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PubMed2001

The use of water structure breakers, urea and guanidium chloride, new mobile phase modifiers in reversed-phase partition high-performance liquid chromatography.

Organic solvent-free mobile-phase systems in ion-pair reversed-phase partition high-performance liquid chromatography (IPRP-HPLC) are demonstrated; using urea at 3.0-7.0 molal (mol kg-1) as a modifier in a mobile phase on an octadecylsilanized silica column, four nitrophenolates and metal 4-(2-pyridilazo)resorcinol (PAR) chelates (in PAR chelates system an aqueous mobile phase with 15 wt% methanol was used) were separated rapidly within 6 min at no sacrifice to the separation efficiency. On the addition of urea in the mobile phase, reduced retention times of nitrophenolates and naphthalenesulfonates and also diminution of the height equivalent to a theoretical plate were observed. The addition of urea and guanidium chloride (GuCl) in the mobile phase gave rise to a decrease in the mobile phase volume; in turn, this meant an increased volume of the stationary phase. As the concentration of urea and GuCl in the mobile phase increased, the volume of the mobile phase in the column decreased within about 70% and 40% at 7.0 molal of urea and GuCl, respectively. A decrease in the mobile phase volume suggests an increase in the extent of solvation of the bonded hydrocarbon chain of the stationary phase. The possible explanations for the LC behavior with the urea and GuCl are turned into reduction of hydrophobic interaction in LC processes, solute partitioning and entangling of alkyl chain brushes, with the addition of urea. The water structure breakers, urea and GuCl, most likely affect the solvation states of both solute molecules and the hydrocarboneous stationary phase by changing the nature of the water solvent, which provides a new technique for fine tuning of the LC resolution of the analytes.

Anions↗
PubMed1999

Phase Behavior and Salt Partitioning in Two- and Three-Phase Anionic Surfactant Microemulsion Systems: Part II, Partitioning of Salt.

The partitioning of salt (chlorides of Na, K, Ca, Mg) between an excess water phase and a microemulsion phase were studied for two five-component anionic microemulsion systems at 20 degrees C. Three-phase microemulsions contain two phases that include inorganic salts, that is, excess brine and the microemulsion phase. The aim of the investigation was to describe the partitioning of Na(+), K(+), Ca(++), Mg(++), and Cl(-) between water in the microemulsion phase and water in the excess water phase. Both surfactant systems were anionic with Na(+) as counterion. It was found that Na(+) and Cl(-) partitioned more strongly toward the excess water phase while K(+), Ca(++), and Mg(++) show more preference for the microemulsion phase. The validity of considering (NaCl + water) as pseudo-component brine has been examined. Including a chloride depleted region, caused by electrostatic repulsion from the negatively charged surfactant layer, in the surfactant pseudo-component, is found to describe the NaCl partitioning. When discussing partitioning of K(+), Ca(++), and Mg(++) between the microemulsion phase and the excess water phase in these systems, it is important to consider that there are two cations in the solutions, Na(+) as counterion for the surfactant and either K(+), Ca(++), or Mg(++) for the electrolyte. The preferential partitioning toward the microemulsion phase for K(+), Ca(++), and Mg(++) seems to be caused by an ion exchange of Na(+) in the surfactant layer. Copyright 1999 Academic Press.

Journal Article↗
PubMed1992

Erythrocyte partitioning in dextran-poly(ethylene glycol) aqueous phase systems. Events in phase and cell separation.

Early events in the partitioning process which involve characteristic kinetics of cell- and phase-specific interactions and phase separation have been described previously. This paper reports on red cell-phase droplet interactions pertaining at the time of usual phase sampling (i.e., the time at which a clear bulk interface is first apparent) and beyond in cell partitioning and countercurrent distribution experiments. In non-charge-sensitive phase systems close to the critical point, cells can be free or attached to phase droplets. Cells that are free are virtually completely in the top phase, whereas different cell populations that show essentially complete binding to droplets can nevertheless have different partition ratios and be separated, thus reflecting the effects of the difference in the cells' avidity for the phase droplets during the early, elapsed events in partitioning. At higher polymer concentrations (i.e., higher interfacial tensions), the cell populations, completely bound to phase droplets, partition completely to the interface, and consequently cannot be separated. When such systems are made charge-sensitive by the generation of a Donnan potential between the phases or made into affinity systems by the incorporation of PEG ligands (e.g., PEG-palmitate), there is a decrease in the avidity of the cells for phase droplets. The resulting increase in the ratio of free to droplet-bound red cells in the top phase at the time of sampling correlates with an increase in the partition ratio, P, observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1999

Effects of electrostatic interaction on the phase stability and structures of cubic phases of monoolein/oleic acid mixture membranes.

