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PubMed1986

The dynamics of phase partition. A study of parameters affecting rat liver organelle partitioning in aqueous two-polymer phase systems.

Separation of subcellular organelles by two-phase partition is thought to reflect differential partition of the organelles between the two phases or between one of the phases and the interface. Studies by Fisher and colleagues [Fisher & Walter (1984) Biochim. Biophys. Acta 801, 106-110] suggest that cell separation by phase partition is a dynamic process in which the partition changes with time. This is mainly due to association of the cells with sedimenting droplets of one phase in the bulk of the other. Rat liver organelle partition was studied to determine whether the same dynamic behaviour is observed. Partition was clearly time-dependent during 24 h at unit gravity, and was also affected by altering the volume ratio of the two phases and the duration of phase mixing. These results indicate that, as with cells, the partition of organelles between phases is a dynamic process, and is consistent with the demonstration that organelles adhere to the phase droplet surfaces. Optimization of the volume ratio between phases may lead to significant processing economies. Organelle sedimentation in the upper phase was significantly faster than in the isoosmotic sucrose. Theoretical modelling of apparent organelle sizes indicates that aggregation occurs in the poly(ethylene glycol)-rich upper phase. This phenomenon is likely to limit the use of this technique in organelle separations unless means can be found to decrease aggregation.

Acetylglucosaminidase↗

PubMed1990

Phorbol-ester-induced down-regulation of protein kinase C in mouse pancreatic islets. Potentiation of phase 1 and inhibition of phase 2 of glucose-induced insulin secretion.

The influence of down-regulation of protein kinase C on glucose-induced insulin secretion was studied. A 22-24 h exposure of mouse pancreatic islets to the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA; 0.16 microM) in RPMI 1640 culture medium (8.3 mM-glucose, 0.43 mM-Ca2+) abolished TPA (0.16 microM)-induced insulin secretion and led to a potentiation of phase 1 and a decrease in phase 2 of glucose-induced insulin secretion. Thus, although the total insulin release during 40 min of perfusion with glucose (16.7 mM) (45-85 min) was unaffected, the percentage released during phase 1 (45-55 min) was increased from 12.9 +/- 1.5 (4)% in controls to 35.8 +/- 3.9 (4)% in TPA-treated islets (P less than 0.01), and the percentage released during phase 2 (65-85 min) was decreased from 63.2 +/- 3.9 (4)% to 35.3 +/- 1.4 (4)% (P less than 0.005). In contrast, TPA exposure in TCM 199 medium (5.5 mM-glucose, 1.26 mM-Ca2+) caused a total abolition of both phases 1 and 2 of glucose-induced secretion. However, inclusion of the alpha 2-adrenergic agonists adrenaline (10 microM) or clonidine (10 microM), or lowering of the Ca2+ concentration in TCM 199 during down-regulation, preserved and potentiated phase 1 of glucose-induced secretion. Furthermore, perifusion of islets in the presence of staurosporine (1 microM), an inhibitor of protein kinase C, potentiated phase 1 and inhibited phase 2 of glucose-induced secretion. In addition, down-regulation of protein kinase C potentiated phase 1 and inhibited phase 2 of carbamoylcholine (100 microM)-induced insulin secretion at 3.3 mM-glucose, and abolished the potentiating effect of carbamoylcholine (100 microM) at 16.7 mM-glucose. These results substantiate a role for protein kinase C in insulin secretion, and suggest that protein kinase C inhibits phase 1 and stimulates phase 2 of both glucose-induced and carbamoylcholine-induced insulin secretion.

Alkaloids↗

PubMed1989

Phase determination of the circadian rhythm of conidiation in heterocaryons between two out-of-phase mycelia in Neurospora crassa.

