Unusual Application for Phosphonium Salts and Phosphoranes: Synthesis of Chalcogenides.

A novel strategy for the synthesis of sulfides and selenides from phosphonium salts and thio- or selenesulfonates, commercially available compounds, is described. When phosphoranes were used in the reaction, different products were obtained. The methodology does not require the use of metals, reactive species, or anhydrous conditions to be performed.

Regio- and diastereoselective Pd-catalyzed aminochlorocyclization of allylic carbamates: scope, derivatization, and mechanism.

The regio- and diastereoselective synthesis of oxazolidinones via a Pd-catalyzed vicinal C-N/C-Cl bond-forming reaction from internal alkenes of allylic carbamates is reported. The oxazolidinones are obtained in yields of 44 to 95% with high to excellent diastereoselectivities (from 6 : 1 to >20 : 1 dr) from readily available precursors. This process is scalable, and the products are suitable for the synthesis of useful amino alcohols. A detailed theoretical and experimental mechanistic study was carried out to describe that the reaction proceeds through an anti-aminopalladation of the alkene followed by an oxidative C-Pd(ii) cleavage with retention of the carbon stereochemistry to yield the major diastereomer. The role of Cu(ii) in a C-Cl bond-forming mechanism step has also been proposed.

Metabolomic profile as a noninvasive adjunct tool for the diagnosis of Grades III and IV endometriosis-related infertility.

The aim of the present case-control study was to develop a noninvasive adjuvant tool for the diagnosis of endometriosis. Serum samples from 100 patients undergoing intracytoplasmic sperm injection were split into two groups according to the cause of infertility: an endometriosis group (n = 50), consisting of samples derived from patients with Grade III and IV endometriosis, and a control group (n = 50), comprising samples derived from patients with isolated male factor infertility. The metabolomic profile of each sample was obtained, through mass spectrometry. Partial least squares discriminant analysis was able to clearly classify the endometriosis and control groups. Ten potential biomarkers were selected based on their importance for model prediction. These ions were used to build the receiver-operating characteristic curve, which presented an area under the curve of 0.904 (95% confidence interval: 0.796-0.985). To validate the model, 30 other samples from infertile women without any evidence of endometriosis were tested. Considering these ions as possible biomarkers, the model was able to correctly classify 84% of the patients. Finally, a similar prediction potential was observed in the model validated set, when samples from the disease-free group were tested. Serum metabolomics may be useful as a noninvasive adjunct tool for the selection of patients who must undergo laparoscopy for definitive endometriosis diagnosis.

Structure-activity relationship study of antitrypanosomal chalcone derivatives using multivariate analysis.

Chagas disease represents one of several neglected diseases with a reduced number of chemotherapeutical drugs including the highly toxic compounds benznidazole and nifurtimox. In this sense, natural products represent an import scaffold for the discovery of new biologically active compounds, in which chalcones are promising representatives due to their antitrypanosomal potential. In this work, a series of 36 chalcone derivatives were synthesized and tested against trypomastigotes of Trypanosoma cruzi. In addition, a detailed investigation on their molecular features was performed. The obtained results suggest that certain molecular features are fundamental for an efficient antitrypanosomal potential of chalcones, such as allylic groups, α,ß-unsaturated carbonyl system, and aromatic hydroxyl groups. These results were obtained based on the interpretation of machine-learning and multivariate statistical methods, which revealed the essential characteristics of chalcone prototypes against trypomastigotes of T. cruzi.

Antitrypanosomal Acetylene Fatty Acid Derivatives from the Seeds of Porcelia macrocarpa (Annonaceae).

Chagas' disease is caused by a parasitic protozoan and affects the poorest population in the world, causing high mortality and morbidity. As a result of the toxicity and long duration of current treatments, the discovery of novel and more efficacious drugs is crucial. In this work, the hexane extract from seeds of Porcelia macrocarpa R.E. Fries (Annonaceae) displayed in vitro antitrypanosomal activity against trypomastigote forms of T. cruzi by the colorimetric MTT assay (IC50 of 65.44 µg/mL). Using chromatographic fractionation over SiO2, this extract afforded a fraction composed by one active compound (IC50 of 10.70 µg/mL), which was chemically characterized as 12,14-octadecadiynoic acid (macrocarpic acid). Additionally, two new inactive acetylene compounds (α,α'-dimacro-carpoyl-ß-oleylglycerol and α-macrocarpoyl-α'-oleylglycerol) were also isolated from the hexane extract. The complete characterization of the isolated compounds was performed by analysis of NMR and MS data as well as preparation of derivatives.

