Observing alteration of restriction enzyme activity has been employed frequently to determine the sequence specificity of the binding of many types of molecules to DNAs. Generally, these studies employed restriction enzymes which cut the target DNA several times. The effects of binding to sequences flanking the restriction enzyme cleavage sites may have been obscured. In this study, we report on restriction enzyme activity assays of the binding of the intercalators actinomycin D, ametantrone and ethidium, the groove binder netropsin, and the covalent binding cisplatin to a mixture of supercoiled and relaxed phiX 174 RF DNA using restriction enzymes which cleave this DNA once or twice. Sequence selectivities and topological selectivities were observed for these ligands. In some cases restriction enzymes not containing the reported preferred binding sites had altered activities, suggesting binding to flanking sequences affects activity in neighboring DNA sequences.

This chapter presents the role of cation-chloride transporters in brain ischemia. Ischemic stroke is the inevitable consequence of transient or permanent reduction of regional cerebral blood flow to brain tissue. Because of the high energy demand of brain tissue, a reduction of blood flow below ∼50% will cause serious perturbations in tissue metabolism. The resulting tissue damage will depend on the severity and length of the brain perfusion deficit. The brain damage in the infarct core is considered to be irreversible in focal stroke. Within minutes, there is a complete collapse of energy production, dissipation of ion gradients, cessation of macromolecular synthesis and loss of cell structure. On the other hand, the penumbra, which surrounds the infarct core, retains residual blood flow and can remain viable for hours or even days. The penumbra tissue is the target for therapeutic intervention.

Background and Objectives: Approximately 10 years ago, “bath salts” became popular as legal alternatives to the psychostimulants cocaine and the amphetamines. These products contained synthetic cathinones, including 3,4-methylenedioxypyrovalerone (MDPV), 4-methylmethcathinone (mephedrone), and 3,4-methylenedioxymethcathinone (methylone). Most preclinical investigations have only assessed the effects of these synthetic cathinones independently; however, case reports and Drug Enforcement Administration (DEA) studies indicate that bath salts contain mixtures of these substances. In this study, we examine the pharmacokinetic interactions of the drug combination. We hypothesized that combined exposure to MDPV, mephedrone, and methylone would result in increased drug concentrations and enhanced total drug concentrations when compared to individual administration. Methods: Adolescent male Swiss–Webster mice were injected intraperitoneally with either 10 mg/kg MDPV, 10 mg/kg mephedrone, 10 mg/kg methylone, or 10 mg/kg combined MDPV, mephedrone, and methylone. Following injection, brains and plasma were collected at 1, 10, 15, 30, 60, and 120 min. Drugs were extracted via solid-phase extraction, and concentrations were determined using a previously published high-pressure liquid chromatography–tandem mass spectrometry (HPLC-MS/MS) method. Results: All drugs crossed the blood–brain barrier quickly. For methylone, the maximal concentration (Cmax) and the total drug exposure [as represented by the area under the concentration-time curve (AUC)] were significantly higher when combined with mephedrone and MDPV in both matrices (2.89-fold increase for both Cmax and AUC with combined treatment). For mephedrone, the Cmax was unchanged, but the AUC in brain was increased when in combination by approximately 34%. Interestingly, for MDPV, the Cmax was unchanged, yet the AUC was higher when MDPV was administered individually (there was a 62% decrease in AUC with combined treatment). Conclusions: The pharmacokinetics of methylone, mepedrone, and MDPV are altered when the drugs are used in combination. These data provide insight into the consequences of co-exposure to synthetic cathinones in popular bath salt products.

No description available.

A rapid, sensitive, and reproducible assay is described for the quantitative determination of the monoamine neurotransmitters dopamine, norepinephrine and serotonin, their metabolites, and the internal standard 3,4-dihydroxybenzlyamine hydro-bromide in mouse brain homogenate using high-performance liquid chromatography with electrochemical detection. The method was validated in the following brain areas: frontal cortex, striatum, nucleus accumbens, hippocampus, substantia nigra pars compacta and ventral tegmental area. Biogenic amines and relevant metabolites were extracted from discrete brain regions using a simple protein precipitation procedure, and the chromatography was achieved using a C18 column. The method was accurate over the linear range of 0.300–30 ng/mL (r = 0.999) for dopamine and 0.300–15 ng/mL (r = 0.999) for norepinephrine, 3,4-dihydroxybenzlyamine hydro-bromide, homovanillic acid and 5-hydroxyindolacetic acid, with detection limits of ~0.125 ng/mL (5 pg on column) for each of these analytes. Accuracy and linearity for serotonin were observed throughout the concentration range of 0.625–30 ng/mL (r = 0.998) with an analytical detection limit of ~0.300 ng/mL (12 pg on column). Relative recoveries for all analytes were approximately ≥90% and the analytical run time was <10 min. The described method utilized minimal sample preparation procedures and was optimized to provide the sensitivity limits required for simultaneous monoamine and metabolite analysis in small, discrete brain tissue samples.

