Integrative open workflow for confident annotation and molecular networking of metabolomics MSE/DIA data.

Liquid chromatography coupled with high-resolution mass spectrometry data-independent acquisition (LC-HRMS/DIA), including MSE, enable comprehensive metabolomics analyses though they pose challenges for data processing with automatic annotation and molecular networking (MN) implementation. This motivated the present proposal, in which we introduce DIA-IntOpenStream, a new integrated workflow combining open-source software to streamline MSE data handling. It provides 'in-house' custom database construction, allows the conversion of raw MSE data to a universal format (.mzML) and leverages open software (MZmine 3 and MS-DIAL) all advantages for confident annotation and effective MN data interpretation. This pipeline significantly enhances the accessibility, reliability and reproducibility of complex MSE/DIA studies, overcoming previous limitations of proprietary software and non-universal MS data formats that restricted integrative analysis. We demonstrate the utility of DIA-IntOpenStream with two independent datasets: dataset 1 consists of new data from 60 plant extracts from the Ocotea genus; dataset 2 is a publicly available actinobacterial extract spiked with authentic standard for detailed comparative analysis with existing methods. This user-friendly pipeline enables broader adoption of cutting-edge MS tools and provides value to the scientific community. Overall, it holds promise for speeding up metabolite discoveries toward a more collaborative and open environment for research.

Younger Researchers of the Brazilian Chemical Society: a network fostering representation, communication, and collaboration.

The Younger Researchers of the Brazilian Chemical Society committee supports early career researchers promoting communication, collaboration, education, networking, representation, and career development.

Effect of Extracts, Fractions, and Isolated Molecules of Casearia sylvestris to Control Streptococcus mutans Cariogenic Biofilm.

The effects of extracts, fractions, and molecules of Casearia sylvestris to control the cariogenic biofilm of Streptococcus mutans were evaluated. First, the antimicrobial and antibiofilm (initial and pre-formed biofilms) in prolonged exposure (24 h) models were investigated. Second, formulations (with and without fluoride) were assessed for topical effects (brief exposure) on biofilms. Third, selected treatments were evaluated via bacterium growth inhibition curves associated with gene expression and scanning electron microscopy. In initial biofilms, the ethyl acetate (AcOEt) and ethanolic (EtOH) fractions from Brasília (BRA/DF; 250 µg/mL) and Presidente Venceslau/SP (Water/EtOH 60:40 and Water/EtOH 40:60; 500 µg/mL) reduced ≥6-logs vs. vehicle. Only the molecule Caseargrewiin F (CsF; 125 µg/mL) reduced the viable cell count of pre-formed biofilms (5 logs vs. vehicle). For topical effects, no formulation affected biofilm components. For the growth inhibition assay, CsF yielded a constant recovery of surviving cells (≅3.5 logs) until 24 h (i.e., bacteriostatic), and AcOEt_BRA/DF caused progressive cell death, without cells at 24 h (i.e., bactericidal). CsF and AcOEt_BRA/DF damaged S. mutans cells and influenced the expression of virulence genes. Thus, an effect against biofilms occurred after prolonged exposure due to the bacteriostatic and/or bactericidal capacity of a fraction and a molecule from C. sylvestris.

Untargeted Metabolomics Sheds Light on the Diversity of Major Classes of Secondary Metabolites in the Malpighiaceae Botanical Family.

Natural products produced by plants are one of the most investigated natural sources, which substantially contributed to the development of the natural products field. Even though these compounds are widely explored, the literature still lacks comprehensive investigations aiming to explore the evolution of secondary metabolites produced by plants, especially if classical methodologies are employed. The development of sensitive hyphenated techniques and computational tools for data processing has enabled the study of large datasets, being valuable assets for chemosystematic studies. Here, we describe a strategy for chemotaxonomic investigations using the Malpighiaceae botanical family as a model. Our workflow was based on MS/MS untargeted metabolomics, spectral searches, and recently described in silico classification tools, which were mapped into the latest molecular phylogeny accepted for this family. The metabolomic analysis revealed that different ionization modes and extraction protocols significantly impacted the chemical profiles, influencing the chemotaxonomic results. Spectral searches within public databases revealed several clades or genera-specific molecular families, being potential chemical markers for these taxa, while the in silico classification tools were able to expand the Malpighiaceae chemical space. The classes putatively annotated were used for ancestral character reconstructions, which recovered several classes of metabolites as homoplasies (i.e., non-exclusive) or synapomorphies (i.e., exclusive) for all sampled clades and genera. Our workflow combines several approaches to perform a comprehensive evolutionary chemical study. We expect it to be used on further chemotaxonomic investigations to expand chemical knowledge and reveal biological insights for compounds classes in different biological groups.

