Reverse Micellar Extraction of Bromelain from Pineapple (Ananas comosus L. Merryl) Waste: Scale-up,Reverse Micelles Characterization and Mass Transfer Studies

Scale-up studies for phase transfer mode of reverse micellar extraction are attempted for the separation and primary purification of bromelain (EC 3.4.22.33) from pineapple (Ananas comosus L. Merryl) waste. Characterization of reverse micelles and mass transfer studies for the real system has been attempted for the first time. Scale-up of the extraction process employing commercial grade surfactant cetyltrimethylammonium bromide (CTAB) and solvent isooctane resulted in purification of 2.43 fold with an activity recovery 81.3%. The reverse micellar size estimated using empirical and geometrical models indicated that the reverse micelles are large enough (R_m = 7.2–9.6 nm) to host bromelain molecules that are relatively smaller in size (～1.67 nm). The studies on the kinetics of mass transfer indicated a relatively slower rate (by ～34%) of mass transfer in case of back extraction compared to forward extraction. Process scale-up did not significantly affect the extraction efficiency whereas purity of phase components played a major role. The mass transfer across the phases was high in the initial period of mixing for both forward and back extractions.     

Activity of horseradish peroxidase in aqueous and reverse micelles and back‐extraction from reverse‐micellar phases

Studies on the activity of the enzyme horseradish peroxidase (HRP) have been carried out in micellar as well as reverse‐micellar media. The activity of the enzyme was studied in the presence of different classes of surfactants – ionic as well as non‐ionic. In aqueous media, the activity of the enzyme varied depending on whether the concentration of the surfactant used was above or below the critical micellar concentration (CMC). The enzyme was also studied in reverse‐micellar systems. HRP was introduced into the reverse micellar phase by the injection method and its activity within the reverse micelles was determined. The effect of water to surfactant ratio (W_o) on activity within reverse micelles was studied, and an almost two‐fold increase in activity was seen when the enzyme was encapsulated within reverse micelles of aqueous phase fractional hold‐up (?) of 0.0072 (v/v) consisting of sodium bis‐(2‐ethylhexyl) sulfosuccinate (AOT) in isooctane at a W_o of 20. The activity of HRP was measured over a wide range of AOT concentrations having different W_o values. Back‐extraction of HRP from these reverse micelles was carried out at varying ionic strengths of phosphate buffer. Back extraction was found to be highest at pH 7.0 in 40 mol m^?3 phosphate buffer and 100 mol m^?3 sodium chloride. © 2001 Society of Chemical Industry     

Purification of recombinant human interferon gamma from fermentation broth using reverse micellar extraction: A process optimization study

In the present investigation, we have shown a single step purification of IFN-γ from the fermentation broth of recombinant Kluyveromyces lactis (K. lactis) strain, using cationic surfactant based reverse micellar extraction (RME) system. The forward and back extraction processes were optimized in order to improve the overall extraction process. 78%, 93% and 98% forward extraction efficiencies (FEE) were obtained after the process by using aqueous phase pH 12, 150 mM CTAB and 0.2 M NaCl respectively. Afterwards, back extraction efficiency (BEE) was maximized by the optimization of stripping phase pH, isopropyl alcohol (IPA) and potassium chloride (KCl) concentration by employing Taguchi’s method of orthogonal array. Stripping phase pH 7, 15% IPA & 0.8 M KCl were found to be the best possible parameters producing 83% BEE. The usage of IPA has proven to reduce the effect of micellar-micellar based interaction, which was reflected as an improvement in BEE. This study demonstrates that the reverse micellar system is a highly promising and efficient tool for the direct extraction of recombinant proteins from fermentation broth. Moreover, this system holds a high potential to be an integral part of the downstream process used in biopharmaceutical industry.     

