Evaluation of Closed System Transfer Devices in Preventing Chemotherapy Agents Contamination During Compounding Process-A Single and Comparative Study in China.

Aim: We performed a comparative study to investigate the efficacy of closed system transfer devices (CSTDs) on the safe handling of injectable hazardous drugs (HDs). Methods: The exposure assessments of cyclophosphamide and cytarabine were performed under traditional or CSTDs. For preparation activity, chemotherapy contamination samples on protective equipment (such as gloves and masks) were collected. The contamination analysis was performed by liquid chromatography with tandem mass spectrometry (LC-MS/MS). A 6-item form was distributed monthly (form M1-M6, total 6 months) to assess the pharmacists' experience on ergonomics, encumbrance, and safety impression. Results: Totally, 96 wiping samples were collected throughout the study. The numbers of contaminated cyclophosphamide samples reduced under CSTD were -37.8, -41.6, -67.7, -47.3, and -22.9% and cytarabine were -12.3, -12.1, -20.6, -69.6, and -56.7% for left countertop, right countertop, medial glass, air-intake vent and door handle, as compared to traditional devices. The reduction was similar to pharmacist devices, i.e., -48.2 and -50.0% for masks and gloves cyclophosphamide contamination, -18.0 and -42.4% for cytarabine. This novel system could improve contamination on dispensing table, transfer container, and dispensing basket by -16.6, -6.0, and -22.3% for cyclophosphamide and -28.5, -22.5, and -46.2% for cytarabine. A high level of satisfaction was consistently associated with ergonomics for CSTD during the compounding process. Meanwhile, a slightly decreased satisfaction on ergonomics, encumbrance, and safety impression was observed for the traditional system between M2 and M3. Conclusion: Closed system transfer devices are offering progressively more effective alternatives to traditional ones and consequently decrease chemotherapy exposure risk on isolator surfaces.

Occupational exposure to antineoplastic drugs: what about hospital sanitation personnel?

OBJECTIVE: Occupational exposure to antineoplastic drugs (ANPs) occurs mainly through dermal contact. Our study was set up to assess the potential exposure of hospital sanitation (HS) personnel, for whom almost no data are available, through contamination of surfaces they regularly touch. METHODS: In the oncology departments of two hospitals around Montreal, surface wipe samples of 120-2000 cm2 were taken at 10 sites cleaned by the HS personnel and five other sites frequently touched by nursing and pharmacy personnel. A few hand wipe samples were collected to explore skin contamination. Wipes were analyzed by ultra-performance liquid chromatography tandem-mass spectrometry for 10 ANPs. RESULTS: Overall, 60.9% of 212 surface samples presented at least one ANP above the limits of detection (LOD). Cyclophosphamide and gemcitabine were most often detected (52% and 31% of samples respectively), followed by 5-fluorouracil and irinotecan (15% each). Highest concentrations of five ANPs were found in outpatient clinics on toilet floors (5-fluorouracil, 49 ng/cm2; irinotecan, 3.6 ng/cm2), a perfusion pump (cyclophosphamide, 19.6 ng/cm2) and on a cytotoxic waste bin cover (gemcitabine, 4.97 ng/cm2). Floors in patient rooms had highest levels of cytarabine (0.12 ng/cm2) and methotrexate (6.38 ng/cm2). Hand wipes were positive for two of 12 samples taken on HS personnel, seven of 18 samples on nurses, and two of 14 samples on pharmacy personnel. CONCLUSIONS: A notable proportion of surfaces showed measurable levels of ANPs, with highest concentrations found on surfaces cleaned by HS personnel, who would benefit from appropriate preventive training. As potential sources of worker exposure, several hospital surfaces need to be regularly monitored to evaluate environmental contamination and efficacy of cleaning.

Development and validation of a liquid chromatography tandem mass spectrometry quantification method for 14 cytotoxic drugs in environmental samples.

