Mechanism of the Long-Term Effects of Electromagnetic Radiation on

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Mechanism of the Long-Term Effects of Electromagnetic Radiation on Solutions and Suspended Colloids Miroslav Colic*,†,‡ and Dwain Morse† R&D Division, ZPM Inc., 5770 Thornwood Dr., Suite C, Goleta, California 93117, and Materials Department, University of California at Santa Barbara, Santa Barbara, California 93106 Received August 29, 1997. In Final Form: November 25, 1997 The mechanisms of the electromagnetic water treatment have been studied with a multitude of techniques and it has been found that the gas/water interface is essential to perturb water and suspended colloids. Perturbation of the gas/liquid interface results in nonequilibrium conditions which require hours to relax. Certain magnetic and electric fields also produce small amounts of ozone, superoxide, hydrogen peroxide, or atomic hydrogen. The addition of hydrogen peroxide or hydrogen did not produce equivalent effects without additional gas/liquid interface perturbations with electromagnetic fields (EMFs). Hydrophobic gases such as argon or carbon dioxide which promote clathrate-like structuring of water appear to be affected more significantly through the action of EMFs.

Introduction It has been reported that electromagnetic water treatment can enhance the growth rate of plants and animals, accelerate the healing of broken bones and soft tissue, descale or inhibit the scaling of metallic surfaces, and improve the compressive strength of cement.1-6 Most relaxation phenomena in water, aqueous solutions, and suspensions occur on a time scale of picoseconds to seconds, yet these startling results occur even when the water is treated with magnets, electromagnets, radio frequency (rf) radiation, or microwaves and used hours after the treatment.7 The mechanism by which the long-term effects are stored in water is largely unknown despite recent contributions from different laboratories which have quantified magnetic effects on solutions, suspensions, and even pure water.8-14 However, no one has attempted to explain the physicochemical reasons for the long-term effects of EMF water treatment. * To whom correspondence should be addressed: email, [email protected]; fax, (805)-893-8971. † ZPM Inc. ‡ University of California at Santa Barbara. (1) Dushkin, S. S.; Ievstratov, V. N. In Magnetic Water Treatment in Chemical Undertaking; Khymia: Moscow, 1986. (2) Nakashima, K.; Yamamoto, H. J. Toyota College Technol. 1987, 20, 67. (3) Lin, J. J.; Yotvat, J. J. Magn. Magn. Mater. 1990, 83, 525. (4) Sisken, S. B.; Walker, J. In Electromagnetic Fields: Biological Interactions and Mechanisms; Blank, M., Ed.; American Chemical Society: Washington, DC, 1995. (5) Basset, C. A. In Electromagnetic Fields: Biological Interactions and Mechanisms; Blank, M., Ed.; American Chemical Society: Washington, DC, 1995. (6) Walleczek, J. In Electromagnetic Fields: Biological Interactions and Mechanisms; Blank, M., Ed.; American Chemical Society: Washington, DC, 1995. (7) Baker, J. S.; Judd, S. J. Water Res. 1996, 30, 247. (8) Chibowski, E.; Gopalkrishnan, S.; Busch, M. A.; Busch, K. W. J. Colloid Interface Sci. 1990, 139, 43. (9) Chibowski, E.; Holysz, L.; Wojcik, W. Colloids Surf. 1994, 92, 79. (10) Holysz, L.; Chibowski, E. J. Colloid Interface Sci. 1994, 165, 243. (11) Chibowski, E.; Holysz, L. Colloids Surf. 1995, 101, 99. (12) Higashitani, K.; Kage, A.; Kotamura, S.; Imai, K.; Hatade, S. J. Colloid Interface Sci. 1993, 156, 90. (13) Higashitani, K.; Okuhara, K.; Hatade, S. J. Colloid Interface Sci. 1992, 152, 125. (14) Higashitani, K.; Iseri, H.; Okuhara, K.; Kage, A.; Hatade, S. J. Colloid Interface Sci. 1995, 172, 383.

