Effect of an aqueous extract of Phaseolus vulgaris on the properties of tail tendon collagen of rats with streptozotocin-induced diabetes

Changes in the structural and functional properties of collagen caused by advanced glycation might be of importance for the etiology of late complications in diabetes. The present study was undertaken to investigate the influence of oral administration of aqueous pod extract (200 mg/kg body weight) of Phaseolus vulgaris, an indigenous plant used in Ayurvedic Medicine in India, on collagen content and characteristics in the tail tendon of streptozotocin-diabetic rats. In diabetic rats, collagen content (117.01 ± 6.84 mg/100 mg tissue) as well as its degree of cross-linking was increased, as shown by increased extent of glycation (21.70 ± 0.90 µg glucose/mg collagen), collagen-linked fluorescence (52.8 ± 3.0 AU/µmol hydroxyproline), shrinkage temperature (71.50 ± 2.50ºC) and decreased acid (1.878 ± 0.062 mg hydroxyproline/100 mg tissue) and pepsin solubility (1.77 ± 0.080 mg hydroxyproline/100 mg tissue). The alpha/ß ratio of acid- (1.69) and pepsin-soluble (2.00) collagen was significantly decreased in streptozotocin-diabetic rats. Administration of P. vulgaris for 45 days to streptozotocin-diabetic rats significantly reduced the accumulation and cross-linking of collagen. The effect of P. vulgaris was compared with that of glibenclamide, a reference drug administered to streptozotocin-diabetic rats at the dose of 600 µg/kg body weight for 45 days by gavage. The effects of P. vulgaris (collagen content, 64.18 ± 1.97; extent of glycation, 12.00 ± 0.53; collagen-linked fluorescence, 33.6 ± 1.9; shrinkage temperature, 57.0 ± 1.0; extent of cross-linking - acid-soluble collagen, 2.572 ± 0.080, and pepsin-soluble collagen, 2.28 ± 0.112) were comparable with those of glibenclamide (collagen content, 71.5 ± 2.04; extent of glycation, 13.00 ± 0.60; collagen-linked fluorescence, 38.9 ± 2.0; shrinkage temperature, 59.0 ± 1.5; extent of cross-linking - acid-soluble collagen, 2.463 ± 0.078, and pepsin-soluble collagen, 2.17 ± 0.104). In conclusion, administration of P. vulgaris pods had a positive influence on the content of collagen and its properties in streptozotocin-diabetic rats.

Immunomodulatory properties of Alternanthera tenella Colla aqueous extracts in mice

Plants from the genus Alternanthera are thought to possess antimicrobial and antiviral properties. In Brazilian folk medicine, the aqueous extract of A. tenella Colla is used for its anti-inflammatory activity. The present study investigated the immunomodulatory property of A. tenella extract by evaluating the antibody production in male albino Swiss mice weighing 20-25 g (10 per group). The animals received standard laboratory diet and water ad libitum. The effect of A. tenella extract (5 and 50 mg/kg, ip) was evaluated in mice immunized with sheep red blood cells (SRBC 10%, ip) as T-dependent antigen, or in mice stimulated with mitogens (10 µg, Escherichia coli lipopolysaccharide, LPS, ip). The same doses (5 and 50 mg/kg, ip) of A. tenella extract were also tested for antitumor activity, using the Ehrlich ascites carcinoma as model. The results showed that 50 mg/kg A. tenella extract ip significantly enhanced IgM (64%) and IgG2a (50%) antibody production in mice treated with LPS mitogen. The same dose had no effect on IgM-specific response, whereas the 5 mg/kg treatment caused a statiscally significant reduction of anti-SRBC IgM-specific antibodies (82%). The aqueous extract of A. tenella (50 mg/kg) increased the life span (from 16 ± 1 to 25 ± 1 days) and decreased the number of viable tumor cells (59%) in mice with Ehrlich ascites carcinoma. The present findings are significant for the development of alternative, inexpensive and perhaps even safer strategies for cancer treatment.

