Evaluating the effects of anticoagulant rodenticide bromadiolone in Wistar rats co-exposed to vitamin K: impact on blood-liver axis and brain oxidative status.

The aim of this study was to investigate the beneficial effects of vitamin K relate to protection against detrimental effects of bromadiolone. Wistar rats (n = 30) were divided in three groups (n = 10): control group and two groups treated with bromadiolone (0.12 mg/kg) and bromadiolone + vitamin K (0.12 mg/kg + 100 mg/kg) over the period of four days. The main findings in the bromadiolone-exposed rats, such as damaged hepatocytes, high levels of globulin, total proteins and lymphocytes, and altered albumin/globulin ratio, collectively indicate an acute inflammatory process. Morphological changes in erythrocytes include microcytosis, hypochromia, hyperchromia, hemolysis, stomatocytosis, and spherocytosis. Significantly low values of RBC, Hct, and hemoglobin concentrations indicate impairments of the hematopoietic pathway causing combined anemia. The selected dose of bromadiolone caused a non-significant increase of catalase activity and a significant increase of the total protein content in brain tissue homogenates. Vitamin K supplementation reduced many of the harmful effects of bromadiolone. The cytoprotective role of vitamin K was proved to be of great importance for the preservation of structural changes on the membranes of hepatocytes and erythrocytes, in addition to the known role in the treatment of coagulopathies. The results of the study suggest valuable properties of vitamin K in the prevention and treatment of various types of anemia caused by bromadiolone toxicity. Future research is necessary to determine the adequate dose and treatment duration with vitamin K in disorders caused by the cumulative action of bromadiolone and possibly other pesticides.

Comparative pharmacokinetics of difethialone stereoisomers in male and female rats and mice: development of an intra- and inter-species model to predict the suitable formulation mix.

The ecotoxicity of anticoagulants used for rodent pests' management is a major concern, particularly with second generation anticoagulants, which are more persistent in the body of rodents and therefore more likely to cause secondary exposure in their predators. One of the solutions envisaged to mitigate this risk is to use stereoisomers of these anticoagulants, each of which has particular pharmacokinetics. However, the few studies published to date have considered only one species and one sex. Here, we study the pharmacokinetics of the 4 stereoisomers of 3.4 mg/kg of difethialone in rats (Rattus norvegicus) and 3 mg/kg in mice (Mus musculus) in both sexes and propose a model to choose the optimal stereoisomer efficacy/ecotoxicity mixture for the management of all these animals. Our results show that while the most persistent stereoisomer (E3-cis) is common to both species and sexes, the pharmacokinetics of the other stereoisomers show marked differences between sexes and species. Thus, the area under curve (AUC) of E4-trans in male rats is four times lower than in females or mice, making it a priori unusable in male rats. Conversely, our modeling seems to show that the E1-trans stereoisomer seems to offer the best compromise AUC persistence. In conclusion, we highlight that studies on anticoagulants must necessarily integrate research on the effect of gender and species both on efficacy and with regard to the ecotoxicity of these molecules.

Mass spectrometry characterization of anticoagulant rodenticides and hydroxyl metabolites.

RATIONALE: Anticoagulant rodenticides (ARs) are used worldwide for rodent population control to protect human health and biodiversity, and to prevent agricultural and economic losses. Rodents may develop a metabolic resistance to ARs. In order to help understand such metabolic resistance, mass spectrometry was used to position the hydroxylated group of hydroxyl metabolites of second-generation ARs (SGARs). METHODS: Most AR pesticides are derived from the 4-hydroxycoumarin/thiocoumarin family. We used low-resolution and high-resolution mass spectrometry to understand the fragmentation pathways of the ARs and their respective metabolites, and to better define the structure of their tandem mass spectrometry product ions. RESULTS: Seven specific product ions were evidenced for five ARs, with their respective chemical structures. Those ions were obtained as well from the mass spectra of the hydroxyl metabolites of four SGARs, difenacoum (DFM), brodifacoum (BFM), difethialone (DFTL) and flocoumafen (FLO), with different positions of the hydroxyl group. CONCLUSIONS: The differences in chemical structure between DFM on the one hand and BFM, FLO and DFTL on the other could explain the differences in bioavailability between these two groups of molecules. The defined product ions will be used to investigate the part played by the metabolic issue in the field resistance of SGARs.