Neurospora grows vegetatively as a syncytium in which multiple nuclei exist within a connected cytoplasm. Because of the ability of separate and distinct mycelia to fuse, the possibility exists of generating heterocaryotic cultures in which the nuclei and cytoplasms of two different strains are comingled into the same syncytium. We have used such heterocaryons, in which the component parts differed with respect to their circadian clock phase, to examine whether or not clock-dominant phases exist in the circadian cycle. To this end, the phase subsequent to the formation of heterocaryons by pairs of mycelial discs that are initially at different circadian phases was examined in Neurospora crassa. The resulting phase was an average of the parent phases in many cases, but was sometimes observed to correspond more closely to just one of the original parental phases. In these cases, we did not observe any dominant phases in the circadian cycle; the phase of a particular parent disc was more dominant in the heterocaryon when the proportion of the nuclei from that parent was greater in the heterocaryon. In some instances, which occurred mostly when the difference in phase of the parental discs was large, the resultant phase could not be related in a simple way to the parental phases. An interpretation based on a limit cycle model of the circadian oscillation is possible.

Cell Cycle↗

PubMed1987

Phase separation and hexagonal HII phase formation by gramicidins A, B and C in dioleoylphosphatidylcholine model membranes. A study on the role of the tryptophan residues.

The role of the tryptophan-residues in gramicidin-induced HII phase formation was investigated in dioleoylphosphatidylcholine (DOPC) model membranes. 31P-NMR and small angle X-ray diffraction measurements showed, that gramicidin A and C (in which tryptophan-11 is replaced by tyrosine) induce a similar extent of HII phase formation, whereas for gramicidin B and synthetic analogs in which one tryptophan, either at position 9 or 11 is replaced by phenylalanine, a dramatic decrease of the HII phase inducing activity can be observed. Modification of all four tryptophans by means of formylation of the indole NH group leads to a complete block of HII phase formation. Sucrose density centrifugation experiments on the various peptide/lipid samples showed a quantitative incorporation of the peptide into the lipid. For all samples in a 1/10 molar ratio of peptide to lipid distinct bands were found, indicative of a phase separation. For the gramicidin A'/DOPC mixture these bands were analyzed and the macroscopic organization was determined by 31P-NMR and small-angle X-ray diffraction. The results demonstrate that a quantitative phase separation had occurred between a lamellar phase with a gramicidin/lipid ratio of 1/15 and a hexagonal HII phase, which is highly enriched in gramicidin. A study on the hydration properties of tryptophan-N-formylated gramicidin in mixtures with DOPC showed that this analog has a similar dehydrating effect on the lipid headgroup as the unmodified gramicidin. In addition both the hydration study and sucrose density centrifugation experiments showed that, like gramicidin also its analogs have a tendency to aggregate, but with differences in aggregation behaviour which seemed related to their HII phase inducing activity. It is proposed that the main driving force for HII phase formation is the tendency of gramicidin molecules to self-associate and organize into tubular structures such as found in the HII phase and that whether gramicidin (analogs) form these or other types of aggregates depends on their tertiary structure, which is determined by intra- as well as intermolecular aromatic-aromatic stacking interactions.

Gramicidin↗

PubMed1988

Cell cycle phase-specific cDNA libraries reflecting phase-specific gene expression of Ehrlich ascites cells growing in vivo.

