High tacrolimus blood concentrations early after lung transplantation and the risk of kidney injury.

PURPOSE: Lung transplant recipients often develop acute kidney injury (AKI) evolving into chronic kidney disease (CKD). The immunosuppressant tacrolimus might be associated with the emergence of AKI. We analyzed the development and recovery of kidney injury after lung transplantation and related AKI to whole-blood tacrolimus trough concentrations and other factors causing kidney injury. METHODS: We retrospectively studied kidney injury in 186 lung-transplantation patients at the UMC Utrecht between 2001 and 2011. Kidney function and whole-blood tacrolimus trough concentrations were determined from day 1 to 14 and at 1, 3, 6, and 12 months postoperative. Systemic inflammatory response syndrome (SIRS), septic shock, and nephrotoxic medications were evaluated as covariates for AKI. We analyzed liver injury and drug-drug interactions. RESULTS: AKI was present in 85 (46%) patients. Tacrolimus concentrations were supra-therapeutic in 135 of 186 patients (73%). AKI in the first week after transplantation was related to supra-therapeutic tacrolimus concentrations (OR 1.55; 95% CI 1.06-2.27), ≥3 other nephrotoxic drugs (OR 1.96; 95% CI 1.02-3.77), infection (OR 2.48; 95% CI 1.31-4.70), and cystic fibrosis (OR 2.17; 95% CI 1.16-4.06). Recovery rate of AKI was lower than expected (19%), and the cumulative incidence of severe CKD at 1 year was 15%. CONCLUSIONS: After lung transplantation, AKI is common and often evolves into severe CKD, which is a known cause of morbidity and mortality. Supra-therapeutic whole-blood tacrolimus trough concentrations are related to the early onset of AKI. Conscientious targeting tacrolimus blood concentrations might be vital in the early phase after lung transplantation. What is known about this subject? • Lung transplant recipients often develop acute kidney injury evolving into chronic kidney disease increasing both morbidity and mortality. • To date, the pathophysiology of kidney injury after lung transplantation has not been fully elucidated. • The immunosuppressant tacrolimus is difficult to dose, especially in the unstable clinical setting, and is nephrotoxic. WHAT THIS STUDY ADDS: • For the first time, supra-therapeutic whole-blood tacrolimus trough concentrations are related to the emergence of acute kidney injury in the first days after lung transplantation. • Supra-therapeutic whole-blood tacrolimus trough concentrations often occur early after lung transplantation. • AKI after lung transplantation shows low recovery rates.

Neurotoxicity screening of (illicit) drugs using novel methods for analysis of microelectrode array (MEA) recordings.

Annual prevalence of the use of common illicit drugs and new psychoactive substances (NPS) is high, despite the often limited knowledge on the health risks of these substances. Recently, cortical cultures grown on multi-well microelectrode arrays (mwMEAs) have been used for neurotoxicity screening of chemicals, pharmaceuticals, and toxins with a high sensitivity and specificity. However, the use of mwMEAs to investigate the effects of illicit drugs on neuronal activity is largely unexplored. We therefore first characterised the cortical cultures using immunocytochemistry and show the presence of astrocytes, glutamatergic and GABAergic neurons. Neuronal activity is concentration-dependently affected following exposure to six neurotransmitters (glutamate, GABA, serotonin, dopamine, acetylcholine and nicotine). Most neurotransmitters inhibit neuronal activity, although glutamate and acetylcholine transiently increase activity at specific concentrations. These transient effects are not detected when activity is determined during the entire 30min exposure window, potentially resulting in false-negative results. As expected, exposure to the GABAA-receptor antagonist bicuculline increases neuronal activity. Exposure to a positive allosteric modulator of the GABAA-receptor (diazepam) or to glutamate receptor antagonists (CNQX and MK-801) reduces neuronal activity. Further, we demonstrate that exposure to common drugs (3,4-methylenedioxymethamphetamine (MDMA) and amphetamine) and NPS (1-(3-chlorophenyl)piperazine (mCPP), 4-fluoroamphetamine (4-FA) and methoxetamine (MXE)) decreases neuronal activity. MXE most potently inhibits neuronal activity with an IC50 of 0.5µM, whereas 4-FA is least potent with an IC50 of 113µM. Our data demonstrate the importance of analysing neuronal activity within different time windows during exposure to prevent false-negative results. We also show that cortical cultures grown on mwMEAs can successfully be applied to investigate the effects of different (illicit) drugs on neuronal activity. Compared to investigating multiple single endpoints for neurotoxicity or neuromodulation, such as receptor activation or calcium channel function, mwMEAs can provide information on integrated aspects of drug-induced neurotoxicity more rapidly. Therefore, this approach could contribute to a faster insight in possible health risks and shorten the regulation process.

