The "Information Desk"
Aim:
To answer questions of the authorities and Flemish official agencies
on
-danger associated with a particular agent
- risk associated with a particular
exposure
To provide authorities with up to date independent advice and
insights concerning environmental (low dose) exposures and their
biological and health effects.
In this presentation:
List of reports of the Information Desk
List of publications of the Information Desk
PCBs and dioxins in Belgian food: background concentrations, contamination episodes ,body burdens
and sources of contamination
Importance of endogenous DNA damage and relative efficiency of exogenous DNA damage
Low dose effects of mutagenic agents: mechanisms involved in supralinear responses
Benzo(a)pyrene adducts and effects : dose-response and risk of cancer
Environmental exposures occur in doses below maximal induction of DNA repair: increased risk
Low dose effects of receptor binding epigenetic carcinogens
Biomarkers for genotoxicity are insufficiently sensitive for the evaluation of cancer risk from lifelong
exposures starting in utero
Some potentially interesting data or conclusions concerning topics in answer to questions from
authorities and to various suggestions
Reports from the “Information desk”
Health effects of residing near landfills
Benzene: risk of cancer
Long-term effects of pollution with fine particles (PM2,5)
Acrylamide
Health effects of sun creams
Health effects of radiation from GSM’s and GSM-antenna’s
High tension Cables
Nitrates and nitrites
Health effects of brominated flame retardants
Acrylamide update
Migration of Bisphenol A from packaging material to food
Environmental medical risk analysis of Monomorium pharaonis (farao ant)
Health effects of pesticides
Transmission of human pathogens through aerosols from Water Purification Installations
Health effects of residing near an airport
Health risks from the Marly fire
Health effects of perfluoro octane sulfonate (PFOS) en perfluoro octanoic acid (PFOA)
Criteria for the indoor environment
Action plan drinking water norms
Health effects of air-freshners
Health effects of UMTS
Action plan drinking water norms (cadmium, pesticides, tri- en tetrachloorethene).
Stabilized hydrogen peroxide as desinfectant in swimming and whirlpools.
Biomonitoring: DDE, HCB, PCB’s, PCDD’s en PCDF’s in cord blood and in adolescents. International data.
Fytofarmaceutical substances used in the Flemish fruit cultivation:Short note in preparation of a biomonitoring study
Epigenetic mechanisms in carcinogenesis: points of action of tumorpromotors and for chemoprevention
Important receptor binding substances active at low doses
Publications of the “Information Desk”
Mechanisms of Carcinogenesis. A short overview. N. van Larebeke In "Proceedings of the Technical Working Group on
Environment and Health, "
Cancer as an Environmental Disease. P. Nicolopoulo-Stamati, L. Hens, C.V. Howard and N. van Larebeke (eds):
Environmental Science and technology Library Vol 20, Kluwer Academic Publishers, Dordrecht/Boston/London.
Endogenous DNA damage in humans : a review of quantitative data. Rinne De Bont and Nik van Larebeke: Mutagenesis
vol. 19 no. 3 pp. 169-185, May 2004.
A survey of 3 PCB and dioxin contamination episodes. From contamination of food items to body burdens." Rinne De
Bont1, Marc Elskens2, Willy Baeyens2, Luc Hens3, Nik van Larebeke1 Reviews in Food and Nutrition Toxicity, Vol 2,
p301-342.
“Hormesis”—An Inappropriate Extrapolation from the Specific to the Universal. DEBORAH AXELROD, MD, KATHY
BURNS, PHD, DEVRA DAVIS, PHD, MPH, NICOLAS VON LAREBEKE, MD, MPH. INT J OCCUP ENVIRON HEALTH
2004;10:335–339
Omgevingsconcentraties van mutagene agentia als oorzaak van kanker: mechanistische inzichten. N. van Larebeke en R
De Bont.Tijdschrift voor Geneeskunde, 62 (4) 319-329
Unrecognised / Potential Risk Factors for Childhood Cancer. N. van Larebeke, Linda Birnbaum , Marc Bracke , Marc
Boogaerts , Devra Davis , David DeMarini , Jos Kleinjans , Marvin Legator , Greet Schoeters and Kirsi Vähäkangas In
Proceedings of the "Technical Working Group on Environment and Health, ". INT J OCCUP ENVIRON HEALTH
2005;11:199–201
PCBs AND PCDD/FS IN FISH AND FISH PRODUCTS AND THEIR IMPACT ON THE HUMAN BODY BURDEN IN
BELGIUM. W. Baeyens1, M. Leermakers1, M. Elskens1, N. Van Larebeke2, R. De Bont2, H. Vanderperren3, A. Fontaine3,
J-M Degroodt4, L. Goeyens1,4, V. Hanot4, I. Windal4. Accepted for publication, A R C H I V E S O F Environmental
Contamination a n d Toxicology
Adducts, SCE’s and Mutations following Benzo(a)pyrene Exposure: a Review of quantitative Data followed by some
Considerations regarding Risk. Rinne De Bont, Nik van Larebeke .