RATIONALE: Cytotoxic drug preparation in hospital pharmacies is associated with chronic occupational exposure leading to a risk of adverse effects. The objective was to develop and validate a quantification method for the following cytotoxic drugs in environmental wipe samples: cyclophosphamide, ifosfamide, cytarabine, dacarbazine, docetaxel, paclitaxel, doxorubicin, epirubicin, etoposide, 5-fluorouracil, gemcitabine, irinotecan, methotrexate and pemetrexed. METHODS: The quantification method was developed using liquid chromatography coupled to tandem mass spectrometry and a wiping technique using viscose swabs. Linearity, accuracy, precision, limit of quantification, specificity and stability were assessed, from swab desorbed solution, to validate the analytical method, with respect to ICH guidelines. Environmental samples were collected by wiping five work surfaces of 225 cm2 with viscose swabs, during three days. RESULTS: The quantification method was linear over the calibration range with a lower limit of quantification ranging from 0.5 to 5.0 ng mL-1 depending on the cytotoxic drug. The intra-day and inter-day relative biases were below 1.5% and 13.5%, respectively. This method was successfully applied to surface-wipe sampling and environmental contaminations ranged from 0.7 to 1840.0 ng cm-2 for the most contaminated areas. CONCLUSIONS: This quantification method for 14 cytotoxic drugs was successfully applied to environmental contamination monitoring and could therefore be a useful tool for monitoring and toxicological studies.

HPLC-MS/MS method for the simultaneous determination of MB07133 and its metabolites, cytarabine and arabinofuranosyluracil, in rat plasma.

MB07133 is an intravenously administered cytarabine mononucleotide (araCMP) prodrug, for the treatment of hepatocellular carcinoma (HCC). A simple, selective and sensitive HPLC-MS/MS method using high pressure liquid chromatography (HPLC) coupled to triple-quadrupole mass spectrometer, was developed and validated for the detection of prodrug MB07133 and its metabolites, cytarabine (araC) and arabinofuranosyluracil (araU) in rat plasma. Protein precipitation using 3% trichloroacetic acid (TCA) was employed to extract analytes from 100µL rat plasma. Adequate separation of araC and araU from their endogenous compounds was achieved on the Synergi(®) fusion-RP column (150mm×4.6mm, 4µm) by a gradient-elution with a mobile phase consisting of ammonium formate (1mM) and methanol at a flow rate of 1mL/min. Multiple reaction monitoring mode (MRM) was applied in the detection of MB07133, araC, araU and Ganciclovir (internal standard) with ion pairs 441.2/330.2, 244.2/112.2, 245.2/113.2 and 256.1/152.2, respectively. The assays were validated with respect to specificity, linearity (100-50000ng/mL for MB07133, 2-1000ng/mL for araC and araU), accuracy and precision, extraction recovery, matrix effect and stability. The validated method has been successfully applied to an intravenous bolus pharmacokinetic study of MB07133 in male Sprague-Dawley rats (18mg/kg i.v.).

Simultaneous determination of 1-ß-d-Arabinofuranosylcytosine and two metabolites, 1-ß-d-Arabinofuranosyluracil and 1-ß-d-Arabinofuranosylcytosine triphosphate in leukemic cell by HPLC-MS/MS and the application to cell pharmacokinetics.

A specific and reliable HPLC-MS/MS method was developed and validated for the simultaneous determination of 1-ß-d-Arabinofuranosylcytosine (ara-C), 1-ß-d-Arabinofuranosyluracil (ara-U) and 1-ß-d-Arabinofuranosylcytosine triphosphate (ara-CTP) in the leukemic cells for the first time. The analytes were separated on a C18 column (100mm×2.1mm, 1.8µm) and a triple-quadrupole mass spectrometry equipped with an electrospray ionization (ESI) source was used for detection. The ion-pairing reagent, NFPA, was added to the mobile phase to retain the analytes in the column. The cell homogenates sample was prepared by the simple protein precipitation. The calibration curves were linear over a concentration range of 3.45-3450.0ng/mL for ara-C, 1.12-1120.0ng/mL for ara-U and 4.13-4130.0ng/mL for ara-CTP. The intra-day and inter-day precision was less than 15% and the relative error (RE) were all within ±15%. The validated method was successfully applied to assess the disposition characteristics of ara-C and support cell pharmacokinetics after the patients with leukemia were intravenously infused with SDAC and HiDAC. The result of the present study would provide the valuable information for the ara-C therapy.

