Guido Mastrantonio

Development of an ionic liquid-based dispersive liquid–liquid microextraction method for the determination of nifurtimox and benznidazole in human plasma

Dispersive ionic liquid-liquid microextraction combined with liquid chromatography and UV detection was used for the determination of two antichagasic drugs in human plasma: nifurtimox and benznidazole. The effects of experimental parameters on extraction efficiency-the type and volume of ionic liquid and disperser solvent, pH, nature and concentration of salt, and the time for centrifugation and extraction-were investigated and optimized. Matrix effects were detected and thus the standard addition method was used for quantification. This microextraction procedure yielded significant improvements over those previously reported in the literature and has several advantages, including high inter-day reproducibility (relative standard deviation=1.02% and 3.66% for nifurtimox and benznidazole, respectively), extremely low detection limits (15.7 ng mL(-1) and 26.5 ng mL(-1) for nifurtimox and benznidazole, respectively), and minimal amounts of sample and extraction solvent required. Recoveries were high (98.0% and 79.8% for nifurtimox and benznidazole, respectively). The proposed methodology offers the advantage of highly satisfactory performance in addition to being inexpensive, simple, and fast in the extraction and preconcentration of these antichagasic drugs from human-plasma samples, with these characteristics being consistent with the practicability requirements in current clinical research or within the context of therapeutic monitoring.

Population Pharmacokinetic Study of Benznidazole in Pediatric Chagas Disease Suggests Efficacy despite Lower Plasma Concentrations than in Adults

Introduction Chagas disease, caused by the parasite Trypanosoma cruzi, can lead to long term cardiac morbidity. Treatment of children with benznidazole is effective, but no pediatric pharmacokinetics data are available and clinical pharmacology information on the drug is scarce. Patients and Methods Prospective population pharmacokinetic (PK) cohort study in children 2–12 years old with Chagas disease treated with oral benznidazole 5–8 mg/kg/day BID for 60 days. (clinicaltrials.gov #NCT00699387). Results Forty children were enrolled in the study. Mean age was 7.3 years. A total of 117 samples were obtained from 38 patients for PK analysis. A one compartment model best fit the data. Weight-corrected clearance rate (CL/F) showed a good correlation with age, with younger patients having a significantly higher CL/F than older children and adults. Simulated median steady-state benznidazole concentrations, based on model parameters, were lower for children in our study than for adults and lowest for children under 7 years of age. Treatment was efficacious in the 37 patients who completed the treatment course, and well tolerated, with few, and mild, adverse drug reactions (ADRs). Discussion Observed benznidazole plasma concentrations in children were markedly lower than those previously reported in adults (treated with comparable mg/kg doses), possibly due to a higher CL/F in smaller children. These lower blood concentrations were nevertheless associated to a high therapeutic response in our cohort. Unlike adults, children have few adverse reactions to the drug, suggesting that there may be a direct correlation between drug concentrations and incidence of ADRs. Our results suggest that studies with lower doses in adults may be warranted. Trial Registration ClinicalTrails.gov NCT00699387

Hydrogen cyanide and carbon monoxide in blood of convicted dead in a polyurethane combustion: a proposition for the data analysis

Carbon monoxide is a well-known toxic component in fire atmospheres. However, the importance of hydrogen cyanide as a toxic agent in fire causalities is under discussion. A tragic polyurethane mattress fire provoked death of 35 convicts in a prison (Unit I, Olmos, Penitenciary Service of Buenos Aires Province, Argentina), in 1990. There is no report of any investigation carried out with such a large amount of victims in Argentina. Carboxihemoglobin (COHb) and hydrogen cyanide (HCN) were quantified in victims blood to elucidate the cause of the death. Saturation of COHb ranged between 4 and 18%, and HCN 2.0-7.2mg/l. These latter values were higher than the lethal levels reported in literature. Other toxic components routinely measured (ethanol, methanol, aldehydes and other volatile compounds) gave negative results on the 35 cases. Neither drugs of abuse nor psychotropics were detected. Statistical chi(2) analysis was applied to find differences between HCN and COHb concentrations. Saturation of COHb and HCN in blood were not independent variables (chi(2)=8.25). Moreover, the ratio COHb/HCN was constant (0.47+/-0.04). In order to evaluate the contribution of each toxic to the diagnosis, a lethal index was defined for each toxic (LI(CO) and LI(HCN)). The most probable cause of death could be inferred by a suitable plot of both indexes. The results indicated that death in the 35 fire victims was probably caused by HCN, generated during the extensive polyurethane decomposition provoked by a rapid increase of temperature.

