Application of stir bar sorptive extraction for wine analysis

Stir bar sorptive extraction (SBSE) coupled with gas chromatography/mass spectrometry (GC/MS) was used to analyse wine samples for three applications: flavour and compositional analysis; 2,4,6-trichloroanisole (TCA), a common off-aroma in wine; and agrochemicals. SBSE was found to be orders of magnitude more sensitive than modern conventional methodology, allowing for lower detection and quantitation levels, and improved confirmation of identity; SBSE often gave better signal to noise in scan mode than other methods in selective ion monitoring (SIM) mode. With the help of their characteristic mass spectra all agrochemicals could be identified unambiguously at concentrations of 10 microg L(-1) in wine and a further 100 constituents were detected in a Cabernet Sauvignon sample. Thus it is now possible to analyse complex samples such as wine by scan mode, with better confirmation of identity, and without sacrificing sensitivity, where previously SIM methodology had to be used.     

Tobacco protein separation by aqueous two-phase extraction

Tobacco has long been considered as a host to produce large quantity of high-valued recombinant proteins. However, dealing with large quantities of biomass is a challenge for downstream processing. Aqueous two-phase extraction (ATPE) has been widely used in purifying proteins from various sources. It is a protein-friendly process and can be scaled up easily. In this paper, ATPE was studied for its applicability to recombinant protein purification from tobacco with egg white lysozyme as the model protein. Separate experiments with poly(ethylene glycol) (PEG)-salt-tobacco extract and PEG-salt-lysozyme were carried out to determine the partition behavior of tobacco protein and lysozyme, respectively. Two-level fractional factorial designs were used to study the effects of factors such as, PEG molecular mass, PEG concentration, the concentration of phase forming salt, sodium chloride concentration and pH, on protein partitioning. The results showed that, among the studied systems, PEG-sodium sulfate system was most suitable for lysozyme purification. Detailed experiments were conducted by spiking lysozyme into the tobacco extract. The conditions with highest selectivity of lysozyme over native tobacco protein were determined using a response surface design. The purification factor was further improved by decreasing the phase ratio along the tie line corresponding to the phase compositions with the highest selectivity. Under selected conditions the lysozyme yield was predicted to be 87% with a purification factor of 4 and concentration factor of 14. From this study, ATPE was shown to be suitable for initial protein recovery and partial purification from transgenic tobacco.     

Observation of a quadrupole plasmon mode for a colloidal solution of gold nanoprisms

The synthesis and optical properties of single crystalline gold nanoprisms have been investigated. A three-step mediated seed growth process in an aqueous solution generated gold nanoprisms with a relatively homogeneous size distribution. The purity of these nanostructures has allowed us to observe a weak quadrupole resonance in addition to a strong dipole resonance associated with these novel structures. The experimental optical spectra agree with discrete dipole approximation calculations that have been modeled from the dimensions of gold nanoprisms produced in this synthesis.     

The use of novel water-in-oil microemulsions in microemulsion electrokinetic chromatography

We describe the novel use of water-in-oil (W/O) microemulsions to achieve unique separations in microemulsion electrokinetic chromatography (MEEKC). The choice and concentration of the buffer type, surfactant and co-surfactant were all examined and optimized. Separations of a range of neutral and acidic analytes was shown to be markedly different to that obtained by (oil-in-water) O/W MEEKC. Neutral solutes are separated by virtue of their solubility (log P) values in O/W MEEKC with the more water-insoluble solutes migrating last. This separation process does not occur in W/O, as neutral solutes are not separated in order of log P.     

PHOTOSENSITIZATION WITH BACTERIOCHLORINS

Abstract— Biophysical and photobiological properties of a group of bacteriochlorins were compared with efficacy of these products for photodynamic therapy of murine tumors. Predictive factors for selective photosensitization in vivo include affinity binding to lipoproteins greater than albumin, extinction coefficient at the wavelength of irradiation and tumor/skin distribution. Efficacy was correlated with circulating plasma levels of the different sensitizers but not with the photodynamic therapy response in cell culture.     

