The Detection Of Oxygen And pH Using The Luminescence Method

Theory of optical oxygen and pH sensing

Figure 1: The Jablonski diagram illustrates the electronic transitions and energy level relaxations of a molecule after absorbing light.

Optical sensing, particularly using fiber optic techniques, has emerged as a powerful tool for measuring critical chemical parameters like oxygen concentration (O2) and pH levels. This approach offers several advantages over traditional methods, making it increasingly attractive for diverse applications in industry, biomedicine, and environmental monitoring. Here, we delve into the theoretical principles and background behind optical O2 and pH sensing.

1. Principles of Optical O₂ Sensing:

Excitation: Light from an external source excites the luminescent molecule (fluorophore or phosphorescent molecule) to its excited singlet state (S₁) (Fig. 1). This is depicted by an upward arrow in the Jablonski diagram.

Relaxation Pathways: From S₁, the excited molecule can relax back to the ground state (S₀) in many ways:

• Fluorescence: In the absence of oxygen, the molecule can return to S₀ by emitting a photon (light) with a longer wavelength (lower energy) than the absorbed light. This emission process is shown as a downward wavy arrow on the diagram.
• Phosphorescence (O₂ - independent): In some molecules, a transition to a lower-lying triplet state (T₁) can occur through a process called intersystem crossing. Relaxation from T₁ to S₀ can also involve light emission (phosphorescence), but with a longer delay and lower intensity compared to fluorescence. This process is independent of oxygen concentration.
• Non-radiative Relaxation: The molecule might lose its excess energy through collisions with surrounding molecules or other non-radiative pathways, returning to S₀ without emitting light.

• Oxygen Quenching: When oxygen molecules are present, they can interact with the excited molecule in S₁ through collisions. This interaction transfers the excitation energy to the oxygen molecule, preventing the fluorophore from returning to S₀ via fluorescence. The oxygen molecule does not emit light itself, but the overall luminescence intensity (fluorescence) is reduced. This quenching effect of oxygen is represented by a dashed arrow in the Jablonski diagram. The degree of quenching depends on the oxygen concentration and the specific properties of the fluorophore.

2. Principles of Optical pH Sensing:

• Indicator Dyes: Here, a pH-sensitive dye is used. The dye's chemical structure and electronic configuration change depending on the surrounding pH.

  pH-Dependent Absorption: Depending on the pH, the dye absorbs light at specific wavelengths. This absorption process promotes the dye molecule to an excited electronic state.

  Emission and pH Dependence: The excited dye molecule can then relax back to the ground state by emitting light (fluorescence). The key aspect is that the emitted light's intensity or spectrum (wavelength distribution) varies depending on the pH-induced changes in the dye's structure. By analyzing these changes in emission properties, we can determine the pH of the solution.

  RESULT

  

  Figure 2: The fluorescence spectral of a hydrophilic photosensitizer (Rose Bengal) with different oxygen concentrations.

  Figure 2 illustrates the influence of oxygen concentration on the luminescence properties of a molecule called RB (Rose Bengal). All measurements used the same concentration of a photosensitizer molecule. Under excitation light of 523 nm, RB displayed fluorescence peaks at specific wavelengths. However, phosphorescence, another form of light emission from excited molecules, was absent under normal and high oxygen conditions. Remarkably, when oxygen concentration was significantly reduced, a distinct phosphorescence peak emerged. This finding aligns with previous research. The figure underscores the critical role of oxygen, as its concentration heavily impacts both fluorescence and phosphorescence emission in RB. This variation translates to a change in the overall light emission profile. Notably, the intensity of phosphorescence increases as the dissolved oxygen concentration decreases. Figure 2 reveals how oxygen levels can significantly influence how RB emits light.

  The figure presents the fluorescence spectra of fluorescein measured in aqueous solutions at different indicated pH values.

  

  Figure 3: The fluorescence spectra of fluorescein measured in aqueous solutions at different indicated pH values.

  Like a pH indicator, fluorescein's light absorption properties change based on the surrounding environment's acidity or alkalinity, as shown in Figure 3. At high pH (basic conditions), fluorescein primarily exists as a dianion (two negative charges) with a peak absorption around 490 nm. As the environment becomes more acidic (lower pH), the dianion concentration decreases, and other ionic forms of fluorescein become more prominent. These include anions (one negative charge) with absorption maxima around 474 nm and 453 nm, a neutral form absorbing maximally at 433 nm, and a cation (one positive charge) with a weak absorption spectrum. These findings align well with previous research, solidifying fluorescein's potential as a pH sensor due to its shifting absorption profile.

  Oxygen and pH monitoring system

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1. Oxygen (O₂) sensing setup:

Figure 4: Phase fluorometer system from Ocean Optics NeoFox and Bifurcated fiber-bundle.

NeoFox Phase Fluorometer:

• The NeoFox is a benchtop instrument specifically designed for measuring fluorescence lifetime, phase, and intensity. This advanced capability makes it ideal for applications where sensitivity to environmental factors and system stability are critical.
• Fluorescence Lifetime Measurement: This technique goes beyond simply measuring the intensity of emitted light. It analyzes the average time a molecule stays in its excited state before releasing light. This lifetime can be sensitive to the surrounding environment, including O₂ concentration.
• Phase Shift Detection: NeoFox can detect the phase shift between the excitation light and the emitted fluorescence. This phase shift can also be influenced by the presence of oxygen molecules, providing additional information for accurate O₂ sensing.
• Advantages: Compared to traditional intensity-based methods, NeoFox offers enhanced sensitivity and reduced susceptibility to interferences, making it a valuable tool for demanding applications.
• Applications: The NeoFox finds use in various fields, including environmental monitoring (measuring dissolved oxygen in water), bioprocess monitoring (analyzing oxygen levels in cell cultures), and material research (investigating oxygen diffusion in polymers).

BIFBORO-1000-2 Fiber-Bundle:

• This specialized fiber acts as a light guide with two distinct branches. One branch efficiently delivers the excitation light from the NeoFox to the tip of a fiber-optic sensor, while the other branch collects the emitted light and guides it back to the NeoFox for detection.
• Benefits: The bifurcated design ensures efficient light delivery and collection, minimizing signal loss within the setup. Additionally, the BIFBORO-1000-2 offers:
  • Versatility: It can be used with various fiber-optic O₂ sensors with different diameters (from exceedingly small to quite large), providing flexibility for diverse applications.
  • Durability: These fiber bundles are typically constructed from numerous single optical fibers, offering a robust and reliable light delivery system.

2. pH sensing setup:

• For measuring the emitted light, the Ocean SR spectrometer is a powerful tool. Its high spectral resolution allows for detailed analysis of emitted light, leading to highly accurate pH measurements. Additionally, Ocean SR enables real-time monitoring and holds potential for remote sensing, providing a comprehensive picture of ocean acidification dynamics in crucial areas. This innovative combination offers a promising approach for effective oceanographic research and environmental management.

  

  Figure 5: Ocean SR spectrometer.

  Conclusion

  Oxygen (O₂) and pH are fundamental for life and industry, requiring precise monitoring. This blog explored how optical-fiber sensors, leveraging luminescence, revolutionize O₂ and pH detection. These sensors offer non-invasive, real-time measurements and potential for miniaturization. We discussed fluorescence quenching and phosphorescence for O₂ sensing, and indicator dyes for pH sensing, all based on luminescence interactions. The concluding figure highlights the importance of calibration for accurate pH measurements. Overall, luminescence-based optical-fiber sensors provide a powerful and versatile tool for O₂ and pH detection, with applications in environmental monitoring, bioanalysis, and medical diagnostics.