Biological effects of radiofrequency

Influence of sub lethal microwave radiation on bacterial growth, enzyme activity, and exopolysaccharide production


Scientific Study, 2013

82 Pages, Grade: A


Excerpt


Index

List of tables

List of figures

1. Prologue
1.1 Preamble
1.2 The Importance of the Study
1.3 Statement of the Problem
1.4 Rationale of the Research Work
1.5 Objectives

2. Literature Review
2.1 Interactions of Microwave with Biological Materials
2.2 Athermal (Non-thermal) Mechanisms of Interaction
2.3 Thermal versus athermal effects
2.4 Factors affecting Microwave effects
2.5 Biological effects of Microwave radiation
2.6 Microwave mutagenesis
2.7 Effect of Microwave on higher organisms
2.8 Microwave and cellphones
2.9 Effect of Microwave on growth and enzyme activity
2.10 Amylase
2.11 Pectinase
2.12 β-galactosidase
2.13 Glucose-6-phosphatase
2.14 Xanthan gum

3. Effect of low power microwave on growth, enzyme activity (amylase and pectinase) and EPS production on different bacteria
3.1 Materials and Methods
3.1.1 Culture maintenance
3.1.2 Culture activation
3.1.3 Inoculum preparation
3.1.4 Microwave oven and its maintenance
3.1.5 Microwave treatment to inoculums
3.1.6 Growth measurement
3.1.7 Amylase estimation
3.1.8 Pectinase estimation
3.1.9 EPS quantification
3.1.10 Statistical Analysis
3.2 Results and Discussion

4. Effect of low power microwave on growth, intra- and extracellular protein and intracellular enzymes (glucose-6-phosphatase and β-galactosidase)34-
4.1 Materials and Methods
4.1.1 Test organisms
4.1.2 Experimental outline
4.1.3 Estimation of intra- and extracellular protein content
4.1.4 Estimation of G6P
4.1.5 Estimation of β-galactosidase
4.2 Results and discussion

5. To investigate mutagenic effect of MW on exopolysaccharide production in X. campestris
5.1 Materials and Methods
5.1.1 Test organisms
5.1.2 Microwave treatment to inoculums
5.1.3 Experimental outline
5.2 Results and discussion

6. Epilogue

7. Appendices
Appendix I
Appendix II
Appendix III
Appendix IV
Appendix V
Appendix VI

8. References

List of Tables

Table 3.1 Experimental conditions

Table 3.2 Effect of MW on growth and amylase activity in B. subtilis

Table 3.3 Effect of MW on growth and amylase activity in S. mutans

Table 3.4 Effect of MW radiation on growth and pectinase activity in P. carotovora

Table 3.5 Effect of MW radiation on growth and pectinase activity in B. subtilis

Table 3.6 Effect of MW on growth and EPS production in X. campestris

Table 3.7 Effect of MW on growth and EPS production in S. mutans

Table 4.1 Effect of MW on growth, and protein synthesis in B. subtilis

Table 4.2 Effect of MW radiation on glucose-6-phosphatase and β-galactosidase activity in B. subtilis

Table 4.3 Effect of MW radiation on growth, and protein synthesis in L. acidophilus

Table 4.4 Effect of MW radiation on glucose-6-phosphatase and β-galactosidase activity in L. acidophilus

Table 4.5 Effect of MW radiation on growth, and protein synthesis in E. coli

Table 4.6 Effect of MW radiation on G6P and β-galactosidase activity in E.coli

Table 4.7 Comparison of effect of different MW exposure on growth, protein synthesis, and enzyme activity in the test organisms

Table 5.1 Effect of MW (90 W) on growth and xanthan produced by X. campestris

Table 5.2 Growth and xanthan gum production by three different isolates selected randomly from plate corresponding to 6 min MW treatment

Table 5.3 Effect of MW (90 W) on growth and xanthan production by X. campestris

Table 5.4 Effect of MW (450 W) on growth and xanthan produced in X. campestris..

