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One step purification of corilagin and ellagic acid from Phyllanthus urinaria using high-speed countercurrent chromatography.

High-speed countercurrent chromatography (HSCCC) has been successfully applied to the preparative separation of corilagin and ellagic acid in one step from the Chinese medicinal plant Phyllanthus urinaria L. by use of direct and successive injections of a crude methanolic extract. Some aspects concerning the practical use of this technique in the described application are considered.

Countercurrent Distribution↗

High-speed countercurrent chromatography as a valuable tool to isolate C-glycosylflavones from Cecropia lyratiloba Miquel.

A new apigeninglycoside, apigenin 6-C-galactosyl-6"-O-beta-galactopyranoside (1), isoorientin, and a mixture of orientin and isovitexin were isolated from leaves of Cecropia lyratiloba by high-speed countercurrent chromatography using a solvent system containing ethyl acetate, butanol, methanol and water. The structural elucidation of 1 was based on NMR spectroscopy.

Apigenin↗

Application of atmospheric pressure chemical ionisation mass spectrometry in the analysis of barbiturates by high-speed analytical countercurrent chromatography.

Four barbiturates (barbital, allobarbital, phenobarbital and butalbital) were analysed using high-speed analytical countercurrent chromatography (HSACCC) and high-performance liquid chromatography (HPLC) interfaced with mass spectrometry, using negative mode atmospheric pressure chemical ionisation (APCI). The polar biphasic solvent system of butyronitrile/acetonitrile/water (1:1:1) was used, in the upper-stationary, lower-mobile mode of operation, at a flow rate of 1 mL/min and a rotational speed of 1200 rpm, equating to an applied "g"-field of 177 g. The fractional stationary phase retention (S(F)) was 0.58. Representative mass spectral data are presented from the HPLC and the HSACCC analyses. Structural information was obtained using source-induced fragmentation at increased source block voltages. The effect of increasing g-field on chromatographic resolution is illustrated using the binary base system of butyronitrile/water (1:1), under electrospray ionisation.

Algorithms↗

Separation and characterisation of five polar herbicides using countercurrent chromatography with detection by negative ion electrospray ionisation mass spectrometry.

Five polar herbicides were separated and characterised using high-speed analytical countercurrent chromatography (HSACCC) in conjunction with online electrospray mass spectrometry (ESI-MS). The countercurrent chromatography used a standard isocratic biphasic solvent system of hexane/ethyl acetate/methanol/water in reverse phase to effect the separation of these five environmentally important compounds. The chromatograph was coupled to a triple quadrupole mass spectrometer via a standard electrospray liquid chromatography interface that was able to give mass spectra in negative ion mode of each compound. Limits of detection are reported for this series of compounds along with representative negative ion ESI-MS data and calibrations for the separation.

Chromatography, High Pressure Liquid↗

O2 gradients and countercurrent exchange in the cat vitreous humor near retinal arterioles and venules.

Recessed cathode O2 microelectrodes were used to measure spatially detailed oxygen tension (PO2) gradients in the vitreous humor near the cat retina. Measurement sites (n = 41 in 8 cats) included single arterioles and venules and parallel vessel pairs. Mean vitreous PO2 was 37.9 +/- 1.5 (SE) Torr. Close to the retinal surface (approximately 200 microns), PO2 was found to be both higher and lower than the vitreous PO2, depending on the proximity of the microelectrode tip to retinal vessels. Both positive (inward) and negative (outward) O2 fluxes (JO2) were measured, consistent with the anatomy and expected boundary conditions in the eye. The PO2 at the closest approach above arterioles was 55.2 +/- 2.3 Torr, significantly higher than in the vitreous (P < 0.0001). All arterioles had outward JO2 with an overall mean of -2.58.10(-6) ml O2/sec/cm2. Some of the venules were also losing O2, but at much lower rates than arterioles. Several venules were gaining O2. Countercurrent transport (A-V shunting) was also seen between vessel pairs. Our experimental results allow theoretical predictions to be made for the axial drop in blood PO2 along an arteriole as a function of blood flow.

Animals↗

Enzyme-enzyme interaction in the chloroplast: glyceraldehyde-3-phosphate dehydrogenase, triose phosphate isomerase and aldolase.

Apparent physical interaction between pea chloroplast (Pisum sativum L.) glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.13) and aldolase (EC 4.1.2.13) is seen in phase-partitioning, fluorescent-anisotropy and isoelectric-focusing experiments. Similarly, results obtained in phase-partitioning and isoelectric-focusing experiments indicate physical interaction between aldolase and triose-phosphate isomerase (EC 5.3.1.1). Kinetic experiments suggest that both aldolase-bound glyceraldehyde-3-phosphate can act as substrate for glyceraldehyde-3-phosphate dehydrogenase. These results are consistent with the notion that there is interaction between these three enzymes both during photosynthetic CO2 fixation and during glycolysis in the chloroplast.

