Heavy metals (Zn, As, V, Cu, Co, Ni, U, Pb, and Cr) in archived human

samples NKS NORDIC ICP seminar

26th September, 2017

Susanna Salminen-Paatero & Jussi Paatero

susanna.salminen-paatero@helsinki.fi

Heavy metals in environment and man • Both from natural and anthropogenic origins

• soil erosion and weathering of the crust, acidification of environment, mining, industrial effluents and other wastes, sewage discharge, pesticides, microplastics in oceans…

• Some heavy metals are nutrients having essential biological functions in animals and plants (Fe, Mn, Co, Cu, Zn, Cr, V, Ni…), but they are poisonous in high concentrations

• Biological effect depends on heavy metal and its chemical form

• malfunctioning of cells, leading to DNA damage

• Enrichment to different organs (for example, uranium to kidneys , bones, etc.) → damaged organ

• Pb, Cd, Hg are very poisonous

• eventually all heavy metals are toxic and/or carcinogenic in high concentrations

Project ”Food and health security” (EU, 2013-2015), Finnish Meteorological Institute was one of the participants

• WP 1: Human exposure assessment and identification of dietary sources of exposure across border areas

• WP 2: Contaminants in relevant food items – geographical differences and trend data

• WP 3: Health effects of contamination in the region – status and future predictions

• WP 4: The socioeconomic consequences of contamination and food safety in the region

• WP 5: Public awareness and informed policy decisions – addressing human security

Part of this work became ”preliminary study of human samples: heavy metals and plutonium in man”

Heavy metals – health risks – Lapland – what samples should we study, where to get the samples? • The work start

was delayed for a year for other duties •During that

year, an old freezer was found while emptying a sample storage

Frozen whole lungs, ribs, liver pieces, lymph nodes, etc. had been stored since the 1970s, after a completed Ph.D. project. There were many plastic bags full of organs!

Background of the samples • People who died in accidents, seizures/attacks, or in other unclear

circumstances (murder, suicide) in 1976-1979

→ Need for an autopsy at Department of Forensic Medicine

• Northern Finland: 5 men, Southern Finland: 67 men, women and children

• liver, lungs and bone (ribs) were taken within each autopsy, occasionally also blood, urine, lymph nodes, collarbones, testicles (plus testes), etc.

• Helena Mussalo-Rauhamaa (Ph. D. work in 1981): ”Accumulation of plutonium from fallout in southern Finns and Lapps”

* The differences in radionuclide concentrations between habitats of Northern and Southern Finland were explained with different radioactivity levels in surface air (inhalation) and diet – people in Lapland ate mainly reindeer and

fish at their lifetime

Ideas for using these archived human samples (if funding!)

• Heavy metals in lung, liver, lymph nodes, flesh and bone (ribs)

• Strontium in bones

• Plutonium in lungs, liver and bones, isotope ratios of plutonium in different organs

• Americium in different organs → 241Pu

• Differences between enrichment of heavy metals and radionuclides in different organs

• Differences between Northern and Southern Finland

• Only preliminary method for these samples has been tested!

Compromises made for simultaneous determination of heavy metals and radionuclides

• Both heavy metals and radionuclides were determined from the same sample, to get as similar sample as possible for these two determinations (heavy metals and radionuclides) representing the whole organ

• Very big sample amount was needed due to low radionuclide content of the samples (Pu-act. conc. ~µBq/g wet weight)

→ The whole lung or set of ribs was used!

→ Microwave oven could not be used

→ A lot of disturbing matrix, like iron, calcium, phosphates…

→ After preliminary tests, maybe two subsamples with separate dissolution procedures for heavy metals and radionuclides would be good enough…

Sample set for preliminary tests

• liver (2)

• lung (16)

• lymph nodes (7)

• ribs (3)

• flesh (2)

• collarbone (1)

Procedure

• Drying at 105 °C for 48 hours

• Ashing at 450 °C (not for flesh and lymph nodes)

• Wet-ashing with HNO3, ≥ 6 hours, H2O2 in the end

• Filtration

• Fraction for heavy metal determination

• Filtration

• Dilution to 5%-33% HNO3 with mQ-water

• Determination of heavy metal concentrations

A frozen collarbone

A big lung, weight 2100 g.

Tracheobronchial lymph nodes (collected from the junction of tracheal and bronchial tubes)

Ribs

BBQ-time: The samples are dried in 105 °C for 48 h. Observation: human ribs smell like pork meat.

Flesh is peeled off from dried ribs

Peeled and cut-up ribs are ashed in 450 °C overnight

Dried human flesh – it was directly wet-ashed without ashing

Traditional wet-ashing on a hot plate and in Kjeldahl flasks

Problems with the method: protein (?) precipitation

• Some lung and liver samples were very bloody, giving dark colour to final measurement samples (high iron content): after filtration and dilution with water, red-brown precipitation occurred immediately • Further dilution of the samples did not help much •Higher HNO3-concentration (5% → 33%) of measuring

samples prevented precipitation in some cases → it was not possible to use ICP-MS due to clogging, instead, MP-AES was used for determining heavy metals

Determination of heavy metals from sample solutions

• Agilent 4100 Microwave Plasma-Atomic Emission Spectrometer

• The samples were measured at Department of Geography, UH

• Samples, blanks, and QC samples were measured

Comparison of different instrument types (borrowed from the web)

Heavy metals in human organs determined with MP-AES (µg/g wet wt.)