Focused ultrasound-induced blood-brain barrier disruption enhances the delivery of cytarabine to the rat brain.

To investigate the feasibility of using focused ultrasound (FUS) with microbubbles for targeted delivery of cytarabine to the brain. Sprague-Dawly rats (weighing 200-250 g) received focused ultrasound with intravenous injection microbubbles. At 0, 2, 4, 8, and 24 hours (n=5 for each time point) after sonication, animals received intravenous administration of cytarabine at a normal dose of 4 mg/kg body weight. Additional five rats were given with a high dose (50 mg/kg body weight) of cytarabine alone. Blood-brain barrier (BBB) permeability and cerebral cytarabine were determined. FUS in conjunction with microbubbles caused a transient BBB opening. Sonication exposure promoted cytarabine accumulation at the sonicated site. Animals injected with a normal dose of cytarabine 2 hours after sonication had similar concentrations of cerebral cytarabine compared to those with higher cytarabine without sonication. FUS can temporarily open the BBB and thus facilitate the penetration of systemic cytarabine into the brain.

Wipe sampling procedure coupled to LC-MS/MS analysis for the simultaneous determination of 10 cytotoxic drugs on different surfaces.

A simple wipe sampling procedure was developed for the surface contamination determination of ten cytotoxic drugs: cytarabine, gemcitabine, methotrexate, etoposide phosphate, cyclophosphamide, ifosfamide, irinotecan, doxorubicin, epirubicin and vincristine. Wiping was performed using Whatman filter paper on different surfaces such as stainless steel, polypropylene, polystyrol, glass, latex gloves, computer mouse and coated paperboard. Wiping and desorption procedures were investigated: The same solution containing 20% acetonitrile and 0.1% formic acid in water gave the best results. After ultrasonic desorption and then centrifugation, samples were analysed by a validated liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) in selected reaction monitoring mode. The whole analytical strategy from wipe sampling to LC-MS/MS analysis was evaluated to determine quantitative performance. The lowest limit of quantification of 10 ng per wiping sample (i.e. 0.1 ng cm(-2)) was determined for the ten investigated cytotoxic drugs. Relative standard deviation for intermediate precision was always inferior to 20%. As recovery was dependent on the tested surface for each drug, a correction factor was determined and applied for real samples. The method was then successfully applied at the cytotoxic production unit of the Geneva University Hospitals pharmacy.

Biochemical modulation of aracytidine (Ara-C) effects by GTI-2040, a ribonucleotide reductase inhibitor, in K562 human leukemia cells.

GTI-2040 is a potent antisense to the M2 subunit of the ribonucleotide reductase (RNR), an enzyme involved in the de novo synthesis of nucleoside triphosphates. We hypothesized that combination of GTI-2040 with the cytarabine (Ara-C) could result in an enhanced cytotoxic effect with perturbed intracellular deoxynucleotide/nucleotide (dNTP/NTP) pools including Ara-C triphosphate (Ara-CTP). This study aims to provide a direct experimental support of this hypothesis by monitoring the biochemical modulation effects, intracellular levels of Ara-CTP, dNTPs/NTPs following the combination treatment of Ara-C, and GTI-2040 in K562 human leukemia cells. GTI-2040 was introduced into cells via electroporation. A hybridization-ligation ELISA was used to quantify intracellular GTI-2040 concentrations. Real-time PCR and Western blot methods were used to measure the RNR M2 mRNA and protein levels, respectively. 3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt assay was used to measure the cytotoxicity following various drug treatments. A non-radioactive HPLC-UV method was used for measuring the intracellular Ara-CTP, while a LC-MS/MS method was used to quantify intracellular dNTP/NTP pools. GTI-2040 was found to downregulate M2 mRNA and protein levels in a dose-dependent manner and showed significant decrease in dNTP but not NTP pool. When combining GTI-2040 with Ara-C, a synergistic cytotoxicity was observed with no further change in dNTP/NTP pools. Importantly, pretreatment of K562 cells with GTI-2040 was found to increase Ara-CTP level for the first time, and this effect may be due to inhibition of RNR by GTI-2040. This finding provides a laboratory justification for the current phase I/II evaluation of GTI-2040 in combination with Ara-C in patients with acute myeloid leukemia.

