Study summary · research use only
Some pitfalls in the measurement of blood glutathione
Plain-language summary
Paraphrased from the published abstract below — not a verdict on whether anything works.
This methodological study in healthy human subjects reported pitfalls in measuring whole blood and plasma glutathione. Blood was collected by brachial vein puncture using a tourniquet and a 21-gauge needle, and total glutathione (reduced and oxidized) was analysed by a spectrophotometric recycling method using the glutathione reductase/NADPH couple. Oxidized glutathione was measured after treating aliquots with 2-vinylpyridine or N-ethylmaleimide. The abstract reports that with separated plasma, 2-vinylpyridine yielded oxidized glutathione values twice as high as N-ethylmaleimide, with a greater discrepancy in whole blood not resolved by deproteinization with 5-sulphosalicylic acid. Using N-ethylmaleimide, oxidized glutathione was less than 1% of total in whole blood, and lysed erythrocytes contributed on average 25% to plasma glutathione.
Abstract
1. We report some pitfalls in the measurement of whole blood and plasma glutathione in man. 2. Using a simple tourniquet to the forearm and a 21-gauge needle, blood samples were collected by brachial vein puncture from healthy subjects. Whole blood and plasma were analysed for total glutathione, including the reduced and oxidized forms, by a spectrophotometric recycling method involving the glutathione reductase/NADPH couple. 3. The concentration of oxidized glutathione was determined after treatment of aliquots with either 2-vinylpyridine or N-ethylmaleimide to trap reduced glutathione. Reduced glutathione in the native samples could then be obtained by subtraction. 4. When the reagents were added to separated plasma, 2-vinylpyridine yielded values for oxidized glutathione that were twice as high as with N-ethylmaleimide. In whole blood studies the discrepancy was even greater, and the problem was not resolved by deproteinization of samples with 5-sulphosalicylic acid. Using N-ethylmaleimide, levels of oxidized glutathione were less than 1% of total glutathione in whole blood. 5. Despite attempts to minimize haemolysis, lysed erythrocytes contributed on average 25% to the 'plasma' glutathione concentration.
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