Dissociation of deprotonated microcystin variants by collision-induced dissociation following electrospray ionization.

Microcystins (MC) are a family of hepatotoxic cyclic heptapeptides produced by a number of different cyanobacterial species. Considering the recent advances in the characterization of deprotonated peptides by mass spectrometry, the fragmentation behavior of four structurally related microcystin compounds was investigated using collision-induced dissociation (CID) experiments on an orbitrap mass spectrometer. It is demonstrated in this study that significant structural information can be obtained from the CID spectra of deprotonated microcystins. A predominant ring-opening reaction at the isoMeAsp residue, as well as two major complementary fragmentation pathways, was observed, reducing the complexity of the product ion spectra in comparison with spectra observed from protonated species. This proposed fragmentation behavior was applied to characterize [Leu(1)]MC-LR from a cyanobacterial cell extract. In conclusion, CID spectra of microcystins in the negative ion mode provide rich structurally informative mass spectra which greatly enhance confidence in structural assignments, in particular when combined with complementary positive ion CID spectra.

Determination of chlorpheniramine in human plasma by HPLC-ESI-MS/MS: application to a dexchlorpheniramine comparative bioavailability study.

In the present study a fast, sensitive and robust validated method to quantify chlorpheniramine in human plasma using brompheniramine as internal standard (IS) is described. The analyte and the IS were extracted from plasma by LLE (diethyl ether-dichloromethane, 80:20, v/v) and analyzed by HPLC-ESI-MS/MS. Chromatographic separation was performed using a gradient of methanol from 35 to 90% with 2.5 mm NH(4)OH on a Gemini Phenomenex C(8) 5 microm column (50 x 4.6 mm i.d.) in 5.0 min/run. The method fitted to a linear calibration curve (0.05-10 ng/mL, R > 0.9991). The precision (%CV) and accuracy ranged, respectively: intra-batch from 1.5 to 6.8% and 99.1 to 106.6%, and inter-batch from 2.4 to 9.0%, and 99.9 to 103.1%. The validated bioanalytical procedure was used to assess the comparative bioavailability in healthy volunteers of two dexchlorpheniramine 2.0 mg tablet formulations (test dexchlorpheniramine, Eurofarma, and reference Celestamine, Schering-Plough). The study was conducted using an open, randomized, two-period crossover design with a 2 week washout interval. Since the 90% confidence interval for C(max) and AUC ratios were all within the 80-125% interval proposed by ANVISA and FDA, it was concluded that test and reference formulations are bioequivalent concerning the rate and the extent of absorption.

Development and validation of a rapid and sensitive LC-ESI-MS/MS method for ondansetron quantification in human plasma and its application in comparative bioavailability study.

The validation of a high throughput and specific method using a high-performance liquid chromatography coupled to electrospray (ES+) ionization tandem triple quadrupole mass spectrometric (LC-ESI-MS/MS) method for ondansetron quantification in human plasma is described. Human plasma samples were extracted by liquid-liquid extraction (LLE) using methyl tert-butyl ether and analyzed by LC-ESI-MS/MS. The limit of quantification was 0.2 ng/mL and the method was linear in the range 0.2-60 ng/mL. The intra-assay precisions ranged from 1.6 to 7.7%, while inter-assay precisions ranged from 2.1 to 5.1%. The intra-assay accuracies ranged from 97.5 to 108.2%, and the inter-assay accuracies ranged from 97.3 to 107.0%. The analytical method was applied to evaluate the relative bioavailability of two pharmaceutical formulations containing 8 mg of ondansetron each in 25 healthy volunteers using a randomized, two-period crossover design. The geometric mean and respective 90% confidence interval (CI) of ondansetron test/reference percent ratios were 90.15% (81.74-99.44%) for C(max) and 93.11% (83.01-104.43%) for AUC(0-t). Based on the 90% confidence interval of the individual ratios (test formulation/reference formulation) for C(max) and AUC(0-inf), it was concluded that the test formulation is bioequivalent to the reference one with respect to the rate and extent of absorption of ondansetron.

Quantification of chlordesmethyldiazepam by liquid chromatography-tandem mass spectrometry: application to a cloxazolam bioequivalence study.