Assessment of primary, oxidative and excision repaired DNA damage in hospital personnel handling antineoplastic drugs.

The International Agency for Research on Cancer has classified several antineoplastic drugs in Group 1 (human carcinogens), among which chlorambucil, cyclophosphamide (CP) and tamoxifen, Group 2A (probable human carcinogens), among which cisplatin, etoposide, N-ethyl- and N-methyl-N-nitrosourea, and Group 2B (possible human carcinogens), among which bleomycins, merphalan and mitomycin C. The widespread use of these mutagenic/carcinogenic drugs in the treatment of cancer has led to anxiety about possible genotoxic hazards to medical personnel handling these drugs. The aim of the present study was to evaluate work environment contamination by antineoplastic drugs in a hospital in Central Italy and to assess the genotoxic risks associated with antineoplastic drug handling. The study group comprised 52 exposed subjects and 52 controls. Environmental contamination was assessed by taking wipe samples from different surfaces in preparation and administration rooms and nonwoven swabs were used as pads for the surrogate evaluation of dermal exposure, 5-fluorouracil and cytarabine were chosen as markers of exposure to antineoplastic drugs in the working environment. The actual exposure to antineoplastic drugs was evaluated by determining the urinary excretion of CP. The extent of primary, oxidative and excision repaired DNA damage was measured in peripheral blood leukocytes with the alkaline comet assay. To evaluate the role, if any, of genetic variants in the extent of genotoxic effects related to antineoplastic drug occupational exposure, the study subjects were genotyped for GSTM1, GSTT1, GSTP1 and TP53 polymorphisms. Primary DNA damage significantly increased in leukocytes of exposed nurses compared to controls. The use of personal protective equipment (i.e. gloves and/mask) was associated with a decrease in the extent of primary DNA damage.

A bioluminescent microbial biosensor for in vitro pretreatment assessment of cytarabine efficacy in leukemia.

BACKGROUND: The nucleoside analog cytarabine (Ara-C [cytosine arabinoside]) is the key agent for treating acute myeloid leukemia (AML); however, up to 30% of patients fail to respond to treatment. Screening of patient blood samples to determine drug response before commencement of treatment is needed. This project aimed to construct and evaluate a self-bioluminescent reporter strain of Escherichia coli for use as an Ara-C biosensor and to design an in vitro assay to predict Ara-C response in clinical samples. METHODS: We used transposition mutagenesis to create a cytidine deaminase (cdd)-deficient mutant of E. coli MG1655 that responded to Ara-C. The strain was transformed with the luxCDABE operon and used as a whole-cell biosensor for development an 8-h assay to determine Ara-C uptake and phosphorylation by leukemic cells. RESULTS: Intracellular concentrations of 0.025 µmol/L phosphorylated Ara-C were detected by significantly increased light output (P < 0.05) from the bacterial biosensor. Results using AML cell lines with known response to Ara-C showed close correlation between the 8-h assay and a 3-day cytotoxicity test for Ara-C cell killing. In retrospective tests with 24 clinical samples of bone marrow or peripheral blood, the biosensor-based assay predicted leukemic cell response to Ara-C within 8 h. CONCLUSIONS: The biosensor-based assay may offer a predictor for evaluating the sensitivity of leukemic cells to Ara-C before patients undergo chemotherapy and allow customized treatment of drug-sensitive patients with reduced Ara-C dose levels. The 8-h assay monitors intracellular Ara-CTP (cytosine arabinoside triphosphate) levels and, if fully validated, may be suitable for use in clinical settings.

