Fernando Armani Aguiar

Characterization and pharmacological evaluation of febrile response on zymosan-induced arthritis in rats

The present study investigated the febrile response in zymosan-induced arthritis, as well as the increase in PGE(2) concentration in the cerebrospinal fluid (CSF), along with the effects of antipyretic drugs on these responses in rats. Zymosan intra-articularly injected at the dose of 0.5 mg did not affect the body core temperature (Tc) compared with saline (control), whereas at doses of 1 and 2 mg, zymosan promoted a flattened increase in Tc and declined thereafter. The dose of 4 mg of zymosan was selected for further experiments because it elicited a marked and long-lasting Tc elevation starting at 3 1/2 h, peaking at 5 1/2 h, and remaining until 10 h. This temperature increase was preceded by a decrease in the tail skin temperature, as well as hyperalgesia and edema in the knee joint. No febrile response was observed in the following days. In addition, zymosan-induced fever was not modified by the sciatic nerve excision. Zymosan increased PGE(2) concentration in the CSF but not in the plasma. Oral pretreatment with ibuprofen (5-20 mg/kg), celecoxib (1-10 mg/kg), dipyrone (60-240 mg/kg), and paracetamol (100-200 mg/kg) or subcutaneous injection of dexamethasone (0.25-1.0 mg/kg) dose-dependently reduced or prevented the fever during the zymosan-induced arthritis. Celecoxib (5 mg/kg), paracetamol (150 mg/kg), and dipyrone (120 mg/kg) decreased CSF PGE(2) concentration and fever during zymosan-induced arthritis, suggesting the involvement of PGE(2) in this response.

Capillary electrophoresis method for the determination of isradipine enantiomers: Stability studies and pharmaceutical formulation analysis

A simple enantioselective method based on CE using CD as chiral selector was developed and validated for the determination of isradipine (IRD) enantiomers in a pharmaceutical formulation and for the determination of IRD enantiomers in degradation studies. After optimization, the best results were obtained using 15 mM borate buffer at pH 9.3 and sulfobutyl ether‐β‐cyclodextrin (2.5%, w/v) as chiral selector. The applied voltage was +30 kV, and the sample injection was performed in the hydrodynamic mode. All analyses were carried out in a fused‐silica uncoated capillary with an id of 50 μm and total length of 60.0 cm. Under these conditions, a complete separation between IRD enantiomers was achieved in less than 7 min. Linearity was obtained in the range 50–300 μg/mL for both enantiomers (r≥0.9978). The RSD (%) and relative errors (%) obtained in precision and accuracy studies (intra‐day and inter‐day) were lower than 5%. Therefore, this method was found to be appropriate for controlling pharmaceutical formulations containing IRD enantiomers and the assay was considered to be stability indicating. The drug was subjected to oxidation, hydrolysis and photolysis. In all stress conditions the drug presented considerable degradation when compared with a fresh sample (zero time).

Dipyrone metabolite 4-MAA induces hypothermia and inhibits PGE2-dependent and -independent fever while 4-AA only blocks PGE2-dependent fever

The antipyretic and hypothermic prodrug dipyrone prevents PGE2‐dependent and ‐independent fever induced by LPS from Escherichia coli and Tityus serrulatus venom (Tsv) respectively. We aimed to identify the dipyrone metabolites responsible for the antipyretic and hypothermic effects.

Quantitative Determination of Lercanidipine Enantiomers in Commercial Formulations by Capillary Electrophoresis

An enantioselective method based on capillary electrophoresis (CE) using cyclodextrin (CD) as chiral selector was developed and validated for determination of lercanidipine (LER) enantiomers, a drug calcium channel blocker which exerts antihypertensive effects of long duration, in a pharmaceutical formulation. Optimum separation of LER enantiomers was obtained on a 50 cm × 50 μm id capillary using a sodium acetate buffer solution 200 mmol/L pH 4.0 containing 10 mmol/L of 2,3,6-o-methyl-β-cyclodextrin (TM-β-CD) as background electrolyte. The capillary temperature and voltage were 15°C and 25 kV, respectively, hydrodynamic injection and detection at 237 nm. Linearity was obtained in the range 12.5–100 μg/mL for both enantiomers (r ≥ 0.995). The RSD (%) and relative errors (E, %) obtained in precision and accuracy studies (intraday and interday) were lower than 5%. After validation, the method was applied to quantify the enantiomers of LER in commercial tablets and the results were satisfactory in terms of accuracy and precision, both less than 5%. Therefore, this method was found to be appropriate for enantioselective quality control of LER enantiomers in pharmaceutical formulations.

Quercetin does not alter lipopolysaccharide-induced fever in rats.

Fever is considered an important component of the acute phase response of the body in defence against invading organisms such as bacteria. Quercetin, an important representative of the flavonoid class, has been extensively studied as an anti‐inflammatory agent. In the present study, we investigated the effect of quercetin, administered orally (5, 25 and 50 mg kg−1) or intraperitoneally (50 mg kg−1), on the febrile response induced by either intraperitoneally (50 μg kg−1) or intravenously (5 μg kg−1) injected lipopolysaccharide (LPS from Escherichia coli) in rats. In contrast with the well known anti‐inflammatory activity of quercetin, the results demonstrate that quercetin, at the doses used, did not alter the fever induced by LPS, regardless of the route of administration.

