Variability of microelements and antioxidants in maize hybrids and their parental inbred lines

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1 ORIGINAL SCIENTIFIC PAPER Variability of microelements and antioxidants in maize hybrids and their parental inbred lines Snežana MLADENOVIĆ DRINIĆ, Vesna DRAGIČEVIĆ, Milomir FILIPOVIĆ, Dragan KOVAČEVIĆ Maize Research Institute Zemun Polje, Belgrade, Slobodana Bajića 1, Zemun Polje, Serbia, ( Abstract Adequate macro- and microelements concentration in seed is very important for maize productivity and nutritive value of product. The aim of this study was comparison of five microelements, β carotene, and tocopherol content of 8 maize hybrids and their parental lines. The concentration of Mg varied from mg kg -1 to mg kg -1 ; Fe from mg kg -1 to mg kg -1, Zn from mg kg -1 to mg kg -1, Ca from to mg kg -1, and Mn from 1.78 mg kg -1 to 2.90 mg kg -1. Hybrids have higher content of Ca and Fe and lower Mg, Mn, and Zn than inbred lines. Hybrid ZP684 has the highest value of Ca content, high Fe content and the lowest Mg content. Out of eight hybrids four have lower content of Mg, Ca, Fe, Mn than inbred lines. Average content of α tocopherol was higher in parental lines ( ) than hybrids ( ) but hybrids have higher content of γ tocopherol ( ) than parental lines ( ). Hybrids have average content of β carotene mg kg -1 and parental lines 14.6 mg kg -1. Hybrid ZP427 has the highest β carotene content (21.90 mg kg -1 ) and hybrid ZP684 the lowest one (11.36 mg kg -1 ). Increased levels of microelements and carotenoids and tocopherols in maize grain should increase the nutritive value of maize. Key words: microelements, tocopherols, β carotene, maize Introduction Maize is one of the most important crops due to its high productivity and its multiple uses as a food source for humans, livestock feed and as raw material in various industries. Among metal elements Fe, Mn, Mg, Ca, Mo, Ni and Zn are considered plant micronutrients because they are essential for physiological processes in trace amounts. Deficiency of micronutrients in plants is a global nutritional problem as the major food staples are highly susceptible to such deficits. The development of an efficient breeding program to increase minerals concentration in maize depends on the presence of genetic variability in this species (Menkir, 2008). The microelements content in grain is a complex trait affected by a number of factors, including genotype, soil properties, environmental conditions and nutrient interactions (House, 1999). As a consequence, results reported in literature about micronutrient concentrations in maize grain varied substantially. Significant genetic variation in grain microelements has been reported in maize (Bänziger and Long, 2000, Brkić et al., 2003; Brkić et al., 2004; Šimić et al., 2004, and Menkir, 2008). Maize as a major crop has been investigated for decades for metal accumulation, particularly iron (Fe) and zinc (Zn) accumulation in grain or biofortification purposes (Ortiz-Monasterio et al., 2007). Carotenoids and tocopherols are lipid soluble antioxidants associated with decreased risk of several degeneative diseases. Both vitamins occur naturally in maize grain. Carotenoids are located in endosperm and tocopherols, a component of oil, in the germ (Weber, 1987). Several studies have shown significant differences among maize inbreds for carotenoid and tocopherol levels (Egesel, 2001). Enhanced grain micronutrient content for both vitamins and minerals is rapidly emerging as the next suite of seed quality traits to be improved by breeding and biotechnology. The aim of this study was comparison of five microelements, β carotene, and tocopherol content of 8 maize hybrids and their parental lines. Proceedings. 50 th Croatian and 10 th International Symposium on Agriculture. Opatija. Croatia ( ) Section 3. Genetics, Plant Breeding and Seed Production 247

