无花果果皮中花青素的提取工艺的建立

本研究使用单因素方法考察了无花果(Ficus carica L.)果皮中花青素的最佳提取条件,并考察了7种参数对花青素提取率的影响。参数设置如下:溶剂性质(水,甲醇,乙醇和丙酮)、提取次数(1~3次)、固液比(1/50,1/100,1/150和1/200)、提取时间(60 min,120 min,180 min和240 min)、甲醇浓度(0,20%,40%,60%,80%和100%)、酸类型(盐酸,乙酸,柠檬酸和酒石酸)和酸浓度(0,1%,2%,5%和10%)。使用pH-示差法测量无花果果皮中单体花色素的含量。研究显示,无花果果皮中花青素的最佳提取条件为:溶剂为甲醇溶剂,提取次数为2次,固液比为1/100,提取时间为180 min,甲醇浓度为80%,酸类型为柠檬酸,柠檬酸浓度为5%。该最佳提取条件下的花青素的提取率达到最高(345.62 mg/100g DS)。     

EVALUATION OF PIGMENT EXTRACTION METHODS AND A RECOMMENDED PROTOCOL FOR PERIPHYTON CHLOROPHYLL a DETERMINATION AND CHEMOTAXONOMIC ASSESSMENT1

Many methods have been proposed to extract and quantify algal pigments. Comparative studies have found that pigment extraction efficiency varies among solvent and mechanical disruption protocols due to differential cellular resistance, thereby, leading to potential misinterpretation of pigment data. When the type or resistance of algae are unknown, a method is required that efficiently extract pigments from all taxonomic groups. The objective of this study was to develop a simple and efficient one stage periphyton pigment extraction protocol by comparing the extractability of four solvents (acetone, methanol, methanol/acetone, and methanol/acetone/N,N‐dimethylformamide), the effects of grinding, and the effects of freeze‐drying. The best overall extraction was obtained using freeze‐dried samples extracted with methanol/acetone/DMF/water (MAD). Eighty‐six percent more chlorophyll was extracted when the sample was freeze‐dried relative to fresh/frozen samples extracted with 90% acetone. Freeze‐drying greatly improved the extraction of both polar and non‐polar (lipophilic/hydrophobic) pigments while MAD increased the extractability of polar pigments and improved peak resolution of all pigments. Chemotaxonomic assessment differed between samples that were fresh/frozen or freeze‐dried before extraction. The relative abundance of cyanobacteria was greater for freeze‐dried material compared with fresh/frozen due to the improved extractability of cyanobacterial pigments. Based on the results of this study, the traditional approach of 90% acetone as a solvent is not recommended for periphyton samples containing cyanobacteria or when the composition of the mat is unknown. The combination of freeze‐drying and MAD was sufficient for the extraction of pigments from a periphyton mat containing filamentous cyanobacteria, green algae, and diatoms.     

Rapid methods to determine procyanidins, anthocyanins, theobromine and caffeine in rat tissues by liquid chromatography-tandem mass spectrometry

Rapid, selective and sensitive methods were developed and validated to determine procyanidins, anthocyanins and alkaloids in different biological tissues, such as liver, brain, the aorta vein and adipose tissue. For this purpose, standards of procyanidins (catechin, epicatechin, and dimer B(2)), anthocyanins (cyanidin-3-glucoside and malvidin-3-glucoside) and alkaloids (theobromine, caffeine and theophylline) were used. The methods included the extraction of homogenized tissues by off-line liquid-solid extraction, and then solid-phase extraction to analyze alkaloids, or microelution solid-phase extraction plate for the analysis of procyanidins and anthocyanins. The eluted extracts were then analyzed by ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry, using a triple quadrupole as the analyzer. The optimum extraction solution was water/methanol/phosphoric acid 4% (94/4.5/1.5, v/v/v). The extraction recoveries were higher than 81% for all the studied compounds in all the tissues, except the anthocyanins, which were between 50 and 65% in the liver and brain. In order to show the applicability of the developed methods, different rat tissues were analyzed to determine the procyanidins, anthocyanins and alkaloids and their generated metabolites. The rats had previously consumed 1g of a grape pomace extract (to analyze procyanidins and anthocyanins) or a cocoa extract (to analyze alkaloids) per kilogram of body weight. Different tissues were extracted 4h after administration of the respective extracts. The analysis of the metabolites revealed a hepatic metabolism of procyanidins. The liver was the tissue which produced a greater accumulation of these metabolites.     

