Extraction Of Aglycones Research Articles

Ultrahigh pressure enhances the extraction efficiency, antioxidant potential, and hypoglycemic activity of flavonoids from Chinese sea buckthorn leaves

Ultrahigh pressure extraction (UHPE) is an emerging non-thermal extraction technology, but whether this technology can be used for extracting flavonoids from sea buckthorn leaves remains unelucidated. This study optimized the processing conditions for flavonoid extraction from Chinese sea buckthorn leaves through UHPE and compared the results obtained with those of ultrasonic-assisted extraction (UAE) and conventional solvent extraction (CSE). The optimum parameters for UHPE were as follows: ethanol concentration, 50%; liquid-solid ratio, 41:1; pressure, 143 MPa; temperature, 44 °C; and holding time, 3 min. Compared with the UAE and CSE extracts the UHPE extracts had a higher content of total flavonoids and flavonol monomers, and the advantages of UPHE were primarily observed in the extraction of flavonoid monosaccharide chain glycosides and flavonoid aglycones. In total, 36 flavonoids were identified in the three extracts, with 33, 30, and 28 flavonoids identified in UHPE, UAE and CSE, respectively. Additionally, the UHPE extracts had a greater ferric reducing antioxidant power (FRAP), oxygen radical absorption capacity (ORAC), α-amylase inhibitory activity, and α-glucosidase inhibitory activity. In conclusion, the efficiency of UHPE to extract flavonoids from sea buckthorn leaves was significantly higher than those of UAE and CSE.

Simultaneous Extraction of Flavonoid Glycosides and Flavonoid Aglycones from Discarded Apple Branches by Enzyme-assisted Micelle-mediated Extraction with Cloud Point Enrichment Method

Simultaneous Extraction of Flavonoid Glycosides and Flavonoid Aglycones from Discarded Apple Branches by Enzyme-assisted Micelle-mediated Extraction with Cloud Point Enrichment Method

Multistep Optimization of β-Glucosidase Extraction from Germinated Soybeans (Glycine max L. Merril) and Recovery of Isoflavone Aglycones.

Epicotyls from germinated soybeans (EGS) have great potential as sources of endogenous β-glucosidase. Furthermore, this enzyme may improve the conversion of isoflavones into their corresponding aglycones. β-Glucosidase may also increase the release of aglycones from the cell wall of the plant materials. Therefore, the aim of this work was to optimize both the extraction of β-glucosidase from EGS and to further examine its application in defatted soybean cotyledon to improve the recovery of aglycones, which were evaluated by ultra-high performance liquid chromatography (UHPLC). A multistep optimization was carried out and the effects of temperature and pH were investigated by applying a central composite design. The linear effect of pH and the quadratic effect of pH and temperature were significant for the extraction of β-glucosidase and recovery aglycones, respectively. Optimum extraction of β-glucosidase from EGS occurred at 30 °C and pH 5.0. Furthermore, the maximum recovery of aglycones (98.7%), which occurred at 35 °C and pH 7.0–7.6 during 144 h of germination, increased 8.5 times with respect to the lowest concentration. The higher bioaccessibility of aglycones when compared with their conjugated counterparts is well substantiated. Therefore, the data provided in this contribution may be useful for enhancing the benefits of soybean, their products, and/or their processing by-products.

Deep eutectic solvents as green media for extraction of flavonoid glycosides and aglycones from Platycladi Cacumen

Deep eutectic solvents (DESs) are emerging as alternatives to conventional ionic liquids and organic solvents due to their unique advantages. In the present study, the tuneability of DESs as tailor-made solvents to efficiently extract polar and non-polar bioactive compounds from Platycladi Cacumen was detailedly investigated. Totally 12 types of choline chloride-, betaine-, and l-proline-based DESs were synthesized for initial screening, and extraction conditions was optimized by single-factor experiment. Experiments with different DESs and principal components analysis demonstrated that the extractability of both flavonoid glycosides and aglycones was greater with certain designed DESs than conventional solvents. In addition, the water content in DESs led to significantly different extraction yields of flavonoid compounds. The target compounds were recovered from DESs by macroporous resin LX-38 with a satisfactory yield between 77.44% and 98.92%. The knowledge acquired in this study could contribute to further DES application in extraction of bioactive compounds from natural sources.

