1) Zinc and iron deficiencies affect approximately 2 billion people globally and are a major cause of malnutrition, particularly in developing countries where cereal-based diets provide most calories. 2) Biofortification through agronomic practices such as fertilization can increase micronutrient concentrations in crops and provide a potentially sustainable solution. Application of zinc-containing fertilizers to the soil and foliage has been shown to reliably increase zinc levels in grains across multiple countries. 3) Foliar application of zinc at different crop growth stages affects the distribution and concentration of zinc deposited in different grain tissues. Higher nitrogen fertilization can also influence grain zinc and iron concentrations.

1. SYMPOSIA:
Biofortification through
Agronomic Practices

2. Zinc
Estimated
2 billion
Zn, Fe, Se and I Deficiencies:
Global Malnutrition Problem
www.harvestplus.org
Iron
Estimated 2
billion

3. Children particularly sensitive
>450,000 deaths/year
children under 5 – 4.4%
attributed to Zn
deficiency
Black et al. 2008
The Lancet Maternal and Child
Undernutrition Series

4. “ZINC SAVES KIDS”
Campaign
International Zinc Association

5. Major Reason: Low Dietary Intake
High Consumption Cereal Based Foods
with Low Micronutrient Concentrations
In number of developing countries,
cereals contributes nearly 75 % of
the daily calorie intake.

6. Solutions to Micronutrient Deficiencies
• Supplementation
• Food Fortification
(not affordable in rural regions)
Golden Wheat Fortfied with Zn

7. Increase in Concentration of
Non-Nutritive Elements
(especially Cadmium and
Arsenic) in Food Crops is a
Growing Concern

8. Agricultural Solutions
(Breeding and Fertilizer Approaches)
•Breeding
•Agronomy/Fertilizers

9. Biofortification through
Agronomic Practices
Short-Term Solution
F. Zhang

10. SYMPOSIA SPEAKERS
Ismail Cakmak (Sabanci
University, Istanbul)
• Fertilizer Strategy for
Improving Grain Zinc and
Iron Concentrations
Maha V. Singh (Indian Institute
of Soil Science -ICAR)
• Detrimental Effects of Soil
Zinc Deficiency on Crop
Production and Human
Nutrition in India
Graham Lyons (University of
Adelaide Waite Campus)
• Agronomic Biofortification
to Reduce Selenium
Deficiency in Human
Populations: Achievements
and Challenges
Cynthia Grant (Agriculture and
Agri-Food Canada)
• Agronomic Practices to
Reduce Non-Nutritive
Elements in Food Crops
Rapporteur: Rufus Chaney,
Research Agronomist, USDA

11. Fertilizer Strategy for Improving Grain
Zinc and Iron Concentrations
Ismail Cakmak
Sabanci University, Istanbul

12. Zn Deficiency:
Global Micronutrient Deficiency in Soils
Alloway, 2007. IZA Publications, Brussels
Widespread
Medium

13. +Zn
-Zn
When Zn is deficient in soil or plant
Grain Zn:
12 mg kg-1
Grain Zn:
35 mg kg-1

14. A successful breeding program for biofortifying
cereals with micronutrients greatly depends
on the amount of plant-available pools of
micronutrients in the soil and/or leaf tissue for
translocation into grain
On soils with low plant available Zn: 8-15 ppm
On soils with adeqate plant available Zn: 20-35 ppm
Cakmak et al., 2010 Cereal Chem.
Range of Grain Zn Concentration
in Wheat in Turkey:

15. For a better Zn and Fe nutrition of human
beings, cereal grains should contain around
40-60 mg Zn or Fe kg-1
Current Situation:
10-30 mg kg-1

16. Human Zinc Deficiency
Moderate
Not sufficient data available
Low
High
http://www.izincg.org/
Widespread Zn Deficiency
Medium Zn Deficiency
Alloway, 2004. IZA Publications, Brussels
Soil Zinc Deficiency
Soil and Human Zn Deficiency: geographical overlap

17. Application of Micronutrient
Fertilizers
Application of Zn- or Fe-containing fertilizers
offers a rapid solution to the problem, and
represents an important complementary
approach to on-going breeding programs for
developing new genotypes with high Zn or Fe
density in grain.