HPLC method development and validation for simultaneous detection of Arabinoside-C and doxorubicin.

This paper describes a new validated high performance liquid chromatography (HPLC) method for the simultaneous determination of two anti-cancer drugs, Arabinoside-C (Ara-C) and doxorubicin hydrochloride (DOX). A simultaneous determination method saves cost and time as both drugs can be injected into a single HPLC system without the need to change or re-equilibrate with a new mobile phase. The objective of the study is to develop a simultaneous determination method of two anti-cancer drugs, Ara-C and DOX. The mobile phase consisted of a mixture (45:55) of acetonitrile:ammonium hydrogen phosphate aqueous solution (0.01 M) at pH 6.2 at a flow rate of 0.3 ml/min, with UV detection at 252 nm. Separation was achieved on a C-18 column (5 µm: 250 mm × 4.6 mm) maintained at 30°C in a column oven. The method was linear between 325 ng/ml and 10 µg/ml for Ara-C and 625 ng/ml and 20 µg/ml for DOX. The limit of detection (LOD) was 20 ng/ml for Ara-C and 60 ng/ml for DOX. The developed HPLC method achieved good precision and accuracy as well as limit of quantitations. The developed and validated method is suitable to be used for routine analysis of Ara-C and DOX.

A new, simple method for quantifying gemcitabine triphosphate in cancer cells using isocratic high-performance liquid chromatography.

A deoxycytidine analog, gemcitabine (dFdC), is effective for treating solid tumors and hematological malignancies. After being transported into cancer cells, dFdC is phosphorylated to dFdC triphosphate (dFdCTP), which is subsequently incorporated into the DNA strand, thereby inhibiting DNA synthesis. Intracellular dFdCTP is the critical determinant for dFdC cytotoxicity, so therapeutic drug monitoring or in vitro testing of the capability of cancer cells to accumulate dFdCTP may be informative for optimizing dFdC administration. We have developed a new isocratic-elution high-performance liquid chromatography method for quantifying dFdCTP in cancer cells. Samples (500 microL) were eluted isocratically using 0.06 M Na(2)HPO(4) (pH 6.9) containing 20% acetonitrile, at a constant flow rate of 0.7 mL/min and at ambient temperature. Separation was carried out using an anion-exchange column (TSK gel DEAE-2SW; 250 mm x 4.6 mm inside diameter, particle size 5 microL) and monitored at 254 nm. The standard curve was linear with low within-day and interday variability. The lower detection limit (20 pmol) was as sensitive as that of the previous gradient-elution method. dFdCTP was well separated from other nucleoside triphosphates. The method could measure dFdCTP in cultured or primary leukemic cells treated in vitro with dFdC. The method was also applicable to simultaneous determination of dFdCTP and cytarabine triphosphate, the results of which demonstrated ara-CTP production augmented by dFdC pretreatment. Thus, our isocratic high-performance liquid chromatography assay method will be of great use because of its sensitivity and simplicity as well as its applicability to biological materials.

Evaluation of early DNA damage in healthcare workers handling antineoplastic drugs.

