We have developed the NBS (2-nitrobenzenesulfenyl) method, a quantitative proteome analysis method utilizing stable isotope labeling followed by mass spectrometry. The potential of this method was reported previously, and the procedure has now been further optimized. Here, we describe a procedure utilizing urea or guanidine hydrochloride as a protein denaturant, in conjunction with an improved chromatographic enrichment method for the NBS-labeled peptides using a phenyl resin column. By using this new protocol, both sample loss throughout the protocol and the elution of unwanted unlabeled peptides can be minimized, improving the efficiency of the analysis significantly.

A real-time gas monitoring system based on optical absorption spectroscopy is proposed for localized carbon dioxide (CO2) measurement in respiratory tracts. In this system, a small gas cell is attached to the end of a hollow optical fiber that delivers mid-infrared light with small transmission loss. The diameters of the fiber and the gas cell are smaller than 1.2 mm so that the probe can be inserted into a working channel of common bronchoscopes. The dimensions of the gas cell are designed based on absorption spectra of CO2 standard gases in the 4.2 μm wavelength region, which are measured using a Fourier-transform infrared spectrometer. A miniature gas cell that is comprised of a stainless-steel tube with slots for gas inlet and a micro-mirror is fabricated. A compact probing system with a quantum cascade laser (QCL) light source is built using a gas cell with a hollow optical fiber for monitoring CO2 concentration. Experimental results using human breaths show the feasibility of the system for in-situ measurement of localized CO2 concentration in human airways.

A novel acidic morpholine-derivative containing glyceride (M-glyceride) was isolated from the cells of two strains of the thraustochytrid, Aurantiochytrium. The glyceride accounted for approximately 0.1 -0.4% of the lyophilized cells. The glyceride consisted of peaks I (85%) and II (15%). The structures of the intact and acetylated glycerides were elucidated by liquid chromatography-quadrupole time-of-flight chromatograph mass spectrometer (LC–Q/TOF) and NMR spectroscopy. The hydrate type of M-glyceride was detected as a minor component by LC–MS/MS. By 2D-NMR experiments, peaks I of the intact M-glyceride were elucidated as 1,2-didocosapentaenoyl-glyceryl-2′-oxy-3′-oxomorpholino propionic acid, and peak II was estimated 1,2-palmitoyldocosapentaenoyl- and/or 1,2-docosapentaenoylpalmitoyl-glyceryl-2′-oxy-3′-oxomorpholino propionic acid. The double bond position of docosapentaenoic acid was of the ω − 6 type (C22 = 5.ω − 6). The M-glyceride content varied by the cell cycle. The content was 0.4% of lyophilized cells at the mid logarithmic phase, and decreased to 0.1% at the mid stationary phase. When cells were grown in 1.0 µM M-glyceride-containing growth media, cell growth was stimulated to 110% of the control. With 0.1 µM acetyl M-glyceride, stimulation of 113% of the control was observed. Finding morpholine derivatives in biological components is rare, and 2-hydroxy-3-oxomorpholino propionic acid (auranic acid) is a novel morpholine derivative.

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Remarkable advances in performance of recent mass spectrometers promote dramatically the proteome analysis as well as the application of mass spectrometry to glycan analysis, and mass spectrometry has become one of the main analytical methods in glycan study. However, although many researchers can operate mass spectrometers easily these days due to the increase of user-friendliness of the instruments, mass spectrometry is still developing in theˆeld of glycan analysis, and it is very important not only to understand the principle of mass spectrometers and the related technologies but also to interpret obtained mass spectrum properly. In this paper, the glycan analysis using MALDI and ESI mass spectrometers is explained with some actual examples and useful notes. A. IntroductionMass spectrometry (MS) is, as the name indicates, an analytical technique that is used to measure mass. The information obtained through the mass spectrometry of glycans and glycoproteins covers a broad range that, in addition to mass, includes structural information, glycan binding site information, and quantitative information. The development of``matrix assisted laser desorption/ionization'' (MALDI) (1,2) and``electrospray ionization'' (ESI) (3) has enabled the ionization of biological macromolecules, and advances have been made in the application of mass spectrometry to glycan analysis. As a result of this, and because of its ability to achieve highsensitivity, rapid analysis, mass spectrometry is becoming an indispensable analytical technique for glycan analysis. In this report, while explaining some general points about measurement, some examples of the analysis of glycans and glycoproteins performed using MALDI mass spectrometry and ESI mass spectrometry will be presented.