Horizontal Flow-through Coil Planet Centrifuge Research Articles

Studies on the peparative capability of the horizontal flow-through coil planet centrifuge and high-performance liquid chromatography in the separation of polar compounds from Oxytropis ochrocephala Bunge

Horizontal flow-through planet centrifuge (CPC) and high-performance liquid chromatography (HPLC) techniques were used for separation of polar compounds from a crude ethanol extract of Oxytropic ochrocephala Bunge, a poisonous legume plant widely distributed in northwestern China. The performance of these two chromatographic methods was compared in terms of column efficiency, peak resolution, separation time, sample loading capacity, etc. The results indicated that two polar compounds in the crude extract were equally well separated by these two methods. HPLC gave comparable peak resolution in shorter separation time while its sample loading capacity was limited to the mg range. The CPC method required a long separation time, but yielded a higher purity of fractions with a much greater capacity.

Protein separation with aqueous—aqueous polymer systems by two types of counter-current chromatographs

Two different types of counter-current chromatographs, the cross-axis coil planet centrifuge ( X-axis CPC) and horizontal flow-through coil planet centrifuge (horizontal CPC), were evaluated for protein separation with an aqueous-aqueous two-phase polymer system. The sample solution, containing 10–200 mg each of cytochrome c, myoglobin, ovalbumin and hemoglobin in 2 ml of each phase was eluted with the lower phase. In both instruments, the effects of the flow-rate, revolution speed, and parameter β (helical diameters of the multilayer coil) on the protein separation were investigated. The best results were obtained from the X-axis CPC operated at 750 rpm and a flow-rate of 2.0 ml/min using a multilayer coil with a small helical diameter (β = 0.25–0.60). Four protein samples were well resolved in less than 5 h.

Chromatography of peptides on a multi-coil counter-current chromatograph

A modified horizontal flow-through coil planet centrifuge has been constructed that has the potential for preparative choromatography. The multi-coil counter-current chromatograph, equipped with a set of four multi-layer coils, has have improved performance with polar solvents suitable for peptide elution. Separations of a group of dipeptides and purification of a cholecystokinin fragment and an undecapeptide were achieved with n-butanol—acetic acid—water at high flow-rates with good resolution.

Separation of Flavonoids in Crude Extract from Sea Buckthorn by Countercurrent Chromatography with Two Types of Coil Planet Centrifuge

Abstract Flavonoid constituents in a crude ethanol extract from dried fruits of sea buckthorn (Hippophae rhamnoides) were subjected to countercurrent chromatography with a two-phase solvent system composed of chloroform, methanol and water at a 4:3:2 volume ratio. Separations were performed with two different types of the coil planet centrifuge: One is called the multilayer coil planet centrifuge and the other, the horizontal flow-through coil planet centrifuge. Both instruments permit continuous elution through the rotating column without the use of rotary seals. Although the horizontal flow-through coil planet centrifuge produced efficient peak resolution for five flavonoid components, the multilayer coil planet centrifuge yielded much superior results in terms of partition efficiency, separation time and sample loading capacity.

Large Scale Purification of Apocarotenoids from Cochlospermum Tinctorium by Countercurrent Chromatography

Abstract Carotenoids are well-known for their instability so as usual analytical methods such as HPLC or preparative TLC applied to the separation of the constituents of Cochlospermum tinctorium failed to the isolation of pure pigments for further chemical characterization. Thus high-speed CCC using a horizontal flow-through coil planet centrifuge was used to conveniently and efficiently achieve this separation for quantities up to 500mg of the crude plant extract.

Horizontal flow-through coil planet centrifuge equipped with a set of multilayer coils around the column holder: Counter-current chromatography of proteins with a polymer-phase system

Horizontal flow-through coil planet centrifuge equipped with a set of multilayer coils around the column holder: Counter-current chromatography of proteins with a polymer-phase system

Preparative Purification of Peptides by Countercurrent Chromatography on the Ito Coil Planet Centrifuges

Abstract For the purification of up to 1000 mg of synthetic peptides by countercurrent chromatography, the coil planet centrifuges have proven useful in the research laboratory. Besides the earlier described horizontal flow-through coil planet centrifuge which chromatographs substances in all solvent systems at room temperature, the multi-layer coil planet centrifuge affords more rapid chromatography. However, separations using n-butanol, especially suited for peptides, have to be conducted at elevated temperatures. A new machine that has characteristics of both of the foregoing instruments is the compact horizontal flow-through coil planet centrifuge which promises rapid chromatography with full retention of the stationary phase.