Table 5.5 Growth and xanthan gum production by four different isolates selected randomly from plate corresponding to 3 min MW treatment

Table 5.6 Effect of MW (450 W) on growth and xanthan production by X. campestris

List of Figures

Figure 1.1 Position of microwave radiation in electromagnetic spectrum

Figure 2.1 The orientation of dipoles in absence (a) and in presence (b) of electric field

Figure 2.2 The structures of amylose (A) and amylopectin (B)

Figure 2.3 The action of β-gal on lactose converting it into galactose and glucose subunits

Figure 2.4 The conversion of G-6-P to glucose

Figure 2.5 Structure of xanthan gum

Figure 3.1 Comparison of effect of different duration of MW exposure on growth and amylase activity in B. subtilis.

Figure 3.2 Comparison of different duration of MW on growth and amylase activity

in S. mutans

Figure 3.3 Comparison of different duration of MW on growth and pectinase activity in P. carotovora.

Figure 3.4 Comparison of different duration of MW on growth and pectinase activity in B. subtilis.

Figure 3.5 Comparison of effect of different duration of MW exposure on growth and EPS production in X. campestris

Figure 3.6 Comparison of effect of different duration of MW exposure on growth and EPS production in S. mutans

Figure 4.1 Comparison of effect of different duration of MW exposure on growth, protein synthesis, glucose-6-phosphatase and β-galactosidase activity in B. subtilis

Figure 4.2 Comparison of effect of different duration of MW exposure on growth, protein synthesis, glucose-6-phosphatase and β-galactosidase activity in L. acidophilus

Figure 4.3 Comparison of effect of different duration of MW exposure on growth, protein synthesis, glucose-6-phosphatase and β-galactosidase activity in E. coli

Figure 5.1 Outline of the experiment to study the mutagenic effect of MW in X. campestris

1. Prologue

1.1. Preamble

Since many years, scientists are interested in studying the interaction of electromagnetic fields (EMFs) and various bio-system and its bioprocesses. All biological systems are electrochemical in nature so EMF may influence them. Attention has been focused on different frequency range waves, of which microwave (MW) is an important part. Microwaves (MW) are non-ionizing electromagnetic waves in 1mm to 1m wavelength range, with a wide frequency band between 300 MHz and 300 GHz [Banik et al., 2003]. They are a very important component of the electromagnetic spectrum as demonstrated by the increasing scope of applications. They have relatively short wavelengths and high frequencies compared to the extremely low frequency fields. They therefore have a greater energy which is sufficient to cause heating in conductive materials (Figure 1.1). Unlike the X-rays and gamma rays, which are ionizing, MW interaction with matter does not result in removal of orbital electrons. Rather, such interactions are known to cause effects like atomic excitations, increased atomic and molecular vibrations, rotations, and heat production [Davis and VanZandt, 1988]. Because of the nature of MW, its interaction mechanisms and the biological effects have previously not been associated with production of free radicals, oxidative modification of cell membranes, lipid peroxidation. These mechanisms are associated with ionizing radiations. Interest in the study of MW interaction with biological systems has been sustained for several decades. The first person to explore the bioeffects of MW fields was Antonin Gosset in 1924, when he and his co-workers used short waves to destroy tumors in plants with no damage to the plant itself [Bren, 1996]. During the 1930s, physicists, engineers, and biologists studied the effects of low frequency electromagnetic waves on biological materials. Studies of the effects of microwaves on bacteria, viruses and DNA were performed in the 1960s and included research on heating, biocidal effects, dielectric dispersion, mutagenic effects, etc. [Yaghmaee and Durance, 2005]. In recent year, the industrial MW applications have grown considerably, apart from the usage of domestic MW ovens. Some of the MW applications in industries are tampering of frozen product, thawing, blanching, baking, drying/dehydration/vacuum drying/freeze-drying, pasteurization, sterilization, cooking, etc. In modern world, MW radiations are used in FM radio, television, RADAR (Radio Detection and Ranging), and satellite-to-earth communications. Apart from its use in domestic ovens, many other applications of MW in different areas have been identified.