Differential Isotope Labeling of 38 Dietary Polyphenols and Their Quantification in Urine by Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometry.

A large number of polyphenols are consumed with the diet and may contribute to the prevention of chronic diseases such as cardiovascular diseases, diabetes, cancers, and neurodegenerative diseases. More comprehensive methods are needed to measure exposure to this complex family of bioactive plant compounds in epidemiological studies. We report here a novel method enabling the simultaneous measurement in urine of 38 polyphenols representative of the main classes and subclasses found in the diet. This method is based on differential (12)C-/(13)C-isotope labeling of polyphenols through derivatization with isotopic dansyl chloride reagents and on the analysis of the labeled polyphenols by tandem mass spectrometry. This derivatization approach overcomes the need for costly labeled standards. Different conditions for enzyme hydrolysis of polyphenol glucuronides and sulfate esters, extraction, and dansylation of unconjugated aglycones were tested and optimized. Limits of quantification varied from 0.01 to 1.1 μM depending on polyphenols. Intrabatch coefficients of variation varied between 3.9% and 9.6%. Interbatch variations were lower than 15% for 31 compounds and lower than 29% for 6 additional polyphenols out of the 38 tested. Thirty seven polyphenols were validated and then analyzed in 475, 24 h urine samples from the European Prospective Investigation on Cancer and Nutrition (EPIC) study. Thirty four polyphenols could be detected and successfully estimated and showed large interindividual variations of concentrations (2-3 orders of magnitude depending on the compound), with median concentrations spanning from 0.01 to over 1000 μM for all 34 compounds.

Alleviating exercise-induced muscular stress using neat and processed bee pollen: oxidative markers, mitochondrial enzymes, and myostatin expression in rats

BackgroundThe current study was designed to investigate the influence of monofloral Indian mustard bee pollen (MIMBP) and processed monofloral Indian mustard bee pollen (PMIMBP) supplementation on chronic swimming exercise-induced oxidative stress implications in the gastrocnemius muscle of Wistar rats. MethodsMIMBP was processed with an edible lipid-surfactant mixture (Captex 355:Tween 80) to increase the extraction of polyphenols and flavonoid aglycones as analyzed by UV spectroscopy and high performance liquid chromatography-photo diode array. Wistar rats in different groups were fed with MIMBP or PMIMBP supplements at a dose of 100mg/kg, 200mg/kg and 300mg/kg individually, while being subjected to chronic swimming exercise for 4 weeks (5 d/wk). Various biochemical [superoxide dismutase (SOD), glutathione (GSH), malonaldehyde (MDA), nitric oxide (NO), and total protein content], mitochondrial (Complex I, II, III, and IV enzyme activity), and molecular (myostatin mRNA expression) parameters were monitored in the gastrocnemius muscle of each group. ResultsAdministration of both MIMBP (300mg/kg) and PMIMBP (100mg/kg, 200mg/kg, and 300mg/kg) wielded an antioxidant effect by significantly improving SOD, GSH, MDA, NO, and total protein levels. Further MIMBP (300mg/kg) and PMIMBP (200mg/kg and 300mg/kg) significantly improved impaired mitochondrial Complex I, II, III, and IV enzyme activity. Significant down-regulation of myostatin mRNA expression by MIMBP (300mg/kg) and PMIMBP (200mg/kg and 300mg/kg) indicates a muscle protectant role in oxidative stress conditions. ConclusionThe study establishes the antioxidant, mitochondrial upregulatory, and myostatin inhibitory effects of both MIMBP and PMIMBP in exercise-induced oxidative stress conditions, suggesting their usefulness in effective management of exercise-induced muscular stress. Further, processing of MIMBP with an edible lipid-surfactant mixture was found to improve the therapeutic efficiency of pollen.