Simultaneous determination of dipyrone metabolites in rat hypothalamus, cerebrospinal fluid and plasma samples by LC-MS/MS

BACKGROUND After oral administration dipyrone is rapidly hydrolyzed to 4-methylaminoantipyrine, which is absorbed and further metabolized to 4-formylaminoantipyrine and to 4-aminoantipyrine, which is acetylated by a polymorphic N-acetyltransferase system to 4-acetylaminoantipyrine. To evaluate the presence of dipyrone metabolites in different rat matrices after intraperitoneal administration, an analytical method was developed and validated. METHODOLOGY The four main dipyrone metabolites were extracted from plasma, cerebrospinal fluid and hypothalamus samples by LLE prior to LC-MS/MS. RESULTS Standard calibration graphs for all metabolites were linear (r > 0.99). The intra- and inter-day precision and accuracy values were both inferior to 15%. CONCLUSION This method is simple and specific for studying dipyrone metabolites after intraperitoneal administration.

Development, validation, and application of a capillary electrophoresis method for analysis of cytokine interferon alpha-2a in pharmaceutical formulations

A simple CE based method was developed and validated for the determination of interferon alpha-2a in a pharmaceutical formulation. After optimization, the best results were obtained using 30 mmol L−1 tetraborate buffer at pH 8.50 with 50 mmol L−1 of sodium dodecyl sulfate, and using a diode array detector at 200 nm. The applied voltage was +25 kV, and the sample injection was performed in the hydrodynamic mode. All analyses were carried out in a fused-silica uncoated capillary with an i.d. of 75 μm and an effective length of 50.0 cm. Under these conditions, the analysis was achieved in less than 12 min. Linearity was obtained in the range 0.41–1.54 MIU mL−1 (r ≥ 0.997). The RSD (%) and relative errors (%) obtained in precision and accuracy studies (intra-day and inter-day) were lower than 5%. Therefore, this method was found to be appropriate for controlling pharmaceutical formulations containing interferon alpha-2a.

Phytochemical Study of Tapirira guianensis Leaves Guided by Vasodilatory and Antioxidant Activities

The aim of this research was to perform a phytochemical study of the methanol leaves extract of T. guianensis (MET) guided by vasodilatory and antioxidant activities. The chemical profile of MET and the ethyl acetate fraction (EA fraction) was determined by HPLC-UV-MS and EA fraction guided fractionation by reverse-phase chromatography. The vasorelaxant effects of MET, fractions, sub-fractions and constituents were assessed on rat aorta pre-contracted with phenylephrine. Antioxidant activity was evaluated by using a DPPH assay. The results show that MET-induced vasodilation was dependent on NO/cGMP; and that the PI3K/Akt pathway seems to be the main route involved in eNOS activation. The EA fraction showed greater vasodilatory and antioxidant potency and was submitted to further fractionation. This allowed the isolation and characterization of quercetin, quercetin 3-O-(6″-O-galloyl)-β-d-galactopyranoside and 1,4,6-tri-O-galloyl-β-d-glucose. Also, galloyl-HHDP-hexoside and myricetin deoxyhexoside were identified by HPLC-UV-MS. These compounds are being described for the first time for T. guianensis. 1,4,6-tri-O-galloyl-β-d-glucose and quercetin 3-O-(6″-O-galloyl)-β-d-galactopyranoside showed no vasodilatory activity. Quercetin and myricetin glycoside seems to contribute to the MET activity, since they have been reported as vasodilatory flavonoids. MET-induced vasodilation could contribute to the hypotensive effect of T. guianensis previously reported.

DETERMINATION OF DICLOFENAC SODIUM IN RABBIT PLASMA BY HPLC/UV: EVALUATION AND CHARACTERIZATION OF ITS CRYSTALLINE FORMS, ANHYDROUS AND HYDRATE, ON THE ANTIPYRETIC EFFECT

Diclofenac sodium (DS) is a non-steroidal anti-inflammatory drug that is widely prescribed for the treatment of rheumatoid arthritis and post-surgery analgesia. The active pharmaceutical ingredient is the anhydrous form; however, it can also exist in hydrate form. In this context, knowing the properties of the solid state is important and relevant in the pharmaceutical area because they have a significant impact on the solubility, bioavailability, and chemical stability of the drugs. In the present study, data from XRPD, FTIR spectroscopy, and thermal analysis were used for the identification and characterization of DS forms (anhydrous and hydrate). An HPLC method was optimized to evaluate the plasma concentration of DS in rabbits. The optimized method exhibited good linearity over the range 0.1–60 µg/mL with correlation coefficients of >0.9991. The mean recovery was 100%. Precision and accuracy were determined within acceptable limits. Finally, to compare the pharmacological properties of anhydrous and hydrate DS forms, we investigated their effects in the febrile response induced by lipopolysaccharide from E. coli in rabbits. The results show that the antipyretic effect of anhydrous and hydrate DS forms are similar.