[Study on the pharmacokinetics of bromadiolone in oral poisoning rabbits].

Objective: To investigate the variations of bromadiolone concentration in blood and its metabolism in rabbits after oral administration of bromadiolone, and to provide reference for the study of bromadiolone metabolism. Methods: Designed absolute alcohol (1 g/kg) reagent control group, high dose (0.3 mg/kg) and low dose group (0.05 mg/kg) , there were 6 rabbits in each group. Blood samples were collected from the rabbit central auricular artery at regular intervals as 1 h, 2 h, 4 h, 8 h, 12 h, 16 h, 24 h, 36 h, 48 h, 72 h, 168 h, 336 h, 504 h after oral administration. The samples were centrifuged within 1 h. Prothrombin time (PT) , activated partial thromboplastin time (APTT) and concentrations of bromadiolone in plasma were tested. Metabolic kinetics data was analyzed by DAS 3.0.2 software. Results: Bromadiolone had no significant effect on the body weight of the experimental rabbits during the experimental period (P>0.05) . PT and APTT were significantly abnormal in different dose groups, but for occurrence of exception, PT was earlier than APTT. The concentration of bromadiolone in plasma reached the peak value 12 h after gavage in both high-dose and low-dose groups. The absorption time of t(1/2Ka) in high-dose group was 4.34 h, the clearance time of t(1/2) was 81.52 h, the absorption time of t(1/2Ka) in low-dose group was 6.90 h, and the elimination time of t(1/2) was 56.38 h. The atrioventricular model of bromadidone was three compartment model in rabbits. Conclusion: Bromadiolone can be absorbed rapidly by oral administration, but its metabolism is slow. The change of bromadiolone in vivo accords with the three compartment model.

Recovery of brodifacoum in vomitus following induction of emesis in dogs that had ingested rodenticide bait.

AIM: To assess the benefit of inducing emesis in dogs that have ingested rodenticide bait containing brodifacoum (BDF), by determining the amount of BDF in bait recovered from the vomitus relative to the estimated amount consumed. METHODS: Between 2014 and 2015 samples of vomitus from seven dogs that ingested rodenticide baits containing BDF were submitted by veterinarians in New Zealand. All seven dogs had been given apomorphine by the veterinarian and vomited within 1 hour of ingesting the bait. Some or all of the bait particles were retrieved from each sample and were analysed for concentrations of BDF using HPLC. Based on estimations of the mass of bait consumed, the concentration of BDF stated on the product label, and the estimated mass of bait in the vomitus of each dog, the amount of BDF in the vomited bait was calculated as a percentage of the amount ingested. RESULTS: For five dogs an estimation of the mass of bait ingested was provided by the submitting veterinarian. For these dogs the estimated percentage of BDF in the bait retrieved from the vomitus was between 10-77%. All dogs were well after discharge but only one dog returned for further testing. This dog had a normal prothrombin time 3 days after ingestion. CONCLUSIONS AND CLINICAL RELEVANCE: The induction of emesis within 1 hour of ingestion can be a useful tool in reducing the exposure of dogs to a toxic dose of BDF. The BDF was not fully absorbed within 1 hour of ingestion suggesting that the early induction of emesis can remove bait containing BDF before it can be fully absorbed.

Development of an Ecofriendly Anticoagulant Rodenticide Based on the Stereochemistry of Difenacoum.