Accepted for publication in "Chromosomal Alterations: Importance in Human Health", edited by Gunter Obe and
Vijayalaxmi (Springer Verlag).
PCB’s and dioxins: background concentrations in Belgian Food
A study based on measurements obtained from the Federal Agency for the Safety of the Food Chain and
on biomonitoring by the Flemish Center of Expertise on Environment and Health
Background concentrations measured in 2001
Crisis-related PCB concentrations May-August 1999
PCB and dioxin intake
Calculated and measured PCB and dioxin body burdens for female adolescents and adult
women.
High PCB concentrations in Belgian meat stem for a large part from recycling of fat in animal
feed
Measured PCB body burdens in Belgian women before and after the contamination crisis early
in 1999.
Non-crisis-related PCB concentrations 2001
PCBs
Concentration
(ng/g fat)
Maximum level
(ng/g fat)
Percent of samples (ng/g
fat)
≥200
≥1,000
71.0
None
None
25.1
35.0
None
None
35.0 ± 0.5
35.0
35.0
None
None
345
32.8 ± 8.1
30.8
80.0
None
None
295
35.2 ± 3.5
35.0
81.0
None
None
45
35.0 ± 0.0
35.0
35.0
None
None
Animal
feed
8,692
35.3 ± 10.1
34.8
586.0
0.05
None
Animal
feed
308
39.3 ± 23.3
36.9
304.0
0.3
None
Other
prod.
9
35.0 ± 0.0
35.0
35.0
None
None
Sheep
5
21.8 ± 12.2
19.2
35.0
None
None
Ostrich
4
32.7 ± 4.5
32.5
35.0
None
None
Horse
5
91.6 ± 162.6
31.7
382.0
Rabbit
5
35.0 ± 0.0
35.0
35.0
None
None
15
35.0 ± 0.0
35.0
35.0
None
None
N
AM±SD
GM
Beef
312
34.2 ± 6.2
33.3
Cow’s
milk
Poultry
190
29.5 ± 11.3
302
Eggs
Pork
Fat
(unspec)
Baby
20
None
Non-crisis-related dioxin concentrations september-december 1999
Dioxins
N
Beef
Concentration
(pg TEQ/g fat)
N
Maxi
mum
level
(pg/g
fat)
Percent of samples
(pg/g fat)
≥2
≥5
AM±SD
GM
9
2.9 ± 2.5
1.9
17
8.3
66.7
5.6
6
2.6 ± 1.3
2.4
8
13.7
87.5
25.0
40
1.5 ± 4.7
0.5
50
29.6
4.0
4.0
Milk
Poultry
Eggs
Pork
Crisis-related PCB concentrations May-August 1999
PCBs
N
Concentration (ng/g fat)
AM±SD
N
Maximum
level
(ng/g fat)
GM
Percent of samples
(ng/g fat)
≥200
≥1,000
Beef
224
61.0 ± 103.6
37.8
373
2,170.0
7.2
Cow’s milk
356
39.7 ± 43.7
28.7
721
336.0
0.6
Poultry
328
421.3 ± 2,084.8
59.6
1,403
56,856.0
10.8
3.0
135.5 ± 270.9
56.1
290
46,000.0
17.9
5.5
383.8 ± 1,063.3
100.9
4,998
39,700.0
30.6
11.4
37.2
768
4,092.0
0.9
0.5
Eggs
