Optimization of fatty acid extraction from Phaeodactylum tricornutum UTEX 640 biomass

Fatty acids in the microalga Phaeodactylum tricornutum were isolated using an optimized three-step method: extraction of crude fatty acid potassium salts made by direct saponification of lipids in the microalgal biomass with KOH/ethanol (96%, vol/vol), separation of unsaponifiable lipids by extraction with hexane, and final purification of fatty acids by acidification of the alcoholic solution of potassium soaps followed by extraction of fatty acid into hexane. Direct saponification was carried out in ethanol (96%, vol/vol) using 2.09 mL ethanol (96%) per gram of wet biomass (10 mL/g of dry biomass) mixed with 0.4 g KOH/g of biomass. Under these conditions the fatty acid yield was 87%. The optimal water content of the alcoholic solution for extraction of the unsapononifiables was established as 40%, w/w. Data on equilibrium carotenoid distribution between the alcoholic (40%, w/w water) and hexane phases were determined. These data allow prediction of the carotenoid yields with different volumes of hexane in several extraction steps. The optimal pH of the alcoholic solution before extracting the purified fatty acid was established as pH 6, and the equilibrium fatty acid distribution between the alcoholic and hexane phases was determined. This optimized method permited a 20% reduction in the production costs of highly purified eicosapentaenoic acid (EPA) in the three-step preparative process (extraction of fatty acid, concentration of polyunsaturated fatty acids by the urea method, and EPA fractionation through preparative high-performance liquid chromatography) previously developed by the authors.     

Supercritical carbon dioxide extraction of cottonseed with co-solvents

Extraction of cottonseed lipids with supercritical carbon dioxide (SC-CO₂) was conducted with and without a cosolvent, ethanol or 2-propanol (IPA). At 7000 psi and 80°C, the reduced pressure, temperature and density of SC-CO₂ was at 6.5, 1.17 and 1.85, respectively; the specific gravity was 0.87. Under these conditions, CO₂ is denser than most liquid extraction agents such as hexane, ethanol and IPA. The extraction of cottonseed with SC-CO₂ gave a yield of more than 30% (moisture-free basis). This is comparable to yields obtained by the more commonly used solvent, hexane. The crude cottonseed oil extracted by SC-CO₂ was visually lighter than refined cottonseed oil. This was substantiated by colorimetric measurements. No gossypol was detected in the crude oil. However, crude oil extracted by SC-CO₂, to which less than 5% of ethanol or IPA as co-solvent was added, containedca. 200 ppm of gossypol, resulting in the typical dark color of cottonseed crude oil with gossypol. CO₂ extracted a small amount of cottonseed phosphatides, about one-third of that extracted by pure ethanol, IPA or hexane. A second extraction with 100% ethanol or IPA after the initial SC-CO₂ extraction produced a water-soluble lipid fraction that contained a significant amount of gossypol, ranging between 1500 and 5000 ppm. Because pure gossypol is practically insoluble in water, this fraction is believed to be made up of gossypol complexed with polysaccharides and phosphatides. Partially presented at the AOCS 1993 Annual Meeting & Expo in Anaheim, California.     

营养元素对三角褐指藻生长和脂类积累的影响

研究了不同营养元素(氮、磷、硅、铁)在不同培养浓度条件下,对三角褐指藻生长以及细胞脂类积累的影响作用。结果显示:藻细胞生长随氮含量的增加而增加,但超过一定浓度后作用不明显,最适合三角褐指藻生长的氮含量为 1.76 mmol/L。缺氮和低氮(0.22~0.44 mmol/L)培养能够明显提高藻细胞脂类含量。缺磷培养能有效促进藻株脂类积累,但是藻细胞生长受到明显抑制。缺硅导致藻细胞的生长减缓,但是对藻细胞的油脂积累影响不显著。在缺铁和高铁(0.048 mmol/L)条件下,藻的总脂含量均高于正常的培养条件。实验证明通过两步培养的方法,先在营养充足的培养基中进行藻细胞生长,然后将藻细胞转移到海水中培养,可以明显提高藻细胞的油脂含量。最终本实验确定海水为诱导三角褐指藻脂类积累的最佳诱导培养基。     

Hexane elimination from soybean oil by continuous packed tower processing with supercritical CO2

