6. After a line of best fit is produced, this relationship is then used to calculate NO from the area under curve of the sample.

2.5. Analysis of Data The AUC typically is measured because it provides the total NO release/detected rather than a time course for NO release. The presence of certain biological matrices within the purge vessel will likely affect the "detectability" of the released NO by the NOA. In principle, this should only serve to broaden the signal detected so long as all the NO is ultimately released by the cleavage reagent from the subsequent products formed (this means that although signal peak height may be influenced the AUC should remain unchanged). This is particularly important with regard to the measurement of NO in samples, where there is a high concentration of heme groups available to rebind the released NO.

An example that we have studied in great detail in the tri-iodide cleavage reagent is NO linked to hemoglobin. At low hemoglobin concentrations, relative to NO (high NO-hemoglobin ratio), the hemoglobin in the reaction chamber simply broadens the NO signal (signal height is significantly reduced but the area under curve remains unaffected: see Fig. 2.3A).

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