Holistic food safety evaluation of herbs: methods for the determination of organic and inorganic trace contaminants in Moringa stenopetala as a case study.

Moringa stenopetala is considered a superfood due to the many bioactive compounds that it provides to the diet. However, like all edible plants, it is mandatory to guarantee food safety. Thus it is necessary to develop analytical methods that can rapidly and accurately determine hazardous pollutants, to evaluate compliance with food regulations. In this regard, two multi-component procedures were developed trying to cover some of the main organic and inorganic potential contaminants. A microwave-assisted digestion followed by electrothermal atomic absorption spectrometry was used for arsenic, cadmium, and lead determination, while a modification of the QuEChERS protocol followed by gas chromatography-tandem mass spectrometry was employed for the determination of 55 pesticides from different families. Both analytical methods were thoroughly validated according to international guidelines. The analyzed samples obtained from the Uruguayan market showed compliance with both, national and international, food regulations. The holistic approach employed in this research is not commonly presented in the literature, thus constituting a novel way to face food safety.

Metallomics: An Essential Tool for the Study of Potential Antiparasitic Metallodrugs.

Metallomics is an emerging area of omics approaches that has grown enormously in the past few years. It integrates research related to metals in biological systems, in symbiosis with genomics and proteomics. These omics approaches can provide in-depth insights into the mechanisms of action of potential metallodrugs, including their physiological metabolism and their molecular targets. Herein, we review the most significant advances concerning cellular uptake and subcellular distribution assays of different potential metallodrugs with activity against Trypanosma cruzi, the protozoan parasite that causes Chagas disease, a pressing health problem in high-poverty areas of Latin America. Furthermore, the first multiomics approaches including metallomics, proteomics, and transcriptomics for the comprehensive study of potential metallodrugs with anti-Trypanosoma cruzi activity are described.

Biomonitoring of arsenic, lead, manganese and mercury in hair from a presumably exposed Uruguayan child population.

Aim: To perform an exposure assessment of arsenic, manganese, mercury and lead levels in hair samples from children from poor neighborhoods. Materials & methods: A total of 38 Caucasian children were recruited with the consent of their parents or tutors. Determinations were performed by atomic absorption spectrometry. Results & conclusion: Results were 0.045-0.12 µg/g-1 (arsenic), 0.56-2.05 µg/g-1 (manganese) and 0.34-27.8 µg/g-1 (lead). Lead results did not correlate with those previously reported in blood from the same individuals, suggesting that hair is not useful for exposure assessment of this contaminant. Mercury was determined for the first time in Uruguayan children showing levels <0.083 µg/g-1. Results revealed low-to-moderate metal exposure, except for some high lead findings.

Biomonitoring of Potentially Toxic Elements in Dyed Hairs and Its Correlation with Variables of Interest.

Hair is good bioindicator of exposure, due to its ability to store and retain trace elements for long periods of time. But it can be especially useful when hair dyes are used since they may contain potentially toxic salts in their composition. In this context, analytical methods for the determination of bismuth, cadmium, lead, and silver in scalp human hair by electrothermal atomic absorption spectrometry were successfully validated. A total of 60 samples obtained from women between 18 and 60 years were analyzed: 34 dyed hairs and 26 untreated hairs (control). Average results expressed in dry weight (dyed/control) for each element were 2.34/0.49 µg g-1 (silver), 0.142/0.139 µg g-1 (bismuth), 0.055/0.054 µg g-1 (cadmium), and 2.09/0.99 µg g-1 (lead), respectively. These results agreed with those previously reported for non-exposed populations. A statistically significant higher Ag concentration in dyed hairs was observed, suggesting the bioaccumulation of this element. The associations between metal concentration and variables of interest (age, education, smoking habit, dye brand, use of dietary supplements) were investigated. A strong Pearson correlation was found for the pair Ag/Pb (r = 0.494, p < 0.05). Also, strong associations between lead levels and all the selected variables were observed (p < 0.05), while strong associations between silver levels with age and dye brand and association between cadmium levels and smoking habit were found. Furthermore, several commercial hair dye brands were analyzed to verify compliance with cosmetic regulations. This constitutes the first study of such characteristics performed in Uruguay, with worldwide relevance.

Distribution of Inorganic Contaminants Along the Coast of Ciudad de la Costa, Uruguay.