Asynchronous populations of Ehrlich ascites tumor cells grown in vivo were separated by centrifugal elutriation into fractions of G1-, S-, and G2/M-phase cells with less than 10% cross-contamination. Cytoplasmic mRNA from phase-synchronous cells was used to prepare cDNA which was ligated with bacteriophage lambda gt10 arms and amplified in Escherichia coli C600 hfl-. EcoRI digests of DNA isolated from the sublibraries (G1, S, G2/M) were submitted to Southern hybridizations with radiolabeled probes either (a) for genes whose phase-specific expression is clearly documented, thymidine kinase, dihydrofolate reductase, and thymidylate synthase, or (b) for genes whose change of expression during the cell cycle is likely, lamin C, beta-actin, alpha- and beta-tubulin, c-myc, c-fos, p53. The cDNA sequences for genes of group (a) were found to be significantly enriched in DNA of the S-phase library indicating that the cell cycle phase-specific patterns of the respective mRNA levels are conserved in the sublibraries. Sequences belonging to group (b) were also found to be enriched in DNA isolated from the sublibraries: c-fos in G1 phase, lamin C, beta-actin, tubulins, c-myc in S phase, and p53 in G1/S phase. The unexpected prevalence of c-myc and alpha-tubulin in the S-phase library is supported by Northern analysis of RNA from phase-synchronous cells. Non-phase-specific, randomly chosen sequences hybridized equally strong with DNA isolated from the different sublibraries. No significant changes of the patterns of hybridization signals were observed with DNA from different amplifications of the sublibraries when analyzed with the same DNA probe indicating that the cDNA complexities are well conserved during amplifications. Consequently, the sublibraries are useful to obtain information about the cell cycle phase-specific expression of mRNAs for other genes of interest. Since the sublibraries reflect mRNA levels of the cells growing in vivo they supply data on the physiological in vivo pattern of gene expression undisturbed by potentially unphysiological in vitro conditions.

Animals↗

PubMed1992

Cubic phases of lipid-containing systems. The structure of phase Q223 (space group Pm3n). An X-ray scattering study.

The hexagonal (H) and the cubic (Q223) phases of the systems dodecyltrimethylammonium chloride-water and palmitoyllysophosphatidy choline-water have been studied by X-ray scattering techniques. The signs of the reflections of phase H were determined by a systematic study as a function of the water content, those of phase Q223 were assessed using a pattern recognition approach based upon the axiom that the histograms of the electron density maps of phases Q223 and H, extrapolated to the same concentration and properly normalized in scale and shape, are very similar to each other. In the case of phase Q223, all the sign combinations (the phi-sets) compatible with the observed reflections were generated, and each of the corresponding histograms was compared with the histogram of the map of phase H. One novelty of this work is the use of a highly sensitive criterion to estimate the similarity of the histograms, namely the distance in the six-dimensional space of the moments [mean value of (delta rho)n]1/n, for 3 greater than or equal to n greater than or equal to 8. In the two systems, the use of this criterion has led to the unambiguous choice of one electron density map. The maps show that the structure of phase Q223 consists of disjointed micelles (of type I), belonging to two different classes: those of one class are quasi-spherical in shape and are centered at the points a, those of the other class are disc-shaped and are centred at the points c. The results of this work rule out a structure formed by a cage-like distribution of rods enclosing a set of quasi-spherical micelles and is consistent with previous proposals. This is the second example, after that of phase Q227, of a micellar cubic phases in lipid-containing systems; all the known examples of phase Q223 are of type I, those of phase Q227 of type II.

Freeze Fracturing↗

PubMed1992

Biomechanical properties of duodenal wall and duodenal tone during phase I and phase II of the MMC.

We used a new method, impedance planimetry, to look at variations in compliance, tone, and distension-induced peristaltic activity during phase I and phase II of the migrating myoelectric complex (MMC) in the human duodenum. A balloon was inflated stepwise with pressures up to 30 cmH2O in the duodenum, while the pressure and balloon cross-sectional area (CSA) were measured simultaneously. The biomechanical wall parameters were calculated from these measurements. Nine duodenal phase IIIs were recorded in six subjects. A balloon pressure of 20 cmH2O induced a smaller CSA in early phase I [266 (236-324) mm2] than in late phase II [385 (276-474) mm2] (P < 0.05). Balloon distensions elicited no contractions in phase I, whereas they increased contractile activity 60% (P < 0.05) proximal to the balloon and 4% distal to the balloon in late phase II. Step distensions in phase I with balloon pressures between 10 and 30 cmH2O increased the CSA from 40 (30-81) to 645 (603-704) mm2. It increased circumferential wall tension from 35 (28-63) to 429 (402-466) mm x cmH2O and the pressure elastic modulus from 9.7 (9.0-14.7) to 33.8 (27.6-33.8) cmH2O, respectively. Thus compliance differs from phase I to phase II. This is most likely caused by increased smooth muscle tone during phase I. Duodenal wall stiffness increases with the balloon pressure applied.