45
Pork
3,081
48.0 ± 36.1
1.1
None
Animal fat
unspecified
329
Fat
unspecified
240
98.7 ± 168.0
59.4
423
3,080.0
7.6
1.7
Animal
feed
181
994.5 ± 9,447.9
38.8
615
336,000.0
12.2
3.6
Other food
products
402
36.8 ± 26.7
32.4
1,096
464.0
0.5
Waste oils
248
1,716.2 ± 16,672.9
101.6
274
245,992.0
5.5
Butter
46
41.6 ± 22.0
37.4
47
157.5
Animal
feed
92
80.3 ± 101.3
52.0
615
1,262.0
None
2.9
None
4.0
None
0.4
Crisis-related dioxin concentrations May-August 1999
N
Concentration
(pg TEQ/g fat)
AM±SD
Beef
Cow’s milk
N
Maximum
level
(pg/g fat)
Percent of samples (pg/g
fat)
≥2
GM
≥5
8
5.6 ± 2.1
5.3
71
19.5
94.1
47.1
11
2.8 ± 1.1
2.6
72
27.8
47.2
2.8
14.9
39
2,613.4
64.7
38.2
14.9 ± 1.3
Poultry
2
Eggs
4
2.4 ± 1.4
2.2
41
713.1
68.3
36.6
Pork
118
2.1 ± 2.9
1.2
184
62.8
32.1
10.3
Animal fat
unspec
Fat unspec
Animal
feed
2
9.4 ± 12.6
3.1
11
45.1
90.9
90.9
10
144.6 ± 91.1
114.7
13
6,144.0
100
100
73.9
5
2,379.4
100
100
Other food
products
Waste oil
5
519.8 ± 1,041.4
PCB and dioxin intake adolescents (14-18 years) and adults (non-crisis related values of the year
1999 (September-December)
Food
product
Adolescents
Adults
PCB intake
(ng/kg/day)
Dioxin intake
(pg TEQ/kg/day)
PCB intake (ng/kg/day)
Dioxin intake (pg
TEQ/kg/day)
Chicken
1.790
0.070
2.104
0.082
Pork
7.951
0.298
7.070
0.265
Beef
2.994
0.243
2.296
0.186
Egg
1.900
0.106
1.375
0.077
Milk
6.519
0.498
5.791
0.442
Fish
4.456
0.310
6.553
0.455
Vegetables
7.964
0.070
7.533
0.059
Sheep
0.258
0.008
0.150
0.005
Horse
0.367
0.029
0.165
0.013
Total
34.200
1.631
33.038
1.584
Table 14. Calculated
and measured PCB and dioxin body burdens for female adolescents and
adult women.
BODY BURDEN
Calculated
Female
adolescents
Female adults
bw body weight
Measured
PCB (g/kg
bw)
Dioxins (ng TEQ/kg
bw)
PCB (g/kg bw)
Dioxins (ng
TEQ/kg bw)
75.48
3.67
57.12
8.82
146.95
7.07
121.63
14.66
High PCB concentrations in Belgian meat stem for a large part
from recycling of fat in animal feed
PCB concentrations (7 marker) in ng/g fat in meat
From grain in
animal feed
From fish
flour in
animal feed
Pork
15.4
1.1
35.2 ± 3.5
Chicken
14.0
6.2
35.0 ± 0.5
Based on Schepens et al., 2001
Observed
Non-crisis-related
2001 (AM±SD)
Measured PCB body burdens in Belgian women before and after the contamination crisis early
in 1999.