Systematic Approach to Identify Novel Antimicrobial and Antibiofilm Molecules from Plants' Extracts and Fractions to Prevent Dental Caries.

Natural products provide structurally different substances, with a myriad of biological activities. However, the identification and isolation of active compounds from plants are challenging because of the complex plant matrix and time-consuming isolation and identification procedures. Therefore, a stepwise approach for screening natural compounds from plants, including the isolation and identification of potentially active molecules, is presented. It includes the collection of the plant material; preparation and fractionation of crude extracts; chromatography and spectrometry (UHPLC-DAD-HRMS and NMR) approaches for analysis and compounds identification; bioassays (antimicrobial and antibiofilm activities; bacterial "adhesion strength" to the salivary pellicle and initial glucan matrix treated with selected treatments); and data analysis. The model is simple, reproducible, and allows high-throughput screening of multiple compounds, concentrations, and treatment steps can be consistently controlled. The data obtained provide the foundation for future studies, including formulations with the most active extracts and/or fractions, isolation of molecules, modeling molecules to specific targets in microbial cells and biofilms. For example, one target to control cariogenic biofilm is to inhibit the activity of Streptococcus mutans glucosyltransferases that synthesize the extracellular matrix' glucans. The inhibition of those enzymes prevents the biofilm build-up, decreasing its virulence.

Metabolomics to Characterize Adaptive and Signaling Responses in Legume Crops under Abiotic Stresses.

Legume species are an important source of protein and other nutrients for human and livestock consumption, playing a central role in food security. Besides, legumes benefit agriculture because of their ability to establish symbiotic interactions with nitrogen-fixing bacteria, providing nitrogen for subsequent crops, which is very much appreciated for sustainable agricultural practices. However, like other food crops, legumes are highly vulnerable to climate variations, water stresses being the main constraint that negatively affects both crop quality and productivity. Because of this, the development of strategies to improve the tolerance of such cultivars against water stresses, as well as the study of effective approaches to monitor these improvements, have gained special attention during the last years. Among these strategies, metabolomics has been considered one of the most promising approaches for the detection and/or quantification of primary and secondary stress-responsive metabolites in abiotic stresses. In plant science, many research groups have been using metabolomics to evaluate the success of genetic modifications by the analysis of chemical markers that can be altered in breeding programs. In addition, metabolomics is a powerful tool for the evaluation and selection of wild specimens with desirable traits that can be used in the development of improved new cultivars. Therefore, the aim of the present paper is to review the recent progress made in the field of metabolomics and plant breeding, especially concerning the adaptive responses of legume species to abiotic stresses as well as to point out the key primary and secondary metabolites involved in the adaptation and sensing mechanisms.

Antimicrobial and antibiofilm activities of Casearia sylvestris extracts from distinct Brazilian biomes against Streptococcus mutans and Candida albicans.

BACKGROUND: Dental caries is a biofilm-diet-dependent worldwide public health problem, and approaches against microorganisms in cariogenic biofilms are necessary. METHODS: The antimicrobial and antibiofilm activities of 12 Casearia sylvestris extracts (0.50 mg/mL) from different Brazilian biomes (Atlantic Forest, Cerrado, Caatinga, Pampa, and Pantanal) and varieties (sylvestris, lingua, and intermediate) were tested against two species found in cariogenic biofilms (Streptococcus mutans and Candida albicans). The extracts effective against S. mutans were used to evaluate the "adhesion strength" of this bacterium to the salivary pellicle and initial glucan matrix and the S. mutans-GtfB activity. Also, the antimicrobial activity against S. mutans of three fractions (methanol, ethyl acetate, and hexane; 0.25 mg/mL) from the extracts was evaluated. RESULTS: Three extracts from the Atlantic Forest variety sylvestris (FLO/SC, GUA/CE, PRE/SP) reduced ≥50% (> 3 logs) S. mutans viable population (p < 0.0001 vs. vehicle), while two extracts from the same biome and variety (PAC/CE, PRE/SP) decreased ≥50% of the viable counts of C. albicans (p < 0.0001 vs. vehicle). For S. mutans biofilms, three extracts (GUA/CE, PAC/CE, PRE/SP) reduced the biomass by ≥91% (p > 0.0001 vs. vehicle) and 100% of the microbial population (p < 0.0001 vs. vehicle). However, for the fungal biofilm, two extracts (PAC/CE, PRE/SP) reduced the viable counts by ≥52% (p < 0.0001 vs. vehicle), but none reduced biomass. The extracts with higher antimicrobial and antibiofilm activities presented higher content of clerodane-type diterpenes and lower content of glycosylated flavonoids than the less active extracts. The extracts had no effect on the removal of cells adhered to the pellicle (p > 0.05 vs. vehicle) while promoted the detachment of a larger number of S. mutans cells from GtfB-glucan matrix (p < 0.0031 vs. vehicle), and FLO/SC, GUA/CE and PRE/SP reduced the quantity of glucans (p < 0.0136 vs. vehicle). Only the ethyl acetate fractions reduced the microbial population of S. mutans (p < 0.0001 vs. vehicle), except for one (PAC/CE). Among the ethyl acetate fractions, three from var. lingua (two from Cerrado, and one from Cerrado/Caatinga) reduced ≥83% of the microbial population. CONCLUSIONS: C. sylvestris extracts from Atlantic Forest var. sylvestris and ethyl acetate fractions from Cerrado and Cerrado/Caatinga var. lingua may be used as a strategy against cariogenic microorganisms.