18. Global Zinc Fertilizer Project

19. Canada Germany
Brazil
Zimbabwe
Zambia
Mozambique
South Africa
Ethiopia
Pakistan
India
Kazakhstan
Iran
Laos
China
Thailand
Australia
Turkey
International Zinc Assoc.
Coordinating Institution: Sabanci University
Global Zinc Fertilizer Project
Mexico

20. Project objectives
• to test Zn-containing N-P-K fertilizers for increasing root
Zn uptake and improving grain Zn concentration
• to identify the effective foliar Zn application for promoting
Zn accumulation in the grain
• to determine an effective combination of soil and foliar
application of Zn fertilizers for increasing Zn
concentration in the grain
• to characterize seed deposition of the leaf-applied Zn
• to achieve capacity building through close cooperation
and dissemination activities among scientists,
agronomists, extension staff and local farmers in the
target countries

21. 1) Standard/ Local Farmers’ Application (LS)
2) LS + Soil Zn (50 kg ZnSO4/ha)
3) LS +Foliar Zn (0.5 % for wheat and rice, 0.3 % for maize).
4) LS + Soil Zn+ Foliar Zn
5) LS + Foliar OMEX-Type-I
6) LS+ OMEX-Type-II
7) LS+Urea-Zn + adjustments (same rates of N, P, K
8) LS+Mosaic MESZ-Zn (and + adjustments)
9) LS+KALI KornKali-Zn (and + adjustments)
10) LS+KALI Korn Kali-Zn + Foliar Zn (+ adjustments)
11) LS+ Mosaic MESZ-Zn + Foliar Zn (+ adjustments)
12) LS+Urea MESZ+Kali+ foliar Zn
Fertilizer Treatments

22. Effect of Soil and/or Foliar Applied
ZnSO4 on Grain yield and Grain Zn
Concentrations in Wheat
Soil Zn Application : 25 to 50 kg ZnSO4.7H2O ha-1
Foliar Zn Application: Generally 2 times: before and
after flowering (1 to 4 kg ZnSO4 ha-1)

23. Clinton Global Initiative highlighted the importance of
Zn-containing fertilizers to alleviate Zn deficiency
problem in human populations at 5th Annual Event in
September 24, 2009

24. Rice Trials in Thailand

25. Maize Trials in Zambia

26. Wheat Trials in India

28. Maize Trials in Zimbabwe

29. Trials in Pakistan

30. Maize Trials in Mozambique

31. Wheat trials in China, Yanglin-Xian

32. 0
2
4
6
8
10
0
20
40
60
80
INDIA - Punjab State
(Partner: Punjab Agricultural University)
Grainyield,tonha-1
GrainZn,mgkg-1
1. Control
3. Foliar Appl.
4. Soil + Foliar Appl.
Zn Applications
2. Soil Appl.
Location-I Location-II
25
61
49
65

33. 0
2
4
6
8
0
10
20
30
40
50
60
Grainyield,tonha-1GrainZn,mgkg-1
1. Control
3. Foliar Appl.
4. Soil + Foliar Appl.
Zn Applications
2. Soil Appl.
Location-I Location-II
27
48
28
44
PAKISTAN
(Partner: Pakistan Atomic Energy Comission)

34. 0
1
2
3
4
0
10
20
30
40
Control (No Zn)
Booting + Milk
Milk + Dough
Foliar Zn Applications
Stem + Booting
No Soil Zn Appl.
With Soil Zn Appl.
12
27
22
29
Grainyield,tonha-1GrainZn,mgkg-1
-Zn +Zn
-Zn +Zn
Cakmak et al., 2010,
J. Agric. Food. Chem.
TURKEY
(Partner: Ministry of Agriculture)

35. 0
10
20
30
40
50
60
70
GrainZn,mgkg-1
1. Control
3. Booting
4. Tillering + Booting
Foliar Zn Spray
2. Tillering
5. Milk
Cultivar
Akmola
Cultivar
Trizo
KAZAKHSTAN
(Partner: CIMMYT Kazakhstan)
WHEAT
19
54
18
44

36. Grain Zn concentration in different countries with and without zinc fertilization
Average of all countries -Zn: 26 +Zn:50
Country/Location -Zn +Zn
India
Varanasi 29 47
PAU-I 25 81
PAU-II 28 77
PAU-III 26 61
PAU-IV 49 65
IARI 33 45
Kazakhstan
Loc-I 19 54
Loc-II 28 73
Pakistan
Loc-I 27 48
Loc-II 28 44
Loc-III 30 40
Loc-IV 29 60
mg kg
-1
Country/Location -Zn +Zn
Mexico
Year-I 21 45
Year-II 36 60
Turkey
Konya 12 29
Adana 32 57
Samsun 23 49
Eskisehir 22 43
China
Loc-I 28 54
Loc-II 19 26
Australia
Loc-I 18 39
Germany
Average 20 32
Iran
Average 17 28
Brazil
Average 30 52
mg kg
-1