Applications of MW that are based primarily on its thermal effects include those in food processing industry [Decareau and Kenyon, 1970], disinfection and sterilization [Bhattacharjee et al., 2009; Kothari et al., 2011], moisture removal [Ozbek and Dadali, 2007], waste treatment [Beszedes et al., 2010], etc.

illustration not visible in this excerpt

Figure 1.1 Position of microwave radiation in electromagnetic spectrum

(http://www.chem.ucalgary.ca/courses/350/Carey5th/Ch13/ch13-1.html)

MW irradiation is also used to disinfect household products such as sponges, kitchen utensils, syringes and medicinal devices where bacterial viability is reduced by MW treatment. Microwave assisted extraction (MAE) has emerged as a promising method for preparation of bioactive plant extracts. It is considered suitable for fast extraction of phenolic compounds, and also for extraction of heat-labile phytoconstituents [Gupta et al. 2012; Kothari et al., 2012; Mandal et al., 2007]. Research on non-thermal effects can open the door for new applications which may be based either solely on non-thermal effect, or both thermal and non-thermal effects. Either low power or high power (for very short duration, and with a provision of cooling water circulation to avoid heating) MW can be used for mutagenesis in plants [Jangid et al., 2010] and microorganisms [Lin et al., 2012; Li et al., 2009], protein unfolding [George et al., 2008], tumor detection based on electrical properties of tissues [Stuchly and Stuchly, 1983], rate enhancement of biochemical reactions [Bose et al., 2002; Sun et al., 2011], enzyme immobilization [Wang et al., 2011], and reduction of pathogenic population [Barnabas et al., 2010].

1.2. The Importance of the Study

Sub- lethal MW-microbes interaction: may change level of

- Growth rate [Rebrova et al., 1992; Golant et al., 1994; Shub et al., 1995]
- Metabolite activity [Dreyfuss et al., 1980]
- Plasmid amplification [Otludil et al., 2005]
- Rapid Strain identification [Spencer et al., 1985]
- Transformation efficiency [Fregel et al., 2008]
- Porosity of the cell membrane that allow uptake of drugs into MW-treated cell
[Shamis et al., 2011]
- Secondary metabolites such as antibiotics [Himabindu et al., 2007]
- Lose of virulence of virulent strain [Moore et al., 1979; Tsuji and Yokoigawa 2011]
- Mutation [Lin et al., 2012; Li et al., 2009]

1.3. Statement of the Problem

Increasing applications of MW radiation has led to concerns globally due to the suspected bio effects associated with its exposure. Effect of MW, thermal and/or athermal, is inconclusive, complex, and controversial in literature. Thermal effect causes thermogenic effect while athermal effects are other than heat and such effects reported as somatic effect and/or genetic effect.

This study basically deals with the athermal effects and is aimed at investigating the hypothesis that the exposure of microbial cells to MW (low power) may cause athermal effect, which affect on growth of microbes, enzyme activity, and production of exopolysaccharides. Furthermore, we have also checked the effect of different intracellular enzymes on MW treated bacteria. Our study also gives information that MW athermal effects causes changes at genetic level and can be passed on to next generation.

1.4. Rationale of the Research Work

There are numerous and increasing applications of MW energy and technology in the industries, in homes, in medical, research institutions etc., and there is greater awareness and concern of the public over the suspected potential health hazards associated with such exposures [ICNIRP Guidelines, 1998]. There is therefore, a need for deeper understanding of the bio-effects of exposure to this radiation. Due to the ease of handing them in laboratory, microorganisms can be conveniently used to study the effect of MW on living systems. Besides, employing mutagenic frequencies of MW radiation for microbial strain improvement can be of considerable industrial significance.

1.5. Objectives:

1. To investigate the effect of low power MW on,
a. Growth
b. Extracellular enzyme (amylase and pectinase) activity in Bacillus subtilis, Streptococcus mutans and Pectobacterium carotovora.
c. Exopolysaccharide (EPS) in S. mutans and Xanthomonas campestris.