Adult↗

PubMed1990

Signal-to-noise in phase angle reconstruction: dynamic range extension using phase reference offsets.

The dynamic range of phase-reconstructed magnetic resonance images is compared to that of magnitude-reconstructed images. From analysis of propagation of errors, the phase angle noise is phase-independent and given in radians by sigma ([I])/[I], the noise-to-signal ratio of the corresponding magnitude-reconstructed image. As the phase can range from minus pi to pi, the phase angle dynamic range is 2 pi times that of the signal magnitude. These results agree with experiment, verifying that the noise in the two receiver channels is uncorrelated. An artifact-free technique is presented for correcting phase spillover, which further extends the phase angle dynamic range. The reconstruction-based reference phase is adjusted on a local basis so that the boundary of phase wraparound is reconstructed near the center of the [- pi, pi] interval. For a particular flow study, the phase signal-to-noise was extended over twofold by spillover correction, to a value 15 times that of the magnitude signal-to-noise.

Algorithms↗

PubMed1985

Phase transitions and fatty acid spin label behavior in interdigitated lipid phases induced by glycerol and polymyxin.

Glycerol and polymyxin have been shown by X-ray diffraction to induce interdigitated bilayers in phosphatidylcholine (PC) and phosphatidylglycerol (PG), respectively (McDaniel, R.V., et al. (1983) Biochim. Biophys. Acta 731, 97-108; Ranck, J.-L. and Tocanne, J.-F. (1982) FEBS Lett. 143, 175-178). In the present study we have investigated the phase behavior of PC and PG in the presence of glycerol and polymyxin by differential scanning calorimetry and the use of fatty acid spin labels. Interdigitation causes a large increase in the order parameter of a fatty acid spin labeled near the terminal methyl, 16-doxylstearate, so that it was similar to that of a fatty acid labeled much closer to the polar head group region, 5-doxylstearate. Thus interdigitation abolishes the fluidity gradient found in a non-interdigitated bilayer. 16-Doxylstearate may be useful in detecting interdigitation of lipid bilayers caused by other substances. The different samples all went through two transitions on heating or cooling, or both. However, use of the fatty acid spin label showed that the molecular events during these transitions varies for different samples. The results suggested that PC-glycerol freezes from the liquid-crystalline phase into a non-interdigitated gel phase. This subsequently becomes interdigitated upon lowering the temperature a few degrees, in a low enthalpy transition. PG-polymyxin shows a similar behavior except that the enthalpy of the non-interdigitated gel to interdigitated phase transition is greater and the transition is reversible on heating. Thus on heating PG-polymyxin first goes through a transition from the interdigitated phase to a non-interdigitated gel phase and then, in a separate transition, to the liquid-crystalline phase. This occurs because the fatty acid chains in the presence of polymyxin become too disordered with increase in temperature to maintain the interdigitated state. PG-glycerol goes into the interdigitated state less readily than the other mixtures. If cooled rapidly, PG-glycerol freezes into a metastable phase which is more disordered than the interdigitated phase. It goes into the interdigitated phase in an exothermic transition on heating. An increase in fatty acid chain length causes greater steric hindrance to interdigitation but also increases the stabilizing energy gained by interdigitation.

Calorimetry, Differential Scanning↗

PubMed1985

Phase 1 and Phase 2 drug metabolism in isolated epidermal cells from adult hairless mice and in whole human hair follicles.

