Advancing Integrating-Sphere-Assisted Resonance Synchronous Spectroscopy for Quantitative Absorbance Analysis in...

Area of Science:

• Analytical Chemistry
• Spectroscopy
• Biophysical Chemistry

Background:

• Conventional UV-vis spectrophotometry struggles with turbid solutions due to scattering.
• Integrating-Sphere-Assisted Resonance Synchronous (ISARS) spectroscopy offers an alternative for turbid samples.
• Existing ISARS methods had limitations in spectral range, dynamic range, and sample volume sensitivity.

Purpose of the Study:

• To enhance the capabilities of ISARS spectroscopy for broader applications.
• To overcome limitations of spectral range, linear dynamic range, and sample volume variations.
• To enable accurate absorbance quantification in optically complex media.

Main Methods:

• Expanded spectral range to 250-750 nm using neutral-density-filter (NDF) strategy.
• Extended linear dynamic range using short-path cuvettes and direct "first-strike" geometry.
• Developed polynomial models and a Python GUI for data analysis and conversion to double-beam values.

Main Results:

• Achieved a significantly expanded usable spectral range (250-750 nm).
• Enabled reliable quantification of protein absorbance in undiluted commercial milks (optical density > 150).
• Demonstrated ISARS's unique capability in solutions with extremely high scattering.

Conclusions:

• The enhanced ISARS method overcomes critical limitations of the original technique.
• ISARS is now a powerful and accessible platform for absorbance analysis in optically complex media.
• This advancement opens new possibilities for analyzing challenging biological and industrial samples.