[show abstract][hide abstract] ABSTRACT: Despite the fact that more than 5000 safety-related studies have been published on bisphenol A (BPA), there seems to be no resolution of the apparently deadlocked controversy as to whether exposure of the general population to BPA causes adverse effects due to its estrogenicity. Therefore, the Advisory Committee of the German Society of Toxicology reviewed the background and cutting-edge topics of this BPA controversy. The current tolerable daily intake value (TDI) of 0.05 mg/kg body weight [bw]/day, derived by the European Food Safety Authority (EFSA), is mainly based on body weight changes in two- and three-generation studies in mice and rats. Recently, these studies and the derivation of the TDI have been criticized. After having carefully considered all arguments, the Committee had to conclude that the criticism was scientifically not justified; moreover, recently published additional data further support the reliability of the two- and three-generation studies demonstrating a lack of estrogen-dependent effects at and below doses on which the current TDI is based. A frequently discussed topic is whether doses below 5 mg/kg bw/day may cause adverse health effects in laboratory animals. Meanwhile, it has become clear that positive results from some explorative studies have not been confirmed in subsequent studies with higher numbers of animals or a priori defined hypotheses. Particularly relevant are some recent studies with negative outcomes that addressed effects of BPA on the brain, behavior, and the prostate in rodents for extrapolation to the human situation. The Committee came to the conclusion that rodent data can well be used as a basis for human risk evaluation. Currently published conjectures that rats are insensitive to estrogens compared to humans can be refuted. Data from toxicokinetics studies show that the half-life of BPA in adult human subjects is less than 2 hours and BPA is completely recovered in urine as BPA-conjugates. Tissue deconjugation of BPA-glucuronide and -sulfate may occur. Because of the extremely low quantities, it is only of minor relevance for BPA toxicity. Biomonitoring studies have been used to estimate human BPA exposure and show that the daily intake of BPA is far below the TDI for the general population. Further topics addressed in this article include reasons why some studies on BPA are not reproducible; the relevance of oral versus non-oral exposure routes; the degree to which newborns are at higher systemic BPA exposure; increased BPA exposure by infusions in intensive care units; mechanisms of action other than estrogen receptor activation; and the current regulatory status in Europe, as well as in the USA, Canada, Japan, New Zealand, and Australia. Overall, the Committee concluded that the current TDI for BPA is adequately justified and that the available evidence indicates that BPA exposure represents no noteworthy risk to the health of the human population, including newborns and babies.
Critical Reviews in Toxicology 04/2011; 41(4):263-91. · 6.25 Impact Factor
[show abstract][hide abstract] ABSTRACT: During the last two decades, substantial efforts have been made towards the development and international acceptance of alternative methods to safety studies using laboratory animals. In the EU, challenging timelines for phasing out of many standard tests using laboratory animals were established in the seventh Amending Directive 2003/15/EC to Cosmetics Directive 76/768/EEC. In continuation of this policy, the new European Chemicals Legislation (REACH) favours alternative methods to conventional in vivo testing, if validated and appropriate. Even alternative methods in the status of prevalidation or validation, but without scientific or regulatory acceptance may be used under certain conditions. Considerable progress in the establishment of alternative methods has been made in some fields, in particular with respect to methods predicting local toxic effects and genotoxicity. In more complex important fields of safety and risk assessment such as systemic single and repeated dose toxicity, toxicokinetics, sensitisation, reproductive toxicity and carcinogenicity, it is expected that the development and validation of in silico methods, testing batteries (in vitro and in silico) and tiered testing systems will have to overcome many scientific and regulatory obstacles which makes it extremely difficult to predict the outcome and the time needed. The main reasons are the complexity and limited knowledge of the biological processes involved on one hand and the long time frame until validation and regulatory acceptance of an alternative method on the other. New approaches in safety testing and evaluation using "Integrated Testing Strategies" (ITS) (including combinations of existing data, the use of chemical categories/grouping, in vitro tests and QSAR) that have not been validated or not gained wide acceptance in the scientific community and by regulatory authorities will need a thorough justification of their appropriateness for a given purpose. This requires the availability of knowledge and experience of experts in toxicology. The challenging deadlines for phasing out of in vivo tests in the Cosmetics Amending Directive 2003/15/EC appear unrealistic. Likewise, we expect that the application of validated alternative methods will only have a small or moderate impact on the reduction of in vivo tests under the regimen of REACH, provided that at least the same level of protection of human health as in the past is envisaged. As a consequence, under safety aspects, it appears wise to consider established in vivo tests to be indispensable as basic tools for hazard and risk assessment with respect to systemic single and repeated dose toxicity, sensitisation, carcinogenicity and reproductive toxicity, especially regarding quantitative aspects of risk assessment such as NOAELs, LOAELs and health-related limit values derived from them. Based on the overall evaluation in this review, the authors are of the opinion that in the short- and mid-term, the strategy of the development of alternative methods should be more directed towards the refinement or reduction of in vivo tests. The lessons learnt during these efforts will provide a substantial contribution towards the replacement initiatives in the long-term.
