Multivariate calibrations in gas chromatographic profiles for prediction of several physicochemical parameters of Brazilian commercial gasoline

Several Brazilian commercial gasoline physicochemical parameters, such as relative density, distillation curve (temperatures related to 10%, 50% and 90% of distilled volume, final boiling point and residue), octane numbers (motor and research octane number and anti-knock index), hydrocarbon compositions (olefins, aromatics and saturates) and anhydrous ethanol and benzene content was predicted from chromatographic profiles obtained by flame ionization detection (GC-FID) and using partial least square regression (PLS). GC-FID is a technique intensively used for fuel quality control due to its convenience, speed, accuracy and simplicity and its profiles are much easier to interpret and understand than results produced by other techniques. Another advantage is that it permits association with multivariate methods of analysis, such as PLS. The chromatogram profiles were recorded and used to deploy PLS models for each property. The standard error of prediction (SEP) has been the main parameter considered to select the "best model". Most of GC-FID-PLS results, when compared to those obtained by the Brazilian Government Petroleum, Natural Gas and Biofuels Agency - ANP Regulation 309 specification methods, were very good. In general, all PLS models developed in these work provide unbiased predictions with lows standard error of prediction and percentage average relative error (below 11.5 and 5.0, respectively). (C) 2007 Elsevier B.V. All rights reserved.

Screening Brazilian C gasoline quality: Application of the SIMCA chemometric method to gas chromatographic data

A total of 2400 samples of commercial Brazilian C gasoline were collected over a 6-month period from different gas stations in the São Paulo state, Brazil, and analysed with respect to 12 physicochemical parameters according to regulation 309 of the Brazilian Government Petroleum, Natural Gas and Biofuels Agency (ANP). The percentages (v/v) of hydrocarbons (olefins, aromatics and saturated) were also determined. Hierarchical cluster analysis (HCA) was employed to select 150 representative samples that exhibited least similarity on the basis of their physicochemical parameters and hydrocarbon compositions. The chromatographic profiles of the selected samples were measured by gas chromatography with flame ionisation detection and analysed using soft independent modelling of class analogy (SIMCA) method in order to create a classification scheme to identify conform gasolines according to ANP 309 regulation. Following the optimisation of the SIMCA algorithm, it was possible to classify correctly 96% of the commercial gasoline samples present in the training set of 100. In order to check the quality of the model, an external group of 50 gasoline samples (the prediction set) were analysed and the developed SIMCA model classified 94% of these correctly. The developed chemometric method is recommended for screening commercial gasoline quality and detection of potential adulteration. (c) 2007 Elsevier B.V. All rights reserved.

Electrophoretic and chromatographic profile for S-like hemoglobin

Hemoglobin variants originate mainly by simple amino acid substitutions, the result of nucleotide sequence changes. Recently, the number of known abnormal hemoglobins has increased due to improvement in analysis methodologies; however, many laboratories are not prepared to correctly identify mutants. Hb S is a very well-characterized hemoglobin variant that varies in prevalence in different regions of Brazil. However, there is a type of Hb that presents electrophoretic migration in alkaline pH similar to Hb S, named S-like Hb, which can be incorrectly diagnosed; therefore, its frequency is underestimated. We obtained reference ranges for Hb S by HPLC, and we examined the electrophoretic and chromatographic profiles of S-like Hb. Hb Hasharon, Hb D-Los Angeles, Hb Korle-Bu, Hb Lepore, Hb D-Iran, Hb G-like, Hb Queens, Hb Montgomery, and Hb Q-India were found. Cases of association between two beta chain mutants were also found. Electrophoresis in alkaline and acid pH was utilized to initially screen these Hb variants, and globin chain electrophoresis at both high and low pH was performed to identify the globin chain mutant. Chromatographic analysis permitted the identification of the hemoglobin variant and also facilitated the quantification of these variants. Therefore, an association of classical laboratory diagnostic methodologies is fundamental for the correct identification of suspect Hb variants. The S and S-like hemoglobin profiles determined in this study will help in the diagnosis of these variants in health care services.