反胶团技术应用研究进展

反胶团具有独特的优良性能,有着十分广阔的应用前景,近年来正越来越多地受到关注。该文介绍了反胶团理论的最新研究进展,综述了反胶团技术在酶催化反应、萃取分离、纳米粒子的制备等方面的应用研究进展, 提出了反胶团技术目前存在的问题,并对其应用前景作了展望。引用文献22篇。     

Selective reverse micellar extraction of penicillin acylase from E coli

The recovery of penicillin acylase from E coli by a new reverse micellar treatment is described. The results are compared with the cell disruption by ultrasound followed by reverse micellar extraction. The process gave selective extraction of penicillin acylase directly from the periplasmic space of E coli without disrupting the cells. Unlike ultrasonication which breaks open the cells entirely, making subsequent processing difficult and expensive, reverse micellar treatment of cells gave a moderate recovery of 60% of enzyme activity in a highly pure form. © 2001 Society of Chemical Industry     

反向微胶团萃取氨基酸的进展

综述了反向微胶团萃取氨基酸研究的发展过程。     

A comprehensive study of extraction of hyperoside from Hypericum perforatum L. using CTAB reverse micelles

BACKGROUND: Hyperoside is a valuable natural pharmaceutical that has many biomedical functions. By the traditional method of alcohol extraction, only hyperoside solution of very low concentration can be obtained, so a new method is urgently needed to produce pure hyperoside more effectively. Reverse micellar extraction has been widely used in the purification of biological macromolecules. Theoretically, this method could also be used to purify materials with small molecules. Therefore it would seem appropriate to consider the extraction of hyperoside, a material with small molecules, using reverse micelles. In this study the factors affecting hyperoside extraction using cetyl trimethylammonium bromide (CTAB) reverse micelles were comprehensively investigated. RESULTS: The results showed that the partition coefficient increases with an increase in aqueous phase pH or CTAB concentration before reaching a saturated state. The existence of anions in the system can affect the extraction in several ways. For example, SO and PO ions can form hydrogen bonds with hyperoside and thereby improve the extraction significantly. CONCLUSION: This study proved that extraction using CTAB reverse micelles is a very effective method to separate hyperoside from Hypericum perforatum L. Under optimal conditions the extraction efficiency can reach 68.3% and the single‐strip‐extraction efficiency can reach 85.9%. The proportion of hyperoside among the flavonoids left in the strip‐extract can reach 93.3 wt%. This finding is very significant, because it demonstrates that reverse micellar extraction is a promising way to separate and purify materials with small molecules. Copyright © 2008 Society of Chemical Industry     

微波辅助胶团提取甘草中的甘草酸和甘草甙

The feasibility of emploving non-ionic surfactant （Triton X-100） as an alternative and effective solventfor the microwave-assisted extraction of glycyrrhizic acid （GA） and liquiritin （LQ） from licorice root was studied.The optimal extraction parameters based on the microwave-assisted micellar extraction technique were determined.Under the optimal conditions, i.e. 5% （by volume） Triton X-100, microwave-assisted extraction for 3--5min at 373K, the percentage extraction of active ingredients reached the highest value. The preconcentration tactor for GA and L＇Q （about 13.5） and the extraction efficiency for these two ingredients approached 100% showed the coupling of microwave-assisted extraction and cloud-point extraction could be employed as a new and. effective techniquefor the rapid extraction and preconcentration of pharrnacologically active ingredients from medicinal plants SUCh aslicorice root without disturbing chromatographic analysis.     

十二烷基聚氧乙烯(9)醚双水相体系萃取苯酚

采用十二烷基聚氧乙烯醚(AEO9)双水相胶束体系进行苯酚的萃取,考察Na2SO4和NaCl对质量浓度为50 g/L AEO9水溶液体系浊点的影响。结果表明,Na2SO4对浊点下降的影响比NaCl大,当盐的浓度均为0.6 mol/L,添加Na2SO4时浊点为28.5℃,而添加NaCl时为36.5℃。研究了无机盐(Na2SO4和NaCl)、普通离子型表面活性剂(十六烷基三甲基溴化铵CTAB和十二烷基硫酸钠SDS)和gemini型酯基季铵盐阳离子表面活性剂(Ⅱ-12-6)对苯酚萃取率E的影响。结果表明,盐的加入会使E下降,但Na2SO4对E的影响比NaCl要小;添加SDS使E下降,并随着n(SDS)∶n(AEO9)的增加而下降;而添加CTAB或Ⅱ-12-6均使E明显提高,且随着n(CTAB)∶n(AEO9)、n(Ⅱ-12-6)∶n(AEO9)的增加而进一步提高;但在同等条件下添加Ⅱ-12-6的E高于添加CTAB。     

