Polymerized glutaraldehyde has been used as a fixative agent for electron microscopy and to immobilize proteins for decades (1-5). The solution chemistry of glutaraldehyde has been reviewed (2). The UV Ratio is a parameter that has been used classically to describe the extent of polymerization of the material (3,4,7,8). An additional analytical method was developed to better characterize the polymeric mixture and correlate to this absorbance ratio. Development data from a low molecular weight SEC method resulted in nearly the same molecular weight values for all four of the HTGA samples examined (Mp = 377 to 384 Da) which had UV Ratios of from 2 to 20. This suggested that the UV Ratio (degree of polymerization) was not simply molecular weight dependent but was also dependent on the structures of the oligomers present. The existing literature seems to support this (1,3,6-9). Direct infusion mass spectra of the HTGA preparations were acquired. A simple weighted average molecular weight was calculated for the samples (based on the masses seen and their relative abundances in each sample) and was found to not correlate to the UV Ratio, as was seen for the SEC results. From the MS data the structures were assigned for the various types of oligomers present. Based on these structures and the published values for the mass extinction coefficients of the chromophores (1) in the oligomers a MS-based UV Ratio was calculated for the bulk HTGA.
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