Separation of S-Triazine Herbicides by Countercurrent Chromatography

Abstract Countercurrent chromatography (CCC) has been successfully applied for the separation of four chlorinated s-triazine derivatives, namely, simazine, atrazine, propazine, and trietazine, which are widely used as herbicides. Of several solvent systems investigated, n-hexane-ethyl acetate-methanol-water (8:2:5:5) gave the best range of the partition coefficient values because of the satisfactory solubility for all samples and therefore was used for the separation of s-triazines. Two types of countercurrent chromatographs, high speed CCC, and horizontal flow-through coil planet centrifuge, were used for s-triazines separation, and the identity of the separated fractions was unequivocally established by mass spectrometry. The potential practical application of CCC to herbicide analysis is discussed.

Large-Scale Preparative Counter-Current Chromatography with a Coil Planet Centrifuge

Abstract Development of the large-scale preparative countercurrent chromatographic schemes has been continued by increasing the diameter of the separation column. A 0.55 cm i.d. FEP tube was coaxially coiled around the holder (7.5 cm, 10 cm or 15 cm in diameter) of a horizontal flow-through coil planet centrifuge (15 cm revolutional radius). Performance of each column was evaluated on the separation of dinitrophenyl amino acid samples with a two-phase solvent system composed of chloroform, acetic acid, and 0.1N hydrochloric acid (2:2:1) by using both aqueous and nonaqueous phases as the mobile phase. Experiments with the short preliminary columns (114 ml capacity) revealed that the hydrodynamic distribution of the two solvent phases was sensitively affected by the helical diameter of the column. However, by choosing the proper elution mode of the mobile phase, satisfactory results were obtained with the helical diameters of 7.5 cm and 15 cm at a high flow rate of 500 ml/h under a moderate revolutional spe...

Chromatography of AC-ASP-TYR-MET-GLY-TRP-MET-ASP-NH2on the Horizontal flow-Through Coil Planet Centrifuge and the High-Speed Multi-Layer Coil Planet Centrifuge

Abstract The peptide Ac-Asp-Tyr-Met-Gly-Trp-Met-Asp-NH2 was purified by countercurrent chromatography in the horizontal flow-through coil planet centrifuge. The solvent system used was ammonium acetate, pH 8.5 and n-butanol (1:1 by volume). The high pH served to maintain the peptide in solution. When the upper phase was utilized as the mobile phase better separation of the peptide from impurities resulted. The peptide was also chromatographed in a new apparatus, the high-speed multi-layer coil planet centrifuge. With the lower phase mobile and at a higher temperature, the peptide was fractionated very rapidly in 30 min compared to 7 hr on the other instrument.

Gram Quantity Separation of DNP (Dinitrophenyl) Amino Acids with Multi-Layer Coil Countercurrent Chromatography (CCC)

Abstract The efforts have been successfully made to extend the preparative capability of the high-speed CCC scheme with a multi-layer large capacity coiled column. The apparatus is a table top model of a horizontal flow-through coil planet centrifuge which produces a synchronous planetary motion of the column holder. The separation column was prepared from a single piece of 70 m long, 2.6 mm i.d., PTFE tubing coiled around the spool-shaped holder to form multiple layers of the coil with a total capacity of about 400 ml. The performance of the apparatus was assessed with a standard set of DNP amino acid samples and a two-phase solvent system composed of chloroform, acetic acid and 0.1N HCl (2:2:1). Preparative capability of the method was evaluated in terms of the retention level of the stationary phase and peak resolution for various sample size ranging from 0.05g to 2g. The effects of sample volume, sample concentration and the choice of the sample diluent on the separation were studied. The results indi...

Continuous extraction of urinary anthracycline antitumor antibiotics with the horizontal flow-through coil planet centrifuge

Extraction of doxorubicin (adriamycin) and daunorubicin and their metabolites from human urine was attempted utilizing the horizontal flow-through coil planet centrifuge. Partition coefficients of the drugs for various combinations of non-aqueous phases and aqueous salt solutions were determined. Optimal coefficients for adriamycin and daunorubicin were achieved with n-butanol—0.3 M disodium hydrogen phosphate. Extraction efficiencies of the drugs from human urine comparable to those obtained by standard resin column techniques could be realized by employing the n-butanol—urine (containing 0.3 M disodium hydrogen phosphate) system in the coil planet centrifuge, at flow-rates of 500—600 ml/h, and at 650 rpm revolutional speed. Small quantities of drugs and metabolites could be continuously concentrated into small volumes of the n-butanol phase from large volumes of salted urine. The versatility of the technique was demonstrated by its application to extraction of aclacinomycin A, a novel anthracycline antitumor agent, and its metabolites from human urine.

