Supercritical fluid extraction of lycopene from tomato processing byproducts

Tomato seeds and skins acquired from the byproduct of a local tomato processing facility were studied for supercritical fluid extraction (SFE) of phytochemicals. The extracts were analyzed for lycopene, beta-carotene, alpha-carotene, alpha-tocopherol, gamma-tocopherol, and delta-tocopherol content using high-performance liquid chromatography-electrochemical detection and compared to a chemically extracted control. SFEs were carried out using CO(2) at seven temperatures (32-86 degrees C) and six pressures (13.78-48.26 MPa). The effect of CO(2) flow rate and volume also was investigated. The results indicated that the percentage of lycopene extracted increased with elevated temperature and pressure until a maximum recovery of 38.8% was reached at 86 degrees C and 34.47 MPa, after which the amount of lycopene extracted decreased. Conditions for the optimum extraction of lycopene from 3 g of raw material were determined to be 86 degrees C, 34.47 MPa, and 500 mL of CO(2) at a flow rate of 2.5 mL/min. These conditions resulted in the extraction of 61.0% of the lycopene (7.19 microg lycopene/g).     

Effects of solvent and temperature on pressurized liquid extraction of anthocyanins and total phenolics from dried red grape skin

Pressurized liquid extraction (PLE) was used to extract anthocyanins from the freeze-dried skin of a highly pigmented red wine grape with six solvents at 50 degrees C, 10.1 MPa, and 3 x 5 min extraction cycles. Temperature (from 20 to 140 degrees C in 20 degrees C increments) effects on anthocyanin recovery by acidified water and acidified 60% methanol were also studied. Acidified methanol extracted the highest levels of total monoglucosides and total anthocyanins, whereas the solvent mixture (40:40:20:0.1 methanol/acetone/water HCl) extracted the highest levels of total phenolics and total acylated anthocyanins. Acidified water extracts obtained by PLE at 80-100 degrees C had the highest levels of total monoglucosides, total acylated anthocyanins, total anthocyanins, total phenolics, and ORAC values. Acidified methanol extracts obtained by PLE at 60 degrees C had the highest levels of total monoglucosides and total anthocyanins, whereas extracts obtained at 120 degrees C had the highest levels of total phenolics. High-temperature PLE (80-100 degrees C) using acidified water, an environmentally friendly solvent, was as effective as acidified 60% methanol in extracting anthocyanins from grape skins.     

Subcritical water extraction of antioxidant compounds from rosemary plants

Subcritical water extraction at several temperatures ranging from 25 to 200 degrees C has been studied to selectively extract antioxidant compounds from rosemary leaves. An exhaustive characterization of the fractions obtained using subcritical water at different temperatures has been carried out by LC-MS, and the antioxidant activities of the extracts have been measured by a free radical method (DPPH). Results indicate high selectivity of the subcritical water toward the most active compounds of rosemary such as carnosol, rosmanol, carnosic acid, methyl carnosate, and some flavonoids such as cirsimaritin and genkwanin. The antioxidant activity of the fractions obtained by extraction at different water temperatures was very high, with values around 11.3 microg/mL, comparable to those achieved by SFE of rosemary leaves. A study of the effect of the temperature on the extraction efficiency of the most typical rosemary antioxidant compounds has been performed.     

Supercritical carbon dioxide extraction of essential oils from Perovskia atriplicifolia Benth