The aim of this work was to study the distribution of relevant inorganic contaminants in environmental samples from Ciudad de la Costa, the second most populated city of Uruguay. For this task aluminum (Al), cadmium (Cd), chromium (Cr), nickel (Ni) and lead (Pb) levels were determined in sand, water, and plant material from six beaches that are located in the area. Analytical methods were successfully validated. Accuracy was ensured by using certified reference materials and spiking procedures. Average metal concentrations in sand and water samples were compared with the maximum limits established by international or national regulations (when available), being far below these limits in all cases. This may indicate that the inorganic contamination in the studied area is rather low, despite the rapid increase in anthropic activities in recent years. To give a deeper characterization of the ecosystem, metal accumulation and mobility within a common plant of the zone, Ammophila arenaria, were studied. Results showed that concentrations in roots were one order of magnitude higher in comparison to aerial parts. In addition, the bioaccumulation coefficient (BAC) showed that the plant accumulates Cd, Cr and Ni, while a low accumulation for Pb was observed. Finally, a Cr(VI) speciation analysis in Ammophila arenaria was performed, showing Cr(VI) percentages below 4.6%, which is relevant from the (eco)toxicological point of view due to the high environmental mobility and bioavailability of this valence state. This work constitutes the first environmental surveillance report of this kind, regarding this populated area of the country.

Platinum and Palladium Organometallic Compounds: Disrupting the Ergosterol Pathway in Trypanosoma cruzi.

Current treatment for Chagas' disease is based on two drugs, Nifurtimox and Benznidazol, which have limitations that reduce the effectiveness and continuity of treatment. Thus, there is an urgent need to develop new, safe and effective drugs. In previous work, two new metal-based compounds with trypanocidal activity, Pd-dppf-mpo and Pt-dppf-mpo, were fully characterized. To unravel the mechanism of action of these two analogous metal-based drugs, high-throughput omics studies were performed. A multimodal mechanism of action was postulated with several candidates as molecular targets. In this work, we validated the ergosterol biosynthesis pathway as a target for these compounds through the determination of sterol levels by HPLC in treated parasites. To understand the molecular level at which these compounds participate, two enzymes that met eligibility criteria at different levels were selected for further studies: phosphomevalonate kinase (PMK) and lanosterol 14-α demethylase (CYP51). Molecular docking processes were carried out to search for potential sites of interaction for both enzymes. To validate these candidates, a gain-of-function strategy was used through the generation of overexpressing PMK and CYP51 parasites. Results here presented confirm that the mechanism of action of Pd-dppf-mpo and Pt-dppf-mpo compounds involves the inhibition of both enzymes.

Multifunctional organometallic compounds for the treatment of Chagas disease: Re(I) tricarbonyl compounds with two different bioactive ligands.

Chagas' disease (American Trypanosomiasis) is an ancient and endemic illness in Latin America caused by the protozoan parasite Trypanosoma cruzi. Although there is an urgent need for more efficient and less toxic chemotherapeutics, no new drugs to treat this disease have entered the clinic in the last decades. Searching for metal-based prospective antichagasic drugs, in this work, multifunctional Re(I) tricarbonyl compounds bearing two different bioactive ligands were designed: a polypyridyl NN derivative of 1,10-phenanthroline and a monodentate azole (Clotrimazole CTZ or Ketoconazol KTZ). Five fac-[Re(CO)3(NN)(CTZ)](PF6) compounds and a fac-[Re(CO)3(NN)(KTZ)](PF6) were synthesized and fully characterized. They showed activity against epimastigotes (IC50 3.48-9.42 µM) and trypomastigotes of T. cruzi (IC50 0.61-2.79 µM) and moderate to good selectivity towards the parasite compared to the VERO mammalian cell model. In order to unravel the mechanism of action of our compounds, two potential targets were experimentally and theoretically studied, namely DNA and one of the enzymes involved in the parasite ergosterol biosynthetic pathway, CYP51 (lanosterol 14-α-demethylase). As hypothesized, the multifunctional compounds shared in vitro a similar mode of action as that disclosed for the single bioactive moieties included in the new chemical entities. Additionally, two relevant physicochemical properties of biological interest in prospective drug development, namely lipophilicity and stability in solution in different media, were determined. The whole set of results demonstrates the potentiality of these Re(I) tricarbonyls as promising candidates for further antitrypanosomal drug development.

New multifunctional Ru(II) organometallic compounds show activity against Trypanosoma brucei and Leishmania infantum.