Pediatric Clinical Pharmacology Studies in Chagas Disease

Chagas disease is a neglected parasitic disease endemic in the Americas. It mainly affects impoverished populations and the acute phase of the infection mostly affects children. Many cases have also been detected in nonendemic countries as a result of recent migratory trends. The chronic phase is relatively asymptomatic, but 30% of patients with chronic infection eventually develop cardiac and digestive complications that commonly lead to death or disability. Only two drugs are available for the treatment of Chagas disease, benznidazole and nifurtimox. These drugs have been shown to be effective in the treatment of both acute and early chronic phases in children, but the pharmacokinetics of these drugs have never been studied in this population. We have set out to conduct a pharmacokinetics study of benznidazole in a pediatric population with Chagas disease. The results of this study are expected to allow better estimation of the optimal doses and schedule of pharmacotherapy for Chagas disease in children.

Spectroscopic and conformational comparative study of trimethyl chalcogenphosphates

Abstract Geometrical, conformational and spectroscopical properties of three chalcogenphosphates were studied comparatively. Rotational isomerism and conformational flexibility were evaluated for trimethyl phosphate, O , O , O -trimethyl thiophosphate and O , O , O -trimethyl selenophosphate by studying the vibrational and NMR spectroscopy. Semi empirical and ab initio calculations have been carried out. Experimental and theoretical results show similar conformational behavior, founding equivalent symmetries for conformations of minimal energies, but a decrease in the conformational flexibility is observed when going from O, to S and to Se species. This result would represent a new factor to explain the higher ability of interaction of O-organophosphorus compounds in some biological active sites, in esterases for example, with respect to the analogous S-organophosphorus compounds and Se-organophosphorus compounds.

Development of UV/HPLC Methods for Quantitative Analysis of Benznidazole in Human Plasma and Urine for Application in Pediatric Clinical Studies

Chagas disease constitutes a major public health problem in Latin America. Correctly designed pharmacokinetic, safety, and bioequivalence studies are desirable in order to fill the knowledge gaps that presently exist on available drugs. It is necessary to develop accurate, simple, reproducible, and sensitive high‐performance liquid chromatography (HPLC)/UV methods for the quantization of benznidazole (BNZ) in human plasma and urine for clinical applications, specially in pediatric patients.

Pharmacokinetic and pharmacodynamic responses in adult patients with Chagas disease treated with a new formulation of benznidazole

Pharmacological treatment of Chagas disease with benznidazole (BNZ) is effective in children in all stages, but it is controversial in chronically infected adults. We report the pharmacokinetics and pharmacodynamics in six adult patients with Chagas disease treated with the new BNZ formulation (ABARAX®) in doses between 2.5-5.5 mg/Kg/day. All but one patient had plasmatic BNZ concentrations within the expected range. All patients finalised treatment with nondetectable Trypanosoma cruziquantitative polymerase chain reaction, which remained nondetectable at the six month follow-up. Our data suggests parasitological responses with the new BNZ and supports the hypothesis that treatment protocols with lower BNZ doses may be effective.

A simple and efficient HPLC method for benznidazole dosage in human breast milk.