A rapid, sensitive and specific LC-MS/MS method was developed and validated for quantifying chlordesmethyldiazepam (CDDZ or delorazepam), the active metabolite of cloxazolam, in human plasma. In the analytical assay, bromazepam (internal standard) and CDDZ were extracted using a liquid-liquid extraction (diethyl-ether/hexane, 80/20, v/v) procedure. The LC-MS/MS method on a RP-C18 column had an overall run time of 5.0 min and was linear (1/x weighted) over the range 0.5-50 ng/mL (R > 0.999). The between-run precision was 8.0% (1.5 ng/mL), 7.6% (9 ng/mL), 7.4% (40 ng/mL), and 10.9% at the low limit of quantification-LLOQ (0.500 ng/mL). The between-run accuracies were 0.1, -1.5, -2.7 and 8.7% for the above mentioned concentrations, respectively. All current bioanalytical method validation requirements (FDA and ANVISA) were achieved and it was applied to the bioequivalence study (Cloxazolam -- test, Eurofarma Lab. Ltda and Olcadil -- reference, Novartis Biociências S/A). The relative bioavailability between both formulations was assessed by calculating individual test/reference ratios for Cmax, AUClast and AUC0-inf. The pharmacokinetic profiles indicated bioequivalence since all ratios were as proposed by FDA and ANVISA.

Serum metabolites as predictive molecular markers of ovarian response to controlled stimulation: a pilot study.

OBJECTIVE: This study aimed to look into the use of serum metabolites as potential biomarkers of response to controlled ovarian stimulation (COS) in patients undergoing intracytoplasmic sperm injection (ICSI) cycles. METHODS: This case-control study analyzed serum samples from 30 patients aged <36 years undergoing COS for ICSI in a university-affiliated assisted reproduction center from January 2017 to August 2017. The samples were split into three groups based on response to COS as follows: poor responders: <4 retrieved oocytes (PR group, n=10); normal responders: ≥ 8 and ≤ 12 retrieved oocytes (NR group, n=10); and hyper-responders: >25 retrieved oocytes (HR, n=10). The metabolic profiles of the serum samples were compared between the groups through Principal Component Analysis (PCA). Receiver Operating Characteristic (ROC) curves were built to assess the power of the model at predicting response to COS. RESULTS: PCA clearly distinguished between PR, NR and HR, and 10 ions were chosen as potential biomarkers of response to COS. These ions were more specific for PR than for NR. The ROC curve considering PR and NR had an area under the curve of 99.6% (95% CI: 88.9 - 100%). CONCLUSION: The preliminary evidence discussed in this study suggests that serum metabolites may be used as predictive molecular markers of ovarian response to controlled stimulation. The integration of clinical and "omics" findings may allow the migration toward an era of personalized treatment in reproductive medicine.

Multiple sclerosis has a distinct lipid signature in plasma and cerebrospinal fluid.

OBJECTIVE: The diagnosis of multiple sclerosis (MS) has changed over the last decade, but remains a composite of clinical assessment and magnetic resonance imaging to prove dissemination of lesions in time and space. The intrathecal synthesis of immunoglobulin may be a nonspecific marker and there are no plasma biomarkers that are useful in the diagnosis of MS, presenting additional challenges to their early detection. METHODS: We performed a preliminary untargeted qualitative lipidomics mass spectrometry analysis, comparing cerebrospinal fluid (CSF) and plasma samples from patients with MS, other inflammatory neurological diseases and idiopathic intracranial hypertension. RESULTS: Lipid identification revealed that fatty acids and sphingolipids were the most abundant classes of lipids in the CSF and that glycerolipids and fatty acids were the main class of lipids in the plasma of patients with MS. The area under the curve was 0.995 (0.912-1) and 0.78 (0.583-0.917), respectively. The permutation test indicated that this ion combination was useful for distinguishing MS from other inflammatory diseases (p < 0.001 and 0.055, respectively). CONCLUSION: This study concluded that the CSF and plasma from patients with MS bear a unique lipid signature that can be useful as a diagnostic biomarker.

Evaluation of Ca2+ Binding Sites in Tacrolimus by Infrared Multiple Photon Dissociation Spectroscopy.