Chibowski and co-workers8-11 studied the effects of electromagnetic radiation in the rf range (44 MHz) on the pH, conductivity, and zeta potential of colloidal particles of different oxides. Oscillations in zeta potentials were observed for hours after rf treatment, while no change in the zeta potentials was observed with nontreated samples. Higashitani and co-workers also observed the “magnetic water memory” effect on the precipitation of calcium carbonate from solutions of calcium nitrate and sodium carbonate.12 Calcium chloride and sodium carbonate solutions were magnetically treated and stored separately and then mixed after extended periods of time (hours to days). Nontreated solutions yielded mostly calcite, while magnetically treated solutions produced mixtures of calcite and aragonite. The magnetic treatment also increased the size of the calcium cabonate precipitates. Higashitani and co-workers13,14 observed the long-term effects after magnetic water treatment with colloid stability, electrophoretic mobility, and interparticle force measurements on treated suspensions. These researchers also observed a “magnetic memory effect” on the molecular level.15 Changes in the fluorescence of hydrophobic and hydrophilic probes dissolved in water were followed after magnetic treatment. While no change in the fluorescence of hydrophilic probes was observed, a statistically significant change occurred in the fluorescence of the hydrophobic probes. The long-term effects lasted for up to 8 h, not days as in the case of treated sodium carbonate solutions. Fesenko and co-workers16-18 performed a series of experiments on the effects of electromagnetically treated water on biological systems. First, cells were irradiated with microwave radiation and a significant change in the frequency of the opening of potassium channels was measured. In a second set of experiments, water was treated with microwaves and added to the same type of cells after different times of storage. Similar changes in (15) Higashitani, K.; Oshitani, J.; Ohmura, N. Colloids Surf. 1996, 109, 167. (16) Geltyuk, V. I.; Kazachenko, V. N.; Chemeris, N. K.; Fesenko, E. E. FEBS Lett. 1995, 359, 85. (17) Fesenko, E. E.; Geletyuk, V. I.; Kazachenko, V. N.; Chemeris, N. K. FEBS Lett. 1995, 366, 49. (18) Fesenko, E. E.; Gluvstein, A. Y. FEBS Lett. 1995, 367, 53.

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the frequency of opening of the potassium channels were observed when compared to cells that were directly irradiated. Finally, the treated and nontreated waters were analyzed through the relaxation of short dc capacitive pulses. This method directly showed a change in water response, and possibly structure. Fesenko and co-workers used triply distilled deionized water but noted that the water was always in contact with ambient air. Consequently, a correlation likely exists between effects on the gas/water interface and long-term effects on aqueous solutions and suspensions after EMF treatment. The concept of treating the gas/water interface with different stimuli to produce unusual effects has received considerable attention in recent years due to significant interest in the sonoluminesecence phenomena. Sonoluminescence19-22 is an emission of light resulting from resonant ultrasonic treatment of water containing bubbles. The generation of visible light from 24 kHz ultrasound is an amazing amplification in frequency of 14 orders of magnitude. Interestingly, while the duration of ultrasound pulses is in microseconds, the emitted visible light pulses burst in picoseconds.20 The mechanisms of the sonoluminescence phenomenon are still largely unknown. Eberlein recently proposed a quantum vacuum radiation theory of sonoluminescence23 which predicts an oscillating EMF to strongly influence the gas/water interface. In this model, the two interfaces with different polarizabilities (water and bubbles) are treated as a possible two-photon state source during excitation by electromagnetic fields. The virtual two-photon state becomes real when the dielectric moves due to an EMF: the magnetic field causes the paramagnetic species to move inside and the diamagnetic species to move outside the field. An oscillating EMF can influence the bubble/liquid interface by simply causing oscillations. Sonoluminescence results in the breakup of water molecules and production of hydrogen and hydroxyl free radicals, which yield hydrogen peroxide upon disproportionation. Experimental Materials and Methods The radio frequency (rf) signal was delivered with a helical resonator source described in ref 24 . The rf antenna was a gold-coated copper sphere with a diameter of 1 cm. The antenna was enclosed in a plastic box to prevent dielectric and conductivity losses that result from direct contact with water. The treatment was performed in the water-filled bucket. The source was positioned ca. 10 cm from closed polypropylene bottles with samples to be treated, as shown in Figure 1. The possibility of any contamination was minimized with such a setup. The outgassing of solutions (when indicated) was performed as follows. The solutions were placed in Erlenmeyer flasks closed with Teflon stopers and outgassed on vacuum (vacuum pump) for 30 min. Solutions were used immediately after outgassing. For electrophoretic mobility measurements, suspensions of 10 mg of oxide in 100 mL of solution were prepared in 0.001 M of NaNO3 and equilibrated overnight before rf treatment and zeta potential measurements. Zeta Meter 3.0 and Malvern Zeta Sizer 3 instruments were used to follow zeta potential changes with time. The kinetics of calcium carbonate precipitation was followed with the method developed by Higashitani et al.12 Solutions of 5 mM calcium nitrate and sodium carbonate were treated with the rf for 15 min as shown in Figure 1 and then mixed on the (19) Hiller, R. A.; Barber, B. P., Sci. Am. 1995, 272 (Feb), 96. (20) Barber, B. P.; Putterman, S. J. Nature 1991, 352, 318. (21) Hiller, R.; Weninger, K.; Putterman, S. J.; Barber, B. P. Science 1994, 266, 248. (22) Hiller, R.; Putterman, S. J.; Barber, B. P. Phys. Rev. Lett. 1992, 69, 1182. (23) Eberlein, C. Phys. Rev. A 1996, 53, 2772. (24) Morse, D. E.; Cook, J. H.; Matherly, T. G.; Ham, H. M. U.S. Patent 5,606, 723, 1997.