2 Snežana MLADENOVIĆ DRINIĆ, Vesna DRAGIČEVIĆ, Milomir FILIPOVIĆ, Dragan KOVAČEVIĆ Material and methods А set of 8 hybrids and their parental inbred lines was used. Hybrids ZP341 (L1 L5) belonged to FAO maturity group 300; ZP 427 (L9 L2) belonged to FAO maturity group 400, ZP555 (L3 L5) and ZP560 (L7 L6) belonged to FAO maturity group 500, ZP600 (L7 L8); ZP606 (L3 L8), ZP666 (L3 L6) and ZP684 (L8 L4) belonged to FAO maturity group 600. An experiment set up as randomized block design with three replications at experimental field of MRI. The content of Mg, Fe, Ca, Mn and Zn was determined by Inductively Coupled Plasma - Optical Emission Spectrometry in the laboratory of Scientific Institute Vinča, Belgrade. The content of tocopherols was determined by the HPLC method. A HPLC system with the Waters M600 E pump, thermostat and Rheodyne 7125 injector was used. The separation of tocopherols was performed on the Nucleosil 50-5 C18 column (250 4 mm, i.d., 5 µm) at flow rate of 1.0 ml min -1. The mobile phase consisted of 95% methanol. The detection was performed with the Shimadzu RF-535 fluorescence detector at an excitation wavelength of 295 nm and an emission wavelength of 330 nm. Identified peaks were confirmed and quantified by data acquisition and spectral evaluation using the Clarify chromatographic software. The content of tocopherols is expressed as μg per 100g of d.m. Β-carotene was determined according to AACC (1995) procedure after extraction with saturated butanol. Results and discussion Microelements content made significant contribution towards the genetic divergence of the genotypes, indicating the presence of considerable variability for these traits in the inbred lines and hybrids. A wide range of genetic variation was found among genotypes for all the grain microelements (Table 1): Mg ( mg kg mg kg -1 ), Fe (10.21 mg kg mg kg -1 ), Zn (17.46 mg kg mg kg -1 ), Ca ( mg kg -1 ), and Mn (1.78 mg kg mg kg -1 ). The content of Mg varied from mg kg -1 (ZP606) to mg kg -1 (ZP427) in hybrids, average mg kg -1 and from mg kg -1 ( ZPL7) to mg kg -1 (ZPL1) in inbred lines average mg kg -1, Table 1. Four hybrids ZP341, ZP555, ZP606, and ZP684 have lower content of Mg than their parental inbred lines. The lowest content of Ca have hybrid ZP341 (56.88 mg kg -1 ) and the highest hybrid ZP684 ( mgkg -1 ). Out of eight hybrids four have lower content of Ca (ZP427, ZP560, ZP666, ZP684) than parental lines. The content of Fe varied from mg kg -1 to mgkg -1 in inbred lines, average 16,79 mg kg -1 and from mgkg -1 to mg kg -1 for hybrids, average 18,83 mg kg -1. Hybrid ZP341 have the highest content of Mn but the lowest of Zn, and hybrid ZP666 have the lowest content of Mn but the highest content of Zn. Hybrid ZP684 has the highest value of Ca content, high Fe content and the lowest Mg content and hybrid ZP427 the highest Fe, high Zn and Mn content. Menkir (2008) found similar variation in Zn and Fe levels in 278 maize inbred lines evaluated in five environments and showed that there were highly significant effects of maize genotypes in mineral contents, but location effect was not significant on the concentration of any kernel minerals. Simple correlation analysis was computed to assess the association between mineral elements. Negative correlation was obtained between content of Mn and Mg with Ca (r=-0,44, r=-0,02), respectively. The highest positive correlation was between Mg and Zn (r=0,58). Fe had positive significant correlation with Zn (r=0,57) and Ca (r=0,54) and not significant with Mg (r= 0,28) and Mn (r=0,22). Zn had positive and significant correlations with Mg, Fe, Ca, but its association with Mn was not significant (r= 0,07). This is in agreement of results of Menkir (2008), who obtained significant correlation between Fe and Zn as well with Mg and P. Simic et al (2004) obtained generally positive correlation between Fe, Mn, Mg and Zn th Croatian and 10 th International Symposium on Agriculture

3 Variability of microelements and antioxidants in maize hybrids and their parental inbred lines Table 1. Contents of microelements in maize hybrids and parental line (mg kg -1 ) Genotypes Mg Ca Fe Mn Zn ZP ZP ZP ZP ZP ZP ZP ZP ZPL ZPL ZPL ZPL ZPL ZPL ZPL ZPL ZPL Average content of α tocopherol was higher in parental lines ( µg kg -1 ) than hybrids ( µg kg -1 ) but hybrids have higher content of γ tocopherol ( µg kg -1 ) than parental lines ( µg kg -1 ) (Table 2). Content of α tocopherol varied from to µg kg -1 for hybrids and from µg kg -1 to µg kg -1 in parental lines. Two of eight hybrid and all inbred lines except ZPL5 has content of δ tocopherol <20 µg kg -1. The highest α tocopherol content have hybrid ZP666 and inbred line ZPL6 ( µg kg -1 ) and the lowest ZPL 1 ( µg kg -1 ). γ tocopherol content range from µg kg -1 to µg kg -1 for hybrids and from µg kg -1 to µg kg -1 for parental lines. Inbred line with the highest α tocopherol content has the lowest γ tocopherol content. Hybrid ZP684 has the lowest α tocopherol content but the highest γ and δ tocopherol content. Hybrids ZP606 and ZP666 with ZPL3 as common parent have high α tocopherol content and hybrids ZP600, ZP606 and ZP684 with ZPL8 as common parent have high γ tocopherol content. Ratio of α- and γ-tocopherols ranges from 0.38 to 1.04 for hybrids and from 0.4 to 5.5 for parental lines (Table 2). Hybrids ZP427, ZP606 and ZP666 as well as inbred lines ZPL3, ZPL4, ZPL5, ZPL6, ZPL7, ZPL8 and ZPL9 have higher content of α tocopherol than γ tocopherol. Hybrids have average content of β carotene mg kg -1 and parental lines 14.6 mg kg -1. Hybrid ZP427 has the highest β carotene content (21.90 mgkg -1 ) and hybrid ZP684 the lowest one (11.36 mgkg -1 ). Inbred line ZPL 4 has the highest β carotene content (18.61 mgkg -1 ) and ZPL7 the lowest one (8.69 mgkg -1 ). In breeding program for selection of high carotenes maze lines, Safawo et al.(2010) have also determined high variation in β-carotene in maize grains. Section 3. Genetics, Plant Breeding and Seed Production 249