Background: Due to migration, Chagas disease is a significant public health problem in Latin America, and in other nonendemic regions. The 2 drugs currently available for the treatment, nifurtimox and benznidazole (BNZ), are associated with a high risk of toxicity in therapeutic doses. Excretion of drug into human breast milk is a potential source of unwanted exposure and pharmacologic effects in the nursing infant. However, this phenomenon was not evaluated until now, and measurement techniques for both drugs in milk were not developed. Methods: In this work, we described the development of a simple and fast method to quantify BNZ in human milk using a pretreatment that involves acid protein precipitation followed by tandem microfiltration, and reverse phase high-performance liquid chromatography/ultraviolet analysis. It is simple because it takes only 3 steps to obtain a clean extracted solution that is ready to inject into the high-performance liquid chromatography equipment. It is fast because a complete analysis of a sample takes only 36 minutes. Results: Although the human breast milk composition is very variable, and lipids are one of the most difficult compounds to clean up on a milk sample, the procedure has proven to be robust and sensitive with a limit of detection of 0.3 &mgr;g/mL and quantization of 0.9 &mgr;g/mL. Despite a 70% recovery value, which could be considered a relatively low result, this recovery is reproducible (coefficient of variation <10%) and the analytical response under the linear range is very good (r2 = 0.9969 adjusted). Real samples of human breast milk from patients in treatment with BNZ were dosed to support the validation process of the method. Conclusions: The method described is fast, specific, accurate, precise, and sufficiently sensitive in the clinical context for the quantification of BNZ in human milk. For all these reasons, it is suitable for clinical risk evaluation studies.

Interpretation of the mechanism of acetylcholinesterase inhibition ability by organophosphorus compounds through a new conformational descriptor. an experimental and theoretical study

Organophosphorus pesticides are the most common classes involved in poisonings related to pesticides. We used inhibitory ability on enzymatic activity of acetylcholinesterase activity and molecular mechanics or ab initio methods of molecular modelling to perform a theoretical approach of the enzyme interaction mechanism of these compounds. Kinetic values for strong and weak inhibitors were measured in a high amplitude range for affinity (Ka) and phosphorylation constants (Kp). To quantitatively describe the conformational behaviour of these molecules, conformational descriptors of free molecules were developed. Quantitative structure activity relationships (QSARs) were constructed with inhibition kinetic values and their molecular descriptors. The conformational descriptors show a high degree of correlation with the kinetic behaviour of these molecules. A positive correlation between the conformational freedom of the studied molecules with Ka is observed. This study allows us to reinterpret the organophosphorus cholinesterase inhibition mechanism and consequently the ‘thiono’ and ‘thiolo effect’ based on a global ‘chalcogen effect’.

Limited infant exposure to benznidazole through breast milk during maternal treatment for Chagas disease

Background Benznidazole (BNZ) is safe and effective for the treatment of paediatric Chagas disease. Treatment of adults is also effective in many cases, but discouraged in breastfeeding women because no information on BNZ transfer into breast milk is available. We aimed to evaluate the degree of BNZ transfer into breast milk in lactating women with Chagas disease. Patients and methods Prospective cohort study of lactating women with Chagas disease treated with BNZ administered for 30 days. Patients and their breastfed infants were evaluated at admission, the 7th and 30th day of treatment (and monthly thereafter, for 6 months). BNZ was measured in plasma and milk by high performance liquid chromatography. The protocol was registered in ClinicalTrials.gov (#NCT01547533). Results 12 lactating women with chronic Chagas disease were enrolled (median age 28.5 years, range 20–34). Median BNZ dose was 5.65 mg/kg/day twice daily. Five mothers had adverse drug events (45%), but no adverse drug reactions or any untoward outcomes were observed in the breastfed infants. Median milk BNZ concentration was 3.8 mg/L (range 0.3–5.9) and 6.26 mg/L (range 0.3–12.6) in plasma. Median BNZ milk to plasma ratio was 0.52 (range 0.3–2.79). Median relative BNZ dose received by the infant (assuming a daily breast milk intake of 150 mL/kg/day) was 12.3% of the maternal dose per kg (range 5.5%–17%). Conclusions The limited transference of BNZ into breast milk and the reassuring normal clinical evaluation of the breastfed babies suggest that maternal BNZ treatment for Chagas disease during breast feeding is unlikely to present a risk for the breastfed infant. Trial registration number ClinicalTrials.gov NCT01547533.