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Figure 1. Schematic representation of the rf treatment system used in the current work. rf energy delivery system and a bottle with water to be treated are immersed in the same bucket of water for a better rf transmission.

Figure 2. Effect of rf water treatment on zeta potential of suspended rutile with and without evacuation. vortex inside the turbidimeter cell. The turbidity was then followed versus time with a HiScience turbidimeter. The calcium carbonate precipitate was collected, observed under an Olympus microscope equipped with a video camera, and dried before X-ray analysis. X-ray analysis was performed on a Scintag Instrument diffractometer. FTIR analysis was performed with the Digilab FTS 40 FTIR spectrometer. Formation of hydrogen peroxide was measured fluorometrically at an excitation wavelength of 350 nm and emission wavelength of 460 nm by monitoring the oxidation of 1.5 mm of scopoletin in the presence of 6.6 units/mL of horseradish peroxidase, using a Perkin-Elmer LB50 spectrophotometer with a magnetic stirring unit. Small amounts of hydrogen peroxide (0.05-0.3 ppm) were observed after rf treatment of water saturated with air but not after rf treatment of outgassed water.

Results and Discussion All of the observed phenomena occurred in water solutions or suspensions. It is currently believed that the long-term effects after magnetic water treatment are related to changes in hydrogen bonding in water and the hydration of ions, colloids, and adsorbed ions. Results presented in this report indicate that careful evacuation of water solutions and suspensions results in the disappearance of the magnetic water memory effect. Figure 2 presents the results of electrophoretic mobility measurements versus time after rutile suspension treatment. rf treatment was performed for 15 min with the 27 MHz radio frequency generator described in ref 24. The

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Figure 3. Effect of rf water treatment on calcium carbonate precipitation with and without outgassing prior to mixing calcium nitrate and sodium carbonate

nontreated suspension exhibited constant electrophoretic mobility versus time behavior, as expected. Treated solutions showed oscillations in the electrophoretic mobility of rutile particles with time after treatment similar to that observed by Chibowski and co-workers.8 Careful outgassing of solutions before or after the rf treatment produced suspensions with no changes in electrophoretic mobility with time. Figure 3 shows the influence of preliminary rf water treatment on the kinetics of calcium carbonate precipitation with nonevacuated and evacuated solutions. As in Higashitani’s work,12 this was measured by following the turbidity change with time. In our work, the turbidity was observed after mixing 5 mM calcium nitrate and sodium carbonate solutions starting at 15 min after the rf treatment. In the inital few minutes after mixing, the turbidity increases due to nucleation, then levels off because of the onset of precipitation (approximately equal number of nuclei are formed and reacted to yield the precipitate), and finally decreases as larger precipitated particles begin to sediment. The rf treatment lowers the plateau in the turbidity vs time curve, suggesting that fewer particles were formed. Optical microscopy indicated that these particles are larger than those formed in nontreated solutions. This probably occurs because of the improved hydration of the ions, with increasingly hydrated calcium and carbonate ions being more difficult to dehydrate. The dehydration is a crucial process involved in the precipitation into the crystal lattice from the initiated nuclei. When the ions are more difficult to dehydrate, the nuclei will grow larger before the critical size for dehydration and precipitation is reached. Another possible explanation for the EMF effects on calcium carbonate precipitation is the change in solubility and diffusivity of carbon dioxide, which is closely involved in the chemistry of carbonates. As in the case of the electrophoretic mobility measurements, careful outgassing before or after the rf treatment caused the dissappearance of the long-term effects after magnetic water treatment. X-ray diffraction and FTIR studies showed calcite as the majority phase present in nontreated samples, with only traces of vaterite. Treated samples contained some aragonite and small amounts of calcium hydroxide; however, when outgassed, the treated solutions again yielded calcite with no aragonite (data not shown). FTIR spectra of the collected precipitate before and after treatment are presented in Figure 4. The details of the precipitate analysis are described below.

Figure 4. Effect of rf water treatment on calcium carbonate precipitation. FTIR spectra of the collected precipitates.

