Ecotoxicology: Conventional and new topics and methods

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In-vitro measurement of aerosol generated by Respimat SMI penetrating through upper airways

Respimat soft mist inhaler (SMI) is an inhaler producing a soft mist with low velocity. The liquid drug solution is emitted by the mechanical energy of compressed spring, so the device is propellant-free. This paper describes the measurement of particle fraction penetrating through the model of the upper respiratory tract (mouth cavity, pharynx, larynx trachea). The solution inside the cartridge was marked by fluorescein in order to evaluate the regional particle deposition in the model. After the exposition, segments of the model, as well as the filter, were washed out and the deposited fluorescein was extracted into water. The analysis showed that the fraction of particles penetrating through the upper respiratory tract was 64.7±6.7 %. However, the real inhalation pattern was not simulated and measurement was provided with continual “inhalation” without “exhalation”

Environmental and Molecular Mutagenesis / Cytome micronucleus assays with a metabolically competent human derived liver cell line (Huh6) : a promising approach for routine testing of chemicals?

One of the main problems of in vitro genotoxicity tests is the inadequate representation of drug metabolizing enzymes in most indicator cell lines which are currently used. We identified recently a human derived liver cell line (Huh6) which detected induction of DNA damage by representatives of different groups of promutagens without enzyme mix and showed that these cells are more suitable in terms of reproducibility and sensitivity as other currently used liver derived lines. We developed a protocol for micronucleus (MN) cytome assays with these cells and validated the procedure in experiments with representatives of different groups of directly and indirectly acting genotoxic carcinogens (MMS, cisplatin, PhIP, IQ, NDMA, B(a)P, AFB1, etoposide, and H2O2). The optimal cytochalasin B concentration in combination with 48 hr treatment was found to be 1.5 g/mL and leads to a cytokinesis block proliferation index in the range between 1.7 and 2.0. The morphological characteristics of different nuclear anomalies which reflect DNA damage (MN, nuclear bridges, and buds) and their baseline frequencies in untreated cells were characterized, and the rates which are required to cause significant effects were calculated. All compounds caused dose dependent induction of MN when the cells were treated for 24 hr, longer and shorter exposure times were less effective. Experiments with different serum levels (fetal bovine serum [FBS]) showed that 10% FBS in the medium (instead of 4%) causes a substantial increase of the sensitivity of the cells. Our results indicate that the new protocol is a promising approach for routine testing of chemicals.(VLID)341191

Inhalers and nebulizers: basic principles and preliminary measurements

Inhalers are hand-held devices which are used for administration of therapeutic aerosols via inhalation. Nebulizers are larger devices serving for home and hospital care using inhaled medication. This contribution describes the basic principles of dispersion of aerosol particles used in various types of inhalers and nebulizers, and lists the basic physical mechanisms contributing to the deposition of inhaled particles in the human airways. The second part of this article presents experimental setup, methodology and preliminary results of particle size distributions produced by several selected inhalers and nebulizers

Dielectrophoretic classification of fibres: principles and application to glass fibres suspended in air

Particles exposed to an electric field experience forces that influence their movement. This effect can be used for filtration of air, or for size classification of aerosols. The motion of charged particles in a non-uniform electric field is called electrophoresis. Two processes are involved in this phenomenon: 1) charging of particles and 2) electrical mobility separation. If fibres are exposed to electrophoresis, they are separated on the basis of two parameters: diameter and length. Regrettably, as naturally occurring fibres are polydisperse both in diameter and length, the electrophoresis is not very efficient in length classification. In contrast, dielectrophoresis is the motion of electrically neutral particles in a non-uniform electric field due to the induced charge separation within the particles. As deposition velocity of fibres induced by dielectrophoretic force strongly depends on length and only weakly on diameter, it can be used for efficient length classification. Principles of length classification of conducting and non-conducting fibres are presented together with design of a fibre classifier. Lastly, images of motion of fibres recorded by high-speed camera are depicted

Dielectrophoretic classification of fibres: principles and application to glass fibres suspended in air

Particles exposed to an electric field experience forces that influence their movement. This effect can be used for filtration of air, or for size classification of aerosols. The motion of charged particles in a non-uniform electric field is called electrophoresis. Two processes are involved in this phenomenon: 1) charging of particles and 2) electrical mobility separation. If fibres are exposed to electrophoresis, they are separated on the basis of two parameters: diameter and length. Regrettably, as naturally occurring fibres are polydisperse both in diameter and length, the electrophoresis is not very efficient in length classification. In contrast, dielectrophoresis is the motion of electrically neutral particles in a non-uniform electric field due to the induced charge separation within the particles. As deposition velocity of fibres induced by dielectrophoretic force strongly depends on length and only weakly on diameter, it can be used for efficient length classification. Principles of length classification of conducting and non-conducting fibres are presented together with design of a fibre classifier. Lastly, images of motion of fibres recorded by high-speed camera are depicted

The flow of micrometre-sized glass fibres in a replica of the human trachea

Inhaled fibres can potentially cause inflammation of the lung tissue and interstitium which, after long-term exposure, may lead to lung cancer, malignant mesothelioma or pulmonary and pleural fibrosis. For risk reduction and correct setting of occupational hygiene regulations, it is important to be able to precisely calculate the fate of inhaled fibres depending on their physical characteristics and inhalation conditions. As there is a lack of experimental data on the orientation of fibres, a new test rig has been assembled for visualization and recording of flowing fibres in a replica of the human trachea. Fibres prepared from regular glass fibres produced commercially for blown thermal insulation have been processed, dispersed and introduced into the glass tube with dimensions of the trachea. Visualization was performed using a powerful LED light and a high-speed camera. Angles of the fibres have been evaluated for six different flowrates and the dependence of the angles on the flow Reynolds number was searched for. The angles of fibres agreed with expected values, i.e. only vertically and horizontally oriented fibres were recorded. However, the number of vertically and horizontally oriented fibres did not seem to be correlated with the flow Reynolds number

Genotoxic properties of XLR-11, a widely consumed synthetic cannabinoid, and of the benzoyl indole RCS-4

Aim of this study was the investigation of the genotoxic properties of XLR-11 [1-(5-fluoropentyl)-1H-indol-3-yl](2,2,3,3-tetramethylcyclopropyl)methanone, a widely consumed synthetic cannabinoid (SC), and of the benzoyl indole RCS-4 (4-methoxyphenyl)(1-pentyl-1H-indol-3-yl)methanone). We characterized the DNA-damaging properties of these drugs in different experimental systems. No evidence for induction of gene mutations was detected in bacterial (Salmonella/microsome) tests, but clear dose-dependent effects were found in in vitro single cell gel electrophoresis (SCGE) assays with human lymphocytes and with buccal- and lung-derived human cell lines (TR-146 and A-549). These experiments are based on the determination of DNA migration in an electric field and enable the detection of single- and double-strand breaks and apurinic sites. Furthermore, we found that both drugs induce micronuclei which are formed as a consequence of chromosomal aberrations. The lack of effects in SCGE experiments with lesion-specific enzymes (FPG, Endo III) shows that the DNA damage is not caused by formation of oxidatively damaged bases; experiments with liver enzyme homogenates and bovine serum albumin indicate that the drugs are not converted enzymatically to DNA-reactive intermediates. Furthermore, results with buccal- and lung-derived human cells show that gaseous treatment of the cells under conditions which reflect the exposure situation in drug users may cause damage of the genetic material in epithelia of the respiratory tract. Since DNA instability is involved in the etiology of cancer, these findings can be taken as an indication that consumption of the SCs may cause tumors in the respiratory tract of consumers.(VLID)348927