Direct evaluation of glycated and glyco-oxidized globins by matrix-assisted laser desorption/ionization mass spectrometry

Abstract

Matrix-assisted laser desorption ionization (MALDI) mass spectrometry (MS) has been applied to achieve a qualitative and quantitative evaluation of glycated globins on a wide number of healthy and diabetic subjects. The method allowed us to establish that both alpha- and beta-globins are glycated and that, in addition to simply glycated products, other species are detected. Investigations by different sample treatments and by analysis of the glycated beta-globin fraction obtained by preparative chromatography indicated that these species correspond to glyco-oxidized globins. Consequently MALDI-MS can be validly employed to evaluate not only the glycation level, but also the degree of oxidative stress. The percentages of glycated and glycooxidized species were calculated from the related MALDI spectra by the measurement of the related peak areas, without any other treatment of data. A linear relationship between HbA1c values and the total percentage of glycated and glyco-oxidized globins has been found, and its slope (< 1) has been rationalized by the uncorrected evaluation of glycated globins content in the standard samples employed for HbA1c measurements.

Impaired Function of Antibodies to Pneumococcal Surface Protein A but Not to Capsular Polysaccharide in Mexican American Adults with Type 2 Diabetes Mellitus

“…The nonenzymatic glycation of immunoglobulins as a result of hyperglycemia could result in low antigen and receptor binding capacities of these antibodies. It is therefore likely that despite comparable titers, the functionality of the antibody is compromised in diabetics compared to nondiabetics (25,26). Contrary to our findings with PspA, we observed comparable titers of antibodies to all eight capsular polysaccharides tested in individuals with and individuals without diabetes.…”

The complexity of non-enzymatic glycation product sets of human globins

“…Apart from peaks due to protonated α and β globins, the MALDI spectra in our study showed peaks due to glycated α and β globins, confirming our previous findings [6,7] that both globins are glycated in vivo, in contrast to the usual thinking, as reported in the literature, on the definition of HbA 1 c and its measurement [2]. HbA 1 c measurements represent the integral of all glycated α and β globins, as proven by the relatively good linear plot.…”

“…For MALDI analysis, blood samples were treated as already described [6,7]. MALDI time of flight [8] spectra were obtained using a Voyager-DE PRO instrument (Applied Biosystems, Foster City, Calif., USA) that was operating in positive ion linear mode [6,7].…”

The role of mass spectrometry in the study of non-enzymatic protein glycation in diabetes

“…The application of mass spectrometry to evaluation of glycated globins has the merit of demonstrating that both globins are glycated to similar extents (Lapolla et al, 1997b). MALDI can reveal the presence of glycooxidated products, which cannot be detected in ESI conditions 1999).…”