Abstract
Neutral Red (NR), a eurhodin dye, is often used for staining living cells, but we demonstrated for the first time that NR can also serve as a CO2 sensor, because of NR’s unique optical properties, which change with dissolved carbon dioxide (dCO2) concentrations. In the present study, optical sensitivity of NR was quantified as a function of changes in absorption and emission spectra to dCO2 in a pH 7.3 buffer medium at eight dCO2 concentrations. NR exhibited a response time of two minutes for equilibration in pure N2 to 100% CO2 with an ~200% percent change (%∆) in emission intensity and >400%∆ in absorbance, with full reversibility. Important to its application to biological systems, NR exhibited zero sensitivity to dissolved oxygen, which has routinely caused interference for CO2 measurements. NR exhibited pH sensitive emission and excitation energies with dual excitation wavelengths at 455 nm and 540 nm, and a single emission at 640 nm. The CO2 sensing properties of NR were benchmarked by a comparison to pyranine (8-hydroxypyrene-1, 3,6-trisulfonic acid trisodium salt) (HPTS). Future studies will evaluate the feasibility of NR as an intracellular in vivo pCO2 sensor in aquatic organisms critically impacted by increasing global CO2 levels.
Highlights
The development of optical carbon dioxide (CO2 ) sensors is an expanding area of analytical science because of the demand for inexpensive and efficient devices for use in engineering and medicine [1]
While Neutral Red (NR) has been known in the past primarily as a biological cellular staining agent, results from the present study were the first to demonstrate NR’s ability to exhibit pH-induced emission and absorbance changes when exposed to different concentrations of dissolved CO2
An added advantage of the NR system is that it has an emission wavelength that is significantly red-shifted compared to the most established pH-induced, fluorescent CO2 sensors in use (e.g., HPTS)
Summary
The development of optical carbon dioxide (CO2 ) sensors is an expanding area of analytical science because of the demand for inexpensive and efficient devices for use in engineering and medicine [1]. CO2 sensors are essential for the detection of environmental changes related to global temperature increases such as ocean acidification and for use in monitoring dissolved CO2 levels in stagnant rivers and lakes [1,2,3,4]. While there are several CO2 sensitive optical materials available most of them have limitations, which include high cost, sensitivity to electrical interference, sensitivity to pH changes and acidic or basic gases, and often have slow response and recovery times [1]. Indirect sensing of CO2 involves the measurement of changes in CO2 concentrations and NR, a cellular stain, responds to changes in dissolved (dCO2 ) under different pHs and external CO2 concentrations. Measuring changes in CO2 concentration based on the changes in pH of the medium has been a method used primarily by the Severinghaus electrode [1,7]
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