2. To study the effect of low power MW on,
a. Growth
b. Protein synthesis
c. Intracellular enzyme (Glucose-6-phosphatase and β- galactosidase) activity

3. To investigate mutagenic effect of MW on EPS production in X. campestris.

2. Literature Review

2.1. Interactions of MW with Biological Materials

The interactions can be considered at various levels of organization of a living organism: atomic, molecular, subcellular, cellular, entire organism. These interactions with biosystem arise because of three processes: (i) penetration by electromagnetic waves and their propagation into the living system, (ii) primary interaction of the waves with biosystem; and (iii) possible secondary effects arising from the primary interaction. One of the first fundamental steps in evaluating the effects of a certain exposure to radiation in a living organism is determination of the induced internal electromagnetic field and its spatial distribution. Further, various possible biophysical mechanisms of interaction can be applied. Any such interactions, which may be considered primary, elicit one or more secondary reactions in the living system. For instance, when MW energy absorption results in a temperature increase within cell (a primary interaction), the activation of the thermoregulatory, compensatory mechanism is a possible secondary interaction [Czerski, 1975; Czerski, 1975] While the primary interactions are becoming better understood, there is still insufficient attention devoted to the interaction mechanisms involving molecular level. Studies on the biological effects of MWs reveal several areas of established effects and mechanisms on the one hand and speculative effects on the other. There are known thermal and athermal interaction mechanisms of MWs with biological systems.

2.2. Athermal (Non-thermal) Mechanisms of Interaction

MW radiation seems to affect system in a manner, which cannot be explained by thermal effects alone [Spencer et al., 1985]. MW has ability to destroy bacterial cells at specific parameters without causing heating of the substrate [Barnabas et al., 2010]. MW plays role in dielectric saturation [Hyland, 1988], formation of oxidative stress [Sokolovic et al., 2008], protein unfolding [George et al., 2008], changing the structures by differentially partitioning the ions [Asadi et al., 2011], others chemical transformation of small molecules such as chemical bond cleavage [Oslen, 1966], vibrational resonance in DNA molecules [Edwards et al., 1985]. The oscillating EMF of MW couples energy into large biomolecules with several oscillations. When a large number of dipoles are present in one molecule (DNA, protein, RNA etc.) and kept under MW, enough energy can be transferred to the molecules, which would be able to break the bond.

2.3. Thermal versus athermal effects

Biological effects of MW radiation can be divided into two categories: thermal effects and non-thermal effects. Thermal effect is the one in which the MW energy is converted into heat energy in the living systems. These effects can be macroscopic where whole organisms or major portions of them participate in the heat transfer process, or microscopic where a cellular component like bound water is vaporized by the selective application of the MW heating [Richmond, 1969]. The dielectric effect of MW on polar molecules has been known for more than a century [Debye, 1922]. Polar molecules are present in the cells in the form of water, DNA, and proteins and they respond to an electromagnetic field by rotating. This rotation creates an angular momentum which results in friction with neighboring molecules, thereby developing a linear momentum (vibrational energy) [Saifuddin et al., 2009]. In this way, radiation energy is converted into thermal energy. Effect generated from vibrational energy is thermal effect which occurs in a biosystem due to penetration of electromagnetic waves (MW) into biological materials and heating up the intra- and extra- cellular fluids by transfer of vibrational energy [Tahir et al., 2009]. However, MW thermal effect is different from conventional heating effect. Dipolar polarization and rotation of molecules in an attempt to align the dipoles with applied MW field (Fig. 2.1) produces effects which cannot be achieved by conventional heating [Zelentsova et al., 2004].

