Measurement of Opaque Liquids with UV-Vis and ATR

Importance of Topic

The analysis of highly pigmented or turbid liquids such as paints, inks, food dyes, sunscreens, and emulsions presents significant challenges for conventional UV-Vis spectroscopy using 10 mm cuvettes. Extensive dilution often leads to increased solvent consumption, possible sample alteration, and dilution errors. Attenuated Total Reflection (ATR) fiber-optic probes, when combined with a UV-Vis spectrophotometer, enable direct measurement of these opaque samples without dilution, enhancing speed, accuracy, and sample integrity.

Study Objectives and Overview

This application note demonstrates how pairing an Agilent Cary 60 UV-Vis spectrophotometer with a Hellma Katana XP 6 three-bounce ATR fiber-optic probe allows for rapid, dilution-free analysis of opaque liquids and in situ monitoring of reaction kinetics. The study compares ATR-based absorbance measurements to theoretical 10 mm cuvette values, illustrating the practical benefits of the combined system.

Methodology and Instrumentation

• Instrumentation
  • Agilent Cary 60 UV-Vis spectrophotometer with focused beam optics and xenon flash lamp (10-year guarantee)
  • Hellma Katana XP 6, three-bounce ATR fiber-optic probe with sapphire crystal
  • Agilent Cary WinUV software for spectral acquisition and processing
• Methodology
  • Record baseline spectrum with a clean ATR crystal
  • Immerse probe tip directly into sample without dilution
  • Acquire absorbance spectra from 800 to 250 nm, 1.5 nm bandwidth, 0.4 s averaging, 2 nm interval, 150 nm/min scan speed
  • Between samples: clean probe tip, re-measure baseline, then analyze next sample

Main Results and Discussion

• Paint Samples: Yellow, red, and blue paints produced clear absorbance peaks (e.g., yellow paint peak at 440 nm with 0.15 Abs), equivalent to ~250 Abs in a 10 mm cuvette (≈200× dilution required).
• Food Coloring: Royal blue food dye showed 1.27 Abs at 650 nm (effective path ≈8.5 µm), corresponding to ~1500 Abs in a cuvette (≈1000× dilution needed).
• Sunscreen Formulations: SPF 15, 30, and 50 samples displayed decreasing absorbance with higher protection factors. Reflectance spectra derived from absorbance confirmed increased reflectivity in the UVA/UVB/UVC regions as SPF rose.
• Reaction Kinetics: In situ monitoring of Fmoc deprotection in peptide synthesis (280 nm) tracked reaction progress in a turbid slurry. Absorbance increased by 0.05 Abs (≈140 Abs in cuvette terms) over ~6 minutes, demonstrating real-time kinetic analysis without sample preparation issues.

Benefits and Practical Applications

• Eliminates extensive sample dilution, reducing solvent use and contamination risk
• Allows direct measurement of turbid and highly absorbing liquids
• Enables real-time monitoring of heterogeneous reaction kinetics
• Applicable to quality control, R&D, industrial analytics, and educational laboratories

Future Trends and Potential Uses

• Integration with automated sampling for high-throughput analysis
• Expansion to solid or semi-solid samples using specialized ATR probes
• Development of portable remote spectroscopic systems for on-site monitoring
• Advanced chemometric models tailored to ATR UV-Vis data interpretation

Conclusion

The combination of the Agilent Cary 60 UV-Vis spectrophotometer with a Hellma Katana XP 6 ATR fiber-optic probe offers a robust, user-friendly solution for direct, dilution-free analysis of opaque liquids and real-time reaction monitoring. This approach streamlines workflows, conserves materials, and delivers reliable quantitative data across diverse analytical applications.

References

• Agilent Technologies. Cary 60 UV-Vis Spectrophotometer Datasheet.
• Hellma UK Ltd. Katana XP ATR Probe Application Notes.