Produção in vitro, análise fitoquímica e avaliação da atividade antineoplásica de metabólitos de Hovenia dulcis Thunb. (Rhamnaceae) obtidos por diferentes estratégias biotecnológicas

Hovenia dulcis Thunberg, natural da Ásia Oriental, é cultivada no Brasil onde é conhecida como uva-do-japão. A espécie possui várias indicações na medicina popular e alguns estudos apontam o seu potencial antineoplásico, tripanocida e hepatoprotetor. Metabólitos secundários são substâncias não essenciais para a sobrevivência celular, mas que fornecem vantagens adaptativas aos vegetais, sendo atribuído, para algumas delas, atividades biológicas importantes. Substâncias de interesse medicinal têm sido obtidas por técnicas da cultura de tecidos vegetais, como a calogênese e a cultura de células em suspensão, que permitem a síntese de matéria-prima de forma contínua e homogênea, independentemente de fatores ambientais e sazonais. O presente estudo objetivou o estabelecimento de culturas in vitro de H. dulcis, visando à produção de metabólitos de interesse, com vistas à avaliação do seu potencial antineoplásico sobre células K562. Foram testados protocolos para o estabelecimento de diferentes sistemas, como culturas de calos, de células em suspensão (CCS) e compact callus clusters (CCC) e ainda a avaliação do uso de elicitores na otimização de metabólitos produzidos in vitro. Foi verificado que a adição dos fitorreguladores KIN e TDZ, substituindo o BAP, não foi capaz de induzir a formação de calos friáveis, bem como a manutenção das culturas em ausência de luz. O uso do nitrato de prata promoveu a friabilidade de calos em todas as concentrações testadas, considerando-se 2,0 mg.L-1 a melhor concentração. Foram alcançadas taxas de 100% de formação de CCS tanto na presença, quanto em ausência de AgNO3. O maior acúmulo de biomassa foi verificado na concentração mais baixa de PIC (0,625 mg.L-1). A análise dos espectros de RMN indicou a presença de (+)-dihidromiricetina, (+)-galocatequina, hovenitina II, hovenosideo G, hodulosideo III, hodulosideo IV, hodulosideo I e hovenidulciosideo B1 nas culturas de calos friáveis. No estabelecimento de culturas CCC, observou-se a formação de calos compactos verdes em todas as concentrações de ANA testadas. O aumento da velocidade de rotação para 135 rpm aumentou a dispersão das células com consequente formação dos agregados celulares desejados. A seleção de linhagens celulares demonstrou ser um método eficiente na uniformização do tamanho desses agregados e tal uniformidade se manteve estável por mais de cinco subcultivos em 100% das culturas. Uma fração rica em saponinas foi obtida a partir dos agregados celulares, correspondendo a 1,46% da massa seca. A análise por RMN sugeriu a presença das saponinas Hovenosideo G e dos hovenidulciosideos A2 e B2. O uso de elicitores em cultura de calos mostrou-se adequado à produção de metabólitos secundários, sem alterações morfológicas nos mesmos. A elicitação alterou o perfil cromatográfico analisado por HPLC. Na elicitação com 5,0 mg.L-1 de extrato de levedura foi verificado um aumento de quase três vezes (12,280 3,396 equivalentes de quercetina/mg de extrato) na síntese de flavonoides. Finalmente, os estudos de ação antitumoral in vitro demonstraram citotoxicidade dos extratos de calos não elicitados de H. dulcis sobre linhagem de leucemia mieloide crônica (IC50 de 74,05 μg.mL-1.) e inibição do crescimento de tais células (K562), sugerindo o potencial antineoplásico para um produto biotecnológio (calo) desta espécie.

Product monitoring and quantitation of oligosaccharides composition in agar hydrolysates by precolumn labeling HPLC

Since the discovery of multiple bioactivities for agarobiose oligomers, a quantitative method has been in great need to monitor the agarobiose oligomers. This report demonstrates that agarobiose oligomers can be separated with high resolution in HPLC after introducing a-naphthylamine into compounds. Agarobiose oligomers ranged from biose to decaose were isolated by Sephadex column. HPLC analysis indicated that each oliomer could be quantified with good linearity and a low detection limit of 0.1-4 mug/ml. The chromatographic profiles of agaro-oligosaccharides with different hydrolysis modes (hydrochloride, citric acid, solid acid, and hydroxyl radical degradation) showed that agarobiose could be obtained more than 57.8% using solid acid mediated hydrolysis, while hydrochloride acid could degrade agar into a series of agaro-oligosaccharides from biose to decaose. The yield of oligosaccharides was low if hydrolyzed by citric acid. The Fenton degradation can increase the speed of hydrolysis, but the product was complex. (C) 2004 Elsevier B.V. All rights reserved.

