Evaluation carotenoids content in South African potato, tomato and wheat cultivars Charlotte Mashaba Eugenia Barros Introduction • Malnutrition - 50% of deaths in children • Food.
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Evaluation carotenoids content in South African potato, tomato and wheat cultivars Charlotte Mashaba Eugenia Barros Introduction • Malnutrition - 50% of deaths in children < 5 • Food insecurities - Community-based intervention projects • Screening for high micronutrient crops © CSIR 2007 www.csir.co.za Carotenoids • Lipid soluble molecules – produced by plants and • photosynthetic organism Disease prevention and reduction Carotenoids Disease/condition References β-carotene, βcryptoxanthin, lycopene and lutein Reduce risk of Breast cancer developing by up to 50% Dorgan et al., 1998; Toniolo et al., 2001 Lycopene Reduced risk of coronary heart disease Sesso et al., 2004 Lutein and zeaxanthin Protective role against AMD Gale et al., 2003 α-carotene, βcarotene, βcryptoxanthin Pro-vitamin A surface linings, bone growth, immune system Semba, 1998 During and Harrison, 2007 Unlu et al., 2005 © CSIR 2007 www.csir.co.za Choice of Crops • Why them? → Wheat → potato → Tomato © CSIR 2007 www.csir.co.za Objectives • Optimize method • Quick protocol © CSIR 2007 www.csir.co.za Platforms used for analysis Crude extraction • Quantify known carotenoid HPLC © CSIR 2007 www.csir.co.za Results - Standard curves Fig 1 Standard curves to determine carotenoids content © CSIR 2007 www.csir.co.za Potato Crop Cultivar (n=3) Lutein Zeaxanthin Canthaxanthin β-Carotene Lycopene Potato Mondial 0.265 ± 0.008 0.441 ± 0.01 ND 0.0201 ± 0.004 ND BP1 0.333 ± 0.003 0.120 ± 0.003 ND 0.0142 ± 0.005 ND Salad 10.801 ± 0.028 0.303 ± 0.008 0.065 ± 0.006 28.940 ± 0.55 387.000 ± 5 Roma 10.867 ± 0.056 0.212 ± 0.001 0.080± 0.01 25.197 ± 1.37 443.090 ± 8.908 Mathabas 0.544 ± 0.017 0.138 ±0.004 ND 0.034± 0.003 ND Elands 0.268 ± 0.017 0.154±0.011 ND ND ND Moni 0.302±0.0339 0.082±0.006 ND ND ND Orania 0.307±0.008 0.036±0.001 ND ND ND Tomato Wheat © CSIR 2007 www.csir.co.za Tomato Crop Cultivar (n=3) Lutein Zeaxanthin Canthaxanthin β-Carotene Lycopene Potato Mondial 0.265 ± 0.008 0.441 ± 0.01 ND 0.0201 ± 0.004 ND BP1 0.333 ± 0.003 0.120 ± 0.003 ND 0.0142 ± 0.005 ND Salad 10.801 ± 0.028 0.303 ± 0.008 0.065 ± 0.006 28.940 ± 0.55 387.000 ± 5 Roma 10.867 ± 0.056 0.212 ± 0.001 0.080± 0.01 25.197 ± 1.37 443.090 ± 8.908 Mathabas 0.544 ± 0.017 0.138 ±0.004 ND 0.034± 0.003 ND Elands 0.268 ± 0.017 0.154±0.011 ND ND ND Moni 0.302±0.0339 0.082±0.006 ND ND ND Orania 0.307±0.008 0.036±0.001 ND ND ND Tomato Wheat © CSIR 2007 www.csir.co.za Wheat Crop Cultivar (n=3) Lutein Zeaxanthin Canthaxanthin β-Carotene Lycopene Potato Mondial 0.265 ± 0.008 0.441 ± 0.01 ND 0.0201 ± 0.004 ND BP1 0.333 ± 0.003 0.120 ± 0.003 ND 0.0142 ± 0.005 ND Salad 10.801 ± 0.028 0.303 ± 0.008 0.065 ± 0.006 28.940 ± 0.55 387.000 ± 5 Roma 10.867 ± 0.056 0.212 ± 0.001 0.080± 0.01 25.197 ± 1.37 443.090 ± 8.908 Mathabas 0.544 ± 0.017 0.138 ±0.004 ND 0.034± 0.003 ND Elands 0.268 ± 0.017 0.154±0.011 ND ND ND Moni 0.302±0.0339 0.082±0.006 ND ND ND Orania 0.307±0.008 0.036±0.001 ND ND ND Tomato Wheat © CSIR 2007 www.csir.co.za