Application of supercritical CO2 for extraction of polyisoprenoid alcohols and their esters from plant tissues

In this study, a method of supercritical fluid extraction (SFE) with carbon dioxide of polyisoprenoids from plant photosynthetic tissues is described. SFE was an effective extraction method for short- and medium-chain compounds with even higher yield than that observed for the “classical extraction” method with organic solvents. Moreover, SFE-derived extracts contained lower amounts of impurities (e.g., chlorophylls) than those obtained by extraction of the same tissue with organic solvents. Elevated temperature and extended extraction time of SFE resulted in a higher rate of extraction of long-chain polyisoprenoids. Ethanol cofeeding did not increase the extraction efficiency of polyisoprenoids; instead, it increased the content of impurities in the lipid extract. Optimization of SFE time and temperature gives the opportunity of prefractionation of complex polyisoprenoid mixtures accumulated in plant tissues. Extracts obtained with application of SFE are very stable and free from organic solvents and can further be used directly in experimental diet supplementation or as starting material for preparation of semisynthetic polyisoprenoid derivatives, e.g., polyisoprenoid phosphates.     

Selective extraction and separation of oxymatrine from Sophora flavescens Ait. extract by silica-confined ionic liquid

This study highlighted the application of a two-stepped extraction method for extraction and separation of oxymatrine from Sophora flavescens Ait. extract by utilizing silica-confined ionic liquids as sorbent. The optimized silica-confined ionic liquid was firstly mixed with plant extract to adsorb oxymatrine. Simultaneously, some interference, such as matrine, was removed. The obtained suspension was then added to a cartridge for solid phase extraction. Through these two steps, target compound was adequately separated from interferences with 93.4% recovery. In comparison with traditional solid phase extraction, this method accelerates loading and reduces the use of organic solvents during washing. Moreover, the optimization of loading volume was simplified as optimization of solid/liquid ratio.     

The extraction of leukotrienes (LTC4, LTD4, and LTE4) from tissue fluids: the metabolism of these mediators during IgE-dependent hypersensitivity reactions in lung

The metabolites of arachidonic acid known as the leukotrienes are a class of lipid mediators which have potent and diverse biological effects in pulmonary tissue. Leukotrienes C, D, and E (LTC4, LTD4, and LTE4) are known to be principal mediators of immunoglobulin E (IgE)-mediated hypersensitivity reactions in lung tissue. It is therefore important to develop reliable and quantitative isolation techniques for estimating levels of these mediators in tissue. In this study, LTC4, LTD4, and LTE4 were separated from other arachidonate metabolites by organic extraction procedures. 5-Hydroxyeicosatetraeonic acid and leukotriene B4 extract efficiently into the organic layer of aqueous:ether or aqueous:chloroform extractions, whereas arachidonate metabolites containing conjugated peptides (e.g., LTC4, LTD4, and LTE4) failed to extract into these organic solvents. An extraction step was therefore developed that affords quantitative extraction of LTC4, LTD4, and LTE4 into the organic phase of an isopropanol:ether:H2O mixture. This step is the key for a two-step extraction method that isolates histamine, LTC4, LTD4, and LTE4 with a recovery of 100, 85, 75, and 57%, respectively. One advantage of this separation procedure for obtaining these mediators by organic extraction is an ability to expediently process many samples. Furthermore, the leukotriene content of extracted samples can be analyzed using the guinea pig ileum bioassay without interference from vasoamines or platelet-activating factor. These later substances are eliminated from leukotriene-enriched fractions by this extraction process. When histamine and LTC4 were added to supernatant fluids recovered from isolated lung tissue, they were quantitatively recovered using this extraction method.(ABSTRACT TRUNCATED AT 250 WORDS)     

The use of buttermilk as a source of phospholipids

A method for obtaining phospholipids from butter-milk, a by-product of butter making is described. The method consists in coagulating the fat globule membranes with mineral or organic acid, separating the resulted precipitate by centrifugation or filtering, lipid extraction by organic solvents mixture and isolation of phospholipids from the extract by acetone treatment or by column chromatography. Butter-milk, a new source of biological membranes, is characterized by a low cost and accessibility.     

