A rapid and sensitive sub-micro phosphorus determination

Böttcher, C.; Gent, C.M. van; Pries, Caitlin Hicks

doi:10.1016/0003-2670(61)80041-x

Cited by 970 publications

(421 citation statements)

References 3 publications

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“…Phospholipids were quantitated by measuring the inorganic phosphate (Bartlett, 1959), after hydrolysis of extracts at 180 OC in 70% HCIOl (Bottcher et al, 1961). Cholesterol in lipid extracts was assayed by the Lieberman-BGrchard reaction (Huang et al, 1961).…”

Section: Methodsmentioning

confidence: 99%

Modulation of sarcoplasmic reticulum calcium pump activity by membrane fluidity

Almeida¹,

Vaz

Zachariasse

et al. 1984

Biochemistry

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Intramolecular excimerization of 1,3-di-l-pyrenylpropane [Py(3)Py] was used to assess the fluidity of sarcoplasmic reticulum membranes (SR); on the basis of the spectral data, the probe incorporates completely inside the membrane probably somewhere close to the polar head groups of phospholipid molecules, however not in the very hydrophobic core. The excimerization rate is very sensitive to lipid phase transitions, as revealed by thermal profiles of dimyristoylphosphatidylcholine (DMPC) and dipalmitoylphosphatidylcholine (DPPC) bilayers. Cholesterol abolishes pretransitions and broadens the thermal profiles of the main transitions which vanish completely at 50 mol % sterol. Excimer formation in liposomes of SR total lipid extracts does not show any sharp transitions, as in the case of DMPC and DPPC. However, the plots display discontinuities at about 20 OC which are broadened by cholesterol and not observed at 50 mol % sterol.Sarcoplasmic reticulum membranes (SR) have been extensively characterized in their structural and functional aspects (Weber et al., 1973;Hasselbach, 1979;Tada et al., 1978). The major protein, the Ca++-pump enzyme, is intrinsically associated with membrane lipids which greatly influence the enzyme activity (Martonosi et al., 1971;Bennett et al., 1980; Johannsson et al., 1981a). Lipids in contact with the ATPase enzyme modulate its function through physical interactions (Bennett et al., 1980; Johannsson et al., 1981a), including changing membrane fluidity. Thus, CaZ+ translocation across S R membrane and molecular mechanisms associated to energy transductions between the electroosmotic energy of Ca2+ gradients and chemical energy of ATP may be modulated by lipid-protein interactions, presumably affected by membrane fluidity. Cholesterol has a condensing effect on the acyl chains of bilayers in the fluid state (Houslay & Stanley, 1982). When forced to interact with some membrane proteins, it has a strong inhibitory effect on their function (Warren et al., 1975). Cholesterol is normally excluded from direct contact with the Ca2+-ATPase enzyme (Bennett et al., 1975; Johannsson et al., 1981a;Simmonds et al., 1982) Also cholesterol has been incorporated in native SR membranes by an exchange technique allowing progressive enrichment without changing the phospholipid/protein molar ratio. As in liposomes, discontinuities of excimer formation at 20 OC are broadened by cholesterol enrichment. The full activity of uncoupled Ca*+-ATPase is only affected by cholesterol above a molar ratio to phospholipid of 0.4. However, a significant decrease in activity (about 20%) is only noticed at a ratio of 0.6 (the highest technically achieved); at this ratio, about 28 lipid molecules per CaZ+-ATPase are expected to be relatively free from cholesterol interaction. The vesicle structure is still intact at this high ratio, as judged from the absence of basal activity (not Ca2+ stimulated). However, the sterol significantly decreases to about 60% the energetic efficiency of Ca2+ pumping (CaZ+/ATP ratio).

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Section: Methodsmentioning

confidence: 99%

Modulation of sarcoplasmic reticulum calcium pump activity by membrane fluidity

Almeida¹,

Vaz

Zachariasse

et al. 1984

Biochemistry

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“…The phospholipid contents of the virosome preparations were determined by phosphate analysis 34 after degradation of the lipids with perchloric acid. 35 Determination of fusion activity The fusion activity of virosomes was evaluated by monitoring low pH-dependent dilution of pyrPC into erythrocyte ghost target membranes. 21 Right-side-out erythrocyte ghosts were prepared from hypotonically lysed red blood cells.…”