PCB 138 (ng/g
fat)
PCB 153 (ng/g
fat)
PCB 180
(ng/g fat)
1996-1998
106 infertile women (aged 24-42)(mean age
31.9)a
69.9
94.5
72.0
Second half on 1999
120 girls (mean age 17.4)b
75.9
101.6
55.5
125.4
171.1
123.1
33.6
33.5
10.2
Second half of 1999
197 women aged 50-65 (mean age 58.5)b
Body burden increase in 1999, after
correction for agec, as percent of the body
burden found during 1996-1998 in infertile
women
Most DNA damage is endogenous
- Oxidative processes: between 0.07 and 145.25 8-oxo-dG adducts/106 bases. 15 studies on 17 above 1 adduct/ 10 6 bases
-Lipid peroxidation: up to 0.9 M1G adducts /106 bases
-Genotoxic substances derived from DNA oxidation: base propenals
-Endogenous oestrogens
-Endogenous alkylating agents: up to 2.27 7-alkylguanine adducts /106 bases and 4.24 O4- ethylthymine adducts /106 bases
-DNA hydrolysis leading to abasic sites: of the order of 8 to 9 /106 bases
-Hydrolytic deamination
- Carbonyl stress (e.g. methylglyoxal from glucose metabolism)
This damage is repaired swiftly and efficiently mostly by Base Exision Repair (BER), MGMT (O6-methylguanine
DNAmethyltransferase) and MMR (mismatch repair)
Exogenous DNA damage :
- Larger adducts removed preferentially by nucleotide excision repair (NER ), more complex and slower
- Double strand breaks repaired by homologous recombination and mainly by error-prone non-homologous end-joining
Exogenous adducts are often more efficient in the induction of mutations and cancer
than endogenous adducts
Adducts/106 nucleotides
8-oxo-7,8-dihydro-2’deoxyguanosine
Human brain (nDNA)
2.39-5.17
MalondialdehydeM1G (lipid
peroxidation)
Human Breast
0.30
7-methylguanine
Human Pancreas
0.40-1.39
Sum of 9 important
endogenous
adducts
Human peripheral blood cells
4.97-69.05
Benzo(a)pyreendiolepoxide
Cervix Niet rooksters
Cervix rooksters
0,019
0,035
Sum of 9 important
endogenous
adducts
In vivo Rat organs including
liver
Spontaneous liver tumor
incidence is 1-2%
2.18-137.3
Aflatoxine
Rat environmental exposure
Dose for 50 % liver
tumorincidence
0,00091
0,85
Initiation of carcinogenesis: mainly from endogenous mutations.
Initiation < often mutation in repair mechanism for exogenous damage (mutation in repair for
endogenous damage often lethal)
Initiated cells often more susceptible to exogenous agents
Additional mutations often from exogenous agents
Mutational spectra in the HPRT gene of normal lymphocytes from different human populations:
similar, < endogenous mechanisms of mutagenesis
Mutation spectra of the p53 anti oncogene in cells from different types of cancers (e.g. lung vs brain):
substantial differences, < different exogenous agents
Following mechanisms can lead to a relatively higher
mutagenic efficiency of low dose exposures
1. Adaptation: Intense exposure induces DNA repair mechanisms. This
results in less and other types of mutations. A very low dose, or a low
dose spread over a large time interval, hits the cells while these repair
mechanisms are not fully induced.
2. Bystander effect:cells that are not hit by an agent also can show
effects. Proven for ionising radiation and gene therapy
3. Induced genetic instability. A form of induced genetic instability
has been described that acts as an on/off phenomenon in which the
genetic instability does not increase with the dose of the inducing agent. It
acts as if a program for genetic instability is expressed. Probably a critical
treshold value exists.It is not known when this phenomenon occurs
Benzo(a)pyrene: dose-effect relation
Supralinear exposure effect curves were found for:
BaP adducts in human cells exposed in vitro (11 studies totalling 23 measured adduct levels):
mean adduct level < 5 µM: 2.47/106 nucl per µM
>= 5 µM: 0.104/ 106 nucl per µM
p= 0.0054 ( in anova with confounders)
BaP adducts in animal cells exposed in vitro (6 studies totalling 11 measured adduct levels) :
mean adduct level < 5 µM: 1.46/106 nucl per µM
>= 5 µM: 0.11/ 106 nucl per µM
Sister chromatid exchanges in human cells in vitro (5 studies) and in animal cells in vitro (3 studies) totalling 24 meas.:
mean amount of SCEs in human cells: < 5 µM:1.48per cell per µM
>=5 µM: 0.18 per cell per µM
p= 0.046 for the entire dataset
mean amount of SCEs in animal cells: < 5 µM:13.32 per cell per µM
anova with confounders
>=5 µM: 0.97 per cell per µM
Mutations in vitro in chinese hamster ovary cells
mean mutation frequency in HGPRT or XPRT gene: < 5 µM:4.30 per 10 5 per µM
>=5 µM: 0.88 per 105 per µM
Infralinear exposure effect curve found for:
Adduct formation in vitro following BPDE exposure (5 studes) showed a nonsignificant infralinear
exposure effect curve
BaP adducts in human tissues following BaP exposure in vitro:
doses < 5μM (left), doses ≥ 5 μM (right).
