A reversed phase HPLC assay for the simultaneous quantitation of non-ionic and ionic surfactants in bioprocess intermediates.

This report describes a rapid and accurate reversed phase HPLC method for the simultaneous quantitation of multiple surfactants in various bioprocess solution matrices including cell lysates. Separation and quantitation of a mixture of the cationic detergent domiphen bromide from the non-ionic detergent Triton X-100 in crude cell mixtures can be achieved within 15 min using a TSK-gel C18-NPR reversed phase column and an aqueous mobile phase gradient of acetonitrile:water with the reagent PIC-B8 as ion-pairing modifier. The linear dynamic range for quantitation of domiphen bromide (DB) and Triton in this assay extends from 20 to 2000 microM. Linear regression analyses from the standard curve determinations showed an R2 of > or = 0.990. The assay does not show any interferences from proteins or other cellular contaminants such as nucleic acids. The assay has been used to evaluate clearance of these compounds throughout the purification process of an adenovirus-based vaccine candidate, as well as to determine the effects of process changes on detergent clearance.

Structure of the capsular polysaccharide of pneumococcal serotype 11A reveals a novel acetylglycerol that is the structural basis for 11A subtypes.

We have undertaken a structural assessment of Streptococcus pneumoniae 11A polysaccharide as well as two clinical isolates related to 11A. The clinical isolates were labeled 11Aalpha and 11Abeta. The result of our experiments is a revision to the old structure for S. pneumoniae 11A polysaccharide. The new structure differs from the old structure in both the primary connectivities and acetylation pattern. We also show that 11A contains an acetylglycerol-PO4 moiety, a substitution that is heretofore unknown in the bacterial polysaccharide literature. The two clinical isolates were also structurally characterized. 11Aalpha was determined to be identical to 11A. 11Abeta is a new serotype, which differs from 11A in the absence of the acetylation of the glycerol-PO4 moiety and a different acetylation pattern of the saccharides. Thus, we propose that the acetylglycerol is the structural basis for 11Aalpha and 11Abeta subtypes.