Tacrolimus (TAC) is an efficient immunosuppressant used in organ transplantation procedures. There is an intrinsic correlation between TAC and Ca2+ because of the dependence of its action mechanism on calcium and calcineurin, and the role of ion coordination on TAC identification and quantitation. To depict the Ca2+ binding sites in TAC, this work carried out gas-phase vibrational infrared multiple photon dissociation spectroscopy of [Ca(TAC)]2+ and of three other TAC mimetic molecules (probes 1-3). Density functional theory (DFT) and Monte Carlo (MC) simulations were also used to support the experimental data assignment, and natural bond orbital (NBO) analysis was carried out to depict the coordination sphere. PM3 and B3LYP/6-31G(d) levels of theory displayed similar trends during the MC simulations, suggesting that PM3 is a viable alternative to more expensive DFT calculations, at least during the conformational analysis step. Infrared spectroscopy of the [Ca(probe X)1]2+ and [Ca(probe X)3]2+ ( X = 1-3) complexes allowed for a useful guide for building guess geometries and for the band assignment of the [Ca(TAC)]2+ complex. Nevertheless, the MC approach was particularly useful for exploring the potential energy surface. The lowest energy conformation for [Ca(TAC)]2+ was found by MC simulations and is 32.92 kJ mol-1 lower in energy than the one found by comparing the results obtained for Ca2+ coordination in probes, despite the calculated spectra being virtually identical. Both approaches are good ways to depict the coordination sites, and these results suggest that using small molecules as models is a reliable approach to depict the geometry or coordination sites of extensive ions, yielding a robust correlation between experimental and theoretical spectra. Furthermore, MC survey produced a lower energy conformation with a good match to the experimental results. Both methods depict the Ca2+ coordination sphere as a hexacoordinated environment where the main coordination centers are carbonyl groups.

Quantification of carvedilol in human plasma by high-performance liquid chromatography coupled to electrospray tandem mass spectrometry: application to bioequivalence study.

A rapid, sensitive and specific method to quantify carvedilol in human plasma using metoprolol as the internal standard (IS) is described. The analyte and the IS were extracted from plasma by liquid-liquid extraction using a diethyl-ether solvent. After removed and dried the organic phase, the extracts were reconstituted with a fixed volume of acetonitrile-water (50/50; v/v). The extracts were analyzed by a high performance liquid chromatography coupled to electrospray tandem mass spectrometry (HPLC-MS/MS). Chromatography was performed isocratically on Alltech Prevail C18 5 microm analytical column, (150 mm x 4.6 mm i.d.). The method had a chromatographic run time of 3.5 min and a linear calibration curve over the range 0.1-200 ng ml(-1) (r2>0.997992). The limit of quantification was 0.1 ng ml(-1). This HPLC-MS/MS procedure was used to assess the bioequivalence of two carvedilol 25 mg tablet formulations (carvedilol test formulation from Laboratórios Biosintética Ltda and Coreg from Roche Químicos e Farmacêuticos S.A standard reference formulation). A single 25 mg dose of each formulation was administered to healthy volunteers. The study was conducted using an open, randomized, two-period crossover design with a 2-week wash-out interval. Since the 90% CI for C(max) and AUCs ratios were all inside the 80-125% interval proposed by the US Food and Drug Administration Agency, it was concluded that carvedilol formulation elaborated by Laboratórios Biosintética Ltda is bioequivalent to Coreg formulation for both the rate and the extent of absorption.

Chemical composition and in vitro cytotoxic and antileishmanial activities of extract and essential oil from leaves of Piper cernuum.

Fractionation of the MeOH extract from leaves of Piper cernuum Vell. (Piperaceae) afforded six phenylpropanoid derivatives: 3',4'-dimethoxydihydrocinnamic acid (1), piplaroxide (2), methyl 4'-hydroxy-3',5'-dimethoxy cinnamate (3), 3',4',5'-trimethoxydihydrocinnamic acid (3), dihydropiplartine (5), and piplartine (6). The structures of isolated metabolites were characterized by NMR and MS spectral data analysis. The chemical composition of essential oil from the leaves was determined using GC/LREIMS followed by the determination of Kovats indexes. This procedure allowed the identification of nineteen terpenoids, with ß-elemene (7), bicyclogermacrene (8), germacrene D (9), and (E)-caryophyllene (10) as the main compounds. Compounds 1 and 3-6 displayed no in vitro cytotoxicity against cancer cell lineages B16F10-Nex2, U87, HeLa, HL-60, HCT, and A2058 while 2 showed moderate activity against B16F10-Nex2 and HL-60 lines. Otherwise, compounds 7-10 displayed high cytotoxic activity. Evaluation against non-tumorigenic HFF cells indicated a reduced selectivity of compounds 7-10 to tumoral cells. No antileishmanial activity on macrophages infected with L. (L.) amnazonensis was found for the crude MeOH extract and compounds 1-6. The crude essential oil and compounds 7-10 reduced parasitism and eliminated the majority of infected and non-infected cells at 50 µg/mL.

