下列九個類黃酮化合物(1至9)係分離自菲律賓楠(Machilus
philippinensis),其分離方法/步驟如下: The EtOH extract
(160 g) of the dried leaves of M. philippinensis (774 g) was partitioned
into fractions soluble in CH2Cl2 (38.30 g), EtOAc (10.51 g), n-BuOH
(31.02 g) and H2O (74.12 g) by a liquid-liquid partitioning process. Part
of the EtOAc-soluble fraction (6.03 g out of 10.51 g) was fractionated on a
Sephadex LH-20 column (815 × 35 mm, MeOH) to give six fractions (E1~6). An
aliquot of fraction E2 (45.0 mg out of 500.7 mg) was separated by a
semi-preparative RP-18 HPLC column (Phenomenex® Prodigy ODS-3, 250 × 10 mm, 5
μm), each run 5.0 mg, delivered by 16% MeCN in H2O for 30 min, to 25%
MeCN in 40 min by a linear gradient mode, and then MeCN for 15 min, with a flow
rate of 2.5 mL/min and detection at 300 nm. After nine runs, the fractions
containing pure compounds were evaporated under reduced pressure to give 3 (2.9 mg, tR =
21.39 min), 4 (10.0
mg, tR
= 22.61 min), 6 (7.3 mg, tR = 30.86 min), 7 (4.8 mg, tR =
36.36 min), and 9
(1.9 mg, tR = 42.33 min), respectively. An aliquot of fraction E3
(273.8 mg out of 1.20 g) was separated on a Lobar (low-pressure) RP-18 column
(LiChroprep RP-18, size B, 310 × 25 mm; 40-63 μm, Merck), delivered by a
stepwise gradient of MeOH―H2O from 30:70 to 100:0, to give six
subfractions (E3-1~6). Fractions E3-3 (24.7 mg) and -5 (2.0 mg) were pure 5 and 8, respectively. An
aliquot of fraction E6 (201.0 mg out of 756.0 mg) was separated on the same Lobar
RP-18 column, delivered by a stepwise gradient of MeOH―H2O from
35:65 to 85:15, to give five subfractions (E6-1~5). Fractions E6-2 (14.4 mg)
and -4 (2.3 mg) were pure 1
and 2,
respectively.
(一) 以上圖右之方式表達化合物1至9之分離流程。(10分)
(二) 試由分離之結果簡述所使用的層析法之分離原理與特性。(16分)