Zinc biofortification strategies for wheat grown on calcareous Vertisols in southern Spain: application method and rate



The aims of this work were (i) to find a soil indicator to predict durum wheat yield response to Zn fertilization, (ii) to compare the effect of various Zn fertilization strategies on wheat yield and Zn biofortification in calcareous Vertisols of southern Spain, and (iii) to assess the effect of these Zn fertilization strategies on crop P uptake (durum and bread wheat).


Different Zn fertilization strategies, soil application (0.3–10 kg ha−1) and foliar spraying (two rates, different growth stages), were tested in wheat crops under field conditions in the period 2012–2019.


A simple soil indicator failed to predict durum wheat response to Zn fertilization. Only one of the combinations tested increased wheat yield in the 11 field experiments carried out. Zinc foliar spraying (1.28 kg ha−1) was effective for wheat biofortification when applied at early booting (durum wheat) or flowering, and also when splitting this application between stem elongation and flowering stages (bread wheat). The foliar treatments produced the highest zinc use efficiencies (6–19%) and soil applications the lowest (0.2–1.3%). Moreover, foliar treatments increased grain Zn concentrations by 12–51% while soil application increased such concentrations by only 4–13%. None of the Zn fertilization strategies altered P uptake.


No yield increase in wheat is expected from Zn fertilization for the application methods and rates used here and the soils studied (calcareous Vertisols under Mediterranean climate). However, foliar applications at and after early booting stage are promising for durum and bread wheat biofortification.

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Data availability

The data are available under request.



electrical conductivity


organic carbon by rapid dichromate oxidation

POlsen :

Available soil P


labile Zn in soil, extracted with diethylenetriaminepentaacetic acid


Zn use efficiency


  1. Ahsin M, Hussain S, Rengel Z, Amir M (2020) Zinc status and its requirement by rural adults consuming wheat from control or zinc-treated fields. Environ Geochem Health 42:1877–1892. https://doi.org/10.1007/s10653-019-00463-8

    CAS  Article  PubMed  Google Scholar 

  2. Alloway BJ (2009) Soil factors associated with zinc deficiency in crops and humans. Environ Geochem Health 31:537–548. https://doi.org/10.1007/s10653-009-9255-4

    CAS  Article  PubMed  Google Scholar 

  3. Alloway BJ (2008) Zinc in soils and crop nutrition, 2nd edn. IZA and IFA, Brussels, p 139

    Google Scholar 

  4. Brown KH, Rivera JA, Bhutta Z et al (2004) International zinc nutrition consultative group (IZiNCG) technical document #1. Assessment of the risk of zinc deficiency in populations and options for its control. In: food and nutrition bulletin. United Nations University press, Tokyo

  5. Cakmak I (2008) Enrichment of cereal grains with zinc: agronomic or genetic biofortification? Plant Soil 302:1–17. https://doi.org/10.1007/s11104-007-9466-3

    CAS  Article  Google Scholar 

  6. Cakmak I, Kalayci M, Kaya Y, Torun AA, Aydin N, Wang Y, Arisoy Z, Erdem H, Yazici A, Gokmen O, Ozturk L, Horst WJ (2010) Biofortification and localization of zinc in wheat grain. J Agric Food Chem 58:9092–9102. https://doi.org/10.1021/jf101197h

    CAS  Article  PubMed  Google Scholar 

  7. Cakmak I, Kutman UB (2018) Agronomic biofortification of cereals with zinc: a review. Eur J Soil Sci 69:172–180. https://doi.org/10.1111/ejss.12437

    Article  Google Scholar 

  8. Cakmak I, Yilmaz A, Kalayci M, Ekiz H, Torun B, Erenoglu B, Braun HJ (1996) Zinc deficiency as a critical problem in wheat production in Central Anatolia. Plant Soil 180:165–172. https://doi.org/10.1007/BF00015299