The supercritical fluid extraction (SFE) of the aerial parts of Perovskia atriplicifolia Benth. was studied. The effect of different parameters such as pressure, temperature, modifier identity, and modifier volume on the SFE of the plant was investigated. The extracts were analyzed by GC and GC-MS and compared with the essential oil obtained from P. atriplicifolia Benth by steam distillation. The supercritical extracts and the steam-distilled products had very different compositions. The main constituents of the oil obtained by steam distillation were 1,8-cineole, limonene, camphor, beta-caryophyllene, alpha-pinene, camphene, and alpha-humulene. On the other hand, the major components of SFE extracts were 1,8-cineole, limonene, camphor, beta-caryophyllene, gamma-cadinene, alpha-pinene, and alpha-terpinyl acetate. The results showed that increasing the temperature from 35 to 65 degrees C (at a constant pressure of 100 atm) drastically reduced the number of extracted components. Also, the number of extracted constituents and the percent of main analytes increased when lower pressures were used. Using different modifiers (e.g., methanol, ethanol, dichloromethane, and hexane) for the extraction of the plant at low pressure (100 atm) and temperature (35 degrees C) showed that hexane was more selective than the other modifiers.     

Evaluation of the light-sensitive cytotoxicity of Hypericum perforatum extracts, fractions, and pure compounds

Hypericum perforatum (Hp) is known for possessing antidepressant and antiviral activities. Despite its use as an alternative to conventional antidepressants, the identification of the cytotoxic chemicals derived from this herb is incomplete. In this study, the cytotoxicity of Hp extracts prepared in solvents ranging in polarity, fractions of one extract, and purified compounds were examined in three cell lines. All extracts exhibited significant cytotoxicity; those prepared in ethanol (no hyperforin, 3.6 microM hypericin, and 134.6 microM flavonoids) showed between 7.7 and 77.4% cell survival (p < 0.0001 and 0.01), whereas the chloroform and hexane extracts (hyperforin, hypericin, and flavonoids not detected) showed approximately 9.0 (p < 0.0001) and 4.0% (p < 0.0001) survival. Light-sensitive toxicity was observed primarily with the ethanol extracts sequentially extracted following removal of material extracted in either chloroform or hexane. The absence of light-sensitive toxicity with the Hp extracts suggests that the hypericins were not playing a prominent role in the toxicity of the extracts.     

Online RP-HPLC-DPPH screening method for detection of radical-scavenging phytochemicals from flowers of Acacia confusa

Acacia confusa is traditionally used as a medicinal plant in Taiwan. In this study, phytochemicals and antioxidant activities of extracts from flowers of A. confusa were investigated for the first time. In addition, a rapid screening method, online RP-HPLC-DPPH system, for individual antioxidants in complex matrices was developed. Accordingly, six antioxidants including gallic acid ( 1), myricetin 3-rhamnoside ( 2), quercetin 3-rhamnoside ( 3), kaempferol 3-rhamnoside ( 4), europetin 3-rhamnoside ( 5), and rhamnetin 3-rhamnoside ( 6) were detected using the developed screening method. Of these, compounds 2, 3, and 5 were found to be major bioactive phytochemicals, and their contents were determined as 11.3, 6.7, and 8.7 mg/g of crude extract, respectively. By comparison with quercetin, a well-known antioxidant, these compounds had the order of compound 2 > compound 5 > quercetin > compound 3 for DPPH radical-scavenging activity. Their IC 50 values were 3.0, 3.2, 4.5, and 7.4 microM, respectively. Moreover, the same order was observed for superoxide radical-scavenging activity, and their IC50 values were 2.6, 2.7, 4.3, and 5.3 microM, respectively. However, for lipid peroxidation, quercetin, an aglycon, showed the best inhibitory activity. The IC50 values of quercetin, compound 2, compound 5, and compound 3 were 46.7, 88.5, 90.7, and 124.6 microM, respectively. These results indicated that a rhamnoside at the C3 position of flavonoids had a negative effect on radical-scavenging activity and antilipid peroxidation. In contrast, the number of hydroxyl groups on the B-ring exhibited a positive relationship with their inhibitory activities.     

Novel field sampling procedure for the determination of methiocarb residues in surface waters from rice fields.