Adsorption of polycyclic aromatic hydrocarbons (PAHs) on Rhizopus oryzae cell walls: application of cosolvent models for validating the cell wall-water partition coefficient

The cell wall-cosolvent partition coefficients (Km) of polycyclic aromatic hydrocarbons (PAHs) were determined for Rhizopus oryzae cell walls by controlling the volume fraction of methanol (f) ranging from 0.1 to 0.5. Five cosolvent models were employed for extrapolating the cell wall-water partition coefficients (Kw) in pure water. The extrapolated Kw values of four PAHs on R. oryzae cell walls were ranged from 2.9 to 5.1. Comparison of various Kw values of pyrene generated from extrapolation and the QSPR model, together with predicted different (PD), mean percentage deviations (MPD), and root mean square errors (RSE), revealed that the performance of the LL and Bayesian models were the best among all five tested cosolvent models. This study suggests that R. oryzae cell walls play an important role in the partitioning of PAHs during bioremediation because of the high Kw of fungal cell walls.     

Isolation of vindoline from Catharanthus roseus by supercritical fluid extraction.

Vindoline was extracted from the leaves of Catharanthus roseus over the ranges of 35-70 degrees C and 100-300 bar using supercritical carbon dioxide with and without the addition of 3 wt % ethanol as a cosolvent. The vindoline contents in the extracts were determined by HPLC and identified by LC/MS. The remarkable highest vindoline concentration, 58 wt %, was obtained at the lowest temperature, 35 degrees C, and the highest pressure, 300 bar, of this study. The use of a cosolvent only slightly improved the extraction yields or selectivities at some experimental conditions.     

Evaluation of green solvents: Oil extraction from oleaginous yeast Lipomyces starkeyi using cyclopentyl methyl ether (CPME)

Cyclopentyl methyl ether (CPME) was evaluated for extracting oil or triacylglycerol (TAG) from wet cells of the oleaginous yeast Lipomyces starkeyi. CPME is a greener alternative to chloroform as a potential solvent for oil recovery. A monophasic system of CPME and biphasic system of CPME:water (1:0.7) performed poorly having the lowest TAG extraction efficiency and TAG selectivity compared to other monophasic systems of hexane and chloroform and the biphasic Bligh and Dyer method (chloroform:methanol:water). Biphasic systems of CPME:water:alcohol (methanol/ethanol/1‐propanol) were tested and methanol achieved the best oil extraction efficiency compared to ethanol and 1‐propanol. Different biphasic systems of CPME:methanol:water were tested, the best TAG extraction efficiency and TAG selectivity achieved was 9.9 mg/mL and 64.6%, respectively, using a starting ratio of 1:1.7:0.6 and a final ratio of 1:1:0.8 (CPME:methanol:water). Similar results were achieved for the Bligh and Dyer method (TAG extraction efficiency of 10.2 mg/mL and TAG selectivity of 66.0%) indicating that the biphasic CPME system was comparable. The fatty acid profile remained constant across all the solvent systems tested indicating that choice of solvent was not specific for any certain fatty acid. This study was able to demonstrate that CPME could be used as an alternative solvent for the extraction of oil from the wet biomass of oleaginous yeast. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1096–1103, 2017     

A novel procedure for the isolation of glycolipids from Bifidobacterium adolescentis 94 BIM using supercritical carbon dioxide

