Baseline noise and measurement uncertainty in liquid chromatography.

The stochastic properties of baseline noise in HPLC systems with a UV photo-diode array, photo-multiplier and gamma-ray detector were examined by dividing the noise into auto-correlated random process (Markov process) and an independent process (white noise). The present work focused on the effect of the stochastic noise properties on a theoretical estimation of the standard deviation (SD) of area measurements in instrumental analyses. An estimation theory, called FUMI theory (Function of Mutual Information), was taken as an example. A computer simulation of noise was also used. It was shown that the reliability (confidence intervals) of theoretical SD estimates mainly depends on the following factors: the ratio of the white noise and Markov process occurring in the baselines; the number of data points used for the estimation; the width of a target peak for which the SD is estimated.     

Novel fluorescent asparaginyl-N-acetyl-d-glucosamines (Asn-GlcNAc) for the resolution of oligosaccharides in glycopeptides, based on enzyme transglycosylation reaction

Several fluorescent asparaginyl-N-acetyl-d-glucosamines (Asn-GlcNAcs), i.e., DBD-Asn-GlcNAc, NBD-Asn-GlcNAc, NDA-Asn-GlcNAc, PS-Asn-GlcNAc, FITC-Asn-GlcNAc, DMEQ-Asn-GlcNAc and DBD-PZ-Boc-Asn-GlcNAc, were synthesized as the acceptors for the resolution of oligosaccharides in glycopeptides. The resolution of oligosaccharides is based on the transglycosylation reaction with end-β-N-acetylglucosaminidase (Endo-M), isolated from Mucor hiemalis. The synthesis of each fluorescent acceptor was carried out in a one-pot reaction of Asn-GlcNAc and the corresponding fluorescent tagging reagent. The transglycosylation reaction using Endo-M proceeds at RT in neutral phosphate buffer (pH 6.0) and reached maxima at around 30 min. When Fmoc-Asn-GlcNAc (acceptor) was enzymatically reacted with Disialo-Asn (donor) in the presence of Endo-M, the ratio of Disialo-Asn-Fmoc (transglycosylation product) was almost comparable with the decreasing ratio of Fmoc-Asn-GlcNAc. Therefore, the transglycosylation activity of Endo-M from Disialo-Asn (donor) and fluorescent-Asn-GlcNAc (acceptor) was calculated from the decreasing ratio of fluorescent-Asn-GlcNAc. The order was NDA-Asn-GlcNAc > DBD-Asn-GlcNAc Fmoc-Asn-GlcNAc > NBD-Asn-GlcNAc DMEQ-Asn-GlcNAc > DNS-Asn-GlcNAc > PS-Asn-GlcNAc > FITC-Asn-GlcNAc. On the other hand, the activity with a fluorescent acceptor (DBD-PZ-Boc-Asn-GlcNAc), labeled to a carboxylic acid group in the Asn residue, was the strongest among the synthesized acceptors.     

Training humans in non-native phoneme perception using a monkey psychoacoustic procedure

Humans were trained to categorize problem non-native phonemes using an animal psychoacoustic procedure that trains monkeys to greater than 90% correct in phoneme identification [Sinnott and Gilmore, Percept. Psychophys. 66, 1341-1350 (2004)]. This procedure uses a manual left versus right response on a lever, a continuously repeated stimulus on each trial, extensive feedback for errors in the form of a repeated correction procedure, and training until asymptotic levels of performance. Here, Japanese listeners categorized the English liquid contrast /r-l/, and English listeners categorized the Middle Eastern dental-retroflex contrast /d-D/. Consonant-vowel stimuli were constructed using four talkers and four vowels. Native listeners and phoneme contrasts familiar to all listeners were included as controls. Responses were analyzed using percent correct, response time, and vowel context effects as measures. All measures indicated nativelike Japanese perception of /r-l/ after 32 daily training sessions, but this was not the case for English perception of /d-D/. Results are related to the concept of "robust" (more easily recovered) versus "fragile" (more easily lost) phonetic contrasts [Burnham, Appl. Psycholing. 7, 207-240 (1986)].     

Nonlinear nanodevices using magnetic flux quanta

All devices realized so far that control the motion of magnetic flux quanta employ either samples with nanofabricated spatially-asymmetric potentials (which strongly limit controllability), or pristine superconductors rectifying with low-efficiency time-asymmetric oscillations of an external magnetic field. Using layered Bi2Sr2CaCu2O8+delta materials, here we fabricate and simulate two efficient nonlinear superconducting devices with no spatial asymmetry. These devices can rectify with high-efficiency a two-harmonic external current dragging vortices in target directions by changing either the relative phase or the frequency ratio of the two harmonics.     

A novel approach for LC-MS/MS-based chiral metabolomics fingerprinting and chiral metabolomics extraction using a pair of enantiomers of chiral derivatization reagents

Chiral metabolites are found in a wide variety of living organisms and some of them are understood to be physiologically active compounds and biomarkers. However, the overall analysis of chiral metabolomics is quite difficult due to the high number of metabolites, the significant diversity in their physicochemical properties, and concentration range from metabolite-to-metabolite. To solve this difficulty, we developed a novel approach for chiral metabolomics fingerprinting and chiral metabolomics extraction, which is based on the labeling of a pair of enantiomers of chiral derivatization reagents (i.e., DMT-(S,R)-Pro-OSu and DMT-3(S,R)-Apy) and precursor ion scan chromatography of the derivatives. The multivariate statistics is also required for this strategy. The proposed procedures were evaluated by the detection of a diagnostic marker (i.e., d-lactic acid) using the saliva of diabetic patients. This method was used for the determination of biomarker candidates of chiral amines and carboxyls in Alzheimer's disease (AD) brain homogenates. As the results, l-phenylalanine (L-Phe) and l-lactic acid (L-LA) were identified as the decreased and increased biomarker candidates in the AD brain, respectively. Therefore, the proposed approach seems to be helpful for the determination of non-target chiral metabolomics possessing amines and carboxyls.     

