Research

In this page, the research topics on which shimabukuro is working are shown below. All research topics have been started since 2016. I am in charge of all of this research such as design, experiment, analysis, and writing a paper. In order to improve the research quality, I discuss the experimental results with extramural researcher at the academic meeting.


In this research topic, reactions between electrically neutral chemically reactive species and biomolecules have been being surveyed. Our ultimate goal is building a novel medical infrastructure system utilizing mass spectrometry (MS) to diagnose diseases from a drop of blood at anywhere in the world. This research is started from April, 2016 as a joint research with Koichi Tanaka Mass Spectrometry Research Laboratory, Shimadzu Corporation.

Background

The base sequence of DNA are translated to linear sequence of amino acids of peptides. The structured peptides are modified or integrated to the Post-translational modification (PTM) that realizes the specific function of the peptides. Identification of which functional groups attached to which amino acid requires new method for chemical analysis. Recently, some PTMs are shown to be determined by cleaving peptides in the mass spectrometer preserving fragile functional groups with ECD (Electron Capture Dissociation) and ETD (Electron Transfer Dissociation).

These electron induced dissociation techniques enable the analyses of PTM, however target ions are limited to highly charged positive ions. Therefore, developing of new fragmentation technique which does not depend upon the charged state and mass of target ions preserving PTMs is important.

Contrary to the electron based analyses, our method called Hydrogen Attachment/Abstraction Dissociation or HAD enables analysis of negative or singly charged positive ions. In HAD, electrically neutral hydrogen radical attaches to peptide ion or abduct atomic hydrogen from peptide ion to cause fragmentation of peptide ions preserving PTMs. Since the hydrogen radical is electrically neutral, HAD method does not depend on the charged state of the target ions. Therefore, the HAD method has the advantage over electron based analyses in terms of target ions to be analyzed. In this original study, a thermal cracking source was utilized as an atomic hydrogen source. The designed thermal cracking cell produces hydrogen radicals with a high degree of dissociation by the catalytic effect on heated tungsten surface maintained about 2300 K. However, thermal cracker does not have a capability to generate atoms reactive to high temperature tungsten. The heating filament does not last long in the system even with a trace amount of oxygen. Thus, a long-life time versatile radical source based upon the microwave plasma generator is being developed.

In order to expand the analyzable biomolecules by opening the new fragmentation methods using a various kinds of low-energy radicals, versatile radical beam sources has been developed in this study. Thermal dissociation of typical molecules except for hydrogen is impossible due to the high bond-dissociaiton energies. Thus, chemically reactive radicals should be produced by discharge of gases or liquids. A radio frequency (RF) plasma source can dissociate many kinds of reactive gases, such as oxygen, water vapor, hydrochloric acid, etc., and is suitable as a radical source for operation with chemically reactive species because the RF source can generate plasma in the vacuum vessel with electrode less configuration. Especially in the application of gas-phase mass spectrometry, gas introduction to the reaction chamber of the system with high flow rate induces poor S/N ratio, insufficient mass separation, and damage to the detector. Thus, the radical source must be operated under a low gas pressure condition to realize high sensitivity measurement. In general, microwave excited plasma can ignite and sustain plasma at the lower gas pressure condition than typical RF plasma source. Moreover, the combination with electron cyclotron resonance (ECR) can enhance the density and the degree of dissociation by effective electron acceleration. In order that the reactive radicals realize the biomolecular structural analyses with sufficient reaction rate, microwave driven radical beam sources were developed to satisfy high atomic flux, high degree of dissociation, low operating pressure, and less heat loss.

(1)Development of microwave driven radical sources

Discharge type radical sources such as RF discharges at 13.56, 27.12, 40.68 MHz have the possibility to overcome problems of thermal cracking type atomic beam source. These kinds of RF plasma sources have been utilized in the field of surface sciences. However, the entire system is relatively large due to the external matching unit and a large antenna depending upon the wavelength. Besides these spatial problems, the RF plasma requires power input and gas input higher than the microwave plasma. We have been developing two types of neutral beam sources utilizing microwave discharge: the capacitively coupled plasma (CCP) type with a needle shaped electrode, and the localized inductively coupled plasma (LICP) type that couples the input microwave power to the plasma by winding a coil around a small diameter dielectric tube.

CCP type radical beam sources (1st gen. – 4th gen.)
We developed four CCP type radical beam sources. In the CCP configuration, a plasma is ignited and sustained by an intense electric field concentration at the source tip. A copper tube and a copper capillary are surrounded by a tapered quartz glass vessel and a grounded electrode. The grounded electrode has a small beam aperture to intensify the local electric field confronting to the copper capillary. A plasma plume – plasma leaking of the exit aperture due to the high density – is formed under the high input power high pressure condition.

LICP type radical beam sources (1st gen. – 2nd gen.)
We developed two LICP type radical beam sources. In the LICP configuration, a 2.45 GHz microwave power source supplies the microwave to a 0.3 mm thickness copper spiral antenna tightly wound around a 6 mm outer diameter 4 mm inner diameter alumina or quartz glass tube via N-type connector without any external impedance matching unit. The system that does not expose any metallic parts from the gas inlet to the atom exit contributing to reduce contaminants in plasma and recombination of atomic species at the metallic wall. Two groups of Nd-Fe magnets from the magnetic field which satisfies the ECR condition in the axial direction.

(2) Development of new biomolecular structural analysis techniques

 

 

Research background

One main application area of negative hydrogen (H) ion sources is the field of the thermo nuclear fusion research. According to the Lawson’s criterion, the ion temperature in the fusion plasma to optimize the D(deuterium)-T(tritium) reaction has to be higher than several tens of keV. Heating of plasma is a necessary process to meet this criterion by injection energy from the outside of the reactor. One of the H ion production mechanism is believed due to the low-energy (1 eV) atomic hydrogen injection to the plasma grid surface. However, the phenomenon has yet to be observed in the actual ion source operating condition. In this chapter, developed three types of atomic hydrogen sources are utilized for low-energy H0 production to see the conversion effect of atomic hydrogen to negative ion as a part of the radical source performance evaluation.

 


The surface production of H ion by atomic hydrogen reflection has not been directly confirmed in the actual ion source operating condition yet.  In this study, the atomic source which is developed for biopolymer analysis is attached to a cesiated multicusp type H ion source to see if the additional plasma operation enhances the extracted H ion current in an actual hydrogen plasma condition. A change in H ion current extracted from a cesiated H ion source was observed by injecting an amplitude modulated hydrogen atomic beam. A 2.45 GHz microwave power excited hydrogen plasma for injecting atoms to the plasma grid of the multicusp type H ion source.