Optimizing a High-Throughput Solid-Phase Microextraction System to Determine the Plasma Protein Binding of Drugs in Human Plasma.

Plasma protein binding refers to the binding of a drug to plasma proteins after entering the body. The measurement of plasma protein binding is essential during drug development and in clinical practice, as it provides a more detailed understanding of the available free concentration of a drug in the blood, which is in turn critical for pharmacokinetics and pharmacodynamics studies. In addition, the accurate determination of the free concentration of a drug in the blood is also highly important for therapeutic drug monitoring and in personalized medicine.

Modification of polyacrylate sorbent coatings with carbodiimide crosslinker chemistry for sequence-selective DNA extraction using solid-phase microextraction.

Selective DNA extraction is immensely useful for the isolation and detection of low-abundance sequences. Oligonucleotide-modified substrates are often used to capture sequences of interest for downstream analysis. In this study, we explore the chemical modification of commercial-available polyacrylate solid-phase microextraction fibers for selective DNA analysis using carbodiimide crosslinker chemistry.

Analysis of polynuclear aromatic hydrocarbons in olive oil after solid-phase extraction using a dual-layer sorbent cartridge followed by high-performance liquid chromatography with fluorescence detection.

A simple and easy direct solid-phase extraction (SPE) method was developed for the analysis of polynuclear aromatic hydrocarbons (PAHs) in olive oil using a dual-layer cartridge containing activated Florisil and a mixture of octadecyl (C18)-bonded and zirconia-coated silicas. Undiluted olive oil was applied directly to the SPE cartridge, and the sample was eluted with acetonitrile solvent. Background in the extract was found to be low enough for either gas chromatography-mass spectrometry (GC-MS) or high-performance liquid chromatography with fluorescence detection (HPLC-FLD) analysis.

Nontargeted analysis of DNA adducts by mass-tag MS: reaction of p-benzoquinone with DNA.

Using a method in which DNA adducts are discovered based on their conversion in a nucleotide form to phosphorimidazolides with isotopologue benzoylhistamines (or p-bromobenzoylhistamine) prior to detection by MALDI-TOF-MS, we have profiled the adducts that form when calf thymus DNA is reacted in vitro with p-benzoquinone (BQ). We find, as relative values normalized to 100% of adducts observed, 79% BQ-dCMP, 21% BQ-methyl-dCMP (a new DNA adduct), and trace amounts of BQ-dAMP and BQ-dGMP. Because mC is 5% of C in this DNA, the reaction of BQ with DNA in vitro is about five times faster at methyl-C than C.

Novel pre-fractionation method of trans fatty acids by gas chromatography with silver-ion cartridge column.

We developed a novel pre-separation method of trans fatty acids (TFAs) using a silver-ion cartridge column and GC. As a preliminary study, a mixture of fatty acid methyl esters consisting of saturated, cis-unsaturated, and trans-unsaturated fatty acids was dissolved in dichloromethane and loaded onto a Bond Elut SCX ion-exchange cartridge column that was converted to the silver-ion form. The column was then eluted with dichloromethane to obtain the saturated fatty acids, dichloromethane/ethyl acetate (90/10) for the trans mono-ene, dichloromethane/ethyl acetate (65/35) for the cis mono-ene, dichloromethane/acetone (60/40) for the trans di-ene, and acetone/acetonitrile (80/20) for the others.

Determination of malachite green residues in fish using molecularly imprinted solid-phase extraction followed by liquid chromatography-linear ion trap mass spectrometry.

This paper describes the development of an analytical procedure to determine malachite green (MG) residues in salmon samples using molecularly imprinted polymers (MIPs) as the extraction and clean-up material, followed by liquid chromatography-linear ion trap mass spectrometry (LC-QqQLIT-MS/MS). MG and two structurally related compounds, crystal violet (CV) and brilliant green (BG) were employed for the selectivity test. The imprinted polymers exhibited high binding affinity for MG, while CV and BG showed less binding capacity: 47% and 34%, respectively.

Development of an improved method for trace analysis of chloramphenicol using molecularly imprinted polymers.