Human African trypanosomiasis (sleeping sickness) and leishmaniasis are prevalent zoonotic diseases caused by genomically related trypanosomatid protozoan parasites (Trypanosoma brucei and Leishmania spp). Additionally, both are co-endemic in certain regions of the world. Only a small number of old drugs exist for their treatment, with most of them sharing poor safety, efficacy, and pharmacokinetic profiles. In this work, new multifunctional Ru(II) ferrocenyl compounds were rationally designed as potential agents against these trypanosomatid parasites by including in a single molecule 1,1'-bis(diphenylphosphino)ferrocene (dppf) and two bioactive bidentate ligands: 8-hydroxyquinoline derivatives (8HQs) and polypyridyl ligands (NN). Three [Ru(8HQs)(dppf)(NN)](PF6) compounds were synthesized and fully characterized. They showed in vitro activity on bloodstream Trypanosoma brucei (IC50 140-310 nM) and on Leishmania infantum promastigotes (IC50 3.0-4.8 µM). The compounds showed good selectivity towards T. brucei in respect to J774 murine macrophages as mammalian cell model (SI 15-38). Changing hexafluorophosphate counterion by chloride led to a three-fold increase in activity on both parasites and to a two to three-fold increase in selectivity towards the pathogens. The compounds affect in vitro at least the targets of the individual bioactive moieties included in the new chemical entities: DNA and generation of ROS. The compounds are stable in solution and are more lipophilic than the free bioactive ligands. No clear correlation between lipophilicity, interaction with DNA or generation of ROS and activity was detected, which agrees with their overall similar anti-trypanosoma potency and selectivity. These compounds are promising candidates for further drug development.

Application of microwave plasma atomic emission spectrometry in bioanalytical chemistry of bioactive rhenium compounds.

Five newly synthetized fac-Re(I) tricarbonyl compounds were explored as prospective antitrypanosomal agents. The biological activity of the whole series was evaluated preliminarily against the epimastigote form of Trypanosoma cruzi. All compounds showed activity against epimastigotes with IC50 values in the low micromolar range. The most active compound [fac-Re(I)(CO)3(tmp)(CTZ)](PF6), with CTZ = clotrimazole and tmp = 3,4,7,8-tetramethyl-1,10-phenantroline, showed good selectivity towards the parasites and thus was selected to carry out further metallomic studies. For this task, a newly bioanalytical method based on microwave plasma atomic emission spectrometry (MP-AES) was developed and validated. The accuracy of the method was ensured by testing a certified reference material. Results of rhenium elemental analysis by MP-AES agreed with the proposed formula of the studied compounds, contributing to the overall validation of the method, which was then applied to evaluate the percentage of rhenium uptaken by the parasites and the association of the compounds with parasite biomacromolecules. Metallomics results showed low total rhenium percentage uptaken by parasites (∼1.2%) and preferential accumulation in the soluble proteins fraction (∼82.8%). Thus, the method based on MP-AES turned out to be an economical and green alternative for metallomics studies involving potential rhenium metallodrugs. Moreover, a comparison against rhenium determination by electrothermal atomic absorption spectrometry (ET-AAS) was included.

Fe(III)-Complex-Imprinted Polymers for the Green Oxidative Degradation of the Methyl Orange Dye Pollutant.

One of the biggest problems worldwide is the pollution of natural water bodies by dyes coming from effluents used in the textile industry. In the quest for novel effluent treatment alternatives, the aim of this work was to immobilize Fe(III) complexes in molecularly imprinted polymers (MIPs) to produce efficient Fenton-like heterogeneous catalysts for the green oxidative degradation of the methyl orange (MO) dye pollutant. Different metal complexes bearing commercial and low-cost ligands were assayed and their catalytic activity levels towards the discoloration of MO by H2O2 were assessed. The best candidates were Fe(III)-BMPA (BMPA = di-(2-picolyl)amine) and Fe(III)-NTP (NTP = 3,3',3″-nitrilotripropionic acid), displaying above 70% MO degradation in 3 h. Fe(III)-BMPA caused the oxidative degradation through two first-order stages, related to the formation of BMPA-Fe-OOH and the generation of reactive oxygen species. Only the first of these stages was detected for Fe(III)-NTP. Both complexes were then employed to imprint catalytic cavities into MIPs. The polymers showed catalytic profiles that were highly dependent on the crosslinking agent employed, with N,N-methylenebisacrylamide (MBAA) being the crosslinker that rendered polymers with optimal oxidative performance (>95% conversion). The obtained ion-imprinted polymers constitute cheap and robust solid matrices, with the potential to be coupled to dye-containing effluent treatment systems with synchronous H2O2 injection.

