Hemolysis of Human Red Blood Cells by Riboflavin-Cu(II) System: Enhancement by Azide

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I. Ali^1*, N. Sakhnini¹, and I. Naseem²

¹Faculty of Pharmacy, Applied Science University, Amman, Jordan; E-mail: iyad74@yahoo.com

²Department of Biochemistry, Faculty of Life Sciences, Aligarh Muslim University, Aligarh 202002, India

^* To whom correspondence should be addressed.

Received August 24, 2004; Revision received November 4, 2004

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Photoactivated riboflavin in the presence of Cu(II) generates reactive oxygen species (ROS) which can hemolyze human red blood cells (RBC). In the present work we examined the effect of sodium azide (NaN₃) on RBC in the presence of riboflavin and Cu(II). The addition of NaN₃ to the riboflavin-Cu(II) system enhanced K⁺ loss and hemolysis. The extent of K⁺ loss and hemolysis were time and concentration dependent. Bathocuproine, a Cu(I)-sequestering agent, inhibited the hemolysis completely. Among various free radical scavengers used to identify the major ROS involved in the reaction, thiourea was found to be the most effective scavenger. Thiourea caused almost 85% inhibition of hemolysis suggesting that ^*OH is the major ROS involved in the reaction. Using spectral studies and other observations, we propose that when NaN₃ is added to the riboflavin-Cu(II) system, it inhibits the photodegradation of riboflavin resulting in increased ^*OH generation. Also, the possibility of azide radical formation and its involvement in the reaction could not be ruled out.

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KEY WORDS: riboflavin, copper, sodium azide, reactive oxygen species, RBC, hemolysis

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