    CAS  Article  Google Scholar 

  9. Catlett KM, Heil DM, Lindsay WL, Ebinger MH (2002) Soil chemical properties controlling Zinc2+ activity in 18 Colorado soils. Soil Sci Soc Am J 66:1182–1189. https://doi.org/10.2136/sssaj2002.1182

    CAS  Article  Google Scholar 

  10. Chattha MU, Hassan MU, Khan I, Chattha MB, Mahmood A, Chattha MU, Nawaz M, Subhani MN, Kharal M, Khan S (2017) Biofortification of wheat cultivars to combat zinc deficiency. Front Plant Sci 8:1–8. https://doi.org/10.3389/fpls.2017.00281

    Article  Google Scholar 

  11. Chen Y, Barak P (1982) Iron nutrition of plants in calcareous soils. Adv Agron 35:217–240. https://doi.org/10.1016/S0065-2113(08)60326-0

    CAS  Article  Google Scholar 

  12. del Campillo MC, Torrent J (1992) Predicting the incidence of Iron Chlorosis in Cal-Careous soils of southern Spain. Commun Soil Sci Plant Anal 23:399–416. https://doi.org/10.1080/00103629209368598

    Article  Google Scholar 

  13. El-Dahshouri MF, El-Fouly MM, Khalifa RKM, El-Ghany HMA (2017) Effect of zinc foliar application at different physiological growth stages on yield and quality of wheat under sandy soil conditions. Agric Eng Int CIGR J Special issue:193–200

  14. Erdal I, Yilmaz A, Taban S, Eker S, Torun B, Cakmak I (2002) Phytic acid and phosphorus concentrations in seeds of wheat cultivars grown with and without zinc fertilization. J Plant Nutr 25:113–127. https://doi.org/10.1081/PLN-100108784

    CAS  Article  Google Scholar 

  15. Gee GW, Bauder JW (1986) Particle-size analysis. In: Klute A (ed) Methods of soil analysis, 2nd edn. American Society of Agronomy and Soil Science Society of America, Madison, Madison, Wisconsin, pp 383–412

    Google Scholar 

  16. Graham RD, Welch RM (1997) Trace element intervention strategies in animal and human nutrition a strategy for breeding staple-food crops with high micronutrient density. 447–450

  17. IUSS Working Group WRB (2014) World reference base for soil resources 2014. International soil classification system for naming soils and creating legends for soil maps.

  18. Joy EJM, Stein AJ, Young SD, Ander EL, Watts MJ, Broadley MR (2015) Zinc-enriched fertilisers as a potential public health intervention in Africa. Plant Soil 389:1–24. https://doi.org/10.1007/s11104-015-2430-8

    CAS  Article  Google Scholar 

  19. Jurowski K, Szewczyk B, Nowak G, Piekoszewski W (2014) Biological consequences of zinc deficiency in the pathomechanisms of selected diseases. J Biol Inorg Chem 19:1069–1079. https://doi.org/10.1007/s00775-014-1139-0

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  20. Lindsay WL, Norvell WA (1978) Development of a DTPA soil test for zinc, iron, manganese and copper. Soil Sci Soc Am J 42:421–428. https://doi.org/10.2136/sssaj1978.03615995004200030009x

    CAS  Article  Google Scholar 

  21. Liu DY, Liu YM, Zhang W, Chen XP, Zou CQ (2019) Zinc uptake, translocation, and remobilization in winter wheat as affected by soil application of Zn fertilizer. Front Plant Sci 10:1–10. https://doi.org/10.3389/fpls.2019.00426

    Article  Google Scholar 

  22. Liu DY, Zhang W, Yan P, Chen XP, Zhang FS, Zou CQ (2017) Soil application of zinc fertilizer could achieve high yield and high grain zinc concentration in maize. Plant Soil 411:47–55. https://doi.org/10.1007/s11104-016-3105-9

    CAS  Article  Google Scholar 

  23. Loneragan JF, Webb MJ (1993) Interactions between zinc and other nutrients affecting the growth of plants. In: Zinc in soils and plants. Developments in plant and soil sciences, vol 55. Springer, Dordrecht. https://doi.org/10.1007/978-94-011-0878-2_9

