Metachromatic effect

The content of long-chain PolyPs may be estimated by measuring the metachromatic effect in the absorption spectrum of toluidine blue (Chernysheva et al., 1971 Leitao et al., 1995 Lorenz and Schroder, 1999). Toluidine blue in an aqueous solution exhibits a concentration-dependent absorption spectrum due to a monomer (A.max, 632 nm)-dimer O.max, 590 nm) equilibrium. The PolyP induced the maximal shift of the absorption spectrum to 545 nm. Nucleic acids also induce metachromasia, but with a shift of about 570 nm with DNA and 590 nm with RNA. Figure 2.3 demonstrates the typical absorption spectra of toluidine blue and toluidine blue with different preparations of PolyPs (Chernysheva etal., 1971). 

The method used for determination of PolyP, which is based on the Mn2+-induced quenching of the fluorescence of the calcium indicator Fura-2, has been described (Lorenz et al., 1997a). The effect of Mn2+ ions on the Fura-2 fluorescence is gradually removed in the presence of increasing PolyPs concentrations this allows the quantification of PolyPs isolated from tissues or cells. The described method has some advantages when compared with the conventional detection procedures based on the metachromatic effect. It can be applied to the determination of pyrophosphate, tripolyphosphate and other short-chain PolyPs not detectable by toluidine blue (Lorenz et al., 1997a). 

S. P. Damle and P. S. Krishnan (1954). Studies on the role of metaphosphate in moulds. I. Quantitative studies on the metachromatic effect of melaphosphate. Arch. Biochem. Biophys., 49, 58-70. 

The immisdbility of CF2- and CHi-chains was also utilized for the preparation of unsymmetric vesicle membranes. The hydrophobic parts of bolaamphiphile 5 with fatty acid and fluorocarbon sulfonate halves do not mix and all the fluorosulfonate halves were on the outer side of the monolayered vesicle membrane (Figure 4.7). The sulfonate head group was once more localized by the metachromatic effect, the hydrophobic parts with F- and H-substituted spin labels. 

With bilayer lipid membranes it is not possible to achieve a fully asymmetric arrangement of head groups or chains. There is no apparent reason why all the molecules of two independent layers should only concentrate in one layer. Nevertheless, a little asymmetric distribution is found in vesicles made of lipid mixtures. Cerebroside sulfate, an anionic monoglycosyl ceramide was, for example, added exclusively to the outer surface of a performed DPPC vesicle (see Scheme 2.2) which was quantitized by the metachromatic effect of acridine orange. 

A mixture of negatively charged phosphatidic acid and phosphatidylcholine was cosonicated. This system was used to quantify the metachromatic effects of... 

Figure 2.5.14 Asymmetrical monolayered vesicle membranes have been obtained from the two bolaamphiphiles shown, (a) All large headgroups are on the outer surface, all small headgroups at the inside of the vesicle. This phenomenon is by no means universal. It has to be tested for each individual asymmetrical bolaamphiphile. In most cases there is only a small difference in the localization of different headgroups. In (b) the metachromatic effect of polyanions on methylene blue aggregation on the sulfonated membrane outside is indicated (see text above).

A model [22] of how heparin acts specifically in many biological systems in modifying activities of complex ions may be provided by the metachromatic effect on dyes referred to earlier. The dye. Azure A, shows maximum light absorption at 610 nm. This is decreased when heparin is added, and a new absorption band at 505 mu develops. Heparins and heparinoids are able to produce this color change at very low concentrations and under conditions unfavorable to other metachromatic inducing substances. However, little attention has been paid to the numerous experimental observations reported on metachromasia with heparin and heparinoids, of practical importance to those using this color reaction in studies on heparin and mast cells. In... 

The metachromatic effect is presumably due to the association of the dye molecules on binding with the polyanion which may involve both electrostatic and hydrophobic interaction. The destruction of metachromatic effect may occur on addition of low molecular weight electrolytes, alcohols, or urea. The destruction of metachro-masy by alcohols and urea is attributed to the involvement of hydrophobic bonding, as has already been established (Browning and Holtzer, 1961 Frank and Evans, 1945 Frank and Quist, 1961 Kauzmznn et al, 1959 Mukherjee and Ray, 1963 ... 

The interaction was studied by the spectrophotometric method employed by Peacocke and Skerrett (3) to investigate the interaction between proflavine and nucleic acids and depicted by Blake and Peacocke (4) to describe the interaction of aminoacridines with nucleic acids. This method was particularly suitable for studies on 9-Amino-acridine derivatives - DNA complex because of the large metachromatic effect on the spectrum of the dyes. 

Matzner, U., Hartmann, D., Lullmann-Rauch, R., Coenen, R., Rothert, F., Mansson, J. E., Fredman, P., D Hooge, R., De Deyn, P. P. and Gieselmann, V. (2002). Bone marrow stem cell-based gene transfer in a mouse model for metachromatic leukodystrophy Effects on visceral and nervous system disease manifestations. Gene Ther. 9, 53-63. 

When the sulphate groups are removed using the procedures described by Araki (A2), the adsorption effects as well as the electro-osmotic flow are considerably reduced. As a consequence, P-lipoproteins move at nearly the same rate as in paper-electrophoresis experiments. Moreover, sulphate-free agar is very suitable to electrophoretic analysis of mucopolysaccharides, as the background of electrophoresis diagrams stained by metachromatic techniques is practically colorless (R.E.P.A. Ballieux, personal communication). As stained electrophoresis diagrams... 

Mast cells with their metachromatic granules (0.3 to 1.0 [tm in diameter) as their hallmarks produce histamine, which has marked vasoactive effects and also seems to enhance oxygen radical formation by phagocytes (Friedl et al. 1989). 

Numerous publications, particularly by Stone [38, 39], describe the induced Cotton effects in dye complexes with acidic polysaccharides such as heparin, chondroitin sulfate, carboxy me thy 1-cellulose, pectic acid and hyaluronic acid [40—42]. Methylene blue is usually used for these investigations, as its metachromasy depends to a high degree on the polysaccharide structure, As an example of a 4-1 linked amino sugar, heparin with methylene blue has a metachromatic absorption maximum of X = 565 nm. It its ORD a trough appears at 587 nm ( 3), and the CD also shows an exciton split with a zero crossover at 578 nm. These observations indicate dipole coupling and a helical arrangement of the bound dye molecules. 

Chondroitin sulfate, a 3-1 linked amino sugar, in the presence of methylene blue shows only one unsplit positive Cotton effect at 577 nm. The metachromatic absorption maximum and the ORD crossover conform well at 575 and 573 nm respectively. Here too we can assume that the dye cations are bound in strict geometrical order dictated by the secondary... 

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