This study describes a novel isolation procedure for major glycolipids from Bifidobacterium adolescentis 94 BIM. The procedure consists of the use of supercritical carbon dioxide (scCO(2)) with hydro-methanolic solution as co-solvent. The major glycolipids were isolated using the following operating conditions: pressure, 30 MPa, co-solvent concentration, 10% (9:1, methanol/water, v/v), CO(2) flow rate, 5 g/min, extraction time and temperature, 2h and 55 degrees C, respectively. The reference glycolipids sample was prepared by classical organic solvent extraction followed by chromatographic purification. All isolates were characterized by TLC and the major glycolipids additionally by enzyme linked immunosorbent (ELISA). Sixty milligrams of glycolipids with similar immunoreactivity as the reference glycolipids were isolated from 1g of freeze-dried biomass (6% of yield).     

Comparison of Soil VOCs measured by soil gas,heated headspace,and methanol extraction techniques

Comparisons of soil volatile organic compound (VOC) measurement techniques and soil properties expected to influence these measurements were performed at two dissimilar sites. A total of 41 soil gas, 52 heated headspace, and 51 methanol extraction/purge‐and‐trap measurements were obtained on collocated samples. Contaminants present at both sites included cis‐1,2‐dichloroethene, 1, 1, 1‐trichloroethane, trichloroethene, and tetrachloroethene. Heated headspace offered the highest sensitivity, as indicated by the greatest percentage of detections per number of analyses. The statistical regression between headspace concentrations and methanol extraction concentrations was highly significant (p < 0.001) with r² = 0.53. Headspace concentrations (range, 7 to 4250 ng/g) ran approximately 20 to 30% of the methanol extraction concentrations (range, 260 to 7300 ng/g), indicating that the methanol was able to extract significantly more of the chlorinated hydrocarbons (CHCs) than the headspace extraction, even in soils with relatively low organic carbon contents (< 0.25%). None of the soil properties (gravimetric moisture content, organic carbon content, percent sand, and percent clay) significantly improved the regression fit. The soil gas responses were unlike either headspace or methanol extraction data. CHC measurements by vapor extraction/soil gas could not be used to predict soil CHC concentrations at these sites.     

Effects of Cosolvent and Temperature on the Desorption of Polynuclear Aromatic Hydrocarbons from Contaminated Sediments of Chien-Jen River,Taiwan

This study evaluated the effects of the water-miscible cosolvent and temperature on the sorption-desorption of polynuclear aromatic hydrocarbons (PAHs) from contaminated sediments in Chien-Jen River, Taiwan. Sediment samples from five sampling stations of downstream section were utilized in this study. Phenanthrene and anthracene were selected as target compounds. The cosolvent effect on sorption of phenanthrene and anthracene was examined by the addition of various volume fractions of methanol (i.e., 0.3, 0.5, 0.7, and 0.9, respectively) in the sediment/water systems. The utility of the log-linear cosolvency model for predicting PAH sorption from solvent mixtures was evaluated. An inverse relationship was observed for sorption coefficients of phenanthrene and anthracene as a function of increasing cosolvent. The effect of temperature on sorption of phenanthrene and anthracene was conducted at temperature from 10°C to 40°C. The use of elevated temperatures in desorption experiments increased the PAH release from sediments. It was observed that sorption of phenanthrene and anthracene onto sediments decreased when temperature increased. The decrease of sorption coefficient of phenanthrene was more sensitive than that of anthracene. The magnitude of decreased sorption was attributed by the increased desorption rate constant, solubility, and heterogeneities of sediments.     