To elucidate effects of electrostatic interactions due to the surface charges on structures and phase stability of cubic phases of lipids, the mixture membranes of monoolein (MO) and oleic acid (OA) (OA/MO membrane) have been investigated by small-angle X-ray scattering method. As increasing OA concentration in the OA/MO membrane, a phase transition from Q(224) to Q(229) phase occurred, and above 1.0 mol% OA, OA/MO membranes were in the Q(229) phase. However, when NaCl concentration in the bulk phase is large, the Q(224) phase is more stable than the Q(229) phase in the OA/MO membranes containing high concentrations of OA. These results indicate that the electrostatic repulsion between the headgroups of these membranes makes the Q(229) phase more stable than the Q(224) phase. We have also found that lowering pH induces a phase transition from Q(229) to Q(224), and at lower pH a phase transition from Q(224) to H(II). These results are discussed in terms of the spontaneous curvature of the monolayer membrane and a lipid packing parameter of the membrane.

Glycerides↗
PubMed1994

Studies of the thermotropic phase behavior of phosphatidylcholines containing 2-alkyl substituted fatty acyl chains: a new class of phosphatidylcholines forming inverted nonlamellar phases.

We have synthesized a number of 1,2-diacyl phosphatidylcholines with hydrophobic substituents adjacent to the carbonyl group of the fatty acyl chain and studied their thermotropic phase behavior by differential scanning calorimetry, 31P-nuclear magnetic resonance spectroscopy, and x-ray diffraction. Our results indicate that the hydrocarbon chain-melting phase transition temperatures of these lipids are lower than those of the n-saturated diacylphosphatidylcholines of similar chain length. In the gel phase, the 2-alkyl substituents on the fatty acyl chains seem to inhibit the formation of tightly packed, partially dehydrated, quasi-crystalline bilayers (Lc phases), although possibly promoting the formation of chain-interdigitated bilayers. In the liquid-crystalline state, however, these 2-alkyl substituents destabilize the lamellar phase with respect to one or more inverted nonlamellar structures. In general, increases in the length, bulk, or rigidity of the alkyl substituent result in an increased destabilization of the lamellar gel and liquid-crystalline phases and a greater tendency to form inverted nonlamellar phases, the nature of which depends upon the size of the 2-alkyl substituent. Unlike normal non-lamella-forming lipids such as the phosphatidylethanolamines, increases in the length of the main acyl chain stabilize the lamellar phases and reduce the tendency to form nonlamellar structures. Our results establish that with a judicious choice of a 2-alkyl substituent and hydrocarbon chain length, phosphatidylcholines (and probably most other so-called "bilayer-preferring" lipids) can be induced to form a range of inverted nonlamellar structures at relatively low temperatures. The ability to vary the lamellar/nonlamellar phase preference of such lipids should be useful in studies of bilayer/nonbilayer phase transitions and of the molecular organization of various nonlamellar phases. Moreover, because the nonlamellar phases can easily be induced at physiologically relevant temperatures and hydration levels while avoiding changes in polar headgroup composition, this new class of 2-alkyl-substituted phosphatidylcholines should prove valuable in studies of the physiological role of non-lamella-forming lipids in reconstituted lipid-protein model membranes.

Biophysical Phenomena↗
PubMed2000

Mechanisms of phase behaviour and protein partitioning in detergent/polymer aqueous two-phase systems for purification of integral membrane proteins.

Detergent/polymer aqueous two-phase systems are studied as a fast, mild and efficient general separation method for isolation of labile integral membrane proteins. Mechanisms for phase behaviour and protein partitioning of both membrane-bound and hydrophilic proteins have been examined in a large number of detergent/polymer aqueous two-phase systems. Non-ionic detergents such as the Triton series (polyoxyethylene alkyl phenols), alkyl polyoxyethylene ethers (C(m)EO(n)), Tween series (polyoxyethylene sorbitol esters) and alkylglucosides form aqueous two-phase systems in mixtures with hydrophilic polymers, such as PEG or dextran, at low and moderate temperatures. Phase diagrams for these mixtures are shown and phase behaviour is discussed from a thermodynamic model. Membrane proteins, such as bacteriorhodopsin and cholesterol oxidase, were partitioned strongly to the micelle phase, while hydrophilic proteins, BSA and lysozyme, were partitioned to the polymer phase. The partitioning of membrane protein is mainly determined by non-specific hydrophobic interactions between detergent and membrane protein. An increased partitioning of membrane proteins to the micelle phase was found with an increased detergent concentration difference between the phases, lower polymer molecular weight and increased micelle size. Partitioning of hydrophilic proteins is mainly related to excluded volume effects, i.e. increased phase component size made the hydrophilic proteins partition more to the opposite phase. Addition of ionic detergent to the system changed the partitioning of membrane proteins slightly, but had a strong effect on hydrophilic proteins, and can be used for enhanced separation between hydrophilic proteins and membrane protein.