In vitro evaluation of Cu and Fe bioavailability in cashew nuts by off-line coupled SEC-UV and SIMAAS

In this work Cu and Fe bioavailability in cashew nuts was evaluated using in vitro method. Extractions with simulated gastric and intestinal fluids and dialysis procedures were applied for this purpose. The proteins separation and quantification were performed by size exclusion chromatography (SEC) coupled on-line to ultra-violet (UV) and off-line to simultaneous multielement atomic absorption spectrometry (SIMAAS). The SEC-UV and SIMAAS profiles of the protein fractions obtained by alkaline extraction (NaOH) and precipitation with HCl indicated the presence of high and low molecular weight species in the range between >75 kDa and 9.3 kDa. Almost 83% of Cu and 78% of Fe were extracted during cashew nut digestion and 90% of both elements were dialyzed. With these results it is possible to assume that 75% of Cu and 70% of Fe present in cashew nut could be bioavailable. The SEC-UV and SIMAAS chromatographic profiles obtained after in vitro gastrointestinal digestion reveal that Cu and Fe not dialyzed can be associated to a compound of 9.2 kDa. (C) 2010 Elsevier B.V. All rights reserved.

Improving peak shapes with counter gradients in two-dimensional high performance liquid chromatography

To achieve the greatest peak capacity in two-dimensional high performance liquid chromatography (2D-HPLC) a gradient should be operated in both separation dimensions. However, it is known that when an injection solvent that is stronger than the initial mobile phase composition is deleterious to peak performance, thus causing problems when cutting a portion from one gradient into another. This was overcome when coupling hydrophilic interaction with reversed phase chromatography by introducing a counter gradient that changed the solvent strength of the second dimension injection. It was found that an injection solvent composition of 20% acetonitrile in water gave acceptable results in one-dimensional simulations with an initial composition of 5% acetonitrile. When this was transferred to a 2D-HPLC separation of standards it was found that a marked improvement in peak shape was gained for the moderately retained analytes (phenol and dimethyl phthalate), some improvement for the weakly retained caffeine and very little change for the strongly retained n-propylbenzene and anthracene which already displayed good chromatographic profiles. This effect was transferred when applied to a 2D-HPLC separation of a coffee extract where the indecipherable retention profile was transformed to a successful application multidimensional chromatography with peaks occupying 71% of the separation space according to the geometric approach to factor analysis.

New Nanostructured Silica Adjuvant (SBA-15) Employed to Produce Antivenom in Young Sheep Using Crotalus durissus terrificus and Apis mellifera Venoms Detoxified by Cobalt-60

Equine antivenom is considered the only treatment for animal-generated envenomations, but it is costly. The study aimed to produce Apis mellifera (Africanized honeybee) and Crotalus durissus terrificus (C.d.t.) antivenoms using nanostructured silica (SBA-15) as adjuvant and cobalt-60 (60Co)-detoxified venoms utilizing young sheep. Natural and 60Co-irradiated venoms were employed in four different hyperimmunization protocols. Thus, 8 groups of 60- to 90-d-old sheep were hyperimmunized, enzyme-linked immunosorbent assay (ELISA) serum titers collected every 14 d were assessed clinically daily, and individual weight were measured, until d 84. Incomplete Freund's (IFA) and nanostructured silica (SBA15) adjuvants were compared. The lethal dose (LD50) for both venoms was determined following intraperitoneal (ip) administration to mice. High-performance liquid chromatography on reversed phase (HPLC-RP) was used also to measure the 60Co irradiation effects on Apis venom. At the end of the study, sheep were killed in a slaughterhouse. Kidneys were histologically analyzed. LD50 was 5.97 mg/kg Apis and 0.07 mg/kg C.d.t. for native compared to 13.44 mg/kg Apis and 0.35 mg/kg C.d.t. for irradiated venoms. HPLC revealed significant differences in chromatographic profiles between native and irradiated Apis venoms. Native venom plus IFA compared with SBA-15 showed significantly higher antibody titers for both venoms. Apis-irradiated venom plus IFA or SBA-15 displayed similar antibody titers but were significantly lower when compared with native venom plus IFA. Weight gain did not differ significantly among all groups. 60Co irradiation decreased toxicity and maintained venom immunogenic capacity, while IFA produced higher antibody titers. SBA-15 was able to act as an adjuvant without producing adverse effects. Hyperimmunization did not affect sheep weight gain, which would considerably reduce the cost of antiserum production, as these sheep were still approved for human consumption even after being subjected to hyperimmunization.

