Cabral Pavei

The effect of aqueous extract of gross and commercial yerba mate (Ilex paraguariensis) on intra-abdominal and epididymal fat and glucose levels in male Wistar rats

This study analyzed the plasma lipid profile, glucose levels and fat deposits in male rats treated with aqueous extract of gross yerba mate, commercial yerba mate or water. Yerba mate treatment did not change body weight gain and lipid profile. The consumption of gross yerba mate significantly increased blood glucose (6.6 mmol/L) as compared to the water (4.8 mmol/L) and commercial group (5.2 mmol/L) and decreased epididymal and intra-abdominal deposits (10.1mg/g and 23.7 mg/g of weight) as compared to the water (15.4 mg/g and 36.9 mg/g of weight) and commercial group (12.5mg/g and 28 mg/g of weight). The results suggest that gross yerba mate reduces fat more efficiently but produces a greater increase in blood glucose when compared to commercial yerba mate and water groups.

Solid state evaluation of some thalidomide raw materials.

Thalidomide presents polymorphism and is a problematic drug due to its poor solubility and difficult tablet processability, which is the dosage form available in Brazil. In most cases, the pharmacopoeias specify do not address solid state characterization of drugs precisely. In this work, different thalidomide commercial samples were characterized by infrared spectroscopy, particle size analysis, scanning electron microscopy, and X-ray diffraction. In addition, the polymorphic forms were quantified for Rietveld analysis and their intrinsic dissolution rates were evaluated. The results demonstrated the market availability of different raw materials which lack of homogeneity due to differences related to crystalline constitution, crystal habit and intrinsic dissolution rate.

HPLC-PDA method for quinovic acid glycosides assay in Cat's claw (Uncaria tomentosa) associated with UPLC/Q-TOF–MS analysis

Uncaria tomentosa (Willd.) is a medicinal plant largely used in folk medicine due to its wide range of biological activities, many of which are usually ascribed to the two main classes of secondary metabolites, namely, alkaloids and quinovic acid glycosides. In this work, a reversed phase HPLC-PDA method was developed and validated for the assay of quinovic acid glycosides in crude and dried extracts of Uncaria tomentosa (Cats claw) bark. The validation comprised tests of specificity, accuracy, linearity, intermediate precision, repeatability and limits of detection and of quantification. Alpha-hederin was used as the external standard. High coefficients of determination with lower R.S.D. were achieved for both external standard and crude extract. The structural characterization of the main quinovic acid glycosides presented in the crude extract was carried out through UPLC/Q-TOF-MS. The identities of the compounds were obtained through the comparison of their fragmentation patterns with those reported in the literature. The analytical method was successfully applied for quantifying quinovic acid glycosides in two different dried extracts from U. tomentosa and in one quinovic acid glycosides purified fraction.

COMPARISON OF METHYLXANTHINE, PHENOLICS AND SAPONIN CONTENTS IN LEAVES, BRANCHES AND UNRIPE FRUITS FROM ILEX PARAGUARIENSIS A. ST.-HIL (MATE)

The contents of the methylxanthines, saponins, and phenolics compounds were determined in different parts of Ilex paraguariensis A. St. Hil., in order to investigate the chemical differences among some well explored plant parts, leaves and branches, with an innovative raw source as the unripe fruits of the specie. Three specific LC methods were employed. In the plant raw material, the highest contents of methylxanthines (caffeine and theobromine) were found in leaves (0.86% and 0.15%), while the lowest content was observed in the fruits (0.04% and 0.04%). In the same manner the content of phenolic compounds (chlorogenic acid and rutin) was greater in leaves (0.14% and 1.09%) and lowest in unripe fruits (0.03% and not detectable). In contrast, the unripe fruits presented the highest saponin content (12.30%), followed by the leaves (4.14%), and the branches (0.94%). In a mate commercial beverage the high amounts of phenolic compounds (3.82% and 0.51%) and methylxanthines (0.79% and 0.30%) were observed.