Horizontal flow-through coil planet centrifuge: some practical applications of countercurrent chromatography

A horizontal flow-through coil planet centrifuge was constructed that holds a pair of coiled separation columns made of PTFE tubing of different inner diameters. With this scheme, samples from a few to hundreds of milligrams can be separated in each operation. The experimental methods are discussed from three aspects: the selection of the two-phase solvent systems, the determination of the optimal operational conditions and the general separation procedures. The capabilities of this apparatus were demonstrated with separations of several biological samples such as antibiotics, agricultural chemicals and medicinal herbs.

Experimental observations of the hydrodynamic behavior of solvent systems in high-speed counter-current chromatography : I. Hydrodynamic distribution of two solvent phases in a helical column subjected to two types of synchronous planetary motion

Hydrodynamic distribution of two-phase solvent systems in a rotating helical column subjected to centrifugal fields produced by two different types of synchronous planetary motion has been studied by the use of the combined horizontal flow-through coil planet centrifuge. With continuous elution of the mobile phase, the simpler type of motion resulted in low retention of the stationary phase in the column whereas a more complex motion, which produces a quasi-radial centrifugal field varying in both intensity and direction, yielded high stationary phase retention for commonly used solvent systems having a wide range of hydrophobicity. These solvent systems display highly complex modes of hydrodynamic interaction in the coil according to their particular physical properties.

Purification of a cholecystokinin fragment peptide in the horizontal flow-through coil planet centrifuge

Purification of a cholecystokinin fragment peptide in the horizontal flow-through coil planet centrifuge

Counter-current chromatography of diaziquone by a horizontal flow-through coil planet centrifuge

Diaziquone is an active antitumor agent in animals that is unstable in aqueous solution. The chemical hydrolysis products of this antitumor agent have been identified and we have investigated the usefulness of the coil planet centrifuge in separating these products from the parent compound. We were able to separate the reactive diaziquone from the two reaction products using a solvent system containing dichloroethane, methanol, and sodium phosphate buffer. The identities of the products separated on the coil planet centrifuge were confirmed by thin-layer chromatography and mass spectrometry. This technique may be useful in separating metabolites contained in specimens obtained from diaziquone-treated patients.

Isolation of daunorubicin derivatives by counter-current chromatography with the horizontal flow-through coil planet centrifuge

Isolation of daunorubicin derivatives by counter-current chromatography with the horizontal flow-through coil planet centrifuge

Separation of prostaglandins using the new horizontal flow-through coil planet centrifuge

Prostaglandins and their metabolites have been separated using the new horizontal flow-through coil planet centrifuge to perform countercurrent chromatography. The system is a continuous-flow system which provides separations similar to column and thin-layer chromatography without the use of solid supports. Either phase of a two-phase solvent system can be used to elute compounds with the low pressure produced by a metering pump. Separations were produced in PTFE tubing, 0.55 mm i.d. (wound in 0.68 cm helical diameter) with a total capacity of 24 ml. A solvent system of chloroform/acetic acid/water (2 : 2 : 1, v/v) was used at flow rates of 2.4 and 6 ml/h and at a rotation of 500 rev./min. Microgram quantities of prostaglandins E 2, F 2α, A 2, and B 2 and thromboxane B 2 can be separated. This method of can be scaled up to 260-ml columns and thromoboxane B 2 can be separated. This method can be scaled up to 260-ml columns with comparable results.

New horizontal flow-through coil planet centrifuge for counter-current chromatography : III. Separation and purification of dinitrophenyl amino acids and peptides

The new type of horizontal flow-through coil planet centrifuge enables both preparative and analytical separations with partition efficiencies comparable to liquid chromatography. This counter-current chromatographic method was evaluated by the separation and purification of DNP-amino acids and various kinds of peptides using common two-phase solvent systems. The peptide samples separated include commercially available oligopeptides and two synthetic peptides of five and fifteen amino acids. A partition efficiency of up to several thousand theoretical plates was obtained with DNP-amino acids. Purification of the synthetic peptides, which produced problems with conventional liquid chromatography due to contamination and poor solubility, was successfully performed on a preparative scale with a reasonable degree of recovery.

New horizontal flow-through coil planet centrifuge for counter-current chromatography : II. The apparatus and its partition capabilities

The new horizontal flow-through coil planet centrifuge allows continuous elution through a pair of coiled separation columns without the use of rotating seals. Design of the apparatus enables each column to undergo a specific mode of synchronous planetary motion which produces a charateristic pattern of the acceleration field as described in Part I. Using typical two-phase solvent systems, retention of the stationery phase and partition efficiency were studied for each column under various revolutional speeds and flow-rates. The results indicate that one column enables preparative-scale separations and the other, analytical-scale separations both with a high partition efficiency comparable to that obtained refined liquid chromatography.