The non-thermal effect of MW is highly controversial and has been a matter of debate in scientific community. Non-thermal effects are postulated to result from a direct stabilizing interaction of electric field with specific (polar) molecules in reaction medium with no rise in temperature [Herrero et al., 2007]. Thermal effects solely cannot explain the manner in which the MW affects biological systems. Several studies have revealed that MW radiation can kill microbial cells; however it is still not clear if non-thermal effects of MW have any contribution to this. One of the mechanisms involved in killing of microorganisms by MW is by altering the permeability of the cell membrane. The alteration in the permeability of the membrane of the cells is reflected by cell shape changes observed under electron microscopy or by detecting leakage of intracellular protein or DNA using spectroscopy [Chen et al., 2007]. Due to the paucity of information regarding the exact mechanism involved in ‘nonthermal microwave effects’ (also referred to as athermal effects or MW specific effects), their existence is highly controversial. Kozempel et al. (2000) attempted at detection of nonthermal effects of MW energy on microbes at low temperature in various test fluids. To separate thermal and nonthermal effects in a system, they developed a continuous experimental microwave process combining rapid energy input to the food system using microwave, with rapid removal of thermal energy utilising an efficient heat exchanger design. A continuous MW treatment (7 kW, 2450 MHz) was given to the test organisms in various test fluids like water, liquid egg, beer, apple juice, and tomato juice. They concluded that MW energy in the absence of other stresses did not kill microorganisms at low temperatures and there was no convincing evidence that MW energy could kill microorganisms without thermal energy. Dreyfuss and Chipley (1980) studied the effect of MW at sub-lethal temperature on Staphylococcus aureus and suggested the existence of a phenomenon different from thermal heating resulting in altered activity of various metabolic enzymes. There has been a plenty of reports favoring the existence of non-thermal effects [Dreyfuss and Chipley, 1980; Copty et al., 2006; Carta and Desogus, 2012], but the studies refusing the possibility of athermal effects [Vela and Wu, 1979; Kozempel et al., 2000; Shazman et al. 2007] can also not be neglected.

illustration not visible in this excerpt

Figure 2.1 The orientation of dipoles in absence (a) and in presence (b) of electric field

[Williams J.M., 2009]

2.4. Factors affecting MW effects

Interaction of MW with biological entities is influenced by multiple factors viz., MW power and frequency, far-field versus near-field location, duration of exposure, polarization, etc. Reports suggest that continuous and pulsed MW treatments have different effects on cells. Human diploid fibroblasts and rat granulosa cells were exposed to intermittent and continuous waves. With the help of alkaline and neutral comet assay, DNA strand breaks were determined. Single and double strand breaks occurred in both cell types, but the intermittent exposure showed a stronger effect in the comet assay than continuous exposure [Diem et al., 2005]. Normal human lymphocytes isolated from the peripheral blood were exposed for 5 days to 2450 MHz microwave radiation in both continuous and pulsed form. Spontaneous lymphoblastoid transformation was determined with an image analysis system and it was found that pulsed wave exposure enhanced transformation to a greater extent than continuous wave exposure [Czerska et al., 1992].

Duration of MW exposure seems to be a major determinant of MW effect on living cell. The time of exposure and power density are correlated in a way that decrease in power density (PD) could be compensated by increase in duration of exposure. Cells of Escherichia coli K12 AB1157 were irradiated with millimetre waves within the PD range of 10−20 to 10 4 W/cm 2 and it was found that decrease in PD could be compensated by increase in exposure time to achieve the same changes in chromatin conformation [Belyaev et al., 1992]. There have been reports suggesting that the length of post-treatment time following MW exposure is also important in determining response of living cells to MW radiation. Sub-lethal MW radiation studies on E. coli revealed that cell-surface undergo modifications which are electrokinetic in nature, and cells revert back to original stat after 10 min of exposure [Shamis et al. 2011]. Low level MW radiation of 10 GHz frequency, 0.58 mW/cm 2 intensity, applied for 30-120 min caused loss of virulence in Agrobacterium tumefaciens strain B6, where 30-60% decrease in their ability to produce tumor and turnip disk in plant was observed. However, this loss in virulence was reversible within 12 h [Moore et al., 1979].

The medium/matrix in which the cells are embedded during MW exposure can also have its impact. The efficiency with which different solvents absorb microwaves and pass it on as heat to the surrounding molecules is indicated by dissipation factor (tan δ), expressed as:

tan δ = ε’’ / ε’,

where, ε’’ is the dielectric loss which indicates the efficiency of conversion of microwave energy into heat; ε’ is the dielectric constant which is the measure of the ability of the material to absorb microwave energy. A reaction medium with a high tan δ at the particular operating frequency of a microwave synthesis reactor is required for good absorption [Herrero et al., 2007]. Different cells, organs and tissues of biological entities have varying dielectric properties, and thus are affected differently by MW radiation.