Intraspecific variability of flavonoid glycosides and styrylpyrones from leaves of Cryptocarya mandioccana Meisner (Lauraceae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Development and validation of a HPLC method for standardization of herbal and commercial extracts of Myrcia uniflora

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Influence of temperature on the properties of the xylanolytic enzymes of the thermotolerant fungus Aspergillus phoenicis

This study reports on the effects of growth temperature on the secretion and some properties of the xylanase and beta-xylosidase activities produced by a thermotolerant Aspergillus phoenicis. Marked differences were observed when the organism was grown on xylan-supplemented medium at 25 degreesC or 42 degreesC. Production of xylanolytic enzymes reached maximum levels after 72 h of growth at 42 degreesC; and levels were three- to five-fold higher than at 25 degreesC. Secretion of xylanase and beta-xylosidase was also strongly stimulated at the higher temperature. The optimal temperature was 85 degreesC for extracellular and 90 degreesC for intracellular beta-xylosidase activity, independent of the growth temperature. The optimum temperature for extracellular xylanase increased from 50 degreesC to 55 degreesC when the fungus was cultivated at 42 degreesC. At the higher temperature, the xylanolytic enzymes produced by A. phoenicis showed increased thermo stability, with changes in the profiles of pH optima. The chromatographic profiles were distinct when samples obtained from cultures grown at different temperatures were eluted from DEAE-cellulose and Biogel P-60 columns.

High-performance gel filtration patterns of venoms from three species of wasps of the genus Polybia (hymenoptera, vespidae).

The profiles of high-performance gel filtration of venoms from Polybia paulista, Polybia ignobilis and Polybia occidentalis occidentalis showed 13 peaks distributed among the three species. These profiles presented similarities that permitted the chromatographic characterization of the genus Polybia and differences that permitted the identification of each species studied. Thus, the comparative analysis of chromatographic profiles of high-performance gel filtration of venoms may be used as an auxiliary tool in taxonomic studies of Polybia wasps.

Assessment of the chemical profile, polyphenol content and antioxidant activity in extracts of Psidium Guajava L. fruits

Fruits and vegetables that are rich in polyphenolic compounds, especially flavonoids, may be used to benefit human health by reducing the incidence of cancers and cardiovascular diseases. Previous studies have demonstrated the antioxidant activity of guava, a fruit widely available in Brazil, possibly due to the presence of these polyphenolic compounds. The aim of this study was to analyze the total phenolic and flavonoid contents of various guava extracts, assay their antioxidant activity and record the chromatographic profiles of these extracts, to determine a simple and low way of extracting these compounds efficiently from guava. The results confirmed the presence of polyphenols in guava, including flavonoids, and its antioxidant activity. Furthermore, it was demonstrated that the 70% ethanol (by volume) was the most effective solvent to extract these compounds from the fruit, among those tested.

Determination of disease biomarkers in Eucalyptus by comprehensive two-dimensional gas chromatography and multivariate data analysis

In this paper is reported the use of the chromatographic profiles of volatiles to determine disease markers in plants - in this case, leaves of Eucalyptus globulus contaminated by the necrotroph fungus Teratosphaeria nubilosa. The volatile fraction was isolated by headspace solid phase microextraction (HS-SPME) and analyzed by comprehensive two-dimensional gas chromatography-fast quadrupole mass spectrometry (GC. ×. GC-qMS). For the correlation between the metabolic profile described by the chromatograms and the presence of the infection, unfolded-partial least squares discriminant analysis (U-PLS-DA) with orthogonal signal correction (OSC) were employed. The proposed method was checked to be independent of factors such as the age of the harvested plants. The manipulation of the mathematical model obtained also resulted in graphic representations similar to real chromatograms, which allowed the tentative identification of more than 40 compounds potentially useful as disease biomarkers for this plant/pathogen pair. The proposed methodology can be considered as highly reliable, since the diagnosis is based on the whole chromatographic profile rather than in the detection of a single analyte. © 2013 Elsevier B.V..

Desenvolvimento, aperfeiçoamento e validação de método cromatográfico para previsão da qualidade e propriedades físico-químicas do óleo diesel tipo B

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)