37. Country/Location -Zn +Zn
India
Varanasi 29 47
PAU-I 25 81
PAU-II 28 77
PAU-III 26 61
PAU-IV 49 65
IARI 33 45
Kazakhstan
Loc-I 19 54
Loc-II 28 73
Pakistan
Loc-I 27 48
Loc-II 28 44
Loc-III 30 40
Loc-IV 29 60
mg kg
-1
Country/Location -Zn +Zn
Mexico
Year-I 21 45
Year-II 36 60
Turkey
Konya 12 29
Adana 32 57
Samsun 23 49
Eskisehir 22 43
China
Loc-I 28 54
Loc-II 19 26
Australia
Loc-I 18 39
Germany
Average 20 32
Iran
Average 17 28
Brazil
Average 30 52
mg kg-1
Average Concentrations
of Grain Zn
(10 Countries with 32 locations)
-Zn: 26 ppm
+Zn: 50 ppm
Grain Zn concentration in different countries with and without zinc fertilization

38. Staining/Localization of Zinc in
Wheat Grain (red color)
EMBRYO
ENDOSPERM
ALEURONE
ALEURONE
Cakmak et al., 2010
Cereal Chemistry, 77: 10-20

39. Localization of Zn in grain
after foliar application?

40. LA-ICP-MS Tests on Seeds
0
100
200
300
400
500
600
700
800
900
1000
0 500 1000 1500 2000 2500 3000 3500 4000
Distance (µm)
Znconcentration(mg/kg)
0.0
2.0
4.0
6.0
8.0
10.0
12.0
0 500 1000 1500 2000
Distance (µm)
Znconcentration(mg/kg)
Cakmak et al., 2010,
J. Agric. Food. Chem.
White arrow: Zn in
entire cross section
Black arrow: Zn in
endosperm section
entire cross section
endosperm section

41. 0
200
400
600
800
1000
1200
1400
1600
1800
2000
0 500 1000 1500 2000 2500 3000 3500 4000
Distance (µm)
Znconcentration(mg/kg)
0
200
400
600
800
1000
1200
1400
1600
1800
2000
0 500 1000 1500 2000 2500 3000 3500
Distance (µm)
Znconcentration(mg/Kg)
0
200
400
600
800
1000
1200
1400
1600
1800
2000
0 500 1000 1500 2000 2500 3000 3500
Distance (µm)
Znconcentration(mg/kg)
0
5
10
15
20
25
0 200 400 600 800 1000 1200 1400 1600
Distance (µm)
Znconcentration(mg/kg)
0
5
10
15
20
25
0 500 1000 1500 2000
Distance (µm)
Znconcentration(mg/kg)
0
5
10
15
20
25
0 200 400 600 800 1000 1200 1400
Distance (µm)
Znconcentration(mg/Kg)
No Foliar Zn Application
Foliar Zn Application at Stem Elongation and Booting Stages
Foliar Zn Application at Milk and Dough Stages
B
cr
cr
cr
Endosperm
Endosperm
Endosperm
LA-ICP-MS Tests
Cakmak et al., 2010, J. Agric. Food. Chem. 58:9092-9102

42. No Zn
Zn applied at stem
elongation and boot
Zn applied at milk and
and dough stages
0
5
10
15
20
25
0 200 400 600 800 1000 1200 1400 1600
Distance (µm)
Znconcentration(mg/kg)
0
5
10
15
20
25
0 500 1000 1500 2000
Distance (µm)
Znconcentration(mg/kg)
0
5
10
15
20
25
0 200 400 600 800 1000 1200 1400
Distance (µm)
Znconcentration(mg/Kg)
Changes in Endosperm Zinc Concentrations
Cakmak et al., 2010,
J. Agric. Food. Chem.

43. 0
20
40
60
80
1. Control (No Zn)
3. Booting + Milk
4. Milk + Dough
2. Stem + Booting
GrainZn,mgkg-1
Turkey
32
57
23
49
Location
Adana
Location
Samsun
0
20
40
60
80
37
64
29
55
High Nitrogen
Low Nitrogen
58
51
Grain Zn Concentrations as Affected from Soil N
Fertilization and Foliar Zn Applications in Turkey
Cakmak et al., 2010
J. Agric Food Chem

44. Changes in shoot and grain
concentrations of Fe and Zn
depending on N supply
Nitrogen Effect

45. 10
15
20
25
30GRAINZn(ppm)
20
30
40
50
60
ZnYIELD(g/ha)
0 40 80 120
N (kg/ha)
Zn YIELD (g/ha)
GRAIN Zn (ppm)
EFFECTS of N FERTILIZATION on GRAIN Zn
CONCENTRATIONS and GRAIN Zn-YIELD
Cakmak et al., 2010b

46. Possible Nitrogen/Protein Effects
on Zn/Fe Concentration of Seeds
Seed Effect
•protein synthesis
•storage proteins
•sink activity
Re-translocation/Phloem Loading
•N-containing chelators
•Transporter proteins
Transport
•N-containing chelators
•Proteins contributing to xylem loading
•…
Mobilization & Uptake
•Transporter proteins mediating uptake
•Root exudation (e.g., phytosiderophores)
•Root growth
•Microbial activity

