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ARS Home » Northeast Area » Beltsville, Maryland (BARC) » Beltsville Agricultural Research Center » Genetic Improvement for Fruits & Vegetables Laboratory » Research » Publications at this Location » Publication #343986

Title: An improved method for fast and selective separation of carotenoids by UPLC-MS

item ABATE-PELLA, DANIEL - University Of Minnesota
item FREUND, DANA - University Of Minnesota
item Slovin, Janet
item HEGEMAN, ADRIAN - University Of Minnesota
item COHEN, JERRY - University Of Minnesota

Submitted to: Journal of Chromatography
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/25/2017
Publication Date: 9/28/2017
Publication URL:
Citation: Abate-Pella, D., Freund, D.M., Slovin, J.P., Hegeman, A.D., Cohen, J.D. 2017. An improved method for fast and selective separation of carotenoids by UPLC-MS. Journal of Chromatography.

Interpretive Summary: Carotenoids consist of a large group of chemical compounds that are important for human health and sight, as well as for coloration of flowers and animals. They are powerful antioxidants made by plants, fungi and bacteria, but they are not made by most animals and humans have a nutritional requirement for them. Their chemical nature and low amounts in biological samples make them difficult to identify and measure which is important for crop improvement because increases in carotenoids add value. In this paper we report the development of an improved method to separate and unambiguously identify different carotenoids. We show that the method is capable of analyzing a wide range of samples, including strawberry leaves, chicken feed supplements, and bacterial cultures. This work will be used by nutritionists, plant breeders, and plant molecular biologists interested in human health and improving the quality of plant products.

Technical Abstract: Carotenoids are a large class of compounds that are biosynthesized by condensation of isoprene units in plants, fungi, bacteria, and some animals. They are characteristically highly conjugated through double bonds, which lead to many isomers as well susceptibility to oxidation and other chemical modifications. Carotenoids are important because of their potent antioxidant activity and are the pigments responsible for color in a wide variety of foods. Human consumption is correlated to many health benefits including prevention of cancer, cardiovascular disease, and age-related disease. Extreme hydrophobicity, poor stability, and low concentration in biological samples make these compounds difficult to analyze and difficult to develop analytical methods for aimed towards identification and quantification. Examples in the literature frequently report the use of exotic stationary phases, solvents, and additives, such as ethyl acetate, dichloromethane, and methyl tert-butyl ether that are incompatible with liquid chromatography mass spectrometry (LC-MS). In order to address these issues, we implemented the use of LC-MS friendly conditions using a low-hydrophobicity cyano-propyl column (Agilent Zorbax SB-CN). We successfully differentiated between isomeric carotenoids by optimizing two gradient methods and using a mixture of 11 standards and ultra performance liquid chromatography (UPLC) coupled to mass spectrometry (MS) in positive ionization mode. Three complex biological samples from strawberry leaf, chicken feed supplement, and the photosynthetic bacterium Chloroflexus aurantiacus were analyzed and several carotenoids were resolved in these diverse backgrounds. Our results show this methodology is a significant improvement over other alternatives for analyzing carotenoids because of its ease of use, rapid analysis time, high selectivity, and, most importantly, its compatibility with typical LC-MS conditions.