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Rapid, accurate and routine HPLC method for large-scale screening of pro-vitamin A carotenoids in oilseeds

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Abstract

Plant carotenoids, the precursors of vitamin A display several important biological functions as antioxidants and anti-carcinogens. The oilseed crops, owing to their high oil content, form a good matrix for the bioavailability of β-carotene, thereby providing potential targets for biofortification to combat vitamin A deficiency (VAD). However, the screening and characterization of these crops, that otherwise contain very low levels of pro-vitamin A carotenoids has been difficult owing to their poor recovery and strong binding to the oil matrix. Here, we report a rapid method for high volume HPLC analysis involving the extraction and determination of β-carotene in four oilseed crops (peanut, soybean, sunflower and mustard). This included a comprehensive study of the factors that potentially influence the qualitative and quantitative yields of β-carotene in these crops. This is the first crop-independent HPLC method for the quantification of pro-vitamin A carotenoids that shows excellent recovery and reproducibility (>90 percentage recovery in oil) using small tissue sample and is capable of processing up to 30 samples per day. The protocol is sensitive, and enables better detection and separation of individual carotenoids by reducing artefacts during extraction, purification and chromatography that can be used for routine screening of oilseeds.

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Abbreviations

BHT:

Butylated hydroxyltoluene

DE:

Diethyl ether

HPLC:

High-performance liquid chromatography

PE:

Petrolium ether

PDA:

Photodiode array detector

PTFE:

Polytetrafluoroethylene

THF:

Tetrahydrofuran

VAD:

Vitamin A deficiency

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Acknowledgments

This research was supported by grants from HarvestPlus Challenge Program and the Department of Biotechnology, Government of India. We thank the Directorate of Oilseed Research, Hyderabad, India and the Peanut Breeding unit of ICRISAT for providing the seeds for this research. The technical support received from Mohd. Yousuf, D. Pandary and C. Lakshminarayana is duly acknowledged.

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Authors and Affiliations

  1. International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Patancheru, 502 324, Andhra Pradesh, India

    Madhurima Bhatnagar-Panwar, Pooja Bhatnagar-Mathur, Venkata VijayAnand Bhaaskarla, Srinivas Reddy Dumbala & Kiran K. Sharma

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  1. Madhurima Bhatnagar-Panwar
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  2. Pooja Bhatnagar-Mathur
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  3. Venkata VijayAnand Bhaaskarla
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  4. Srinivas Reddy Dumbala
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  5. Kiran K. Sharma
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Correspondence to Kiran K. Sharma.

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Bhatnagar-Panwar, M., Bhatnagar-Mathur, P., Bhaaskarla, V.V. et al. Rapid, accurate and routine HPLC method for large-scale screening of pro-vitamin A carotenoids in oilseeds. J. Plant Biochem. Biotechnol. 24, 84–92 (2015). https://doi.org/10.1007/s13562-013-0239-1

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