Fast Partitioning Research Articles

Fast Centrifugal Partitioning Chromatography (FCPC) towards the recovery of secondary metabolites coming from roots of Argemone mexicana

Mexican poppy (Argemone mexicana L.) is an annual thorny herb which belongs to Papaveraceae family. It is commonly spread in Mexico and in south-western North America where it is growing in the wasteland. Argemone mexicana contains high variety of isoquinoline alkaloids which influence its activity (e.g. cholagogue, hipotensive, antifebrile, antimalarial, spasmolytic and depressive towards the CNS).

Preparative Isolation, Fast Centrifugal Partition Chromatography Purification and Biological Activity of Cajaflavanone from Derris ferruginea Stems

The Derris genus is known to contain flavonoid derivatives, including prenylated flavanones and isoflavonoids such as rotenoids, which are generally associated with significant biological activity. To develop an efficient preparative isolation procedure for bioactive cajaflavanone. Fast centrifugal partition chromatography (FCPC) was optimised to purify cajaflavanone from Derris ferruginea stems in a single step as compared to fractionation from the cyclohexane extract by successive conventional solid-liquid chromatography procedures. The purification yield, purity, time and solvent consumption per procedure are described. The anti-fungal, anti-bacterial, anti-leishmanial, anti-plasmodial, anti-oxidant activities and the inhibition of advanced glycation end-products (AGEs) by cajaflavanone accumulation are described. FCPC enabled cajaflavanone purification in a single separation step, yielding sufficient quantities to perform in vitro biological screening. Interestingly, cajaflavanone had an inhibitory effect on the formation of AGEs, without displaying any in vitro anti-oxidant activity. A simple and efficient procedure, in comparison with other preparative methods, for bioactive cajaflavone purification has been developed using FCPC.

Intensified extraction of ionized natural products by ion pair centrifugal partition extraction

The potential of centrifugal partition extraction (CPE) combined with the ion-pair (IP) extraction mode to simultaneously extract and purify natural ionized saponins from licorice is presented in this work. The design of the instrument, a new laboratory-scale Fast Centrifugal Partition Extractor (FCPE300 ®), has evolved from centrifugal partition chromatography (CPC) columns, but with less cells of larger volume. Some hydrodynamic characteristics of the FCPE300 ® were highlighted by investigating the retention of the stationary phase under different flow rate conditions and for different biphasic solvent systems. A method based on the ion-pair extraction mode was developed to extract glycyrrhizin (GL), a biologically active ionic saponin naturally present in licorice ( Glycyrrhiza glabra L., Fabaceae) roots. The extraction of GL was performed at a flow rate of 20 mL/min in the descending mode by using the biphasic solvent system ethyl acetate/ n-butanol/water in the proportions 3/2/5 (v/v/v). Trioctylmethylammonium with chloride as a counter-ion (Al336 ®) was used as the anion extractant in the organic stationary phase and iodide, with potassium as counter-ion, was used as the displacer in the aqueous mobile phase. From 20 g of a crude extract of licorice roots, 2.2 g of GL were recovered after 70 min, for a total process duration of 90 min. The combination of the centrifugal partition extractor with the ion-pair extraction mode (IP-CPE) offers promising perspectives for industrial applications in the field of natural product isolation or for the fractionation of natural complex mixtures.

Preparative Isolation of Bioactive Nitrile Glycoside “Niazirin” from the Fruits of Moringa oleifera Using Fast Centrifugal Partition Chromatography

Fast centrifugal partition chromatography was successfully applied in the separation of bioactive constituent niazirin directly from the chloroform extract of Moringa oleifera. The experiment was performed with a two-phase solvent system composed of ethyl acetate/BuOH/water (6:0.5:4 v/v/v) in which the upper organic layer was used as stationary phase and lower aqueous phase was used as mobile phase. From 1 g of crude extract, 70 mg of niazirin was obtained in 94.8% purity as determined by HPLC. The total yield recovery was >94%. The isolated nitrile glycoside (niazirin) was characterized on the basis of its 1H, 13C-NMR and ESI-MS data.