Heteroleptic Oxidovanadium(V) Complexes with Activity against Infective and Non-Infective Stages of Trypanosoma cruzi.

Five heteroleptic compounds, [VVO(IN-2H)(L-H)], where L are 8-hydroxyquinoline derivatives and IN is a Schiff base ligand, were synthesized and characterized in both the solid and solution state. The compounds were evaluated on epimastigotes and trypomastigotes of Trypanosoma cruzi as well as on VERO cells, as a mammalian cell model. Compounds showed activity against trypomastigotes with IC50 values of 0.29-3.02 µM. IN ligand and the new [VVO2(IN-H)] complex showed negligible activity. The most active compound [VVO(IN-2H)(L2-H)], with L2 = 5-chloro-7-iodo-8-hydroxyquinoline, showed good selectivity towards the parasite and was selected to carry out further biological studies. Stability studies suggested a partial decomposition in solution. [VVO(IN-2H)(L2-H)] affects the infection potential of cell-derived trypomastigotes. Low total vanadium uptake by parasites and preferential accumulation in the soluble proteins fraction were determined. A trypanocide effect was observed when incubating epimastigotes with 10 × IC50 values of [VVO(IN-2H)(L2-H)] and the generation of ROS after treatments was suggested. Fluorescence competition measurements with DNA:ethidium bromide adduct showed a moderate DNA interaction of the complexes. In vivo toxicity study on C. elegans model showed no toxicity up to a 100 µM concentration of [VVO(IN-2H)(L2-H)]. This compound could be considered a prospective anti-T. cruzi agent that deserves further research.

Development of an Ozone-Assisted Sample Preparation Method for the Determination of Cu and Zn in Rice Samples.

A green analytical method for the determination of Cu and Zn in rice samples was developed. This method was based on an ozone-assisted extraction (OAE) in diluted acid media. A novel closed system was designed for this purpose that allowed four simultaneous sample treatments being safe for the laboratory environment. The method consisted in 0.5 g of the sample, 15 minutes of ozonation, and 3 minutes of centrifugation. The obtained supernatant was ready for Cu and Zn determinations by flame atomic absorption spectrometry. Detection limits were 0.20 and 0.08 mg kg-1 for Cu and Zn, respectively, with a precision (RSD) better than 5% for both elements. A certified reference material of rice flour was analyzed for trueness evaluation, and the mean recoveries (%) were 100.4 (Cu) and 95.9 (Zn). Several commercial rice samples were analyzed using this method, and the results were compared with those obtained using traditional microwave-assisted digestion (MAE). Both methods yielded comparable results. Cu and Zn levels were in accordance with reported values in other regions. The OAE resulted to be simple and economical and with results equivalent with those obtained using traditional sample preparation procedures as MAE with the advantage of being in good agreement with the principles of green analytical chemistry.

New Pd-Fe ferrocenyl antiparasitic compounds with bioactive 8-hydroxyquinoline ligands: a comparative study with their Pt-Fe analogues.

In the search for a more effective chemotherapy for the treatment of Human African Trypanosomiasis, a disease caused by the parasite Trypanosoma brucei, the development of ferrocenyl compounds has arisen as a promising strategy. In this work, five new Pd-Fe heterobimetallic [PdII(L)(dppf)](PF6) compounds, including 8-hydroxyquinolyl derivatives HL1-HL5 as bioactive ligands and dppf = 1,1'-bis(diphenylphosphino)ferrocene as the organometallic co-ligand, were synthesized and fully characterized in the solid state and in solution. Molecular structures of three compounds were solved by single crystal X-ray diffraction methods. The compounds displayed submicromolar or micromolar IC50 values against bloodstream T. brucei (IC50: 0.33-1.2 µM), and good selectivity towards the pathogen (SI: 4-102) with respect to mammalian macrophages (cell line J774). The new Pd complexes proved to be 2-fold to 45-fold more potent than the drug nifurtimox but most of them are less active than their Pt analogues. Potential molecular targets were studied. The complexes interact with DNA but they do not alter the intracellular thiol-redox homeostasis of the parasite. In order to understand and predict the main structural determinants on the anti-T. brucei activity, a search of quantitative structure-activity relationships (QSAR) was performed including all the [M(L)(dppf)](PF6) complexes, where M = Pd(ii) or Pt(ii), currently and previously developed by us. The correlation obtained shows the relevance of the electronic effects, the lipophilicity and the type of metal. According to the QSAR study, compounds with electron-withdrawing ligands, higher lipophilicity and harboring Pt would result in higher T. brucei cytotoxicity. From the whole series of [M(L)(dppf)](PF6) compounds developed, where M = Pt(ii) or Pd(ii) and HL = 8-hydroxyquinolyl derivatives, Pt-dppf-L4 (IC50 = 0.14 µM, SI = 48) was selected to perform an exploratory pre-clinical study in infected mice. This hit compound lacks acute toxicity when applied to animals in the dose/regimen described and exerts an anti-proliferative effect on parasites, which extends animal survival but is not curative.

