTICLE | OPEN ACCESS DOI: 10.23937/2469-5807/1510083

Influence of Anticoagulants on Determination of H2O2 Levels in Blood: Comparison of Citrate and EDTA

Nwose EU1,2*, Bwititi PT3 and Chalada MJ2

1School of Community Health, Charles Sturt University, Australia

2School of Clinical and Psychological Sciences, Charles Darwin University, Australia

3School of Biomedical Sciences, Charles Sturt University, Australia



H2O2 is supposed to be part of the laboratory test panel for assessment of oxidative stress. This molecule contributes to oxidative stress by reacting with Fe2+ ions to produce hydroxyl radicals. Anticoagulants, including citrate and EDTA chelate Fe2+, possibly have implications in oxidative stress studies especially when H2O2 is measured.


This brief commentary is on the measurability of H2O2 in citrated and EDTA blood samples.


Laboratory evaluation was performed using blood samples of 30 sheep collected in EDTA and citrate tubes. Samples were centrifuged, and plasma separated. H2O2 was determined using the Biotech H2O2 560 assay protocol on plasma. The ratio of EDTA to iron was also estimated to help interpret levels of H2O2 in EDTA blood.


There was measurable H2O2 reaction in citrate samples, but not in the EDTA blood. Based on brand sterile hematology tubes containing 1.8 mg EDTA per mL blood; approximately 4:1 ratio of EDTA to iron is estimated. Presuming this calculated ratio is a reflection of the true iron levels in the sheep's blood, the test results support the suggestion that ratio of EDTA to iron is greater than 1:1.


Evidence is hereby presented to advance that EDTA anticoagulant is unsuitable for laboratory testing of H2O2 in plasma. Further studies may be needed to ascertain the effects of different anticoagulants on H2O2 in whole blood and haemolysate to see how these can be compared to plasma. This is relevant to establish a standard protocol for integration of H2O2 test for assessment of full oxidative stress panel in clinical practice.