Extraction of Human IgG4 Monoclonal Antibodies Using AOT- and HDEHP-Isooctane Reverse Micelles

The extraction of 0.05 and 1 mg mL^?1 human IgG4 using reverse micelles (RMs) formed with anionic surfactants AOT or HDEHP in isooctane was evaluated. For both surfactants the use of 1 mg mL^?1 IgG4 resulted in higher forward extraction (FE), and generally better backward extraction (BE) yields than 0.05 mg mL^?1 IgG4, achieving optimum FE and BE yields at FE pHs of 5 at 3.13 mM AOT and 6 at 1.56 mM HDEHP. IgG4 precipitation at the interface was observed at the lower pHs during FE which appeared to cause low overall extraction yields. Water content analysis revealed AOT-RMs were much bigger than HDEHP-RMs.     

飞天蜈蚣七皂苷提取动力学研究

采用球形模型,以Fick第二定律为基础,建立飞天蜈蚣七皂苷的提取动力学模型.对飞天蜈蚣七皂苷进行乙醇回流提取,在不同的液料比(体积质量比)、提取温度及提取时间等条件下,对提取液中皂苷的质量浓度进行检测.再根据该球形模型方程分别推算出了提取速率常数、活化能、相对萃余率、有效扩散系数等动力学函数值.根据该模型方程推算出的提...     

Ionic liquid-assisted micellar extraction for the quantitative determination of sesquiterpenic acids in Valeriana officinalis L. (Caprifoliaceae)

ABSTRACT

A series of hydrophilic ionic liquids (ILs) were investigated as extractants for the quantitative determination of valerenic and acetoxyvalerenic acids in Valerianae radix. The influence of different factors, e.g. type of cation and anion, alkyl chain length, concentration, temperature, extraction time and solid–liquid ratio, was investigated. The results obtained showed that the extraction yield is strongly dependent on the cation type. The best outcome was obtained with IL capable of forming micelles in aqueous solution, e.g. 1-decyl-3-methylimidazolium chloride. As a result, an alternative protocol for the quantification of sesquiterpenic acids in valerian was developed and validated.     

以枸骨叶为原料提取熊果酸工艺的研究

采用正交实验筛选出从枸骨叶中提取熊果酸的最佳工艺。当使用 85%～ 95%乙醇提取两次、每次 80min时 ,对干枸骨叶的提取率达到 90 %。在后处理中使用 2 .5倍于沉淀物质量的石油醚洗涤 3次 ,最后使用乙醇重结晶得到产品。     

Back extraction of platinum metals from liquid membranes in an electric field

A new method for back extraction of platinum metals from the salts of tertiary amines and quaternary onium bases in the organic phase is proposed. This method allows the back extraction of platinum metals into diluted solutions of mineral acids over one stage. The method involves the passing of direct current through a system in which the extract containing a platinum metal is a liquid membrane. This method allows a practically complete back extraction of the anions PtCl₆ ^2-, RuNOCl₅ ^2- and Ru₂OCl₁₀ ^4- (the latter passes into the aqueous phase without breaking the dimer anion). It was found that when PtCl₆ ^2-, PdCl₄ ^2- and Ru₂OCl₁₀ ^4- are in a liquid membrane based on Oct₃NHCl in 1.2-dichloroethane, under the action of an electric field all these anions are transferred into aqueous 1 M HCl (E > 85%). To provide maximum back extraction of platinum metal anions, it is necessary to use small concentrations of extractants in 1.2-dichloroethane (0.1 M for Oct₃NHCl and 0.03–0.04 M both for Oct₄NCl and Ph₄PCl) over the period of the back extraction. Cyclic voltametry has demonstrated the determining role of chloroions in current transport through the interface of phases I and II.     

