Determination Method for Pyrroloquinoline Quinone in Food Products by HPLC-UV Detection Using a Redox-Based Colorimetric Reaction.

We have developed an HPLC-UV method for the determination of pyrroloquinoline quinone (PQQ), which utilizes a redox-based colorimetric reaction. In the proposed colorimetric reaction, the redox reaction between PQQ and dithiothreitol generates superoxide anion radicals that can convert 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyl-2H-tetrazolium chloride (INT) to formazan dye. After PQQ separation on an octadecyl silica column, it was mixed online with dithiothreitol and INT, and the formed formazan dye was monitored by absorbance at 490 nm. The detection limit (S/N = 3) of the proposed method was 7.6 nM (152 fmol/injection). The proposed method could selectively detect PQQ in food products without any clean-up procedures.

Succinimidyl (3-[(benzyloxy)carbonyl]-5-oxo-1,3-oxazolidin-4-yl)acetate on a triazole-bonded phase for the separation of dl-amino-acid enantiomers and the mass-spectrometric determination of chiral amino acids in rat plasma.

Changes in the levels of amino-acid enantiomers are associated with some serious diseases; consequently, amino acid monitoring in peripheral blood can be used to diagnose and predict the onset of disease. Herein, we report the design and synthesis of a new chiral derivatization reagent, namely succinimidyl (4S)-(3-[(benzyloxy)carbonyl]-5-oxo-1,3-oxazolidin-4-yl)acetate ((S)-COXA-OSu), for the separation of dl-amino-acid enantiomers. The usefulness of (S)-COXA-OSu was examined as a derivatization reagent for LC-MS/MS following certification of its total optical purity (>99%). The enantiomeric separations of amino-acid derivatives tagged with the reagent were examined using a triazole-bonded phase. (S)-COXA-OSu enabled the simultaneous enantiomeric separation of more than 40 α-amino acids. (S)-COXA-amino-acid derivatives were efficiently converted into their product ions, from which formaldehyde (CH2O) was eliminated [M-30] from the oxazolidinone moiety of COXA by collision-induced dissociation during LC-MS/MS. Limits of detection were in the 0.0138-0.518 pmol/injection range. For precise and accurate quantitation, we synthesized and used a stable-isotope-labeled (S)-COXA-OSu that was used as an internal standard in LC-MS/MS-determination experiments. Finally, changes in plasma amino-acid levels in rats, following administration of S-methyl-l-cysteine, an alanine-serine-cysteine transporter-1 (Asc-1) inhibitor, were successfully detected by LC-MS/MS using (S)-COXA-OSu.

Ultrasensitive determination of pyrroloquinoline quinone in human plasma by HPLC with chemiluminescence detection using the redox cycle of quinone.

A fast, accurate, and ultrasensitive high-performance liquid chromatography method with chemiluminescence detection (HPLC-CL) was optimized and validated for the determination of pyrroloquinoline quinone (PQQ) concentration in human plasma following solid-phase extraction (SPE). This method is based on the redox cycle of the reaction between PQQ and dithiothreitol, which generates reactive oxygen species that can be detected using luminol as a CL probe. The isocratic HPLC system comprised an ODS column and 4.0mM tetra-n-butylammonium bromide in Tris-HNO3 buffer (pH 8.8; 50mM)-acetonitrile (7:3, v/v) as mobile phase. A novel, rapid, and simple SPE method was also developed providing excellent %recovery (≥95.2%) for PQQ from human plasma samples. The proposed method was linear over the range of 4.0-400nmol/L plasma of PQQ with a lower detection limit (S/N=3) of 1.08 nmol/L plasma (0.27nM). The method was successfully implemented to determine PQQ concentration in the plasma of healthy individuals after administration of PQQ supplements.

Identification of muscle synergies associated with gait transition in humans.

There is no theoretical or empirical evidence to suggest how the central nervous system (CNS) controls a variety of muscles associated with gait transition between walking and running. Here, we examined the motor control during a gait transition based on muscle synergies, which modularly organize functionally similar muscles. To this end, the subjects walked or ran on a treadmill and performed a gait transition spontaneously as the treadmill speed increased or decreased (a changing speed condition) or voluntarily following an experimenter's instruction at constant treadmill speed (a constant speed condition). Surface electromyograms (EMGs) were recorded from 11 lower limb muscles bilaterally. We then extracted the muscle weightings of synergies and their activation coefficients from the EMG data using non-negative matrix factorization. As a result, the gait transition was controlled by approximately 9 muscle synergies, which were common during a walking and running, and their activation profiles were changed before and after a gait transition. Near a gait transition, the peak activation phases of the synergies, which were composed of plantar flexor muscles, were shifted to an earlier phase at the walk-to-run transition, and vice versa. The shifts were gradual in the changing speed condition, but an abrupt change was observed in the constant speed condition. These results suggest that the CNS low-dimensionally regulate the activation profiles of the specific synergies based on afferent information (spontaneous gait transition) or by changing only the descending neural input to the muscle synergies (voluntary gait transition) to achieve a gait transition.