The shrinking behavior, mechanism and anti-shrinkage resolution of an electrospun PLGA membrane.

The deformation shrinkage of a poly(lactide-co-glycolide) (PLGA) fibrous material seriously affects its biomedical application. To demonstrate the underlying shrinking mechanism and to find a method to prevent the shrinkage of an electrospun PLGA membrane, we investigated the shrinking behavior of PLGA electrospun membranes under various test conditions and discussed the underlying shrinking mechanism. The results indicated that the shrinkage of the electrospun PLGA membrane was mainly regulated by the glass transition of its polymer fiber; the temperature and liquid environment were found to be the two main factors leading to the shrinkage of the electrospun PLGA membrane through affecting its glass transition. Then a heat stretching (HS) technique was proposed by us to stabilize the electrospun PLGA membrane. After HS treatment, the glass transition temperature (Tg) of the electrospun PLGA membrane could increase from 48.38 °C to 54.55 °C. Our results indicated that the HS-treated membranes could maintain a high area percentage of 90.89 ± 2.27% and 84.78 ± 3.36% after immersion respectively in PBS and blood at 37 °C for 2 hours. Further experiments confirmed that the HS technique could also stabilize the dimensional structure of the electrospun PDLLA membrane in PBS and blood at 37 °C. This study provides an effective strategy for preventing the shrinkage of electrospun polyester biomaterials in a physiological environment that may benefit both the material structural stability and the in vivo biological performance.

Evaluation of three physiologically based pharmacokinetic (PBPK) modeling tools for emergency risk assessment after acute dichloromethane exposure.

INTRODUCTION: Physiologically based pharmacokinetic (PBPK) models may be useful in emergency risk assessment, after acute exposure to chemicals, such as dichloromethane (DCM). We evaluated the applicability of three PBPK models for human risk assessment following a single exposure to DCM: one model is specifically developed for DCM (Bos) and the two others are semi-generic ones (Mumtaz and Jongeneelen). MATERIALS AND METHODS: We assessed the accuracy of the models' predictions by simulating exposure data from a previous healthy volunteer study, in which six subjects had been exposed to DCM for 1h. The time-course of both the blood DCM concentration and percentage of carboxyhemoglobin (HbCO) were simulated. RESULTS: With all models, the shape of the simulated time course resembled the shape of the experimental data. For the end of the exposure, the predicted DCM blood concentration ranged between 1.52-4.19mg/L with the Bos model, 1.42-4.04mg/L with the Mumtaz model, and 1.81-4.31mg/L with the Jongeneelen model compared to 0.27-5.44mg/L in the experimental data. % HbCO could be predicted only with the Bos model. The maximum predicted % HbCO ranged between 3.1 and 4.2% compared to 0.4-2.3% in the experimental data. The % HbCO predictions were more in line with the experimental data after adjustment of the Bos model for the endogenous HbCO levels. CONCLUSIONS: The Bos Mumtaz and Jongeneelen PBPK models were able to simulate experimental DCM blood concentrations reasonably well. The Bos model appears to be useful for calculating HbCO concentrations in emergency risk assessment.

Induction of hepatic CYP2E1 by a subtoxic dose of acetaminophen in rats: increase in dichloromethane metabolism and carboxyhemoglobin elevation.