High Throughput Approaches to Unravel the Mechanism of Action of a New Vanadium-Based Compound against Trypanosoma cruzi.

Treatment for Chagas disease, a parasitosis caused by Trypanosoma cruzi, has always been based on two drugs, nifurtimox and benznidazole, despite the toxic side effects described after prolonged prescription. In this work, we study a new prospective antitrypanosomal drug based on vanadium, here named VIVO(5Brsal)(aminophen). We found a good IC50 value, (3.76 ± 0.08) µM, on CL Brener epimastigotes. The analysis of cell death mechanism allowed us to rule out the implication of a mechanism based on early apoptosis or necrosis. Recovery assays revealed a trypanostatic effect, accompanied by cell shape and motility alterations. An uptake mostly associated with the insoluble fraction of the parasites was deduced through vanadium determinations. Concordantly, no drastic changes of the parasite transcriptome were detected after 6 h of treatment. Instead, proteomic analysis uncovered the modulation of proteins involved in different processes such as energy and redox metabolism, transport systems, detoxifying pathways, ribosomal protein synthesis, and proteasome protein degradation. Overall, the results here presented lead us to propose that VIVO(5Brsal)(aminophen) exerts a trypanostatic effect on T. cruzi affecting parasite insoluble proteins.

Dispersive liquid-liquid microextraction as a preconcentration alternative to increase ETAAS sensitivity in the analysis of molybdenum in bovine meat and pasture samples.

A new procedure combining dispersive liquid-liquid microextraction and electrothermal atomic absorption spectrometry was proposed for the determination of molybdenum in plants and animal tissue. The optimized conditions for the microextraction procedure were: 150 µL of 0.3 mol L-1 potassium ethyl xanthate (pH 1.4) and 1000 µL of acetonitrile and 50 µL of carbon tetrachloride as dispersive and extractant solvent, respectively. A limit of detection of 0.03 µg kg-1 and a limit of quantification of 0.11 µg kg-1, as well as an enrichment factor of 200, were achieved employing this procedure. Standard reference materials were used for trueness and precision evaluation. No significant differences were found at the 95% confidence level between certified and obtained values, and the precision expressed as repeatability (sr %) was 4.1% for animal tissue and 4.7% for plant material. The method was successfully applied for the determination of Mo in bovine meat and pasture samples from different livestock establishments.

Arsenic levels in groundwater and its correlation with relevant inorganic parameters in Uruguay: A medical geology perspective.

The aim of this work was to perform a groundwater quality monitoring in Uruguay, from the Medical Geology perspective, focusing attention on arsenic levels and its correlations with other relevant inorganic parameters. For this purpose, a total of 46 groundwater samples from private wells, out of the scope of the state-run water utility company, were analyzed. The accuracy of the analytical methods was ensured by using certified reference materials. Arsenic concentration range was 1.72-120.5 µg L-1, half of the samples being above the limit of 10 µg L-1 recommended by WHO for drinking water, with the corresponding risks for human health. Pearson correlations were performed, resulting in strong positive correlations for the pairs As/Cl-, As/F-, As/Na and As/V. These relationships between arsenic and other inorganic parameters in groundwater should be deeply studied, to prevent long-term health effects.

Bactericidal effect of pyridine-2-thiol 1-oxide sodium salt and its complex with iron against resistant clinical isolates of Mycobacterium tuberculosis.