    Google Scholar 

  24. Mabesa RL, Impa SM, Grewal D, Johnson-Beebout SE (2013) Contrasting grain-Zn response of biofortification rice (Oryza sativa L.) breeding lines to foliar Zn application. F Crop Res 149:223–233. https://doi.org/10.1016/j.fcr.2013.05.012

    Article  Google Scholar 

  25. McDonald GK, Genc Y, Graham RD (2008) A simple method to evaluate genetic variation in grain zinc concentration by correcting for differences in grain yield. Plant Soil 306:49–55. https://doi.org/10.1007/s11104-008-9555-y

    CAS  Article  Google Scholar 

  26. Murphy J, Riley JP (1962) A modified single solution method for the determination of phosphate in natural waters. Anal Chim Acta 27:31–36. https://doi.org/10.1016/S0003-2670(00)88444-5

    CAS  Article  Google Scholar 

  27. Nriagu J (2019) Zinc deficiency in human health. In: Encyclopedia of environmental health 2th edition. Elsevier, pp 489–499. https://doi.org/10.1016/B978-0-444-52272-6.00674-7

  28. Olsen SR, Cole CV, Watanabe FS, Dean LA (1954) Estimation of available phosphorus in soils by extraction with sodium bicarbonate. USDA Circ 18:45. https://doi.org/10.2307/302397

    Article  Google Scholar 

  29. Pfeiffer WH, McClafferty B (2007) HarvestPlus: breeding crops for better nutrition. Crop Sci 47:S-88–S-105. https://doi.org/10.2135/cropsci2007.09.0020IPBS

    Article  Google Scholar 

  30. Prasad AS (2013) Discovery of human zinc deficiency: its impact on human health and disease. Adv Nutr 4:176–190. https://doi.org/10.3945/an.112.003210

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  31. Ram H, Rashid A, Zhang W, Duarte AP, Phattarakul N, Simunji S, Kalayci M, Freitas R, Rerkasem B, Bal RS, Mahmood K, Savasli E, Lungu O, Wang ZH, de Barros VLNP, Malik SS, Arisoy RZ, Guo JX, Sohu VS, Zou CQ, Cakmak I (2016) Biofortification of wheat, rice and common bean by applying foliar zinc fertilizer along with pesticides in seven countries. Plant Soil 403:389–401. https://doi.org/10.1007/s11104-016-2815-3

    CAS  Article  Google Scholar 

  32. Ryan J, Ibrikci H, Delgado A et al (2012) Significance of phosphorus for agriculture and the environment in the West Asia and North Africa region. Adv Agron 114:91–143. https://doi.org/10.1016/B978-0-12-394275-3.00004-3

    CAS  Article  Google Scholar 

  33. Sacristán D, González-Guzmán A, Barrón V et al (2019) Phosphorus-induced zinc deficiency in wheat pot-grown on noncalcareous and calcareous soils of different properties. Arch Agron Soil Sci 65:208–223. https://doi.org/10.1080/03650340.2018.1492714

    CAS  Article  Google Scholar 

  34. Sánchez-Rodríguez AR, del Campillo MC, Torrent J (2017) Phosphorus reduces the zinc concentration in cereals pot-grown on calcareous Vertisols from southern Spain. J Sci Food Agric 97:3427–3432. https://doi.org/10.1002/jsfa.8195

    CAS  Article  PubMed  Google Scholar 

  35. van Wesemael JC (1955) De bepaling van het calciumcarbonaatgehalte van gronden. Chem Weekbl 51:35–36

    CAS  Google Scholar 

  36. Walkley A, Black IA (1934) An examination of the degtjareff method for determining soil organic matter, and a proposed modification of the chromic acid titration method. Soil Sci 37:29–38. https://doi.org/10.1097/00010694-193401000-00003

