Accuracy of 3D Tooth Movements in the Fabrication of Manual Setup Models for Aligner Therapy.

Background: The clinical outcome of aligner therapy is closely related to the precision of its setup, which can be manually or digitally fabricated. The aim of the study is to investigate the suitability of manual setups made for aligner therapy in terms of the precision of tooth movements. Methods: Six dental technicians were instructed to adjust each of eleven duplicate plaster casts of a patient models as follows: a 1 mm pure vestibular translation of tooth 11 and a 15° pure mesial rotation of tooth 23. The processed setup models were 3D scanned and matched with the reference model. The one-sample Wilcoxon signed-rank test (p < 0.05) was used for evaluation. Results: The overall precision of the translational movement covers a wide range of values from 0.25 to 2.26 mm (median: 1.09 mm). The target value for the rotation of tooth 23 was achieved with a median rotation of 9.76° in the apical-occlusal direction. Unwanted movements in the other planes also accompanied the rotation. Conclusions: A manual setup can only be fabricated with limited precision. Besides the very high variability between technicians, additional unwanted movements in other spatial planes occurred. Manually fabricated setups should not be favored for aligner therapy due to limited precision.

A plea for the integration of Green Toxicology in sustainable bioeconomy strategies - Biosurfactants and microgel-based pesticide release systems as examples.

A key aspect of the transformation of the economic sector towards a sustainable bioeconomy is the development of environmentally friendly alternatives for hitherto used chemicals, which have negative impacts on environmental health. However, the implementation of an ecotoxicological hazard assessment at early steps of product development to elaborate the most promising candidates of lowest harm is scarce in industry practice. The present article introduces the interdisciplinary proof-of-concept project GreenToxiConomy, which shows the successful application of a Green Toxicology strategy for biosurfactants and a novel microgel-based pesticide release system. Both groups are promising candidates for industrial and agricultural applications and the ecotoxicological characterization is yet missing important information. An iterative substance- and application-oriented bioassay battery for acute and mechanism-specific toxicity within aquatic and terrestrial model species is introduced for both potentially hazardous materials getting into contact with humans and ending up in the environment. By applying in silico QSAR-based models on genotoxicity, endocrine disruption, skin sensitization and acute toxicity to algae, daphnids and fish, individual biosurfactants resulted in deviating toxicity, suggesting a pre-ranking of the compounds. Experimental toxicity assessment will further complement the predicted toxicity to elaborate the most promising candidates in an efficient pre-screening of new substances.

Designed to Be Green, Economic, and Efficient: A Ketone-Ester-Alcohol-Alkane Blend for Future Spark-Ignition Engines.

Model-based fuel design can tailor fuels to advanced engine concepts while minimizing environmental impact and production costs. A rationally designed ketone-ester-alcohol-alkane (KEAA) blend for high efficiency spark-ignition engines was assessed in a multi-disciplinary manner, from production cost to ignition characteristics, engine performance, ecotoxicity, microbial storage stability, and carbon footprint. The comparison included RON 95 E10, ethanol, and two previously designed fuels. KEAA showed high indicated efficiencies in a single-cylinder research engine. Ignition delay time measurements confirmed KEAA's high auto-ignition resistance. KEAA exhibits a moderate toxicity and is not prone to microbial infestation. A well-to-wheel analysis showed the potential to lower the carbon footprint by 95 percent compared to RON 95 E10. The findings motivate further investigations on KEAA and demonstrate advancements in model-based fuel design.

Green toxicological investigation for biofuel candidates.

To avoid potential risks of biofuels on the environment and human, ecotoxicity investigation should be integrated into the early design stage for promising biofuel candidates. In the present study, a green toxicology testing strategy combining experimental bioassays with in silico tools was established to investigate the potential ecotoxicity of biofuel candidates. Experimental results obtained from the acute immobilisation test, the fish embryo acute toxicity test and the in vitro micronucleus assay (Chinese hamster lung fibroblast cell line V79) were compared with model prediction results by ECOSAR and OECD QSAR Toolbox. Both our experimental and model prediction results showed that 1-Octanol (1-Oct) and Di-n-butyl ether (DNBE) were the most toxic to Daphnia magna and zebrafish among all the biofuel candidates we investigated, while Methyl ethyl ketone (MEK), Dimethoxymethane (DMM) and Diethoxymethane (DEM) were the least toxic. Moreover, both in vitro micronucleus assay and OECD QSAR Toolbox evaluation suggested that the metabolites present higher genotoxicity than biofuel candidates themselves. Overall, our results proved that this green toxicology testing strategy is a useful tool for assessing ecotoxicity of biofuel candidates.

Using a high-throughput method in the micronucleus assay to compare animal-free with rat-derived S9.