Isoflavone extraction from okara using water as extractant

We here report on the use of water as a ‘green’ extraction solvent for the isolation of isoflavones from okara, a by-product of soymilk production. At a low liquid-to-solid ratio of 20 to 1 and 20°C, 47% of the isoflavones that can be extracted with 70% aqueous ethanol were extracted. The malonyl-glucosides were fully recovered with a ratio of 20 to 1, while β-glucosides were recovered with an increased liquid-to-solid ratio of 40 to 1. The extraction of aglycones was better at higher ratios, but leveled off before reaching a 100% yield. Temperature hardly affected the total amount of isoflavones. At a 20 to 1 ratio, 20°C, and pH 10, there was no significant difference (p>0.05) between isoflavone extraction in water and in 70% aqueous ethanol. The results suggest that water may be used as a green alternative for separation of isoflavones from okara.

Recovery of Aromatic Aglycones from Grape Pomace Winemaking By-Products by Using Liquid-Liquid and Pressurized-Liquid Extraction

The potential of winemaking grape pomace by-products as a source of glycosidic aroma precursors that under enzymatic hydrolysis might release aroma compounds has been evaluated. Two different extraction methodologies, liquid–liquid and pressurized liquid extraction (LLE and PLE) were employed. Solid phase extraction (SPE)-GC-MS analysis of the hydrolyzed LLE glycosidic extract revealed 22 aroma compounds belonging to different chemical families (terpenes, C13 norisoprenoids, vanillines, etc.). Response surface methodology was employed to study the effect of the most significant PLE experimental variables (temperature and solvent composition) on the extraction of aromatic aglycones. The parameters of the model were estimated by multiple linear regressions. Most of the aroma compounds showed an adequate fit to the calculated model (18 compounds from 22 with R 2 > 0.8). The application of the optimized PLE conditions (50 % of ethanol in the hydroalcoholic solution) and 90 °C showed higher extraction yield of aglycones when comparing with the extraction yield obtained by LLE.

Molecular imprinted polymer for solid‐phase extraction of flavonol aglycones from Moringa oleifera extracts

Molecular imprinted polymer produced using quercetin as the imprinting compound was applied for the extraction of flavonol aglycones (quercetin and kaempferol) from Moringa oleifera methanolic extracts obtained using heated reflux extraction method. Identification and quantification of these flavonols in the Moringa extracts was achieved using high performance liquid chromatography with ultra violet detection. Breakthrough volume and retention capacity of molecular imprinted polymer SPE was investigated using a mixture of myricetin, quercetin and kaempferol. The calculated theoretical number of plates was found to be 14, 50 and 8 for myricetin, quercetin and kaempferol, respectively. Calculated adsorption capacities were 2.0, 3.4 and 3.7 μmol/g for myricetin, quercetin and kaempferol, respectively. No myricetin was observed in Moringa methanol extracts. Recoveries of quercetin and kaempferol from Moringa methanol extracts of leaves and flowers ranged from 77 to 85% and 75 to 86%, respectively, demonstrating the feasibility of using the developed molecularly imprinted SPE method for quantitative clean-up of both of these flavonoids. Using heated reflux extraction combined with molecularly imprinted SPE, quercetin concentrations of 975 ± 58 and 845 ± 32 mg/kg were determined in Moringa leaves and flowers, respectively. However, the concentrations of kaempferol found in leaves and flowers were 2100 ± 176 and 2802 ± 157 mg/kg, respectively.