Calcium carbonate samples for FTIR measurements were prepared by mixing 500 mL of 0.005 M calcium nitrate and 500 mL of 0.005 M sodium carbonate. Solutions were left for 1 h to complete the precipitation and then filtered through a 0.22 µm Millipore filter. The precipitate was dried overnight at room temperature. One milligram of the precipitate was mixed with 50 mg of KBr, and diffuse reflectance FTIR spectra were taken with a FTIR spectrometer. In the FTIR spectra of precipitate collected from solutions which were not rf treated, absorption bands characteristic of calcite appeared at 877 and 1435 cm-1. X-ray diffraction measurements confirmed the existence of pure calcite with trace amounts of vaterite. FTIR spectra of precipitates produced from the rf-treated samples indicated smaller amounts of calcite and the appearance of aragonite (band at 1085 cm-1) and calcium hydroxide (broad hydroxy band centered at 3450 cm-1). X-ray diffraction results confirmed the presence of calcite, aragonite, and calcium oxide in the rf-treated sample. Typically, around 16 mg of precipitate was collected after mixing solutions which were not rf treated. Outgassed samples yielded ca. 20 mg of precipitate. This is not unexpected since calcium carbonate is much less soluble than calcium hydrogen carbonate, which is initially precipitated in the presence of carbon dioxide. No change in yield was observed for the rf-treated samples. On the other hand, ca. 13 mg of precipitate was collected from nonoutgassed rf- treated solutions. The decrease in yield in this case is probably caused by higher solubility of aragonite and calcium hydroxide, which were identified in the rf-treated samples. Only calcite was identified in the nontreated samples. Average particle size of the calcite rhombohedra precipitated from the outgassed or nonoutgassed solutions which were not rf treated was around 7 µm. Calcite

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rhombohedra precipitated after rf treatment of solutions were larger (11 µm average) and more polydisperse. Aragonite particles precipitated from rf-treated solutions were rectangular and very polydisperse. Calcium hydroxide dots were too small to measure with optical microscope. Scanning electron microscope pictures identified that those particles were polydisperse spheres with radii between 100 and 800 nm. Another interesting phenomenon was observed when argon-saturated solutions were treated with rf: a significant overpressure developed due to a decrease in the argon solubility in water. When argon-saturated water bottles were closed airtight and outfitted with a pressure sensor, overpressures up to 94 kPa were observed. This pressure change lasted up to 8 h. With the knowledge that the gas/liquid interface is being affected by magnetic treatment, several other issues remain to be resolved. First, are there any free radicals formed by the water treatment as in sonoluminescence? Second, why would magnetic treatment influence dissolved gas (in sonoluminescence the gas/liquid interface produces the effect, not dissolved gas). The first issue has been resolved by our work as well as the recent work of Hayashi and co-workers.25 The amount of hydrogen peroxide in treated water was measured (peroxide is the long-term stable product of hydroxyl or superoxide radicals formed in water solutions). Indeed we were able to identify nanomolar to micromolar amounts of hydrogen peroxide in water treated by placing a 0.42 T magnet around a test tube containing water. Electromagnetic treatment with oscillating fields also yields nanomolar amounts of hydrogen peroxide when the electrodes are submerged into the water during treatment. No hydrogen peroxide was identified when working with rf frequencies above 50 MHz. Hayashi and co-workers25 recently made a remarkable discovery when treating water with an electrical field delivered with platinum-coated titanium electrodes: the presence of atomic hydrogen was detected in water. The atomic hydrogen was directly detected by the standard analytical technique (hydrothermal reaction with tungsten trioxide) and was stable in water for months. Harsh treatments such as repeated freezing and thawing or boiling in closed containers did not remove the atomic hydrogen. In subsequent biologically relevant experiments with superoxide production with the xantine oxidase/xantine system, they realized the powerful ability of atomic hydrogen to quench reactive oxygen species (hydroxyl radicals, hydrogen peroxide, superoxide, and singlet oxygen). Direct measurements also showed a tremendous capability of atomic hydrogen to prevent oxidative DNA damage. Direct additon of hydrogen gas showed much smaller effects. The gas/liquid system produced through the perturbation with an EMF was needed to yield the observed effects.25 The second question concerning the effect on dissolved gases is more difficult to resolve. How are effects seen so similar to those observed in the sonoluminescence phenomena if all of the gas molecules are dissolved in water, and thus there is no gas/liquid interface present. That question might be answered by the work of Ninham, Bunkin, and co-workers who identified small nanosized bubbles (tens of nanometers diamater) in nonoutgassed samples studied with the aid of neutron diffraction, light (25) Shirata, S.; et al. Biochem. Biophys. Res. Commun. 1997, 234, 269.