A Spectrophotometric Analysis of Pigments in Apples

Methods of pigment extraction using traditional polar organic solvents (acetone or methanol) were compared to those employing a chloroform–methanol mixture. We found that, for spectrophotometric pigment analysis in the apple peel, the cuticular lipids must be preliminarily extracted from the samples with chloroform and MgO must be added during homogenization to prevent pigment degradation. The traditional extraction did not result in the complete extraction of intact pigments, and such extracts contained a considerable amount of light-absorbing impurities. The application of chloroform–methanol extraction allowed us to markedly reduce the content of such impurities and to increase the accuracy and sensitivity of the measurement of the content of chlorophylls and carotenoids. In addition, this extraction method proved useful for the analysis of phenolic substances (anthocyanins and flavonoids) in the water-methanol fraction of the extracts.     

新疆棕色棉纤维中花色苷的提取及初步鉴定

以新疆深棕色棉品系‘棕86’的成熟纤维为材料,用甲醇溶液(VMeOH∶VHCl=99∶1)避光冷浸提取15d后,浓缩成浸膏.用适量蒸馏水溶解浸膏,乙酸乙酯萃取除杂,水相用HP-20大孔树脂纯化色素组分;采用紫外-可见吸收光谱分析,并结合高效液相色谱方法,对天然彩色棉纤维色素组分进行鉴定,为彩色棉花新色彩的分子改良提供参考.结果显示,棕色棉纤维提取液中含有花色苷,且至少含有4种不同的花色苷成分.对棕色棉纤维提取液中花色苷的降解分析显示,棕色棉中的色素物质很不稳定.研究表明,花色苷是棕色棉纤维色素中的一种重要组成成分,进一步补充了彩棉色素的物质基础.     

Comparison of steam and ammonia pretreatment for enzymatic hydrolysis of cellulose

Summary Aspenwood, wheat straw, wheat chaff and alfalfa stems were treated under pressure with either steam or ammonia. The material was then water or methanol/water extracted. The extent of enzymatic hydrolysis of the cellulose portion of the treated substrates was compared using two different cellulases, a commercial preparation, Celluclast, and those from the fungus Trichoderma harzianum. Both steam and ammonia treatment enhanced the accessibility of the cellulose as measured by hydrolysis. Methanol extraction of steamed material generally reduced the access of the enzyme to the cellulose, whereas methanol extraction of ammonia-treated material increased accessibility. The optimum combinations of pretreatment and extraction method depended on the substrate and on the enzyme system; no treatment suitable for all situations could be selected.     

Supercritical carbon dioxide extraction of lipids from Eucalyptus globulus wood

Various typical lipid components of wood extractives have been isolated from Eucalyptus globulus wood by supercritical carbon dioxide modified with methanol. The influence of various extraction parameters on the yield and qualitative composition of the extracts have been studied. The extracts were analyzed by gas chromatography-mass spectrometry and compared with those obtained by Soxhlet extraction with acetone, the standard method for the determination of wood extractives. The qualitative and quantitative results obtained by both methods were in good agreement. The experimental planning to asses the influence of pressure, temperature and percentage of methanol and their interactions on the extraction efficiency was carried out with a factorial design, followed by multiple linear regression algorithm.     

Biodiesel production using Aspergillus niger as a whole‐cell biocatalyst in a packed‐bed reactor

Enzymatic lipase transesterification of palm oil to biodiesel in a packed‐bed reactor (PBR) using a novel strain of the fungus Aspergillus niger, immobilized within polyurethane biomass support particles (BSPs), was investigated. A three‐step addition of methanol was used to reduce lipase inhibition by immiscible methanol. The influence of water content and PBR flow rate was investigated. FAME yield was enhanced with an increase of PBR flow rate in the range of 0.15–30 L h^?1, where inefficient mixing of the reaction mixture at lower flow rates resulted in low conversion rates i.e. 69% after 72‐h reaction. Adding the third mole equivalent of methanol resulted in lipase inhibition due to methanol migration into the accumulated glycerol layer. Glutaraldehyde (GA) solution (0.5 vol.%) was used to stabilize lipase activity, which led to a high FAME yield (>90%) in the PBR after 72‐h of reaction time at a flow rate of 15 L h^?1, and a water content of 15%. Moreover, a high conversion rate (>85%) was maintained after four palm oil batch conversion cycles in the PBR. In contrast, lipase activity of non‐GA‐treated cells decreased with each PBR batch cycle, where only 70% FAME was produced after the forth PBR cycle. Transesterification of palm oil in a PBR system using BSPs‐immobilized A. niger as a whole‐cell biocatalyst is a viable process for enzymatic biodiesel production.     