Results for different metals, brief comparison • No cobalt in any of the samples

• lead only in 9 lung samples (tobacco smokers), <DL – 0.99±0.20 µg/g (wet wt.)

• Tobacco smokers have generally elevated concentrations of lead, nickel, cadmium (excluded from our study) and arsenic

Co: -, Cr: rib -, collarbone 0.24±0.05, liver -, lung <DL - 14±3, Ni: rib -, collarbone 1.36±0.27, liver 1.31±0.26 and 1.61±0.32, lung 3.74±0.75 (µg/g)

Cu: rib -, collarbone 1.88±0.38, liver 6.4±1.3 and 7.9±1.6, lung 1.96±0.39 (µg/g)

Zn: rib 15±3, collarbone 99±20, liver 79±16 and 141±28, lung <DL - 52±10 (µg/g)

Vanadium: concentrations µg/kg (wet wt.) Bone Liver Lung Time and

country Ref.

<1-8 5-19 12-140 Before 1978, former Yugoslavia (?)

[1]

<125 <125 <125 1997-1999, Spain

[2]

190-1400 Poland [3]

3000-4100 2400 and 5200

1300-13000 1976-1979, Finland

This study

Arsenic: concentrations µg/g (wet wt.) Bone Liver Lung Time and

country Ref.

<0.05 <0.05 <0.05 1997-1999, Spain

[2]

0.07-0.16 ~2010, Poland [4]

Ribs:- Collarbone: 0.04

- <DL - 0.11 1976-1979, Finland

This study

Uranium : concentrations µg/kg (wet wt.)

Bone Liver Lung Time and country

Ref.

3.8 0.08-0.24 0.67-4.1 ~1989, USA [5]

7 1997, Global median

[6]

61 1997, China [6]

0.43 ~1986, New York

[7]

3 ~1972, UK [8]

180-460 2900 and 4500

680 – 7300 1976-1979, Finland

This study

Uranium conc. in Finnish samples – high or not?

• Ribs 180-460 µg/kg → 2- 6 Bq/kg, or 2-6 mBq/g

• Liver 2900 and 4500 µg/kg → 36 and 56 mBq/g

• Lung 680-7300 µg/kg → 8 – 91 mBq/g

Average U-conc. in drinking water in Finland: 30 mBq/l, varies from 15 mBq/l (average value for tap water from water supply network, maximum value 1800 mBq/l) to 400 mBq/l (average value for drill well water, maximum value 150 Bq/l)

General population of Colorado, 5 subjects:

Ribs 0.04 mBq/g, Liver 0.007 mBq/g Lung 0.05 mBq/g (M.E. Wrenn et al., 1985)

Future improvements for the dissolution method • All biological material in the samples must be dissolved by a proper

decomposition method

• Small subsample of an organ instead of total organ dissolution – representative sample enough?

→ possibility to use of microwave oven

* Total dissolution of the biological sample

* H2O2 is not needed then (can contain impurities)

• Addition of complexing agent (e.g. EDTA)?

• (Dissolution of biological samples to amino acid?)

→use of ICP-MS instead of MP-AES, lower detection limit and measurement uncertainty!

Conclusions

• Concentration values are mainly at the same level as in references worldwide, but some exceptionally high values occur in Finnish samples

• Comparison between studies is difficult due to generally small sample number in each study and differences between exposure way and exposure level of individuals

• More data (and funding) is needed

• a revised method is needed

• More literature references are needed for producing an article

Acknowledgements

• Dr. Juhani Virkanen (UH, Department of Geography) for measuring the samples with MP-AES

• Project “Food and health security in the Norwegian, Russian and Finnish border regions: linking local industries” (EU´s Kolarctic ENPI CBC 2007-2013 Programme)

References

1. Byrne AR, Kosta L. Vanadium in foods and in human body fluids and tissues.. Sci Total Environ. 1978 Jul;10(1):17-30

2. F. García , A. Ortega , J. L. Domingo & J. Corbella (2001) ACCUMULATION OF METALS IN AUTOPSY TISSUES OF SUBJECTS LIVING IN TARRAGONA COUNTY, SPAIN, Journal of Environmental Science and Health, Part A, 36:9, 1767-1786

3. Lanocha-Arendarczyk et al. Influence of Environmental Factors and Relationships between Vanadium, Chromium, and Calcium in Human Bone. BioMed Research International,Volume 2016, Article ID 8340425

4. Barbara Brodziak-Dopierała, Jerzy Kwapuliński, Jolanta Kowol. Occurrence of Arsenic in Selected Parts of the Human Femur Head. Pol. J. Environ. Stud. Vol. 20, No. 6 (2011), 1633-1636

5. R.L. Kathren, S.Y. Tolmachev. Natural Uranium Tissue Content of Three Caucasian Males. Health Physics 109, 187-197 (2015).

6. L. TANDON, G. V. IYENGAR and R. M. PARR. A Review of Radiologically Important Trace Elements in Human Bones. Appl. Radiat. Isot. Vol. 49, No. 8, pp. 903±910, 1998

7. Fisenne, I. M. and Welford, G. A. (1986) Natural U concentrations in soft tissues and bone of New York City residents. Health Phys. 50: 739–746.

8. Hamilton, E, I. (1972) The concentration of uranium in man and his diet. Health Phys. 22: 149–153.