4 Snežana MLADENOVIĆ DRINIĆ, Vesna DRAGIČEVIĆ, Milomir FILIPOVIĆ, Dragan KOVAČEVIĆ Genotypes Table 2. Tocopherol and beta carotene content α tocopherol γ tocopherol δ tocopherol β carotene mg/100g α / γ tocopherols ratio ZP ZP ZP ZP < ZP ZP ZP < ZP ZPL < ZPL < ZPL < ZPL < ZPL ZPL < ZPL < ZPL < ZPL < Conclusions Considerable variability for microelements and antioxidans in grain of eight hybrids and their parental lines was determined. Hybrids have higher content of Ca and Fe and lower Mg, Mn, and Zn than inbred lines. Significant correlation between content of Mg and Zn and Fe and Zn was observed. A high amount of variation for β-carotine and tocopherol content is present among genotypes as well as for different ratio of α- and γ-tocopherols. Average content of α tocopherol was higher in parental lines than hybrids but hybrids have higher content of γ tocopherol than parental lines. Increased levels of microelemnts as well as carotenoids and tocopherols because of their antioxidant activity, should increase the nutritive value of maize. Acknowledgement This study is supported by the Ministry of Education, Science and technological development of the Republic of Serbia. It is part of a scientific project Improving the quality of maize and soybean by conventional and molecular breeding (Reg. No. TR 31068), as well as COST Action FA 0905 Mineral Improved Crop Production for Healthy Food and Feed. References American Association of Cereal Chemists Method (1995). Approved Methods of the AACC, The association: St. Paul, Minnesota, USA, AACC Method, pp Bänziger, M., Long J. (2000). The potential for increasing the iron and zinc density of maize through plant breeding. Food and Nutrition Bulletin 20: Brkić, I., Šimić, D., Zdunić, Z., Jambrović, A., Ledenčan, T., Kovačević, V., Kadar, I. (2003). Combining abilities of cornbelt inbred lines of maize for mineral content in grain, Maydica, 48, th Croatian and 10 th International Symposium on Agriculture

5 Variability of microelements and antioxidants in maize hybrids and their parental inbred lines Brkić, I., Šimić, D., Zdunić, Z., Jambrović, A. Ledenčan, T. Kovačević, V., Kadar, I. (2004). Genotypic variability of micronutrient element concentrations in maize kernels, Cereal Research Communications 32(1), Egesel, C.O. (2001). Genetic variation among maize genotypes for carotenoid and tocopherol compounds. Ph.D. Thesis (62-08, secb; P3423). University of Illinois at Urbana-Champaign. House, W.A. (1999). Trace element bioavailability as exemplified by iron and zinc. Field Crop Research, 60, Menkir, A. (2008). Genetic variation for grain mineral content in tropical-adapted maize inbred lines. Food Chemistry, v. 110, p Ortiz-Monasterio, I.J., Palacios-Rojas N., Meng E., Pixley K., Trethowan R., Pena R.J.(2007). Enhancing the mineral and vitamin content of wheat and maize through plant breeding. J. Cereal Sci., 46: Šimić, D., Zdunić, Z., Brkić, I., Kadar, I. (2004). Inheritance of mineral concentrations in kernels of elite maize inbred lines. In: Genetic variation for plant breeding. Johann, V. et al. (eds.) 17th EUCARPIA General Congresss, Wienna, Austria BOKU University of Natural Resources and Applied Sciences, 48 Simic, D., Zdunic Z., Jambrovic A., Ledencan T., Brkic I., Duvnjak V. (2009). Relations among six micronutrients in grain determined in a maize population. Poljoprivreda (Agric.) 15: Safawo T., Senthil N., Raveendran M., Vellaikumar S.E., Ganesan K.N., Nallathambi G., Sarnya S., Shobhana V.G., Abirami B., Gowri V. (2010). Exploitation of natural variability in maize for β-carotene content using HPLC and gene specific markers. Electronic Journal of Plant Breeding, 1, 4: Weber, E. J. (1987). Carotenoids and Tocols of Corn Grain Determined by Hplc. Journal of the American Oil Chemists Society 64, sa2015_p0311 Section 3. Genetics, Plant Breeding and Seed Production 251