VALIDATION OF A LC METHOD FOR POLYPHENOLS ASSAY IN CAT'S CLAW (UNCARIA TOMENTOSA)

A reversed-phase LC method was developed and validated for the separation and assay of the main polyphenols in extracts from barks of Uncaria tomentosa. The LC method consists of a RP-18 column, in gradient mode (trifluoroacetic acid-methanol) using chlorogenic acid, caffeic acid, and rutin as externals standards and UV-detection at 325 nm. The method showed a good specificity, linearity, precision, and accuracy for standards compounds and for the five major peaks from bark extract. Calibration curves were linear with determination coefficients higher than 0.99. The repeatability and intermediary precision for the five major peaks ranged from 1.09 to 5.60% and 1.25 to 6.28%, respectively. The accuracy values for chlorogenic acid, caffeic acid, and rutin in the bark extract were 97.17, 98.84, and 101.78%, respectively. The LC method was applied successfully to one commercial spray-dried and four different freeze-dried extracts produced from barks and roots of U. tomentosa.

Development and Validation of an HPLC Method for the Characterization and Assay of the Saponins from Ilex paraguariensis A. St.‐Hil (Mate) Fruits

Abstract Two improved HPLC methods for the characterization and assay of the saponin content in green fruits of Ilex paraguariensis (erva‐mate) are described. Both HPLC methods consisted of isocratic separation at room temperature, using a RP‐18 column with UV‐Vis detection. The first system was intended for the characterization of the main I. paraguariensis saponin fraction, while the second one was specifically developed and validated for its quantitation. Matesaponin‐3 was used as external standard. From the freeze dried extract, ten peaks were characterized as saponins by TLC and HPLC analysis. The matesaponin‐3 content was 5.13 g%, and the total saponin content, calculated by the sum of the areas related to the peaks previously characterized as saponins, was 30.48 g%.

LC-UV assay method and UPLC/Q-TOF-MS characterisation of saponins from Ilex paraguariensis A. St. Hil. (mate) unripe fruits.

INTRODUCTION Ilex paraguariensis A. St. Hil. (mate) is known in several South American countries because of the use of its leaves in stimulant herbal beverages. High saponin contents were reported in mate leaves and unripe fruits that possess a dissimilar composition. Two LC-UV methods previously reported for mate saponins assay focused on mate leaves and the quantification of the less polar saponin fraction in mate fruits. OBJECTIVE To develop and validate a LC-UV method to assay the total content of saponins in unripe mate fruits and characterise the chemical structure of triterpenic saponins by UPLC/Q-TOF-MS. METHODOLOGY From unripe fruits of mate a crude ethanolic extract was prepared (EX40) and the mate saponin fraction (MSF) purified by solid phase extraction. The LC-UV method was validated using ilexoside II as external standard. UPLC/Q-TOF-MS was adjusted from the LC-UV method to obtain the fragmentation patterns of the main saponins present in unripe fruits. RESULTS Both LC-UV and UPLC/Q-TOF-MS methods indicate a wide range of Ilex saponins polarity. The ilexoside II and total saponin content of EX40 were 8.20% (w/w) and 47.60% (w/w), respectively. The total saponin content in unripe fruits was 7.28% (w/w). The saponins present in MSF characterised by UPLC/Q-TOF-MS are derived mainly from ursolic/oleanolic, acetyl ursolic or pomolic acid. CONCLUSION The validated LC-UV method was shown to be linear, precise, accurate and to cover several saponins previously isolated from Ilex species and could be applied for the quality control of unripe fruit saponins.