2.5. Biological effects of MW radiation

Research is being done worldwide on thermal and athermal effects of MW on different biological systems. Phormidium spp. Kutzing ISC31 (a cyanobacterium) grown in BG-11 medium was treated at a frequency of 2450 MHz using a microwave oven by combining five different intensities (180, 360, 540, 720 and 900 W/cm 2 ) and three pretreatments (10, 20 and 30 s). The content of chlorophyll a decreased with increase in intensity and exposure time. Synthesis of phycobiliproteins, phycocyanin, phycoerythrin, and allophycocyanin increased in all exposures except in 720 and 900 W/cm 2 (30 s). Photosynthetic rate compared to nitrogenase activity increased by all microwave exposures except at 180 W (10 s) and 720 W (10 s) as compared to control [Asadi et al., 2011]. Studies on E. coli and S. aureus suggested that physical damage caused by MW led to alterations in membrane permeability and consequently influx of extracellular Ca 2+ . An increase in cell permeability of upto 89.8% and 19.7% was obtained after MW treatment in S. aureus and E. coli respectively [Chen et al. 2007]. Effect of MW on transformation efficiency was studied by Fregel et al. (2008), where calcium chloride competent cells of E. coli were given MW treatment at 180 W for 1 min, and the transformation efficiency was increased three-fold as compared to the classical method. Otludil et al., (2004) studied the effect of microwave (2450 MHz, 55 W) on the cellular differentiation of Bacillus subtilis YB 886 and its Rec derivatives YB 886 A4. It was found that organism’s growth and amount of DNA decreased after MW exposure by 4% and 27% respectively, whereas the amount of RNA and plasmid was enhanced by 6.5% and 21% respectively. They noted an increase in the amount of specific protein synthesized during DNA damage by SOS repair system, and its binding to din C promoter region, following MW exposure in rec+ bacteria.

2.6. Microwave Mutagenesis

A number of studies reveal that effects of MW can reach up to genetic level and can even result in stable mutation. Genetically stable mutant strains with higher nitrogenase activity and phosphate solubilizing capabilities of Klebsiella pneumoniae RSN19 were obtained by microwave (250W, 36 s) mutagenesis [Li et al., 2011]. Mutagenic potential of MW has also been demonstrated with respect to cellulase production in Trichoderma viride, wherein a compound mutagenesis by MW (700W, 15-195 s) and ultraviolet radiation was employed, and the mutants were found to be stable up to 9 generations [Li et al., 2009]. Lin et al. (2012) studied the effect of MW (400 W for 3 min) on Lactobacillus rhamnosus which induced >50% increase in L-lactic acid production than the parent strain, and the mutant generated was found to be stable up to 9 generations. Such studies together indicate the potential of MW as a substantial tool for strain improvement through mutagenesis. MW till now has not been exploited as widely UV has been as a mutagenic radiation. Identifying the frequencies, power range and exposure duration in MW region, which are most suitable for mutagenesis among microbes will certainly be of interest to fermentation industries.

2.7. Effect of MW on higher organisms

A study on effects of low power MW radiation on germination and growth rate in seeds of wheat (Triticum aestivum), bengal gram (Cicer arietinum), green gram (Vigna radiata), and moth bean (Vigna aconitifolia) showed that different treatments stimulated the germination and seedling vigour. Increase in power density resulted in reduced rate of germination [Ragha et al., 2011]. Effect of different MW power was studied on potato tuber biomass and it was found that for seed potatoes irradiated with microwaves (100 W) at frequency 2.45 GHz for 10 s, tuber weight was 7.9% higher compared to check sample. No significant change was observed at 38 GHz, 46 GHz, and 54 GHz [Jakubowski, 2010]. Treatment with low-level MW (35 GHz; surface power density 30 μW/cm 2 ) radiation for 10 s has been reported to induce chromatin condensation (increase in number of heterochromatin granules) in human cells and increased membrane permeability. However, the number of heterochromatin granules decreased to its initial level and membrane permeability was recovered after a few hours [Shckorbatov et al., 2011]. Low-intensity MW radiation effectively changed membrane functions in striated muscle and cardiac pacemaker cells in rats (Chernyakov et al., 1989). Exposure at 0.1-0.15 mW/cm 2 for 90 s or lesser time (frequencies between 54-78 GHz) either decelerated the natural loss of transmembrane potential in myocytes, or even increased it by 5-20 mV. Low intensity MW was also found to suppress and alter the T-peak on electrocardiography of in situ exposed myocardium, enhance respiration, alter membrane calcium binding, and reduce the contractibility of cardiomyocytes. Drop in blood pressure of rats caused by MW pulses persisted for several weeks indicating a stable effect. The properties of blood plasma like dielectric permittivity and absorption coefficient could be altered by microwave radiation [Lu et al., 1999]. Hybridization profile of brain and testis DNA of mice exposed to low power MW (1 mW/cm 2 ; 2450 MHz) showed an additional band suggesting amplification of tandem sequences in particular region [Sarkar et al., 1994]. Elder (2003) concluded that exposure of rabbit eye to 2450 MHz MW at 150 W/kg for more than 30 min can induce cataracts. Induction of cataracts via thermal effects of high-power MW radiation is well established. Whether low-power MW are cataractogenic remains unclear. Yu and Yao (2010) reviewed non-thermal cellular effects of low-power microwave radiation on the lens and lens epithelial cells.

