Achieving Molecular Recognition of Structural Analogues in Surface-Enhanced Raman Spectroscopy: Inducing Charge and Geometry Complementarity to Mimic Molecular Docking

Molecular recognition of complex isomeric biomolecules remains challenging in surface-enhanced Raman scattering (SERS) spectroscopy due to their small Raman cross-sections and/or poor surface affinities. To date, the use of molecular probes has achieved excellent molecular sensitivities but still suffers from poor spectral specificity. Here, we induce “charge and geometry complementarity” between probe and analyte as a key strategy to achieve high spectral specificity for effective SERS molecular recognition of structural analogues. We employ 4-mercaptopyridine (MPY) as the probe, and chondroitin sulfate (CS) disaccharides with isomeric sulfation patterns as our proof-of-concept study. Our experimental and in silico studies reveal that “charge and geometry complementarity” between MPY's binding pocket and the CS sulfation patterns drives the formation of site-specific, multidentate interactions at the respective CS isomerism sites, which “locks” each CS in its analogue-specific complex geometry, akin to molecular docking events. Leveraging the resultant spectral fingerprints, we achieve > 97 % classification accuracy for 4 CSs and 5 potential structural interferences, as well as attain multiplex CS quantification with < 3 % prediction error. These insights could enable practical SERS differentiation of biologically important isomers to meet the burgeoning demand for fast-responding applications across various fields such as biodiagnostics, food and environmental surveillance.     

Kinetic analysis of superoxide anion radical-scavenging and hydroxyl radical-scavenging activities of platinum nanoparticles

There are few reports on the physiological effects of metal nanoparticles (nps), especially with respect to their functions as scavengers for superoxide anion radical (O2(.-)) and hydroxyl radical (.OH). We tried to detect the scavenging activity of Pt nps using a hypoxanthine-xanthine oxidase system for O2(.-) and using a Fenton and a UV/H2O2 system for .OH. Electron spin resonance analysis revealed that 2 nm particle size Pt nps have the ability to scavenge O2(.-) and .OH. The calculated rate constant for the O2(.-)-scavenging reaction was 5.03 +/- 0.03 x 10(7) M (-1) s (-1). However, the analysis of the Fenton and UV/H 2O 2 system in the presence of Pt nps suggested that the .OH-scavenging reaction cannot be determined in both systems. Among particle sizes tested from 1 to 5 nm, 1 nm Pt nps showed the highest O2(.-)-scavenging ability. Almost no cytotoxicity was observed even after adherent cells (TIG-1, HeLa, HepG2, WI-38, and MRC-5) were exposed to Pt nps at concentrations as high as 50 mg/L. Pt nps scavenged intrinsically generated reactive oxygen species (ROS) in HeLa cells. Additionally, Pt nps significantly reduced the levels of intracellular O2(.-) generated by UVA irradiation and subsequently protected HeLa cells from ROS damage-induced cell death. These findings suggest that Pt nps may be a new type of antioxidant capable of circumventing the paradoxical effects of conventional antioxidants.     

Integrating sensitive quantification of hepatic metabolic functions by capillary electrophoresis with laser-induced fluorescence detection

We report the establishment of capillary electrophoresis with laser-induced fluorescence (CE-LIF) detection as a common analytical platform for sensitive quantification of both phase I and II metabolism in various hepatic in vitro models.     

Ultra-performance liquid chromatography/time-of-flight mass spectrometry based metabolomics of raw and steamed Panax notoginseng

At present, metabolite profiling is of growing importance in herbal medicine fields such as breeding, formulation, quality control and clinical trials. This preliminary study indicated that ultra-performance liquid chromatography/time-of-flight mass spectrometry (UPLC/TOFMS)-based metabolomics allows direct detection of down-stream derivatives of metabolites, arising from the herbal formulation process. This analytical approach allows the discrimination and tentative authentication of unique biomarkers related to different herbal extracts using unsupervised multivariate principal component analysis (PCA). The tentative identification of biomarkers is complemented significantly by the accurate mass measurement of TOFMS and the high resolution and high retention time reproducibility rendered by UPLC. The application of this approach in herbal extract discrimination and ginsenoside biomarker discovery of raw and steamed Panax notoginseng (Burk.) F.H. Chen is demonstrated and discussed.     