A sensitive fluorimetric assay to determine both Phase 1 (oxidation) and Phase 2 (conjugation) drug metabolism in epidermal cells isolated from hairless mice, using ethoxycoumarin as a model substrate, is described. Ethoxycoumarin was metabolized by isolated epidermal cells via dealkylation to 7-hydroxycoumarin (7-OHC) and subsequent conjugation. Phase 1 metabolites were extracted in ether from the aqueous incubation media, back extracted into sodium hydroxide and determined fluorimetrically. Conjugated metabolites remaining in the aqueous phase were hydrolysed by the action of beta-glucuronidase and extracted and determined in a similar manner. The production of free 7-OHC by isolated epidermal cells was biphasic at all substrate concentrations tested, exhibiting an initial linear increase followed by a plateau phase. The plateau phase was attributable to the conjugation of 7-OHC produced in situ. Metabolism was inhibited by SKF 525A, carbon monoxide, and alpha-naphthoflavone. Endogenous supplies of reducing equivalents in the form of NADPH were adequate to attain maximal rates of metabolism. With human hair follicles both Phase 1 and Phase 2 activity was detectable in 7 out of 11 subjects. The assay has the advantages of being sensitive, producing single defined metabolites from both Phase 1 and Phase 2 metabolism; is readily adaptable to human skin samples.

Animals↗

PubMed1986

Inverted micellar intermediates and the transitions between lamellar, cubic, and inverted hexagonal amphiphile phases. III. Isotropic and inverted cubic state formation via intermediates in transitions between L alpha and HII phases.

Inverted cubic and isotropic phases have been observed in phospholipid and glycolipid systems. These phases exhibit characteristic morphologies in freeze-fracture electron micrographs, isotropic 31P-NMR resonances and (in some cases) cubic X-ray diffraction patterns. It is proposed here that these phases may form from the same intermediates that are involved in lamellar/inverted hexagonal (L alpha/HII) phase transitions, and that it is possible that these cubic and isotropic phases are metastable. According to a kinetic theory of L alpha/HII phase transitions, intermediates in such transitions can form structures known as interlamellar attachments (ILAs). It is shown that ILAs should form in large numbers during L alpha/HII transitions in systems like those reported to form inverted cubic or isotropic structures. ILAs cannot readily assemble into either the HII phase or well-ordered arrays of L alpha phase bilayers, and represent a kinetic trap for intermediates in L alpha/HII transitions (although it is possible that they are marginally more stable in a thermodynamic sense than the L alpha phase in a small temperature range below TH). It is also shown that arrays of ILAs should form metastable arrays with the same morphology and isotropic 31P-NMR resonances that are observed in isotropic and inverted cubic states. In particular, under some circumstances ILAs will assemble into a structure identical to the bicontinuous inverted cubic phase previously described in monoglycerides and very similar in morphology to structures observed in phospholipid systems. Finally, since isotropic and cubic states form from ILAs, which also can mediate fusion of unilamellar vesicles, unilamellar vesicles should fuse to at least some extent under the same conditions in which multilamellar samples of the same lipid form isotropic or inverted cubic states. This correlation has been observed.

Glycolipids↗

PubMed1987

Leishmania braziliensis: protein, carbohydrate, and antigen differences between log phase and stationary phase promastigotes in vitro.

When Leishmania species are grown in vitro, parasites from the stationary phase differ from those in log phase growth in being more infective and more resistant to complement and macrophage mediated killing. In the present study, log phase and stationary phase promastigotes of Leishmania braziliensis panamensis were compared at the molecular level. Differences in polypeptide and glycoprotein composition and antigenicity between log and stationary phase promastigotes of L. b. panamensis were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting; the former showed that two polypeptides were unique to log phase promastigotes and one was unique to stationary phase promastigotes. There were also differences in surface lectin binding characteristics of log and stationary phase promastigotes. Live stationary phase promastigotes bound more concanavalin and lentil lectin than log phase promastigotes, indicating a greater number of mannose residues on their surfaces.