Tomato Chromatograms DAD1 A, Sig=450,10 Ref=400,10 (CAR0412\CARO0027.D) Norm. 30 Zeaxanthin A Mondial Lutein 40 20 10 Beta-carotene DAD1 A, Sig=450,10 Ref=400,10 5 10 20 25 30 35 min Sig=450,10 Ref=400,10 (CAR1812\CARO0018.D) 1000 17.5 B A 800 BP 1 10 600 Lutein 15 12.5 400 7.5 Zeaxanthin 5 10 15 20 25 30 35 0 DAD1 A, Sig=450,10 5 Norm. Ref=400,10 (CAR1812\CARO0028.D) 10 15 10 15 20 25 30 20 25 30 min Rt -Retention time (minutes) min 200 B 150 (mAU) 35 Lycopene 5 Canthaxanthin Absorb ance 0 Zeaxanthin Beta-carotene 200 2.5 100 Lutein Beta-carotene 50 DAD1 A, Sig=450,10 Ref=400,10 (CAR0412\CARO0029.D) 0 Norm. Lutein 40 5 A tr -Retention time (minutes) Bread wheat 30 Roma 10 ACranthaxanthin ea :1 2. 86 1 0 DAD1 A, Sig=450,10 Ref=400,10 (CAR0412\CARO0031.D) Norm. 80 10 15 20 25 30 35 min Lutein B Durum 60 40 wheat 20 Canthaxanthin (mAU) 5 100 Beta-carotene Absorba nce Zeaxanthin 20 Zeaxanthin (mAU) A, Norm. (CAR0412\CARO0028.D) 15 Lycopene DAD1 Beta-carotene 0 Norm. Lutein Absorb ance 0 5 10 15 20 25 30 35 tr -Retention time (minutes) © CSIR 2007 www.csir.co.za min 35 min Platforms used for analysis Crude extraction • Identify unknown MALDIget pic © CSIR 2007 www.csir.co.za Identification of ‘unknown’ peaks Potato Theoretical mass 574.4750 596.4590 598.4750 Fomula C40H62O2 C42H60O2 C42H62O2 600.4906 716.3359 780.5693 C42H64O2 C40H53NaO8S C52H76O5 Tomato Theoretical mass 552.4331 582.4073 596.4593 598.4022 Matched carotenoid Chloroxanthin Sprilloxanthin Dihydrosprilloxanthin Tetrahydrospirilloxanthin Ketonostoxanthin 3-Sulfate Siphonaxanthin Formula C40H56O C40H54O3 C42H60O2 C40H54O4 Wheat Theoretical mass 574.4750 582.4073 598.4022 © CSIR 2007 Fomula C40H62O2 C40H54O3 C40H54O4 www.csir.co.za Matched carotenoid Chloroxanthin 4-Ketozeaxanthin; Oxosaproxanthin Halocynthiaxanthin; Matched carotenoid Cryptoxanthin Adonixanthin; Sprilloxanthin Ertythroxanthin Conclusion • Methods used robust → More carotenoids were separated →Sensitive to quantify low amounts → Can be used for more crops →Develop SOP © CSIR 2007 www.csir.co.za Thank you! Acknowledge • Judy Reddy • Dr Stoyan Stoychec AIM Theoratical mass 534.4226 536.4382 536.4382 540.4695 542.4851 550.4175 550.4175 552.4331 568.428 582.4073 596.4593 598.4022 Formulae C40H54 C40H56 C40H56 C40H60 C40H62 C40H54O C40H54O C40H56O C40H56O2 C40H54O3 C42H60O2 C40H54O4 Matched carotenoid Didehydrolycopene; alhpa/beta -Carotene; Lycopene alhpa/beta/ -Carotene; Lycopene Tetrahydro-beta-Carotene; Tetrahydrolycopene Phytofulene tetradehydolycopene 4-Keto-gamma-carotene; tetradehydolycopene -cryptoxanthin Zeaxanthin; Lutein; Adonixanthin; Sprilloxanthin Ertythroxanthin © CSIR 2007 www.csir.co.za Current & future work • Analyze more cultivars and increase nutrients © CSIR 2007 www.csir.co.za Comparison of Carotenoids content in Mondial and BP1 potato cultivarss Potato DAD1 A, Sig=450,10 Ref=400,10 (CAR0412\CARO0027.D) Norm. A Mondial Zeaxanthin 40 Lutein 30 20 0 DAD1 A, Norm. 17.5 15 5 B 10 15 20 25 30 35 min BP 1 12.5 10 7.5 Zeaxanthin 5 Beta-carotene (mAU) Sig=450,10 Ref=400,10 (CAR0412\CARO0028.D) Lutein Absorbance Beta-carotene 10 2.5 0 5 10 15 © CSIR 2007 20 25 www.csir.co.za Rt -Retention time (minutes) 30 35 min