Archive für Toxikologie 05/2008; 82(4):211-36. · 5.22 Impact Factor
[show abstract][hide abstract] ABSTRACT: Currently, comprehensive toxicological data are available only for a small percentage of the 30,000 substances produced in volumes of 1–100 tons per year in the EU. Substances with inadequate safety data sets may pose a risk to employees, consumers and the environment. To improve this unsatisfactory situation the European Commission put forward a draft concept that will probably become law in 2006. The acronym of this concept is REACH standing for Registration, Evaluation and Authorization of Chemicals.
The aim of REACH is to systematically evaluate the risk of approximately 30,000 chemical substances produced, used or imported in quantities of 1–100 tons per year. From a practical point of view the testing requirements for these chemicals are one of the most
important parts of the REACH proposal. The latter progressively increase with the volume of chemical substances, including, e.g., acute, subchronic and chronic toxicity tests. Without doubt REACH will provide an important contribution to health protection for
workers and consumers. But perhaps even more importantly,REACHoffers an opportunity to optimize and innovate testing strategies for chemicals. Such novel techniques are in particular RNA expression profiling, proteome analysis and metabonomics to describe
alterations in gene or protein expressions patterns or in metabolite concentrations in response to toxic stimuli. Promising data have been published indicating that these techniques might identify hepato- or nephrotoxic compounds or even carcinogens differentiating between genotoxic and non-genotoxic substances. However, so far only a relatively small number of selected typical substances with well known toxic mechanisms has been tested. Therefore, the most promising innovative techniques should be optimized and validated by investigating a series of other typical but also untypical substances. In a further step a supplementary research program to REACH should be launched including promising innovative techniques (e.g. genomics, proteomics, metabonomics) but also other alternative methods (e.g. in vitro or QSAR), concentrating on the same substances that have to be tested by conventional animal studies in the mandatory part of REACH. In the present review we summarize key features of REACH, and discuss possibilities for the development of improved techniques and integrated strategies for toxicity testing.
[show abstract][hide abstract] ABSTRACT: Genetic polymorphisms of important xenobiotic metabolizing enzymes will be presented and the significance in toxicology, occupational health and environmental medicine will be discussed.
At present, only a very few associations have been established for particular polymorphisms and diseases, such as the slow acetylator phenotype or genotype and bladder cancer in smokers or occupationally exposed subjects. Furthermore, the GSTM1 null genotype is associated with a modest increase in the risk of bladder cancer. There is no scientific evidence for the assumption, that a screening for the known polymorphisms of xenobiotic metabolising enzymes is necessary for individual safety measures against environmental pollutants.
Umweltmedizin in Forschung und Praxis 01/2002; 7(4):232-246.