Hexane elimination is the most energy-consuming step in the industrial extraction of soybean oil. It utilizes three sets of equipment: two evaporation stages in series followed by a stripper at a pressure of about 0.07 bar. The final hexane residue in the oil is about 1000 ppm. We propose an alternative to the present process for hexane elimination, based on the extraction of the soybean oil/hexane mixture with supercritical CO₂ in a continuous countercurrent packed tower. In this work, we tested a soybean oil/hexane mixture feed containing 10% by weight of hexane. Various pressures and temperatures of the column were tested to reduce hexane residue in the oil. The extraction process was demonstrated to be very effective for hexane separation. Indeed, at the bottom of the column we recovered soybean oil containing quantities of hexane as low as 20 ppm when we operated at 120 bar, 40°C. The effect of process parameters is also discussed.     

超临界CO2流体从紫菜中萃取EPA和DHA的研究

采用超临界CO2 流体萃取技术从紫菜中萃取EPA和DHA ,研究了紫菜粉碎粒度、萃取温度、萃取压力和萃取时间对萃取率的影响。结果表明 :超临界CO2 流体从紫菜中萃取EPA和DHA的工艺可行 ,最佳的萃取条件为 :紫菜粉碎粒度为 2 0～ 80目 ,萃取温度 3 5℃ ,萃取压力 2 5MPa ,萃取时间 1 .5h。     

Eicosapentaenoic acid (20∶5n-3) from the marine microalgaPhaeodactylum tricornutum

Eicosapentaenoic acid (EPA, 20∶5n-3) was obtained from the marine microalgaePhaeodactylum tricornutum by a three-step process: fatty acid extraction by direct saponification of biomass, polyunsaturated fatty acid (PUFA) concentration by formation of urea inclusion compounds, and EPA isolation by semipreparative high-performance liquid chromatography (HPLC). Alternatively, EPA was obtained by a similar two-step process without the PUFA concentration step by the urea method. Direct saponification of biomass was carried out with two solvents that contained KOH for lipid saponification. An increase in yield was obtained because the problems associated with emulsion formation were avoided by separating the biomass from the soap solution before adding hexane for extraction of insaponifiables. The most efficient solvent, ethanol (96%) at 60°C for 1 h, extracted 98.3% of EPA. PUFA were concentrated by the urea method with a urea/fatty acid ratio of 4∶1 at a crystallization temperature of 28°C and by using methanol and ethanol as urea solvents. An EPA concentration ratio of 1.73 (55.2∶31.9) and a recover yield of 78.6% were obtained with methanol as the urea solvent. This PUFA concentrate was used to obtain 93.4% pure EPA by semipreparative HPLC with a reverse-phase, C₁₈, 10 mm i.d.×25-cm column and methanol/water (1% acetic acid), 80∶20 w/w, as the mobile phase. Eighty-five percent of EPA loaded was recovered, and 65.7% of EPA present inP. tricornutum biomass was recovered in highly pure form by this three-step downstream process. Alternatively, 93.6% pure EPA was isolated from the fatty acid extract (without the PUFA concentration step) with 100% EPA recovery yield. This two-step process increases the overall EPA yield to 98.3%, but it is only possible to obtain 20% as much EPA as that obtained by three-step downstream processing.     

间十五烷基酚乙醇溶液以及正己烷溶液密度与粘度的测定与关联

在常压条件下,利用比重瓶和乌氏粘度计,测定了间十五烷基酚分别与乙醇和正己烷形成的溶液在不同组成下的密度与粘度,并用VTF方程对所得到的数据进行关联,间十五烷基酚乙醇溶液的密度和粘度数据的标准偏差分别为0.15%和1.22%,间十五烷基酚正己烷溶液的密度和粘度数据的标准偏差分别为0.08%和0.15%,基本符合工程设计需要。     