Dichloromethane (DCM) is metabolically converted to carbon monoxide mostly by CYP2E1 in liver, resulting in elevation of blood carboxyhemoglobin (COHb) levels. We investigated the effects of a subtoxic dose of acetaminophen (APAP) on the metabolic elimination of DCM and COHb elevation in adult female rats. APAP, at 500 mg/kg i.p., was not hepatotoxic as measured by a lack of change in serum aspartate aminotransferase, alanine aminotransferase, and sorbitol dehydrogenase activities. In rats pretreated with APAP at this dose, the COHb elevation resulting from administration of DCM (3 mmol/kg i.p.) was enhanced significantly. Also blood DCM levels were reduced, and its disappearance from blood appeared to be increased. Hepatic CYP2E1-mediated activities measured with chlorzoxazone, p-nitrophenol, and p-nitroanisole as substrates were all induced markedly in microsomes of rats treated with APAP. Aminopyrine N-demethylase activity was also increased slightly, but significantly. Western blot analysis showed that APAP treatment induced the expression of CYP2E1 and CYP3A proteins. Neither hepatic glutathione contents nor glutathione S-transferase activity was changed by the dose of APAP used. The results indicate that, contrary to the well known hepatotoxic effects of this drug at large doses, a subtoxic dose of APAP may induce CYP2E1, and to a lesser degree, CYP3A expression. This is the first report that APAP can increase cytochrome P450 (P450)-mediated hepatic metabolism and the resulting toxicity of a xenobiotic in the whole animal. The pharmacological/toxicological significance of induction of P450s by a subtoxic dose of APAP is discussed.

Determination of dichloromethane, trichloroethylene and perchloroethylene in urine samples by headspace solid phase microextraction gas chromatography-mass spectrometry.

A method for the determination of volatile chlorinated hydrocarbons, namely dichloromethane (DCM), trichloroethylene (TCE), and perchloroethylene (PCE), in urine samples was developed using headspace solid phase microextraction (HS-SPME) gas chromatography-mass spectrometry (GC-MS). HS-SPME was performed using a 75 microm Carboxen-polydimethylsiloxane fiber. Factors, which affect the HS-SPME process, such as adsorption and desorption times, stirring, salting-out effect, and temperature of sampling have been evaluated and optimized. The highest extraction efficiency was obtained when sampling was performed at room temperature (22 degrees C), from samples saturated with salt and under agitation. Linearity of the HS-SPME-GC-MS method was established over four orders of magnitude and the limit of detection was 0.005 microg/l for all the compounds. Precision, calculated as %R.S.D. at three different concentration levels, was within 1-8% for all intra- and inter-day determinations. The method was applied to the quantitative determination of TCE and PCE in human urine samples from exposed (TCE, n=5; median, 9.32 microg/l and PCE, n=39; median, 0.58 microg/l) and non-exposed individuals (n=120; median concentrations, 0.64, 0.22 and 0.11 microg/l for DCM, TCE and PCE, respectively. In addition, two cases of acute accidental exposure to DCM are reported, and the elimination kinetics in blood and urine was followed up. The calculated half-lives of urinary and blood DCM were, respectively, 7.5 and 8.1 h for one subject and 3.8 and 4.3 h for the other.

Lead and methylene chloride exposures among automotive repair technicians.

Potential exposures among repair technicians engaged in vehicle resurfacing operations prior to spray painting have not been thoroughly characterized. Environmental and personal air monitoring conducted in the State of Rhode Island have shown that automotive repair technicians may be exposed to metal particulates in sanding dust and methylene chloride vapors during vehicle paint removal operations. Hand wipe samples demonstrated that metals in sanding dust adhered to the hands of workers throughout the duration of the work day and were available for incidental ingestion from the handling of food/nonfood items and hand-to-mouth contact. A blood lead (PbB) screening effort among 21 workers at 2 facilities showed that 4 non-/less-exposed workers had mean PbB levels at the U.S. geometric mean of 2.8 microg/dL, while 2 out of 9 (22%) dedicated vehicle repair technicians had PbB levels at or above 30 microg Pb/dL whole blood--the level for potential adverse reproductive effects. Methylene chloride exposures were also found to exceed the Occupational Safety and Health Administrations (OSHA) 8-hr time-weighted average (TWA) action level and permissible exposure limit (PEL) in a limited number of samples (120 and 26 ppm, integrated work shift samples). Our findings suggest that thousands of professional technicians and vocational high school students may be at increased risk of adverse reproductive and/or other systemic effects.