[show abstract][hide abstract] ABSTRACT: The tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) induces primarily lung tumors, which are assumed to derive from malignant transformation of alveolar type II (AII) cells within the lung. To elicit its carcinogenic effects, NNK requires metabolic activation by cytochrome P-450 (CYP)-mediated alpha-hydroxylation. Therefore, in this study the metabolism of NNK and expression of the NNK-activating CYP isoform CYP2B1 were investigated in primary cultures of rat AII cells. Although basal expression of CYP2B1 decreased in a time-dependent manner during culture of AII cells, substantial CYP2B1 protein expression was observed in AII cell cultures after the first 24 h. When AII cells were incubated with 0. 05 microM [5-(3)H]NNK, N-oxidation of NNK, which is thought to represent a detoxification pathway, was predominant (42%). alpha-Hydroxylated metabolites resulting from metabolic activation of NNK amounted to 35% of all detected metabolites. However, the proportion of alpha-hydroxylated metabolites decreased to 17% of all detected metabolites when AII cells were incubated with a 100-fold higher concentration of NNK (5 microM). In summary, this study indicates a remarkable activity of cultured AII cells to metabolize NNK, leading to substantial metabolic activation of NNK, which was more pronounced in incubations at low NNK concentration. Because exposure to NNK via cigarette smoking is thought to lead to very low plasma NNK concentrations (1-15 pM), these data suggest that metabolic activation of NNK in cigarette smokers might occur to a larger extent than would be expected according to previous metabolic studies performed with high (micromolar) NNK concentrations.
Drug Metabolism and Disposition 03/2000; 28(2):180-5. · 3.36 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mammalian liver exhibits expression of members of the family of multidrug resistance (mdr) transporters (P-glycoproteins). P-glycoprotein isoforms encoded by mdr1 genes participate in extrusion of an array of xenobiotics into the bile. Induction of mdr1b mRNA expression has been shown to occur in rat hepatocytes in response to hepatotrophic growth factors. As the cytokine tumor necrosis factor alpha (TNF-alpha) is known to exert a direct mitogenic effect on hepatocytes, its influence on mdr1b expression was investigated. In primary rat hepatocytes cultured in the absence of TNF-alpha, a time-dependent increase in basal expression of mdr1b mRNA and in immunodetectable P-glycoprotein was observed. In cells treated with TNF-alpha (4,000 U/ml) for 3 days, expression of mdr1b mRNA and of immunodetectable P-glycoprotein was induced approximately twofold. Moreover, intracellular steady-state levels of the mdr1 substrate rhodamine 123 were decreased in cells pretreated with TNF-alpha in comparison to controls, indicating an increase in functional transporter(s) mediating dye extrusion. Treatment of hepatocytes with antioxidants (1 mM ascorbic acid and 2% dimethyl sulfoxide) for 3 days markedly suppressed mdr1b mRNA and P-glycoprotein expression both in cells cultured in the presence of TNF-alpha and in the absence of the cytokine, but did not fully abolish mdr1b mRNA induction by TNF-alpha, supporting the notion that reactive oxygen species participate in regulation of basal mdr1b gene expression during hepatocyte culture. In conclusion, the present data indicate that by inducing mdr1b expression in hepatocytes, TNF-alpha may affect the capacity of the liver for extrusion or detoxification of endogenous or xenobiotic mdr1 substrates.