超临界CO2萃取大黄总蒽醌工艺研究

对大黄游离蒽醌的超临界萃取工艺进行了优化,确定最优萃取工艺条件为:静萃取时间60 m in,动萃取时间30 m in,夹带剂乙醇相对大黄用量3 mL/g,萃取温度45℃,萃取压力45 MPa。然后分别采用无机酸和生物酶对大黄结合蒽醌进行水解,以提取总蒽醌。结果表明:当盐酸浓度为5 mol/L,淀粉酶质量浓度为2 g/L,纤维素酶质量浓度为1 g/L时,大黄结合蒽醌的糖苷键达到最优水解,此时总蒽醌质量分数分别为2.46%、2.33%和2.23%。     

Solvent extraction of bacteriocins from model solutions and fermentation broths

A novel method for the extraction and purification of bacteriocins (small peptides with antimicrobial properties) from fermentation broths is described. Model solutions of nisin were prepared from Nisaplin®, a commercial whey‐based powder containing about 2.5% nisin and sold as a food preservative. These model solutions and fermentation broths from a culture of nisin‐producing Lactococcus lactis subsp lactis were cross‐flow filtered and stirred with toluene to obtain a white suspension in the toluene layer, which was back extracted with 10 mmol dm⁻³ HCl. Yields of the order of 90% for a single stage extraction were obtained followed by similar yields on back extraction. Yields dropped with decreasing pH, going through a minimum at pH 2. In larger scale experiments (11 dm³) at pH 6, a gel layer was formed which, on centrifugation and drying, gave a white solid containing about 50% nisin, the remainder being water and salt. Further process development would almost certainly bring this figure closer to the small‐scale value. The phase transfer technique was also applied to solutions containing variacin and carnocin (produced by Micrococcus varians and Carnobacterium piscicola, respectively). The mechanism of bacteriocin extraction probably involves concentration of the peptides, which are surface active, at the interface until their solubility is exceeded and they form reverse submicelles, which aggregate into reverse micelles to give a gel. © 2000 Society of Chemical Industry     

贯叶连翘中金丝桃素提取分离方法研究

研究了从贯叶连翘中提取分离金丝桃素,探讨了提取剂、料液比、提取次数、提取温度、提取时间等因素对金丝桃素提取率的影响,并通过正交实验对提取工艺进行了优化。实验表明:提取最佳条件为将原料在45℃水浸泡2 h,以75%的甲醇为提取剂,超声强化20 m in(功率1000 W),按料液比1∶6和1∶4各提取1次,每次3 h,LSI-106型大孔树脂分离,以甲醇洗脱(洗脱速度1~2 mL/m in),提取率为96.36%,得率为5.33 mg/10 g,纯度达到94.71%,同时测得金丝桃素的稳定性(RSD=0.586%)良好。     

超声波提取海红果中总多酚的工艺研究

采用超声波辅助甲醇提取法从海红果中提取总多酚,研究了提取温度、提取时间、料液比等对总多酚提取量的影响。结果表明,超声提取海红果中总多酚的最佳提取工艺条件为:提取液甲醇浓度60%,料液比(g/mL)为1∶12,在55℃下提取50 min。     

反胶团萃取磷酸溶液中镁的动力学

采用恒界面池研究了二壬基萘磺酸(DNNSA)-煤油-磷酸体系萃取镁离子的动力学.考察了搅拌转速和传质界面积对萃取速率的影响,实验结果表明:磷酸中镁离子的萃取速率在200 r·min~(-1)时出现与搅拌强度无关的化学反应动力学"坪区",此时萃取速率正比于两相接触面积,说明萃取过程为界面化学反应控制过程.在动力学"坪区",镁离子萃取速率正比于萃取剂浓度和水相镁离子浓度,随着温度升高萃取速率增加;萃取反应活化能为70.01 kJ·mol~(-1),并得到了DNNSA萃取磷酸中镁离子的萃取动力学方程.