Physiologically based pharmacokinetic modeling of arterial - antecubital vein concentration difference.

BACKGROUND: Modeling of pharmacokinetic parameters and pharmacodynamic actions requires knowledge of the arterial blood concentration. In most cases, experimental measurements are only available for a peripheral vein (usually antecubital) whose concentration may differ significantly from both arterial and central vein concentration. METHODS: A physiologically based pharmacokinetic (PBPK) model for the tissues drained by the antecubital vein (referred to as "arm") is developed. It is assumed that the "arm" is composed of tissues with identical properties (partition coefficient, blood flow/gm) as the whole body tissues plus a new "tissue" representing skin arteriovenous shunts. The antecubital vein concentration depends on the following parameters: the fraction of "arm" blood flow contributed by muscle, skin, adipose, connective tissue and arteriovenous shunts, and the flow per gram of the arteriovenous shunt. The value of these parameters was investigated using simultaneous experimental measurements of arterial and antecubital concentrations for eight solutes: ethanol, thiopental, 99Tcm-diethylene triamine pentaacetate (DTPA), ketamine, D2O, acetone, methylene chloride and toluene. A new procedure is described that can be used to determine the arterial concentration for an arbitrary solute by deconvolution of the antecubital concentration. These procedures are implemented in PKQuest, a general PBPK program that is freely distributed http://www.pkquest.com. RESULTS: One set of "standard arm" parameters provides an adequate description of the arterial/antecubital vein concentration for ethanol, DTPA, thiopental and ketamine. A significantly different set of "arm" parameters was required to describe the data for D2O, acetone, methylene chloride and toluene - probably because the "arm" is in a different physiological state. CONCLUSIONS: Using the set of "standard arm" parameters, the antecubital vein concentration can be used to determine the whole body PBPK model parameters for an arbitrary solute without any additional adjustable parameters. Also, the antecubital vein concentration can be used to estimate the arterial concentration for an arbitrary input for solutes for which no arterial concentration data is available.

Capillary gas chromatography with cryogenic oven temperature for headspace samples: analysis of chloroform or methylene chloride in whole blood.

A new and sensitive gas chromatography (GC) method for measurement of chloroform or methylene chloride in whole blood is presented. Trace levels of these analytes present in the headspace of samples were cryogenically trapped prior to on-line GC analysis. After heating of a blood sample containing chloroform and methylene chloride (internal standard, and vice versa) in a 7.0-mL vial at 55 degrees C for 20 min, 5 mL of the headspace vapor was drawn into a glass syringe. All vapor was introduced into an Rtx-Volatiles middle-bore capillary column in the splitless mode at -30 degrees C oven temperature to trap the entire analytes, and the oven temperature was programmed up to 280 degrees C for detection of the compounds and for cleaning of the column. The present conditions gave sharp peaks for both chloroform and methylene chloride and very low background noises for whole blood samples. As much as 11.5 and 20.0% of chloroform and methylene chloride, respectively, which had been added to whole blood in a vial, could be introduced into the GC column. The calibration curves showed linearity in the range of 0.05-5.0 micrograms/0.5 mL of whole blood. The detection limit was estimated to be about 2 ng/0.5 mL. The coefficients of intraday and interday variations were 1.31 and 8.90% for chloroform and 1.37 and 9.03% for methylene chloride, respectively. The data on chloroform or methylene chloride in rat blood after inhalation of each compound were also presented.

Effect of a single administration of benzene, toluene or m-xylene on carboxyhaemoglobin elevation and metabolism of dichloromethane in rats.