Molecular dynamics simulations of the native and partially folded states of ubiquitin: influence of methanol cosolvent, pH, and temperature on the protein structure and dynamics

A series of explicit-solvent molecular dynamics simulations of the protein ubiquitin are reported, which investigate the effect of environmental factors (presence of methanol cosolvent in the aqueous solution, neutral or low pH value, room or elevated temperature) on the structure, stability, and dynamics of the protein. The simulations are initiated either from the native structure of the protein or from a model of a partially folded state (A-state) that is known to exist at low pH in methanol-water mixtures. The main results of the simulations are: (1) The ubiquitin native structure is remarkably stable at neutral pH in water; (2) the addition of the methanol cosolvent enhances the stability of the secondary structure but weakens tertiary interactions within the protein; (3) this influence of methanol on the protein structure is enhanced at low pH, while the effect of lowering the pH in pure water is limited; and (4) the A-state of ubiquitin can be described as a set of relatively rigid secondary structure elements (a native-like beta-sheet and native-like alpha-helix plus two nonnative alpha-helices) connected by flexible linkers.     

Improving Product Specificity of Whole‐Cell Alkane Oxidation in Nonconventional Media: A Multivariate Analysis Approach

Two‐liquid‐phase reaction media have long been used in bioconversions to supply or remove hydrophobic organic reaction substrates and products to reduce inhibitory and toxic effects on biocatalysts. In case of the terminal oxyfunctionalization of linear alkanes by the AlkBGT monooxygenase the excess alkane substrate is often used as a second phase to extract the alcohol, aldehyde, and acid products. However, the selection of other carrier phases or surfactants is complex due to a large number of parameters that are involved, such as biocompatibility, substrate bioavailability, and product extraction selectivity. This study combines systematic high‐throughput screening with chemometrics to correlate physicochemical parameters of a range of cosolvents to product specificity and yield using a multivariate regression model. Partial least‐squares regression shows that the defining factor for product specificity is the solubility properties of the reaction substrate and product in the cosolvent, as measured by Hansen solubility parameters. Thus the polarity of cosolvents determines the accumulation of either alcohol or acid products. Whereas usually the acid product accumulates during the reaction, by choosing a more polar cosolvent the 1‐alcohol product can be accumulated. Especially with Tergitol as a cosolvent, a 3.2‐fold improvement in the 1‐octanol yield to 18.3 mmol L^?1 is achieved relative to the control reaction without cosolvents.     

The effect of cosolvents on the formulation of nanoparticles from low-molecular-weight poly(l)lactide

The aim of this study was to formulate nanoparticles from poly(l)lactide by a modified nanoprecipitation method. The main focus was to study the effect of cosolvent selection on the shape, size, formation efficiency, degree of crystallinity, x-ray diffraction (XRD) reflection pattern, and zeta potential value of the particles. Low-molecular-weight (2000 g/mol) poly(l)lactide was used as a polymer, and sodium cromoglycate was used as a drug. Acetone, ethanol, and methanol were selected as cosolvents. Optimal nanoparticles were achieved with ethanol as a cosolvent, and the formation efficiency of the particles was also higher with ethanol as compared with acetone or methanol. The particles formulated by ethanol and acetone appeared round and smooth, while with methanol they were slightly angular. When the volume of the inner phase was decreased during the nanoprecipitation process, the mean particle size was also decreased with all the solvents, but the particles were more prone to aggregate. The XRD reflection pattern and the degree of crystallinity were more dependent on the amount of the solvents in the inner phase than on the properties of the individual cosolvents. The zeta potential values of all the particle batches were slightly negative, which partially explains the increased tendency toward particle aggregation.     