Modulation of B lymphocyte function by an aqueous fraction of the ethanol extract of Cissampelos sympodialis Eichl (Menispermaceae)

Cissampelos sympodialis Eichl species are used in folk medicine for the treatment of asthma, arthritis and rheumatism. In the present study, we investigated the immunomodulatory effect of an aqueous fraction of a 70% (v/v) ethanol extract of C. sympodialis leaves on B lymphocyte function. The hydroalcoholic extract inhibited the in vitro proliferative response of resting B cells induced by LPS (IC50 = 17.2 µg/ml), anti-delta-dextran (IC50 = 13.9 µg/ml) and anti-IgM (IC50 = 24.3 µg/ml) but did not affect the anti-MHC class II antibody-stimulated proliferative response of B cell blasts obtained by stimulation with IL-4 and anti-IgM. Incubation with the hydroalcoholic extract used at 50 µg/ml induced a 700% increase in intracellular cAMP levels. IgM secretion by resting B cells (obtained from normal mice) and polyclonally activated B cells (obtained from Trypanosoma cruzi-infected animals) was inhibited by the hydroalcoholic extract. The latter were more sensitive to the hydroalcoholic extract since 6.5 µg/ml induced a 20% inhibition in the response of cells from normal mice while it inhibited the response of B cells from infected animals by 75%. The present data indicate that the alcoholic extract of C. sympodialis inhibited B cell function through an increase in intracellular cAMP levels. The finding that the hydroalcoholic extract inhibited immunoglobulin secretion suggests a therapeutic use for the extract from C. sympodialis in conditions associated with unregulated B cell function and enhanced immunoglobulin secretion. Finally, the inhibitory effect of the hydroalcoholic extract on B cells may indicate an anti-inflammatory effect of this extract.

Effect of an aqueous extract of Scoparia dulcis on blood glucose, plasma insulin and some polyol pathway enzymes in experimental rat diabetes

The effects of an aqueous extract of the plant Scoparia dulcis (200 mg/kg) on the polyol pathway and lipid peroxidation were examined in the liver of streptozotocin adult diabetic male albino Wistar rats. The diabetic control rats (N = 6) presented a significant increase in blood glucose, sorbitol dehydrogenase, glycosylated hemoglobin and lipid peroxidation markers such as thiobarbituric acid reactive substances (TBARS) and hydroperoxides, and a significant decrease in plasma insulin and antioxidant enzymes such as glutathione peroxidase (GPx), glutathione-S-transferase (GST) and reduced glutathione (GSH) compared to normal rats (N = 6). Scoparia dulcis plant extract (SPEt, 200 mg kg-1 day-1) and glibenclamide (600 µg kg-1 day-1), a reference drug, were administered by gavage for 6 weeks to diabetic rats (N = 6 for each group) and significantly reduced blood glucose, sorbitol dehydrogenase, glycosylated hemoglobin, TBARS, and hydroperoxides, and significantly increased plasma insulin, GPx, GST and GSH activities in liver. The effect of the SPEt was compared with that of glibenclamide. The effect of the extract may have been due to the decreased influx of glucose into the polyol pathway leading to increased activities of antioxidant enzymes and plasma insulin and decreased activity of sorbitol dehydrogenase. These results indicate that the SPEt was effective in attenuating hyperglycemia in rats and their susceptibility to oxygen free radicals.

DNA extraction and quantification from touch and scrape preparations obtained from autopsy liver cells

The objective of the present study was to develop a simplified low cost method for the collection and fixation of pediatric autopsy cells and to determine the quantitative and qualitative adequacy of extracted DNA. Touch and scrape preparations of pediatric liver cells were obtained from 15 cadavers at autopsy and fixed in 95% ethanol or 3:1 methanol:acetic acid. Material prepared by each fixation procedure was submitted to DNA extraction with the Wizard® genomic DNA purification kit for DNA quantification and five of the preparations were amplified by multiplex PCR (azoospermia factor genes). The amount of DNA extracted varied from 20 to 8,640 µg, with significant differences between fixation methods. Scrape preparation fixed in 95% ethanol provided larger amount of extracted DNA. However, the mean for all groups was higher than the quantity needed for PCR (50 ng) or Southern blot (500 ng). There were no qualitative differences among the different material and fixatives. The same results were also obtained for glass slides stored at room temperature for 6, 12, 18 and 24 months. We conclude that touch and scrape preparations fixed in 95% ethanol are a good source of DNA and present fewer limitations than cell culture, tissue paraffin embedding or freezing that require sterile material, culture medium, laboratory equipment and trained technicians. In addition, they are more practical and less labor intensive and can be obtained and stored for a long time at low cost.

