Time-of-Flight Mass Spectrometry Of Underivatized Single-Stranded DNA Oligomers by Matrix-Assisted Laser Desorption

Abstract

Matrix-assisted laser desorption with concomitant ionization (MALDI) in conjunction with time-of-flight mass spectrometry (TOF-MS) has been used to analyze underivatized random-base single-stranded DNA (ssDNA) oligomers ranging from 10 to 89 nucleotides in length by embedding them in a solid matrix of 3-hydroxypicolinic acid. At 355-nm desorption wavelength, mass spectra of positive and negative ions measured by reflecting and linear time-of-flight mass spectrometers are compared. Results from the linear system show the ionization yield is approximately equal for each polarity. Metastable ion decay is significant for the larger ssDNA oligomer ions, which results in a decrease in signal intensity and the broadening of mass peaks. In order to obtain an acceptable signal-to-noise ratio on a reflecting TOF system, a higher laser irradiance is needed, which consequently causes further degradation of mass resolution. With the apparent advantages of better sensitivity and mass resolution, it is concluded that a linear TOF system is better suited for the mass spectrometric analysis of ssDNA oligomers larger than about a 25-mer. The current system permits one-base resolution up to about a 40-mer. Mass accuracy for a 20-mer or smaller is within +/- 0.05%. Comparison of mass spectra from 5-ns and 35-ps pulse widths at the same energy density shows no significant differences. Mechanisms for oligonucleotide ion production in these experiments are discussed.

Negative electrospray ionization mass spectrometry of synthetic and chemically modified oligonucleotides

“…The first studies using fast atom bombardment (FAB) or liquid secondary ion mass spectrometry (LSIMS) were limited to small oligonucleotides (4; < 5000 Da). With the development of new methods of ionization such as electrospray ionization (ESI; 5) or matrix-assisted laser desorption ionization (MALDI; [6][7][8], which can produce intact gaseous ions from highly polar molecules, it has become possible to analyze higher molecular weight oligonuleotides (> 10 000 Da) with a precision comparable with that obtained for proteins (9,10).…”

Gas-phase conformations of deprotonated trinucleotides (dGTT−, dTGT−, and dTTG−): the question of zwitterion formation

“…New matrices have shown some success in decreasing the amount of fragmentation [5][6][7] as have adding ammonium salts to the DNA sample [8 -11] and the use of IR-MALDI ioniziation sources [12,13], which have yielded mass spectra of polynucleotides containing more than 1000 bases. Other studies, however, have focused on smaller oligonucleotides to determine the exact mechanism of the fragmentation.…”

Fragmentation pathway studies of oligonucleotides in matrix-assisted laser desorption/ionization mass spectrometry by charge tagging and H/D exchange

“…A large number of studies have been undertaken to better understand the fragmentation process [16 -34]. Generally, the severity of fragmentation is matrix-dependent, and a matrix of choice is 3-hydroxypicolinic acid (3-HPA) [35,36] (or the two component matrix-3HPA/picolinic acid [5]) because it results in a lesser amount of fragmentation and enables the analysis of larger oligonucleotides. The popular matrices for peptides and proteins, such as sinapinic acid (SA) instrument facility tend to produce significant fragmentation when used for oligonucleotide analysis.…”