Animals↗

PubMed1986

Role of head group structure in the phase behavior of amino phospholipids. 1. Hydrated and dehydrated lamellar phases of saturated phosphatidylethanolamine analogues.

Analogues of dimyristoylphosphatidylethanolamine (DMPE) have been prepared with head groups modified by N-alkylation, alkylation of carbon 2 of the ethanolamine group, or interposition of extra methylene segments between the phosphoryl and amino groups. The phases formed by these lipids in aqueous dispersions have been examined by high-sensitivity differential scanning calorimetry and Raman spectroscopy. All of the DMPE analogues examined, excepting N-methyl-DMPE but including N-ethyl-DMPE, form hydrated gel phases that are metastable with respect to a dehydrated "high-melting" solid phase that has been observed previously for DMPE itself. The properties and the conditions of formation of this high-melting phase are qualitatively distinct from those of the "subgel" phase, which is observed for dipalmitoylphosphatidylcholine and for some of the DMPE analogues examined in this study. The high-melting phases of different DMPE analogues all exhibit similarly tight packing of the acyl chains, which however do not pack according to a single type of subcell that can be universally and specifically associated with this phase. Increasing the size of the PE head group invariably decreases the melting temperature of the hydrated gel phase, even when the normal hydrogen-bonding capability of the head group is preserved. By contrast, addition of larger alkyl substituents to either the amino group or carbon 2 of the ethanolamine moiety substantially increases the transition temperature of the high-melting solid phase, indicating that the contributions of the head group to the energies of the hydrated gel and the high-melting phases are fundamentally different. Our results suggest that the head group structural requirements for a neutral phospholipid to form stable hydrated bilayers are rather stringent, a fact that may explain the overwhelming predominance of only a few such head group structures in most natural membranes.

Calorimetry, Differential Scanning↗

PubMed1990

[Phase 3 and phase 4 block in the accessory pathway].

Pacemaker activity at a site proximal to the area of a conduction disturbance has been postulated as depressing conduction in late diastole (phase 4 block). To elucidate the correlation between the site of phase 4 depolarization and that of a conduction disturbance, we examined seven patients with intermittent Wolff-Parkinson-White syndrome who had tachycardia (phase 3) and bradycardia (phase 4) dependent on block in the accessory pathway. In each patient, antegrade conduction over the accessory pathway was absent at the sinus rate. During premature atrial stimulation a "window" of accessory pathway conduction was identified in each patient. The outer limit of the window ranged from 420 to 670 ms; the inner limit, from 330 to 620 ms. The duration of the window ranged from 20 to 160 ms. Four patients with orthodromic atrioventricular reentrant tachycardia exhibited preserved retrograde accessory pathway conduction. In one patient with unsustained orthodromic atrioventricular reentrant tachycardia, retrograde accessory pathway conduction also exhibited phase 3 and phase 4 blocks. The remaining two patients had no retrograde accessory pathway conduction. In two patients, retrograde concealed conduction in the accessory pathway induced by ventricular stimulation eliminated a phase 4 block in antegrade accessory pathway conduction. These results suggest that 1) an antegrade phase 4 block may have phase 4 depolarization in the accessory pathway and block at the ventricular insertion of the accessory pathway; 2) a retrograde phase 4 block may have a conduction disturbance at the atrial insertion of the accessory pathway; and 3) a complete retrograde block may occur at the atrial insertion of the accessory pathway.

Adolescent↗

PubMed1992

The inverted hexagonal phase is more sensitive to hydroperoxidation than the multilamellar phase in phosphatidylcholine and phosphatidylethanolamine aqueous dispersions.

