TY Or High-selenium wheat : biofortifÏcation better health Graham Henry Lyons Thesis by pubtication submitted to The University of Adelaide for the degree of Doctor of Philosophy Discipline of Plant and Pest Science School of Agriculture and Wine Faculty of Science The lJniversity of Adelaide June 2004 1.1 lurnooucrloN 1 l.l.l. Selenium: the essential metalloid 1 1.1.2. Selenium in soils: ubiquitous and variable.... 2 1.1.3. Selenium in plants: important sourcefor animals and humans 4 1.1.3.1. Se levels in plants and their effects.. 4 1.I.3.2. Factors that effect Se uptake by plants 4 1. 1.3.3. Short-distance transport 5 1.1.3.4. Long-distance transport and storage 6 1.1.4. Selenium in animals: more likely too little than too much . 7 1.2. SprBNtuM: ESSENTIAL FoR HUMAN HEALTH.... 8 1.2.1. Introduction I I . 2. 2. Selenoprotein activities... 9 l. 2. 3. Selenium deficiency diseases 9 1.2.4. Immunefunction l. 2. 5. Thyroid function 1.2.6. Cancer............... 1.2.7. Viral diseases.... I . 2. 8. Cardiovascular disease 1. 2.9. Other oxidative stress/inflammatory conditions . 1.2.10. Fertility 1.2.11. Detoxffication of heavy metals....... 1.3. Huvt¡.N SELENILIM INTAKE: vARIABLE BUT MosrLY Too Low..........'..'. 1.3.L Selenium intake in humans 1.3.2. Human blood concentrations of selenium: the global view........'... 1.3.3. Selenium levels in the Australian population 1.3.4 Selenium toxicity: garlic breath and cracked nails 1.3.5. Optimal selenium intake 1.4. SrnerEGrES To INCREASE HUMAN SELENIUM INTAKE... 1.4.1.Increased consumption of higher-seleniumfoods through education..... I .4. 2. Individual supplementation 1.4. 3. Food þrtification................ 1.4.4. Selenium supplementation of livestock 1 . 4. 5. Selenium fertilis ation of crops ................ I . 4. 6. Pl ant breeding for enhanced s el enium accumulation ........'..'. 1.5. WHB¡.I: AN IMpoRTANT SELENIUM souRCE FoRHUMANS.... 1 . 5 . I . Selenium concentrations in wheat grain......... 1.5.1. 1. The global vrew ............ 1.5.1.2. Selenium in Australian wheat... 1 . 5 . 2. High bio availability of wheat-s elenium 1.5.3. Effects of post-harvest processing and cooking on selenium 1.5.3.1. Milling 1.5.3.2. Cooking.......... 1.6. SIn¡,TEGIES To INCREASE SELENruM IN wHEAT 1.6.l. Introduction l. 6. 2. Selenium fertilisation......... 1.6.2.1. Se form; method, timing and rate of application... 1.6.2.2. Effect of plant nutrients on Se uptake....... Sulphur... Nitrogen Phosphorus........... I .6.2.3 . Se toxicity to wheat: effects on germination and early growth . 1.6.3. Weat breeding to increase grain selenium density...... 1.6.3.1. The extent of human micronutrient malnourishment..... 1.6.3.2. Breeding for higher nutrient density in staple crops......... 1.6.3.3. Screening and selection; the importance of genotype-environment interaction 42 1.7. CoNcrusloN.. 44 2. LrST OF RESEARCH QUESTTONS FOR THIS THESrS... .................45 3. ARTrCLES.......... ........46 3.1. High-selenium wheat: biofortification for better health. 3.2. Nutriprevention of disease with high-selenium wheat. 3.3. Trends in selenium status of South Australians. 3.4. Trend in selenium status, and current blood levels of mineral nutrients, of healthy S outh Australian residents. 3.5. High-selenium wheat: agronomic biofortification strategies to improve human nutrition. 3.6. Exploiting micronutrient interaction to optimize biofortification programs. 3.7. Selenium concentration in wheat grain: is there sufficient genotypic variation to use in breeding? 3.8. Distribution of selenium and other minerals in wheat grain, and the effect of processing on wheat selenium content. 3.9. Tolerance of wheat (Triticum aestivum L.) to high soil and solution selenium levels. 4. GENERAL DISCUSSION........... ........47 5. REFERENCES FOR LITERATURE REVIE\ü & GENERAL DISCUSSION....57 Abstract Selenium (Se) is an essential micronutrient for humans and animals, but is defrcient in at least a billion people worldwide, and in some regions appears to be declining in the food chain. V/heat (Triticum aestivum L.) is a major dietary source of Se in many countries, including Australia. The largest survey to date of Se status of Australians found a mean plasma Se concentration of 103 p"/l in 288 Adelaide residents, just above the nutritional adequacy level. In the total sample analysed (six surveys from 1977-2002; n: 834), plasma Se was higher in males and increased with age. This study showed that many South Australians consume inadequate Se to maximise selenoenzyrne expression and cancer protection, and indicated that levels had declined around 20Yo ftom the 1970s. No significant genot¡pic variability for grain Se concentration was observed in modern wheat cultivars, but the diploid wheat Aegilops tauschii L. and rye (Secale cereale L.) were higher. Grain Se concentrations ranged 5-720 þglkg and it was apparent that this variation was determined mostly by available soil Se level. Field trials, along with glasshouse and growth chamber studies, were used to investigate agronomic