    CAS  Article  Google Scholar 

  37. Welch RM, Graham RD, Cakmak I (2013) Linking agricultural production practices to improving human nutrition and health. In: expert paper written for ICN2 second international conference on nutrition preparatory technical meeting. Rome, Italy, pp 13–15

    Google Scholar 

  38. Xue Y, Xia H, Christie P, Zhang Z, Li L, Tang C (2016) Crop acquisition of phosphorus, iron and zinc from soil in cereal/legume intercropping systems: a critical review. Ann Bot 117:363–377. https://doi.org/10.1093/aob/mcv182

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  39. Zadoks JC, Chang TT, Konzak CF (1974) A decimal code for the growth stages of cereals. Weed Res 14:415–421. https://doi.org/10.1111/j.1365-3180.1974.tb01084.x

    Article  Google Scholar 

  40. Zasoski RJ, Burau RG (1977) A rapid nitric-perchloric acid digestion method for multi-element tissue analysis. Commun Soil Sci Plant Anal 8:425–436. https://doi.org/10.1080/00103627709366735

    CAS  Article  Google Scholar 

  41. Zhang W, Liu D, Li C, Cui Z, Chen X, Russell Y, Zou C (2015) Zinc accumulation and remobilization in winter wheat as affected by phosphorus application. F Crop Res 184:155–161. https://doi.org/10.1016/j.fcr.2015.10.002

    Article  Google Scholar 

  42. Zia MH, Ahmed I, Bailey EH, Lark RM, Young SD, Lowe NM, Joy EJM, Wilson L, Zaman M, Broadley MR (2020) Site-Specific Factors Influence the Field Performance of a Zn-Biofortified Wheat Variety Front Sustain Food Syst:4. https://doi.org/10.3389/fsufs.2020.00135

  43. Zou C, Du Y, Rashid A et al (2019) Simultaneous biofortification of wheat with zinc, iodine, selenium, and iron through foliar treatment of a micronutrient cocktail in six countries. J Agric Food Chem 67:8096–8106. https://doi.org/10.1021/acs.jafc.9b01829

    CAS  Article  PubMed  Google Scholar 

  44. Zou CQ, Zhang YQ, Rashid A, Ram H, Savasli E, Arisoy RZ, Ortiz-Monasterio I, Simunji S, Wang ZH, Sohu V, Hassan M, Kaya Y, onder O, Lungu O, Mujahid MY, Joshi AK, Zelenskiy Y, Zhang FS, Cakmak I (2012) Biofortification of wheat with zinc through zinc fertilization in seven countries. Plant Soil 361:119–130. https://doi.org/10.1007/s11104-012-1369-2

    CAS  Article  Google Scholar 

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The authors appreciate the help and knowledge provided by farmers on the studied fields (especially Juan Carlos Moreno and Paco Moreno) and technicians of the Soil Science Unit of the University of Córdoba (Juan Manuel Delgado and Mercedes Castro) and Agrupación Cordobesa de Agricultores (S.A.T.) (particularly Rafael Carranza and Miguel Sánchez).

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This work was co-funded by the Spanish Ministry of Economy and Competitiveness and the European Regional Development Fund [Project AGL2014–57835-C2–2-R], and also by a scholarship awarded to A. González-Guzmán (BES-2015-073507). A. R. Sánchez-Rodríguez acknowledges additional funding by the Spanish Ministry of Science, Innovation and Universities under the “Juan de la Cierva–Incorporación” programme [Grant Number, IJCI-2016-27,388].

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Correspondence to Antonio Rafael Sánchez-Rodríguez.

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Sánchez-Rodríguez, A.R., Marín-Paredes, M., González-Guzmán, A. et al. Zinc biofortification strategies for wheat grown on calcareous Vertisols in southern Spain: application method and rate. Plant Soil (2021). https://doi.org/10.1007/s11104-021-04863-7

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  • P and Zn interaction
  • Zn uptake
  • Foliar spraying
  • Soil application
  • Biofortification
  • Zinc use efficiency