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RUSSIAN ACADEMY OF SCIENCE
N.M. EMANUEL INSTITUTE FOR BIOCHEMICAL PHYSICS

Official seal: Russian Academy of Science* Russian Academy of Science N.M. Emanuel Institute for Biochemical Physics (IBHF RAN)* OGRN (illegible)* INN 7736043895* OKPO 40241274*
APPROVED BY:
Russian Academy of Science
N.M. Emanuel Institute for Biochemical Physics
Deputy Director

(Signed)     A.N. GOLOSHCHAPOV,
Candidate of Medicine

REPORT titled:

"A Study of Effects Produced by Physico-Chemical Properties of
Water upon the Structural State of Erythrocyte Membranes"

Purpose. This was a study of effects produced by various water samples, such as are having dissimilar properties dependent on the origin of such samples, upon structural features of erythrocyte membranes. It is known that a change in water structure can entail changes in the existing biomembrane conformations and, hence, in activity levels of biological processes taking place in a cell, because cellular membrane is top sensitive to any changes in external and internal environments. It has also been shown that various water activation methods and phase changes can give rise to water molecule homolytical decomposition mechano-chemical reactions combined with appearance of hydrogen radical and hydroxyl radical. Such radicals, having become hydrated, can exist for a large span of time; they make up chemically active ions and radicals, including active oxygen forms, and this produces a wide range of biological effects. Therefore, there are reasons to expect manifestations of biological activity in water put through various stages of treatment.

Many diseases are accompanied with intensification in lipid peroxide oxidation and changes in membrane structures. Today, studies of biological membrane structural state are successfully practiced with use of electronic paramagnetic resonance spectroscopy (EPR spectroscopy) well designed to make readings of any slight changes in biophysical parameters of cellular membranes - both in case of their normal functioning and in case of any chemical or physical exposures.

Sample description. The following three water samples were used as test objects:

  1. Pipe-water sample taken out of Moscow City Western Administrative Circuit water supply system (Sample W);
  2. The same water purified with use of water purification household system manufactured by ZEPTER, Series: EE-RO-6PF (Sample F);
  3. Water Sample F later treated with use of Aquator field generator designed to produce specific effects upon structural state of the water (Sample T).

Test methods. 0.01M tris-HCL buffer, where mouse erythrocytes were put as a 5% suspension, was prepared on the three under-test water samples. The buffer so prepared was tested for erythrocyte membrane viscosity rates. Viscosity of various membrane areas was evaluated in reliance upon rotational correlation time of spinal probes incorporated into the membrane. The following stable iminoxyl radicals were used as such probes: 2,2,6,6-tetramethyl-4-capryloil-oxypiperidine-1-oxyl (Probe 1) and 5,6-benzo-2,2,6,6-tetramethyl-1,2,3,4-tetrahydro--3-oxyl (Probe 2). The two radicals are mutually dissimilar in their hydrophobic properties. It is known that, predominantly, Probe 1 is located in the lipid bi-layer, and Probe 2 - in enzyme-proximate lipids of the membrane. Iminoxyl radicals were infused into the 5% erythrocyte suspension as an alcoholic solution 30 minutes before the samples were measured at ER-200D SRC EPR spectrometer manufactured by BRUKER of the FRG. Basing on EPR spectrums obtained, rotational correlation time (), which means, essentially, the period of re-orientation of the radical at /2 angle, was calculated.

Results obtained and discussion. Results obtained in the tests are shown in Table 1.

Table 1. Erythrocyte membrane micro-viscosity, as dependent on exposure to various water samples (, 10-10 sec)

Water In Absolute Units In %, as related to Sample W
Probe 1 Probe 2 Probe 1 Probe 2
W Sample 0.40 0.77 100 100
F Sample 0.46 0.82 115 106
T Sample 0.41 0.77 103 100

It is obvious from the Table that the largest increase in lipid bi-layer micro-viscosity and in enzyme-proximate lipids of the erythrocytes takes place when Sample F is in use, that is, the membrane grows "harder". The growing viscosity of the membrane indicates an intensification of free-radical oxidative reactions taking place in it. The resultant active oxygen forms are highly aggressive and entail cellular impairments giving rise to within-cell metabolic disturbances and to development of various pathologies.

Water T produces, virtually, no changes in erythrocyte membrane viscosity as compared to the initial Sample W, which means that its physiological parameters are close in their rates to the control rates.

Therefore, the obtained data prove the supposed differences in the levels of biological activity of the under-test water samples. It follows from the test conditions that differences in the extent of effects produced upon biomembrane viscosity rates are associated with physico-chemical properties of the under-test samples, which properties have been given rise to by technological treatment of the water.

Therefore, the following conclusion can be made: Water T - filtered and treated with use of Aquator appliance - is the best in quality, as compared to Water F, because it does not act to disturb the structure of erythrocyte membranes.

The testing was performed by Candidate of Biology, Senior Research Officer of Laboratory for Physico-Chemical Fundamentals of Biological System Regulation at: Russian Academy of Science, N.M. Emanuel Institute for Biochemical Physics

(Signed)      /L.D. FATKULLINA/
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