Amino Acid Sequence↗

The role of the cell cycle in differentiation of the cellular slime mould Dictyostelium discoideum.

During development and differentiation of the cellular slime mould Dictyostelium discoideum there appears to be a relationship between the cell cycle and cell fate: amoebae halted in G2 phase during early development differentiate into spores whereas stalk cells are formed from amoebae halted in G1 phase. It is proposed that this is because a major effect of the cell cycle is to generate heterogeneity in the cell surface properties of the developing amoebae.

Cell Cycle↗

The effects of pressure on the water permeability of the descending limb of Henle's loops of rabbits.

Descending limbs of Henle's loops from rabbits were perfused in vitro. Using techniques where the collecting pipets permitted cannulation of the tubule, we were able to maintain reasonable flow rates at lower perfusion reservoir heights than are required with a conventional "Sylgard seal" pipet. The bath was either isosmotic to the perfusate, or was made 300 mOsm hyperosmotic using urea. Net water reabsorption did not occur in tubules perfused at low pressure (average reservoir height = 26 cm H2O) even when the bath was hyperosmotic: delta Jv = -0.06 +/- 0.18 nl/min (n = 7). Observed increases in sodium concentration and osmolality of collected fluid, when the bath was made hyperosmotic, were 16 +/- 8 mM (n - 7) and 254 +/- 38 mOsm (n = 7), respectively. Presumably the large increase in osmolality of the collected fluid was due to entrance of urea. When the "Sylgard seal" collecting end was utilized higher perfusion reservoir heights had to be used to maintain flow (mean height 66 cm H2O). These tubules were highly permeable to water as reported by others for this tubule segment. In the presence of a hyperosmotic bath water extrusion resulted in a dramatic increase in the osmolality of the collected fluid (312 +/- 5 mOsm; 7 tubules) which was almost completely accounted for by an increase in sodium concentration (153 +/- 8 mmole/l; 6 tubules). The 14C urea permeability (measured lumen to bath) of descending limbs in a 300 mOsm bath was 0.64 x 10(-7) cm2 . s-1 +/- 0.23 x 10(-7) (11 tubules). When the bath was made hyperosmotic using urea or raffinose the 14C urea permeability increased significantly.

Animals↗

[New chromatography. Technics for the isolation and separation of natural products].

Besides high-performance liquid chromatography (HPLC) which is now a well-established and currently used technique, several emerging methods for the isolation and separation of natural products are receiving considerable attention. Centrifugal thin-layer chromatography is a very rapid technique, but limited in resolution. Of special interest are the recently developed support-free liquid-liquid chromatography methods such as droplet counter-current chromatography (DCCC) and rotation locular counter-current chromatography (RLCC). This latter method was applied to the separation of the enantiomers of (+/-)-norephedrine.

Chromatography↗

Fixation with even small quantities of glutaraldehyde affects red blood cell surface properties in a cell- and species-dependent manner. Studies by cell partitioning.

Partitioning in charge-sensitive dextran-poly(ethylene glycol) aqueous phase systems reveals that fixation with even small concentrations of glutaraldehyde (e.g., 0.1% w/v) changes the surface properties of cells. While fixation with larger concentrations of glutaraldehyde (i.e., 1.85%) increases erythrocyte partition ratios, the effect of lower glutaraldehyde concentrations on the partition ratios appears to be species-specific. The differential effect of glutaraldehyde on rat reticulocytes and erythrocytes indicates that fixation is also cell-dependent. These data, together with the previous report that glutaraldehyde fixation does not change the characteristic relative partition ratios of rat mature erythrocytes of different cell ages, suggest that the nature and extent of glutaraldehyde alteration of cell surfaces must, in each case, be empirically evaluated.

Aldehydes↗

Analysis by partitioning in aqueous two-phase systems of the loss of transferrin-binding capacity during maturation of rat reticulocytes.

A decrease in the number of binding sites for 125I-transferrin, without an apparent modification of the association constant, has been observed during the maturation of reticulocytes into erythrocytes. As an experimental model, different red cell populations from phenylhydrazinic anaemic rates (95% to 12% reticulocyte-rich) have been used. The fractionation by multiple partition in two-phase systems of these red cell populations has been applied here to show the relationship between number of transferrin receptors and rate of reticulocyte maturation.

Anemia↗