Methiocarb was extracted from surface water samples collected at experimental rice field sites in Louisiana and Texas. The sampling system consisted of a single-stage 90-mm Empore extraction disk unit equipped with a battery-powered vacuum pump. After extraction, the C-18 extraction disks were stored in an inert atmosphere at -10 degrees C and shipped overnight to the laboratory. The disks were extracted with methanol and the extracts analyzed by reversed-phase high-performance liquid chromatography with a methanol/water mobile phase. Methiocarb was detected by ultraviolet absorption at 223 nm and quantified with the use of calibration standards. Recoveries from control surface water samples fortified at 5.0, 10, 50, and 100 ng/mL methiocarb averaged 92 +/- 7%. A method limit of detection for methiocarb in rice field surface water was estimated to be 0.23 ng/mL at 223 nm.     

Investigation of Australian olive mill waste for recovery of biophenols

Olive mill waste is a potential source for the recovery of phytochemicals with a wide array of biological activities. Phytochemical screening of hexane, methanol, and water extracts revealed a diversity of compounds, perhaps overlooked in previous studies through intensive cleanup procedures. Methanol and water extracts contained large amounts of biophenols, and further testing of polar extraction solvents, including ethyl acetate, ethanol, propanol, acetone, acetonitrile, and water/methanol mixtures, highlighted the latter as the solvent of choice for extraction of the widest array of phenolic compounds. Stabilization of the resulting extract was best achieved by addition of 2% (w/w) sodium metabisulfite. Quantitative data are reported for nine biophenols extracted using 60% (v/v) methanol in water with 2% (w/w) sodium metabisulfite. Six compounds had recoveries of greater than 1 g/kg of freeze-dried waste: hydroxytyrosol glucoside, hydroxytyrosol, tyrosol, verbascoside, and a derivative of oleuropein.     

Isolation and partial characterization of rennet-like proteases from Australian cardoon (Cynara cardunculus L.)

The yield, protein content, proteolytic activity, and substrate specificity of crude and partially purified extracts from dried and fresh Australian cardoon (Cynara cardunculus L.) flowers were determined. Crude water extracts had high yield but low protein content and proteolytic activity, whereas citric acid extracts had low yield but high protein content and proteolytic activity. Fresh flower extracts gave higher yield and proteolytic activity but lower protein content in comparison with dried flower extracts. Purification with ammonium sulfate resulted in significantly increased proteolytic activity for water extracts from both fresh and dried cardoon flowers, whereas the proteolytic activity of citric acid extracts did not change significantly after purification. Irrespective of extraction method, all extracts had higher proteolytic activity against ovine whole and kappa-caseins compared to their bovine counterparts, showing optimal activity at 37 degrees C and pH 6.0. Separation of purified extracts by ion-exchange liquid chromatography yielded three active fractions, each of which when assayed with sodium dodecyl sulfate capillary electrophoresis revealed two subunits with molecular masses of 15.5 and 33.1 kDa, respectively.     

Advantages of soxflo extractions for phytochemical analysis and bioassay screening. 1. Terpenoids

The Soxflo technique was evaluated for the rapid extraction of plant materials (<90 min) at room temperature. Qualitatively similar chromatograms were obtained by gas chromatography and thin layer chromatography (TLC) with Soxflo (SoF) and Soxhlet (SoL) extracts. Sequential solvent extractions by SoF gave slightly higher yields (132%) of five major sesquiterpenoids. TLC revealed that SoF extractions at room temperature were more selective as extracts contained compounds with a narrower range of R(f)() values. This means that the SoF technique offers the potential for one-step extractions and partial fractionation. This study also showed that there were large differences in the volatile composition of dried and fresh Piper fruits: dried fruits had predominantly sesquiterpenoids while fresh fruits had considerable quantities of both mono- and sesquiterpenoids. This is the first report of alpha-guaiene and alpha- and beta-selinene in Piper guineense fruits. It is suggested that the SoF technique can be useful for the screening of large numbers of plants for phytochemicals or for the preparation of plant extracts for subsequent bioassay studies.     

