A Sensitive Determination of Ammonia and Nitrite in Exhaled Breath Condensate of Healthy Humans by Using Berthelot Reaction

Abstract

Background: Exhaled breath has recently been identified as an alternative matrix for the detection of drugs and chemical compounds. By keeping in mind that different chemical and biological compounds have already been determined in the exhaled air, the Exhaled Breath Condensate (EBC) was found to be a suitable sample for non-invasive assessment of the respiratory system. Thus, the aim of this work is determination of ammonia and nitrite in EBC samples. Methods: The determination of the analytes is based on the derivatization of ammonia using Berthelot method. The absorbance of colored compound was determined at 655 ± 3 nm. Nitrite ion was determined after its reduction to ammonia by using Zn/HCl reagent. Results: Good calibration curves were obtained for ammonia with linearity in the range of 10– 620 ng mL -1 , and correlation coefficients (r2) of more than 0.999. The Limit of Detection and Quantification (LOD, LOQ) and Lower Limit of Quantification (LLOQ) were found to be 1.5, 4.5 and 10 ng mL -1 , respectively. The recoveries of the method were calculated to be 94 to 102.5% and the intra- and interday precisions were lower than 1.4 and 3.4%, respectively. Conclusion: The results presented here demonstrate that the concentrations of ammonia and nitrite in samples of healthy humans can be determined with good accuracy, high sensitivity, and simple operation. The proposed EBC methodology possessed several advantages like simplicity and low cost, especially if more sophisticated techniques such as HPLC are not available. Keywords: Ammonia, Berthelot reaction, Exhaled Breath Condensate (EBC), nitrite, spectrophotometry, absorbance.