The objective of this study was to determine the activity of pyridine-2-thiol 1-oxide sodium salt (Na mpo) and its complex with iron [Fe(mpo)3] against Mycobacterium tuberculosis. The compounds were tested against a standard strain of M. tuberculosis H37Rv (ATCC 27294), with minimal inhibitory concentrations (MIC90) of 7.20 and 1.07 µM to Na mpo and [Fe(mpo)3], respectively, and against three clinical isolates with different genotypic profiles, with MIC values ranging from 0.74 to 6.52 and 0.30 to 2.25 µM to Na mpo and [Fe(mpo)3], respectively. [Fe(mpo)3] was more effective against susceptible strains but both compounds were effective in inhibiting MDR and XDR-TB clinical strains. The profile activity was determined through the methodology of a time-kill curve against standard and clinical strains of M. tuberculosis. Time-kill studies indicated that Na mpo had an early bactericidal activity against H37Rv and clinical isolates, with sterilizing effects observed in 5 and 7 days, respectively, at its MIC90. The anti MDR and XDR-M. tuberculosis activity and bactericidal effect of Na mpo and [Fe(mpo)3] demonstrate their potential as new compounds for the treatment of tuberculosis.

Characterization of the effects involved in ultrasound-assisted extraction of trace elements from artichoke leaves and soybean seeds.

Ultrasonication is known to induce cavitation and can thus accelerate extraction, dissolution, digestion, and other processes, while the intense mixing caused by the propagation of ultrasound waves enhances analyte transfer to the extraction medium. Consequently, ultrasound-assisted extraction (UAE) is viewed as a simple procedure well suited for the extraction and subsequent determination of trace elements in food. Herein, we use different techniques to characterize several processes involved in probe- and bath-based UAE, apply them to the determination of trace metals (Cd, Cu, Ni, Pb, and Zn) in globe artichoke leaves and soybean seeds, and closely monitor the extraction rates of selected analytes. The developed UAE-based method is found to comply with the criteria of Green Chemistry and is concluded to be a reliable, simple, and cheap alternative to standard food analysis techniques.

Total arsenic and inorganic arsenic speciation in groundwater intended for human consumption in Uruguay: Correlation with fluoride, iron, manganese and sulfate.

Medical Geology is a growing field in Uruguay and the groundwater quality has been the focus of multiple studies, being As levels one of its main concerns. The aim of this study was the application of analytical methodologies for the assessment of total arsenic and inorganic arsenic species, fluoride, iron, manganese and sulfate in groundwater samples from private wells, used for human consumption, and to evaluate the possible correlations among these parameters. The accuracy of the methods was ensured by using certified reference materials. A total of 48 groundwater samples from Uruguay were analyzed. The concentration ranges found were: tAs (1.72-120.48) µg L-1, F- (0.024-1.528) mg L-1, Fe (0.62-211.38) µg L-1 and Mn (0.11-8.705) µg L-1. Almost half of the samples presented tAs concentration levels above those recommended by WHO for drinking water (10 µg L-1), with the corresponding risks for human health. Results showed higher As(V) levels in the samples, which is in agreement with the oxidant conditions of the wells. Pearson correlations were performed, resulting in strong positive correlations for As/F-, As(V)/F- and As(V)/SO42-. As levels in groundwater and its relationship with other inorganic parameters, should be deeply studied to prevent long-term health effects.

Effect of a new anti-T. cruzi metallic compound based on palladium.

Chagas disease is a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi. It is estimated that 6 million people are infected in Latin America. Current treatment is not effective due to the severe side effects and the limited efficacy towards the chronic phase of the disease. Considering the growing need for specific anti-Trypanosoma cruzi drugs, organometallic Pt and Pd based compounds were previously synthesized. Although the Pt-based compound effects on T. cruzi death have been reported, no mechanism of action has been proposed for the Pd-based analogous compound. In this work, we determined excellent to very good values of IC50 and SI. To analyze the compound mode of action, we measured Pd uptake and its association to the macromolecules of the parasite by electrothermal atomic absorption spectrometry. We found a poor uptake, which reaches only 16% after 24 h of incubation using 10× IC50, being the scarce incorporated metal preferentially associated to DNA. However, this compound has a trypanocidal effect, leading to morphological changes such as shortening of the parasite cell body and inducing necrosis after 24 h of treatment. Furthermore, this compound impairs the parasite development in the host both at the trypomastigote infection process and the intracellular amastigotes replication. In conclusion, our findings support that Pd-dppf-mpo compound constitutes a promising anti-T. cruzi compound effective against the chronic phase of the disease.