Journal of Cellular Physiology 10/1998; 176(3):506-15. · 4.22 Impact Factor
[show abstract][hide abstract] ABSTRACT: The scope of the present study was to investigate whether nicotine or cotinine will affect the metabolism of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in isolated perfused rat lungs and livers and to study the effect of starvation on pulmonary metabolism of NNK. NNK metabolism was investigated in isolated perfused liver and lung of male F344 rats perfused with 35 nM [5-3H]NNK in presence of a 1400-fold excess of the main tobacco alkaloid nicotine and its metabolite cotinine. In perfused rat livers, nicotine and cotinine inhibited NNK elimination and metabolism and led to a substantial increase of elimination half-life from 14.6 min in controls to 25.5 min after nicotine and 36.6 min after cotinine co-administration, respectively. In parallel, the pattern of NNK metabolites was changed by nicotine and cotinine. The pathway of alpha-hydroxylation representing the metabolic activation of NNK was decreased to 77% and 85% of control values, whereas N-oxidation of NNK and glucuronidation of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) was increased 2.6- and 1.2-fold in presence of nicotine and cotinine, respectively. When isolated rat lungs were perfused with 35 nM NNK for 3 h neither the elimination nor the pattern of metabolites were substantially affected due to co-administration of 50 microM nicotine or cotinine. Cytochrome P450 2E1 is known to participate in the activation of NNK and can be induced by starvation. However, isolated rat lungs from male Sprague Dawley rats perfused with [1-14C]NNK at about 2 microM for 3 h, revealed only small differences in pulmonary elimination and pattern of NNK metabolites between fed and starved animals. These results suggest that nicotine and its main metabolite cotinine inhibit the metabolic activation of NNK predominantly in the liver whereas activation in lung, a main target organ of NNK induced carcinogenesis, remained almost unaffected.
Archiv für Experimentelle Pathologie und Pharmakologie 03/1998; 357(3):344-50. · 2.15 Impact Factor
[show abstract][hide abstract] ABSTRACT: The tobacco specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is a strong lung carcinogen in all species tested. To elicit its tumorigenic effects NNK requires metabolic activation which is supposed to take place via alpha-hydroxylation, whereas N-oxidation is suggested to be a detoxification pathway. The differences in the organ specific metabolism of NNK may be crucial for the organotropy in NNK-induced carcinogenesis. Therefore, metabolism of NNK was investigated in the target organ lung and in liver of Fischer 344 (F344) rats using the model of isolated perfused organs. High activity to metabolize 35 nM [5-3H]NNK was observed in both perfused organs. NNK was eliminated by liver substantially faster (clearance 6.9 +/- 1.6 ml/min, half-life 14.6 +/- 1.2 min) than by lung (clearance 2.1 +/- 0.5 ml/min, half-life 47.9 +/- 7.4 min). When the clearance is calculated for a gram of organ or for metabolically active cell forms, the risk with respect to carcinogenic mechanisms was higher in lung than in liver. The metabolism of NNK in liver yielded the two products of NNK alpha-hydroxylation, the 4-oxo-4-(3-pyridyl)-butyric acid (keto acid) and 4-hydroxy-4-(3-pyridyl)-butyric acid (hydroxy acid). In lung, the major metabolite of NNK was 4-(methylnitrosamino)-1-(3-pyridyl-N-oxide)-1-butanone (NNK-N-oxide). Substantial amounts of metabolites formed from methyl hydroxylation of NNK, which is one of the two possible pathways of alpha-hydroxylation, were detected in lung but not in liver perfusion. Formation of these metabolites (4-oxo-4-(3-pyridyl)-butanol (keto alcohol), and 4-hydroxy-4-(3-pyridyl)-butanol (diol) can give rise to pyridyloxobutylating of DNA. When isolated rat livers were perfused with 150 microM NNK, equal to a dosage which is sufficient to induce liver tumors in rat, glucuronidation of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) was increased when compared to the concentration of 35 nM NNK. Nevertheless, the main part of NNK was also transformed via alpha-hydroxylation for this high concentration of NNK.
Archiv für Experimentelle Pathologie und Pharmakologie 03/1998; 357(3):336-43. · 2.15 Impact Factor
[show abstract][hide abstract] ABSTRACT: The mRNA expression of members of two cytochrome P450 (CYP) gene subfamilies involved in carcinogen activation, the CYP1A1/2 and CYP2B1 forms, was determined in primary rat hepatocyte cultures in response to metyrapone and to the inducer phenobarbital or 5,6-benzoflavone (BNF), respectively. Incubation of cells with 0.5 mM metyrapone resulted in accumulation of CYP1A1 and CYP1A2 mRNA and in a marked increase in CYP1A-associated enzymatic activity as determined by deethylation of ethoxyresorufin. Metyrapone and phenobarbital in combination acted synergistically in elevation of ethoxyresorufin O-deethylase activity. In hepatocytes treated with metyrapone or with phenobarbital, accumulation of CYP2B1 mRNA levels preceded an increase in CYP2B-associated, pentoxyresorufin O-depentylase activity. However, CYP2B1 mRNA levels were first detectable after 24 hours of treatment with phenobarbital, whereas metyrapone elicited a substantial increase in mRNA levels within 14 hours, suggesting differing mechanisms leading to accumulation of CYP2B1 mRNA under the two inducers.
