18. Make sure that the absorbance values (AU) for your UV-VIS
measurements are in a linear absorption bandwidth for accurate
quantification (between 0.2 and 0.8). Higher or lower concentrations are not reliable for absorbance measurements. If necessary, stock solutions can be diluted with the same organic
solvent before taking the measurements. Multiple measurements for the same sample may ensure the accuracy of your
calculations. If the A
1%
1cm values for cis-isomers are not available, values belong to all-trans carotenoids can be used as an
alternative if necessary, although not recommended.
19. Calculate the slope where X-axis is defined as μg carotenoid/
20–50 μL volume injection and where Y-axis is peak area
(mAU).
Acknowledgments
This work is supported by Australian Research Council Discovery
Grant DP130102593 (to C.I.C). We acknowledge the Western Sydney University and Hawkesbury Institute for the Environment
for providing a Ph.D. scholarship award to Y.A. and N.D. and financial
support to purchase chemicals required for this research.
Author contributions: Y.A. wrote the methods chapter, prepared
figures and tables. Y.A., N.D. and C.M. optimised methods for
carotenoid extraction and HPLC separation. C.I.C outlined and
edited the methods chapter. C.I.C supervised Y.A. and N.D. All
authors contributed to editing this chapter.
References
1. Baranski R, Cazzonelli CI (2016) Carotenoid
biosynthesis and regulation in plants. In: Carotenoids: nutrition, analysis and technology.
Wiley Blackwell, London, pp 161–190
2. Cuttriss AJ, Chubb AC, Alawady A, Grimm B,
Pogson BJ (2007) Regulation of lutein biosynthesis and prolamellar body formation in Arabidopsis. Funct Plant Biol 34(8):663–672
3. Van Norman JM, Zhang J, Cazzonelli CI, Pogson BJ, Harrison PJ, Bugg TD, Chan KX,
Thompson AJ, Benfey PN (2014) Periodic
root branching in Arabidopsis requires synthesis of an uncharacterized carotenoid derivative.
Proc Natl Acad Sci 111(13):E1300–E1309
4. Dickinson AJ, Lehner K, Mi J, Jia K-P,
Mijar M, Dinneny J, Al-Babili S, Benfey PN
(2019) β-Cyclocitral is a conserved root
growth regulator. Proc Natl Acad Sci 116
(21):10563–10567
5. Al-Babili S, Bouwmeester HJ (2015) Strigolactones, a novel carotenoid-derived plant hormone. Annu Rev Plant Biol 66:161–186
6. Cazzonelli CI, Hou X, Alagoz Y, Rivers J,
Dhami N, Lee J, Marri S, Pogson BJ (2019)
A cisÀcarotene derived apocarotenoid regulates etioplast and chloroplast development.
bioRxiv 528331. https://doi.org/10.1101/
528331
7. Wang JY, Haider I, Jamil M, Fiorilli V, Saito Y,
Mi J, Baz L, Kountche BA, Jia K-P, Guo X
(2019) The apocarotenoid metabolite zaxinone regulates growth and strigolactone biosynthesis in rice. Nat Commun 10(1):810
8. Iskandar AR, Miao B, Li X, Hu K-Q, Liu C,
Wang X-D (2016) β-Cryptoxanthin reduced
lung tumor multiplicity and inhibited lung cancer cells motility by down-regulating nicotinic
acetylcholine receptor α7 expression. Cancer
Prev Res 9(11):875–886
Profiling Carotenoids in Plants Using HPLC
161
measurements are in a linear absorption bandwidth for accurate
quantification (between 0.2 and 0.8). Higher or lower concentrations are not reliable for absorbance measurements. If necessary, stock solutions can be diluted with the same organic
solvent before taking the measurements. Multiple measurements for the same sample may ensure the accuracy of your
calculations. If the A
1%
1cm values for cis-isomers are not available, values belong to all-trans carotenoids can be used as an
alternative if necessary, although not recommended.
19. Calculate the slope where X-axis is defined as μg carotenoid/
20–50 μL volume injection and where Y-axis is peak area
(mAU).
Acknowledgments
This work is supported by Australian Research Council Discovery
Grant DP130102593 (to C.I.C). We acknowledge the Western Sydney University and Hawkesbury Institute for the Environment
for providing a Ph.D. scholarship award to Y.A. and N.D. and financial
support to purchase chemicals required for this research.
Author contributions: Y.A. wrote the methods chapter, prepared
figures and tables. Y.A., N.D. and C.M. optimised methods for
carotenoid extraction and HPLC separation. C.I.C outlined and
edited the methods chapter. C.I.C supervised Y.A. and N.D. All
authors contributed to editing this chapter.
References
1. Baranski R, Cazzonelli CI (2016) Carotenoid
biosynthesis and regulation in plants. In: Carotenoids: nutrition, analysis and technology.
Wiley Blackwell, London, pp 161–190
2. Cuttriss AJ, Chubb AC, Alawady A, Grimm B,
Pogson BJ (2007) Regulation of lutein biosynthesis and prolamellar body formation in Arabidopsis. Funct Plant Biol 34(8):663–672
3. Van Norman JM, Zhang J, Cazzonelli CI, Pogson BJ, Harrison PJ, Bugg TD, Chan KX,
Thompson AJ, Benfey PN (2014) Periodic
root branching in Arabidopsis requires synthesis of an uncharacterized carotenoid derivative.
Proc Natl Acad Sci 111(13):E1300–E1309
4. Dickinson AJ, Lehner K, Mi J, Jia K-P,
Mijar M, Dinneny J, Al-Babili S, Benfey PN
(2019) β-Cyclocitral is a conserved root
growth regulator. Proc Natl Acad Sci 116
(21):10563–10567
5. Al-Babili S, Bouwmeester HJ (2015) Strigolactones, a novel carotenoid-derived plant hormone. Annu Rev Plant Biol 66:161–186
6. Cazzonelli CI, Hou X, Alagoz Y, Rivers J,
Dhami N, Lee J, Marri S, Pogson BJ (2019)
A cisÀcarotene derived apocarotenoid regulates etioplast and chloroplast development.
bioRxiv 528331. https://doi.org/10.1101/
528331
7. Wang JY, Haider I, Jamil M, Fiorilli V, Saito Y,
Mi J, Baz L, Kountche BA, Jia K-P, Guo X
(2019) The apocarotenoid metabolite zaxinone regulates growth and strigolactone biosynthesis in rice. Nat Commun 10(1):810
8. Iskandar AR, Miao B, Li X, Hu K-Q, Liu C,
Wang X-D (2016) β-Cryptoxanthin reduced
lung tumor multiplicity and inhibited lung cancer cells motility by down-regulating nicotinic
acetylcholine receptor α7 expression. Cancer
Prev Res 9(11):875–886
Profiling Carotenoids in Plants Using HPLC
161