Development and validation (according to the 2002/657/EC regulation) of a method to quantify sulfonamides in porcine liver by fast partition at very low temperature and LC-MS/MS.

A novel multi-residue method for the quantification of 15 sulfonamides in porcine liver is described. It involves the application of a liquid-liquid extraction with fast partition at very low temperature (LLE-FPVLT) procedure followed by HPLC-MS/MS (high performance liquid chromatography coupled to tandem mass spectrometry) analysis. By this innovative method, acetonitrile is added to a minced porcine liver sample and the resulting suspension centrifuged and immersed in a container with liquid nitrogen for 15 s. The acetonitrile phase, which remains liquid under these conditions, is isolated, evaporated to dryness, recomposed with formic acid 0.1% v/v, and injected into the liquid chromatograph. The whole analytical procedure was validated according to the European Commission Decision 2002/657/EC and acceptable values (except for sulfanilamide) were obtained for the following parameters: linearity (0.97 < R2 < 0.99), decision limit (107.70 µg kg-1 < CCα < 128.65 µg kg-1), detection capability (115.40 µg kg-1 < CCβ < 157.29 µg kg-1), limit of detection (5.58 µg kg-1 < LOD < 16.75 µg kg-1), limit of quantification (18.41 µg kg-1 < LOQ < 55.26 µg kg-1), accuracy (recovery rates), precision (repeatability, intermediate precision, and measurement uncertainty tests), selectivity, and robustness. Recoveries higher than 70%, at three concentration levels (0.5, 1.0 and 1.5 of the maximum residue limit, MRL), were attained for the majority of the sulfonamides. These very promising results point to the inclusion of the present methodology into the National Residue Control Plan scope of the Ministry of Agriculture, Livestock and Food Supply of Brazil.

Preparative Purification of Delphinidin 3-0-sambubioside from Roselle (Hibiscus sabdariffa L.) Petals by fast Centrifugation Partition Chromatography

Delphinidin 3-0-sambubioside, a Hibiscus anthocyanin, was isolated from MeOH/TFA dried flower of H. sabdariffa. Its purification on preparative scale was obtained by centrifugal partition chromatography (CPC) using the ternary biphasic solvent systems composed of ethyl acetate/1-butanol/water, acidified by 0.1% of TFA. Stationary phase was ethyl acetate/1-butanol/water (5:5:90; v/v). We tested 4 mobile phases and found that the system acetate/1-butanol/water (40:46:14; v/v) was the best to separate anthocyanin mentioned above. This support-free liquid-liquid chromatographic procedure made it possible to isolate delphinidin 3-0-sambubioside from flower of H. sabdariffa. The antiamyloidogenic activity of the isolated stilbenes was evaluated versus b-amyloid fibrils. delphinidin 3-0-sambubioside was found to be active with 67% inhibition at 10 mM.

Identification of novel spinal cholinergic genetic subtypes disclose Chodl and Pitx2 as markers for fast motor neurons and partition cells

Spinal cholinergic neurons are critical for motor function in both the autonomic and somatic nervous systems and are affected in spinal cord injury and in diseases such as amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy. Using two screening approaches and in situ hybridization, we identified 159 genes expressed in typical cholinergic patterns in the spinal cord. These include two general cholinergic neuron markers, one gene exclusively expressed in motor neurons, and nine genes expressed in unknown subtypes of somatic motor neurons. Further, we present evidence that chondrolectin (Chodl) is expressed by fast motor neurons and that estrogen-related receptor beta (ERRbeta) is a candidate marker for slow motor neurons. In addition, we suggest paired-like homeodomain transcription factor 2 (Pitx2) as a marker for cholinergic partition cells.