Difenacoum, an antivitamin K anticoagulant, has been widely used as rodenticide to manage populations of rodents. Difenacoum belongs to the second generation of anticoagulant, and, as all the molecules belonging to the second generation of anticoagulant, difenacoum is often involved in primary poisonings of domestic animals and secondary poisonings of wildlife by feeding contaminated rodents. To develop a new and ecofriendly difenacoum, we explored in this study the differences in properties between diastereomers of difenacoum. Indeed, the currently commercial difenacoum is a mixture of 57% of cis-isomers and 43% of trans-isomers. Cis- and trans-isomers were thus purified on a C18 column, and their respective pharmacokinetic properties and their efficiency to inhibit the coagulation of rodents were explored. Tissue persistence of trans-isomers was shown to be shorter than that of cis-isomers with a half-life fivefold shorter. Efficiency to inhibit the vitamin K epoxide reductase activity involved in the coagulation process was shown to be similar between cis- and trans-isomers. The use of trans-isomers of difenacoum allowed to drastically reduce difenacoum residues in liver and other tissues of rodents when the rodent is moribund. Therefore, secondary poisonings of wildlife should be decreased by the use of difenacoum largely enriched in trans-isomers.

Water vole management - Could anticoagulant rodenticides stereochemistry mitigate the ecotoxicity issues associated to their use?

Cyclic water vole population explosions can be controlled in some European countries with anticoagulant rodenticides leading sometimes to wildlife poisonings due to the toxin's tissue persistence. Here, we analyzed the pharmacokinetics of rodenticide residues in voles and we explored potential ways of improving the mass application of these agents based on the concept of stereoisomers. We demonstrated the dramatic persistence of bromadiolone in vole tissues with a hepatic half-life of about 10-30 days, while the tissue persistence of chlorophacinone is rather short with a hepatic half-life of about one day. The dramatic persistence of bromadiolone is due to the trans-isomer group (the major compound in bromadiolone), while the cis-isomer group has a short half-life. Because of resistance to chlorophacinone, the cis-bromadiolone isomers may constitute an excellent compromise between efficacy and ecotoxicological risk to control voles. A mathematical model is proposed to favor the development of baits mixed with cis-isomer groups.

Effects of vitamin K1 treatment on plasma concentrations of long-acting anticoagulant rodenticide enantiomers following inhalation of contaminated synthetic cannabinoids.

Background: An outbreak of synthetic cannabinoid (SC)-associated coagulopathy and bleeding in Illinois, USA was determined to be due to inhalation of SC contaminated with brodifacoum (BDF), difenacoum (DiF), and bromadiolone (BDL), highly potent long-acting anticoagulant rodenticides (LAARs). Treatment with high-dose vitamin K1 (VK1) prevented mortality; however, plasma LAAR levels were not measured risking recurrence of coagulopathy and bleeding due to premature discontinuation. The goal of this study was to determine if plasma LAAR levels were reduced following standard of care treatment to normalize coagulopathy.Methods: Blood samples were collected from a cohort of 32 patients, and ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) analysis used to quantify plasma LAAR levels including enantiomers.Results: BDF was detected in 31 samples; 30 also contained DiF and 18 contained BDL. Initial plasma levels were 581 ± 87, 11.0 ± 1.9, and 14.9 ± 5.9 ng/mL for BDF, DiF, and BDL, respectively (mean ± SE). At discharge plasma, BDF levels remained elevated at 453 ± 68 ng/mL. Plasma half-lives for BDF, DiF, and BDL were 7.5 ± 1.3, 7.2 ± 1.9, and 1.8 ± 0.3 days, respectively. The half-life for trans-BDF enantiomers (5.7 ± 0.8 days) was shorter than for cis-enantiomers (7.6 ± 1.9 days). BDF half-lives were shorter, and coagulopathy normalized faster in patients receiving intravenous VK1 as compared to oral VK1. Patients prescribed VK1 at discharge had fewer re-admittances.Conclusions: These results demonstrate that plasma LAAR levels at discharge were elevated in poisoned patients despite normal coagulation, and that the route of VK1 administration affected LAAR pharmacokinetics and INR normalization. We propose plasma LAAR levels and coagulation be monitored concomitantly during follow-up of patients with LAAR poisoning. KEY POINTSIn patients treated with high-dose vitamin K1 for LAAR poisoning, plasma levels remained 40-fold above safe levels upon discharge from hospital.LAAR half-lives, normalization of coagulopathy, and readmittances were reduced by treatment with intravenous vitamin K1.

Determination of anticoagulant rodenticides in faeces of exposed dogs and in a healthy dog population.

