An SPE column-teflon sleeve assembly for in-line retention during supercritical fluid extraction of analytes from biological matrices

An in-line collector assembly designed for use in supercritical fluid extraction (SFE) vessels is described. This assembly enables solutes extracted by supercritical fluids (SF) to be retained in-line on standard solid phase extraction (SPE) columns. The assembly consists of a standard 1 mL or 3 mL SPE column fitted into a specially fabricated Teflon© sleeve. The SPE column-Teflon sleeve assembly is inserted into the SFE vessel followed by the sample matrix. This unit forms a leak-tight seal with the vessel's end-cap up to pressures of 680 bar. The choice of sorbents used in the in-line SPE columns is dependent upon the properties of the solute in the supercritical state. After SFE is completed, the SPE column is removed and the solutes are recovered in 1–2 mL of the eluting solvent. No further clean-up is normally required prior to chromatographic analysis of the analyte. A comparison was made of recoveries by in-line and off-line (after SF decompression) techniques for the SFE of three anabolic steroids in fortified chicken liver. The HPLC chromatograms of the steroids from the off-line SPE columns were too complex for quantitation, because of coeluting artifacts, whereas chromatograms obtained from in-line SPE columns were free from UV-absorbing interferences and were easily quantified.     

Rapid determination of actinides in emergency food samples

A new rapid method for the determination of actinides in food samples has been developed at the Savannah River Site Environmental Lab (Aiken, SC, USA) that can be used for emergency response or routine food samples. If a radiological dispersive device or improvised nuclear device event occurs, there will be a urgent need for rapid analyzes of many different environmental matrices, as well as food samples, to support dose mitigation and protect general populations from radioactivity that may enter the food chain. The recent accident at Fukushima nuclear power plant in March, 2011 reinforces the need to have rapid analyzes for radionuclides in environmental and food samples. The new method to determine actinides in food samples utilizes a furnace ashing step, a rapid sodium hydroxide fusion method, a lanthanum fluoride matrix removal step, and a column separation process with stacked TEVA, TRU, and DGA resin cartridges. The furnace ashing and rapid fusion steps are performed in relatively inexpensive, reusable zirconium crucibles. Alpha emitters are prepared using rare earth micro precipitation for counting by alpha spectrometry. The method showed high chemical recoveries and effective removal of interferences. The determination of actinides in food samples can be performed in less than 8 h for 10 g samples with excellent quality for emergency samples using short count times. Larger food samples (100 g) may be processed in 24 h or less. The rapid fusion technique is a rugged sample digestion method that ensures that any refractory actinide particles are effectively digested. This method can be used to meet the derived intervention level guidelines recommended by the U.S. Food and Drug Administrations.     

A semipermanent coating for preventing protein adsorption at physiological pH in kinetic capillary electrophoresis

Protein adsorption to the inner capillary wall hinders the use of kinetic capillary electrophoresis (KCE) when studying noncovalent protein-ligand interactions. Permanent and dynamic capillary coatings have been previously reported to alleviate much of the problems associated with protein adsorption. The characteristic limitations associated with permanent and dynamic coatings motivated us to look at a third type of coating - semipermanent. Here, we demonstrate that a semipermanent capillary coating, designed by Lucy and co-workers, comprised of dioctadecyldimethylammonium bromide (DODAB) and polyoxyethylene (POE) stearate, greatly reduces protein adsorption at physiological pH - a necessary requirement for KCE. The coating (i) does not inhibit protein-DNA complex formation, (ii) prevents the adsorption of the analytes, and (iii) supports an electoosmotic flow required for many applications of KCE. The coating was tested in three physiological buffers using a well-known DNA aptamer and four proteins that severely bind to bare silica capillaries as standards. For every protein, a condition was found under which the semipermanent coating effectively suppresses protein adhesion. While no coating can completely prevent the adsorption of all proteins, our findings suggest that the DODAB/POE stearate coating can have a broad impact on CE at large, as it prevents the absorption of several well studied, highly adhesive proteins at physiological pH.     

Investigating the solid state hosting abilities of homo- and hetero-valent [Co7] metallocalix[6]arenes

A family of homo-valent [Co(II)(7)(OH)(6)(L(1))(6)](NO(3))(2) (1), [(MeOH)(2) is a subset of Co(II)(7)(OH)(6)(L(1))(6)](NO(3))(2) (2) (where L(1)H = 2-iminomethyl-6-methoxyphenol) and hetero-valent [(NO(3))(2) is a subset of Co(III)Co(II)(6)(OH)(6)(L(2))(6)](NO(3))·3MeCN (4) (where L(2)H = 2-iminophenyl-6-methoxyphenol) complexes possess metallic skeletons describing planar hexagonal discs. Their organic exteriors form double-bowl shaped topologies, and coupled with their 3-D connectivity, this results in the formation of molecular cavities in the solid state. These confined spaces are shown to behave as host units in the solid state for guests including solvent molecules and charge balancing counter anions. Magnetic susceptibility measurements on 2 and 4 reveal weak ferro- and ferrimagnetism, respectively. The utilisation of other Co(II) salt precursors gives rise to entirely different species including the mononuclear and trinuclear complexes [Co(II)(L(2))(2)] (5) and [Co(III)(2)Na(I)(1)(L(3))(6)](BF(4)) (6) (where L(3)H = 2-iminomethyl-4-bromo-6-methoxyphenol).     