Metabolic Profiling of Saponin-Rich Ophiopogon japonicus Roots Based on 1H NMR and HPTLC Platforms.

Ideally, metabolomics should deal with all the metabolites that are found within cells and biological systems. The most common technologies for metabolomics include mass spectrometry, and in most cases, hyphenated to chromatographic separations (liquid chromatography- or gas chromatography-mass spectrometry) and nuclear magnetic resonance spectroscopy. However, limitations such as low sensitivity and highly congested spectra in nuclear magnetic resonance spectroscopy and relatively low signal reproducibility in mass spectrometry impede the progression of these techniques from being universal metabolomics tools. These disadvantages are more notorious in studies of certain plant secondary metabolites, such as saponins, which are difficult to analyse, but have a great biological importance in organisms. In this study, high-performance thin-layer chromatography was used as a supplementary tool for metabolomics. A method consisting of coupling 1H nuclear magnetic resonance spectroscopy and high-performance thin-layer chromatography was applied to distinguish between Ophiopogon japonicus roots that were collected from two growth locations and were of different ages. The results allowed the root samples from the two growth locations to be clearly distinguished. The difficulties encountered in the identification of the marker compounds by 1H nuclear magnetic resonance spectroscopy was overcome using high-performance thin-layer chromatography to separate and isolate the compounds. The saponins, ophiojaponin C or ophiopogonin D, were found to be marker metabolites in the root samples and proved to be greatly influenced by plant growth location, but barely by age variation. The procedure used in this study is fully described with the purpose of making a valuable contribution to the quality control of saponin-rich herbal drugs using high-performance thin-layer chromatography as a supplementary analytical tool for metabolomics research.

Hydroalcoholic crude extract of Casearia sylvestris Sw. reduces chronic post-ischemic pain by activation of pro-resolving pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Casearia sylvestris Sw. is widely used in popular medicine to treat conditions associated with pain. AIM OF THE STUDY: The present study investigated the influence of hydroalcoholic crude extract of Casearia sylvestris (HCE-CS) and contribution of pro-resolving mediators on mechanical hyperalgesia in a mouse model of chronic post-ischemia pain (CPIP). METHODS AND RESULTS: Male Swiss mice were subjected to ischemia of the right hind paw (3h), then reperfusion was allowed. At 10min, 24h or 48h post-ischemia/reperfusion (I/R), different groups of animals were treated with HCE-CS (30mg/Kg, orally [p.o]), selected agonists at the pro-resolving receptor ALX/FPR2 (natural molecules like resolvin D1 and lipoxin A4 or the synthetic compound BML-111; 0.1-1µg/animal) or vehicle (saline, 10mL/Kg, s.c.), in the absence or presence of the antagonist WRW4 (10µg, s.c.). Mechanical hyperalgesia (paw withdrawal to von Frey filament) was asseseed together with histological and immunostainning analyses. In these settings, pro-resolving mediators reduced mechanical hyperalgesia and HCE-CS or BML-111 displayed anti-hyperalgesic effects which was markedly attenuated in animals treated with WRW4. ALX/FPR2 expression was raised in skeletal muscle or neutrophils after treatment with HCE-CS or BML-111. CONCLUSION: These results reveal significant antihyperalgesic effect of HCE-CS on CPIP, mediated at least in part, by the pathway of resolution of inflammation centred on the axis modulated by ALX/FPR2.

