Using Tryptophan as an Indicator of pH in Bioaerosol Particle Fluorescence

HUNTER RICHARDS, Emily Hong, Zhenyu Ma, Allen E. Haddrell, Herek L. Clack, University of Michigan

     Abstract Number: 541
     Working Group: Bioaerosols

Abstract
Introduction
While the role of pH in respiratory virus inactivation has recently been discussed [1], measuring the change in pH of aerosols carried in exhaled breath is required to better define this relationship. To better understand the potential effect that an increase or decrease in pH has on the inactivation of certain respiratory viruses, it is first necessary to characterize the change in pH of a bioaerosol over time. Common fluorophores found in bioaerosol particles such as amino acids like tryptophan [2] can be exploited to interrogate pH change in exhaled bioaerosols that are relevant to the transmission of infectious disease. In previous experiments, it was determined that a higher pH correlates with a higher fluorescence intensity for tryptophan. Using tryptophan as a pH-dependent fluorophore, it is possible to model the change in pH of bioaerosols.

In this study, a wideband integrated bioaerosol sensor (WIBS) is used to analyze the fluorescence intensity of aerosolized solution within a closed chamber as the CO₂ concentration rises, using tryptophan as a fluorophore to track pH.

Experimental Procedure
A solution of 5% mass fraction of solute (MFS) NaCl and NaHCO₃ with tryptophan was aerosolized using a handheld nebulizer (Mesh Nebulizer Model YM-3R9) into a closed chamber at an aerosolization rate of approximately 1 mL per minute for 2 minutes. Using a CO₂ sensor (SAF Aranet4), the concentration of CO₂ was monitored within the chamber following aerosolization.

The WIBS instrument sampled from the chamber at a flow rate of 2.1 liters per minute for 10-20 second intervals over a 40 minute period. The WIBS instrument measured bioaerosol fluorescence intensity of the aerosols sampled from the chamber at predetermined time intervals by using a Xenon flash lamp to excite tryptophan at its known absorption wavelength of 280 nm, which produces fluorescent emission at 310-400 nm wavelength.

Results
The recorded fluorescence intensity spectra of sampled aerosols generated from bulk solutions of known pH are plotted in terms of relative abundance in Figure 1. The distinct spectra display substantial separation from the background noise (black spectra, Fig. 1), well beyond 5σ. At a higher bulk solution pH, aerosols generated from the NaCl + NaHCO₃ solution with added tryptophan exhibit a higher fluorescence intensity.

With the introduction to the chamber of the aerosolized NaCl + NaHCO₃ solution, the CO₂ concentration steadily rose.

Pairing the temporal evolution of CO₂ concentration in the chamber with the time-resolved fluorescence intensity of aerosols sampled by the WIBS instrument, tryptophan fluorescence decreased as the CO₂ concentration increased. As pH is reduced, fluorescence intensity shifts to the left and decreases.

The color-coded spectra at indicated time points show a consistent reduction in fluorescence intensity of the solution with time and the increase in CO₂ concentration, suggesting the evolution of CO₂ from the aerosols is correlated with decreasing aerosol pH over time.

Summary and Conclusions
As CO₂ concentration increases over time, a reduction in fluorescence intensity was observed for the solution containing tryptophan as a fluorophore. This indicates a decrease in bioaerosol pH as CO₂ concentrations increase. Using WIBS to monitor fluorescence intensity of bioaerosols mimicking exhaled breaths paired with CO₂ concentration monitoring can help better understand the relationship between pH and virus inactivation.

References
[1] Luo, Beiping, et al. “Expiratory Aerosol pH: The Overlooked Driver of Airborne Virus Inactivation.” Environmental Science & Technology (2023), 486-497.
[2] Pan, Yong-Le. “Detection and characterization of biological and other organic-carbon aerosol particles in atmosphere using fluorescence.” Journal of Quantitative Spectroscopy and Radiative Transfer, Volume 150 (2015), pages 12-35.