Fast and Accurate Exhaled Breath Ammonia Measurement

The overall goal of the following experiment is to measure systemic ammonia levels through exhaled breath. This is achieved by first ensuring that the subject has not consumed any food or drink within the past hour. To maintain an ammonia steady state between the oral cavity and deep lung, next, the subject exhales at a desired flow, allowing for comparison of breath samples between subjects.

Then the breath from the phase three segment of the sample, which contains the ammonia. Most reflective of systemic values is chosen for analysis. Results are obtained that show ammonia in parts per million with carbon dioxide and pressure also recorded.

The main advantage of our breath collection system over current methods like uncontrolled breath collection is that our monitor and collection system produces fast sensitive and reproducible results. The implications of this technique extend towards cirrhosis and other disease and wellness states because ammonia is relevant to protein, nutrition, and hepatic encephalopathy. To begin turn on the external power supply to the ammonia optical sensor platform, the air pump, a custom built control electronics unit or CEU, the breath sampler, the laser diode controller, and a laptop.

Check that both the exhaust and cooling fans of the ammonia sensor are operating. Next, on the desktop of the breath sampler set the inlet tubing and mouthpiece temperature to 55 degrees Celsius by clicking on an icon named change T.The temperature can then be changed by clicking the up or down arrow, followed by clicking update temp. A keyboard attached to the sampler can also be used to type in the desired temperature.

Click exit to return to the desktop. To open the software on the laptop that controls the ammonia sensor, open a folder named NH three Breath Sensor program. On the desktop.

Open the main lab view software folder and select main sequence VI to access the desired interface. Then select Run in the top left corner of the screen. Create a new session on the sampler by opening the program.

Breath sampler found on the desktop of the sampler. Adjust the inlet and mouthpiece temperature before entering the breath sampling program while wearing gloves. To avoid contaminating the mouthpiece with ammonia, insert a new disposable mouthpiece into the inlet pipe.

For this protocol, ensure subjects have consumed no food for about 12 hours upon arriving to the lab and that they have refrained from exercise the morning prior to testing. Also verify that they have not introduced anything into their mouth or brushed their teeth within one hour of data collection seat the subject in front of the ammonia sensor. Instruct the subject to hold the inlet pipe and instruct them not to touch the mouthpiece.

To avoid ammonia contamination, click start on the sampler interface. Then have the subject exhale a single breath for at least 10 seconds into the mouthpiece. When the breath sample is completed, click stop.

To take measurements of the breath sample, select the phase three portion of the sample by dragging the vertical lines on the sampler interface to begin when carbon dioxide plateaus and stop right before the pressure drop of the breath, save the data to the flash drive by selecting store on the sampler touchscreen interface. Once the breath data have been stored, select start to initiate a new breath sample to see the effects of mouth rinse and pH on breath ammonia sample three breaths taken at least five minutes apart to establish the baseline ammonia level. Next, instruct the subject to use 30 milliliters of water to rinse their mouth thoroughly for 60 seconds.

Then within 60 seconds, collect a breath sample and continue to collect samples at least five minutes apart for 15 minutes, have the subject use sodium bicarbonate in water to rinse their mouth for 60 seconds and take another 15 minute series of breath samples. Then after using an acidic solution to rinse for 60 seconds, take a third, 15 minute series of samples from the subject to determine the effects of inlet and transport tubing temperatures on breath ammonia. Set the inlet temperature to 40 degrees Celsius before taking three breath samples over the course of 15 minutes.

Then increase the temperatures of the inlet and transport tubing to 55 degrees Celsius. After allowing the system to reach steady state for at least five minutes, sample three breaths into the heated inlet spaced five minutes apart. Shown here is a typical breath sample that integrates the phase three portion of the breath data.

The green and red vertical lines set the spin in which the line is to be analyzed. In contrast, this panel demonstrates the effect an acidic rinse has on breath ammonia. The conversion of NH three to NH four drastically lowers the amount of detectable ammonia.

The levels then return to baseline as the effects of the rinse wears off. For accurate measurements, the subject must provide enough flow of alveolar air into the sampler. This panel demonstrates how decreased flow, which doesn't allow as much alveolar air to be sampled, results in the reduction of alveolar ammonia in the sample when both phases one and two are integrated into the sample.

As in this panel, the analysis significantly lowers the amount of ammonia and is an inaccurate reflection of systemic ammonia. This technique can be done in approximately one minute for immediate results, enabling real-time data analysis. If done properly, It is important to always record CO2 and pressure to ensure uniform data quality.