OBJECTIVES: This study evaluates by comet assay the induction of early DNA damage in healthcare workers of an oncology hospital regularly handling antineoplastic drug mixtures. The aim was to identify a suitable biomarker of DNA damage by exposure to low levels of such drugs. METHODS: We studied 12 day hospital nurses and 13 oncology ward nurses who performed up to 300 and up to 35 drug administrations per week, respectively, and five pharmacy employees who regularly prepared mixtures of antineoplastic agents. Thirty healthy subjects were selected as controls. For exposure evaluation, we performed environmental monitoring of 5-fluorouracil, cytarabine, gemcitabine, cyclophosphamide, and ifosfamide in selected work areas of pharmacy and day hospital units and biological monitoring of urine for the 5-fluorouracile metabolite, alpha-fluoro-beta-alanine. We evaluated early DNA damage in lymphocytes and exfoliated buccal cells by comet assay measuring tail moment (TM) parameter that indirectly indicates the presence of DNA damage. RESULTS: Environmental monitoring detected cyclophosphamide, 5-fluorouracil and ifosfamide, with higher levels of contamination in day hospital unit. The biological monitoring measured detectable levels of alpha-fluoro-beta-alanine only in three nurses. Comet assay showed an increase on exfoliated buccal cells, even if not statistically significant, of mean TM with respect to controls in day hospital nurses (43.2 vs. 28.6, respectively) while ward nurses and pharmacy technicians did not show differences. Comet assay performed on lymphocytes did not show appreciable differences between exposed and controls. CONCLUSIONS: The employment of the sensitive comet assay, which is able to detect early the effects of a recent exposure to genotoxic substances, allowed us to find a slight DNA damage, only on exfoliated buccal cells of day hospital nurses, the group handling the highest amount of drugs during the administration process. This finding suggests that comet assay on exfoliated buccal cells could represent a useful tool to evaluate early and still repairable genotoxic effects of exposure to antineoplastic drug mixtures and then contribute to the improvement of the hospital safety practices.

Markedly improving Novozym 435-mediated regioselective acylation of 1-beta-D-arabinofuranosylcytosine by using co-solvent mixtures as the reaction media.

A comparative study was made of Novozym 435-catalyzed regioselective acylation of 1-beta-D-arabinofuranosylcytosine with vinyl propionate for the preparation of the 5'-O-monoester in eleven co-solvent mixtures and three pure polar solvents. Novozym 435 displayed low or no acylation activity toward 1-beta-D-arabinofuranosylcytosine in pure polar solvents, although those solvents can dissolve the nucleosides well. When a hexane-pyridine co-solvent system was adopted, both the initial rate and the substrate conversion were enhanced markedly. The polarity of co-solvent mixtures had significant effect on the reaction. Among the solvent mixtures investigated, the higher the polarity of the solvent mixture, the lower the initial reaction rate and the substrate conversion. It was also found that the acylation was dependent on the hydrophobic solvent content, the water activity and the reaction temperature. The most suitable co-solvent, initial water activity, and reaction temperature were hexane-pyridine (28:72, v/v), 0.07, and 50 degrees C, respectively. Under these conditions, the initial rate, the substrate conversion and the regioselectivity were as high as 91.1 mM h(-1), >97% and >98%, respectively, after a reaction time of 6 h. Among the reaction mediums examined, the lowest apparent activation energy was achieved with hexane-pyridine (28:72, v/v), in which Novozym 435 also exhibited good thermal stability.

A sensitive new method for clinically monitoring cytarabine concentrations at the DNA level in leukemic cells.