BACKGROUND: Exposure to anticoagulant rodenticides (ARs) in dogs is among the most common causes of poisoning in small animal practice, but information about toxicokinetic of these rodenticides in dogs is lacking. We analysed blood and faeces from five accidentally exposed dogs and 110 healthy dogs by reversed phase ultra-high performance liquid chromatography-tandem mass spectrometry. The aim of the study was to estimate elimination of brodifacoum, bromadiolone and difenacoum after acute exposure, calculate the half-lives of these rodenticides in dogs, estimate faecal elimination in a litter of puppies born, and further to identify the extent of AR exposure in a healthy dog population. RESULTS: Three dogs were included after single ingestions of brodifacoum; two dogs ingested bromadiolone and one dog ingested difenacoum. Maximum concentrations in faeces were found after day 2-3 for all ARs. The distribution half-lives were 1-10 days for brodifacoum, 1-2 days for bromadiolone and 10 days for difenacoum. Brodifacoum and difenacoum had estimated terminal half-lives of 200-330 days and 190 days, respectively. In contrast, bromadiolone had an estimated terminal half-life of 30 days. No clinical signs of poisoning or coagulopathy were observed in terminal elimination period. In blood, the terminal half-life of brodifacoum was estimated to 8 days. Faeces from a litter of puppies born from one of the poisoned dogs were examined, and measurable concentrations of brodifacoum were detected in all samples for at least 28 days after parturition. A cross-sectional study of 110 healthy domestic dogs was performed to estimate ARs exposure in a dog population. Difenacoum was detected in faeces of one dog. Blood and faecal samples from the remaining dogs were negative for all ARs. CONCLUSIONS: Based on the limited pharmacokinetic data from these dogs, our results suggest that ARs have a biphasic elimination in faeces using a two-compartment elimination kinetics model. We have shown that faecal analysis is suitable and reliable for the assessment of ARs exposure in dogs and a tool for estimating the AR half-lives. Half-lives of ARs could be a valuable indicator in the exposed dogs and provides important information for veterinarians monitoring AR exposure and assessment of treatment length in dogs.

Differences in teratogenicity of some vitamin K antagonist substances used as human therapeutic or rodenticide are due to major differences in their fate after an oral administration.

All vitamin K antagonist active substances used as rodenticides were reclassified in 2016 by the European authorities as active substances "toxic for reproduction", using a "read-across" alternative method based on warfarin, a human vitamin K antagonist drug. Recent study suggested that all vitamin K antagonist active substances are not all teratogenic. Using a neonatal exposure protocol, warfarin evokes skeletal deformities in rats, while bromadiolone, a widely used second-generation anticoagulant rodenticide, failed to cause such effects. Herein, using a rat model we investigated the mechanisms that may explain teratogenicity differences between warfarin and bromadiolone, despite their similar vitamin K antagonist mechanism of action. This study also included coumatetralyl, a first-generation active substance rodenticide. Pharmacokinetic studies were conducted in rats to evaluate a potential difference in the transfer of vitamin K antagonists from mother to fetus. The data clearly demonstrate that warfarin is highly transferred from the mother to the fetus during gestation or lactation. In contrast, bromadiolone transfer from dam to the fetus is modest (5% compared to warfarin). This difference appears to be associated to almost complete uptake of bromadiolone by mother's liver, resulting in very low exposure in plasma and eventually in other peripheric tissues. This study suggests that the pharmacokinetic properties of vitamin K antagonists are not identical and could challenge the classification of such active substances as "toxic for reproduction".

The relative toxicity of brodifacoum enantiomers.