This study presents a high-throughput (HTP) micronucleus assay in multi-well plates with an automated evaluation for risk assessment applications. The evaluation of genotoxicity via the micronucleus assays according to international guidelines ISO 21427-2 with Chinese hamster (Cricetulus griseus) V79 cells was the starting point to develop our methodology. A drawback of this assay is that it is very time consuming and cost intensive. Our HTP micronucleus assay in a 48-well plate format allows for the simultaneous assessment of five different sample-concentrations with additional positive, negative and solvent controls with six technical replicates each within a quarter of the time required for the equivalent evaluation using the traditional slide method. In accordance with the 3R principle, animal compounds should be replaced with animal-free alternatives. However, traditional cell culture-based methods still require animal derived compounds like rat-liver derived S9-fraction, which is used to simulate the mammalian metabolism in in vitro assays that do show intrinsic metabolization capabilities. In the present study, a recently developed animal-free biotechnological alternative (ewoS9R) was investigated in the new high-throughput micronucleus assay. In total, 12 different mutagenic or genotoxic chemicals were investigated to assess the potential use of the animal-free metabolization system (ewoS9R) in comparison to a common rat-derived product. Out of the 12 compounds, one compound did not induce micronuclei in any treatment and 2 substances showed a genotoxic potential without the need for a metabolization system. EwoS9R demonstrated promising potential for future applications as it shows comparable results to the rat-derived S9 for 6 of the 9 pro-genotoxic substances tested. The remaining 3 substances (2-Acetamidofluorene, Benzo[a]pyrene, Cyclophosphamide) were only metabolized by rat-derived S9. A potential explanation is that ewoS9R was investigated with an approx. 10-fold lower enzyme concentration and was only optimized for CYP1A metabolization that may be improved with a modified production procedure. Future applications of ewoS9R go beyond the micronucleus assay, but further research is necessary.

Genotoxicity of three biofuel candidates compared to reference fuels.

Global demand for alternative energy sources increases due to concerns regarding energy security and greenhouse gas emissions. However, little is known regarding the impacts of biofuels to the environment and human health even though the identification of such impacts is important to avoid biofuels leading to undesired effects. In this study mutagenicity and genotoxicity of the three biofuel candidates ethyl levulinate (EL), 2-methyltetrahydrofuran (2-MTHF) and 2-methylfuran (2-MF) were investigated in comparison to two petroleum-derived fuels and a biodiesel. None of the samples induced mutagenicity in the Ames fluctuation test. However, the Micronucleus assay revealed significant effects in Chinese hamster (Cricetulus griseus) V79 cells caused by the potential biofuels. 2-MF revealed the highest toxic potential with significant induction of micronuclei below 20.0 mg/L. EL and 2-MTHF induced micronuclei only at very high concentrations (>1000.0 mg/L). In regard to the genotoxic potential of 2-MF, its usage as biofuel should be critically discussed.

Aquatic toxicity of biofuel candidates on Daphnia magna.

The increasing need for carbon-neutral, low-emission transportation sector has led to the development of advanced biofuels with tailor-made production and combustion processes. Even though the large-scale deployment of these advanced biofuels also increases the risk for their release into the environment, their toxic potency remains largely unknown. To identify hazardous biofuel candidates as early as possible, the fuel development process can be expanded by "Green Toxicology". To demonstrate such early Green Toxicology testing, this study investigates the aquatic toxicity for the two biofuel candidates 2-methyltetrahydrofuran (2-MTHF) and 2-methylfuran (2-MF) on Daphnia magna. We performed the prolonged acute immobilisation assay (96 h) and the D. magna reproduction test. 2-MF induced acute effects on D. magna that were two orders of magnitude stronger than those of 2-MTHF. Furthermore, both substances affected the growth and reproductive output of D. magna in a 21 d reproduction test, with 2-MF already inducing effects with concentrations one to two orders of magnitude lower than those of 2-MTHF. Thus, our assessment of the aquatic toxicity suggests that further biofuel development should focus on 2-MTHF. Furthermore, the acute immobilisation test with D. magna was identified as a promising tool for a rapid and sensitive "Green Toxicology" screening of further biofuel candidates.

Comparative ecotoxicity of potential biofuels to water flea (Daphnia magna), zebrafish (Danio rerio) and Chinese hamster (Cricetulus griseus) V79 cells.

The ecotoxicity of two biofuel candidates (1­octanol and 2­butanone) was investigated by an integrative test strategy using three bioassays: the acute immobilisation test with water flea (D. magna), the fish embryo acute toxicity test with zebrafish (Danio rerio) and the in vitro micronucleus assay with Chinese hamster (Cricetulus griseus) V79 cells. The median effective concentration (EC50) values were 14.9±0.66mgL-1 for 1­octanol, and 2152.1±44.6mgL-1 for 2­butanone in the D. magna test. Both 1­octanol and 2­butanone caused teratogenic and lethal effects on zebrafish embryos, while exposure to 1­octanol significantly induced these effects at concentrations ≥2.0mgL-1. These results indicate that 1­octanol exert much higher ecotoxicity than 2­butanone to D. magna and zebrafish embryos. Moreover, both 1­octanol and 2­butanone did not cause significant genotoxic effects, while their metabolites significantly induced micronuclei in V79 cells. The present study proposed an integrative test approach to evaluate the potential ecotoxicity of biofuels using simple, quick and inexpensive bioassays.