Enhanced solvent extraction of polar lipids associated with rubber particles from Hevea brasiliensis

Biochemical studies of lipids bound to rubber particles have been complicated due to the solubility of polyisoprene chains in most extracting solvents and the rather delicate nature of polar lipids that are often denatured when traditional solvent extraction techniques are employed. In this paper, we describe a traditional technique and accompanying solvents that permit optimal extraction of rubber particle bound lipids. The technique, which is validated after characterizing the lipid extracts by elemental analysis, silica column adsorption and thin layer chromatography, appeared more suitable for extracting total lipids with optimal glycolipid and phospholipid contents. This technique is proposed as an alternative to traditional extraction methods used for solid natural rubber as it offers advantages with respect to ease of application, extract quality, extraction yields and reproducibility.     

A two‐step extraction method to measure fecal steroid hormones in female cynomolgus monkeys (Macaca fascicularis)

We developed a two‐step extraction method for measuring fecal steroid concentrations. In the first step, distilled water was used to extract steroids from fecal samples. In the second step, a mixture of organic solvents (hexane and ether) was used to re‐extract water extracts that had been transferred to a glass tube. A portion of the upper layer of the organic solvents was transferred to separate assay‐tubes for measurement of estradiol (E2) or progesterone (P), and the organic solvents were evaporated in vacuo. After phosphate‐buffered saline was added to each tube, commercially supplied radioimmunoassay (RIA) kits were used to determine the steroids. We demonstrated the advantages and reliability of this method by using it to assay the steroid hormone concentrations in fecal samples and serum samples collected on the same day from female cynomolgus monkeys who showed normal menstrual cycles and from monkeys who had induced hyperfunction of ovarian steroidgenesis. Different fecal samples from each monkey were used to determine the recovery rate of each steroid in water extraction from the fecal samples and the reproductivity of hormone concentrations in the fecal samples. The results demonstrate that this two‐step method is simple and effective for measuring fecal steroids for monitoring the reproductive status of cynomolgus monkeys, without having to collect serum samples. Am. J. Primatol. 48:291–298, 1999. © 1999 Wiley‐Liss, Inc.     

麝香中脂溶性成分的提取与麝香质量鉴别

采用薄层层析和气相色谱技术,比较了超声、冷浸和热回流3种提取方法及乙醇、乙醚、正己烷3种溶剂对麝香提取液中脂溶性成分及麝香酮含量的影响。结果显示用不同提取方法和溶剂提取的麝香脂溶性成分经薄层层析后均呈现6或7个斑点。而经气相色谱分析,其结果却有较大的差别,麝香乙醚提取液的色谱峰较乙醇和正己烷更丰富,超声提取的效果较冷缦和热回流好,色谱峰达到20个。麝香酮的定量分析显示冷浸法提取的麝香酮含量高于超声和热回流提取。通过薄层层析和气相色谱分析,对10个麝香样品的质量进行了鉴别。     

提取方法和溶剂对薏苡仁油提取率的影响

为了提高薏苡仁油的提取效率,考察5种不同极性的溶剂（无水乙醇、丙酮、二氯甲烷、乙酸乙酯、环己烷）在3种不同的提取方法中对薏苡仁油提取率的影响,并分析其原因。结果表明,丙酮在加热回流提取法中提取率最高（8.17%）,无水乙醇在超声辅助提取和微波辅助提取法中得率均较高。3种提取方法中,超声辅助提取法的得油率最高（10.79%）,而微波法虽然提取效率高（0.0678g/min）,但得油率太低（6.78%）,不适于薏苡仁油的提取。     

星点设计-效应面法优化杉木枝叶中双黄酮的提取工艺

采用星点设计-效应面法优化杉木枝叶中穗花杉双黄酮和金松双黄酮的提取工艺.以乙醇体积分数、料液比、提取时间为自变量,穗花杉双黄酮和金松双黄酮转移率的归一化值为因变量,通过对自变量与因变量的二次多项式拟合,采用效应面法选取较优的工艺条件,并进行预测分析.最终确定穗花杉双黄酮和金松双黄酮的最佳提取工艺为:乙醇体积分数为50％,料液比为1∶13,超声提取3次,每次提取60 min,最佳工艺验证结果与模型预测值相差-2.55％.结果表明:星点设计-效应面法优选的穗花杉双黄酮和金松双黄酮的提取工艺,方法简便、可靠.     