Brodifacoum (BDF) is a potent, long-acting anticoagulant rodenticide that can cause fatal poisoning in humans. The chemical structure of BDF includes 2 chiral carbons, resulting in 2 pairs of diastereomers, BDF-cis (R/S and S/R) and BDF-trans (R/R and S/S). However, the relative potency of these molecules is not known. The purpose of this study was to compare the in vitro and in vivo toxic effects of the 2 BDF diastereomer pairs. In adult Sprague-Dawley rats BDF-cis was significantly more toxic than BDF-trans (LD50 values of 219 versus 316 µg/kg, respectively) while racemic BDF had intermediate potency (266 µg/kg). In adult New Zealand white rabbits, BDF-cis had a longer half-life than BDF-trans which could contribute to its observed increased toxicity. Lastly, BDF-cis (10 µM), but not BDF-trans, damaged cultured SH-SY5Y human neuroblastoma cells by attenuating mitochondrial reductive capacity. Taken together, these data suggest that different toxic manifestations of BDF poisoning in mammals could be attributed, in part, to differences in relative enantiomer concentrations present in racemic formulations of this commercially-available toxicant.

Should Cytochrome P450 Inducers be Used to Accelerate Clearance of Brodifacoum from Poisoned Patients?

A recent multi-state outbreak of life-threatening bleeding following inhalation of synthetic cannabinoids has been attributed to contamination with the long-acting anticoagulant rodenticide (LAAR) brodifacoum, a second-generation, highly potent, long-acting derivative of the commonly used blood thinner warfarin. While long-term treatment with high-dose vitamin K1 restores coagulation, it does not affect brodifacoum metabolism or clearance, and, consequently, brodifacoum remains in the human body for several months, thereby predisposing to risk of bleeding recurrence and development of coagulation-independent injury in extrahepatic tissues and fetuses. This has prompted the evaluation of pharmacological measures that accelerate brodifacoum clearance from poisoned patients. Since the induction of certain cytochrome P450 (CYP) enzymes accelerates warfarin metabolism, using CYP inducers, such as phenobarbital, to accelerate brodifacoum clearance seems plausible. However, unlike warfarin, brodifacoum does not undergo significant metabolism in the liver, nor have the effects of phenobarbital on vitamin K1 metabolism been previously determined. In addition, the safety of phenobarbital in brodifacoum-poisoned patients has not been established. Therefore, we propose that CYP inducers should not be used to accelerate the clearance of brodifacoum from poisoned patients, but that alternative approaches such as reducing enterohepatic recirculation of brodifacoum, or using lipid emulsions to scavenge brodifacoum throughout the body, be considered.

Factors associated to adequate time in therapeutic range with oral vitamin K antagonists in Tunisia.

INTRODUCTION:   The quality of chronic anticoagulation and predictor factors of poor anticoagulant control in patients under acenocoumarol were unknown in North Africa. METHODS: It is an observational study, carried out between November 2015 and November 30, 2016. The international normalized ratio (INR) values were prospectively obtained, and TTR was calculated using the Rosendaal method. RESULTS: Overall, 215 patients were included in this study, with a mean age of 63±0,8 years. The prevalence of poor anticoagulation control was 78.1%; 95% CI [72.2-83.2] (168 patients with TTR less than 65%). The median TTR with the Rosendaal method was 44.4%. After multivariate adjustment, variables significantly associated with adequate anticoagulation level were: history of ischemic stroke (Adjusted OR equal to 4.3, 95% CI: 1.4-12.9), associated prescription of antiplatelet therapy (Adjusted OR equal to 3.5, 95% CI: 1.1-11.2), daily prescribed dose of coumarins less than 6 mg (Adjusted OR equal to 6.4, 95% CI: 1.1- 36) and lower risk of bleeding assessed as HAS-BLED score (Adjusted OR: 0.5, 95% CI: 0.3-0.8). CONCLUSION: The quality of anticoagulation management with VKA among outpatients who received acenocoumarol was suboptimal. Strategies should be undertaken by clinicians and patients to improve the quality of anticoagulation, to address challenges to adverse cardiovascular outcomes in individuals treated with chronic anticoagulation.

Pharmacogenomics of 4-hydroxycoumarin anticoagulants.