Detergents↗
PubMed1986

Phase equilibria of membrane lipids from Acholeplasma laidlawii: importance of a single lipid forming nonlamellar phases.

A basis for the reorganization of the bilayer structure in biological membranes is the different aggregate structures formed by lipids in water. The phase equilibria of all individual lipids and several in vivo polar lipid mixtures from acyl chain modified membranes of Acholeplasma laidlawii were investigated with different NMR techniques. All dioleoyl (DO) polar lipids, except monoglucosyldiglyceride (MGDG), form lamellar liquid crystalline (L alpha) phases only. The phase diagram of DOMGDG reveals reversed cubic (III), reversed hexagonal (HII), and L alpha phases. In mixtures of DOMGDG and dioleoyldiglycosyldiglyceride (DODGDG), the formation of an III (or HII) phase is enhanced by DOMGDG and low hydration or high temperatures. For in vivo mixtures of all polar DO lipids, a transition from an L alpha to an III phase is promoted by low hydration or high temperatures (50 degrees C). The phospholipids are incorporated in this III phase. Likewise, III and HII phases are formed at similar temperatures in a series of in vivo mixtures with different extents of acyl chain unsaturation. However, their melting temperatures (Tm) vary in an expected manner. All cubic and hexagonal phases, except the III phase with DOMGDG, exist in equilibrium with excess water. The maximum hydration of MGDG and DGDG is similar and increases with acyl chain unsaturation but is substantially lower than that for, e.g., phosphatidylcholine. The translational diffusion of the lipids in the cubic phases is rapid, implying bicontinuous structures. However, their appearances in freeze-fracture electron microscope pictures are different. The III phase of DOMGDG belongs to the Ia3d space group.(ABSTRACT TRUNCATED AT 250 WORDS)

Acholeplasma laidlawii↗
PubMed1987

Evidence for the existence of three or more slow phases in the refolding of ribonuclease A and some characteristics of the phases.

The slow refolding kinetics of RNase A have been analyzed, by using a nonlinear least-squares program for deconvoluting the kinetic phases and applying statistical tests for quality of fit. It is found that a minimum of three slow phases are required to fit the kinetic data properly, and this is true whether the method of detection is absorbance of fluorescence. Since the number of phases and the relaxation times for each phase are independent of the method of detection, it is concluded that the same three rate-limiting processes are seen by absorbance and fluorescence. These phases correspond to the XY, CT, and ct phases described in our earlier studies. The fact that fluorescence-detected kinetics are somewhat slower than absorbance-detected kinetics is a trivial effect due not to differences in relaxation times but to the fact that the amplitude of the CT phase is enhanced in fluorescence measurements, at the expense of the faster XY phase, because of intrinsic fluorescence changes associated with the isomerization of proline-93. By use of a new double-jump technique [Schmid, F.X., Grafl, R., Wrba, A., & Beintema, J.J. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 872], it is shown that proline-93 isomerizes as the rate-limiting step in only one of the three phases, the CT phase, and that this phase involves only 25-30% of the RNase molecules. There is still no indication as to the molecular events that occur in the large, ammonium sulfate dependent XY phase, which is the pathway for formation of the nativelike intermediate.

Amino Acid Isomerases↗
PubMed2001

Intracellular Ca2+ signaling pathway is involved in light-induced phase advance, but may not be in phase delay, of the circadian melatonin rhythm in chick pineal cell.

Chick pineal cells have photoreceptive, circadian clock and melatonin synthetic capacities, and express circadian oscillation of melatonin release in vitro. Light pulses cause phase-dependent phase shift of the melatonin rhythm. The purpose of this study was to address the questions whether intracellular calcium is involved in both light-induced phase advance and delay. Thapsigargin and cyclopiazonic acid, which deplete the intracellular calcium stores, blocked the light-induced phase advance in a dose-dependent manner. The pulses of ryanodine receptor antagonist (dantrolene sodium or ruthenium red) also blocked the light-induced phase advance. Most agents did not cause a significant phase shift by themselves. On the other hand, all the agents used, failed to block the light-induced phase delay, even if the magnitude of phase delay was decreased using low intensity light. An antagonist of nitric oxide synthase blocked neither light-induced phase advance nor phase delay. These results indicate the following possibilities: (1) the mechanism of light-induced phase advance and delay may be different in chick pineal cells, or (2) if intracellular calcium is involved in both light-induced phase advance and delay, the sensitivity to light and/or agents used in this study may differ according to Zeitgeber time.

Animals↗
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↗
PubMedDate not supplied

Two-phase helical CT for detection of early gastric carcinoma: importance of the mucosal phase for analysis of the abnormal mucosal layer.