Simultaneous determination of naphthalene and anthraquinone derivatives in Rumex nepalensis Spreng. Roots by HPLC: comparison of different extraction methods and validation

Rumex nepalensis contains mainly anthraquinone and naphthalene derivatives. Although HPLC methods have been reported for the analysis of anthraquinones, neither a phytochemical analysis of Rumex species nor the simultaneous determination of anthraquinone and naphthalene derivatives in other samples has been reported so far. To develop and validate a HPLC method for the simultaneous determination of anthraquinone and naphthalene derivatives in R. nepalensis roots. Anthraquinones and naphthalenes were extracted from R. nepalensis roots by three methods (reflux, ultrasonication and pressurized liquid extraction) using methanol. Separation was achieved on an RP C₁₈ column with a gradient mobile phase consisting of 0.05% orthophosphoric acid in water (solvent A) and methanol (solvent B) using a UV detector (254 nm). Small differences were observed in the contents of anthraquinone and naphthalene derivatives extracted by the three methods. Chrysophanol-8-O-β-D-glucopyranoside and nepodin were detected as major constituents. The method showed a good linearity (r² > 0.9992), high precision (RSD < 5%) and a good recovery (97-105%) of the compounds. The lowest detection limit was found to be 0.97 ng and the method was found to be robust. Reflux and ultrasonication were found to be the best suited methods for the extraction of glycosides and aglycones, respectively. The developed and validated HPLC method is simple, precise and accurate; and can hence be recommended as the method of choice for the analysis of anthraquinones and naphthalenes in R. nepalensis and other Rumex species for both quality control as well as routine analytical purposes.

Simultaneous Extraction of Bioactive Limonoid Aglycones and Glucoside from Citrus aurantium L. Using Hydrotropy

Citrus limonoids were demonstrated to possess potential biological activities in reducing the risk of certain diseases. Limonoids are present in citrus fruits in the form of aglycones and glucosides. At present, limonoid aglycones and limonoid glucosides are extracted in multiple steps using different solvents. In order to understand their potential bioactivity, it may be beneficial to isolate and purify these compounds using environment friendly methods. A new method of extraction and purification of limonoids was established using a hydrotrope polystyrene adsorbent resin. Extraction of aglycones and glucosides was achieved in a single step, using an aqueous solution of sodium cumene sulphonate (Na-CuS). Sour orange (Citrus aurantium L.) seed powder was extracted with 2 M Na-CuS solution at 45 degrees C for 6 h. The filtered extract was diluted with water and loaded on an SP 700 adsorbent column. The column was washed with distilled water to remove the hydrotrope and then eluted using water and methanol in different compositions to obtain three compounds. The structures of the isolated compounds were confirmed by NMR spectroscopy as deacetyl nomilinic acid glucoside (DNAG), deacetyl nomilin (DAN) and limonin (LIM).

Hydrotropic extraction of bioactive limonin from sour orange ( Citrusaurantium L.) seeds

Limonoids are potential bioactive compounds present only in citrus among fruits and vegetables. A new process for extraction of limonoid aglycones from sour orange ( Citrus aurantium L.) seeds was investigated using aqueous hydrotropic solutions. The extraction efficiency was dependent on hydrotrope concentration, extraction temperature and percent of raw material loaded. Two hydrotropes such as sodium salicylate (Na-Sal) and sodium cumene sulphonate (Na-CuS) were studied using Box-Behnken experiment design. Response surface analysis (RSA) of data was performed to study the effect of parameters on extraction efficiency. Prominent limonoid aglycone such as limonin was extracted and quantified for process optimization. Both hydrotropes gave maximum limonin yield at 2 M concentration, extraction temperature of 45 °C and 10% solid loading. A maximum limonin yield of 0.65 mg/g seeds was obtained using Na-CuS whereas only 0.46 mg/g seed was obtained using Na-Sal. Using this process, the use of organic solvents can be reduced dramatically to keep the process environmental friendly for the extraction of bioactive compounds.