A Highly Polar Phytocomplex Involving Rutin is Responsible for the Neuromuscular Facilitation Caused by Casearia sylvestris (guaçatonga).

BACKGROUND: Of the various biological activities ascribed to extracts from Casearia sylvestris (guaçatonga), its facilitatory activity, i.e., ability to increase skeletal muscle contractile amplitude, has promising therapeutic applications. In this work, we investigated the components responsible for the previously described neurofacilitation caused by C. sylvestris leaves. METHODS: The methanolic fraction of C. sylvestris leaves was initially fractionated by column chromatography and partitioned in a MeOH:H2O gradient. The resulting fractions were analyzed by analytical HPLC and yielded fraction 5:5 (F55) that was subjected to solid phase extraction and preparative HPLC. Of the seven resulting subfractions, only F55-6 caused muscle facilitation. Subfractions F55-6 and F55-7 (similar in composition to F55-6 by TLC analysis, but inactive) were analyzed by 1H-NMR to identify their constituents. RESULTS: This analysis identified a rutin-glycoside phytocomplex that caused neurofacilitation, a property that commercial rutin alone did not exhibit. CONCLUSION: F55-6 apparently caused neurofacilitation by the same mechanism (presynaptic action) as the methanolic fraction since its activity was also inhibited in tetrodotoxin-pretreated preparations.

Ecological strategies of Al-accumulating and non-accumulating functional groups from the cerrado sensu stricto.

The cerrado's flora comprises aluminum-(Al) accumulating and non-accumulating plants, which coexist on acidic and Al-rich soils with low fertility. Despite their existence, the ecological importance or biological strategies of these functional groups have been little explored. We evaluated the leaf flushing patterns of both groups throughout a year; leaf concentrations of N, P, K, Ca, Mg, S, Al, total flavonoids and polyphenols; as well as the specific leaf area (SLA) on young and mature leaves within and between the groups. In Al-accumulating plants, leaf flushed throughout the year, mainly in May and September; for non-accumulating plants, leaf flushing peaked at the dry-wet seasons transition. However, these behaviors could not be associated with strategies for building up concentrations of defense compounds in leaves of any functional groups. Al-accumulating plants showed low leaf nutrient concentrations, while non-accumulating plants accumulated more macronutrients and produced leaves with high SLA since the juvenile leaf phase. This demonstrates that the increase in SLA is slower in Al-accumulating plants that are likely to achieve SLA values comparable to the rest of the plant community only in the wet season, when sunlight capture is important for the growth of new branches.

Use of Chamomilla recutita in the Prevention and Treatment of Oral Mucositis in Patients Undergoing Hematopoietic Stem Cell Transplantation: A Randomized, Controlled, Phase II Clinical Trial.

BACKGROUND: Oral mucositis is a common inflammatory complication among patients undergoing hematopoietic stem cell transplantation (HSCT). Among its therapeutic properties, Chamomilla recutita has anti-inflammatory effects. OBJECTIVE: The aim of this study was to identify the dosage of the liquid extract of C recutita in mouthwash that is needed to reduce the incidence and intensity of oral mucositis in adult patients undergoing allogenic HSCT. METHODS: In a randomized phase II clinical trial, 40 patients were randomized to receive routine care plus mouthwash containing a liquid extract of C recutita at 0.5%, 1%, or 2% (experimental groups) or standard care alone (control group). Daily evaluation was performed using the measurement scale for oral toxicity defined by the World Health Organization. Statistical analysis was performed, in which the incidence, intensity, and duration of oral mucositis were compared between each experimental group and the control group. RESULTS: The experimental group at the 1% dosage demonstrated reduced incidence, intensity, and duration of oral mucositis compared with the control group. The formulation was well tolerated by patients and was safe, as no moderate or severe adverse effects were identified. CONCLUSIONS: In this study, the use of mouthwash containing 1% C recutita extract can be associated with reduced incidence, intensity, and duration of mucositis in adults patients undergoing allogenic HSCT. IMPLICATIONS FOR PRACTICE: The results of this investigation will help nurses and other professionals in selecting the C recutita dosage used to manage oral mucositis in patients undergoing HSCT.

Inhibition of the human neutrophil oxidative metabolism by Baccharis dracunculifolia DC (Asteraceae) is influenced by seasonality and the ratio of caffeic acid to other phenolic compounds.