PURPOSE: To evaluate the efficacy of two-phase dynamic helical computed tomography (CT), including the gastric mucosal phase, for detection of early gastric carcinoma with typical hyperattenuating and atypical nonhyperattenuating enhancement patterns. METHOD: Two-phase helical CT scanning was performed using the water-filling method as negative oral contrast material for 212 patients with proven adenocarcinoma on endoscopic biopsy. Two gastrointestinal radiologists prospectively analyzed the focal alterations of the inner hyperattenuating mucosal layer and the outer hypoattenuating layer before the information obtained at barium study and pathologic examination was available. The first, so-called mucosal phase was obtained at 38-45 seconds after the start of intravenous injection of contrast material for a total of 150 ml/sec at a rate of 4 ml/sec to obtain maximum enhancement of the inner mucosal layer. The second delayed phase was obtained at 3 minutes. RESULTS: Fifty-four cases of early gastric cancer were suspected on two-phase helical CT preoperatively. Postoperatively, 49 cases of early gastric cancer were pathologically confirmed. The detection rate for the typical hyperattenuating early gastric cancer, that is the type I enhancement pattern defined as the localized thickening of the inner hyperattenuating layer, using two-phase helical CT was 18% (9/49 patients). The type 2 enhancement pattern, defined as the focal interruption of the inner hyperattenuating mucosal layer without abnormal enhancement of the outer hypoattenuating layer on the mucosal phase, was seen in 15 cases. These were pathologically confirmed as early gastric cancer IIb + IIc (three patients), IIc (four patients), IIc + IIa (one patient), IIc + III (three patients), IIb + IIc (one patient), and advanced cancer (T2) lesions (three patients). The type 3 enhancement pattern was defined as the focal polypoid protrusion of the inner hyperattenuating layer without abnormal enhancement of the outer thickened hypoattenuating layer on the mucosal phase, and was seen in seven patients who were pathologically confirmed with early gastric cancer IIb + IIc (three patients), IIc + IIa (one patient), and IIc + lIb (three patients). The lesions became less distinct on the delayed phase. Five T2 advanced gastric cancers were falsely interpreted as early gastric cancer. The detection rate for early gastric cancer after considering type 2 and 3 atypical enhancement patterns was increased to 57% (28/49 patients). CONCLUSION: Helical CT with two-phase scan including the mucosal phase was efficient for identifying the enhancement patterns of early gastric cancer.

Adenocarcinoma↗
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↗
PubMed2000

Polar organic phase liquid chromatography with packed capillary columns using a vancomycin chiral stationary phase

Vancomycin immobilized on silica served as the chiral stationary phase (CSP) in this investigation with polar organic solvents as the mobile phase in liquid chromatography (LC). It was shown that trace amounts of water were beneficial for improving peak shape and efficiency. To regulate the retention and selectivity an acid and/or base were added to the mobile phase where an excess of acid was shown to be preferential for enantioseparation. An unusual increase in selectivity with increasing temperature was shown for the acidic drug, thalidomide. Additionally, nonlinear van't Hoff plots were obtained for metoprolol enantiomers that showed increased retention with increasing temperature. Metoprolol also showed unusual behavior in the polar organic phase when water was added to resemble reversed-phase chromatography, with minimum retention observed at high water or high methanol concentrations. In both instances a high degree of electrostatic interaction between metoprolol and vancomycin was concluded. Metoprolol and ten of its analogs were examined on this CSP to evaluate the enantiorecognition process. A comparison in enantioselectivity for a number of acidic and basic drugs using this CSP was also carried out using the polar organic phase, reversed phase, and normal phase LC which were all compared to the results obtained in supercritical fluid chromatography (SFC). Polar organic phase LC offered a better separation of basic molecules while reversed phase LC was preferred for the resolution of acids. SFC showed the broadest enantioselectivity overall and normal phase LC indicated similar properties, as expected, to SFC but with lower column efficiency. Copyright 2000 Wiley-Liss, Inc.

Journal Article↗
PubMedDate not supplied

Optimal phases of dynamic CT for detecting hepatocellular carcinoma: evaluation of unenhanced and triple-phase images.

BACKGROUND: To determine the optimal phases of dynamic computed tomography (CT) for detecting hepatocellular carcinoma (HCC). METHODS: Fifty-two patients with 85 HCC nodules were examined by means of unenhanced and triple-phase CT images of the whole liver. The time for obtaining the arterial-phase images was 25-55 s after intravenous bolus injection of contrast material, the time for obtaining the portal venous-phase images was 65-100 s, and the time for obtaining late-phase images was 145 s to 4 min. Detectability of the HCC nodules for all phases was statistically compared. RESULTS: The detection rates for the arterial- and late-phase images were significantly higher than for the unenhanced and portal venous-phase images (p < 0.01). The combination of arterial- and late-phase images showed the same number of HCC nodules in the same number of patients as did the combination of unenhanced and triple-phase images. CONCLUSION: The combination of the arterial- and late-phase imagings was best for detecting HCC nodules.

Adult↗
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↗