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scattering, and laser cavitation experiments.26 The bubbles are organized into fractal aggregate structures, and the bubble network interpenetrates with the water network. Changes at the water/bubble interface also influence the water structure and consequently the surface forces between particles, ions, and molecules in water. Hydrophobic surfaces were found to be influenced more significantly,27 which came as no surprise, since it is wellknown that bubbles prefer to accumulate near hydrophobic surfaces. Cavitation studies showed that it was much easier to form cavities near hydrophobic surfaces, suggesting the presence of more nuclei near such surfaces. While those nuclei could have been particulate impurities, outgassing studies showed the dissappearance of the effect suggesting gas micronuclei as the probable source of the cavities. The role of the gas/liquid interface in controlling the forces between hydrophobic surfaces has been directly proven by Pashley and co-workers.28 Dodecane/water mixtures are normally unstable and fully demulsify in seconds in the absence of surface active materials. Pashley and co-workers carefully outgassed water and dodecane before mixing. After mixing, most of dodecane separated from the water, but a small amount remained emulsified in water for hours. One of the numerous reasons for demulsification is the hydrophobic interactions between hydrocarbon molecules. The absence of gas nuclei seemed to somewhat decrease the hydrophobic attractions between the alkyl chains mixed with water. Meagher and coworkers29 and Yoon et al.30 actually measured the influence of outgassing on the forces between hydrophobic surfaces. Indeed it was found that the hydrophobic force of shorter range was present after outgassing. Since hydrophobic forces are involved in lipid aggregation, membranemembrane interactions, and protein-membrane interactions, the modification of hydrophobic forces might be related to the bioeffects of electromagnetic radiation. Ninham and co-workers31 actually observed that neutron radiation can modify hydrophobic forces for much longer than the lifetime of the free hydroxyl radicals produced. Atomic hydrogen produced through radiolysis of water might adsorb at the hydrophobic surfaces and modify the water structure around hydrophobic chains. Our preliminary measurements indicate that rf water treatment influences the behavior of the carbon dioxide/ water and argon/water interfaces much more significantly than oxygen/water interface. The hydrogen/water interface also seems to be modified by an rf field. Atomic hydrogen, argon, and carbon dioxide are hydrophobic gases, known to organize water into clathrate-like, longrange structures.32 It appears that magnetic and EM fields perturb the water structure near hydrophobic interfaces more efficiently. Higashitani and co-workers, for instance, realized that magnetic water treatment could not change the fluorescence spectra of dissolved hydrophilic probes. Conversely, the spectra of hydrophobic probes were modified with a memory of at least 8 h.15 Ozeki et al. observed that magnetic fields affected the adsorption of water at hydrophobic surfaces more strongly than at the (26) Ninham, B. W.; Kurihara, K.; Vinogradova, O. I. Colloids Surf. 1997, 123, 7. (27) Bunkin, N. F.; et al. Langmuir 1997, 13, 3024. (28) Karaman, M. E.; Ninham, B. W.; Pashley, R. M. J. Phys. Chem. 1996, 100, 15503. (29) Meagher, L.; Craig, V. S. J. Langmuir 1994, 10, 2736. (30) Rabinovich, Y. I.; Yoon, R. H. Colloids Surf. 1994, 93, 263. (31) Ninham, B. W.; et al. Submitted to J. Phys. Chem. (32) Hallbrucker, J.; Mayer, E. J. Chem. Soc., Faraday Trans. 1990, 86, 3785.

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hydrophilic surfaces.33 Our spectroscopic studies presented elsewhere34 also indicated a dramatic influence of rf on the structure of the gas/water and hydrophobic surface/water interfaces. Perturbations of such clathratelike water by the EMFs appear to last for hours or longer. Conclusions To conclude, it appears that the long-term effects after magnetic water treatment correlate well to the changes of the gas/water interface. Perturbations of the gas/liquid interface require more time to equilibrate than pure water or aqueous solutions. Small amounts of ozone, superoxide, (33) Ozeki, S.; et al. J. Phys. Chem. 1996, 100, 4205. (34) Colic, M.; Morse, D. Submitted to Phys. Rev. Lett.

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hydrogen peroxide, or atomic hydrogen are also produced during the application of the electromagnetic field. These reactive gaseous oxygen or hydrogen species, when produced with the aid of the electromagnetic or electric fields, influence the reactivity and structure of water despite their presence in miniscule amounts. We are just beginning to understand the fascinating mechanisms of the long-term effects after electromagnetic water treatment. Substantial work with different physicochemical techniques is still needed to shed more light on this research area with significant industrial and biomedical applications already in practice. LA970979A