Antioxidant activity of extracts of black sesame seed (Sesamum indicum L.) by supercritical carbon dioxide extraction

Antioxidant activities of extracts derived from sesame seed by supercritical carbon dioxide (SC-CO(2)) extraction and by n-hexane were determined using alpha,alpha-diphenyl-beta-picylhydrazyl (DPPH) radical scavenging and linoleic acid system methods. The highest extracted yield was given at 35 degrees C, 40 MPa, and a CO(2) flow rate of 2.5 mL min(-1) by an orthogonal experiment. The yields of extracts increased with increasing pressure, and yields at 40 and 30 MPa were higher than that by solvent extraction at 46.50%. Results from the linoleic acid system showed that the antioxidant activity follows the order: extract at 35 degrees C, 20 MPa > BHT > extract at 55 degrees C, 40 MPa > extract at 55 degrees C, 30 MPa > Trolox > solvent extraction > alpha-tocopherol. The SC-CO(2) extracts exhibited significantly higher antioxidant activities comparable to that by n-hexane extraction. The extracts at 30 MPa presented the highest antioxidant activities assessed in the DPPH method. At 20 MPa, the EC(50) increased with temperature, which indicated that the antioxidant activity was decreased in a temperature-dependent manner. The significant differences of antioxidant activities were found between the extracts by SC-CO(2) extraction and n-hexane. However, no significant differences were exhibited among the extracts by SC-CO(2) extraction. The vitamin E concentrations were also significantly higher in SC-CO(2) extracts than in n-hexane extracts, and its concentrations in extracts corresponded with the antioxidant activity of extracts.     

LC-ESI-MS study of the flavonoid glycoside malonates of red clover (Trifolium pratense)

High-performance liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) was applied to the analysis of the flavonoids and their glycoside malonates of the flowers and leaves of red clover (Trifolium pratense). Through LC-MS comparative studies on the plant extracts and their malonate-free extracts, approximately 20 flavonoid glycoside malonates were detected in the flower extract. Eight were identified as genistin 6' '-O-malonate (39), formononetin 7-O-beta-D-glucoside 6' '-O-malonate (40), biochanin A 7-O-beta-D-glucoside 6' '-O-malonate (41), trifoside 6' '-O-malonate (42), irilone 4'-O-beta-D-glucoside 6' '-O-malonate (43), pratensein 7-O-beta-D-glucoside 6' '-O-malonate (44), isoquercitrin 6' '-O-malonate (45), and 3-methylquercetin 7-O-beta-D-glucoside 6' '-O-malonate (46). About 15 other flavonoids and clovamides were proved to be present in this extract. The study also found that the flowers contained flavones as the major flavonoids, whereas the leaves had isoflavones as the major flavonoids. This is the first detection of the six malonates (39 and 42-46) in the extracts of red clover, and among them, 42, 43, and 46 are new compounds.     

Variation of chemical composition of the lipophilic extracts from yellow birch (Betula alleghaniensis) foliage

The occurrence of biologically active compounds identified for the first time in the lipophilic extracts of yellow birch (Betula alleghaniensis Britt.) foliage led to the quantification of the seasonal variation of their concentrations. Yellow birch foliage was collected from late June until late September 2003 in two different regions of Quebec. The extraction yields using hexane as a solvent were determined, and the extracts were analyzed by GC-MS to identify their molecular composition. In terms of both extraction yields and the concentration of the targeted molecules present in the extracts, mid-September has been determined as the best time to collect foliage samples. A total of 14 constituents were identified in these extracts. This is the first report of the presence of all of these constituents in yellow birch foliage and of some of them in the genus Betula. The most important compounds identified in yellow birch foliage extracts are triterpene squalene and aliphatic hydrocarbon tetracosan, aliphatic alcohol phytol, fatty acids hexadecanoic and octadecanoic, pentacyclic triterpenes alpha- and beta-amyrin, and phytosterol stigmast-5-en-3-ol.     