川金丝猴粪尿中类固醇性激素抽提方法比较

本文旨在探讨川金丝猴粪尿中类固醇性激素的抽提方法, 实验比较了3 种尿液抽提溶剂和5 种粪便抽提方法的抽提回收率。结果显示, 以二氯甲烷为溶剂抽提雄猴尿液中的睾酮、雌猴尿液中的雌二醇和孕酮, 回收率分别为79.27%、73.39%和80.28%, 抽提效果较其它两种溶剂为好; 以乙醇加热法抽提雄猴粪便中的睾酮和雌二醇、雌猴粪便中的睾酮、雌二醇和孕酮, 回收率分别为1.33%、85.12%与67.41%、79.58%和77.71%, 抽提效果优于其它4 种方法,且该方法简便易于操作。实验表明, 以二氯甲烷和乙醇加热法分别抽提川金丝猴尿液和粪便中的类固醇性激素是较理想的抽提方法。     

基于天然低共熔溶剂的甜叶菊中甜菊糖绿色提取方法及优化

尝试利用天然低共熔溶剂(NADES)提取甜叶菊(Stevia rebaudiana)中的甜菊糖, 探索一种高效、绿色和环保的甜菊糖提取新方法。以甜叶菊干叶为原料, 对照传统提取溶剂水, 以甜菊糖中甜菊苷和莱鲍迪苷A的提取浓度作为指标, 筛选出最优的NADES提取配方, 然后通过Box-Behnken响应面法对NADES提取甜叶菊中甜菊糖的工艺条件进行筛选优化。结果表明, 提取效率最高的NADES配方为1,2-丙二醇:甘油:水=8:1:1 (v/v/v), 提取的甜菊苷浓度为2.59 mg∙mL^-1, 比水提取高16.40%, 提取的莱鲍迪苷A浓度为1.06 mg∙mL^-1, 比水提取高12.62%; 通过响应面法得到最优提取条件: 提取时间90分钟, 提取温度60°C, 超声功率为80 J∙s^-1, 预测甜菊苷提取浓度为3.49 mg∙mL^-1, 莱鲍迪苷A提取浓度为1.43 mg∙mL^-1, 与实验验证值(甜菊苷浓度为3.48 mg∙mL^-1, 莱鲍迪苷A浓度为1.42 mg∙mL^-1)接近。在最优条件下, 甜菊苷提取浓度比初始条件提高了34.36%, 莱鲍迪甘A提取浓度比初始条件提高了33.96%。NADES绿色环保, 且提取效率高于传统溶剂, 可用于甜叶菊中甜菊糖的绿色提取; 同时, 该提取方法可为后续推广至其它大宗经济植物类天然产物的绿色工业生产提供参考。     

Integrated and convenient procedure for protein extraction from formalin‐fixed,paraffin‐embedded tissues for LC‐MS/MS analysis

Because fresh‐frozen tissue samples associated with long‐term clinical data and of rare diseases are often unobtainable at the present time, formalin‐fixed paraffin‐embedded (FFPE) tissue samples are considered a highly valuable resource for researchers. However, protein extraction from FFPE tissues faces challenges of deparaffinization and cross‐link reversion. Current procedures for protein extraction from FFPE tissue require separate steps and toxic solvents, resulting in inconvenience in protein extraction. To overcome these limitations, an integrated method was developed using nontoxic solvents in four types of FFPE tissues. The average amount of proteins from three replicates of bladder, kidney, liver, and lung FFPE tissues were 442.6, 728.9, 736.4, and 694.7 μg with CVs of 7.5, 5.8, 2.4, and 4.5%, respectively. Proteomic analysis showed that 348, 417, 607, and 304 unique proteins were identified and quantified without specification of isoform by a least two peptides from bladder, kidney, liver, and lung FFPE tissue samples, respectively. The analysis of individual protein CV demonstrated that 97–99% of the proteins were quantified with a CV ≤ 30%, verifying the reproducibility of the integrated protein extraction method. In summary, the developed method is high‐yield, reproducible, convenient, simple, low cost, nonvolatile, nonflammable, and nontoxic.     