Cat's claw oxindole alkaloid isomerization induced by common extraction methods

Cat’s claw oxindole alkaloids are prone to isomerization in aqueous solution. However, studies on their behavior in extraction processes are scarce. This paper addressed the issue by considering five commonly used extraction processes. Unlike dynamic maceration (DM) and ultrasound-assisted extraction, substantial isomerization was induced by static maceration, turbo-extraction and reflux extraction. After heating under reflux in DM, the kinetic order of isomerization was established and equations were fitted successfully using a four-parameter Weibull model (R2 > 0.999). Different isomerization rates and equilibrium constants were verified, revealing a possible matrix effect on alkaloid isomerization.

Study of the specificity of cross-povidone (pvpp) as binding agent in the quantification of polyphenolic compounds

O polimero polivinilpirrolidona (PVPP) proporciona uma alternativa analitica ao inves do po-de-pele e caseina, como um agente complexante na quantificacao de taninos. Neste trabalho foi estudado a especificidade do PVPP em complexar com compostos polifenolicos, na presenca de rutina. Os acidos galico e tânico, catequina e pirogalol foram utilizados como substâncias de referencia (PRS), junto com o extrato aquoso das folhas do Psidium guajava L. A especificidade da complexacao foi avaliada pelo metodo espectrofotometrico e por cromatografia liquida de alta eficiencia (CLAE) com arranjo de diodos (DAD). As analises para a mistura das PRS e rutina, demonstraram claramente que a complexacao com PVPP nao e uma reacao especifica e independe da quantidade de polimero utilizada. As analises por CLAE-DAD do extrato de Psidium guajava revelaram picos caracteristicos de flavonoides, alem de catequina e acido galico. Todos os picos destes flavonoides, catequina e acido galico decresceram quando adicionou-se o PVPP. Isto confirma a falta de especificidade. Desde que a ligacao PVPP-polifenois depende de caracteristicas estruturais particulares, a extensao desses resultados a outras especies vegetais deve ser evitada. Cross-povidone, or polyvinylpyrrolidone (PVPP) affords an analytical alternative instead of hide-powder and casein as binding agent in the content assay of vegetable tannins. In this work we studied the specificity of PVPP to bind polyphenolics in the presence of flavonoids. Gallic acid, tannic acid, catechin and pyrogallol were used as polyphenolic reference substances (PRS), along with an aqueous extract from Psidium guajava L. leaves. The binding specificity was assayed by UV-Vis and High Performance Liquid Chromatography-Photodiode array (HPLC-PDA) methods. The analyses of PRS-rutin mixtures showed clearly that PVPP binding is a non-specific reaction, unrelated to the amount of PVPP used. HPLC-PDA analysis revealed peaks which could be characterized as flavonoids in the Psidium guajava extract, beside catechin and gallic acid. All these flavonoids, catechin and gallic acid peaks decreased as PVPP was added; this confirms the lack of specificity. Since the polyphenolic-PVPP reaction depends on specific structural features, any extensive conclusion should be avoided.

Validation of an LC Method for Polyphenol Assay in Extractive Solutions from Ilex paraguariensis (Mate)

Abstract A liquid chromatography (LC) method was developed and validated for identification and quantification of polyphenols in aqueous extractive solution from Ilex paraguariensis (erva‐mate), in agreement with the ICH requirements for analytical methods. The analysis was performed using a RP 18 column, in gradient solvent. Chlorogenic acid (CLOA) and rutin (RU) were used as external standards. The standard curves for CLOA and RU were linear with correlation coefficients higher than 0.9980. The LC method showed excellent performance in separating seven peaks. The method showed excellent repeatability (R.S.D.<2.0%) and accuracy (CLOA=97.4 and RU=104.0%). Besides CLOA and RU, six other constituents were detected, which were identified by LC‐MS/MS based on their mass spectra in full scan mode and retention times compared with those available in the literature data. The analytical method was successfully applied for quantifying polyphenols in four different extractive solutions from erva‐mate, a decoction (ES), an infusion (ESI), and an extractive solution obtained by turbo extraction with water (TE1) or ethanol 40% (v/v) (TE2). The last one presented the highest polyphenol concentration.