A simple liquid chromatography-tandem mass spectrometry method for determination of plasma fentanyl concentration in rats and patients with cancer pain.

A fentanyl patch is widely used for the treatment of cancer pain. Its few adverse effects include constipation and drowsiness. The absorption volume of transdermally applied fentanyl may differ according to its site of application and variability in patch adhesion. Since fentanyl is predominantly metabolized by the drug-metabolizing enzyme cytochrome P450 (CYP) 3A4 in the liver, its concentration may vary in cases of physiologically reduced CYP3A4 activity in the liver (liver disease and aging) or on co-administration of drugs. The clinical significance of measuring plasma concentration of fentanyl is high, but conventional methods require complicated processes such as solid-phase extraction and liquid-liquid extraction before the sample is injected into an HPLC system. In this study, a simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for determining plasma fentanyl concentrations by deproteinization with acetonitrile. A recovery test was conducted using an absolute calibration curve to confirm the method's linearity and inter- and intra-day reproducibility. The required plasma volume for detection was reduced from 1 mL in the conventional method to 20 µL in the present study, and a good calibration curve was obtained in the concentration range from 0.05 to 5 ng/mL. These findings suggest that the method for sample preparation and quantification developed in this study are appropriate for measuring fentanyl concentration in human plasma in clinical settings.

Single-walled carbon nanotubes downregulate stress-responsive genes in human respiratory tract cells.

Carbon nanotubes (CNTs) are currently key promising materials of nanotechnology. However, elucidation of the possible effects of CNTs on the respiratory tract is urgently needed. The present study aimed to clarify the effect of single-walled CNTs (SWCNTs) on the expression of stress-responsive genes, using primary cultured normal human bronchial epithelial (NHBE), diseased HBE (DHBE) cells, and the human carcinoma cell lines A549 and FaDu. Purified SWCNTs were applied at concentrations of 0.1 or 1.0 mg/mL for 6 h, and a polymerase chain reaction (PCR) array was conducted to examine 84 stress-responsive genes. NHBE cell exposure to SWCNTs resulted in global downregulation of genes involved in inflammation, oxidative stress, and apoptosis. Further analysis using DHBE cells and carcinoma cell lines indicated a similar trend, although differences in sensitivity were observed. Downregulation of stress-responsive genes may be involved in the mechanism by which stress response protects against lung injury.

Mechanism of drug interaction between a Kampo medicine, byakkokaninjinto, and tetracycline in rats.

We have previously reported that concomitant oral administration of the Kampo medicine, byakkokaninjinto (TJ-34), in extract granules, reduced the plasma concentrations of tetracycline (TC) and ciprofloxacin in humans, which might be the result of forming a chelate with Ca(2+). In the present study, we investigated the effect of a chelating agent, ethylenediaminetetraacetic acid (EDTA), on the plasma concentration-time profiles of TC after coadministration of TJ-34 dried extract and TC in rats to clarify whether metal ions contained in the TJ-34 dried extract contribute to this interaction. TJ-34 dried extract significantly reduced the plasma concentration of TC. The values of maximum concentration (C (max)), area under the plasma concentration-time curve and percentage of urinary recovery (f (e)) of TC were reduced to 42%, 40%, and 45%, respectively. On the other hand, treatment with EDTA significantly counteracted the effect of TJ-34 dried extract to reduce absorption of TC, indicating that metal ions mainly account for the interaction. Next, we investigated the effect of staggered administration of TJ-34 dried extract and TC to avoid the drug interaction between them. Administration of TJ-34 dried extract 2 h before TC had no effect on plasma concentrations and pharmacokinetic parameters of TC. These results provide a precise mechanism of the interaction TJ-34 and TC, suggesting a safe and effective dosage regimen to coadminister TJ-34 and TC in clinical use.

Changes in expression of drug-metabolizing enzymes by single-walled carbon nanotubes in human respiratory tract cells.