Effects of solvent on chiral and enantiomeric separation of Koga bases using derivatized amylose chiral stationary phase

The separation of R,R‐, S,S‐, and meso‐Koga bases on derivatized amylose chiral stationary phases (CSP) has been studied using different alcohol and alcohol‐hexane mixtures as eluant. Straight‐chain and branched alcohols with carbon numbers from one to four were investigated. The carbon number and geometry of the alcohol impacts the separation of Koga bases. The optimal separations were obtained using a mixture of methanol with linear or branched alcohol. Also, the elution order of meso‐ and R,R‐Koga base was switched as content of branched alcohol increases in cosolvent. The study of acidic and basic additive effects demonstrated that maintaining analytes in the free base state is crucial in order to achieve retention and separation. TEA alone or TEA and TFA mixture were used in the studies. Chirality 2010. © 2009 Wiley‐Liss, Inc.     

栎属7种植物种子的发芽抑制物质研究

运用系统溶剂法和生物测定法,以栎属7种植物种子为材料,研究了其种皮、胚(胚及周围部分子叶)和子叶(远离胚端2/3子叶)1.0 g/mL和0.5 g/mL浓度甲醇等浸提液以及各有机相对白菜种子萌发的影响。结果表明:栓皮栎、锐齿栎、蒙古栎、沼生栎和麻栎甲醇浸提液均能显著降低白菜种子发芽率、根长和苗高,且对发芽率的抑制作用逐渐降低;夏栎和房山栎甲醇浸提液对白菜种子发芽率没有显著影响,但对根长和苗高有一定的抑制作用;种子不同部位甲醇浸提液的抑制作用表现为胚>子叶>种皮,且甲醇高浓度浸提液的抑制作用大于低浓度的抑制作用;甲醇相对白菜种子发芽率、苗高或根长的抑制作用最强,其次是乙酸乙酯相,其它溶剂相萃取液的抑制作用不明显。栎属种子甲醇浸提液及各有机相对白菜种子苗高和根长的抑制作用强于对发芽率的影响,说明栎属种子中所含抑制物质主要是限制自身根和芽的生长,可能是造成延迟萌发和出苗不整齐的原因。     

Effect of hexanol as a cosolvent on partitioning and mass transfer rate of protein extraction using reversed micelles of CB-modified lecithin

Using the affinity-based reversed micelles composed of Cibacron Blue F3G-A (CB) modified soybean lecithin, the effect of hexanol as a cosolvent on the extraction of lysozyme and bovine serum albumin (BSA) was investigated. The water concentration in the reversed micelles increased significantly with increasing hexanol concentration. The partition coefficient of lysozyme could be increased by over 12-fold by introducing hexanol of higher than 0.5 vol%. However, the transfer of BSA was hardly affected because its high molecular weight resulted in a strong steric hindrance effect. The enzymatic activity of lysozyme was nearly 100% retained after undergoing the extraction process with the CB–lecithin micelles containing 3 vol% hexanol. The partitioning isotherms of lysozyme in the CB–lecithin micelles with and without hexanol addition were expressed by the Langmuir equation. The partitioning capacity of lysozyme was nearly increased twofold by introducing 3 vol% hexanol to the CB–lecithin micelles and reached 2.12 g/l. The cosolvent hexanol revealed insignificant effect on the mass transfer rate, and in both the systems with and without hexanol, the mass transfer rate in back extraction was 5–10 times slower than that in the forward extraction. This phenomenon was similar to that in conventionally employed ionic surfactant systems. The result suggests that in the present affinity-based reversed micelles, the interfacial resistance also played a more important role in back extraction than in forward extraction.