Practical recommendations for calcium channel antagonist poisoning.

Calcium channel antagonists (CCAs) are widely used for different cardiovascular disorders. At therapeutic doses, CCAs have a favourable side effect profile. However, in overdose, CCAs can cause serious complications, such as severe hypotension and bradycardia. Patients in whom a moderate to severe intoxication is anticipated should be observed in a monitored setting for at least 12 hours if an immediate-release formulation is ingested, and at least 24 hours when a sustained-release formulation (or amlodipine) is involved, even if the patient is asymptomatic. Initial treatment is aimed at gastrointestinal decontamination and general supportive care, i.e., fluid resuscitation and correction of metabolic acidosis and electrolyte disturbances. In moderate to severe CCA poisoning, a combined medical strategy might be indispensable, such as administration of vasopressors, intravenous calcium and hyperinsulinaemia/euglycaemia therapy. Especially hyperinsulinaemia/euglycaemia therapy is an important first-line treatment in CCA-overdosed patients in whom a large ingestion is suspected. High-dose insulin, in combination with glucose, seems to be most effective when used early in the intoxication phase, even when the patient shows hardly any haemodynamic instability. Intravenous lipid emulsion therapy should only be considered in patients with life-threatening cardiovascular toxicity, such as refractory shock, which is unresponsive to conventional therapies. When supportive and specific pharmacological measures fail to adequately reverse refractory conditions in CCA overdose, the use of extracorporeal life support should be considered. The efficacy of these pharmacological and non-pharmacological interventions generally advocated in CCA poisoning needs further in-depth mechanistic foundation, in order to improve individualised treatment of CCA-overdosed patients.

Pharmacokinetics and Toxicity of Tacrolimus Early After Heart and Lung Transplantation.

Annually, about 8000 heart and lung transplantations are successfully performed worldwide. However, morbidity and mortality still pose a major concern. Renal failure in heart and lung transplant recipients is an essential adverse cause of morbidity and mortality, often originating in the early postoperative phase. At this time of clinical instability, the kidneys are exposed to numerous nephrotoxic stimuli. Among these, tacrolimus toxicity plays an important role, and its pharmacokinetics may be significantly altered in this critical phase by fluctuating drug absorption, changed protein metabolism, anemia and (multi-) organ failure. Limited understanding of tacrolimus pharmacokinetics in these circumstances is hampering daily practice. Tacrolimus dose adjustments are generally based on whole blood trough levels, which widely vary early after transplantation. Moreover, whole blood trough levels are difficult to predict and are poorly related to the area under the concentration-time curve. Even within the therapeutic range, toxicity may occur. These shortcomings of tacrolimus monitoring may not hold for the unbound tacrolimus plasma concentrations, which may better reflect tacrolimus toxicity. This review focuses on posttransplant tacrolimus pharmacokinetics, discusses relevant factors influencing the unbound tacrolimus concentrations and tacrolimus (nephro-) toxicity in heart and lung transplantation patients.

Evaluation of three physiologically based pharmacokinetic (PBPK) modeling tools for emergency risk assessment after acute dichloromethane exposure.

INTRODUCTION: Physiologically based pharmacokinetic (PBPK) models may be useful in emergency risk assessment, after acute exposure to chemicals, such as dichloromethane (DCM). We evaluated the applicability of three PBPK models for human risk assessment following a single exposure to DCM: one model is specifically developed for DCM (Bos) and the two others are semi-generic ones (Mumtaz and Jongeneelen). MATERIALS AND METHODS: We assessed the accuracy of the models' predictions by simulating exposure data from a previous healthy volunteer study, in which six subjects had been exposed to DCM for 1h. The time-course of both the blood DCM concentration and percentage of carboxyhemoglobin (HbCO) were simulated. RESULTS: With all models, the shape of the simulated time course resembled the shape of the experimental data. For the end of the exposure, the predicted DCM blood concentration ranged between 1.52-4.19mg/L with the Bos model, 1.42-4.04mg/L with the Mumtaz model, and 1.81-4.31mg/L with the Jongeneelen model compared to 0.27-5.44mg/L in the experimental data. % HbCO could be predicted only with the Bos model. The maximum predicted % HbCO ranged between 3.1 and 4.2% compared to 0.4-2.3% in the experimental data. The % HbCO predictions were more in line with the experimental data after adjustment of the Bos model for the endogenous HbCO levels. CONCLUSIONS: The Bos Mumtaz and Jongeneelen PBPK models were able to simulate experimental DCM blood concentrations reasonably well. The Bos model appears to be useful for calculating HbCO concentrations in emergency risk assessment.