Identification of Bisprenylated Benzoic Acid Derivatives from Yerba Santa (Eriodictyon ssp.) Using Sensory-Guided Fractionation

Due to certain off-flavor problems and lacking bitter masking effects with Yerba Santa (Eriodictyon angustifolium and E. californicum) extracts, which are also described as bitter, herbal, medicinal, phenolic, or astringent, methanolic extracts were fractionated and evaluated for their taste properties using a high temperature liquid chromatography (HTLC)-based approach. The taste-guided fractionation led to the identification of a series of novel bisprenylated benzoic acids (erionic acids A (1), B (2), C (3), D (4), E (5), and F (6) and eriolic acids A (7), B (8), C (9), and D (10), respectively), along with the known flavonoids eriodictyol, homoeriodictyol, hesperetin, and chrysoeriol. The new compounds were isolated in larger amounts for characterization from Narrow Leaf Yerba Santa (E. angustifolium) and California Yerba Santa (E. californicum), respectively, using fast centrifugal partition chromatography (FCPC) and HTLC. The structures were elucidated using one and two-dimensional NMR spectroscopy and high resolution mass spectrometry (HR-MS). For E. californicum, data regarding seasonal and climatic variation of the eriolic acid contents and of the flavonoids were collected. The flavor properties of some of the isolated new compounds were evaluated; they showed strong off-flavor characteristics, such as bitter, astringent, phenolic, or woody, and may contribute to the sensory effects observed for crude Yerba Santa extracts. Erionic acid C (3) was not only able to increase the absolute bitterness but also to extinguish the bitter masking effect of homoeriodictyol in a caffeine solution.

Comparing extraction methods to recover ginseng saponins from American ginseng (Panax quinquefolium), followed by purification using fast centrifugal partition chromatography with HPLC verification

A series of experiments were carried out to compare the extraction of ginseng saponins, ginsenosides, from powdered American ginseng ( Panax quinquefolium ) using pressurized hot water and the more conventional ultrasonic-assisted extraction. Three solvents were tested, water, n -butanol-saturated water, and water-saturated n -butanol. Each resulting extract was further purified using fast centrifugal partition chromatography (FCPC) in order to better quantify the contents of the crude plant extract. The pressurized hot water system extracted a greater yield of saponins, 11.2 mg/g (extraction at 110 °C and 440 kPa), than the ultrasonic-assisted method, 7.2 mg/g (extraction at variable temperature with no external pressure). The difference in solvent system for either extraction methods was not significant, and the results gave credence for the use of water as the extraction solvent. n -Butanol-saturated water yielded the most saponins (10.1 mg/g), while water yielded 9.8 mg/g, and water-saturated n -butanol yielded 7.8 mg/g. Since water is an environmentally benign solvent, this result is quite attractive for future work.

Fast Counter Current Chromatography of n-Butanolic Fraction from Senecio giganteus (Asteraceae)

Phenolic derivatives such as quinones, acid-phenols and flavonoids were successfully isolated from a n-butanolic fraction of Senecio giganteus Desf. (Asteraceae) flowers, namely jacaranone (1), 3a-hydroxy-3,3a,7,7a-tetrahydrobenzofuran-2,6-dione (2), chlorogenic acid (3), hyperoside (4), quercetin 3-O-beta-D-robinobioside (5), isorhamnetin-3-O-beta-D-glucuronide (6), quercetin-3-O-beta-D-glucuronide (7), and isorhamnetin-3-O-beta-D-glucuronide-6"-methyl ester (8). These compounds were purified through either classical polyamide filtration followed by fractionation on Si gel, or through fast centrifugal partition chromatography (FCPC). Using FCPC, the major compounds could be readily isolated from the crude n-butanolic fraction. Compounds 1-8 were identified by means of spectroscopic and spectrometric analysis (UV, 1H, 13C and 2D NMR, and MS). This work described for the first time the phytochemical composition of this endemic Algerian plant.