A confirmatory method is described for the determination of the illegal antibiotic chloramphenicol using a specifically developed molecularly imprinted polymer (MIP) as the sample clean-up technique. The newly developed MIP was produced using an analogue to chloramphenicol as the template molecule. Using an analogue of the analyte as the template avoids a major traditional drawback associated with MIPs of residual template leeching or bleeding.

Evaluation of a solid-phase extraction dual-layer carbon/primary secondary amine for clean-up of fatty acid matrix components from food extracts in multiresidue pesticide analysis.

The use of dual-layer solid-phase extraction (SPE), a primary-secondary amine (PSA) in combination with graphitized carbon black (GCB), was evaluated for sample clean-up during multiresidue pesticide screening of agricultural and food products. The retention of fatty acids by the PSA sorbent was quantified and the effect of the elution solvent on the retention of fatty acid on the SPE cartridge was evaluated. The use of stronger elution solvents to elute certain pesticides from graphitized carbon was shown to interfere with the capacity of PSA to bind fatty acids.

Nuclease P1 digestion/high-performance liquid chromatography, a practical method for DNA quantitation.

We have developed a practical method for quantifying DNA. The method is practical in two ways. First, a single enzyme is used to digest the DNA to nucleotides that are then quantified by HPLC under ordinary conditions.

Phenolic extraction of DNA from mammalian tissues and conversion to deoxyribonucleoside-5'-monophosphates devoid of ribonucleotides.

Towards a goal of detecting scaled-up DNA adducts as altered deoxynucleotides by mass spectrometry, we have set up a practical and general method for isolating DNA-derived deoxyribonucleoside-5'-monophosphates devoid of ribonucleotides starting with a 1 g sample of mammalian tissue. The method is practical because costs have been minimized, and it is general because it can be applied to a more difficult sample such as mouse skin or non-fresh calf liver. The procedure, consisting of a series of steps that were largely gleaned and tuned from prior literature, proceeds as follows: (1) homogenize the tissue in sodium dodecyl sulfate; (2) digest with ribonuclease A, ribonuclease TI, alpha-amylase and proteinase K; (3) partition between water and phenol; (4) precipitate the DNA with ethanol followed by redissolving and dialysis; and (5) digest with nuclease P1 and phosphodiesterase I followed by ultrafiltration and boric acid gel chromatography.

Handling and detection of 25 amol of near-infrared dye deoxynucleotide conjugates by capillary electrophoresis with laser-induced fluorescence detection.

Near-infrared dyes are attractive as labeling reagents to enhance sensitivity in trace analysis largely because background fluorescence is low in this spectral region. Here we demonstrate, towards a goal of detecting DNA adducts in small biological samples, that some near-infrared (IR) dye-labeled deoxynucleotides can be separated and detected with high sensitivity by capillary electrophoresis (CE)-laser-induced fluorescence detection (LIF) in a realistic way (handling detection limit of 25 amol) for near-IR dye-labeled deoxynucleotides. This detection limit is achieved by polarity-switching injection of 2.

Handling and detection of 0.8 amol of a near-infrared cyanine dye by capillary electrophoresis with laser-induced fluorescence detection.

We are interested in the detection of DNA adducts and other trace analytes by labeling them with a fluorescent tag followed by use of capillary electrophoresis with laser-induced fluorescence detection (CE-LIF) for high resolution and sensitivity. Towards this goal, here we report the following: (1) synthesis and handling properties of a near-IR, carboxyl-substituted heptamethine cyanine dye; (2) modification of an existing ball lens LIF detector to provide near-LIF detection with excitation at 785 nm for CE; and (3) corresponding handling and detection of as little as 0.8 amol of the dye by enrich-injection of 4.

Monensin mediates a rapid and selective transport of Pb(2+). Possible application of monensin for the treatment of Pb(2+) intoxication.

The carboxylic acid ionophore monensin, known as an electroneutral Na(+) ionophore, an anticoccidial agent, and a growth-promoting feed additive in agriculture, is shown to be highly efficient as an ionophore for Pb(2+) and to be highly selective for Pb(2+) compared with other divalent cations. Monensin transports Pb(2+) by an electroneutral mechanism in which the complex PbMonOH is the transporting species. Electrogenic transport via the species PbMon(+) may also be possible.