The effect of a single administration of aromatic hydrocarbons (AHCs) on the metabolic activity responsible for the biotransformation of dichloromethane (DCM) to carbon monoxide (CO) was investigated using adult female rats. In rats treated orally with benzene (1.5 ml kg-1), toluene (2.0 ml kg-1) or m-xylene (2.0 ml kg-1) 16-24 h prior to DCM (3 mmol kg-1, i.p.), the carboxyhaemoglobin (COHb) level was elevated, reaching peaks in blood at 21%, 16% and 23%, respectively, compared to the peak of ca. 10% in rats treated with DCM only. Their effects on COHb generation were highly dependent on the time interval between each AHC and DCM treatment, since an early administration of m-xylene or toluene decreased the COHb elevation. The half-life of DCM in blood was shortened significantly, indicating that the metabolic degradation of DCM was enhanced by the AHCs. Disulfiram (3.4 mmol kg-1, p.o.) blocked COHb elevation completely, suggesting that the metabolic conversion of DCM to CO is mediated by cytochrome P-450 2E1 (P4502E1). Corresponding increases in the concentration and half-life of DCM in blood were also observed. A single administration of the AHCs did not alter the hepatic glutathione level, suggesting that the increase in DCM-induced COHb elevation was not due to hepatic glutathione depletion. In vitro studies showed that the hepatic microsomal metabolism of nitrosodimethylamine and p-nitrophenol was significantly increased by a single dose of each AHC. Total cytochrome P-450 content and p-nitroanisole demethylase activity were also increased; however, only toluene and m-xylene were effective inducers for aminopyrine N-demethylase. Therefore, benzene appears to be a selective inducer for P4502E1 compared to other alkylbenzenes. The results indicate that even a single dose of benzene, toluene or m-xylene may induce the activity of P4502E1 significantly, which is responsible for the increased generation of COHb from DCM, as demonstrated in the present study.

Sensitivity of physiologically based pharmacokinetic models to variation in model parameters: methylene chloride.

The parameters in a physiologically based pharmacokinetic (PBPK) model of methylene chloride were varied systematically, and the resulting variation in a number of model outputs was determined as a function of time for mice and humans at several exposure concentrations. The importance of the various parameters in the model was highly dependent on the conditions (concentration, species) for which the simulation was performed and the model output (dose surrogate) being considered. Model structure also had a significant impact on the results. For sensitivity analysis, particular attention must be paid to conservation equations to ensure that the variational calculations do not alter mass balance, introducing extraneous effects into the model. All of the normalized sensitivity coefficients calculated in this study ranged between -1.12 and 1, and most were much less than 1 in absolute value, indicating that individual input errors are not greatly amplified in the outputs. In addition to ranking parameters in terms of their impact on model predictions, time-dependent sensitivity analysis can also be used as an aid in the design of experiments to estimate parameters by predicting the experimental conditions and sampling points which will maximize parameter identifiability.

Polymorphism of glutathione conjugation of methyl bromide, ethylene oxide and dichloromethane in human blood: influence on the induction of sister chromatid exchanges (SCE) in lymphocytes.

A hitherto unknown glutathione-S-transferase in human erythrocytes displays polymorphism: three quarters of the population ("conjugators") possess, whereas one quarter ("non-conjugators") lack this specific activity. A standard method for the identification of conjugators and non-conjugators with the use of methyl bromide and gas chromatography (head space technique) is described. Three substrates of the polymorphic enzyme, methyl bromide, ethylene oxide and dichloromethane (methylene chloride), were incubated in vitro with individual whole blood samples of conjugators and non-conjugators. All three substances led to a marked increase of sister chromatid exchanges (SCE) in the lymphocytes of the non-conjugators but not in those of conjugators. A protective effect of the glutathione-S-transferase activity in human erythrocytes for the cytogenetic toxicity of these chemicals in vitro is thus confirmed. Since the enzyme activity is not found in erythrocytes of laboratory animals, species extrapolations for risk assessment of methyl bromide, ethylene oxide and dichloromethane should be reconsidered.

Double fatal inhalation of dichloromethane.