Oral anticoagulants of the 4-hydroxycoumarin class, typified by warfarin, are used worldwide to treat thromboembolic disease. These drugs show the beneficial attributes of high efficacy and low cost, but patient management can be complicated by their narrow therapeutic index and wide inter-individual variability in dosing. Our understanding of the latter complication has improved significantly in recent years due to intense investigation of genetic factors influencing drug pharmacokinetics (CYP2C9) and pharmacodynamic response (VKORC1). In particular, the discovery of polymorphisms in the VKORC1 gene that strongly impact oral anticoagulant dose has heightened expectations that genetic testing for a relatively small cadre of warfarin-response genes might substantially enhance patient care in this area, especially during the initiation phase of therapy. However, enthusiasm for genotype-based dosing of oral anticoagulants must be balanced against the ready availability of both a simple phenotypic test (prothrombin time) and an antidote to over-anticoagulation (vitamin K). Wide-spread acceptance of genetically based tests for establishing therapy with warfarin and its congeners will likely require additional evidence that such an approach offers protection against a variety of negative anticoagulation outcomes, especially severe bleeding, as well as offering utility across many racial populations. This article will review recent events in these and other related areas.

Kinetics of bromadiolone in rodent populations and implications for predators after field control of the water vole, Arvicola terrestris.

We document the kinetics of bromadiolone in two rodent populations after a field control of water voles, and their implications for predator exposure. Water voles and common voles were trapped aboveground and underground from 1 to 135 days after bromadiolone treatment in the field. Livers, digestive tracts, and rests of the body were analyzed separately. Our results indicate that 99.6% of the water voles trapped underground and 41% of the common voles trapped aboveground contain bromadiolone residues. Concentrations were maximal between 3.3 and 6.5 days after treatment, according to the tissues examined and the model applied for water voles, and after 1.3 to 3.7 days for common voles. Water voles appeared available almost exclusively for foraging predators. Common voles, found less likely to be poisoned and exhibiting weaker concentrations, were mainly sampled aboveground. The liver, primarily eaten by some predators and scavengers, contains a larger bromadiolone quantity (59% of the total amount found in water voles). The rejection of the digestive tract by those species may lead to a subsequent consumption of voles with higher bromadiolone concentrations (from +3.8 to +5.8% of concentration) and provide a moderate risk increase. After 135 days, eight of the ten water voles and one of the two common voles exhibited detectable residues. Additionally, one specimen presented higher concentrations than the others, and similar to those measured in Voles trapped between the first 15-20 days. This may have consequences on predator intoxications several months after treatment. These results integrate individual differences for the two main rodent species present in treated areas. Implications for predator exposure were investigated at the end of the study and suggest that, if the risk of secondary poisoning is maximal during the first 15-20 days when the rodent densities remain high, exposure conditions are maintained for at least 135 days.

Differential expression of cytochrome P450 genes between bromadiolone-resistant and anticoagulant-susceptible Norway rats: a possible role for pharmacokinetics in bromadiolone resistance.

BACKGROUND: Anticoagulant resistance in Norway rats, Rattus norvegicus (Berk.), has been suggested to be conferred by mutations in the VKORC1 gene, encoding the target protein of anticoagulant rodenticides. Other factors, e.g. pharmacokinetics, may also contribute to resistance, however. To examine the involvement of pharmacokinetics in bromadiolone resistance in male and female rats, liver expression profiles of seven cytochrome P450 genes from a Danish bromadiolone-resistant rat strain (with an Y139C-VKORC1 mutation) were compared with profiles from an anticoagulant-susceptible strain. RESULTS: In the presence of bromadiolone, the Cyp2e1, Cyp2c13, Cyp3a2 and Cyp3a3 genes were significantly overexpressed, while Cyp2c12 expression was suppressed in resistant female rats compared with susceptible females. Relative to susceptible males, resistant males showed significant overexpression of the Cyp2a1, Cyp2e1, Cyp3a2 and Cyp3a3 genes. On exposure to bromadiolone, females had higher Cyp2e1 expression than males, which possibly explains why female rats are generally more tolerant to anticoagulants than male rats. CONCLUSION: Results suggest that bromadiolone resistance in a Danish strain of Norway rats involves enhanced anticoagulant metabolism catalysed by cytochrome P450-2e1, -3a2 and -3a3. This pharmacokinetically based bromadiolone resistance is to some extent sex differentiated, as female resistance furthermore seems to involve overexpression of cytochrome P450-2c13 and suppression of P450-2c12, whereas male resistance appears to involve P450-2a1 overexpression.

