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Biomedical subjects

Stephen Ferguson

Publications and source records attributed to Stephen Ferguson.

7 recordsLinked to original sources
PubMed2006

Comparison of supercritical fluid chromatography and liquid chromatography for the separation of urinary metabolites of nobiletin with chiral and non-chiral stationary phases.

Nobiletin (NOB), a polymethoxylated flavone found in sweet orange (Citrus sinensis) peel, is currently recognized as a promising anti-inflammatory and anti-tumor agent. It is believed that, by undergoing metabolic biotransformation in vivo, nobiletin is demethylated by hepatic P450 enzymes, yielding multiple hydroxylated metabolites. However, it has not been possible to date to separate the two demethylated nobiletin metabolites, 3'-demethyl-NOB and 4'-demethyl-NOB (regio-isomers) on reversed-phase liquid chromatography (RPLC). Additionally, both display similar mass spectrometric fragmentation, resulting in difficulties to identify the dominant metabolite. A successful separation method was developed by utilizing supercritical fluid chromatography (SFC) with chiral stationary phase. The separation was also attempted with normal-phase liquid chromatography (NPLC) in both chiral and non-chiral modes. Chromatographic separation for the two nobiletin metabolites was superior by SFC than by LC, especially using chiral stationary phase. By comparing the SFC profile of the synthesized standards, the major nobiletin metabolite in mouse urine was identified as 4'-demethyl-NOB, with the concentration of 28.9 microg/mL.

Animals↗
PubMed2003

A novel distal enhancer module regulated by pregnane X receptor/constitutive androstane receptor is essential for the maximal induction of CYP2B6 gene expression.

CYP2B6 plays an important role in the metabolism of a variety of structurally unrelated xenobiotics, including the anticancer drugs cyclophosphamide and ifosfamide. Previous studies have shown that the nuclear receptors constitutive androstane receptor (CAR) and pregnane X receptor (PXR) are involved in the transcriptional regulation of CYP2B genes through the phenobarbital-responsive enhancer module (PBREM). However, for human CYP2B6 the relatively weak response of the PBREM to PXR and CAR activation in transfection assays fails to describe the potent induction observed in primary human hepatocyte cultures. In this report, a novel nuclear receptor response module located -8.5 kilobases upstream from the CYP2B6 encoding region is described. Several potential PXR/CAR binding motifs were identified within the distal regulatory cluster. In electrophoretic mobility shift assays, one DR4 motif showed the strongest binding to both PXR and CAR. Transient transfection assays in HepG2 cells demonstrated that the novel distal response cluster could be activated by PXR and CAR. In primary human hepatocytes, both PBREM and the distal responsive element were activated individually by endogenous nuclear receptors upon exposure to prototypical inducers. However, in both HepG2 cells and primary human hepatocytes maximal reporter activation was observed in a construct containing both PBREM and the distal responsive element. In mouse tail-vein injection experiments, a construct containing both the distal responsive element and the proximal PBREM exhibited a strong synergistic expression in phenobarbital-treated mice. These results show that a novel xenobiotic-responsive enhancer module in the distal region of the CYP2B6 promoter (CYP2B6-XREM) together with the PBREM mediates optimal drug-induced expression of CYP2B6.

Amino Acid Motifs↗
PubMed2003

Development of a high-volume concentrated ambient particles system (CAPS) for human and animal inhalation toxicological studies.

