A 50 Hz, 14 mT magnetic field is not mutagenic or co-mutagenic in bacterial mutation assays

A 50 Hz, 14 mT magnetic field is not mutagenic or co-mutagenic in bacterial...

(Parte 2 de 2)

a The values indicate ratios of revertant colonies per plate (exposure/control). b Tested with rat liver S9 mix. Each test used at least three plates. P< 0:05. P< 0:01. P< 0:001 (Student’s t-test).

the mutagenicity of fluorescent light with and without AO (Table 4). Only the light was mutagenic, and the mutagenicity (caused by the UV component of the light) increased with exposure time. The linear relationships between irradiation time (X) and mutation rate (Y) with and without MF were Y D 1:94X C 83:2 (correlation coefficient, r D 0:966) and Y D 2:12X C 7:6( r D 0:948), respectively. The mutagenicity of AO increased with increasing irradiation time. The linear relationships with and without the MF were, Y D 6:85X C 75:5( r D 0:988) and Y D 7:16X C 72:9( r D 0:992), respectively. The slope generated by AO alone was greater than that generated by light alone. In both experiments, the ratio of the revertants in the MF-exposed group to those in the MF-unexposed groups ranged from 0.9 to 1.2, with and without AO. The differences were not statistically significant.

Table 4 Effect of magnetic field on mutagenicity of light irradiation with and without AO in E. coli WP2 uvrA/pKM101a

Experiment Irradiation time (min)

With AO Ratio (exposure/control)

Without AO Ratio

(exposure/control) Control Exposure Control Exposure

a Each test used at least three plates. Values represent mean S.D. No statistical difference was found in any of the experiments.

The repeated experiment shows that the effect of the

MF shown in 3.2 was an artifact caused by the placement of the sham coil and the exposure coil, which caused a difference of light irradiation time.

4. Discussion

Mutational studies of MFs have generally been negative, especially in the density range of 0.15–5mT [4]. Our results showed that 50Hz, 14mT MFs did not damage DNA in four Salmonella and two E. coli tester strains. (Table 2) that detect point and frameshift mutations (but not DNA strand breaks).

Some authors, however, have reported that MF exposure increases mutation frequency [4–8,13,19,20], and mechanisms suggested to explain that effect include altered radical behavior [7,8,19,20], altered DNA replication [5,6], and enhanced activity of DNA-reactive agents [13]. If such indirect effects do occur, they should be detectable in bacterial mutation assays with high density MFs. For example, an effect on radical behavior or on the activity of DNA-reactive agents should be detected in an assay with radical-generating or DNA-reactive mutagens, respectively, and an effect on DNA replication or repair systems should be detected because bacterial cells employ those systems.

4.1. Effect of MFs on radical species

Altering the way radical species behave is one of the most plausible mechanisms by which MFs could contribute to mutagenesis, and it is supported by some experimental evidence [8,19,20]. Radical lifetimes (a few microseconds or less) are shorter than power frequency MF cycles (16–20ms) [8], and lengthening or shortening the lifetime should change the quantity of reaction products.

Evidence from mutation assays on the effect of MF on radical reactions is contradictory. Lai and Singh reported that in vivo exposure to a 60Hz, 0.1–0.5mT MF affected spontaneous radical reactions in a DNA strand break assay in rat brain cells [7]. Suri et al. reported that a 60Hz, 3mT MF was neither mutagenic nor co-mutagenic with N-methylnitrosourea (which is involved in the formation of free radicals) in transgenic rat embryo fibroblasts [21].

The hydroxyl radical attacks DNA, and it is one of the most biologically reactive compounds known [2]. In this study, the mutagenicity of BH and CH, which generate hydroxyl radicals, was not enhanced by a 48h exposure to a 50Hz, 14mT MF, which suggests that exposure to MFs below 14mT would also have no mutation-enhancing effect. In the light irradiation experiments with AO, photodynamic mechanisms would generate singlet oxygen [2,23], and singlet oxygen would generate hydroxyl radicals [2]. Our negative results suggest that singlet oxygen activity would not be affected by exposure to MFs below 14mT.

