Effective analysis of degree of polymerization of polysialic acids in mass spectrometry by combining novel sample preparation and dynamic instrument optimization methods

This work demonstrates a mass spectrometry technique to improve data reliability when analyzing degree of polymerization (DP) of high-mass polysialic acids (PSAs). Matrix-assisted laser-desorption/ionization (MALDI) time-of-flight mass spectrometry is the technique of choice for analyzing large molecules due to its wide mass working range; however, the observed DP of PSAs using such an instrument is unreliable owing to sensitivity bias towards low-mass ions. A multi-layer MALDI sample preparation protocol is demonstrated in the current study to improve PSA sensitivity, and a dynamic instrument optimization method (DIOM) is employed to minimize detector saturation over a wide mass range. The DP information obtained in the DIOM combines a series of mass spectral data obtained with individually optimized instrument parameters to minimize the problem of sensitivity bias in respective mass ranges. The resultant mass spectra facilitate unambiguous determination of DP in the high-mass range due to significantly improved spectral quality. The main instrument parameters involved in the optimization process include extraction delay in MALDI ion source as well as the cutoff mass of the ion detector. In comparison to conventional methods, the DIOM doubles the maximum DP that can be unambiguously identified by mass spectrometry.