Section: Preparation Of Virosomesmentioning

confidence: 99%

“…36 Erythrocyte ghost phospholipid contents were determined by phosphate analysis 34 after extraction 37 and digestion of the lipids. 35 Ghosts and virosomes were added to a quartz microcuvette containing HBS. After a 2-min incubation period, the medium was acidified to pH 5.6 by the injection of 30 l of 0.10 m acetic acid and 0.10 m 2-[N-morpholino]ethanesulfonic acid (pH 4.2).…”

Section: Preparation Of Virosomesmentioning

confidence: 99%

Gene transfer mediated by fusion protein hemagglutinin reconstituted in cationic lipid vesicles

et al. 1999

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Hemagglutinin, the membrane fusion protein of influenza erythrocyte ghosts. Efficient cell transfection of BHK-21 virus, is known to mediate a low-pH-dependent fusion cells was observed with virosomes containing 30 mol% reaction between the viral envelope and the limiting mem-DODAC and plasmid encoding for ␤-galactosidase (pCMV brane of the endosomal cell compartment following cellular ␤-gal) associated to their surface. The transfection activity uptake of the virus particles by receptor-mediated endoobserved was dependent on the functional activity of hemcytosis. Here we exploited this activity of hemagglutinin to agglutinin. Contrary to DNA/cationic lipid complexes the achieve efficient gene delivery to cultured cells. Hemagglutransfection was not dependent on the cationic lipid to DNA tinin was reconstituted in the presence of the monocationic charge ratio. Importantly, transfection of BHK-21 cells with lipid dioleoyldimethylammonium chloride (DODAC) to perpCMV ␤-gal by DODAC-containing virosomes did not show mit plasmid binding to the virosome surface. Virosomes any significant signs of cytotoxicity that is commonly with 30 mol% DODAC exhibited a distinct binding capacity observed with DNA/cationic lipid complexes. Together with for plasmid without causing aggregation. The virosome the high levels of expression of the transgene this highfusion activity was not affected by the cationic lipid DODAC lights the potential of DODAC-containing virosomes as a as demonstrated by low-pH-dependent lipid mixing with novel approach in nonviral gene transfer.

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“…The drug/total lipid ratio (D/L) (%w/w) was determined before liposomal injection by lipid and BNZ quantitation. Lipid concentration was determined by a colorimetric phosphate micro assay (Bötcher et al, 1961) and encapsulated BNZ was determined after complete disruption of one volume of liposomal sample in 10 volumes of DMSO by reverse-phase HPLC (Morilla et al, 2003).…”

Section: Physical Characterization Of Liposomesmentioning

confidence: 99%

Intravenous liposomal benznidazole as trypanocidal agent: increasing drug delivery to liver is not enough

Morilla

Montanari

Prieto

et al. 2004

International Journal of Pharmaceutics

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With the aim of investigating if delivery of benznidazole (BNZ) to liver could be increased by incorporating the drug in multilamellar liposomes, single bolus of free BNZ or liposomal BNZ formulations (MLV-BNZ) composed of HSPC:DSPG:Chol 2:1:2 (mol/mol/mol) at 0.7% (w/w) drug/total lipid ratio, were injected by intramuscular (i.m.), subcutaneous (s.c.) and intravenous (i.v.) routes, at 0.2 mg BNZ/kg, in rats. The resulting blood concentrations were followed along 9 h post-injection (p.i.) and drug accumulation in liver was determined after 4 and 9 h p.i. Only upon i.v. injection of MLV-BNZ, a threefold higher BNZ accumulation in liver was obtained, together with blood BNZ concentrations of 1.1 g/ml (30% lower than the blood BNZ concentration achieved upon i.v. administration of free drug) occurred 4 h p.i. However, such increased liver uptake of BNZ, raised twice a week had no effect on parasitaemia levels of mice infected with the RA strain of Trypanosoma cruzi. Our results indicate that the relationship between increased selectivity for an infected tissue and therapeutic effect is not always straightforward, at least for the MLV-BNZ regimen used in the present study.

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A rapid and sensitive sub-micro phosphorus determination

Cited by 970 publications

References 3 publications

Modulation of sarcoplasmic reticulum calcium pump activity by membrane fluidity

Modulation of sarcoplasmic reticulum calcium pump activity by membrane fluidity

Gene transfer mediated by fusion protein hemagglutinin reconstituted in cationic lipid vesicles

Intravenous liposomal benznidazole as trypanocidal agent: increasing drug delivery to liver is not enough

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