4.2. Effect of MFs on activity of chemical mutagens

Our previous report [13] showed that exposure to static MF fields (2 or 5T) increased the mutation rates induced by DNA-reactive agents (ENNG, AF-2, 4-NQO, N-methyl-N0-nitro-N-nitrosoguanidine, 2-amino-3-methyl-3H-imidazo[4,5-f]quinoline, and ethylmethanesulfonate). Some DNA adduct formation was due to the generation of radical species by the test chemicals. In the present study, however, the mutagenicity of DNA-reactive chemicals (ENNG, AF-2, and 4-NQO) was not affected by exposure to a 14mT MF. Although the MF frequency and flux density differed in the two studies, the main reason for the disparate results would be the difference in MF flux density. The threshold of the effect was between 14mT and 2T.

4.3. Effect of MF on DNA repair systems

Mutations can be caused by mistakes in DNA replication as well as by damage to DNA [2]. Some investigators have reported that power frequency MFs can alter fidelity of replication via an effect on DNA repair systems. Lai and Singh [6] reported that 60Hz, 0.1–0.5mT magnetic fields inhibit DNA repair in rat cells in vivo. Miyakoshi et al. [5] reported that the higher density 50Hz MF (400mT) affects DNA repair in a human cell line, and that the mutation frequency varies with induced current. MF exposures did not increase mutation rates in any tester strain in the present study. In the light irradiation experiment, DNA damage occurred only during light irradiation. The results also suggest that DNA repair activity was not affected. Alkyltransferase, which is involved in the repair of DNA alkylation, and excision repair enzymes, such as those that repair DNA oxidized by the hydroxyl radical (except in uvrA and uvrB), may not be affected by the exposure.

4.4. Effect of MF on electron transfer

Another possible way that power frequency MFs might enhance mutagenicity is by affecting electron transfer, which is important in P450 promutagenic activating systems. A postulated mechanism is that the MFs induce current that affect charge transfers, such as electron or ion flow. Nossol et al. reported that weak static and 50Hz magnetic fields influence the redox activity of cytochrome-C oxidase [24]. In our study, however, cytochrome-C oxidase activity was not affected by MF exposure, which was tested as effect of the MF on the cell growth (data not shown). The cytochrome P450 redox reaction in rat liver S9 mix in the 2AA experiment was not affected by the MF. These results suggest that mammalian electron transfer systems involved in respiration and drug metabolism are not affected by exposure to power frequency MF below 14mT.

5. Conclusions

A 50Hz, 14mT circularly polarized MF was not mutagenic to S. typhimurium strains TA98, TA100, TA1535, or TA1537 and to E. coli WP2 uvrA or WP2 uvrA/pKM101 under the conditions of the assay. The MFwasalsonotco-mutagenicwithDNA-reactivemutagens or promutagens, base analogs, hydroxyl radicals, UV or active oxygen. We conclude that the same MF would not affect electron transfer reactions (as in S9 mix), radical reactions (as with hydroxyl radical or other active oxygen species), or repair enzymes. If MF enhances mutagenesis in eukaryotic cells, the mechanism of the effect would be specific to those cells.


[1] N. Wertheimer, E. Leeper, Electrical wiring configuration and childhood cancer, Am. J. Epidemiol. 109 (1979) 273–284. [2] C.J. Portier, M.S. Wolfe (Eds.), Assessment of health effects from exposure to power frequency electric and magnetic fields, NIEHS working group report, NIH publication no. 98-3981, 1998. [3] J. McCann, F. Dietrich, C. Rafferty, A.O. Martin, A critical review of the genotoxic potential of electric and magnetic fields, Mutat. Res. 297 (1993) 61–95. [4] J. McCann, F. Dietrich, C. Rafferty, The genotoxic potential of electric and magnetic fields: an update, Mutat. Res. 411 (1998) 45–86. [5] J. Miyakoshi, N. Yamagishi, S. Ohtsu, K. Mohri, H. Takebe,

Increase in hypoxanthine–guanine phosphoribosyl transferase gene mutations by exposure to high-density 50 Hz magnetic fields, Mutat. Res. 349 (1996) 109–114.