萱草花瓣中类胡萝卜素分析样品的制备及UPCC-MS检测方法研究

以大苞萱草（Hemerocallis middendorfii Trautv.et Mey.）和‘原谅’萱草（H.‘Pardon Me’）为研究对象,对萱草属植物花瓣中类胡萝卜素的样品制备方法以及UPCC-MS定性和定量检测方法进行了研究。结果表明：（1）萱草花瓣类胡萝卜素样品制备过程中,不同的提取试剂、振荡方法及皂化方法对类胡萝卜素的提取效率均有显著影响,经过对提取结果的方差分析,确定最佳的样品制备方案为：提取试剂B丙酮：正己烷（3：5/V：V）、温控摇床振荡提取,常温皂化16 h;（2） UPCC-MS技术能在10 min内高效分离萱草花瓣中的类胡萝卜素,且使用的有毒化学试剂少,是检测类胡萝卜素的较好选择;（3）大苞萱草和原谅萱草花瓣中共含有20种类胡萝卜素物质,两者颜色不同,类胡萝卜素的组成和含量也存在差异。     

Influence of the extraction solvent on antioxidant activity of Althaea officinalis L. root extracts

Althaea officinalis (Malvaceae) is a well-known plant that is widely distributed throughout the world. Aqueous and hydroalcoholic extracts from A. officinalis root are used mainly because of their antitussive and expectorant activity. It is well known that these activities are based on the polysaccharide composition, but little is known about the possible antioxidant activity of root extract. The present study evaluated antioxidant activity of root extracts prepared with different extraction solvents applying ABTS^·+ (2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid), hypochlorous acid scavenging assay and iron-induced lipid peroxidation. The results showed that the extract prepared with water as extraction solvent did not possess antioxidant activity, whereas the extracts obtained using ethanol:water as extraction agent showed well pronounced antioxidant activity. In particular, the extracts obtained at low concentration of ethanol in the mixed solvent (50:50 and 70:30, v/v) showed higher scavenging activity for ABTS^·+ radicals and hypochlorite ions than the extract obtained with the higher ethanol concentration (90:10, v/v). These results correlated very well with phenolic and flavonoid content of the extracts. The extracts did not show cytotoxic effect on human BV-173 leukemic cells but may have immunomodulating effects due to their antioxidant properties.     

Preparation of the particles of Acanthopanax senticosus

The main objective of this study is to establish the best preparation technology of the particles of Acanthopanax senticosus. First, take the reflux extraction method extract of Acanthopanax senticosus coarse powder, optimized by orthogonal experimental method, to flavonoids flavonoids extraction extraction rate as the indexes to determine the effects of extraction temperature, ethanol concentration, extraction time on flavonoids content. Then by a wet granulation of thorn slender acanthopanax particles, taste with granules, forming rate, melting rate as index to investigate the influences of materials adding amount of granules effect. The results showed that the ethanol water heating reflux extraction method to extract the temperature of 70 deg, the percentage of ethanol 75%, extraction time 2.5 h, the highest content of total flavonoids in the extract. Join the 5 ml and 10 g in the extract of acacia honey, dextrin, starch, sugar ratio for 3:4:8, the best taste of Acanthopanax granules. In the end, the best preparation technology of the granules is established, and the process is simple, which is suitable for the large-scale production of the factory.     

