An improved HPLC method for determination of carotenoids in human serum.

An HPLC method was developed to determine the various carotenoids in human serum. A C-30 column and a mobile phase of 100% methanol (A) and 100% methylene chloride (B) with the following gradient elution were used: 90% A and 10% B in the beginning, maintained for 5 min, decreased to 78% A at 15 min, 62% A at 30 min, 52% A at 40 min, 41% A at 50 min, 38% A at 55 min, maintained for 3 min, and returned to 100% A at 65 min. A total of 21 carotenoids, including all-trans forms of lutein, zeaxanthin, alpha-cryptoxanthin, beta-cryptoxanthin, alpha-carotene, beta-carotene and lycopene, as well as their 14 cis-isomers were resolved within 51 min at a flow rate of 1.0 mL/min and detection at 476 nm. all-trans-beta-Carotene was found to be present in highest amount (256.3-864.2 ng/mL), followed by all-trans-lycopene (64.4-569.2 ng/mL), all-trans-lutein (137.9-450.3 ng/mL), all-trans-alpha-cryptoxanthin (55.7-188.2 ng/mL), all-trans-beta-cryptoxanthin (43.1-134.5 ng/mL), all-trans-alpha-carotene (20.0-122.1 ng/mL) and all-trans-zeaxanthin (9.1-21.3 ng/mL). Similar trend was observed for cis-isomers of carotenoids.