Evaluation of antioxidant activity and phytochemical analysis of Mentha microphylla C.Koch extracts

The genus Mentha belongs to the Lamiaceae family and it consists of ca. 25–30 species of subcosmopolitan distribution, of which 7 are found in the Greek territory. Members of the genus Mentha are known for their spasmolytic, antibacterial and antigenotoxic properties1. Extracts (dichloromethane, methanol, water) of M. microphylla, M. aquatica, M. longifolia and M. pulegium were obtained with A.S.E (Accelerated Solvent Extraction). Methanolic and aqueous extracts were tested for DPPH scavenging and Total Phenolic Content and showed very strong antioxidant activity. Among them M. microphylla C.Koch was chosen for further phytochemical investigation. The Total Phenolic Content for M.microphylla C.Koch was 315.5mg GAE/g of ethanolic extract and 274.5mg GAE/g of aqueous extract. The DPPH %ΔA values were 100% for the ethanolic and 98% for the aqueous extract relatively. Dichloromethanic, ethanolic and aqueous extracts were received with conventional extraction. From the dichloromethanic extract several monoterpenes (i.e. piperitone, piperitenone oxide), triterpenes (i.e. β-amyrin), phenolic compounds (i.e. thymol) and a luteolin derivative were isolated by classic column chromatography. On the other hand, the ethanolic and aqueous extracts were fractionated with F.C.P.C (Fast Centrifugal Partition Chromatography) technique and were proved to be rich sources of rosmarinic acid and flavonoid compounds (i.e. diosmin). Structural elucidation of all compounds was determined by spectroscopic methods (NMR 1D, 2D).

Phytochemical study and biological evaluation of the stem of Derris ferruginea Bentham

The genus Derris Loureiro belongs to the tribe Milletiae of the Leguminosae. It includes about fifteen species widely found in the tropical areas of Africa and Asia [1]. These plants have been traditionally used over centuries as fish poisoning, insecticide and pesticide, particularly due to their large production of rotenone [2]. Biological activities of Derris species are various: cytotoxic, antibacterial, antifungal, and antioxidant [3,4,5]. Though major secondary metabolites found in the genus are known to be flavonoids, including prenylated flavanones, and isoflavonoids such as rotenoids, very few phytochemical informations were available on D. ferruginea [6]. Different crude extracts were obtained from the stem (2kg, Soxhlet apparatus). They were fractionated using successively MPLC, LC, FCPC® (Fast Centrifugal Partition Chromatography, Kromaton Angers), Sephadex LH-20® gel and finally purified on preparative HPLC. Chemical structures of the isolated compounds were elucidated using 1H and 13C NMR spectrum as well as Mass Spectrometry. Most of these compounds were identified as prenylated flavonoids (flavanones and isoflavonoids). Biological effects of these compounds will be reported here, particularly antimicrobial, antiparasitic activities and inhibition of the formation of AGEs (Advanced Glycation End Products involved in age- and diabetes-related chronic diseases).

Extrapolation Scheme For Fast 3D Ising Field Partition Function Estimation. Application to Bayesian Analysis of Within Subject fMRI Data

Extrapolation Scheme For Fast 3D Ising Field Partition Function Estimation. Application to Bayesian Analysis of Within Subject fMRI Data

Two new isolated epimers seem to contribute to the bitter principle of Leonurus japonicus Houtt.

The aerial parts of Chinese motherwort (Leonurus japonicus Houtt) (Yimucao) are commonly used in traditional Chinese medicine (TCM) to regulate menses and to treat associated conditions. As the herb is more and more used in Europe, it has been started to cultivate the plant in Germany [1]. Since bitterness is an important sensory feature of the plant, the aim of the investigation was to isolate and identify the chemical constituents responsible for the bitter taste of Yimucao. It was found that the bitter principle can be extracted by n-hexane. Therefore, the n-hexane extract of Herba Leonuri has been fractionated using Fast Centrifugal Partition Chromatography (FCPC).This led to a fraction containing a mixture of two compounds which seemed to be responsible for the bitter taste of Yimucao. Structure elucidation by MS and NMR spectroscopy led to two new epimeric diterpenoid compounds 1 and 2 (exact mass 392,2), which are related to furanic labdane type lactones previously found in Leonurus japonicus, Leonurus cardiaca, Leonotis leonurus and Marrubium vulgare [2,3].