A miniaturized and integrated gel post platform for multiparameter PCR detection of herpes simplex viruses from raw genital swabs

Herpes simplex virus (HSV) is one of the most prevalent viruses, with acute and recurrent infections in humans. The current gold standard for the diagnosis of HSV is viral culture which takes 2-14 days and has low sensitivity. In contrast, DNA amplification by polymerase chain reaction (PCR) can be performed within 1-2 h. We here describe a multiparameter PCR assay to simultaneously detect HSV-1 and HSV-2 DNA templates, together with integrated positive and negative controls, with product detection by melting curve analysis (MCA), in an array of semi-solid polyacrylamide gel posts. Each gel post is 0.67 μL in volume, and polymerized with all the components required for PCR. Both PCR and MCA can currently be performed in one hour and 20 min. Unprocessed genital swabs collected in universal transport medium were directly added to the reagents before or after polymerization, diffusing from atop the gel posts. The gel post platform detects HSV templates in as little as 2.5 nL of raw sample. In this study, 45 genital swab specimens were tested blindly as a preliminary validation of this platform. The concordance of PCR on gel posts with conventional PCR was 91%. The primer sequestration method introduced here (wherein different primers are placed in different sets of posts) enables the simultaneous detection of multiple pathogens for the same sample, together with positive and negative controls, on a single chip. This platform accepts unprocessed samples and is readily adaptable to detection of multiple different pathogens or biomarkers for point-of-care diagnostics.     

Synthesis of 4-substituted chlorophthalazines, dihydrobenzoazepinediones, 2-pyrazolylbenzoic acid, and 2-pyrazolylbenzohydrazide via 3-substituted 3-hydroxyisoindolin-1-ones

Herein we describe a general three-step synthesis of 4-substituted chlorophthalazines in good overall yields. In the key step, N,N-dimethylaminophthalimide (8a) directs the selective monoaddition of alkyl, aryl, and heteroaryl organometallic reagents to afford 3-substituted 3-hydroxyisoindolinones 9b, 9i-9am. Many of these hydroxyisoindolinones are converted to chlorophthalazines 1b-1v via reaction with hydrazine, followed by chlorination with POCl(3). We have also discovered two novel transformations of 3-vinyl- and 3-alkynyl-3-hydroxyisoindolinones. Addition of vinyl organometallic reagents to N,N-dimethylaminophthalimide (8a) provided dihydrobenzoazepinediones 15a-15c via the proposed ring expansion of 3-vinyl-3-hydroxyisoindolinone intermediates. 3-Alkynyl-3-hydroxyisoindolinones react with hydrazine and substituted hydrazines to afford 2-pyrazolyl benzoic acids 16a-16d and 2-pyrazolyl benzohydrazides 17a-17g rather than the expected alkynyl phthalazinones.     

Fabrication of thermoplastics chips through lamination based techniques

In this work, we propose a novel strategy for the fabrication of flexible thermoplastic microdevices entirely based on lamination processes. The same low-cost laminator apparatus can be used from master fabrication to microchannel sealing. This process is appropriate for rapid prototyping at laboratory scale, but it can also be easily upscaled to industrial manufacturing. For demonstration, we used here Cycloolefin Copolymer (COC), a thermoplastic polymer that is extensively used for microfluidic applications. COC is a thermoplastic polymer with good chemical resistance to common chemicals used in microfluidics such as acids, bases and most polar solvents. Its optical quality and mechanical resistance make this material suitable for a large range of applications in chemistry or biology. As an example, the electrokinetic separation of pollutants is proposed in the present study.     

Solvothermal syntheses of Cu3P via reactions of amorphous red phosphorus with a variety of copper sources

Polycrystalline Cu₃P was successfully prepared under a wide variety of solvothermal conditions. The reaction of red phosphorus with several copper sources (copper metal, copper (I) iodide, copper (I) chloride and copper (II) chloride) at 150-200 °C for 1-2 days in water produced Cu₃P. Products were examined with powder X-ray diffraction, scanning electron microscopy and energy dispersive spectroscopy. Thermal analyses and optical spectroscopy were also performed. A detailed list of reaction conditions, products and impurity phases (where applicable) are reported.     

A quantitative and comprehensive method to analyze human milk oligosaccharide structures in the urine and feces of infants

Human milk oligosaccharides (HMOs), though non-nutritive to the infant, shape the intestinal microbiota and protect against pathogens during early growth and development. Infant formulas with added galacto-oligosaccharides have been developed to mimic the beneficial effects of HMOs. Premature infants have an immature immune system and a leaky gut and are thus highly susceptible to opportunistic infections. A method employing nanoflow liquid chromatography time-of-flight mass spectrometry (MS) is presented to simultaneously identify and quantify HMOs in the feces and urine of infants, of which 75 HMOs have previously been fully structurally elucidated. Matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance MS was employed for high-resolution and rapid compositional profiling. To demonstrate this novel method, samples from mother–infant dyads as well as samples from infants receiving infant formula fortified with dietary galacto-oligosaccharides or probiotic bifidobacteria were analyzed. Ingested oligosaccharides are demonstrated in high abundance in the infant feces and urine. While the method was developed to examine specimens from preterm infants, it is of general utility and can be used to monitor oligosaccharide consumption and utilization in term infants, children, and adults. This method may therefore provide diagnostic and therapeutic opportunities.