2.8. MW and cell phones

Within last decade, the world population has adopted the use of cell phones in a horrendous way. Increasing use of radiofrequency devices has become a trend as well as a need in a large section of society. Cellular phones transmit radiofrequency waves of very low intensity and there has been a lot of discussion on possible adverse effects of these radiations on health. Effect of electromagnetic radiation from communication towers on human and other forms of biodiversity has been a matter of concern. Panagopoulos and Margaritis (2002) exposed fruit flies to cell phone radiation (Global System for Mobile communication-GSM-900 MHz) at very low levels (3-7 mW/cm 2 ) for 6 min/day before hatching. The exposed groups in their adult life showed a loss in reproductive activity varying between 15-60%. Frey (1962) reported that exposure of heads of volunteers to low level of radar from a radar horn resulted in headaches, and that low level of radar can interact with nerve tissue to cause dysaesthesiae. Hocking and Westerman (2001) reported a neurological abnormality in a patient after accidental exposure of left side of the face to cellular phone radiation, which led to long-lasting loss of tactile sensitivity of the facial skin, unilateral left blurred vision, and pupil constriction. The worker took virtually 6 months to recover.

2.9. Effect of MW on growth and enzyme activity

MW effects on growth and other cellular activities are being studied for atleast more than 50 years. Growth rate of yeast Saccharomycetes cerevisiae either increased upto 15% or decreased up to 29%, when exposed to MW radiation having narrow frequency range of 41.8-42.0 GHz [Grundler et al., 1977]. When S. aureus culture was exposed in a controlled temperature experiment to microwave radiation (24 GHz) for 10, 20, 30, and 40 s, the activity of various enzymes like malate dehydrogenase, cytoplasmic adenosine triphosphatase, glucose-6-phosphate dehydrogenase, and cytochrome oxidase increased in microwave treated cells than microwave non treated cells; whereas membrane adenosine triphosphate, alkaline phosphatase and lactate dehydrogenase activity remained unaffected [Dreyfuss and Chipley, 1980]. The effect of MW radiation on 94 strains of Enterobacteriaceae was studied and it was found that microwave irradiation increased the enzyme activity of bacteria in suspension [Spencer et al., 1985]. Low power MW treatment (2450 MHz; 90 W; 2 min exposure) on Aeromonas hydrophila decreased its total protease activity by 33%. Urease activity and aflatoxin production in S. aureus and Aspergillus parasiticus respectively was completely inhibited by MW exposure [Dholiya et al. 2012]. Low power (100 W, 60 s) microwave radiation reduced not only the cell number but also the acid resistance and verocytotoxin productivity in enterohemorrhagic E. coli [Tsuji and Yokoigawa, 2011]. Komarova et al. (2008) investigated influence of MW on soil bacteria. They observed both the suppression and the stimulation of the growth for different bacterial species under the impact of microwaves. Spore suspensions responded to microwave radiation upon a shorter time of exposure than suspensions of vegetative bacterial cells. The influence of microwave radiation on the biomass accumulation and the intensity of other physiological processes in streptomycetes species led to changes in the number and activity of these microorganisms in the soil microbial complex.