富里酸诱导对三角褐指藻EPA合成积累的影响

探究不同富里酸质量浓度对三角褐指藻二十碳五烯酸(EPA)合成积累的影响。结果表明,在质量浓度20 mg/L富里酸诱导下,三角褐指藻EPA含量和产量均达到最大值,为19.81 g/100 g和738.91 mg/L,较无富里酸诱导EPA含量和产量分别提高1.12和1.50倍。外源富里酸作用提高了三角褐指藻胞内过氧化物酶(POD)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)的酶活,使色素(叶绿素、类胡萝卜素)和总多酚抗氧化组分比例增加,活性氧(ROS)和丙二醛(MDA)水平降低,整体抗氧化能力获得显著提升,有效阻抑EPA的氧化分解;富里酸作用同时还上调了EPA合成通路去饱和酶(Δ⁵-FADS、Δ⁶-FADS、Δ¹²-FADS、Δ¹⁵-FADS、Δ¹⁷-FADS)和延长酶(Δ⁶-ELOVL)等系列关键酶,使其上调表达量较无富里酸作用分别增加0.33、0.94、0.33、0.58、1.15和0.70倍,为EPA高效合成积累提供前驱物和能量。     

甲苯－乙醇－正己烷（四氯化碳）三元体系过量焓

使用流动式微量热计测量了２９８．１５Ｋ下甲苯－乙醇－正己烷和甲苯－乙醇－四氯化碳两组三元体系以及正己烷－乙醇、乙醇－甲苯、正己烷－甲苯、四氯化碳－甲苯和四氯化碳－乙醇五组二元体系过量焓，对实测数据进行了关联，并计算了全浓度范围内过量焓，绘制了三元体系的等过量焓图。     

甲苯－乙醇－正己烷（四氯化碳）三元体系过量焓

使用流动式微量热计测量了２９８．１５Ｋ下甲苯－乙醇－正己烷和甲苯－乙醇－四氯化碳两组三元体系以及正己烷－乙醇、乙醇－甲苯、正己烷－甲苯、四氯化碳－甲苯和四氯化碳－乙醇五组二元体系过量焓，对实测数据进行了关联，并计算了全浓度范围内过量焓，绘制了三元体系的等过量焓图。     

富里酸诱导对三角褐指藻EPA合成积累的影响

探究不同富里酸作用浓度对三角褐指藻二十碳五烯酸（EPA）合成积累的影响。结果表明,在20 mg/L富里酸诱导下,三角褐指藻EPA含量和产量均达到最大值,为19.81 g/100 g和738.91 mg/L,较无富里酸诱导EPA含量和产量分别提高2.01和2.54倍。外源富里酸作用提高了三角褐指藻胞内过氧化物酶（POD）、超氧化物歧化酶（SOD）、过氧化氢酶（CAT）酶活,使得色素和多酚抗氧化组分比例增加,活性氧（ROS）和丙二醛（MDA）水平降低,整体抗氧化能力获得显著提升,有效阻抑EPA的氧化分解;富里酸作用同时还上调了EPA合成通路脱饱和酶（Δ5-FADS、Δ6-FADS、Δ12-FADS、Δ15-FADS、Δ17-FADS）和延长酶（Δ6-ELOVL）等系列关键酶,使其上调表达量较无富里酸作用分别增加1.27、1.81、1.24、1.45、1.43和1.46倍,为EPA高效合成积累提供前驱物和能量。富里酸作用效果协同强化了三角褐指藻EPA合成积累效率。     

Comparison of isopropanol and hexane for extraction of vitamin E and oryzanols from stabilized rice bran

The effects of solvent-to-bran ratio (2∶1 and 3∶1, w/w), extraction temperature (40 and 60°C), and time (5, 10, 15, 20, and 30 min) were studied for hexane and isopropanol extraction. Increasing the solvent-to-bran ratios and extraction temperature increased the amounts of crude oil, vitamin E and oryzanol recovered for both solvents. An extraction time of 15 min was sufficient for optimum crude oil, vitamin E, and oryzanol extraction. Preheated isopropanol (3∶1 solvent/bran ratio and 60°C) extracted less crude oil (P<.05) but more vitamin E (P<.05) and similar amounts of oryzanol (P>.05) relative to preheated hexane. The data suggest that isopropanol is a promising alternative solvent to hexane for extraction of oil from stabilized rice bran.     

海带中褐藻糖胶的酸提法研究

采用酸提法从海带中提取褐藻糖胶,分别采用MgCl、乙醇分级沉淀法进行纯化.主要探讨了提取时间、温度、pH等因素对提取率的影响.结果表明,在提取温度70 ℃,pH 2.5,70%乙醇浓度,提取3 h,提取率为7.3%.乙醇分级沉淀法提取率高于MgCl法.