Coulomb excitation of 74Ge beam

⁷⁴Ge beam was Coulomb-excited on a ^natPb target. Ten E2 matrix elements including diagonal matrix elements for 5 low-lying states have been determined using the least-squares search code GOSIA. The expectation values of the rotational invariants 〈Q ²〉 and 〈cos3δ〉 show the small and triaxial deformation of the two lowest members of the ground-state band , while the 0₂ ⁺ and 2₂ ⁺ states are found to be almost spherical. Received: 31 August 2000 / Accepted: 4 December 2000     

Purity of rubrene single crystals

Liquid chromatography connected with mass spectroscopy reveals that the oxidized form of rubrene is the major impurity in commercial powder of rubrene as well as in rubrene single crystals. One form of rubrene impurity can be transformed into the other. In solution, rubrene undergoes photo-oxidation completely until the red color of the rubrene solution disappears. Single crystals, due to compact packing of molecules and the required molecular shape change during oxidation, oxidize only on the surface.     

Study of the vi13/2-1 band in 189Pt

High-spin levels of 189Pt have been studied with the in-beam γ-spectroscopy method via the 176Yb(18O,5n) reaction at the beam energies of 88 and 95 MeV. The previously known vi13/2-1 band has been confirmed, and its unfavored signature branch extended up to the 31/2+ state. Within the framework of the triaxial particle-rotor model, the vi13/2-1 band is suggested to be associated with the 11/2[615] configuration, and to have triaxial deformation.     

Two-dimensional fluorescence difference gel electrophoresis for comparison of affinity and non-affinity based downstream processing of recombinant monoclonal antibody

Although Staphylococcus Protein A (SpA) affinity chromatography is the state of the art capture step for antibody purification, non-affinity methods are more economical. We used two-dimensional fluorescence difference gel electrophoresis (2-D DIGE) to evaluate the purification of a recombinant IgG₁ antibody from cultured cells, with two different processes: (1) SpA capture followed by cation-exchange chromatography (CEX); and (2) CEX capture, followed by anion exchanger, then hydrophobic interaction chromatography. Efficiencies were similar in sodium dodecylsulphate polyacrylamide gel electrophoresis and size-exclusion chromatography; however, 2-D DIGE revealed higher efficiency with SpA than with CEX capture. Thus, 2-D DIGE is a valuable tool for downstream process development.     

Quantitative analysis of human serum leptin using a nanoarray protein chip based on single-molecule sandwich immunoassay

We report a method for the quantitative analysis of human serum leptin, which is a protein hormone associated with obesity, using a nanoarray protein chip based on a single-molecule sandwich immunoassay. The nanoarray patterning of a biotin-probe with a spot diameter of 150 nm on a self-assembled monolayer functionalized by MPTMS on a glass substrate was successfully accomplished using atomic force microscopy (AFM)-based dip-pen nanolithography (DPN). Unlabeled leptin protein molecules in human serum were detected based on the sandwich fluorescence immunoassay by total internal reflection fluorescence microscopy (TIRFM). The linear regression equation for leptin in the range of 100 zM-400 aM was determined to be y = 456.35x + 80,382 (R = 0.9901). The accuracy and sensitivity of the chip assay were clinically validated by comparing the leptin level in adult serum obtained by this method with those measured using the enzyme-linked immunosorbent assay (ELISA) performed with the same leptin standards and serum samples. In contrast to conventional ELISA techniques, the proposed chip methodology exhibited the advantages of ultra-sensitivity, a smaller sample volume and faster analysis time.     

Sensitive detection of potassium ion using Prussian blue nanotube sensor

A model K⁺ sensor using Prussian blue nanotubes is fabricated by electrochemical deposition of Prussian blue (PB) within the nanochannels of a porous metal-coated membrane with partially covered pore openings. The PB nanotube sensor exhibits excellent stability giving reproducible peak potentials up to 500 measurement cycles, a very low detection limit of 2.0 × 10⁻⁸ M and extremely wide logarithmic linear ranges between 5.0 × 10⁻⁸–7.0 × 10⁻⁴ M and 7.0 × 10⁻⁴–1.0 M. Negligible interferences by Na⁺, Mg²⁺ and Ca²⁺ are observed and a rapid analysis time of 30 s is readily achieved. The ease of electrodeposition, high stability of PB nanotubes and outstanding analytical performance which surpasses conventional PB voltammetric and potentiometric sensors demonstrates potential sensing applications including ion sensors and biosensors using PB and other metal hexacyanoferrate nanotubes.