[RASMB] Absorbance Scan Question

[Peter Edward Prevelige Jr]

Hi All -

I'm doing an equilibrium run on a  ~3 KDa synthetic peptide lacking aromatics in 20% TFE. I  thought I might use multiple wavelengths to extend the concentration range that I could follow.

I thought I'd be clever and obtain the ratio of extinction coefficients by dividing the absorbance at one wavelength (236 nm) by the other  (240 nm) across the cell (rather than using the spectrum).

I calculated the ratio using only the points that were at the same radial distance and plotted them vs radial distance. It is pretty apparent that the ratio is changing across the cell.

My question is whether this is likely to be an optical artifact or whether it most likely represent heterogeneity in the sample. It is probably worth noting that the ratio from the spectrum is 1.43, intermediate between the extremes seen across the cell which to me suggests heterogeneity.

Thanks

Peter

Peter E. Prevelige Jr.
Professor
Dept. of Microbiology, BBRB 416/6
Univ. of Alabama @ Birmingham
845 19th St. South
Birmingham AL. 35294-2170
Phone 205 975-5327
FAX 205 975-5479
prevelig at uab.edu<mailto:prevelig at uab.edu>
http://www.microbio.uab.edu/faculty/prevelige/prevelige-p.htm

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