EXTRACTION OF OIL FROM PRESSED PALM OIL (Elaes guineensis) FIBERS USING SUPERCRITICAL CO2

Residual fibers from palm oil production are a good source of carotene, since they contain more than 5% of the original oil, with about 5000 ppm of carotenoids. As carotenoids are thermosensitive molecules, supercritical CO2 can be used for oil recovery, because this technique employs low temperatures. In this work results of oil extraction experiments from pressed palm oil fibers are shown. Fibers were from AGROPALMA, an industry which is located in Tailândia (Pará, Brazil). Extractions were carried out at 200, 250 and 300 bar and at temperatures of 45 and 55oC. Oil was analyzed by UV/vis spectrophotometry for total carotene determination. Results showed a large increase in extraction rate from 200 to 250 bar and a small variation from 250 to 300 bar. The total amount of carotenes did not increase in the course of extraction at 300 bar, but it showed a large increase at 200 and at 250 bar. Free fatty acids are present in amounts larger than those found in commercial oils.

Effect of blanching time on selective mineral elements extraction from the spinach substitute (Tetragonia expansa) commonly used in Brazil

The true spinach (Spinacia oleracea) does not grow well in warm climates and for that reason is not commercialized in Brazil. Instead, a spinach substitute (Tetragonia expansa), originally from New Zealand, is widely used in the country. There is scant information on the mineral profile and none on the soluble mineral fraction of this vegetable. The solubility of a mineral is one of the important factors for its absorption. For this reason, the calcium, magnesium, iron, manganese, copper, zinc, potassium, and sodium soluble fractions in the raw spinach substitute were determined and the effect of blanching times on the solubility of these minerals was investigated. Blanching times of 1, 5, and 15 minutes were employed. The magnesium, manganese, potassium, and sodium soluble fractions increased sizably with shorter blanching time. Longer blanching time (15 minutes) caused large losses of minerals. The soluble mineral fractions can contribute poorly to diet in terms of potassium, magnesium, manganese, and zinc. The spinach substitute cannot be considered a dietary source of calcium, iron and copper due to the insolubility of these minerals in the vegetable, possibly caused by the large oxalate content.

Head Space Solid Phase Micro-Extraction (HS - SPME) of volatile organic compounds produced by Sporidiobolus salmonicolor (CBS 2636)

The aim of the present study was the assessment of volatile organic compounds produced by Sporidiobolus salmonicolor (CBS 2636) using methyl and ethyl ricinoleate, ricinoleic acid and castor oil as precursors. The analysis of the volatile organic compounds was carried out using Head Space Solid Phase Micro-Extraction (HS - SPME). Factorial experimental design was used for investigating extraction conditions, verifying stirring rate (0-400 rpm), temperature (25-60 ºC), extraction time (10-30 minutes), and sample volume (2-3 mL). The identification of volatile organic compounds was carried out by Gas Chromatography with Mass Spectrum Detector (GC/MSD). The conditions that resulted in maximum extraction were: 60 ºC, 10 minutes extraction, no stirring, sample volume of 2.0 mL, and addition of saturated KCl (1:10 v/v). In the bio-production of volatile organic compounds the effect of stirring rate (120-200 rpm), temperature (23-33 ºC), pH (4.0-8.0), precursor concentration (0.02-0.1%), mannitol (0-6%), and asparagine concentration (0-0.2%) was investigated. The bio-production at 28 ºC, 160 rpm, pH 6,0 and with the addition of 0.02% ricinoleic acid to the medium yielded the highest production of VOCs, identified as 1,4-butanediol, 1,2,2-trimethylciclopropilamine, beta-ionone; 2,3-butanodione, pentanal, tetradecane, 2-isononenal, 4-octen-3-one, propanoic acid, and octadecane.