A two-stage, high-volume, ambient particle concentrator was developed and characterized. This versatile system, depending on its operational parameters, can be used to fractionate and concentrate particles in three size ranges (PM(10-2.5), PM(10-1), PM(2.5-1)). The performance of this concentrated ambient particle system (CAPS), as well as its individual virtual impaction stages, was investigated as a function of several parameters, including minor-to-total flow ratios and acceleration nozzle Reynolds number. During these laboratory tests, performance parameters such as concentration enrichment factor (CF), particle losses, collection efficiency curves, cutpoint, and pressure drop were measured. The main objective of these investigations was to optimize the ability of the system to concentrate ambient PM(2.5-10) and PM(1-10) particles. PM(2.5-10) particles were concentrated by a factor of 70 to 150. The flow rate of the concentrated aerosol can range between 12.5 and 50 LPM (L/min). Other features of the system include relatively low-pressure drops in the major and minor flows, low particle losses, and a compact design. Performance evaluation of the system also confirmed that separation and concentration of the PM(2.5-10) particles occurred without any significant distortion of the size distribution, during the concentration process. Similar results were obtained for the PM(1-10) size range. For this size range, concentration enrichment was 70 times, and again, no particle size distribution distortion was observed. The overall performance of this versatile system makes it suitable for inhalation toxicological studies.

Aerosols↗
PubMed2002

Development and laboratory performance evaluation of a personal cascade impactor.

This paper presents the design and laboratory evaluation of a personal cascade impactor. The system is compact, lightweight, and uses a single battery-operated sampling pump. It operates at a flow rate of 5 L/min and consists of four impaction stages, each equipped with slit-shaped acceleration nozzles, and a backup filter. The impactor was calibrated using polydisperse particles. The 50% cut points of the four stages were 9.6, 2.6, 1.0, and 0.5 microm, respectively. The backup filter is placed downstream of the fourth stage and is used to collect the particles with an aerodynamic diameter smaller than 0.5 microm (dp < 0.5 microm). The major feature of this novel sampler is its ability not only to fractionate the particles with an aerodynamic diameter smaller than 10 microm to the various size fractions, but also to collect them onto relatively small polyurethane foam substrates without using adhesives. Although the impaction substrates are not coated with adhesives such as grease or mineral oil, particle bounce and re-entrainment losses were found to be insignificant. Interstage losses of particles smaller than 0.5 microm were less than 10%; for fine particles, less than 5%; and for coarse particles, less than 12%. The pressure drop across the four stages and the backup filter were 0.015 kPa (0.153 cm H2O),0.025 kPa (0.255 cm H2O), 0.274 kPa (2.794 cm H2O), 0.323 kPa (3.294 cm H2O), and 0.370 kPa (3.773 cm H2O), respectively. Particles can be easily recovered from the foam substrates using aqueous extraction.

Air Movements↗
PubMed2002

Identification and functional characterization of new potentially defective alleles of human CYP2C19.

CYP2C19 is a clinically important enzyme responsible for the metabolism of a number of therapeutic drugs, such as mephenytoin, omeprazole, diazepam, proguanil, propranolol and certain antidepressants. Genetic polymorphisms in this enzyme result in poor metabolizers of these drugs. There are racial differences in the incidence of the poor metabolizer trait, which represents 13-23% of Asians but only 3-5% of Caucasians. In this study, single nucleotide polymorphisms (SNPs) in CYP2C19 were identified by direct sequencing of genomic DNA from 92 individuals from three different racial groups of varied ethnic background, including Caucasians, Asians and blacks. Several new alleles were identified containing the coding changes Arg114 His (CYP2C19*9), Pro227 Leu (CYP2C19*10), Arg150 His (CYP2C19*11), stop491 Cys (CYP2C19*12), Arg410 Cys (CYP2C19*13), Leu17 Pro (CYP2C19*14) and Ile19 Leu (CYP2C19*15). When expressed in a bacterial cDNA expression system, CYP2C19*9 exhibited a modest decrease in the V(max) for 4'-hydroxylation of -mephenytoin, and no alteration in its affinity for reductase. CYP2C19*10 exhibited a dramatically higher K(m) and lower V(max) for mephenytoin. CYP2C19*12was unstable and expressed poorly in a bacterial cDNA expression system. Clinical studies will be required to confirm whether this allele is defective in vivo. CYP2C19*9, CYP2C19*10 and CYP2C19*12 all occurred in African-Americans, or individuals of African descent, and represent new potentially defective alleles of CYP2C19 which are predicted to alter risk of these populations to clinically important drugs.

Alleles↗