Determination of ricin by nano liquid chromatography/mass spectrometry after extraction using lactose-immobilized monolithic silica spin column.

Ricin is a glycosylated proteinous toxin that is registered as toxic substance by Chemical Weapons convention. Current detection methods can result in false negatives and/or positives, and their criteria are not based on the identification of the protein amino acid sequences. In this study, lactose-immobilized monolithic silica extraction followed by tryptic digestion and liquid chromatography/mass spectrometry (LC/MS) was developed as a method for rapid and accurate determination of ricin. Lactose, which was immobilized on monolithic silica, was used as a capture ligand for ricin extraction from the sample solution, and the silica was supported in a disk-packed spin column. Recovery of ricin was more than 40%. After extraction, the extract was digested with trypsin and analyzed by LC/MS. The accurate masses of molecular ions and MS/MS spectra of the separated peptide peaks were measured by Fourier transform-MS and linear iontrap-MS, respectively. Six peptides, which were derived from the ricin A-(m/z 537.8, 448.8 and 586.8) and B-chains (m/z 701.3, 647.8 and 616.8), were chosen as marker peptides for the identification of ricin. Among these marker peptides, two peptides were ricin-specific. This method was applied to the determination of ricin from crude samples. The monolithic silica extraction removed most contaminant peaks from the total ion chromatogram of the sample, and the six marker peptides were clearly detected by LC/MS. It takes about 5 h for detection and identification of more than 8 ng/ml of ricin through the whole handling, and this procedure will be able to deal with the terrorism using chemical weapon.

Preparation and evaluation of lactose-modified monoliths for the adsorption and decontamination of plant toxins and lectins.

A series of sugar-modified porous silica monoliths with different sugar ligands (ß-lactoside, ß-N-acetyllactosaminide, ß-d-galactoside, ß-d-N-acetylgalactosaminide and ß-d-glucoside) and linkers were prepared and evaluated using plant toxins and lectins including ricin and a Ricinus communis agglutinin (RCA(120)). Among these sugar monoliths, a lactose monolith carrying a triethylene glycol spacer adsorbed ricin and RCA(120) with the highest efficiency. The monolith showed no binding with albumin, globulin, and lectins from Jack beans, Osage orange, Amur maackia and wheat germ. All these data support the utility of the lactose-modified monolith as a tool for adsorption and decontamination of plant toxins.

Preparation and supramolecular properties of unadulterated glycosyl liposomes from a bis(alpha-D-mannopyranosyl)-[60]fullerene conjugate.

The bis(alpha-D-mannopyranosyl)-[60]fullerene conjugate 3 was prepared by thermal coupling of C60 and either 2-azidoethyl 2,3,4,6-tetra-O-acetyl- or 2,3;4,6-di-O-isopropylidene-alpha-D-mannopyranoside (Scheme). Compound 3 was found to readily self-assemble. Dynamic-light-scattering (DLS) and atomic-force microscopy (AFM) experiments supported that the amphiphilic compound gives rise to nano-sized supramolecular structures during sugar deprotection (Ac-group removal) performed in MeOH/CH2Cl2 solution. Encapsulation studies with an aqueous suspension of 3 showed that the self-assembling structure envelopes Ba2+ and the fluorescent dye Acridine Red during its formation, which indicates that it resembles a bilayer vesicle or an unadulterated liposome with an inner hollow space. In addition to this notable property, the unique molecular geometry of the spatially arranged mannosyl surface residues of 3 gives rise to strong binding of the carbohydrate-recognizing lectin Con A. Hence, the polar amphiphilic end of 3 mimics the structure of 3,6-branched tri-alpha-D-mannoside (6; Fig. 3), a natural ligand of the Con A protein.

Stereo- and biochemical profiles of the 5-6- and 6-6-junction isomers of alpha-D-mannopyranosyl [60]fullerenes.

The 5-6- and 6-6-junction isomers of alpha-D-mannopyranosyl [60]fullerene were studied by means of circular dichroism (CD), deuterium labeling, 1H-NMR, molecular-dynamics (MD) calculations, and a lectin-binding assay. The CD spectra of the O-acetylated derivatives allowed clear discrimination of the isomers, while the 1H-NMR spectra, with assistance from deuterium labeling and MD calculations, served to disclose the unique conformation and molecular geometry of each acetylated isomer in chloroform solution. The deprotected 5-6- and 6-6-isomers, which gave colloidal suspensions in aqueous mixtures, displayed marked activity in blocking lectin-induced hemagglutination by concanavalin A.

Structure of a missing-caged metallofullerene: La2@C72.

The La2@C72 metallofullerene having the so-called "missing" C72 fullerene cage was structurally elucidated by using 13C NMR and 139La NMR spectroscopy. The obtained structure of La2@C72 does not satisfy fullerene's structural golden rule, that is, the isolated-pentagon rule. The structure is consistent with a non-IPR D2-C72 (#10611) cage structure where each La atom is situated close to one of the two-fused pentagons.

Lanthanoid endohedral metallofullerenols for MRI contrast agents.

Water-soluble multi-hydroxyl lanthanoid (La, Ce, Gd, Dy, and Er) endohedral metallofullerenes (metallofullerenols, M@C(82)(OH)(n)()) have been synthesized and characterized for the use of magnetic resonance imaging (MRI) contrast agents. The observed longitudinal and transverse relaxivities for water protons, r(1) and r(2), of the metallofullerenols are in the range 0.8-73 and 1.2-80 (sec(-1)mM(-1)), respectively, which are significantly higher than those of the corresponding lanthanoid-DTPA chelate complexes. Among these Gd-metallofullerenols, Gd@C(82)(OH)(n)() has exhibited the highest r(1) and r(2) values in consistent with our previous results. The observed large r(1) of the current metallofullerenols can mainly be ascribed to the dipole-dipole relaxation together with a substantial decrease of the overall molecular rotational motion. The large r(2), except for the Gd-metallofullerenols, have been attributed to the so-called Curie spin relaxation. The MRI phantom studies are also performed and are consistent with these results. The metallofullerenols will be an ideal model for future MRI contrast agents with higher proton relaxivities.