酸性浸润干燥辅助低水耗水代法提取亚麻籽油工艺

为解决传统水代法提取亚麻籽油过程中乳状液生成过多,耗水量大等问题,该文在低料液比1:2.5 kg/L的条件下,探究了水代法提取亚麻籽油的工艺。结果表明,酸浸润预处理通过影响亚麻蛋白的溶解度,有效提高水代法中的清油得率,由未处理时的18.95%±0.91%提升至83.27%±0.67%。水代法提取亚麻籽油的工艺优化结果为:pH值9.0、温度50℃、料液比1∶2.5 kg/L、提取时间2 h。在此条件下,清油得率为82.88%±0.30%。在水代法提油后的水相中添加50%原料质量的纯水重复提取渣相后,渣相残油率从3.97%±0.11%降至2.09%±0.04%。剩余乳状液经木瓜蛋白酶破乳后,总清油得率为93.44%±0.29%。水代法得到的亚麻籽油各项指标均符合一级成品亚麻籽油标准。该研究为亚麻籽油的高效提取提供了一种新的思路。     

Suppressive effect of a proanthocyanidin-rich extract from longan (Dimocarpus longan Lour.) flowers on nitric oxide production in LPS-stimulated macrophage cells

Anthocyanidins found in certain flowers have been shown to act as strong antioxidants in various systems, exhibiting multiple biological actions. The antioxidative effects of water extract and ethanolic extract of longan (Dimocarpus longan Lour.) flowers were evaluated by radical scavenging activity and compared to those of gallic acid, myricetin, and epigallocatechin gallate. In this study, the suppressive effects of longan flower extracts on nitric oxide and prostaglandin E2 production were investigated using a lipopolysaccharide-stimulated RAW 264.7 cell model. Abundant levels of phenolic compounds including flavonoids, condensed tannins, and proanthocyanidins were found in water or ethanolic extracts prepared from dried longan flowers. The antioxidative effect of longan flower extract was similar to the effect exhibited by pure antioxidants. Moreover, longan flower extract showed prominent inhibitory effects on prostaglandin E2 production. Significant concentration-dependent inhibition of nitric oxide production was detected when cells were cotreated with lipopolysaccharide and various concentrations of longan flower extracts. These inhibitory effects were further attributed to suppression of inducible nitric oxide synthase protein expression and not to reduced enzymatic activity. These results suggest that longan flower crude extracts, especially ethanolic extract, have antioxidant and anti-inflammatory effects, and the probable mechanism involves inhibition of inflammation by proanthocyanidins. Preliminary observations suggest that longan flower extract, especially alcoholic extract, could be another potential source of natural dietary antioxidant and anti-inflammatory agent.     

Apples prevent mammary tumors in rats

Regular consumption of fruits and vegetables has been consistently shown to be associated with reduced risk of developing chronic diseases such as cancer and cardiovascular disease. Apples are commonly consumed and are the major contributors of phytochemicals in human diets. It was previously reported that apple extracts exhibit strong antioxidant and antiproliferative activities and that the major part of total antioxidant activity is from the combination of phytochemicals. Phytochemicals, including phenolics and flavonoids, are suggested to be the bioactive compounds contributing to the health benefits of apples. Here it is shown that whole apple extracts prevent mammary cancer in a rat model in a dose-dependent manner at doses comparable to human consumption of one, three, and six apples a day. This study demonstrated that whole apple extracts effectively inhibited mammary cancer growth in the rat model; thus, consumption of apples may be an effective strategy for cancer protection.     