Research communications in molecular pathology and pharmacology 11/1996; 94(1):47-61.
[show abstract][hide abstract] ABSTRACT: We observed the in vivo kinetics of bupivacaine in the cardiopulmonary system, particularly in the pulmonary artery, the upper part of the descending aorta and the coronary sinus of anaesthetized sheep, each of which received a high dose infusion into the central vein. In some experiments dilution curves were monitored for the non-extracted dye, indocyanine green. Concentrations of bupivacaine were approximately 20% lower in the aorta than in the pulmonary artery. This gradient of bupivacaine was present across the lung for 5-10 min. Concentrations of bupivacaine in the coronary venous plasma were also markedly lower than at the arterial site. Initially more than 50% of the amount of bupivacaine at the arterial site was removed by the heart. Later, the myocardial extraction ratio decreased and plateaued at a value of 0.30-0.40. At this time, concentrations of bupivacaine in the pulmonary artery were approximately 12 micrograms ml-1. Therefore, approximately 0.3-0.6 mg of bupivacaine were extracted per minute by the sheep heart in vivo. On the other hand, isolated perfused rat hearts did not substantially remove bupivacaine (2 micrograms ml-1) from the medium. Approximately one-third of 14C-bupivacaine was retained in slices of rat and sheep myocardial tissue. However, there was no evidence that metabolism played a substantial role in the cardiac kinetics of bupivacaine.
BJA British Journal of Anaesthesia 09/1996; 77(2):257-64. · 4.24 Impact Factor
[show abstract][hide abstract] ABSTRACT: P-glycoproteins encoded by members of the mdr gene family function as membrane-situated transport proteins, isoforms of which are involved in conferring a form of multidrug resistance by participating in secretion of various xenobiotics. In primary rat hepatocytes maintained in serum-free culture, accumulation of immunodetectable P-glycoprotein and mdr1b mRNA occurred in a time-dependent manner and was accompanied by a substantial decrease in retention of the mdr1 substrate rhodamine 123. However, incubation of cells with epidermal growth factor (EGF) or with insulin-like growth factor I (IGF-I) markedly enhanced time-dependent accumulation of P-glycoprotein and mdr1b mRNA. Furthermore, EGF-treated cells exhibited decreased intracellular rhodamine 123 retention, an effect partially inhibited by the chemosensitizer verapamil. These data suggest that an increase in (a) functional transporter(s) eliciting transport of mdr1 substrates occurs under EGF.
Biochemical and Biophysical Research Communications 11/1995; 215(1):179-85. · 2.41 Impact Factor
[show abstract][hide abstract] ABSTRACT: The pulmonary first-pass kinetics of the amide-linked local anaesthetics prilocaine, mepivacaine and bupivacaine were studied in 33 patients after a single epidural injection. Drug concentrations were monitored before and after lung passage, i.e. in samples withdrawn simultaneously from mixed venous and arterial blood. In most cases, maximum plasma concentrations were observed 10 min after injection (range 2 to 30 min). Two min after injection the local anaesthetics were distinctly extracted by the lung (prilocaine 40%, mepivacaine 20%, and bupivacaine 12%). Prilocaine was retained by the lung more effectively than bupivacaine and mepivacaine. However, a transpulmonary concentration gradient could be observed only for a short time, i.e. maximum 15 min. Altogether, in the case of accidental fast absorption, e.g. inadvertent intravenous injection, arterial peak concentrations of these drugs will be attenuated by passage of the lung. However, the lung will not substantially lower the risk of toxicity by amide-linked local anaesthetics during normal conditions of regional anaesthesia where slow absorption occurs.