1 Two cases of lethal poisoning following acute inhalation of extremely high concentrations of dichloromethane (DCM) are reported. The concentrations of the solvent found in the blood of the two subjects collected at autopsy and analysed by gas chromatography/mass spectrometry (572 and 601 mg l-1) were compatible with those measured in the air a few hours after the discovery of the bodies (up to 168,000 ppm). 2 Extensive brain and lung oedema and congestion, microhaemorrhagic changes of the stomach and congestion in other organs were observed on macroscopic and microscopic examination of both subjects. In addition, and in both cases, high but not lethal carboxyhaemoglobin (COHb) levels (30%) were found in the blood collected at autopsy. 3 Narcosis and respiratory depression due to the effect of DCM on the central nervous system (CNS) appear to have played a critical role in the death of the two men. However, biotransformation of the solvent to toxic metabolites, including carbon monoxide (via oxidative dehalogenation by the cytochrome P450-dependent mixed function oxidase system) or formaldehyde, formic acid, inorganic chloride and carbon dioxide (via the glutathione-S-transferase pathway) may have also contributed significantly to fatal toxicity.

Interaction of dichloromethane and ethanol in rats: toxicokinetics and nerve conduction velocity.

The influence of a single combined administration of ethanol (174 mmol/kg per os) plus dichloromethane (1.6, 6.2, or 15.6 mmol/kg p.o.) on blood concentrations of the tested substances and of carboxyhemoglobin, and on nerve conduction velocity was studied in rats. The blood alcohol concentration was not influenced significantly by dichloromethane. The single high dose of ethanol completely inhibited the carboxyhemoglobin concentration increase due to dichloromethane, but did not prevent the dichloromethane-induced decrease of nerve conduction velocity. It produced initially lower, then higher concentrations of dichloromethane in blood than values seen after administration of dichloromethane per se. Rats treated with ethanol plus dichloromethane showed a more pronounced decrease of nerve conduction velocity compared with rats administered dichloromethane only.

The pharmacokinetics of dichloromethane. I. Disposition in B6C3F1 mice following intravenous and oral administration.

The tissue distribution and metabolism of dichloromethane (DCM; CH2Cl2) was investigated in B6C3F1 mice following iv or oral administration. The route of exposure and the composition of the dosing solution were found to have a significant effect on the pharmacokinetics. Following single iv doses of 10 or 50 mg [14C]DCM/kg dose-dependent metabolism to 14CO2 and 14CO and rapid pulmonary clearance of unchanged 14CH2Cl2 characterized the elimination of DCM from the body. The highest concentrations of 14CH2Cl2 were found in the liver, lung and kidney, with more than 50% of the total radioactivity in these tissues represented by the parent compound. When DCM was administered orally in single gavage doses for 14 consecutive days at treatment levels of 50 mg/kg in water or 500 and 1000 mg/kg in corn oil, rapid absorption and elimination of DCM characterized the treatment in water while distinctly slower trends were found for the doses in corn oil. No observable pharmacokinetic or metabolic effect resulted from repeated oral dosing over the 2-wk treatment period.

The pharmacokinetics of dichloromethane. II. Disposition in Fischer 344 rats following intravenous and oral administration.

The tissue distribution and metabolism of dichloromethane (DCM; CH2Cl2) was investigated in Fischer 344 rats following iv or oral administration. The route and level of exposure were found to have a significant effect on the disposition characteristics. A two-compartment model was used to describe the elimination of DCM from blood following single iv doses. The estimates of t1/2,beta were 11.9 and 23.5 min for doses of 10 and 50 mg/kg, respectively, and the disposition rate constants, beta were found to differ significantly at P less than 0.05. When DCM was administered orally (by gavage) in a daily dose of 50 or 200 mg/kg for 14 consecutive days, rapid absorption and distribution to the tissues characterized the disposition. Dose-dependent metabolism to 14CO2 and 14CO and rapid pulmonary clearance of unchanged 14CH2Cl2 were the dominant routes of elimination of DCM from the body following both iv and oral doses. No observable pharmacokinetic or metabolic effect resulted from repeated oral dosing.