The effect of phase behaviour (hexagonal II phase and lamellar phase) on the peroxidation of membrane phospholipids has been investigated in dilinoleoyl phosphatidylcholine (DLPC)/dilinoleoyl phosphatidylethanolamine (DLPE) aqueous dispersions. Peroxidation was initiated with a water-soluble radical inducer 2,2'-azobis (2-amidino-propane) dihydrochloride (AAPN). The phospholipid morphology was monitored by 31P-nuclear magnetic resonance (NMR). Phospholipid hydroperoxides (PCOOH and PEOOH) were determined by chemiluminescence high-performance liquid chromatography (CL-HPLC). In pH-induced phase transition systems, DLPE in the bilayer state was much less oxidized than in the hexagonal II state. In composition-induced phase transition systems, the formation of total hydroperoxides and the consumption of alpha-tocopherol in the hexagonal II phase were greater than in the bilayer phase. These data suggest that the hexagonal II phase is more sensitive to hydroperoxidation than the bilayer phase in phospholipid aqueous dispersions.

Chromatography, High Pressure Liquid↗

PubMed1992

Interneuronal interaction between members of quadrature phase and anti-phase pairs in the cat's visual cortex.

Interactions between adjacent simple cells recorded simultaneously from the same microelectrode placement were studied by correlational analysis. The receptive fields of pairs of such cells exhibit either 90 degrees (quadrature phase) or 180 degrees (anti-phase) phase relationships. We now show that the majority of quadrature phase pair members do not receive common input from the immediately precedent stage along the visual pathway, nor do these cells interact with each other. The anti-phase pairs show relatively strong mutual inhibition. These results suggest that each of the physically adjacent phase-related simple cells receives excitatory input from a distinct group of pre-cortical cells, and that mutual inhibitions between members of anti-phase pairs are used to construct the inhibitory subzones of these cells. We propose a model which incorporates these new results and provides a parsimonious explanation for the construction of both quadrature phase and anti-phase pairs.

Animals↗

PubMed1988

Effect of rapidity of phase separation on the efficiency of cell fractionation by partitioning in aqueous two-phase systems.

Partitioning in two-polymer aqueous phase systems is an established method for the separation, purification and characterization of biomaterials. Because of the relatively slow settling rates of these phases, a consequence of the slight difference in density between them, effort has been directed to speeding up phase separation by various means (e.g., the development of a thin-layer countercurrent distribution apparatus). This has resulted in the more rapid processing of materials. Unlike soluble materials, biological particulates (e.g., cells) generally partition between one of the bulk phases and the interface. The mechanism of cell partitioning involves cell-specific adsorption to droplets of one phase suspended in the other, subsequent to phase mixing, and the delivery of adsorbed cells to the bulk interface as the droplets settle. In this communication we show, using erythrocytes as a model, that speeding up phase separation is counterproductive when partitioning cells and results in reduced efficiency of their separation or subfractionation. The most likely reason for this result is that increasing the speed of phase settling removes the droplets of one phase suspended in the other more rapidly than cells can attach to them, thereby interfering with the mechanism whereby cells partition.

Animals↗

PubMed1992

Temperature dependence of retention in reversed-phase liquid chromatography. 1. Stationary-phase considerations.

The retention mechanism in reversed-phase liquid chromatography (RPLC) has been investigated by examining the temperature dependence of retention, with emphasis on the role of the stationary phase in the retention process. Both chromatographic temperature studies and differential scanning calorimetry were used to examine the role of alkyl chain bonding density on the retention mechanism in RPLC. Phase transitions of reversed-phase stationary phases were observed at bonding densities greater than 2.84 mumol/m2. Thermodynamic constants for the transfer of a solute from the mobile phase to the stationary phase (delta H degrees and delta S degrees) were calculated for low bonding density columns, and comparison of these values to previously reported values for the partitioning of a nonpolar solute from the bulk organic liquid to water indicated that the chromatographic retention process is not well-modeled by bulk-phase oil-water partitioning processes. In addition, this data showed that the entropic contribution to retention becomes more significant with respect to the enthalpic contribution as the stationary-phase bonding density is increased, providing additional support that partitioning, rather than adsorption, is the relevant model of retention.

Chromatography, High Pressure Liquid↗