Towards the chiral metabolomics: Liquid chromatography–mass spectrometry based dl-amino acid analysis after labeling with a new chiral reagent, (S)-2,5-dioxopyrrolidin-1-yl-1-(4,6-dimethoxy-1,3,5-triazin-2-yl)pyrrolidine-2-carboxylate,and the application to saliva of healthy volunteers

A novel triazine-type chiral derivatization reagent, i.e., (S)-2,5-dioxopyrrolidin-1-yl-1-(4,6-dimethoxy-1,3,5-triazin-2-yl) pyrrolidine-2-carboxylate (DMT-(S)-Pro-OSu), was developed for the highly sensitive and selective detection of chiral amines and amino acids by UPLC–MS/MS analysis. The enantiomers of amino acids were easily labeled with the reagents at room temperature within 40 min in an alkaline medium containing triethylamine. The diastereomers derived from proteolytic amino acids, except serine, were well separated under isocratic elution conditions by reversed-phase chromatography using an ODS column (R_s = 1.2–9.0). dl-Serine was separated by use of an ADME column which has relatively higher polar surface than the conventional ODS column. The characteristic product ions, i.e., m/z 195.3 and m/z 209.3, were detected from all the diastereomers by the collision-induced dissociation of the protonated molecule. A highly sensitive detection on the amol–fmol level was obtained from the selected reaction monitoring (SRM) chromatogram. The chiral amines (e.g., adrenaline and noradrenaline) labeled with DMT-(S)-Pro-OSu were also well separated and sensitively detected by the present procedure. The method using DMT-(S)-Pro-OSu was used for the determination of dl-amino acids in the human saliva from healthy volunteers. Various l-amino acids were identified in the saliva. Furthermore, d-alanine (d-Ala) and d-proline (d-Pro) were also detected in relatively high concentrations (>5%). The ratio was higher in male saliva than in female saliva. However, the difference in the ratio of d-Ala for one day was not very high and the effect of foods and beverage seemed to be negligible. Based on the results using l-Ala-d₃, the d-Ala in saliva seemed to be produced due to the racemization with some enzymes such as racemase. The racemization reaction was reversible, i.e., d-Ala-d₃ was also racemized to l-Ala-d₃ in saliva. Thus, care should be taken during the analysis of dl-amino acids in saliva. The present method using DMT-(S)-Pro-OSu may be applicable for the determination of chiral amine metabolomics, because the resulting derivatives produce the same product ions without relation to the compounds and show highly sensitive detection in the SRM mode of MS/MS. Consequently, DMT-(S)-Pro-OSu seems to be a useful chiral derivatization reagent for the determination of amines and amino acids in biological samples.     

Analysis of natural convection melting inside isothermally heated horizontal rectangular capsule

Melting characteristics of phase change material by natural convection heat transfer inside horizontal rectangular capsules are examined experimentally. The capsules are heated isothermally and three kinds of aspect ratios (qH/W=3, 1 and 1/3) are provided. Octadecane and ice are used, respectively, as the phase change material. A method of analysis applying the empirical correlations for natural convection heat transfer in a vertical or horizontal enclosure to the melting in the rectangular capsules is presented. The predicted results show good agreement with the experimental data. For the melting of octadecane and ice, it is found that the effect of aspect ratio on the melting process is not significant for the range ofB=1/3 to 3.     

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.     

A novel hydrazide type organocatalyst for enantioselective Diels-Alder reactions

The development of a new class of hydrazide type organocatalyst, (4R,5R)-1,3-bis(isopropylamino)-4,5-dihenylimidazolidin-2-one 2a, for enantioselective Diels-Alder reactions between cyclopentadiene and α,β-unsaturated aldehydes are presented. The new organocatalyst 2a promoted the reaction, affording Diels-Alder adducts in good yields with good levels of enantioselectivity.     

An MD simulation of the decoy action of Epstein�CBarr virus LMP1 protein mimicking the CD40 interaction with TRAF3

The Epstein?CBarr virus (EBV) is associated with a variety of malignancies and chronic active EBV infection is a severe systemic disease associated with high rates of mortality and morbidity. In this paper, the dynamics of the interaction of EBV-expressed latent membrane protein 1 (LMP1) with cellular signaling intermediate tumor necrosis factor receptor (TNFR)-associated factor 3 (TRAF3) is simulated using standard classical molecular dynamics (MD) protocols. For comparison, the dynamics of the interaction of TRAF3 with CD40, a TNFR mimicked by LMP1 to effect EBV infection is also calculated under similar conditions. Essential dynamics (ED) analysis is carried out to identify important degrees of vibrational freedom that relate to protein function and virus infection. Both the MD simulation and ED analysis reveal novel interactions that help explain the structural decoy action of LMP1 over CD40. These interactions involve the consensus sequence PXQXTXX shared by CD40 and LMP1. In LMP1, we have found novel interaction of Asp 209 with TRAF3 and the interaction is crucial although the adjacent Asp 210 was suggested to be essential by the X-ray analysis. In CD40, it is found that the hairpin formation is not indispensable for the interaction with TRAF3.