Multiple sclerosis has a distinct lipid signature in plasma and cerebrospinal fluid / Esclerose múltipla tem uma assinatura lipídica distinta no plasma e no líquido cefalorraquiano

ABSTRACT The diagnosis of multiple sclerosis (MS) has changed over the last decade, but remains a composite of clinical assessment and magnetic resonance imaging to prove dissemination of lesions in time and space. The intrathecal synthesis of immunoglobulin may be a nonspecific marker and there are no plasma biomarkers that are useful in the diagnosis of MS, presenting additional challenges to their early detection. Methods We performed a preliminary untargeted qualitative lipidomics mass spectrometry analysis, comparing cerebrospinal fluid (CSF) and plasma samples from patients with MS, other inflammatory neurological diseases and idiopathic intracranial hypertension. Results Lipid identification revealed that fatty acids and sphingolipids were the most abundant classes of lipids in the CSF and that glycerolipids and fatty acids were the main class of lipids in the plasma of patients with MS. The area under the curve was 0.995 (0.912-1) and 0.78 (0.583-0.917), respectively. The permutation test indicated that this ion combination was useful for distinguishing MS from other inflammatory diseases (p < 0.001 and 0.055, respectively). Conclusion This study concluded that the CSF and plasma from patients with MS bear a unique lipid signature that can be useful as a diagnostic biomarker.

RESUMO Embora o diagnóstico da EM tenha se modificado na última década, ainda tem como requisito básico a demonstração da disseminação no tempo e no espaço, através do quadro clínico e do exame de ressonância magnética. A síntese intratecal de imunoglobulina pode ser um marcador inespecífico e não há biomarcadores plasmáticos que sejam úteis no diagnóstico da EM, impondo desafios à sua detecção precoce. Métodos Realizamos uma análise lipidômica preliminar por espectrometria de massas, não direcionada, qualitativa, comparando amostras de LCR e plasma de pacientes com EM, outras doenças neurológicas inflamatórias e hipertensão intracraniana idiopática (HII). Resultados A identificação lipídica revelou que os ácidos graxos e esfingolipídios foram as classes mais abundantes de lipídios no LCR e que glicerolipídios e ácidos graxos foram a principal classe de lipídios no plasma de pacientes com EM. A AUC foi de 0,995 (0,912-1) e 0,78 (0,583-0,917), respectivamente. O teste de permutação indicou que essa combinação de íons foi útil para distinguir a EM de outras doenças inflamatórias (p < 0,001 e 0,055, respectivamente). Conclusão Este estudo sugere que o líquido cefalorraquidiano (LCR) e o plasma de pacientes com EM possuem uma assinatura lipídica única, pode ser útil como um biomarcador diagnóstico.

Pharmacodynamic evaluation and physical/chemical analysis of two formulations of propofol used in target-controlled infusion.

BACKGROUND AND OBJECTIVES: There are several formulations of propofol available to the anesthesiologist for clinical use. The aim of this study was to analyze the physicochemical properties, pharmacodynamic effect, and pharmaceutical and clinical equivalence of the reference drug propofol as well as a similar formulation. METHOD: Sixteen volunteers were enrolled in this randomized, double-blind, and paired study of Diprivan and Propovan formulations. Formulations were given as target-controlled infusion with target concentration of 3.0 µg.mL(-1) for 15 minutes. Variables studied were the area under the curve (AUC) of the bispectral index (BIS) graph regarding time, minimum BIS reached and time to reach it, and recovery time. The two formulations were sent to analysis of particle size of lipid emulsion, surface potential, and active principle quantification. RESULTS: There was no difference between the formulations when comparing AUC, minimum BIS reached and time to reach it. The similar formulation recovery time was lower compared to the reference formulation (eight and 10 min, respectively, p=0.014). Mean particle size of lipid emulsion, surface potential, and active ingredient quantification were similar for both formulations. CONCLUSION: There was no clinically significant difference between the use of propofol, reference Diprivan, and the similar Propovan during infusion. However, the recovery time was longer with the reference drug. Although analysis of both formulations studied show similar results regarding its physicochemical characterization, further studies should be conducted to justify this difference.