Phytochemicals of apple peels: isolation, structure elucidation, and their antiproliferative and antioxidant activities

Bioactivity-guided fractionation of Red Delicious apple peels was used to determine the chemical identity of bioactive constituents, which showed potent antiproliferative and antioxidant activities. Twenty-nine compounds, including triterpenoids, flavonoids, organic acids and plant sterols, were isolated using gradient solvent fractionation, Diaion HP-20, silica gel, and ODS columns, and preparative HPLC. Their chemical structures were identified using HR-MS and 1D and 2D NMR. Antiproliferative activities of isolated pure compounds against HepG2 human liver cancer cells and MCF-7 human breast cancer cells were evaluated. On the basis of the yields of isolated flavonoids (compounds 18- 23), the major flavonoids in apple peels are quercetin-3-O-beta-D-glucopyranoside (compound 20, 82.6%), then quercetin-3-O-beta-D-galactopyranoside (compound 19, 17.1%), followed by trace amounts of quercetin (compound 18, 0.2%), (-)-catechin (compound 22), (-)-epicatechin (compound 23), and quercetin-3-O-alpha-L-arabinofuranoside (compound 21). Among the compounds isolated, quercetin (18) and quercetin-3-O-beta-D-glucopyranoside (20) showed potent antiproliferative activities against HepG2 and MCF-7 cells, with EC 50 values of 40.9 +/- 1.1 and 49.2 +/- 4.9 microM to HepG2 cells and 137.5 +/- 2.6 and 23.9 +/- 3.9 microM to MCF-7 cells, respectively. Six flavonoids (18-23) and three phenolic compounds (10, 11, and 14) showed potent antioxidant activities. Caffeic acid (10), quercetin (18), and quercetin-3-O-beta-D-arabinofuranoside (21) showed higher antioxidant activity, with EC 50 values of <10 microM. Most tested flavonoids and phenolic compounds had high antioxidant activity when compared to ascorbic acid and might be responsible for the antioxidant activities of apples. These results showed apple peel phytochemicals have potent antioxidant and antiproliferative activities.     

Pressurized fluids for extraction of cedarwood oil from Juniperus virginianna

The extraction of cedarwood oil (CWO) using liquid carbon dioxide (LC-CO(2)) was investigated and compared to supercritical fluid extraction, including the effects of extraction pressure and length of extraction. The chemical composition of the extracts was monitored over the course of the extraction as well. The cumulative yields of CWO from cedarwood chips using 80 L of carbon dioxide varied very little treatment to treatment, with all temperature/pressure combinations yielding between 3.55 and 3.88% CWO, and the cumulative yields were statistically equivalent. The rate of extraction was highest under the supercritical extraction conditions (i.e., 100 degrees C and 6000 psi). Under the liquid CO(2) conditions (i.e., 25 degrees C), the extraction rates did not vary significantly with extraction pressure. However, there were differences in the chemical composition of the collected CWO. Extractions at 100 degrees C gave a much lower ratio of cedrol/cedrene than extractions at 25 degrees C. The highest ratio of cedrol/cedrene was obtained using 25 degrees C and 1500 psi. The use of subcritical water was also investigated for the extraction of CWO as well. Although some CWO was extracted using this method, the temperature/pressure combinations that gave the highest weight percentage yields also gave oils with an off odor while those combinations that gave a higher quality oil had very low yields. It appears that the high temperatures and acidic conditions cause a dehydration of the tertiary alcohol, cedrol, to its hydrocarbon analogue, cedrene, during CO(2) or pressurized water extractions of cedarwood.     

Development of a new method for the complete extraction of carotenoids from cereals with special reference to durum wheat (Triticum durum Desf.)

Due to the growing interest in the role of carotenoids in human health, their qualitative and quantitative analysis in foods is becoming more and more important. High-performance liquid chromatography has become the method of choice for the determination of these phytochemicals. A crucial step prior to the chromatographic separation is the quantitative extraction from the food matrix which was proven to be impeded in durum wheat. To optimize the extraction procedure, several factors with influence on extractability of carotenoids were investigated. Finally, it was shown that soaking of samples in water for 5 min prior to extraction with organic solvents had the strongest impact on extraction yield and led to the most rapid and gentle method. Contents of carotenoids in the extracts of several durum wheat and corn samples were doubled by soaking in water before extracting with methanol/tetrahydrofuran (1/1, v/v). In light of these findings, literature data on contents of carotenoids in cereal grains have to be viewed critically regarding the extraction procedures employed.