Cytarabine (ara-C), a major antileukemic agent, is phosphorylated in the cell to cytarabine triphosphate (ara-CTP), which is then partly incorporated into DNA. The drug incorporation into DNA poisons the extending primer against further incorporation of deoxyribonucleotides including dCTP, ultimately inhibiting DNA synthesis. While intracellular ara-CTP concentration has been found to predict clinical outcome, cytotoxicity in vitro is determined primarily by the extent of drug incorporation into DNA. However, clinically appropriate quantitation methods for ara-C at the DNA level have not been available. We developed a sensitive new method for monitoring ara-C incorporated into DNA in vivo. After DNA from leukemic cells was fractionated using the Schmidt-Thannhauser-Schneider method, it was degraded to constituent nucleosides to release ara-C, which was isolated from the nucleosides using HPLC and then measured by radioimmunoassay. Recovery for DNA fractionation, ara-C release by degradation, and ara-C isolation were 92.0+/-6.4%, 90.7+/-9.4%, and 98.5+/-1.4%, respectively. The method was found to determine ara-C incorporation into DNA of ara-C-treated HL 60 cells in vitro with minimal interassay variation. The values determined were compatible with those determined by scintillation counting in parallel experiments using tritiated ara-C. Our method could be used to monitor DNA-incorporated ara-C concentrations during ara-C therapy, together with plasma ara-C and intracellular ara-CTP concentrations. ara-C incorporation into DNA appeared to be associated with intracellular retention of ara-CTP or persistence of plasma ara-C. Thus, the present method is sensitive, accurate, precise, and may permit therapeutic drug monitoring at the DNA level for better individualization of antileukemic regimens.

Simultaneous separation and determination of Tarabine PFS and Adriblastine using micellar electrokinetic chromatography and high performance liquid chromatography. Application to some biological fluids.

The simultaneous determination of Tarabine PFS and Adriblastine by two independent techniques, viz. micellar electrokinetic chromatography (MEKC) and high performance liquid chromatography (HPLC), has been studied. For MEKC analysis, separations and identifications were accomplished using uncoated fused-silica capillaries and injections were performed in the hydrodynamic mode. The running buffer consisted of 0.05 M borate/phosphate pH 8.70, with 0.10 M SDS at an operating voltage of 15.0 kV and the temperature held at 25.0 degrees C. Under these conditions, the migration times of Tarabine PFS and Adriblastine were 2.70 and 6.40 min, respectively. Calibration curves were established for 0.010-0.300 microg/mL (r = 0.99) Tarabine PFS and 8.000-120.0 microg/mL (r = 0.99) Adriblastine. The limit of detection (LOD) was estimated and found to be 0.003 and 3.000 microg/mL of Tarabine PFS and Adriblastine, respectively. The limit of quantitation (LOQ) was found to be 0.009 and 8.000 microg/mL of Tarabine PFS and Adriblastine, respectively. For HPLC analysis, separations and determinations were performed on teicoplanin stationary phase with reversed mobile phase containing methanol:buffer pH 4.05 (20.0:80.0%, v/v) at 285 nm. Calibration curves were established for 3.000-90.00 microg/mL (r = 0.99) Tarabine PFS and for 10.00-120.0 microg/mL (r = 0.99) Adriblastine. LOD and LOQ were estimated and found to be 0.950 and 2.050 microg/mL of Tarabine PFS and 3.130 and 9.250 microg/mL of Adriblastine, respectively. Both MEKC and HPLC methods were applied for the simultaneous determination of analytes in urine samples. It was found that 8.00-10.0% (Tarabine PFS) and 13.0-15.0% (Adriblastine) of the injected dose was recovered in urine samples with 99.5-102% recovery.

Osteryoung square wave stripping voltammetry at mercury film electrode for monitoring ultra trace levels of Tarabine PFS and its interaction with ssDNA.

The electrochemical oxidation and reduction behaviour of adsorbed species of antimetabolic antineoplastic agent Tarabine PFS (Cytosar-U) in Sorensen buffer solution of different pH values at an in situ-mercury film electrode (MFE) is studied using cyclic voltammetry (CV) and Osteryoung square-wave stripping voltammetry (OSWSV). Optimal experimental and operational parameters have been selected for the drug preconcentration and determination in aqueous medium. Based on the adsorption and accumulation of Tarabine PFS using Osteryoung square-wave anodic stripping voltammetry (OSWASV) at MFE, the drug is easily detected as 0.134 ng/ml (5.51 x 10(-10) M). Calibration plots have been constructed at different accumulation times. The standard deviation (n=10) at a concentration level of 6 x 10(-8) M Tarabine PFS is 0.062. The interaction of ssDNA with the drug under the optimal conditions at pH 7.7 has been studied. The formal potentials E degrees and E degrees ' and the equilibrium constants K(1) and K(2) have been calculated for the free form of Tarabine PFS and the bonded form with ssDNA, respectively. It was found that K(2) value for the bonded oxidized form is 298 times than that of K(1) for the bonded reduced form. Therefore, ssDNA has been found to interact strongly with the oxidized form of the drug. The method has been used for the nanogram determination of ssDNA with 1.9% variation coefficient. Detection limit of 3 ng/ml ssDNA has been achieved. Possible interfering organic compounds, cations and anions have been tested. The method has been applied for the drug determination in urine samples, down to 0.23 ng/ml could be easily achieved in such samples.