Slope of the curve: 1.68
Slope of the curve: 0.105
Cancer risk associated with BaP adducts in peripheral blood cells derived from exposure through inhalation
Exposure in
coke ovens
in terms of
BaP
Bap adducts
per 106
nucleotides
in white
blood cells
(Based on 5
studies on
coke oven or
related
industries)
Life time
Cancer risk
only due to
1 µg/m3 BaP
(70 year
lifetime
exposure)
(Based on
BaP unit risk
OEHHA
1994)
Life time Lung Cancer risk from
Coke oven emissions with 1
µg/m3 BaP (70 year lifetime
exposure)
Based on all 22 coke oven, gas
works and aluminium
production studies
Based on 7 coke oven, gas
works and aluminium
production studies with
exposures <=20 µg BaP/ m3
years
Life time Lung & genito-urinary
Cancer risk (Costantino et al. 1995)
from Coke oven emissions with 1
µg/m3 BaP (70 year lifetime
exposure)
Based on all 22 coke oven, gas works
and aluminium production studies
Based on 7 coke oven, gas works and
aluminium production studies with
exposures <=20 µg BaP/ m3 years
Armstrong et al. 2004
Meta-analysis
Armstrong et al. 2004
Meta-analysis
1 µg/m3
0.418
1.1 to 3.3
/1000
39/1000
200/1000
48/1000
245/1000
Environmental exposures do not reach the treshhold level for maximal induction of DNA
repair and have consequently a higher efficiency in the induction of biological and health
effects than high experimental or occupational exposures
Critical dose for induction of adaptive repair by exogenous mutagenic agents: order of 1 mGy
Induction of adaptive response takes in the order of 1 hour
Duration of adaptive response: order of 40 hours
Exposure to 1mGy/40 hours during 70 years equals 15,3 Gy
1Gy (acute) causes 11% lethal cancers (UNSCEAR, 2000) , and a cancer incidence of about 22%.
Chronic exposure might lead to only 50% of the above mentioned risk .
15,3 m Gy in chronic exposure causes of the order of 15,3 x 11%= 168 % cancer incidence in 70
years
The observed cumulative cancer incidence rate at age 70 is of the order of 30%
Consequently total current environmental and other external exposures are well below the level
required for maximal induction of DNA repair and so have a higher efficiency per unit of dose than
high experimental or occupational exposures on which norms are based.
Receptor binding exogenous ligands often show supralinear dose-effect curves and
can have non-monotonous effects and effects that are substantially different from the
endogenous ligands
Saturation of the receptor leads to supralinear dose-effect curves
Binding of a ligand leads to changes in conformation of the receptor. Xeno-estrogens do not necessarily have the
same effects , in qualitative terms, as endogenous estrogens.
100
4 x Kd
80
Respons (% van maximum)
Bezetting (% van maximum)
Bezetting (% van maximum)
100
Kd = 0,1 nM
60
Fout door lineaire
extrapolatie
40
80
60
Fout door lineaire
extrapolatie
40
20
20
0
0
0
0,1
0,2
0,3
0,4
0,5
0
Concentratie (nM) lineaire schaal
Concentratie (nM) lineaire schaal
1
2
3
Concentratie, lineaire schaal
After Welshons et al., 2003
4
5
Biomarkers for genotoxicity have an acceptable sensitivity for
exposures occuring during adult life but are insufficiently
sensitive for the evaluation of cancer risk from lifelong
exposures starting in utero
50
20
30
40
Peripheral cells
10
Stem cel
Birth
20
40
80
Age from conception
Fig. 1
20
40
80
20
30
Birth
Peripheral cells
40
Fig. 2
50
Age from conception
stemm cells control
10
Stem cel
10
Mutant frequency per million cells
12
stemm cells w hen exposed f rom age 30 to 40
Birth
peripheral cells control
20
40
peripheral cells w hen exposed f rom age 30 to 40
8
6
Exposure from age 30 to age 40
An exposure increasing the mutation rate by 10%
will result in an attributable risk percent of cancer amounting
to 7,15% and in an increase of 4,9% in the observable
mutant frequency in peripheral Cells.
Ratio=7.15% / 4.9%=1.46
4
2
0
0
5
10
15
20
25
30
35
40
45
50
number of consecutive stemm cell divisions
Age from conception
Mutant frequency per million cells
Fig. 3
Birth
20
40
80
12
10
8
Lifelong exposure starting In utero
An exposure increasing the mutation rate by 10%
will result in an attributable risk percent of cancer
amounting to 43,5% and in an increase of 10% in
the observable mutant frequency in peripheral cells.
