Multi-wavelength analytical ultracentrifugation of human serum albumin complexed with porphyrin


  • The Analytical Ultracentrifugation (AUC) required an enhanced mode of separation based on distinct optical properties.
  • The Cölfen detector was constrained by its limited sensitivity in the UV spectrum.
  • The Optima AUC's method of capturing wavelengths sequentially, rather than simultaneously, posed complexities in multi-wavelength data analysis.
  • The inherently poor solubility of protoporphyrins in water led them to form non-reactive aggregates.
  • Difficulties arose in quantifying the molar extinction coefficients and in distinguishing the proportion of Human Serum Albumin (HSA) complexed in aqueous solutions.


  • Integrated multi-wavelength detection capabilities into the AUC.
  • Leveraged the Optima AUC instrument, which exhibited enhanced sensitivity in the low UV range.
  • Adapted the UltraScan method to accommodate and decipher data derived from Optima AUC's multi-wavelength process.
  • Perfected the sample preparation process to improve solubility and ensure primary bonding between porphyrins and apo-HSA.
  • Employed MWL-AUC combined with UV/visible spectral analysis to isolate extinction signals, providing a clearer perspective on protein-porphyrin interactions.


  • Multi-wavelength SV experiments can effectively quantify distinct components in mixtures with similar hydrodynamic properties based on unique absorbance spectra.
  • These data help determine extinction coefficients for species hard to obtain in pure form.
  • UltraScan's new AUC analysis tools enhance the analysis of multi-wavelength data from the Beckman Coulter Optima AUC instrument.
  • Multi-wavelength AUC is ideal for characterizing complex mixtures with unique absorbance spectra components.
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