Use of real-time PCR to evaluate two DNA extraction methods from food

The DNA extraction is a critical step in Genetically Modified Organisms analysis based on real-time PCR. In this study, the CTAB and DNeasy methods provided good quality and quantity of DNA from the texturized soy protein, infant formula, and soy milk samples. Concerning the Certified Reference Material consisting of 5% Roundup Ready® soybean, neither method yielded DNA of good quality. However, the dilution test applied in the CTAB extracts showed no interference of inhibitory substances. The PCR efficiencies of lectin target amplification were not statistically different, and the coefficients of correlation (R²) demonstrated high degree of correlation between the copy numbers and the threshold cycle (Ct) values. ANOVA showed suitable adjustment of the regression and absence of significant linear deviations. The efficiencies of the p35S amplification were not statistically different, and all R² values using DNeasy extracts were above 0.98 with no significant linear deviations. Two out of three R² values using CTAB extracts were lower than 0.98, corresponding to lower degree of correlation, and the lack-of-fit test showed significant linear deviation in one run. The comparative analysis of the Ct values for the p35S and lectin targets demonstrated no statistical significant differences between the analytical curves of each target.

Assessment of fructooligosaccharides production from sucrose in aqueous and aqueous-organic systems using immobilized inulinase from Kluyveromyces marxianus NRRL Y-7571

This work investigated the fructooligosaccharides (FOS) synthesis by immobilized inulinase obtained from Kluyveromyces marxianus NRRL Y-7571 in aqueous and aqueous-organic systems using sucrose as substrate. The sequential strategy of experimental design was used to optimize the FOS conversion in both systems. For the aqueous-organic system, a 2(6-2) fractional design was carried out to evaluate the effects of temperature, sucrose concentration, pH, aqueous/organic ratio, enzyme activity, and polyethylene glycol concentration. For the aqueous system, a central composite design for the enzyme activity and the sucrose concentration was carried out. The highest fructooligosaccharides yield (Y FOS) for the aqueous-organic system was 18.2 ± S0.9 wt%, at 40 ºC, pH 5.0, sucrose concentration of 60% (w/w), enzyme activity of 4 U.mL-1, and aqueous/organic ratio of 25/75 wt%. The highest Y FOS for the aqueous system was 14.6 ± 0.9 wt% at 40 ºC, pH 5.0, sucrose concentration of 60 wt%, and enzyme activity of 4.0 U.mL-1.

Physical and chemical characterization of the pulp of different varieties of avocado targeting oil extraction potential

The aim of this study was to evaluate the physicochemical properties of avocado pulp of four different varieties (Avocado, Guatemala, Dickinson, and Butter pear) and to identify which has the greatest potential for oil extraction. Fresh avocado pulp was characterized by moisture, protein, fat, ash, carbohydrates and energy contents were determined. The carotenoids and chlorophyll contents were determined by the organic solvent extraction method. The results showed significant differences in the composition of the fruit when varieties are compared. However, the striking feature in all varieties is high lipid content; Avocado and Dickinson are the most suitable varieties for oil extraction, taking into account moisture content and the levels of lipids in the pulp. Moreover, it could be said that the variety Dickinson is the most affected by the parameters evaluated in terms of overall quality. Chlorophyll and carotenoids, fat-soluble pigments, showed a negative correlation with respect to lipids since it could be related to its function in the fruit. The varieties Avocado and Dickinson are an alternative to oil extraction having great commercial potential to be exploited thus avoiding waste and increasing farmers’ income.