Optimization of a rapid microwave assisted extraction method for the liquid chromatography–electrospray-tandem mass spectrometry determination of isoflavonoid aglycones in soybeans

A very fast chromatographic separation of isoflavonoids genistein, daidzein, formononetin and biochanin A was developed on a C18 high-speed column under isocratic conditions. The method was validated in terms of detection limits, quantitation limits (LOQs), linearity and precision. LOQs in 0.04–0.2 μg/g range were calculated, making feasible the determination of these compounds of nutritional concern at trace levels. Good linearity was demonstrated over three concentration orders of magnitude for each analyte ( r 2 0.990–1.000). The intra-day and inter-day repeatability was evaluated in terms of relative standard deviation (RSD%) at two concentration levels for each analyte (RSD% <9%). An optimization strategy was adopted to find the best conditions for the extraction of isoflavonoid aglycones from yellow soybeans using microwave-assisted extraction. The most relevant parameters resulted to be the microwave power, the extraction time and the acid concentration, optimal values being 600 W, 1 min and 12 M, respectively. When performing sample treatment on a fortified soybean sample, high recovery percentage was obtained for both compounds (94 ± 8% for daidzein and 97 ± 5% ( n = 4) for genistein). The concentration level at which daidzein and genistein were found in the soybean sample were 1.21 ± 0.15 mg/g and 2.38 ± 0.09 mg/g ( n = 4), respectively.

Supercritical fluid extraction of daidzein and genistein isoflavones from soybean hypocotyl after hydrolysis with endogenous β-glucosidases

An optimal condition of supercritical fluid extraction (SFE) for isoflavone aglycones (daidzein and genistein) in soybean hypocotyls previously subjected to thermohydration at pH 5.0 and a temperature of 50 °C for 6, 12 and 18 h was developed. Different temperatures, pressures and cosolvents (methanol, ethanol, and acetonitrile) was tested and compared with solid–liquid extraction using aqueous methanol solution (80% v v −1) conducted in parallel for comparison. The extraction conditions were 50–70 °C, 176–380 bar, adding 0, 5, 10 mol% of cosolvents 80% in water as a modifier. The results from SC–CO 2 showed that the cosolvent and pressure have significant effects in the extraction efficiency. It was found that the extraction conditions promoting the highest extraction of daidzein and genistein were at the temperature of 60 °C, pressure of 380 bar and both static and dynamic extraction of 15 min with the addition of 10% acetonitrile (80% v v −1). The maximum amounts of daidzein and genistein extracted by each method were solid–liquid extraction (70.07 mg 100 g −1) and carbon dioxide–acetonitrile (17.97 mg 100 g −1). The yield of daidzein and genistein achieved by a 30 min SC–CO 2 extraction on soybean hypocotyls after 12 h soaking time was markedly improved by the addition of a modifier (acetonitrile) to the CO 2 fluid. HPLC analysis of the obtained extracts revealed that extraction of isoflavone aglycones by SC–CO 2 was 4.78 and 13.19 mg 100 g −1 for daidzein and genistein, respectively. The contents of daidzein and genistein obtained in the solid–liquid extraction were superior to 86% and 63%, respectively, compared to supercritical extraction.

Citrus Bioactive Limonoid Extraction using Environment-friendly Hydrotropy

Citrus consumption has been shown to promote human health due to presence of several bioactive compounds. In the process of understanding the health benefits of citrus, we need to isolate and characterize these compounds. Limonoids are one of such prominent, but lesser-known phytonutrients that have been shown to prevent cancers of the mouth, skin, lung, breast, and colon. With the growing interest in the health-promoting properties of citrus limonoids, the demand for these bioactives has significantly increased. It has been critical to explore environment-friendly extraction methods rather than using hazardous organic solvents. A water-based hydrotropic extraction of limonoid aglycones from sour orange (Citrus aurantium L.) seeds was developed. Two hydrotropes, sodium salicylate (Na-Sal) and sodium cumene sulfonate (Na-CuS), were studied for extraction efficiency using the Box Behnken experiment design method. The extraction efficiency of prominent aglycone limonin was observed depending on hydrotrope concentration, extraction temperature, and percentage of raw material loading. Response Surface Analysis (RSA) of data predicted the optimum conditions for maximum yield. Recovery of aglycones from filtered extract is also easily achieved by mere dilution using water at pH 3 or 7 or by partitioning the extract with dichloromethane. At optimum conditions, limonin yield of 0.46 mg/g seeds in the case of Na-Sal extraction and 0.65 mg/g seeds in the case of Na-CuS extraction was achieved. The results demonstrated that the hydrotropic extraction process of limonoid aglycones has practical commercial importance. This project is based upon work supported by the USDA–CSREES IFAFS #2001 52102 02294 and USDA–CSREES #2005-34402-14401 “Designing Foods for Health” through the Vegetable and Fruit Improvement Center.