Bromadiolone toxicokinetics: diagnosis and treatment implications.

INTRODUCTION: Ingestion of bromadiolone can lead to prolonged and life-threatening coagulopathy. Traditional treatment of bromadiolone intoxication relies on the coagulation profile. Currently, there is scanty information on bromadiolone elimination kinetics and half-life. CASE REPORT: We report a case of bromadiolone poisoning in a 40-year old female who, by history, ingested four 42.5-gram bags of rat poison (0.005% bromadiolone), equivalent to 8.5 mg bromadiolone (0.17 mg/kg body weight), four days prior to admission. On admission, her prothrombin time was 92.0 seconds, international normalized ratio was 5.7, and activated partial thromboplastin time was 50.2 seconds with no bleeding on clinical examination. The first plasma bromadiolone level (5 days post-ingestion) was 92 ng/mL. Serial measurement of plasma bromadiolone levels confirmed the diagnosis and demonstrated that bromadiolone obeys the elimination kinetic of a two-compartment model with a rapid, fairly steep decline phase (half-life 3.5 days) followed by a slower termination phase (half-life 24 days). Plasma bromadiolone level of less than 10 ng/mL in our patient was associated with a consistently normal coagulation profile without vitamin K1 therapy. CONCLUSIONS: There is a lack of information on the toxicodynamics and toxicokinetics of bromadiolone in humans; further studies are needed before the plasma bromadiolone level can serve as one of the logical and safe therapeutic endpoints for vitamin K1 therapy.

Accumulation of anticoagulant rodenticides (chlorophacinone, bromadiolone and brodifacoum) in a non-target invertebrate, the slug, Deroceras reticulatum.

Anticoagulant rodenticides (ARs) are used worldwide to control populations of agricultural and urban rodents, but these pesticides may be accumulated in and poisoned non-target species of wildlife. Slugs may feed on rodenticide bait following field applications. Thus, it can be assumed that their predators are exposed to rodenticides through food chain transfer. However, AR exposure in the slugs has not been systematically studied. We investigated the accumulation of three ARs (chlorophacinone, bromadiolone or brodifacoum) in the slug Deroceras reticulatum exposed for a period of 5days followed by depuration time of 4days in the laboratory. Moreover, we studied the exposure of slugs to brodifacoum in the field. In the laboratory exposure, the slugs consumed rodenticide baits, but no mortality was observed. After 1day, their concentrations were stable over the time and no differences were detected between the concentrations of the three ARs. After 5days of exposure, mean concentrations in slugs were 1.71, 1.91 and 0.44mg/kg wet weight for chlorophacinone, bromadiolone and brodifacoum respectively. A significant decrease of bromadiolone and brodifacoum in slugs was observed in the post exposure period. In the field study, brodifacoum was detected in >90% of analyzed slugs after application of brodifacoum baits. Then, based on a toxicity-exposure ratio approach, we found that slug consumption may represent a risk of secondary poisoning for three of their predators under acute, repeated or subchronic exposure scenarios. These results suggest that the slugs are not only the potential subject to primary exposure, but also the source of secondary exposure for their predators following application of rodenticide baits.

Brodifacoum poisoning with toxicokinetic data.

The case of a 46-year-old woman who survived after a brodifacoum poisoning is presented. The patient was admitted due to a severe coagulopathy. Initial prothrombin time and activated partial thromboplastin time were both greater than 110 seconds and the patient suffered severe gastric and pulmonary hemorrhage requiring fresh frozen plasma transfusion and parenteral phytonadione administration (up to 100 mg per day). Serum brodifacoum levels were determined by HPLC during seven months. Five days after admission, serum brodifacoum level was 1302 ng/ml. Serum brodifacoum levels decreased till day 209 when became not detectable. Brodifacoum elimination showed a first order kinetic and a 56-day half-life. Investigation of superwarfarin should be considered in any patient with vitamin K dependent coagulation disorder. It would be also useful to obtain periodic brodifacoum levels and build the corresponding elimination curve to help direct phytonadione therapy in poisoned patients.