Increase of baclofen intoxications: risks involved and management.

Baclofen has been increasingly used in the treatment of alcohol withdrawal syndrome (AWS). We present a patient with AWS and psychiatric comorbidity who ingested 700 mg of baclofen. ICU admission was necessary for ventilatory support and symptomatic treatment. The patient was dismissed without sequelae.

Recommendations for the paracetamol treatment nomogram and side effects of N-acetylcysteine.

Treatment of paracetamol intoxication consists of administration of N-acetylcysteine, preferably shortly after paracetamol ingestion. In most countries, the decision to treat patients with N-acetylcysteine depends on the paracetamol plasma concentration. In the literature, different arguments are given regarding when to treat paracetamol overdose. Some authors do not recommend treatment with N-acetylcysteine at low paracetamol plasma concentrations since unnecessary adverse effects may be induced. But no treatment with N-acetylcysteine at higher paracetamol plasma concentrations may lead to unnecessary severe morbidity and mortality. In this review, we provide an overview on the severity and prevalence of adverse side effects after N-acetylcysteine administration and the consequences these side effects may have for the treatment of paracetamol intoxication. The final conclusion is to continue using the guidelines of the Dutch National Poisons Information Centre for N-acetylcysteine administration in paracetamol intoxication.

Management of prolonged QT interval and torsades de pointes in the intoxicated patient.

Many drugs can significantly influence cardiac repolarisation causing an increased duration of this repolarisation phase, challenging the repolarisation reserve. This may set the stage for life-threatening ventricular arrhythmias such as torsades de pointes (TdP). TdP generally occurs in conjunction with a prolonged QT interval (QT) on the electrocardiogram. The Dutch Poisons Information Centre (NVIC) often receives information requests about drugs that can influence the QT already at therapeutic dosages. Drug-induced QT prolongation is dose dependent and hence can be particularly pronounced in overdose situations. Also, additional risk factors for the development of life-threatening arrhythmias are often present in intoxicated patients. This review focuses on identification and management of drug-intoxicated patients who are at risk for a reduction in their repolarisation reserve, measured by their QT interval. The QT interval is strongly dependent on heart rate, which has led to the introduction of different methods to adjust the QT interval, i.e. the QTc. Bazett's formula, which has been used for decades, lacks accuracy concerning QTc calculation at higher and lower heart rates, situations often relevant when dealing with intoxicated patients. Additionally, we highlight drugs with QT-prolonging potential that are commonly associated with an overdose setting in the Netherlands. Finally, standard treatment options specifically pointed toward the intoxicated patient at risk of QT prolongation and TdP will be discussed.

Ethylene glycol or methanol intoxication: which antidote should be used, fomepizole or ethanol?

Ethylene glycol (EG) and methanol poisoning can cause life-threatening complications. Toxicity of EG and methanol is related to the production of toxic metabolites by the enzyme alcohol dehydrogenase (ADH), which can lead to metabolic acidosis, renal failure (in EG poisoning), blindness (in methanol poisoning) and death. Therapy consists of general supportive care (e.g. intravenous fluids, correction of electrolytes and acidaemia), the use of antidotes and haemodialysis. Haemodialysis is considered a key element in the treatment of severe EG and methanol intoxication and is aimed at removing both the parent compound and its toxic metabolites, reducing the duration of antidotal treatment and shortening the hospital observation period. Currently, there are two antidotes used to block ADH-mediated metabolism of EG and methanol: ethanol and fomepizole. In this review, the advantages and disadvantages of both antidotes in terms of efficacy, safety and costs are discussed in order to help the physician to decide which antidote is appropriate in a specific clinical setting.

