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NanoSIMS Analysis of Arsenic NanoSIMS Analysis of Arsenic and Selenium in Cereal Grainsand Selenium in Cereal Grains
Supervisor: Chris GrovenorSupervisor: Chris Grovenor
Katie MooreKatie Moore33rdrd year D.Phil year D.Phil
Department of Materials – University of OxfordDepartment of Materials – University of Oxford
Motivation
Why is a materials scientist looking at plants?
Interdisciplinary collaborations allow critical problems in the life sciences, difficult to solve with traditional analysis techniques, to be explored with established physical science techniques.
The Arsenic Problem• Arsenic contamination
of groundwater
• Contaminated groundwater is used to irrigate rice paddy fields
• Resulting in rice grain containing elevated levels of arsenic
• A major problem in Bangladesh, India, China and America.
Ref: X. Y, Xu et al., Environ. Sci. Technol., 42(15), 2008
• Arsenic is a toxic and carcinogenic element
The Selenium Problem• Between 0.5 and 1
billion people worldwide may be deficient in selenium including populations in developed countries.
• In the UK this is caused by a reduction in the amount of wheat imported from America and a fall in the consumption of cereals
• Selenium is an important trace elementDaily selenium intake in the UK is
now about ½ of the reference amount
M. R. Broadley et al., Proc. Nutr. Soc. (65) 2006M. S. Fan et al., Sci. Total Environ. (389), 2008MAFF, Food Surveillance Information Sheet, (126), 1997
Refs:
Agricultural Solutions
• To increase Se:– Add a selenium fertiliser to the soil (practiced in Finland)
• To decrease As:– Polish the grain to remove the high As parts
• Both of these solutions require knowledge of where the trace elements are located in the grain.
• Determining where these very low concentrations are located with sub-cellular resolution is a serious analytical challenge
Ref: M. H. Eurola et al., J. Sci. Food Agric., (56), 1991
Secondary Ion Mass Spectrometry (SIMS)
• Sample is bombarded by positively charged primary ion beam
• This results in sputtering of the top few atomic layers and ejection of atoms, ions and clusters
• Secondary ions are collected and mass analysed
Image adapted from Ref:http://www.eaglabs.com/training/tutorials/sims_theory_tutorial/index.php
The NanoSIMS 50
The Oxford NanoSIMS
Schematic of the NanoSIMS
Ref: CAMECA, http://www.cameca.fr/doc_en_pdf/ns50_instrumentation_booklet.pdf, Instrumentation booklet, June 2007.
Characteristics of SIMS• SIMS
– High sensitivity (down to ppb for some elements)
– Detection of all elements from Hydrogen to Uranium including all isotopes
– High mass resolution
• NanoSIMS – High lateral resolution (50 nm)– Parallel detection of 5 ionic species
Ref: CAMECA, http://www.cameca.fr/doc_en_pdf/ns50_instrumentation_booklet.pdf, Instrumentation booklet, June 2007.
SIMS Sample Preparation
• Sample needs to be flat, conducting, and dry
• Bulk chemical analysis (ICP-MS) showed
trace levels of 2.5 ppm arsenic in the rice and 17 ppm selenium in the wheat
• Rice samples were
grown at Rothamsted Research
• Wheat samples were grown
in a field trial in Nottingham
Structure of Wheat GrainAleurone layer
Starchy endosperm
80µm Embryo
Cross section
Selenium in Wheat Grain
Max selenium counts: 4Max CN- counts: 105,000
31P16O- 80Se-
32S-12C14N-16O-
SE
30µm
Ref: K. L. Moore et al., New Phytol., (185), 2010
Selenium in Wheat GrainAleurone cell Starch grains
31P16O- 80Se- 32S-
12C14N-16O- 12C14N-16O-
80Se-
Ref: K. L. Moore et al., New Phytol., (185), 2010
Selenium in Wheat Grain
High resolution, sub-cellular, localisation of ppm concentrations
Selenium is localised in the protein region around the starch grains
Selenium hotspots are found in the aleurone cells
Starchgrain
31P16O- 80Se- 32S-
16O- 12C14N-16O-
80Se-
32S-
Ref: K. L. Moore et al., New Phytol., (185), 2010
Arsenic in Rice Grain
Arsenic is localised in the sub-aleurone protein
Ref: K. L. Moore et al., New Phytol., (185), 2010
Rice Roots – Experiment setup
Fe plaque No Fe plaque
Variables:•Arsenate or arsenite•With or without Fe plaque•Wild type or lsi2 mutant
Hydroponically grown rice plants
Lsi2 transporter
Ref: Zhao, F.J., et al., New Phytol., 181(4), 2009
Rice Roots – Fe Plaque
25 µm
EPEPExEx
ScSc
SESE
12C14N-
56Fe16O-
31P-28Si-
75As-
Rice Roots – Lsi2 mutant12C14N-
75As-
31P-
SE
28Si-
25 µm
XyXy
EnEn
Colour merge: Red = As, Green = CN, Blue = Si
Conclusions• The NanoSIMS has successfully been used to provide a detailed
analysis of the distribution of trace elements selenium and arsenic in wheat and rice respectively and the distribution of As in roots.
• Selenium is localised in the protein regions around the wheat starch grains with hotspots in the bran layer
• Arsenic is concentrated in the sub-aleurone protein of the rice rather than in the aleurone.
• The Fe plaque has a strong adsorption affinity for As
• The Lsi2 mutant blocks As uptake in the endodermis
• These experiments have demonstrated the unique potential of state-of-the-art SIMS instrumentation to analyse the distribution of ppm levels of important trace elements with sub-cellular resolution
AcknowledgementsSupervisor: Chris GrovenorNanoSIMS postdoc: Markus Schröder
EPSRC:D.Phil funding
Root Sample Preparation: Barry Martin, Chris Hawes
Collaborators:Fang-jie Zhao, Steve McGrath,Malcolm Hawkesford, Peter Shewry
IOM3:For this opportunity