Toxicity screening of mouthwashes in the pollen tube growth test: safety assessment of recommended dilutions.

This study investigated whether mouthwash dilutions, recommended by the manufacturers, may contribute to oral mucosa irritation. Twenty frequently used mouthwash brands were examined by the pollen tube growth test, an alternative in vitro method for assessment of the irritating potential of ingredients of cosmetic formulations. The test is based on the photometric quantification of pollen tube growth inhibition. This parameter is expressed by IC50 values that characterize the cytotoxic potential of a product. The IC50s clearly revealed that none of the tested mouthwashes should cause acute irritation if used in the recommended dilution. However, at least 4 of the mouthwashes evaluated in this study could probably irritate the oral mucosa acutely if used undiluted (original concentration).

Toxicity assessment of mouthwashes in the pollen tube growth test.

AIM: Mouthwashes are frequently used to treat oral discomfort, halitosis or superficial infections. Adverse effects of mouthwash use are recognized, e.g. mucosal peeling and stomatitis. In addition, mouthwashes have been accused of causing oral cancer. The aim of this study was to analyse the acute toxicity of mouthwashes in an in vitro testing system, the pollen tube growth test (PTG test). MATERIALS AND METHODS: Twenty mouthwash brands were prepared for study. The mouthwashes were added to suspensions of pollen of tobacco plants (Nicotiana sylvestris) that were collected during the early stages of anthesis. The inhibition of the pollen germination and pollen tube growth T(t) was measured photometrically from centrifuged supernatants of the Alcian blue-stained suspensions at a wavelength of 607 nm after 18 hours of incubation. The results were compared to inhibitions that resulted from other toxic substances. RESULTS: In the PTG-test, the acute toxicity of several mouthwashes exceeds the toxic effect of alcohol. Calculated IC50 values for the mouthwashes ranged from 49,326 +/- 4881 mg/l to 541 +/- 137 mg/l. CONCLUSION: Consumers should be informed about the number and concentrations of mouthwash ingredients and their toxic potential. These declarations would provide the basis for further testing the toxic effects of mouthwashes.

Toxicity screening of mouthwashes in the pollen tube growth test: safety assessment of recommended dilutions of twenty brands.

The question of whether mouthwash dilutions, recommended by the producers, contribute to irritation of the oral mucosa was examined. Twenty frequently used mouthwash brands were examined by the pollen tube growth test, an alternative in vitro method for the irritation assessment of ingredients of cosmetic formulations. The test is based on the photometric quantification of pollen tube growth inhibition. This parameter is expressed by IC50 values which characterize the cytotoxic potency of a product. The IC50s clearly revealed that none of the tested mouthwashes should cause acute irritation if used at the recommended dilution. However, 4 to 5 of the mouthwashes could probably irritate the oral mucosa acutely, when used in the form of the original producer concentrations.

Morphology of cell injury: An approach to the EDIT programme by the use of tobacco pollen tubes. Evaluation-guided Development of New In Vitro Test Batteries.

Tobacco pollen tubes were used as a standard in vitro system to investigate cell growth aberrations caused by some of the Multicentre Evaluation of In Vitro Cytotoxicity (MEIC) programme chemicals and other toxic compounds. Changes in cytoskeletal pattern were observed in the tube cells by using tubulin immunofluorescence and rhodamin-phalloidin fluorescence for the localisation of microtubules and actin filaments, respectively. Four different types of cell malformation were found: screw-like growth, isodiametric tip swelling, hook formation, and pollen grain enlargement. We suggest that these malformations resulted from an interference by the chemicals with the cytosolic calcium gradient which controls tip growth and the orientation of the pollen tube. The results may contribute to a general understanding of toxicity-based cell malformations.

Development of an in vitro test battery for the estimation of acute human systemic toxicity: An outline of the EDIT project. Evaluation-guided Development of New In Vitro Test Batteries.

The aim of the Evaluation-guided Development of new In Vitro Test Batteries (EDIT) multicentre programme is to establish and validate in vitro tests relevant to toxicokinetics and for organ-specific toxicity, to be incorporated into optimal test batteries for the estimation of human acute systemic toxicity. The scientific basis of EDIT is the good prediction of human acute toxicity obtained with three human cell line tests (R(2) = 0.77), in the Multicentre Evaluation of In Vitro Cytotoxicity (MEIC) programme. However, the results from the MEIC study indicated that at least two other types of in vitro test ought to be added to the existing test battery to improve the prediction of human acute systemic toxicity - to determine key kinetic events (such as biotransformation and passage through biological barriers), and to predict crucial organ-specific mechanisms not covered by the tests in the MEIC battery. The EDIT programme will be a case-by-case project, but the establishment and validation of new tests will be carried through by a common, step-wise procedure. The Scientific Committee of the EDIT programme defines the need for a specific set of toxicity or toxicokinetic data. Laboratories are then invited to perform the defined tests in order to provide the "missing" data for the EDIT reference chemicals. The results obtained will be evaluated against the MEMO (the MEIC Monograph programme) database, i.e. against human acute systemic lethal and toxicity data. The aim of the round-table discussions at the 19th Scandinavian Society for Cell Toxicology (SSCT) workshop, held in Ringsted, Denmark on 6-9 September 2001, was to identify which tests are the most important for inclusion in the MEIC battery, i.e. which types of tests the EDIT programme should focus on. It was proposed that it is important to include in vitro methods for various kinetic events, such as biotransformation, absorption in the gut, passage across the blood-brain barrier, distribution volumes, protein binding, and renal clearance/accumulation. Models for target organ toxicity were also discussed. Because several of the outlier chemicals (paracetamol, digoxin, malathion, nicotine, paraquat, atropine and potassium cyanide) in the MEIC in vivo-in vitro evaluation have a neurotoxic potential, it was proposed that the development within the EDIT target organ programme should initially be focused on the nervous system.

Adaptation of the Pollen Tube Growth Assay to Cytotoxicity Testing of Cigarette Smoke.

We describe a recently developed in vitro assay for the assessment of the cytotoxicity of gaseous materials. The method is based on the pollen tube growth (PTG) test suited to determine the cytotoxicity of non-gaseous, water-soluble substances (Kristen and Kappler, 1995). Instead of the conventional PTG test, we used a semi-automatic, computer-controlled multi-channel device for the injection of gases and gas mixtures into a battery of incubation tubes containing germinating, suspension-cultured tobacco pollen as indicator of toxic effects. The efficiency of the new PTG gas test device was examined using the gas phase of cigarette smoke. The examination exhibited sufficient sensitivity of the method and high reproducibility of the toxicity data obtained. The assay can be adapted to various sources of exhaust gases.

Toxicity screening of mouthwashes in the pollen tube growth test: safety assessment of recommended dilutions

Este estudo investigou se soluções de bochecho nas diluições recomendadas pelos fabricantes podem contribuir para a irritação das mucosas orais. Vinte soluções de bochecho frequentemente utilizadas foram avaliadas por meio do análise do crescimento do tubo polínico. Este teste constitui um método alternativo in vitro para estimar o potencial irritante dos ingredientes de formulações cosméticas. O teste baseia-se numa medição fotométrica da inibição do crescimento de tubos polínicos. A inibição do crescimento é expressa por valores IC50 que caracterizam o potencial citotóxico de uma dada substância. Os valores de IC50 aqui apresentados demonstraram que nenhuma das soluções de bochecho estudadas deve causar irritação aguda das mucosas orais, desde que utilizadas nas diluições recomendadas pelo fabricante. No entanto, verificou-se que pelo menos quatro dos colutórios podem causar irritação aguda das mucosas orais ao serem utilizados na a forma concentrada.