액체상 크로마토그래피와 MALDI 질량 분석기 연계 전자 분무 시료 증착

Abstract

In chapter 1, the focused electrospray (FES) deposition method is presented for matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. FES ion optics consists of two cylindrical focusing electrodes capped with a truncated conical electrode through which an electrospray emitter passes along the cylindrical axis. A spray of charged droplets is focused onto a sample well on a MALDI target plate under atmospheric pressure. The shape and size distributions of matrix crystals are visualized by scanning electron microscope and the mass spectra are obtained by time-of-flight mass spectrometry. Angiotensin II, bradykinin, and substance P are used as test samples, while α-cyano-4-hydroxycinnamic acid and dihydroxybenzoic acid are employed as matrices. FES of a sample/matrix mixture produces fine crystal grains on a 1&#8211

3 &#61549

m spot and reproducibly yields the mass spectra with little shot-to-shot and spot-to-spot variations. Although FES greatly stabilizes the signals, the space charge due to matrix ions limits the detection sensitivity of peptides. To avoid the space charge problem, we adopted a dual FES/FES mode, which separately deposits matrix and sample by FES in sequence. The dual FES/FES mode reaches the detection sensitivity of 0.88 amol, enabling ultrasensitive detection of peptides by homogeneously depositing matrix and sample under atmospheric pressure. This highly sensitive FES deposition method is applied to couple the liquid chromatography to MALDI-time-of-flight (TOF) mass spectrometry. A mixture of standard peptides as well as tryptic digest of BSA were loaded on LC, deposited by FES, and analyzed by MALDI-TOF. FES provides a highly uniform and concentrated spot of sample crystals with a few micrometer sizes on a MALDI plate, which enables detection of a sample with a great signal stability and reduced shot-to-shot fluctuations. Thus, the FES deposition method is compatible with high throughput LC-MS analysis of complex biomolecules.In chapter 2, experimental and theoretical studies of Li+ solvation by N-donor bases are described. Gas-phase solvation of Li+ with nitrogen-donor bases, such as pyrrolidine, 3-pyrroline, pyrrole, piperidine, piperazine, and pyridine, were studied using Fourier-transform ion cyclotron resonance mass spectrometry and ab initio theoretical calculations were performed to determine the structures of solvated complexes. The rate of one, two and three stepwise solvation was measured and the solvation enthalpies were determined from the van’t Hoff plot. The bond dissociation enthalpy of the Li+&#8226

(solvent)3 complex for solvent = pyridine, pyrrolidine, 3-pyrroline, and piperidine was measured to be &#8211

19.7 ± 1.6, &#8211

17.5 ± 1.5, &#8211

19.3 ± 1.1 and &#8211

17.3 ± 0.9 kcal mol-1, respectively. Theoretical binding energies are in accord with experiments. Of the N-donor molecules, pyridine shows the highest solvation energy to Li+, whereas piperidine shows the weakest solvation energy. The monodentate N-donors, such as pyridine, pyrrolidine, 3-pyrroline, and piperidine coordinate Li+ in end-on geometry.In chapter 3, chiral ammonium ion recognition by tris(oxazoline) receptor was studied by ab initio calculations. The origin of chiral ammonium ion recognition by the C3-symetric tripodal receptor was characterized by examining the difference in rotational barrier of the chiral ammonium ion between R and S complexes. Ab initio calculation shows that the R-complex is energetically more stable than the S- complex, which is in line with experiments showing that the R-form of α-phenylethyl ammonium ion is thermodynamically favored by tris(oxazoline) host. Notably, the rotation barrier of the phenyl group in α-phenylethyl ammonium ion significantly differs between the two diastereomeric host&#8211

guest complexes.In chapter 4, solvation of Li+ by ditropic ionophore was studied by experiments and theory. Trimethylboroxine (TMB) is a six-membered ring compound containing Lewis acidic boron and Lewis basic oxygen atoms that can bind halide anion and alkali metal cation, respectively. Fourier-transform ion cyclotron resonance spectroscopy was employed to study the gas-phase binding of LiBrLi+ and F&#8211

(KF)2 to TMB. TMB forms association complexes with both LiBrLi+ and F&#8211

(KF)2 at room temperature, providing direct evidence for the ditopic binding. Interestingly, the TMB&#8226

F&#8211

(KF)2 anion complex is formed 33 times faster than the TMB&#8226

LiBrLi+ cation complex. To gain insight into the ditopic binding of an ion pair, the structures and energetics of TMB&#8226

Li+, TMB&#8226

, TMB&#8226

LiF (the contact ion pair), and Li+&#8226

TMB&#8226

(the separated ion pair) were investigated using Hartree&#8211

Fock and density functional theory. Theory suggests that F&#8211

binds more strongly to TMB than Li+ and the contact ion-pair binding (TMB&#8226

LiF) is more stable than the separated ion-pair binding (Li+&#8226

). In addition, the ditopic binding of NaCl and KBr to TMB was investigated by theory. The ditopic character of TMB could be useful in extracting alkali halide in solution.