Microscale In Vitro Assays for the Investigation of Neutral Red Retention and Ethoxyresorufin-O-Deethylase of Biofuels and Fossil Fuels.

Only few information on the potential toxic effectiveness of biofuels are available. Due to increasing worldwide demand for energy and fuels during the past decades, biofuels are considered as a promising alternative for fossil fuels in the transport sector. Hence, more information on their hazard potentials are required to understand the toxicological impact of biofuels on the environment. In the German Cluster of Excellence "Tailor-made Fuels from Biomass" design processes for economical, sustainable and environmentally friendly biofuels are investigated. In an unique and interdisciplinary approach, ecotoxicological methods are applied to gain information on potential adverse environmental effects of biofuels at an early phase of their development. In the present study, three potential biofuels, ethyl levulinate, 2-methyltetrahydrofuran and 2-methylfuran were tested. Furthermore, we investigated a fossil gasoline fuel, a fossil diesel fuel and an established biodiesel. Two in vitro bioassays, one for assessing cytotoxicity and one for aryl hydrocarbon receptor agonism, so called dioxin-like activity, as measured by Ethoxyresorufin-O-Deethylase, were applied using the permanent fish liver cell line RTL-W1 (Oncorhynchus mykiss). The special properties of these fuel samples required modifications of the test design. Points that had to be addressed were high substance volatility, material compatibility and low solubility. For testing of gasoline, diesel and biodiesel, water accommodated fractions and a passive dosing approach were tested to address the high hydrophobicity and low solubility of these complex mixtures. Further work has to focus on an improvement of the chemical analyses of the fuel samples to allow a better comparison of any effects of fossil fuels and biofuels.

Evaluations of combined zebrafish (Danio rerio) embryo and marine phytoplankton (Diacronema lutheri) toxicity of dissolved organic contaminants in the Ythan catchment, Scotland, UK.

A wide variety of organic contaminants including pesticides, polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) have previously been detected in surface waters in the river Ythan catchment, North East Scotland UK. While the concentrations detected were below Water Framework Directive Environmental Quality Standards (WFD-EQSs) environmental exposures to the diverse mixtures of contaminants, known and unknown, may pose chronic and/or sublethal effects to non target organisms. The present study assessed the embryo and algal toxicity potential of freely dissolved organic contaminants from the Ythan catchment using silicone rubber passive sampling devices (SR-PSDs) and miniaturised bioassay techniques. Zebrafish (Danio rerio) embryos and marine phytoplankton species (Diacronema lutheri) were exposed to extracts from SR-PSDs deployed at different locations along the river Ythan and an undeployed procedural blank. Statistically significant developmental and algal toxicities were measured in all tests of extracts from deployed samples compared with the procedural blanks. This indicates environmental exposure to, and the combined toxicity potential of, freely dissolved organic contaminants in the catchment. The present and previous studies in the Ythan catchment, coupling SR-PSDs and bioassay techniques, have both helped to understand the interactions and combined effects of dissolved organic contaminants in the catchment. They have further revealed the need for improvement in the techniques currently used to assess environmental impact.

Prolonged conversion of n-butyrate to n-butanol with Clostridium saccharoperbutylacetonicum in a two-stage continuous culture with in-situ product removal.

n-Butanol was produced continuously in a two-stage fermentor system with integrated product removal from a co-feed of n-butyric acid and glucose. Glucose was always required as a source of ATP and electrons for the conversion of n-butyrate to n-butanol and for biomass growth; for the latter it also served as a carbon source. The first stage generated metabolically active planktonic cells of Clostridium saccharoperbutylacetonicum strain N1-4 that were continuously fed into the second (production) stage; the volumetric ratio of the two fermentors was 1:10. n-Butanol was removed continuously from the second stage via gas stripping. Implementing a two-stage process was observed to dramatically dampen metabolic oscillations (i.e., periodical changes of solventogenic activity). Culture degeneration (i.e., an irreversible loss of solventogenic activity) was avoided by periodical heat shocking and re-inoculating stage 1 and by maintaining the concentration of undissociated n-butyric acid in stage 2 at 3.4 mM with a pH-auxostat. The system was successfully operated for 42 days during which 93% of the fed n-butyrate was converted to n-butanol at a production rate of 0.39 g/(L × h). The molar yields Y(n-butanol/n-butyrate) and Y(n-butanol/glucose) were 2.0, and 0.718, respectively. For the same run, the molar ratio of n-butyrate to glucose consumed was 0.358. The molar yield of carbon in n-butanol produced from carbon in n-butyrate and glucose consumed (Y(n-butanol/carbon) ) was 0.386. These data illustrate that conversion of n-butyrate into n-butanol by solventogenic Clostridium species is feasible and that this can be performed in a continuous system operating for longer than a month. However, our data also demonstrate that a relatively large amount of glucose is required to supply electrons and ATP for this conversion and for cell growth in a continuous culture.