Pharmacodynamic evaluation and physical/chemical analysis of two formulations of propofol used in target-controlled infusion.

BACKGROUND AND OBJECTIVES: There are several formulations of propofol available to the anesthesiologist for clinical use. The aim of this study was to analyze the physicochemical properties, pharmacodynamic effect, and pharmaceutical and clinical equivalence of the reference drug propofol as well as a similar formulation. METHOD: Sixteen volunteers were enrolled in this randomized, double-blind, and paired study of Diprivan® and Propovan® formulations. Formulations were given as target-controlled infusion with target concentration of 3.0 µg.mL(-1) for 15 minutes. Variables studied were the area under the curve (AUC) of the bispectral index (BIS) graph regarding time, minimum BIS reached and time to reach it, and recovery time. The two formulations were sent to analysis of particle size of lipid emulsion, surface potential, and active principle quantification. RESULTS: There was no difference between the formulations when comparing AUC, minimum BIS reached and time to reach it. The similar formulation recovery time was lower compared to the reference formulation (eight and 10 min, respectively, p = 0.014). Mean particle size of lipid emulsion, surface potential, and active ingredient quantification were similar for both formulations. CONCLUSION: There was no clinically significant difference between the use of propofol, reference Diprivan®, and the similar Propovan® during infusion. However, the recovery time was longer with the reference drug. Although analysis of both formulations studied show similar results regarding its physicochemical characterization, further studies should be conducted to justify this difference.

Determination of dimethyltryptamine and ß-carbolines (ayahuasca alkaloids) in plasma samples by LC-MS/MS.

BACKGROUND: Ayahuasca is a psychoactive plant beverage originally used by indigenous people throughout the Amazon Basin, long before its modern use by syncretic religious groups established in Brazil, the USA and European countries. The objective of this study was to develop a method for quantification of dimethyltryptamine and ß-carbolines in human plasma samples. RESULTS: The analytes were extracted by means of C18 cartridges and injected into LC-MS/MS, operated in positive ion mode and multiple reaction monitoring. The LOQs obtained for all analytes were below 0.5 ng/ml. By using the weighted least squares linear regression, the accuracy of the analytical method was improved at the lower end of the calibration curve (from 0.5 to 100 ng/ml; r(2)> 0.98). CONCLUSION: The method proved to be simple, rapid and useful to estimate administered doses for further pharmacological and toxicological investigations of ayahuasca exposure.

Antioxidant therapy attenuates oxidative insult caused by benzonidazole in chronic Chagas' heart disease.

Chronic chagasic cardiac patients are exposed to oxidative stress that apparently contributes to disease progression. Benznidazole (BZN) is the main drug used for the treatment of chagasic patients and its action involves the generation of reactive species. 41 patients with Chagas' heart disease were selected and biomarkers of oxidative stress were measured before and after 2 months of BZN treatment (5 mg/kg/day) and the subsequent antioxidant supplementation with vitamin E (800 UI/day) and C (500 mg/day) during 6 months. Patients were classified according to the modified Los Andes clinical hemodynamic classification in groups IA, IB, II and III, and the activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase (GST) and glutathione reductase (GR), as well as the contents of reduced glutathione (GSH), thiobarbituric acid reactive species (TBARS), protein carbonyl (PC), vitamin E and C and nitric oxide (NO), myeloperoxidase (MPO) and adenosine deaminase (ADA) activities were measured in their blood. Excepting in group III, after BZN treatment SOD, CAT, GPx and GST activities as well as PC levels were enhanced while vitamin E levels were decreased in these groups. After antioxidant supplementation the activities of SOD, GPx and GR were decreased whereas PC, TBARS, NO, and GSH levels were decreased. In conclusion, BZN treatment promoted an oxidative insult in such patients while the antioxidant supplementation was able to attenuate this effect by increasing vitamin E levels, decreasing PC and TBARS levels, inhibiting SOD, GPx and GR activities as well as inflammatory markers, mainly in stages with less cardiac involvement.