Diffusive partitioning tracer test for the quantification of nonaqueous phase liquid (NAPL) in the vadose zone: Performance evaluation for heterogeneous NAPL distribution

A partitioning tracer test based on gas-phase diffusion in the vadose zone yields estimates of the residual nonaqueous phase liquid (NAPL) saturation. The present paper investigates this technique further by studying diffusive tracer breakthrough curves in the vadose zone for a heterogeneous NAPL distribution. Tracer experiments were performed in a lysimeter with a horizontal layer of artificial kerosene embedded in unsaturated sand. Tracer disappearance curves at the injection point and tracer breakthrough curves at some distance from the injection point were measured inside and outside of the NAPL layer. A numerical code was used to generate independent model predictions based on the physicochemical sand, NAPL, and tracer properties. The measured and modeled tracer breakthrough curves were in good agreement confirming the validity of important modeling assumptions such as negligible sorption of chlorofluorocarbon (CFC) tracers to the uncontaminated sand and their fast reversible partitioning between the soil air and the NAPL phase. Subsequently, the model was used to investigate different configurations of NAPL contamination. The experimental and model results show that the tracer disappearance curves of a single-well diffusive partitioning tracer test (DPTT) are dominated by the near-field presence of NAPL around the tip of the soil gas probe. In contrast, breakthrough curves of inter-well tracer tests reflect the NAPL saturation in between the probes, although there is no unique interpretation of the tracer signals if the NAPL distribution is heterogeneous. Numerical modeling is useful for the planning of a DPTT application. Simulations suggest that several cubic meters of soil can be investigated with a single inter-well partitioning tracer test of 24-hour duration by placing the injection point in the center of the investigated soil volume and probes at up to 1 m distance for the monitoring of gaseous tracers.

Large-scale production of selected type A trichothecenes: the use of HT-2 toxin and T-2 triol as precursors for the synthesis of d 3-T-2 and d 3-HT-2 toxin

The type A trichothecenes T-2 and HT-2 toxins are toxic secondary metabolites produced by fungi of the Fusarium genus. Their occurrence in cereals, especially in oats, implies health risks for the consumer. Therefore, it is an important task to develop selective and sensitive methods for the analysis of T-2 and HT-2 toxins, and to undertake further studies on their stability and toxicity. Although most toxins are commercially available, their high prices are the limiting factor on the realization of these experiments. Thus, we developed a method for large-scale production of T-2 and HT-2 toxin as well as T-2 triol and T-2 tetraol. T-2 toxin was obtained in gram quantities by biosynthetic production with cultures of F. sporotrichioides. As HT-2 toxin was only formed as a by-product, and T-2 triol and T-2 tetraol were not generated, these compounds were produced by alkaline hydrolysis of T-2 toxin. Separation and isolation of crude toxins was achieved by fast centrifugal partition chromatography (FCPC), which is an efficient tool for the large-scale purification of natural products. Using this fast and yield effective technique, several hundred milligrams of HT-2 toxin, T-2 triol, and T-2 tetraol were obtained. Subsequent, HT-2 toxin and T-2 triol were used for the large-scale synthesis of isotope-labeled T-2 and HT-2 toxin, respectively. Using these standards, an isotope dilution-(ID)-HPLC-MS/MS method for the quantification of T-2 and HT-2 toxin in different matrices was developed.

A FAST MULTISPRITE GENERATOR WITH NEAR-OPTIMUM CODING BIT-RATE

Sprite coding, which can increase the coding efficiency of backgrounds greatly, is a novel technology adopted in MPEG-4 object-based coding. Due to the geometric transformation applied to each nonreference frame in the procedure of sprite coding, the generated sprite is distorted and the available view angles relative to the reference frame are restricted. These problems can be resolved by using multiple sprites. An optimal multisprite generator has been proposed by Farin et al., but the optimal method requires high computation in the sprite coding cost. To treat this disadvantage, a fast multisprite partition algorithm is proposed in this paper based on frame translations and scalings. The proposed algorithm divides a video sequence into several subsequences, and a sprite is generated from each subsequence. The experimental results show that the executing speed of the proposed method is increased by 10 to 190 times than the optimal method, with the total size of generated sprites is slightly higher and the qualities of generated sprites are preserved.