No description available.

Drug based nanoparticle (NP) formulations have gained considerable attention over the past decade for their use in various drug formulations. NPs have been shown to increase bioavailability, decrease side effects of highly toxic drugs, and prolong drug release. Nonsteroidal anti-inflammatory drugs such as diclofenac block cyclooxygenase expression and reduce prostaglandin synthesis, which can lead to several side effects such as gastrointestinal bleeding and renal insufficiency. The aim of this study was to formulate and characterize diclofenac entrapped poly(lactide-co-glycolide) (PLGA) based nanoparticles. Nanoparticles were formulated using an emulsion-diffusion- evaporation technique with varying concentrations of poly vinyl alcohol (PVA) (0.1, 0.25, 0.5, or 1%) or didodecyldimethylammonium bromide (DMAB) (0.1, 0.25, 0.5, 0.75, or 1%) stabilizers centrifuged at 8,800 rpm or 12,000 rpm. The resultant nanoparticles were evaluated based on particle size, zeta potential, and entrapment efficacy. DMAB formulated NPs showed the lowest particle size (108±2.1 nm) and highest zeta potential (-27.71±0.6 mV) at 0.1 and 0.25% respectively, after centrifugation at 12,000 rpm. Results of the PVA based NP formulation showed the smallest particle size (92.4±7.6 nm) and highest zeta potential (-11.14±0.5 mV) at 0.25% and 1% w/v, respectively, after centrifugation at 12,000 rpm. Drug entrapment reached 77.3±3.5% and 80.2±1.2% efficiency with DMAB and PVA formulations, respectively. The results of our study indicate the use of DMAB for increased nanoparticle stability during formulation. Our study supports the effective utilization of PLGA based nanoparticle formulation for diclofenac.

No description available.

Methamphetamine is the most common amphetamine used and, along with 3,4-methylenedioxymethamphetamine (MDMA, Ecstasy), is considered part of a worldwide drug epidemic. Monitoring metham-phetamine levels in the body is important for purposes of drug screening for employment, criminal investigations, and therapeutic drug monitoring. While methamphetamine is suitable for detection using immunoassay techniques, these methods tend to have significant cross reactivity with other compounds. Over the last decade, more than eighty different quantitative, confirmatory analytical methods for measuring methamphetamine in biological samples have been published in the scientific literature. Analytical instrumentation used in these methods includes gas chromatography-mass spectrometry (GC-MS), liquid chromatography-mass spectrometry (LC-MS), capillary electrophoresis (CE), among others. These assays are capable of quantifying methamphetamine concentrations in a variety of biological matrices, including blood, plasma, urine, hair, and fingernails. Some of these techniques can achieve detection as low as 0.1 ng/mL (1 ppb) concentra-tions. The strengths and limitations of these methodologies will be discussed in the context of methamphetamine analysis. Additionally, methods that can simultaneously measure methamphetamine levels as well as metabolites and other drugs of abuse will be highlighted.

The present work focuses on modifying a non-steroid anti-inflammatory drug (NSAID) into an ionic liquid and evaluating the resulting thermal behavior and structural changes of the drug. Naproxen was chosen as the NSAID molecule due to thermal stability and limited examples of its use as an ionic liquid in current literature. Lidocaine was chosen as the counterion based on a screening study of potential ionic liquid formers. The screening included both potential protic and aprotic formation and counterions were included with consideration to pKa, hydrogen bonding ability, molecular size, diffuse charge distribution and functional groups. Analytical techniques used to evaluate the counterions included high performance liquid chromatography (HPLC), differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA). Naproxen and lidocaine were then combined in varying molar ratios to determine the thermal behavior of the mixtures. The samples with equimolar, or higher, ratio of naproxen showed a phase which had a melting point of 82–85 °C. The DSC data was analyzed using a modified Tamman plot, resulting in the unexpected and previously unreported behavior of ionic liquid formation at a 2:1 molar ratio of naproxen to lidocaine, referred to as IL1 in this research. This stoichiometry was confirmed through Fourier Transformed Infrared (FT-IR) and nuclear magnetic resonance (NMR) spectroscopy methods. The samples that contained a higher molar ratio of lidocaine than naproxen, resulted in material more consistent with higher-order complex clusters. Further characterization of IL1 found that the material demonstrated behaviors of an ionic liquid, including weak intermolecular forces and at least partial ionization of the drug and counterion.