Development of obesity and neurochemical backing in aurothioglucose-treated mice.

To clarify the neurochemical backing of aurothioglucose (ATG)-induced obesity in mice, we investigated lesion sites, hypothalamic neurotransmitters and c-Fos-like immunoreactivity (Fos-IR). At day 2 after ATG, tissue loss or cells death was observed in several parts of the ventral area of the ventromedial hypothalamic nucleus (VMH), and the dorsal area of arcuate nucleus and in the nucleus of the solitary tract (NTS). However, the greater part of the VMH was retained. Body weight began to increase in week 1. Hypothalamic serotonin (5-HT) and the metabolites were increased at day 2. The contents of acetylcholine, norepinephrine and dopamine in the hypothalamus showed no significant change. In week 1, the area shown tissue loss was compacted and plugged up. In the control group, most obvious c-Fos-like immunoreactive region was paraventricular nucleus (PVN). At day 2, Fos-IR was observed around destroyed regions in the hypothalamus and NTS, but few Fos-IR was found in the other regions including PVN. The Fos-IR around destroyed regions diminished after week 1. In week 3, Fos-IR in the PVN increased. These results suggest that the development of ATG-induced obesity cannot be attributed to solely VMH destruction. The restoration processes of the neuronal dysfunction involving PVN seem to play an important role in the development of obesity. NTS lesion and 5-HT system might contribute to decrease in food intake for several days after ATG.

Measurement of surface contamination by certain antineoplastic drugs using high-performance liquid chromatography: applications in occupational hygiene investigations in hospital environments.

Within the context of continuing interest in occupational hygiene of hospitals as workplaces, the authors report the results of a preliminary study on surface contamination by certain antineoplastic drugs (ANDs), recently performed in eight cancer departments of two large general hospitals in Milan, Italy. Since reliable quantitative information on the exposure levels to individual drugs is mandatory to establish a strong interpretative framework for correctly assessing the health risks associated with manipulation of ANDs and rationally advise intervention priorities for exposure abatement, two automated analytical methods were set up using reverse-phase high-performance liquid chromatography for the measurement of contamination by 1) methotrexate (MTX) and 2) the three most important nucleoside analogue antineoplastic drugs (5-fluorouracil 5FU, Cytarabin CYA, Gemcytabin GCA) on surfaces such as those of preparation hoods and work-benches in the pharmacies of cancer wards. The methods are characterized by short analysis time (7 min) under isocratic conditions, by the use of a mobile phase with a minimal content of organic solvent and by high sensitivity, adequate to detect surface contamination in the 5-10 micrograms/m2 range. To exemplify the performance of the analytical methods in the assessment of contamination levels from the target analyte ANDs, data are reported on the contamination levels measured on various surfaces (such as on handles, floor surfaces and window panes, even far from the preparation hood). Analyte concentrations corresponding to 0.8-1.5 micrograms of 5FU were measured on telephones, 0.85-28 micrograms/m2 of CYA were measured on tables, 1.2-1150 micrograms/m2 of GCA on furniture and floors. Spillage fractions between 1-5% of the used ANDs (daily use 5FU 7-13 g; CYA 0.1-7.1 g; GCA 0.2-5 g) were measured on the disposable polythene-backed paper cover sheet of the preparation hood.