Review of acute chemical incidents as a first step in evaluating the usefulness of physiologically based pharmacokinetic models in such incidents.

CONTEXT. Acute chemical incidents can have substantial public health consequences in terms of morbidity and mortality. OBJECTIVE. We aimed to characterize acute chemical incidents and near-misses in the Netherlands and compare the results with previous studies. This review is a first step in evaluating whether Physiologically Based Pharmacokinetic (PBPK) models can be of value in acute chemical incidents. MATERIAL AND METHODS. Government, regional, municipal and University Hospital Institutes involved in the management of acute chemical incidents in the Netherlands were contacted, and they provided data between 2008 and 2010 on the characteristics and consequences of the incidents. The study is a retrospective epidemiological study based on data from five institutes. Incidents involving biological agents or radiation were excluded. RESULTS. A total of 764 reports were available which involved 722 incidents after cross-matching the different sources of data. Forty incidents were excluded, leaving 682 incidents for which information was available in accordance with the inclusion criteria. Of the 682 incidents included in this study, most occurred in non-industrial buildings (37%) or industrial sites (34%). The most frequently observed event types were loss of containment (60%) and fire (36%), leading to gas emission (54%), followed by spill of liquid or solid chemicals (36%). The chemicals involved were most often products of combustion (e.g. smoke, soot, particles, 25%) and volatile organic compounds (e.g. solvents, styrene, xylene, 23%), followed by inorganic gases (e.g. carbon monoxide, hydrogen, hydrogen sulphide, 13%). A minimum of 847 people experienced adverse health effects following exposure during a chemical incident, and 10 fatalities were reported. The most frequently reported symptoms were respiratory (27%), due to irritant chemicals. The number of incidents related to fire and the number of injured people were higher in this study than in previous studies; 49% of the injured were transported to hospital. DISCUSSION. This study helps to identify which chemicals are frequently involved in acute chemical incidents in the Netherlands. The results will be used in future to assess whether PBPK models may be useful for risk assessment of chemicals often involved in acute chemical incidents and for which human toxicological and kinetic data are scarce.

Extracorporeal membrane oxygenation in the treatment of poisoned patients.

CONTEXT: Although extracorporeal membrane oxygenation (ECMO) was used in many patients following its introduction in 1972, most hospitals had abandoned this experimental treatment for adult patients. Recently, improvements in the ECMO circuitry rendered it more biocompatible. The surprisingly low mortality in patients with severe acute respiratory distress syndrome who were treated with ECMO in the influenza A/H1N1 pandemic of 2009 resurrected interest in ECMO in many intensive care units around the world. OBJECTIVES: This article reviews the different techniques of ECMO, the indications, contraindications and complications of its use, its role in poisoned patients and the ethics of its use. METHODS: We searched Pubmed, Toxnet, Cochrane database and Embase from 1966 to September 2012 using the search terms (''extracorporeal membrane oxygenation'', 'extracorporeal life support', 'ECMO', 'ECLS', 'assist-device', and 'intox*' or 'poison*'). These searches identified 242 papers of which 116 described ECMO in conditions other than intoxications or were reviews. In total 46 publications selected for this manuscript were case reports or case series involving poisoned patients. ECMO TECHNIQUES: Two types of ECMO are used: veno-venous ECMO (VV-ECMO) or veno-arterial ECMO (VA-ECMO). VV-ECMO is used for patients with severe ARDS to secure adequate oxygenation of the organs while protecting the lungs from harmful ventilation pressures or prolonged inspiratory fraction of oxygen. VA-ECMO can be used whenever the patient remains in shock despite adequate fluid resuscitation and is refractory to administration of inotropes and vasopressors. INDICATIONS: The organ support that can be applied with ECMO makes it especially useful in patients with severe poisoning as the clinical impact of the intoxication is often temporary; ECMO can be used as a 'bridge to recovery'. CONTRAINDICATIONS: Absolute contraindications are uncontrolled coagulopathy and severe intracranial bleeding, which precludes the use of anticoagulation therapy. Relative contraindications to ECMO include advanced age, severe irreversible brain injury, untreatable metastatic cancer, severe organ dysfunction (some suggest a Sequential Organ Failure Assessment (SOFA) score > 15), and high pressure positive pressure ventilation for more than 7 days. COMPLICATIONS: The most common complication of ECMO is either bleeding at the cannulation site (in VV-ECMO) or bleeding at the surgical entry site (in VA-ECMO). Overall bleeding complications currently occur in 10-36% patients, and intracranial haemorrhage is seen in up to 6% of patients. ECMO should be reserved, therefore, for the most severely ill poisoned patients with a high risk of death. ECMO in poisoned patients. There are no randomised trials of ECMO in poisoned patients with refractory shock or who have ARDS caused by an intoxication. VV-ECMO can be considered in patients with type l and ll respiratory failure. In patients with life-threatening haemodynamic instability, VA-ECMO can be considered when shock persists despite volume administration, inotropes and vasoconstrictors, and (sometimes) intra-aortic balloon counterpulsation. Typical examples include poisoning due to calcium channel antagonists, beta-blockers, tricyclic antidepressants, chloroquine and colchicine. ETHICS OF ECMO USE: It is only ethical to use such a costly intervention (£19,252 and US$ 31,000 per quality-adjusted life year) if the treatment has a real purpose such as a 'bridge to recovery', a 'bridge to transplant', or a 'bridge to permanent assist device' (in the case of persistent cardiac failure). CONCLUSIONS: In the last decade, ECMO equipment has improved considerably, rendering it more biocompatible, and it has been used more frequently as an assist device for patients needing oxygenation as well as circulatory support. ECMO is considered a good salvage therapy for patients who are severely poisoned with ARDS or refractory circulatory shock.