Ratio=43,5%/10%=4,35
stemm cells control
stemm cells w hen lif elong exposed
peripheral cells control
6
peripheral cells w hen lif elong exposed
4
2
0
0
5
10
15
20
25
30
35
40
45
50
Number of consecutive stemm cell divisions
80
Age from conception
Correspondence:
Prof. Dr. N. van Larebeke
Study Centre for Carcinogenesis and Primary Prevention of Cancer,
Ghent University
Universitary Hospital 3K3, De Pintelaan 185
B 9000 Gent, Belgium
Tel. 32.(0)2.380.14.10
Fax 32 (0)2 381 16 45
GSM Mobile telephone 32.(0)475.44.99.55
Air-freshners
Following substances were found in some air-freshners:
Benzene, formaldehyde, styrene, acetaldehyde, toluene, chlorobenzene, naphthalene, glycolethers,
ftalates, artificial musks,paradichlorobenzene, benzylacetate, benzylalcohol, dipropyleen glycol, terpenes.
Combined use of ozone generators and air-freshners leads to an increase in the number and
concentration of fine particles
Exposure to air-freshners induced airway irritation and changes in behaviour in mice
Dipropylene glycol was found (National Toxicology Program, 2004) to cause nephropathy, liver toxicity
and decreased body weight in rats
Use of air-freshners was associated with diarea and ear-ache in infants and with headache and
depression in mothers
It was concluded that the use of air-freshners is contraindicated.
Benzene
Mutagenic metabolites and oxidative stress. Has radiomimetic properties
epigenetic action: receptor mediated disruption of stem cell differentiation in bone marrow towards myeloid
precursors
Metabolites hydroquinone and fenol might inhibit GJIC and act as tumorpromotors
In experimental animals benzene causes cancers in many organs, suggesting
There are at least 9 publications in the literature in which benzene was associated with cancers other than
hematopoietic in humans
It seems likely that benzene is recognized as causing mainly or only hematopoietic malignancies, because of its
relative importance as a hematopoietic carcinogen , and because of the fact that its effect on other tissues is masked
by a high background of other carcinogenic influences on these tissues. Formerly, ionising radiation was considered
to be mainly a hematopoietic carcinogen.
Probably benzene causes many different types of cancer in humans, as does ionising radiation.
Benzene risk estimations from the American Petroleum Institute and even those of EPA (2,2 to 7,7 deaths from
hematopoietic malignancies/milion for life time exposure to 1 µg/m3) probably suffer from substantial
underestimation.Our best estimate was 12,3.
Some reasonable worst case estimates (of the order of 50 deaths from hematopoietic malignancies/milion for
life time exposure to 1 µg/m3) were presented.
Gap Junction Intercellular Communication (GJIC)
Important in growth, differentiation, development, homeostasis and carcinogenesis
Tumor-promotion: reversible inhibition of GJIC might contribute to selective expansion of initiated cells
Most tumorpromotors inhibit GJIC.
Tests for detection of tumorpromoting activity might be usefull (as are tests for mutagenic activity)
Simple tests based on GJIC could be used to detect tumorpromoting activity in environmental samples
In collaboration with Dr. Trosko we propose the use of such tests to the Flemish authorities
GSM and UMTS
Typical GSM (data 1999): a SAR= 1.6 W/KG in the brain
Antenna’s: generally a much lower exposure (8 seconds of GSM = 24 h at 100 m of a typical antenna)
Biological effects at SAR=4W/kg
In vitro effects on gene expression at exposures not far above those occuring in GSM users
Epidemiological studies: unlikely to have sufficient sensitivity and sufficient discerning power.
More experimental work, in vitro and on animals, recommended. Interactions with at least some important exposures
occurring in real life should be included.
High Tension Cables
ELF magnetic fields above 0.4 Tesla might increase risk of cancer.
Indoor magnetic fields above 0.2 Tesla almost always due to outdoor sources.
Epidemiological studies: often association between 50 Hz-elektromagnetic fields and leukemia in
children and chronic lymfoid leukemia in occupationaaly exposed adults adults.
Further experimental research is indicated