47. Effect of increasing N supply on root
uptake and root-to-shoot
translocation of Zn in wheat

48. 65Zn uptake rates and root-to-shoot translocation rates
of 22-day-old wheat seedlings precultured with low (0.5
mM), medium (1.0 mM) or high (4.0 mM) N supply.
Erenoglu et al., 2010, New Phytologist
65Zn uptake rates root-to-shoot translocation

49. Grain K Concentration
No Foliar Application
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
0.40
0.45
0.50
Low Adequate High
Soil Zn Supply
Grain[K](%)
Grain K Concentration
Foliar Zn Application
0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
0.40
0.45
0.50
Low Adequate High
Soil Zn Supply
Grain[K](%)
Grain Fe Concentration
Foliar Zn Application
0
10
20
30
40
50
60
70
Low Adequate High
Soil Zn Supply
Grain[Fe](mg.kg
-1
)
Grain Fe Concentration
No Foliar Application
0
10
20
30
40
50
60
70
Low Adequate High
Soil Zn Supply
Grain[Fe](µg.g
-1
)
Grain Zn Concentration
Foliar Zn Application
0
10
20
30
40
50
60
70
80
90
100
110
Low Adequate High
Soil Zn Supply
Grain[Zn](µg.g
-1
)
Grain Zn Concentration
No Foliar Application
0
10
20
30
40
50
60
70
80
90
100
110
Low Adequate High
Soil Zn Supply
Grain[Zn](µg.g
-1
)
Low N
Adequate N
High N
Zinc
Iron
Potassium
Effect of
Increasing
Supply of Zn
and N on Grain
Concentrations
of Zn, Fe and K
Kutman et al., 2010 Cereal Chem

50. 65ZnSO4
65Zn
Effect of N nutrition on transport
and accumulation of 65Zn in grain
after the treatments of the flag
leaves with 65Zn-labelled solution
Growth at 3 N levels in soil
65Zn Flag Leaf
Erenoglu et al., 2010, New Phytol.

51. 0
0.1
0.2
0.3
0.4
0.5
0.6
60 180 540
Grain65Zn/Flagleaf65Zn(%)
N supply, mg kg-1 soil
65Zn-Translocation Efficiency into Grain
Erenoglu et al., 2010, New Phytol. in press

52. Distribution and partitioning of Fe
and Zn among the stems, leaves,
husks and grains in wheat grown with
low and adequate N supply in
greenhouse
Kutman et al. 2010b

53. Shoot Part Low N High N Low N High N
Husks 9 7 10 6
Grains 38 60 59 78
Leaves 48 28 17 8
Stem 5 6 14 7
Fe Zn
Zinc and Iron Partitioning (%) at Maturity
Kutman et al. 2010, in review
Nitrogen Dependent

54. High Protein in Seed:
a Sink for Zn and Fe ?

55. Possible Nitrogen/Protein Effects
on Zn/Fe Concentration of Seeds
Seed Effect
•protein synthesis
•storage proteins
•sink activity
Re-translocation/Phloem Loading
•N-containing chelators
•Transporter proteins
Transport
•N-containing chelators
•Proteins contributing to xylem loading
•…
Mobilization & Uptake
•Transporter proteins mediating uptake
•Root exudation (e.g., phytosiderophores)
•Root growth
•Microbial activity Cakmak et al., 2010 Cereal
Chem, 77: 10-20

56. Cakmak et al., 2010
Cereal Chem, 77: 10-20
Staining of Protein, Zinc and Iron in Wheat Grain
Protein Zinc Iron

57. Cakmak et al., 2010 Cereal
Chem, 77: 10-20
Staining of Protein, Zinc and Iron in Wheat Grain
Protein Zinc Iron
High Protein in Seed:
a Sink for Zn and Fe

58. Int’l Zinc Assoc.
Acknowledgement
Mosaic Co.
K+S Kali
IFA IPNIOmex Agrifluids

59. Sabanci University
Thank you…
Thank You…
Sabanci University

61. Change in Nutrient Composition With Milling Affects
Bioavailability & Warrants Consideration in Breeding
% of Total
in
Unmilled
Grain

62. WHOLE
GRAIN
WHOLE
GRAIN
WHITE
FLOURMILLING
Phytate/Znratio
+Zn
-Zn
No foliar Zn application
After foliar Zn application

63. SEED
ZINC
Increasing
Resistance to
Diseases
Decreasing
Seeding Rate
Better Seed
Viability and
Seedling Vigor
Improving
Abiotic Stress
Tolerance
Improving
Human
Nutrition
Higher Yield
under Zn
Deficiency
Agronomic and human nutritional benefits
resulting from use of Zn-enriched seeds
Cakmak, 2008; Plant and Soil, 302: 1-17