Harmonisation of product notification to Poisons Centres in EU Member States.

INTRODUCTION: In the European Union (EU), notification of product information by industry to poisons centres and/or competent authorities is a legal obligation for mixtures classified as hazardous. However, EU legislation does not specify the precise information needed for this product notification. As a consequence, varying requirements have been developed in different EU Member States. The European Commission (EC) carried out an assessment of whether harmonisation of product notification can be achieved. This manuscript provides an overview of the most important (discussion) points to reach harmonisation. COMPOSITION AND CONCENTRATION OF INGREDIENTS: Discussions have focused mainly on whether non-classified ingredients should be notified only above a concentration threshold and on the use of defined, narrow concentration ranges instead of exact concentrations for hazardous ingredients. ELECTRONIC DATA EXCHANGE FORMAT: All stakeholders agree to the development of an electronic data exchange format for product notification and identify the eXtensible Markup Language (XML) as the most appropriate format. EUROPEAN PRODUCT DATABASE: Instead of multiple notifications to national databases, the EC will analyse the benefits, feasibility and costs of a European product database to provide a centralised portal for companies to upload their product information. Poisons centres and competent authorities need to have access to this information. UNIQUE PRODUCT IDENTIFIER: A Unique Product Identifier (UPI) on the product label can unambiguously identify the product and its formula and links it to the corresponding notified product information. A procedure for the creation of a UPI by companies has already been proposed. PRODUCT CATEGORY SYSTEM: There is broad support for the development of a hierarchical product category system to facilitate statistical analyses and comparability of poisoning incidents in EU Member States. OUTLOOK: Following a 3-year assessment period, the EC concluded that harmonisation of product notification is an achievable goal. In order to draft an Annex to the CLP Regulation concerning this topic, a new working group with representatives of EU Member States, European Association of Poisons Centres and Clinical Toxicologists (EAPCCT) and other stakeholders will attempt to find consensus on harmonisation of product notification.

Modulation of human GABAA receptor function: a novel mode of action of drugs of abuse.

