Studies on the membrane interactions of the cyclotides kalata B1 and kalata B6 on model membrane systems by surface plasmon resonance

In this study we have demonstrated the interactions of kalata B1 and its naturally occurring analogue kalata B6 with five model lipid membranes and have analyzed the binding kinetics using surface plasmon resonance. Two kalata peptides showed a higher affinity for the phosphatidylethanolamine-containing membranes, indicating that the peptides would bind selectively to bacterial membranes. Also we have optimized the procedure for the immobilization of five liposome mixtures and have shown that the procedure provides reproducible levels of immobilized liposomes and could be used to screen the selective binding of putative antimicrobial peptides to model mammalian or microbial phospholipid membranes. (C) 2004 Elsevier Inc. All rights reserved.

Measurement of free drug and clinical end-point by high-performance liquid chromatography-mass spectrometry: Applications and implications for pharmacokinetic and pharmacodynamic studies

Free drug measurement and pharmacodymanic markers provide the opportunity for a better understanding of drug efficacy and toxicity. High-performance liquid chromatography (HPLC)-mass spectrometry (MS) is a powerful analytical technique that could facilitate the measurement of free drug and these markers. Currently, there are very few published methods for the determination of free drug concentrations by HPLC-MS. The development of atmospheric pressure ionisation sources, together with on-line microdialysis or on-line equilibrium dialysis and column switching techniques have reduced sample run times and increased assay efficiency. The availability of such methods will aid in drug development and the clinical use of certain drugs, including anti-convulsants, anti-arrhythmics, immunosuppressants, local anaesthetics, anti-fungals and protease inhibitors. The history of free drug measurement and an overview of the current HPLC-MS applications for these drugs are discussed. Immunosuppressant drugs are used as an example for the application of HPLC-MS in the measurement of drug pharmacodynamics. Potential biomarkers of immunosuppression that could be measured by HPLC-MS include purine nucleoside/nucleotides, drug-protein complexes and phosphorylated peptides. At the proteomic level, two-dimensional gel electrophoresis combined with matrix-assisted laser desorption/ionisation time-of-flight (TOF) MS is a powerful tool for identifying proteins involved in the response to inflammatory mediators. (C) 2003 Elsevier Science B.V. All rights reserved.

Lipid and carbohydrate based adjuvant/carriers in immunology

This review discusses various issues regarding vaccines:what are they and how they work, safety aspects, the role of adjuvants and carriers in vaccination, synthetic peptides as immunogens, and new technologies for vaccine development and delivery including the identification of novel adjuvants for mucosal vaccine delivery. There has been a recent increase of interest, in the use of lipids and carbohydrates as adjuvants, and so a particular emphasis is placed on adjuvants derived from lipids or carbohydrates, or from both. Copyright (C) 2003 European Peptide Society and John Wiley Sons, Ltd.

Miscibility and phase separation in blends of phenolphthalein poly(aryl ether ketone) and poly(ethylene oxide): a differential scanning calorimetric study

Miscibility and phase separation in the blends of phenolphthalein poly(aryl ether ketone) (PPAEK) and poly(ethylene oxide) (PEO) were investigated by means of differential scanning calorimetry (DSC). The PPAEK/PEO blends prepared by solution casting from N,N-dimethylformamide (DMF) displayed single composition-dependent glass transition temperatures (T-g), intermediate between those of the pure components, suggesting that the blend system is miscible in the amorphous state at all compositions. All the blends underwent phase separation at higher temperatures and the system exhibited a lower critical solution temperature (LCST) behavior. A step-heating thermal analysis was designed to determine the phase boundaries with DSC. The significant changes in the thermal properties of blends were utilized to judge the mixing status for the blends and the phase diagram was thus established. (C) 2004 Elsevier B.V. All rights reserved.

Simultaneous liquid chromatographic determination of doxorubicin and its major metabolite doxorubicinol in parrot plasma

A new microscale method is reported for the determination of doxorubicin and its active metabolite, doxorubicinol, in parrot plasma. Sample workup involved acetonitrile protein precipitation, ethyl acetate extraction, followed by back extraction into HCl. Separations were achieved on a phenyl-hexyl column at 30 degrees C using acetonitrile (17%, v/v) in 0.01 M orthophosphoric acid (83%, v/v) delivered via a linear flow program. Fluorometric detection wavelengths were 235 nm (excitation) and 550 nm (emission). Calibration plots were linear (1 2 > 0.999), and recoveries were 71-87% from 20 to 400 ng/mL. Assay imprecision was

Simultaneous determination of morphine, oxycodone, morphine-3-glucuronide, and noroxycodone concentrations in rat serum by high performance liquid chromatography-electrospray ionization-tandem mass spectrometry

An assay using high performance liquid chromatography (HPLC)-electrospray ionization-tandem mass spectrometry (ESI-MS-MS) was developed for simultaneously determining concentrations of morphine, oxycodone, morphine-3-glucuronide, and noroxycodone, in 50 mul samples of rat serum. Deuterated (d(3)) analogues of each compound were used as internal standards. Samples were treated with acetonitrile to precipitate plasma proteins: acetonitrile was removed from the supernatant by centrifugal evaporation before analysis. Limits of quantitation (ng/ml) and their between-day accuracy and precision (%deviation and %CV) were-morphine, 3.8 (4.3% and 7.6%); morphine-3-glucuronide, 5.0 (4.5% and 2.9%); oxycodone, 4.5 (0.4% and 9.3%); noroxycodone, 5.0 (8.5% and 4.6%). (C) 2004 Elsevier B.V. All rights reserved.

