HPLC-based method for the analysis of whole-cell sialic acids

The sialic acids are a group of nine-carbon monosaccharides that are present on the surface of human cells. These acidic sugars take the terminal position on the oligomers that coat the outer surface of the cell membrane and are involved in cellular adhesion, motility and intracellular communication. It has been observed that the overall level of cellular sialylation can vary and is altered in the presence of disease including cancer. There is evidence to show that increased levels of sialylation, or hypersialylation, is linked to an increase in the metastasis of tumors. This evidence, along with work demonstrating the effect of lowering cellular sialylation, demonstrates the potential that inhibiting the transfer of sialic acids to the oligosaccharides that coat the cell surface may one day aid in the treatment of cancer.

This work demonstrates a HPLC-based method for the direct measurement of N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) in samples of cultured human cells. This method avoids the use of a derivative, and the cells may be treated with sialyltranferase inhibitors or other compounds before measurement. Thus, this method may be used to measure the efficacy of sialyltransferase inhibitors on different cell lines, regarding overall sialylation. HeLa and RT-4 cells were cultured in the presence or absence of known sialyltranferase inhibitors, and their Neu5Ac And Neu5Gc levels measured using a modified previously reported method by our group. The LOD and LOQ was 0.00551 mM and 0.01669 mM respectively for Neu5Ac and 0.00726 mM and 0.02199 mM respectively for Neu5Gc. Inter/intra-day precision and accuracy results for the concentration range tested were both within 15%.