2.10. Amylase

Amylase was the first enzyme to be discovered and isolated in 1833 by Anselme Payen as diastase. The first amylase was produced in 1894 from a fungal source which was used as a pharmaceutical aid for digestive disorders. Biodin and Effront were the first to use B. subtilis and B. mesentericus for the production of α- amylase on commercial scale using large fermentors. Amylases are starch-degrading enzymes that are widely distributed in plants, animals and microbial kingdom. Amylase forms complex with iodine to form intense blue colour and this forms the basis of a method for quantitative determination of amylase [Singh et al., 2011].

Starch is a major reserve carbohydrate component of all higher plants and in some cases it accounts for as high as 70% of the undried plant material. It occurs in the form of water insoluble granules but when they are heated in water the hydrogen bonds that hold the granules together begin to weaken and this permits them to swell and gelatinize.

Starch is a heterogeneous polysaccharide composed of two high molecular weight entities called amylose and amylopectin (Fig 2.2). These two polymers have different structures and physical properties. Amylose is composed of linear chains of α-1,4 linked D-glucose residues and therefore extensively degraded by α-amylase. Because of the shape and molecular structure of amylase, it is not stable in aqueous solution and thus precipitates spontaneously.

Amylopectin may account for 75- 85% of most starches. It has molecular weight in excess on 10 7 – 10 8 and has a branched structure composed of chains of about 20 – 25 α-1, 4 linked D-glucose residues. In aqueous solutions, amylopectins are relatively stable due to branched molecules and are not able to form compact aggregates. The hydrolysis of starch results in formation of short-chain polymers called dextrins, then the disaccharide maltose, and ultimately glucose [Souza, 2010].

α- Amylases:

α- Amylases (1,4-α-glucan-glucanohydrolases) are extracellular enzymes which hydrolyze α-1,4-glycosidic bonds. Their action is not affected by α-1,6-glycosidic bonds although they do not split.

Bacteria that can produce α-amylases are: B. subtilis, B. amyloliquefaciens, B. cereus, Lactobacillus, Pseudomonas, Arthrobacter, Escherichia, Proteus, Serratia, etc.

Fungi from the genera Aspergillus, Penicillium, Cephalosporium, Mucor, Candida, Neurospora and Rhizopus can also produce α- amylases.

α- amylases are produced industrially from Bacillus and Aspergillus species. For industrial production, α- amylases are produced either in batch or in fed-batch fermentation. The rate of formation of enzyme is low in exponential growth, but as the growth rate decreases and spore formation begins, amylase production increases.

illustration not visible in this excerpt

Figure 2.2 The structures of amylose (A) and amylopectin (B)

http://www.rsc.org/Education/EiC/issues/2006Sept/MakingMostStarch.asp

β- Amylases:

β- Amylases (α- 1,4-glucan-maltohydrolases) are produced by Bacillus polymyxa, B. cereus, B. megaterium, Streptomyces sp., Pseudomonas sp., and Rhizopus japanicus but usually these are of plant origin. Bacterial β- amylases have a much greater heat resistance (>70%) than plant β- amylases.

Applications [Crueger and Crueger, 1984; Aiyer, 2005]

Removal of starch sizer from textile (desizing)

In textile industry, sizing agent like starch is used in fabric production for fast and secure weaving process. It also prevents the loss of string by friction, cutting and generation of static electricity on the string by giving softness to the surface of string due to laid down warp. From the woven fabric, starch is removed by amylase ensuring that the warp-thread remains intact.

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Details

Title
Biological effects of radiofrequency
Subtitle
Influence of sub lethal microwave radiation on bacterial growth, enzyme activity, and exopolysaccharide production
College
Nirma University  (Institute of Science)
Grade
A
Authors
Year
2013
Pages
82
Catalog Number
V269552
ISBN (eBook)
9783656609681
ISBN (Book)
9783656608608
File size
1607 KB
Language
English
Keywords
Microwave, athermal effects, mutation
Quote paper
Assistant Professor Vijay Kothari (Author)Toshi Mishra (Author)Preemada Kushwah (Author), 2013, Biological effects of radiofrequency, Munich, GRIN Verlag, https://www.grin.com/document/269552

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Title: Biological effects of radiofrequency



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