Mössbauer Spectroscopic Studies of Chlorostannate(II) Complexes in Frozen Aqueous Solutions

Mössbauer parameters of frozen aqueous solutions of Sn(ClO₄)₂ in 0.5 M HClO₄ did not change with the concentration of NaClO₄, so that it was used to keep the ionic strength at the desired level. When NaCl was added into Sn(ClO₄)₂ solutions, isomer shift did not change, but quadrupole splitting did. Therefore, stability constants were calculated from the quadrupole splitting data, which were in good agreement with the published values. The observed spectra could be resolved into components.     

Automatic absolute determination of total sulfur by combustion and coulometric titration

An automatic method for the absolute determination of total sulfur, which overcomes the tendency of previous methods to give low sulfur recoveries on samples with high oxygen demand, is described. The technique combusts the sample in oxygen and sweeps the combustion gases through a hydrogen peroxide trap to remove the sulfur oxides from the gas stream. The trap contents are volatilized and swept with nitrogen through reduced copper heated to 890 °C. Sulfur oxides are quantitatively converted to sulfur dioxide and titrated coulometrically with iodine. Halogens are held by the copper and do not interfere with the sulfur determination. Analysis time is 12 min and automation of the method allows unattended operation. Sample size is 0.5 to 20 mg. Sulfur levels of 0.05 to 100% can be determined.     

Parallel mixing of photolithographically defined nanoliter volumes using elastomeric microvalve arrays

Portable microfluidic systems provide simple and effective solutions for low-cost point-of-care diagnostics and high-throughput biomedical assays. Robust flow control and precise fluidic volumes are two critical requirements for these applications. We have developed a monolithic polydimethylsiloxane (PDMS) microdevice that allows for storing and mixing subnanoliter volumes of aqueous solutions at various mixing ratios. Filling and mixing is controlled via two integrated PDMS microvalve arrays. The volumes of the microchambers are entirely defined by photolithography, hence volumes from picoliter to nanoliter can be fabricated with high precision. Because the microvalves do not require an energy input to stay closed, fluid can be stored in a highly portable fashion for several days. We have confirmed the mixing precision and predictability using fluorescence microscopy. We also demonstrate the application of the device for calibrating fluorescent calcium indicators. Due to the biocompatibility of PDMS, the device will have broad applications in miniaturized diagnostic assays as well as basic biological studies.     

Spectroscopic properties of tyrosyl radicals in dipeptides

Redox-active tyrosine residues play important roles in long-distance electron reactions in enzymes, including prostaglandin H synthase, galactose oxidase, ribonucleotide reductase, and photosystem II. Magnetic resonance and vibrational spectroscopy provide methods with which to study the structures of redox-active amino acids in proteins. In this report, ultraviolet photolysis was used to generate tyrosyl radicals from polycrystalline tyrosinate or dipeptides, and the structure of the radical was investigated with EPR and reaction-induced FT-IR spectroscopy at 77 K. Photolysis at 77 K is expected to generate a neutral tyrosyl radical through oxidation of the aromatic ring. EPR and FT-IR results obtained from (13)C-labeled tyrosine were consistent with that expectation. Surprisingly, labeling of the tyrosyl amino group with (15)N also resulted in isotope-shifted bands in the photolysis spectrum. The force constant of a NH deformation mode increased when the tyrosyl radical was generated. These data suggest an interaction between the pi system of the tyrosyl radical and the amino group. In spectra acquired from the dipeptides, evidence for a sequence-dependent interaction between the tyrosyl radical and the amide bond of the dipeptide was also obtained. We postulate that perturbation of the amino or the amide/imide groups may occur through a spin polarization mechanism, which is indirectly detected as a change in NH force constant. This conclusion is supported by density functional calculations, which suggest a conformationally sensitive delocalization of spin density onto the amino and carboxylate groups of the tyrosyl radical. These experiments provide a step toward a detailed spectral interpretation for protein-based tyrosyl radicals.