[6] H. Lai, N.P. Singh, Acute exposure to a 60 Hz magnetic field increases DNA single and double strand breaks in rat brain cells, Bioelectromagnetics 18 (1997) 156–165. [7] H. Lai, N.P. Singh, Melatonin and N-tert-butyl-a-phenylnitrone block 60 Hz magnetic field-induced single and double strand breaks in rat brain cells, J. Pineal Res. 2 (1997) 152– 162. [8] T. Koana, M.O. Okada, M. Ikehata, M. Nakagawa, Increase in the mitotic recombination frequency in Drosophila melanogaster by magnetic field exposure and its suppression by Vitamin E supplement, Mutat. Res. 373 (1997) 5–60. [9] B.N. Ames, J. McCann, E. Yamazaki, Methods for detecting carcinogens and mutagens with the Salmonella/mammalian-microsome mutagenicity test, Mutat. Res. 31 (1975) 347–363. [10] J. Juutilainen, A. Liimatainen, Mutation frequency in

Salmonella exposed to weak 100 Hz magnetic fields, Hereditas 104 (1986) 145–147. [1] R.L. Moore, Biological effects of magnetic fields: studies with microorganisms, Can. J. Microbiol. 25 (1979) 1145– 1151. [12] M.A. Morandi, C.M. Pak, R.P. Caren, L.D. Caren, Lack of an EMF-induced genotoxic effect in the Ames assay, Life Sci. 59 (1996) 263–271. [13] M. Ikehata, T. Koana, Y. Suzuki, H. Shimizu, M. Nakagawa,

Mutagenicity and co-mutagenicity of static magnetic fields detected by bacterial mutation assay, Mutat. Res. 427 (1999) 147–156. [14] K. Yamazaki, H. Fujinami, T. Shigemitsu, I. Nishimura, Low stray ELF magnetic field exposure system for in vitro study, Bioelectromagnetics 21 (2000) 75–83.

[15] S. Nakasono, H. Saiki, Effect of ELF magnetic fields on protein synthesis in E. coli K12, Radiat. Res. 154 (2000) 208–216. [16] T. Yahagi, M. Nagao, Y. Seino, T. Matsushima, T. Sugimura,

Mutagenicities of N-nitrosamines on Salmonella, Mutat. Res. 48 (1977) 121–129. [17] D.M. Marron, B.N. Ames, Revised methods for the

Salmonella mutagenicity test, Mutat. Res. 113 (1983) 173– 215. [18] R.E. McMahon, J.C. Cline, C.Z. Thompson, Assay of 855 test chemicals in ten tester strains using a new modification of the Ames test for bacterial mutagens, Cancer Res. 39 (1979) 682–693. [19] T.B. Grissom, T.T. Harkins, Magnetic field effects on

B12 ethanolamine ammonia lyase: evidence for a radical mechanism, Science 263 (1994) 958–960. [20] C.B. Grissom, Magnetic field effects in biology: a survey of possible mechanisms with emphasis on radical-pair recombination, Chem. Rev. 95 (1995) 3–24. [21] A. Suri, J. DeBoer, W. Kusser, B. Glickman, A 3 mT, 60

Hz magnetic field in neither mutagenic nor co-mutagenic in the presence of menadione and MNU in a transgenic rat cell line, Mutat. Res. 372 (1996) 23–31. [2] E.C. Friedberg, G.C. Walker, W. Siede, DNA Repair and

Mutagenesis, ASM Press, Washington, DC, 1995. [23] J. Decuyper, N. Houba-Herin, C.M. Calberg-Bacq, A.V. Vorst,

Evidence for the production of singlet oxygen by photoexcited acridine orange, Photochem. Photobiol. 40 (1984) 149–151. [24] B. Nossol, G. Buse, J. Silny, Influence of weak static and 50 Hz magnetic fields on the redox activity of cytochrome-C oxidase, Bioelectromagnetics 14 (1993) 361–372.

(Parte 2 de 2)