Single-walled carbon nanotubes (SWCNTs) have attracted attention for biomedical and biotechnological applications, such as drug delivery. However, there are concerns about the safety of SWCNTs for use in humans. To investigate the potential use of SWCNTs for targeted drug delivery to the lung, we examined their effect on drug-metabolizing enzymes in primary normal human bronchial epithelial (NHBE) cells from two donors and the lung carcinoma A549 cell line. Exposure of NHBE and A549 cells to SWCNTs dysregulated some of the important drug-metabolizing enzymes expressed in the human respiratory organs. Exposure of NHBE cells to SWCNTs for 24 h had a pronounced effect on expression of CYP1A1 and CYP1B1 mRNAs, which were reduced to less than 1% of control cells. These effects were also observed in A549 cells. Exposure of A549, HepG2 hepatic carcinoma cells, and MCF-7 breast carcinoma cells to tetrachlorodibenzo-p-dioxin induced the expression and enzymatic activity of CYP1A1 and CYP1B1, which were also suppressed by SWCNTs, suggesting that SWCNTs down-regulated both basal and induced CYP1A1 and CYP1B1 activities. Chromatin immunoprecipitation assays revealed that the down-regulatory effect of SWCNTs may be due to inhibition of activated aryl hydrocarbon receptor binding to the enhancer regions of the CYP1A1 and CYP1B1 genes. Down-regulation of CYP1A1 and CYP1B1 genes by SWCNTs may affect the defense mechanisms by reducing procarcinogen bioactivation in the human lung.

Differential effects of single-walled carbon nanotubes on cell viability of human lung and pharynx carcinoma cell lines.

Carbon nanotubes (CNTs) are attracting significant attention as a novel material for future innovations. Many in vitro studies have assessed the cytotoxicity of CNTs, but the effects of CNTs differ depending on the cell lines and the synthetic method adopted for fabricating CNTs. In the present study, the differential effects of single-walled CNTs (SWCNTs) on the cell viability of A549 cells from human lung carcinomas and FaDu cells from human head and neck carcinomas were investigated. The SWCNTs used in the present study were synthesized with nickel and yttrium (SO-SWCNTs), and iron (FH-P-SWCNTs) as catalysts. Cell viability was evaluated on the basis of cell-membrane biomass, adenosine triphosphate (ATP) content, and intracellular metabolic capacity. After 24-hr exposure of A549 and FaDu cells to 1.0 mg/ml SO-SWCNTs, the cell-membrane biomass of A549 cells decreased to 43% as compared to the control cells, whereas that of FaDu cells remained over 90%. After 24-hr exposure of A549 and FaDu cells to 1.0 mg/ml SO-SWCNT, the intracellular metabolic capacity decreased to 24% and 37%, respectively, and the ATP content decreased to 40% and 54%, respectively. SWCNTs had a greater impact on the viability values of A549 cells than on those of FaDu cells. In addition, cells exposed to FH-P-SWCNTs exhibited a higher viability than those exposed to SO-SWCNTs. Caspase 3/7 activity was not increased at maximum concentration of 1.0 mg/ml SO-SWCNTs. It was surmised that sensitivity to SWCNTs differs among the 2 cell lines; additionally, SWCNT characteristics may produce different effects on these cell lines.

Effect of a Kampo preparation, byakkokaninjinto, on pharmacokinetics of ciprofloxacin and tetracycline.

The effect on the bioavailability of the antimicrobial agents (ciprofloxacin and tetracycline), which are well known to form chelates with cationic metals such as calcium, was evaluated in 20 healthy male volunteers according to an open, random crossover fashion using a Kampo preparation, byakkokaninjinto (TJ-34) which contains various cationic metals including calcium. Each subject received a single oral dose of tetracycline (250 mg) alone or ciprofloxacin (200 mg) alone along with a single coadministration of one pack (3 g) of the Kampo preparation, at one-week intervals. Concentrations of the drugs in plasma and urine were analyzed by HPLC. Concomitant administration of the Kampo preparation significantly decreased the peak plasma concentration (C(max)) and area under the plasma concentration-time curves (AUC), but not time to reach C(max) (T(max)), of ciprofloxacin and tetracycline. However, the decrease in bioavailability of ciprofloxacin was slight (15%) compared with that of tetracycline (30%). The Kampo preparation significantly decreased the urinary recovery of tetracycline, but not ciprofloxacin, and it had no effect on the renal clearance of either ciprofloxacin or tetracycline. These results indicate that the Kampo preparation tested in this study reduces the extent of bioavailability of ciprofloxacin and tetracycline, but not renal excretion, by decreasing the gastrointestinal absorption due to the formation of insoluble chelates with calcium. We recommend that the dose timing of the Kampo preparation should be carefully controlled to avoid therapeutic failure especially for patients receiving the treatment with tetracycline.