The combined SPE : ToxY-PAM phytotoxicity assay; application and appraisal of a novel biomonitoring tool for the aquatic environment

Mounting concerns regarding the environmental impact of herbicides has meant a growing requirement for accurate, timely information regarding herbicide residue contamination of, in particular, aquatic systems. Conventional methods of detection remain limited in terms of practicality due to high costs of operation and the specialised information that analysis provides. A new phytotoxicity bioassay was trialled for the detection of herbicide residues in filter-purified (Milli-Q) as well as natural waters. The performance of the system, which combines solid-phase extraction (SPE) with the ToxY-PAM dual-channel yield analyser (Heinz Walz GmbH), was tested alongside the traditional method of liquid chromatography-mass spectrometry (LC-MS). The assay methodology was found to be highly sensitive (LOD 0.1 ng L-1 diuron) with good reproducibility. The study showed that the assay protocol is time effective and can be employed for the aquatic screening of herbicide residues in purified as well as natural waters.

A rapid and sensitive microscale HPLC method for the determination of indomethacin in plasma of premature neonates with patent ductus arteriousus

Indomethacin (IND) is the drug of choice for the closure of a patent ductus arteriosus (PDA) in neonates. This paper describes a simple, sensitive, accurate and precise microscale HPLC method suitable for the analysis of IND in plasma of premature neonates. Samples were prepared by plasma protein precipitation with acetonitrile containing the methyl ester of IND as the internal standard (IS). Chromatography was performed on a Hypersil C-18 column. The mobile phase of methanol, water and orthophosphoric acid (70:29.5:0.5, v/v, respectively), was delivered at 1.5 mL/min and monitored at 270 nm. IND and the IS were eluted at 2.9 and 4.3 min, respectively. Calibrations were linear (r > 0.999) from 25 to 2500 mu g/L. The inter- and intra-day assay imprecision was less than 4.3% at 400-2000 mu g/L, and less than 22.1% at 35 mu g/L. Inaccuracy ranged from -6.0% to +1.0% from 35 to 2000 mu g/L. The absolute recovery of IND over this range was 93.0-113.3%. The IS was stable for at least 36 h when added to plasma at ambient temperature. This method is suitable for pharmacokinetic studies of IND and has potential for monitoring therapy in infants with PDA when a target therapeutic range for IND has been validated. (c) 2005 Elsevier B.V. All rights reserved.

Analysis of lipoproteins using 2D diffusion-edited NMR spectroscopy and multi-way chemometrics

This study represents the first application of multi-way calibration by N-PLS and multi-way curve resolution by PARAFAC to 2D diffusion-edited H-1 NMR spectra. The aim of the analysis was to evaluate the potential for quantification of lipoprotein main- and subtractions in human plasma samples. Multi-way N-PLS calibrations relating the methyl and methylene peaks of lipoprotein lipids to concentrations of the four main lipoprotein fractions as well as 11 subfractions were developed with high correlations (R = 0.75-0.98). Furthermore, a PARAFAC model with four chemically meaningful components was calculated from the 2D diffusion-edited spectra of the methylene peak of lipids. Although the four extracted PARAFAC components represent molecules of sizes that correspond to the four main fractions of lipoproteins, the corresponding concentrations of the four PARAFAC components proved not to be correlated to the reference concentrations of these four fractions in the plasma samples as determined by ultracentrifugation. These results indicate that NMR provides complementary information on the classification of lipoprotein fractions compared to ultracentrifugation. (C) 2004 Elsevier B.V. All rights reserved.

Environmental contamination of arsenic and its toxicological impact on humans

Inorganic arsenic compounds are known carcinogens. The human epidemiologic evidence of arsenic-induced skin, lung, and bladder cancers is strong. However, the evidence of arsenic carcinogenicity in animals is very limited. Lack of a suitable animal model until recent years has inhibited studies of the mechanism of arsenic carcinogenesis. The toxicity and bioavailability of arsenic depend on its solubility and chemical forms. Therefore, it is critical to be able to measure arsenic speciation accurately and reliably. However, speciation of arsenic in more complex matrices remains a real challenge. There are tens of millions of people who are being exposed to excessive levels of arsenic in the drinking water alone. The source of contamination is mainly of natural origin and the mass poisoning is occurring worldwide, particularly in developing countries. Chronic arsenicosis resulting in cancer and non-cancer diseases will impact significantly on the public health systems in their respective countries. Effective watershed management and remediation technologies in addition to medical treatment are urgently needed in order to avoid what has been regarded as the largest calamity of chemical poisoning in the world.

Effects of litter and fine root composition on their decomposition in a Rhodic Paleustalf under different land uses

Plant litter and fine roots are important in maintaining soil organic carbon (C) levels as well as for nutrient cycling. The decomposition of surface-placed litter and fine roots of wheat ( Triticum aestivum ), lucerne ( Medicago sativa ), buffel grass ( Cenchrus ciliaris ), and mulga ( Acacia aneura ), placed at 10-cm and 30-cm depths, was studied in the field in a Rhodic Paleustalf. After 2 years, = 60% of mulga roots and twigs remained undecomposed. The rate of decomposition varied from 4.2 year -1 for wheat roots to 0.22 year -1 for mulga twigs, which was significantly correlated with the lignin concentration of both tops and roots. Aryl+O-aryl C concentration, as measured by 13 C nuclear magnetic resonance spectroscopy, was also significantly correlated with the decomposition parameters, although with a lower R 2 value than the lignin concentration. Thus, lignin concentration provides a good predictor of litter and fine root decomposition in the field.