Drugs of abuse are known to mainly affect the dopaminergic and serotonergic system, although behavioral studies indicated that the GABA-ergic system also plays a role. We therefore investigated the acute effects of several commonly used drugs of abuse (methamphetamine, amphetamine, 3,4-methylenedioxymethamphetamine (MDMA), 3,4-methylenedioxyamphetamine (MDA) and meta-chlorophenylpiperazine (mCPP)) on the function of the human α(1)ß(2)γ(2) GABA(A) receptor (hGABA(A)-R), expressed in Xenopus oocytes, using the two-electrode voltage-clamp technique. Although none of the tested drugs acted as full agonist on the hGABA(A)-R, some drugs induced differential modulation of hGABA(A)-R function, depending on the degree of receptor occupancy. Methamphetamine did not affect the GABA-evoked current at high receptor occupancy, but induced a minor inhibition at low receptor occupancy. Its metabolite amphetamine slightly potentiated the GABA-evoked current. MDMA and its metabolite MDA both inhibited the current at low receptor occupancy. However, MDMA did not affect the current at high occupancy, whereas MDA induced a potentiation. mCPP induced a strong inhibition (max. ∼ 80%) at low receptor occupancy, but ∼ 25% potentiation at high receptor occupancy. Competitive binding to one of the GABA-binding sites could explain the drug-induced inhibitions observed at low receptor occupancy, whereas an additional interaction with a positive allosteric binding site may play a role in the observed potentiations at high receptor occupancy. This is the first study to identify direct modulation of hGABA(A)-Rs as a novel mode of action for several drugs of abuse. Consequently, hGABA(A)-Rs should be considered as target for psychiatric pharmaceuticals and in developing treatment for drug intoxications.

Clinical pharmacology of exogenously administered alkaline phosphatase.

PURPOSE: To evaluate the clinical pharmacology of exogenous alkaline phosphatase (AP). METHODS: Randomized, double-blind, placebo-controlled sequential protocols of (1) ascending doses and infusion duration (volunteers) and (2) fixed dose and duration (patients) were conducted at clinical pharmacology and intensive care units. A total of 103 subjects (67 male volunteers and 36 patients with severe sepsis) were administered exogenous, 10-min IV infusions (three ascending doses) or 24-72 h continuous (132.5-200 U kg(-1) 24 h(-1)) IV infusion with/without preceding loading dose and experimental endotoxemia for evaluations of pharmacokinetics, pharmacodynamics, safety parameters, antigenicity, inflammatory markers, and outcomes. RESULTS: Linearity and dose-proportionality were shown during 10-min infusions. The relatively short elimination half-life necessitated a loading dose to achieve stable enzyme levels. Pharmacokinetic parameters in volunteers and patients were similar. Innate immunity response was not significantly influenced by AP, while renal function significantly improved in sepsis patients. CONCLUSIONS: The pharmacokinetics of exogenous AP is linear, dose-proportional, exhibit a short half-life, and are not influenced by renal impairment or dialysis.

[Intoxication with magnesium, a 'forgotten' electrolyte]. / Intoxicatie met magnesium, een 'vergeten' elektrolyt.

Magnesium salts are widely used and are usually regarded as rather harmless. Magnesium intoxication, however, can be lethal. Recently, severe magnesium intoxication was identified in 3 patients, one woman aged 68 and 2 men aged 19 and 26 years respectively, who required treatment in an Intensive Care setting. Initial symptoms of magnesium intoxication went unnoticed due to sedation or paraplegia. Only after developing severe neurological symptoms, respiratory or circulatory failure, magnesium intoxication was diagnosed. Plasma magnesium levels exceeded 6 mmol/l (reference value: 0.70-1.00). Therapy consisted of cessation of magnesium therapy, administration of calcium and enhanced elimination of magnesium by haemodialysis. All patients survived the intoxication. Magnesium intoxication may develop unnoticed when the initial signs and symptoms are masked by clinical conditions. Especially patients with renal failure, inflammatory bowel disease, sedation and paraplegia need careful monitoring when magnesium is administered.

Effects of sub-chronic nitrate exposure on the thyroidal function in humans.

No experimental data exist on the thyroid toxicity of nitrate among humans. We aimed to show that no significant antithyroid effect could be observed after exposure to a three times the acceptable daily intake of nitrate in humans. In a randomized controlled non-inferiority trial, 10 volunteers received 15 mg/kg sodium nitrate during 28 days whereas 10 control participants received distilled water. We performed 5- and 24-h measurements of thyroidal (131)I uptake (RAIU) before and at the end of the exposure period. Thyroid hormone plasma concentrations of T3, rT3, T4, TSH were also measured prior to and after exposure. Differences in RAIU between the intervention and the control groups at 4 weeks were 3.4% (95% confidence interval -0.5 to 7.3, and 4.8% (95% confidence interval -1.4 to 11.0, respectively, for the 5- and 24-h RAIU measurement. Plasma concentrations of thyroid hormones stayed normal. In conclusion, no significant effects on thyroidal (131)I uptake and thyroid hormones plasma concentrations were observed after sub-chronic exposition to 15 mg/kg sodium nitrate among humans.