High-resolution MALDI-TOF MS study on analysis of low-molecular-weight products from photo-oxidation of poly(3-hexylthiophene).

High-resolution matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry (TOF MS) was used for the analysis of the low-molecular-weight products from the photo-oxidation of poly(3-hexylthiophene) (P3HT) in solution and thin film. Eight new peak series were observed in the low-mass range of the mass spectra of the products degraded in solution, and the formulas of the eight components were determined from the accurate mass. From SEC/MALDI-TOF MS, two components were identified as the degraded products, and the other six components were derived from the fragmentation of the degraded products during the MALDI process. A mechanism for the formation of these components was proposed on the basis of the results of MALDI-TOF MS. For the thin film degradation, a part of products in the solution degradation were observed, which supports that the oxidation of P3HT in solution and thin film proceeded in the same mechanism. This study shows that high-resolution MALDI-TOF MS is effective for the analysis of the low-molecular-weight products from P3HT photo-oxidation and expected to be feasible for the degradation analyses of other polymers. Copyright © 2015 John Wiley & Sons, Ltd.

Development of novel active acceptors possessing a positively charged structure for the transglycosylation reaction with Endo-M and their application to oligosaccharide analysis.

With Boc-Asn-GlcNAc as a basic structure, four permanently positively charged kinds of new acceptors (GP-Boc-Asn-GlcNAc, GT-Boc-Asn-GlcNAc, HMP-Boc-Asn-GlcNAc, MPDPZ-Boc-Asn-GlcNAc) and five kinds of similar structure acceptors (2-PA-Boc-Asn-GlcNAc, 3-PA-Boc-Asn-GlcNAc, 4-PA-Boc-Asn-GlcNAc, HP-Boc-Asn-GlcNAc, PDPZ-Boc-Asn-GlcNAc) were synthesized as acceptors for the resolution of oligosaccharides in glycopeptides. The synthesized acceptors enzymatically reacted with Disialo-Asn (donor) in the presence of Endo-M. The reaction yields of each transglycosylation product were not obvious, because we do not have all the authentic Disialo-Asn-Boc-acceptors. Therefore, we used the peak area of the transglycosylation product detected by mass spectrometry and evaluated the utility of each acceptor. Among the Boc-Asn-GlcNAc acceptors, the positively charged MPDPZ derivative peak area was the highest, MPDPZ-Boc-Asn-GlcNAc with a positively charged structure showed about a 2.2 times greater sensitivity of the transglycosylation product compared to the conventional fluorescence acceptor DBD-PZ-Boc-Asn-GlcNAc. As a result, the MPDPZ-Boc-Asn-GlcNAc acceptor was suitable for the transglycosylation reaction with Endo-M. The development of a qualitative determination method for the N-linked oligosaccharides in glycoproteins was attempted by combination of the transglycosylation reaction and semi-micro high-performance liquid chromatography/electrospray ionization quadrupole time-of-flight tandem mass spectrometry (HPLC/ESI-QTOF-MS/MS). The asparaginyl-oligosaccharides in glycoproteins, liberated by treatment with Pronase E, were separated, purified and labeled with positively charged MPDPZ. The resulting derivatives were separated by a semi-micro HPLC system. The eluted N-linked oligosaccharide derivatives were then introduced into a QTOF-MS instrument and sensitively detected in the ESI(+) mode. Various fragment ions based on the carbohydrate units appeared in the MS/MS spectra. Among the peaks, m/z 782.37 corresponding to MPDPZ-Boc-Asn-GlcNAc is the most important one for identifying the asparaginyl-oligosaccharides. Disialo-Asn-Boc-MPDPZ was easily identified by the selected-ion chromatogram at m/z 782.37 by MS/MS detection. Therefore, the identification of N-linked oligosaccharides in glycoproteins seems to be possible by the proposed semi-micro HPLC separations followed by the QTOF-MS/MS detection. Furthermore, several oligosaccharides in ovalbumin and ribonuclease B were successfully identified by the proposed procedure.

Mechanical tuning of molecular machines for nucleotide recognition at the air-water interface.

Molecular machines embedded in a Langmuir monolayer at the air-water interface can be operated by application of lateral pressure. As part of the challenge associated with versatile sensing of biologically important substances, we here demonstrate discrimination of nucleotides by applying a cholesterol-armed-triazacyclononane host molecule. This molecular machine can discriminate ribonucleotides based on a twofold to tenfold difference in binding constants under optimized conditions including accompanying ions in the subphase and lateral surface pressures of its Langmuir monolayer. The concept of mechanical tuning of the host structure for optimization of molecular recognition should become a novel methodology in bio-related nanotechnology as an alternative to traditional strategies based on increasingly complex and inconvenient molecular design strategies.

Mechanical tuning of molecular recognition to discriminate the single-methyl-group difference between thymine and uracil.

Construction of enzyme-like artificial cavities is a complex and challenging subject. Rather than synthesizing complicated host molecules, we have proposed mechanical adaptation of relatively simple hosts within dynamic media to determine the optimum conformation for molecular recognition. Here we have applied this concept to one of the most challenging biomolecular recognition problems, i.e., that of discriminating thymine from uracil. We synthesized the novel cholesterol-armed triazacyclononane as a host molecule and subjected it to structural tuning by compression of its Langmuir monolayers in the absence and in the presence of Li(+) cations in the subphase. Experimental results confirm that the monolayer of triazacyclononane host selectively recognizes uracil over adenine (ca. 7 times based on the binding constant) and thymine (ca. 64 times) under optimized conditions ([LiCl] = 10 mM at surface pressure of 35 mN m(-1)). The concept of mechanical tuning of a host structure for optimization of molecular recognition offers a novel methodology in host-guest chemistry as an alternative to the more traditional molecular design strategies.

Observation of HOPG by STM and contact AFM in various gas atmospheres under pressures up to 1.1 MPa.

Apparatus comprising a scanning tunneling microscopy (STM) and an atomic force microscopy (AFM) has been developed for use under supra-atmospheres. Observations of highly oriented pyrolytic graphite (HOPG) were carried out by STM and contact AFM operating in air and various gas atmospheres (hydrogen, helium, neon and argon) under pressures up to 1.1 MPa. Atomic resolution images of the HOPG were obtained by STM in all the gas atmospheres studied. However, it was found that the presence of water vapor gave rise to a noise current at increased pressures. Using contact AFM, the atomic resolution in an argon atmosphere decreased with increasing pressure, while atomic images were obtained under the other gas atmospheres at 1.1 MPa.

Investigation of influence of gas atmosphere and pressure upon non-contact atomic force microscopy.

Resonance measurements and atomic force microscopy (AFM) observations were carried out by the non-contact AFM operating in various gas atmospheres (hydrogen, helium, nitrogen and argon) over the range of pressures from 0.1 to 1.1 MPa. In each atmosphere, the resonance frequency of the AFM cantilever depended on the pressure of gases studied. The plots of the relative resonance frequency at a constant pressure vs. the gas density gave a straight line. It was found that the characteristic of the resonance frequency for the AFM cantilever were dependent on the density of the gas species. The resolution of the AFM was hardly influenced by the gas atmosphere under the present experimental conditions.