Advantages of a small-volume counter-current chromatography column

Counter-current chromatography (CCC) works with a support-free liquid stationary phase. This allows for preparative separations and purifications. However, there are serious technical constraints because of the need to keep a liquid stationary phase in a column. Centrifugal fields are used. A new commercial hydrodynamic 18 mL column made with a narrow-bore 0.8 mm Teflon tubing was evaluated by comparing it with older hydrodynamic CCC columns and a similar 19 mL column but made with 1.6 mm Teflon tubing. A small-volume CCC column allows for reliable and fast solute partition coefficient determination. When resolution is required, both high efficiency and liquid stationary phase retention are needed. Unfortunately, these two requirements bear technical contradictions. A column coiled with a narrow tubing bore will provide a high chromatographic efficiency while a column containing wider tubing bore will achieve higher stationary phase retention. In all cases, increasing the magnitude of the centrifugal field also increases the stationary phase retention. The solution is to build centrifuges able to produce high fields that will provide acceptable liquid phase retention with narrow-bore tubes. The new 18 mL 0.8 mm tubing bore column is able to rotate as fast as 2100 rpm generating a 240 × g field. The two older CCC columns cannot compete with the new one. However, the small 19 mL column with 1.6 mm bore tubing can be useful when fast results are desired without top resolution.

Separation of Silymarins from Milk Thistle (Silybum Marianum L.) Extracted with Pressurized Hot Water using Fast Centrifugal Partition Chromatography

Fast centrifugal partition chromatography was used to separate a class of flavonolignans called silymarins from both a purchased silymarin powder and a crude pressurized hot water extract of milk thistle (Silybum marianum L.). Initially, a purchased power of a mixture of the six silymarin compounds was separated with a two-phase solvent system consisting of heptane/ethyl acetate/methanol/water (1:4:3:4 v/v/v/v) in order to verify elution times of the compounds by fast centrifugal partition chromatography. Next, a crude pressurized hot water extract from 10 g of ground seeds of Silybum marianum was separated with the same solvent system. The separation from the hot water extract gave yields of silychristin at 70.2% purity, silydianin at 93.7% purity, and a mixture of silybinin and isosilybinin at 96.1% purity.

Purification hemisynthesis of xanthatin derivates and in vitro evaluation of their activity towards farnesyltransferase (PFTase)

Originating from America and introduced a few centuries ago in Europe, Xanthium macrocarpum DC. (Asteraceae) also called „Lampourde a gros fruits“ is a species commonly growing on the edges of the Loire. Xanthanolides (sesquiterpene lactones) found in this plant exhibit interesting biological activities [1]. These activities are usually explained by their alkylating properties due to the presence of the α-methylene-γ-lactone function which is also related to the toxicity of several plants. As recent reports pointed out the potential of dimeric or monomeric sesquiterpene lactones as farnesyltransferase (PFTase) inhibitors, we decided to explore this activity knowing that PFTase is an interesting target to find new effective therapeutic agents for the treatment of cancer. So, the objective was to obtained atoxic hemisynthetic derivates from the natural xanthanolides: xanthatin and xanthinin. These compounds were first isolated in one step from the crude chloroformic extract of the leaves of X. macrocarpum using differents methods of chromatography: silica gel and a 5L pilot scale FCPC® (Fast Centrifungal Partition Chromatograph, Kromaton, Angers, France). We have shown that FCPC® is more efficient for purification: solvent consumption is lowered (divided by 2), with highest purity (5 fold increase on average), high loading capacity (2g of extract/L of organic solvent), and manipulation time reduction (few hours versus few days) [2]. Sixteen derivates were investigated as potent inhibitors of protein farnesyl transferase. These results showed that the α-methylene-γ-lactone function is not required to insure a PFTase inhibitory activity whereas the α-methyl-γ-lactone function is responsible for a non competitive-inhibition.