Influence of the extraction method and storage time on the physicochemical properties and carotenoid levels of pequi (Caryocar brasiliense Camb.) oil

The objective of this study was to analyze the physicochemical properties and carotenoid levels of pequi oil obtained by different extraction methods and to evaluate the preservation of these properties and pigments during storage time. The pequi oil was obtained by solvent extraction, mechanical extraction, and hot water flotation. It was stored for over 180 days in an amber bottle at ambient conditions. Analyses for the determination of the acidity, peroxide, saponification and iodine values, coloration, total carotenoids, and β-carotene levels were conducted. The oil extraction with solvents produced the best yield and carotenoid levels. The oil obtained by mechanical extraction presented higher acidity (5.44 mg KOH.g-1) and peroxide values (1.07 mEq.kg-1). During the storage of pequi oil, there was an increase in the acidity and the peroxide values, darkening of the oil coloration, and a reduction of the carotenoid levels. Mechanical extraction is the less advantageous method for the conservation of the physicochemical properties and carotenoid levels in pequi oil.

Optimization of the extraction of carrageenan from Kappaphycus alvarezii using response surface methodology

This study aims to optimize an alternative method of extraction of carrageenan without previous alkaline treatment and ethanol precipitation using Response Surface Methodology (RSM). In order to introduce an innovation in the isolation step, atomization drying was used reducing the time for obtaining dry carrageenan powder. The effects of extraction time and temperature on yield, gel strength, and viscosity were evaluated. Furthermore, the extracted material was submitted to structural analysis, by infrared spectroscopy and nuclear magnetic resonance spectroscopy (¹H-NMR), and chemical composition analysis. Results showed that the generated regression models adequately explained the data variation. Carrageenan yield and gel viscosity were influenced only by the extraction temperature. However, gel strength was influenced by both, extraction time and extraction temperature. Optimal extraction conditions were 74 ºC and 4 hours. In these conditions, the carrageenan extract properties determined by the polynomial model were 31.17%, 158.27 g.cm-2, and 29.5 cP for yield, gel strength, and viscosity, respectively, while under the experimental conditions they were 35.8 ± 4.68%, 112.50 ± 4.96 g.cm-2, and 16.01 ± 1.03 cP, respectively. The chemical composition, nuclear magnetic resonance spectroscopy, and infrared spectroscopy analyses showed that the crude carrageenan extracted is composed mainly of κ-carrageenan.

Characterization and extraction of volatile compounds from pineapple (Ananas comosus L. Merril) processing residues

The aim of this study was to extract and identify volatile compounds from pineapple residues generated during concentrated juice processing. Distillates of pineapple residues were obtained using the following techniques: simple hydrodistillation and hydrodistillation by passing nitrogen gas. The volatile compounds present in the distillates were captured by the solid-phase microextraction technique. The volatile compounds were identified in a system of high resolution gas chromatography system coupled with mass spectrometry using a polyethylene glycol polar capillary column as stationary phase. The pineapple residues constituted mostly of esters (35%), followed by ketones (26%), alcohols (18%), aldehydes (9%), acids (3%) and other compounds (9%). Odor-active volatile compounds were mainly identified in the distillate obtained using hydrodistillation by passing nitrogen gas, namely decanal, ethyl octanoate, acetic acid, 1-hexanol, and ketones such as γ-hexalactone, γ-octalactone, δ-octalactone, γ-decalactone, and γ-dodecalactone. This suggests that the use of an inert gas and lower temperatures helped maintain higher amounts of flavor compounds. These data indicate that pineapple processing residue contained important volatile compounds which can be extracted and used as aroma enhancing products and have high potential for the production of value-added natural essences.

Extraction of DNA from honey and its amplification by PCR for botanical identification

The physiochemical and biological properties of honey are directly associated to its floral origin. Some current commonly used methods for identification of botanical origin of honey involve palynological analysis, chromatographic methods, or direct observation of the bee behavior. However, these methods can be less sensitive and time consuming. DNA-based methods have become popular due to their simplicity, quickness, and reliability. The main objective of this research is to introduce a protocol for the extraction of DNA from honey and demonstrate that the molecular analysis of the extracted DNA can be used for its botanical identification. The original CTAB-based protocol for the extraction of DNA from plants was modified and used in the DNA extraction from honey. DNA extraction was carried out from different honey samples with similar results in each replication. The extracted DNA was amplified by PCR using plant specific primers, confirming that the DNA extracted using the modified protocol is of plant origin and has good quality for analysis of PCR products and that it can be used for botanical identification of honey.