ETHNOPHARMACOLOGICAL RELEVANCE: The great potential of phytotherapic drugs for treating and preventing inflammatory diseases mediated by increased neutrophil reactive oxygen species (ROS) generation has guided the search for new natural products with antioxidant and immunomodulatory properties. Baccharis dracunculifolia D.C. (Asteraceae), the main botanical source of Brazilian green propolis, is a native plant from Brazil widely used in folk medicine as anti-inflammatory. This study aims: (a) to determine the influence of seasonality on the chemical profile and biological activity of Baccharis dracunculifolia (Asteraceae) leaf extracts (BdE); (b) to analyze the correlation between the major compounds and the ability of BdE to modulate the superoxide anion and total ROS generation by human neutrophils. MATERIALS AND METHODS: The extracts were obtained from leaf samples collected monthly during one year. The superoxide anion and total ROS generation were assessed by the lucigenin (CL-luc)- and luminol (CL-lum)-enhanced chemiluminescence assays. RESULTS: Seasonality influenced more the quantitative than the qualitative chemical profile of B. dracunculifolia, and affected its biological activity. The major compounds identified were caffeic acid, p-coumaric acid, aromadendrin-4'-methyl ether (AME), isosakuranetin and artepillin C. The IC50 values obtained for CL-lum and CL-luc inhibition by BdE ranged from 8.1-15.8 and 5.8-13.3µgmL(-1), respectively, and correlated positively with caffeic acid concentration. CL-luc inhibition correlated negatively with the concentration of artepillin C, AME, isosakuranetin and total flavonoids. The BdE sample from May/07 inhibited CL-lum and CL-luc the most strongly (IC50=8.1 ± 1.6 and 5.8 ± 1.0 µg mL(-1), respectively), and contained the highest ratio of caffeic acid to the other isolated compounds; so, this ratio could be employed as chemical marker for this biological activity of B. dracunculifolia. CONCLUSION: The ability of B. dracunculifolia to inhibit the neutrophil ROS generation depends more on the type and ratio of phenolic compounds and flavonoids than on their high absolute concentrations. Together, our results help select the most appropriate plant material for the production of phytotherapic drugs to be used in the treatment of inflammatory diseases mediated by increased neutrophil activation.

Baccharis dracunculifolia, the main source of green propolis, exhibits potent antioxidant activity and prevents oxidative mitochondrial damage.

Baccharis dracunculifolia DC (Asteraceae) is the main botanical source used by honeybees to produce Brazilian green propolis whose hepatoprotective properties have been already described. In this work we investigated the protective effects of the glycolic extract of B. dracunculifolia (GEBd) against oxidative stress in isolated rat liver mitochondria (RLM). The GEBd was prepared by fractionated percolation using propylene glycol as solvent. The total phenols and flavonoids, which are substances with recognized antioxidant action, were quantified in GEBd and the phytochemical analysis was carried out by HPLC. GEBd exhibited significant scavenger activity towards DPPH radicals and superoxide anions in a concentration-dependent manner, and also a Fe2+ chelating activity. GEBd decreased the basal H2O2 generation and the Fe2+- or t-BuOOH-induced ROS production in isolated mitochondria. Lipid oxidation of mitochondrial membranes, protein thiol groups and GSH oxidation were also prevented by GEBd. This shows that B. dracunculifolia exhibit potent antioxidant activity protecting liver mitochondria against oxidative damage and such action probably contribute to the antioxidant and hepatoprotective effects of green propolis.

A validated capillary gas chromatography method for guaco (Mikania glomerata S) quality control and rastreability: from plant biomass to phytomedicines / Método validado por cromatografia gasosa capilar para o controle de qualidade e rastreabilidade de guaco (Mikania glomerata S): da matéria-prima ao fitoterápico