Optimization of micellar extraction for the pre-purification of paclitaxel from<Emphasis Type="Italic">Taxus chinensis</Emphasis>

Currently, there are many reports in the literature regarding technological methods for paclitaxel purification. However, there have been few reports on the purification of paclitaxel using a micellar system. This method is based on the transfer of paclitaxel within the crude extract to an aqueous surfactant solution as a micelle, allowing the use of organic solvents to be used for the removal of lipids and non-polar impurities. In this study, we optimized the important process parameters of micellar extraction to obtain a high purity and yield of paclitaxel in a pre-purification step. The optimal surfactant (N-cetylpyridinium chloride, CPC) concentration, initial crude paclitaxel concentration, organic solvents (methylt-butyl ether/hexane) ratio, extraction temperature, and extraction time were 7.5% (w/v), 16.4 mg/mL, 1.5/1 (v/v), 25°C, and 30 min, respectively. The crude extracts from the liquid-liquid extracts were efficiently pre-purified by micellar extraction, increasing in purity from 6% to over 21%, with a yield of 92%. Overall, the use of micellar extraction in the pre-purification process allowed for rapid and efficient separation of paclitaxel from interfering compounds, and dramatically increased the yield and purity of the crucle paclitael for subsequent purification steps.     

Aqueous extraction of gibberellins from pea

Summary Aqueous buffers were used to extract gibberellin-like substances from pea tissue. The method possesses several distinct advantages when compared with extraction methods using organic solvents. Aqueous buffer extracts can be prepared more rapidly and produce extracts which are free of pigments and other alcohol soluble materials.Extraction of pea with aqueous buffer has indicated the presence of two gibbrellin-like substances in addition to those previously described.Work was supported by National Science Foundation Grant No. GB-5863     

Review of methods used in lunar organic analysis: Extraction and hydrolysis techniques

Extraction, hydrolysis and crushing procedures have proven useful in analyzing for some of the carbon-containing compounds which are present in Apollo 11 and 12 lunar samples. Three main extraction methods employed with aqueous and nonaqueous solvents were refluxing in open and closed systems, Soxhlet extraction, and sonication. With water and acids, refluxing was the method of choice. Of the various nonaqueous solvent systems used, benzene: methanol mixtures were most often selected, and sonication was favored over Soxhlet extraction. Extraction of lunar samples with water followed by acid hydrolysis of the water extract was found to be superior to direct acid hydrolysis of lunar material in the search for amino acids or their precursors. Direct acid hydrolysis of lunar materials did demonstrate however, that carbides or carbide-like compounds are present on the moon. Hydrolysis with deuterated acids and bases showed that lunar samples contain indigenous methane and ethane and confirmed the presence of carbide-like materials. Crushing experiments also showed that gaseous hydrocarbons can be released from lunar samples.     

Purification of dog VIP from a single animal

VIP, a potent vasodilator peptide, is reported to be identical in pig, cow, human and rat but to differ in four amino acids in chicken. This report describes the purification of dog VIP from the small intestine of a single animal. The purification method is based on tissue extraction with a sequence of organic solvents. The extracted VIP is concentrated onto cation-exchange cellulose and brought to purity by three HPLC steps. A 30% final yield of pure VIP was obtained from the original extract. Dog VIP was found to have the following sequence: His-Ser-Asp-Ala-Val-Phe-Thr-Asp-Asn-Tyr-Thr-Arg-Leu-Arg-Lys-Gln-Met-Ala -Val-Lys-Lys-Tyr-Leu-Asn-Ser-Ile-Leu-Asn. Thus the amino acid sequence of dog VIP is identical with all the mammalian VIP's which have been reported. This suggests that a high degree of conservation throughout the molecule may be required for VIP bioactivity.     

Identification of a Pea Component Stimulatory for Heat-Stressed Putrefactive Anaerobe 59-123 Spores

Pea extract contains a factor which improves recovery counts of heat-stressed putrefactive anaerobe spores in a complex medium up to threefold. The factor is heat-stable and nondialyzable. Most of the active principle is found in the precipitate which forms during storage of pea extract at 4 C. The precipitate disperses upon heating, is high in starch content, and retains activity after extraction with organic solvents and water. Treatment of pea extract with alpha-amylase results in complete destruction of the active principle. These observations indicate that starch is the factor in pea extract responsible for increased recovery counts of heat-stressed putrefactive anaerobe spores.