HPLC identification and determination of myricetin, quercetin, kaempferol and total flavonoids in herbal drugs

A new and rapid HPLC method for identification and determination of myricetin, quercetin, kaempferol and total flavonoids in ten herbal drugs of Macedonian origin is presented. Preparation of samples (Uvae ursi folim, Pruni spinosae flos, Sambuci flos, Betulae folim, Primulae flos, Herniariae herba, Centaurii herba, Tiliae flos, Robiniae pseudoacaciae flos, Bursae pastoris herba) included hydrolysis of glycosides and extraction of total aglycones with ethyl acetate. HPLC analysis with UV-diode array detection was carried out on RP C18 column, using 5% acetic acid and acetonitrile in agradient elution mode and column temperature of 30 o C. The monitoring of the elution is performed in the whole UV-range and the acquisition of data for quantitative analysis at 367 nm. Screening of the extracts showed presence of quercetin in nine, kaempferol in seven and myricetin in only one sample. The quantitative analysis showed that the content of quercetin ranged from 0.026-0.506 % (m/m), while for kaempferol it was from traces to 1.246 %. Uvaeursi folium and Pruni spinosae flos were rich in content of quercetin (0.482 % and 0.506 %, respectively), while Pruni spinosae flos and Robiniae pseudoaccaciae flos contained the highest amounts of kaempferol (1.246 % and 0.892 %, respectively). Myricetin was identified and determined only in Betulae folium (0.102 %). The content of total flavonoids in the investigated samples expressed in terms of quercetin ranged from 0.040 to 1.680 %. The proposed HPLC method is convenient for use in routine analysis of myricetin, quercetin and kaempferol, as well as for estimation of total flavonoids content in herbal drugs.

Validation of a high-performance liquid chromatographic method for determination of isoflavonoids in soybeans. Study of the extraction procedure by experimental design

An improved analytical method, reversed-phase high-performance liquid chromatography on a narrow-bore C18 column, has been developed for the simultaneous determination of genistein, daidzein, formononetin, and biochanin A. The method was validated in terms of detection limits, quantitation limits (LOQ), linearity, and precision.LOQ in the 0.04–0.1 μg mL−1 range were calculated, enabling determination of these compounds of nutritional concern at trace levels. Good linearity was demonstrated over three orders of magnitude of concentration for each analyte (r 2=0.998–1.000). The intra-day repeatability was evaluated in terms ofRSD (%) at two concentration levels for each analyte (RSD (%) 0.05 at the 95% confidence level). The method was applied to the determination of genistein and daidzein in yellow soybeans, after optimization of the method for extraction of isoflavonoid aglycones from soybeans by experimental design, i.e. central composite design. Extraction recoveries up to 87±4% were obtained when the corresponding glycosidic forms (genistin and daidzin) were added to soybean samples.

Modified method for tuber glycoalkaloid and leaf glycoalkaloid analysis

A widely employed, comprehensive method exists for the determination of tuber total glycoalkaloids (TTGA). This method, which consists of a bisolvent extraction, preparation, and purification of aglycones and subsequent titration, was not acceptable for quantification of leaf total glycoalkaloids (LTGA). A modification of an existing extraction procedure was developed and tested. When the original bisolvent and the newly developed acetic acid extractions were contrasted percent recovery of a-solanine from leaf tissue was 19–27% and 91–97%, respectively. This new procedure was easily adapted for TTGA analysis and demonstrated improved recovery of α-solanine from tuber tissue when compared with the bisolvent extraction.