Este trabalho descreve a validação completa de metodologia analítica empregando cromatografia gasosa capilar com padronização interna para quantificação da cumarina (1,2-benzopirona) em produtos contendo guaco (Mikania glomerata Spreng - Asteraceae): xarope, planta e extrato padronizado, além do estudo de estabilidade do fitoterápico em questão. Utilizou-se uma coluna capilar HP-5 (30 m x 0,32 mm x 0,25 µm), hidrogênio a 1,8 mL/min e rampa de temperatura de 100 ºC a 250 ºC, a 15 ºC/min. A temperatura do injetor (split 1:20) foi de 250 ºC, enquanto a do detector foi de 270 ºC. Os tempos de retenção dos padrões foram: 2,86 minutos para o 1, 2, 3, 4-tetrametilbenzeno, 4,45 minutos para o piperonal (padrões internos) e 5,36 minutos para a cumarina. Após o procedimento de extração da planta in natura, a recuperação da cumarina foi de 101,6 por cento, enquanto que para o xarope esta foi de 100,8 por cento. Os limites de detecção e quantificação foram 0,5 µg/mL e 1,5 µg/mL, respectivamente. A precisão, determinada para todas as amostras, apresentou desvios padrões relativos menores que 2,5 por cento. Os teores de cumarina presentes nas folhas, extrato e xarope foram de 0,38 por cento m/m, 1,33 mg/mL e 0,143 mg/mL, respectivamente.

This work describes a full validation of a capillary gas chromatography analytical methodology using internal standardization for the quantification of coumarin (1,2-benzopyrone) in guaco (Mikania glomerata Spreng - Asteraceae) products: syrup, plant and its extract, including the stability study of the phytomedicine. For the analysis, it was used an HP-5 capillary column (30 m x 0.32 mm x 0.25 µm), hydrogen at a flow rate of 1.8 mL/min and the increasing temperature gradient was: 100 ºC to 250 ºC, 15 ºC/min. The temperature of injector (split 1:20) and detector were kept at 250 ºC and 270 ºC, respectively. The retention times of the standards for the above conditions were 2.86 minutes for 1, 2, 3, 4-tetramethylbenzene, 4.45 min for piperonal (internal standards), and 5.36 minutes for coumarin. After extraction procedure, the recovery of coumarin determined for plant raw material was 101.6 percent, while for syrup it was 100.8 percent. Detection and quantification limits were 0.5 µg/mL and 1.5 µg/mL, respectively. Precision was determined for all samples and the results were lower than 2.5 percent. The total amount of coumarin in plant raw material, its extract and syrup were 0.38 percent w/w, 1.33 mg/mL and 0.143 mg/mL, respectively.

A validated reverse-phase HPLC analytical method for the quantification of phenolic compounds in Baccharis dracunculifolia.

INTRODUCTION: Baccharis dracunculifolia, which has great potential for the development of new phytotherapeutic medicines, is the most important botanical source of the southeastern Brazilian propolis, known as green propolis on account of its color. OBJECTIVE: To develop a reliable reverse-phase HPLC chromatographic method for the analysis of phenolic compounds in both B. dracunculifolia raw material and its hydroalcoholic extracts. METHODOLOGY: The method utilised a C(18) CLC-ODS (M) (4.6 x 250 mm) column with nonlinear gradient elution and UV detection at 280 nm. A procedure for the extraction of phenolic compounds using aqueous ethanol 90%, with the addition of veratraldehyde as the internal standard, was developed allowing the quantification of 10 compounds: caffeic acid, coumaric acid, ferulic acid, cinnamic acid, aromadendrin-4'-methyl ether, isosakuranetin, drupanin, artepillin C, baccharin and 2,2-dimethyl-6-carboxyethenyl-2H-1-benzopyran acid. RESULTS: The developed method gave a good detection response with linearity in the range 20.83-800 microg/mL and recovery in the range 81.25-93.20%, allowing the quantification of the analysed standards. CONCLUSION: The method presented good results for the following parameters: selectivity, linearity, accuracy, precision, robustness, as well as limit of detection and limit of quantitation. Therefore, this method could be considered as an analytical tool for the quality control of B. dracunculifolia raw material and its products in both cosmetic and pharmaceutical companies.

A reliable quantitative method for the analysis of phenolic compounds in Brazilian propolis by reverse phase high performance liquid chromatography.

A sensitive and reliable RP-HPLC method was developed using a C18 CLC-ODS (M) - 4.6x250 mm(2)column and gradient elution for the analysis of phenolic compounds in propolis raw material and its products. A procedure for the extraction of phenolic compounds using aqueous ethanol (90%) with the addition of veratraldehyde as the internal standard (IS) was developed allowing to quantify ten compounds: caffeic acid, coumaric acid, ferulic acid, cinnamic acid, aromadendrin-4'-methyl ether (AME), isosakuranetin, drupanin, artepellin C, baccharin, and 2,2-dimethyl-6-carboxyethenyl-2H-1-benzopyran acid (DCBEN). The developed method gave good detection response and linearity in the range of 20.83-533.33 microg/mL.