MALDI Spectra Research Articles

Molecular and Structural Characterization of Polydisperse Polymers and Copolymers by Combining MALDI-TOF Mass Spectrometry with GPC Fractionation

Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) allows detection of large molecules such as those present in synthetic and natural macromolecules. Until recently, it was reported that MALDI-TOF measurements can provide correct molecular weight (MW) averages only for samples with a narrow MW distribution (M w /M n ≤ 1.20). We have now developed a methodology for polydisperse samples. We recorded the GPC trace of two polydisperse polymeric samples, namely, poly(butylene adipate)(PBA) and poly(butylene adipate-co-butylene succinate)(PBAS), collecting about 40-50 fractions per run. Selected fractions were analyzed by MALDI-TOF, and the average MW of each fraction was determined, allowing calibration of the GPC curves against absolute MW. The two calibrated GPC traces were then used to compute average MW and molecular weight distributions (MWD) of the unfractionated samples. End group analysis from MALDI-TOF spectra revealed that the PBA sample is composed of seven different types of chains. For the copolymer sample, PBAS, analysis of the MALDI spectra established a random sequence distribution of comonomer units. The succinate/adipate molar ratio calculated from the MALDI spectra is in good agreement with the molar ratio found by NMR.

Matrix-assisted laser desorption ionisation time-of-flight mass spectrometry of synthetic polyesters

We report the analysis of aliphatic and aromatic polyesters by matrix-assisted laser desorption-ionisation time-of-flight mass spectrometry (MALDI-TOF-MS) using 2,5-dihydroxybenzoic acid and t-3-indoleacrylic acid as matrices. Broad asymmetrical distributions of linear oligomer related peaks (protonated and cationised molecules) were observed with both matrices. Peaks corresponding to cyclic oligomers were observed for the aliphatic polyester. The oligomer molecular weight distribution derived from the MALDI spectra exhibits a certain variability with both laser fluence and matrix material. This effect probably originates from different solubilities of the various oligomers. In addition, for those polycondensates which are characterised by a very broad distribution with a tail in the high mass range, a discrimination against the higher mass oligomers is likely to exist; the agreement between the average molecular weights determined by MALDI and size exclusion chromatography is not as good as for poly(ethylene glycol) or poly(methyl methacrylate).

Characterization of polymers by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry: Molecular weight estimates in samples of varying polydispersity

AbstractMatrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) allows the detection of large molecules such as those present in synthetic and natural macromolecules. Synthetic polymers may show a wide range of molecular weight distributions (MWD), according to the synthetic method used in their preparation. We have studied several poly(methylmethacrylate) (PMMA), poly(styrene) (PS) and poly(ethyleneglycol) (PEG) samples with varying MWD, and also a number of condensation polymers such as Nylon‐6, poly(carbonate) and poly(ester). Measurements of MALDI‐TOF spectra in the linear mode were used in our work to estimate the molecular weight (MW) and MWD of the polymer samples. Our results show that the molecular weight estimates provided by MALDI‐TOF measurements agree with the values obtained by conventional techniques, such as gel‐permeation chromatography (GPC), only in the case of polymer samples with very narrow molecular weight distribution. However, when the polydispersity reaches values around 1.10 the difference between the MW measured by GPC and measured by MALDI may amount to up to about 20%. At higher dispersions, the MALDI spectra fail to yield reliable MW values.

Mass spectrometric characterization of a series of adenosylated peptides acting as bisubstrate analogs of protein kinases

We are currently developing strategies to synthesize bisubstrate analogs as potential inhibitors of serine and tyrosine protein kinases; several such analogs have been synthesized. The initial target proteins were the cAMP dependent protein kinase (cAPK) and the Ca +2/calmodulin dependent protein kinase (CaM kinase II). These bisubstrate analogs were based on either known peptide substrates such as kemptide, a seven amino acid peptide substrate of cAPK, or on inhibitory peptides such as a seventeen amino acid peptide encompassing the autoinhibitory domain of CaM kinase II. Peptides containing a single phosphoserine group were first synthesized and then adenosine 5′-monophosphate (AMP), denosine 5′-diphosphate (ADP), or adenosine 5′-triphosphate (ATP) was coupled through the serine phosphate with prior activation by 1,1-carbonyldiimidazole using either a solution or solid phase reaction scheme. In this current study, we report the characterization of the bisubstrate analogs by liquid secondary ionization mass spectrometry (LSIMS), matrix-assisted laser desorption mass spectometry (MALDI), and tandem mass spectometry (MS/MS). In the positive-ion mode, the LSIMS spectra of the bisubstrate analogs yielded a series of molecular ions containing mono-, di-, and trivalent cation adducts. Cation adducts were absent in the negative-ion mode where the dominant species were deprotonated molecular ions, [M − H] −, making this latter technique more useful for confirming product identity and assessing purity. Analysis of these compounds by MALDI in both the positive- and negative-ion modes yielded molecular ions which also contained metal ion adducts, although they were limited primarily to Fe +2 adducts. Unlike LSIMS, the MALDI spectra showed no evidence for the elimination of the phosphoadenosine or other structural moieties. When these compounds were subjected to high energy collision-induced dissociation (CID), the dominant fragmentation pathways under positive-ion MS/MS conditions resulted from cleavage of the phosphate linkages to the adenosine moiety with charge retention on the peptide, although a major peak fo 5′-deoxyadenosine was also seen at m/z 250. Charge retention in the negative-ion mode was most pronounced for ion fragments containing the highly acidic phosphate moieties and yielded phosphoadenosine related ions, for example, (AMP-H) −, (AMP-H-H 2O) −, (ADP-H), etc., as well as ions originating from the phosphate linker such as PO − 3, H 2PO − 4, HP 2O 6, H 3P 2O − 7, and H 2P 3O − 9. The largest phosphoadenosine ion in the negative-ion CID spectra for each bisubstrate analog, for example, m/z 426 (ADP-H) −, m/z 506 (ATP-H) −, or m/z 586 (AP 4H) −, indicated that the desired covalent modification had been formed between the phosphoserine and AP n moieties.