losartan-potassium and 2-5-dihydroxybenzoic-acid

2,5-Dihydroxybenzoic acid (DHB) has been demonstrated to be a more suitable matrix than 3,5-dimethoxy-4-hydroxycinnamic acid (sinapinic acid, SA) to obtain reliable molecular mass values of intact glycoproteins because it prevents sugar fragmentation. Lack of spot homogeneity during the crystallization step was prevented by drying the sample-matrix mixture under vacuum conditions. Nevertheless, this sample-matrix preparation procedure requires a specific experimental setup and may be time-consuming. In this work, we investigated the effectiveness of different ionic liquid matrices (ILMs) with SA and DHB on the ionization of a set of intact glycoproteins with several degrees of glycosylation. The obtained results demonstrate that some of the tested ILMs allow detection of the studied intact glycoproteins. Furthermore, the selected optimum conditions solve the reproducibility issue of using the DHB as a solid matrix without the vacuum drying method and, surprisingly, avoid sugar fragmentation when both SA and DHB were used as ILMs.

Topics: alpha-Fetoproteins; Animals; Cattle; Coumaric Acids; Erythropoietin; Gentisates; Glycoproteins; Humans; Ionic Liquids; Orosomucoid; Recombinant Proteins; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Transferrin

Oligosaccharides derivatized with 4-aminobenzoic acid 2-(diethylamino) ethyl ester (ABDEAE) can be analyzed by ESI (Yoshino, K.; et al. Anal. Chem. 1995, 67, 4028-4031) and MALDI (Takao, T.; et al. Rapid Commun. Mass Spectrom. 1996, 10, 637-640) mass spectrometry. In this study, oligosaccharides derived from the enzymatic cleavage of the sugar chains of glycoproteins ribonuclease B, erythropoietin, and transferrin were subjected to ABDEAE derivatization, prior to analysis on a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MALDI-TOF MS) for high-resolution mass measurement and a postsource decay (PSD) experiment. In the mass measurement of ABDEAE derivatives, quasi-molecular ion species have been observed in monoisotopic resolution using 2,5-dihydroxybenzoic acid as the matrix from spots that contain 50-200 fmol of sample; in the PSD analyses from the spots contained 500 fmol-1 pmol of sample, the predominant backbone ion series which covers the entire mass range for all the derivatives, the internal ion series which reflect the branched trimannosyl core structure of N-glycans, and the low m/z fingerprint ion of ABDEAE were consecutively observed, permitting structure elucidation of the oligosaccharides. Given the effectiveness of this derivatization in terms of its high sensitivity and resolution with respect to MALDI-TOF MS, current methodology is clearly applicable to the sensitive detection and accurate structural analysis of N-glycans.

Topics: Carbohydrate Sequence; Erythropoietin; Gentisates; Hydroxybenzoates; Molecular Sequence Data; Oligosaccharides; Procaine; Reproducibility of Results; Ribonucleases; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Transferrin