| Summary: | 碩士 === 輔仁大學 === 食品科學系 === 99 === Taraxacum formosanum, a traditional Chinese herb, has been reported to possess pharmacological functions like antioxidative, anti-inflammatory and antimutagenic activities. In recent years the physiological studies have focused mainly on flavonoids, however, the other functional components like carotenoids and chlorophylls may also play a vital role. The objectives of this study were to determine the variety and content of carotenoids and chlorophylls in T. formosanum by using high performance liquid chromatography-diode array detection-mass spectrometry methods with atmospheric pressure chemical ionization mode (HPLC-DAD-APCI-MS) and develop a preparative column chromatographic method for separation of carotenoids and chlorophylls.
Initially, both carotenoids and chlorophylls were extracted with hexane-ethanol-acetone-toluene (10:6:7:7, v/v/v/v). A total of 25 carotenoids were separated within 66 min by employing a YMC C30 column and a gradient mobile phase of methanol-acetonitrile-water (79:14:7, v/v/v)(A) and methylene chloride(B) with the following condition: 95% A in the beginning, maintained for 9 min, decreased to 85% A in 23 min, 83% A in 33 min, 71% A in 35 min, 70% A in 45 min, and 66% A in 66 min with flow rate at 1 mL/min and detection wavelength at 450 nm. All-trans-canthaxanthin was used as an internal standard to quantify all the carotenoids. All-trans-β-carotene and its cis isomers was present in largest amount (413.6 μg/g), followed by all-trans-violaxanthin and its cis isomers (225.9 μg/g), all-trans-lutein and its cis isomers (212.4 μg/g), all-trans-neoxanthin and its cis isomers (134.7 μg/g), antheraxanthin (16.5 μg/g), all-trans-β-cryptoxanthin and its cis isomers (5.8 μg/g), all-trans-zeaxanthin (3.6 μg/g).
Likewise, a total of 11 chlorophylls were separated within 30 min by a HyPURITY C18 column and a quaternary solvent system of water(A), methanol(B), acetonitrile(C) and acetone(D) in gradient mode was developed: 70% A and 30% B in the beginning, changed to 45% A and 55% B in 0.3 min, 100% B in 4 min, 38% B and 62% C in 6 min, 50% B and 50% C in 10 min, 51% B and 49% C in 15 min, 57% B and 43% C in 22 min, 60% B and 40% D in 26 min, 45% B and 55% D in 30 min with flow rate at 1 mL/min and detection wavelength at 660 nm. Fast Green FCF was used as an internal standard to quantify all the chlorophylls. Chlorophyll a and its isomer (1389.6 μg/g) was present in the largest amount in T. formosanum, followed by chlorophyll b and its isomer (561.2 μg/g), pheophytin a and its isomer (31.8 μg/g), hydroxychlorophyll b (26.5 μg/g), hydroxychlorophyll a and its isomer (9.8 μg/g), chlorophyllide a and its isomer (0.4 μg/g).
For preparation of carotenoid and chlorophyll fractions, 3 mL of hexane extract was poured into a column containing 52 g of MgO-diatomaceous earth (1: 3, w/w). With a flow rate of 10 mL/min, 300 mL of ethyl acetate could elute carotenoids including all-trans- violaxanthin, all-trans-lutein, all-trans-β-carotene and their cis isomers, whereas 800 mL of 50% ethanol in acetone eluted the chlorophyll fraction containing hydroxychlorophyll a, hydroxychlorophyll b and their derivatives. The carotenoid fraction was present in largest amount (490.5 μg/mL) and followed by chlorophyll fraction (400.9 μg/mL).
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