[show abstract][hide abstract] ABSTRACT: Prilocaine is assumed to undergo significant elimination by extrahepatic organs and to differ in this respect from other commonly used local anaesthetics. In order to clarify whether the lung may play an important role as a site of elimination of prilocaine, the kinetic parameters were studied in isolated perfused rat lungs and were compared to those of isolated livers. Furthermore, the structurally related compounds bupivacaine and mepivacaine were also investigated in this system. Prilocaine was dispersed into a relatively large apparent distribution volume in perfused rat lung (139 ml versus 97 ml in controls). In single-pass perfused lungs the observed maximum of concentration was decreased by about 60% compared to controls. The mean residence time was prolonged by about 40%. These observations suggest that prilocaine is substantially retained by rat lung and that this effect occurs particularly during first-pass. However, the ability of rat lung to degrade prilocaine was relatively low. The clearance values were about 0.3 ml/min equal to about 20% of the hepatic capacity calculated per g of tissue. Thus it must be assumed that prilocaine is only transiently retained by the lung and will gain systemic availability later on. In rat lungs the kinetics of prilocaine elimination were not substantially different from those of bupivacaine and mepivacaine (16 and 12%). These observations do not support the assumption that especially prilocaine undergoes extrahepatic elimination.
Archiv für Experimentelle Pathologie und Pharmakologie 02/1995; 351(1):93-8. · 2.15 Impact Factor
[show abstract][hide abstract] ABSTRACT: The ability of rat lung to remove the local anaesthetic drug bupivacaine from the blood was studied in isolated organs which were perfused either in an open (single-pass mode) or in a closed system (recirculating medium). Isolated perfused rat lungs exhibited a very low capacity to metabolize bupivacaine within 3 h during which the drug circulated continuously through the organ. The clearance values differed only by 0.2 ml/min from the control parameters in sham perfusions. The calculated extraction ratio was 0.2% and the elimination half-life was about 210 min. The volume of distribution of bupivacaine was 133 ml which remarkably surmounted the reference values obtained for sham perfusions. The distribution of bupivacaine into the pulmonary tissue was investigated applying the multiple indicator dilution technique to isolated lungs perfused in the single-pass mode. The mean elimination time of model compounds for distribution into the intravascular space, 14C-insulin, and the total water space, 3H-water, were 68 and 75 s at a flow rate of 6 ml/min. The volume of distribution was 5.9 ml for inulin and 6.5 ml for water. The mean transit time for concomitantly injected bupivacaine was 221 s and the volume of distribution was 14.4 ml. The respective parameters of sham perfusions performed without an isolated organ were substantially lower, i.e. mean elimination time 50, 50 and 61 s and distribution volume 4.9, 5.0 and 6.1 ml for inulin, water and bupivacaine.2+ f1p4
Archiv für Experimentelle Pathologie und Pharmakologie 02/1995; 351(1):99-106. · 2.15 Impact Factor
[show abstract][hide abstract] ABSTRACT: The effect of the new phosphodiesterse inhibitor R80122 (E)-N-cyclohexyl-N-methyl-2-[[[phenyl(1,2,3,5-tetrahydro-2-oxoimidazo [2,1-b]-quinazolin-7-yl)methylene] amino]oxy]acetamide, (CAS 133718-29-3) on haemodynamic parameters and myocardial oxygen consumption were intraindividually compared with those of enoximone, a clinically established phosphodiesterase inhibitor. In 12 anaesthetised sheep the drugs were given in randomized order as i. v. infusions for 6 min at each setting (10, 20 and 30 micrograms.kg-1.min-1 (R80122) and 32, 64 and 96 micrograms.kg-1.min-1 (enoximone)). R 80122 as well as enoximone caused a significant increase in cardiac inotropism with a simultaneous increase of myocardial oxygen consumption. The peripheral resistance was significantly decreased by both drugs. The haemodynamic effects elicited by the application of equieffective doses of R80122 and enoximone did not show any differences.