biofortification of wheat. Se applied as sodium selenate at rates of 4-120 {ha Se increased grain Se concentration progressively up to 133-fold when sprayed on soil at seeding and up to 2O-fold when applied as a foliar spray after flowering. A threshold of toxicity of around 325 mglkg Se in leaves of young wheat plants was observed, a level that would not normally be reached with Se fertilisation. On the other hand suiphur (S) applied at the low rate of 30 kg/ha at seeding reduced grain Se concentrationby 160/o. This study showed that, although Se concentration was highest in the embryo of wheat grain, Se and S were more evenly distributed throughout the glain, and hence a lower proportion was removed in the milling residue, than for other mineral nutrients. Post- milling processing of wheat, including baking and toasting, is unlikely to result in reduction of Se content. Agronomic biofortification could be used by food companies as a cost-effective method to produce high-Se wheat products that contain most Se in the desirable selenomethionine iv form. Further studies are needed to assess the functionality of high-Se wheat, for example short-term clinical trials that measure changes in genome stability, lipid peroxidation and immunocompetence. Increasing the Se content of wheat is a food systems strategy that could increase the Se intake of whole populations. V Declaration This work contains no material which has been accepted for the award of any other degree or diploma in any university or other l'rufüary institution and, to the best of my knowledge and beliet contains no material previously published or written by another person, except where due reference has been made in the text. I give consent to this copy of my thesis, when deposited in the University Library, being available for loan and photocopylng. Graham Lyons V1 Acknowledgements I wish to sincerely thank the Grains Research and Development Corporation for funding and the University of Adelaide, School of Agriculture and Wine for hosting this PhD study. ln addition, I would like to express my gratitude to the following people: . My supervisors, Professor Robin Graham and Dr James Stangoulis for their inspirational guidance, insight, support, patience and friendship throughout this study. Also thankyou to Robin for funding me to attend the International Plant Nutrition Colloquium, Hanover, 2001 and the International Symposium on Trace Elements and Minerals in Medicine and Biology, Munich,2004. 'Waite o Analytical Services (Teresa Fowles, L¡mdon Palmer, Nick Dykshoorn, Carolyn Jones, Deidre Cox and Kirstie Mclaren) for accurate and timely analyses of selenium and other micronutrients. o Jim Lewis for his expert management of field trials, maintenance of growth chambers, and advice. My colleagues and friends in the Plant Nutrition Group, in particular Yusuf Genc, Julia Humphries, Nick Dykshoom, Lam Huynh Bao and Julie Kitchen for their support and frequent valuable advice and technical assistance. My co-authors, including Robin Graham, James Stangoulis, Geoff Judson, Ivan Ofüz- Monasterio, Yusuf Genc and Lyndon Palmer. ¡ Bob Holloway and Dot Brace and their colleagues at the Minnipa Agricultural Centre for managing the selenium freld trial. o Michelle Lorimer and Janine Jones, BiometricsSA for their statistical advice and analyses. o Graeme Mclntosh, CSIRO Human Nutrition for stimulating discussion on selenium. o Michael Mackay, Greg Grimes (Australian Winter Cereals Collection, Tamworth), Gil Hollamby, Jim Lewis, Wayne Veitch, Rob Wheeler, Ursula Langridge, Steve Jefferies, Jeremy Lemon, David Cooper, Alan Fishie, Yusuf Genc, Steve Hughes, Carolyn Schultz, Andrew Rogers, David Gibeaut and Kath Cooper for supplying seed. o Peter Smith, Andrea Hensing, Leanne Mosionek, Marie Vawser, Kolumbina Mrva, Rajinder Sharma and Daryl Mares for assistance with milling and grain quality issues. o The V/aite library staff for locating and providing numerous articles. o Kathleen Doherty and Sue Heatley (Australian Red Cross Blood Service, Adelaide) for assistance with blood collection. o Kath Cooper for funding the purchase of high-selenium triticale and wheat from Frances, SE. o Geoff Judson (SARDD for financial assistance with conducting blood selenium analyses. o Margaret Cargill, Dr Margaret Rayrnan and Professors Gerry Combs, Ross Welch and Gunnar Gissel-Nielsen for valuable technical advice. o Finally, and most importantly, I would like to thank my family: my wife Susan and daughters, Georgina and Henrietta, for their love, patience and support throughout this study. 1 1. Literature Review 1.1 Introduction A review of literature on selenium (Se) in soils, plants, animals and humans is presented. The importance of Se in human health is discussed, followed by Se intake by humans, with a focus on Australia. Strategies to increase Se intake are discussed briefly. The review then examines wheat as a major source of bioavailable Se, and discusses Se fertilisation and plant breeding, two strategies to increase Se density in wheat grain.