Supporting information
The extraction and isolation processes of MHBFC
9 kg cut crude drugs ofMillettiapulchra (Benth.) Kurz var. Laxior (Dunn)Z. Wei(Papilionaceae) was extracted with 72 L of 60% EtOH by microwave-assisted extraction for 20 min. x 2 times at 55°C as reported [1]. The filtered solvents were combined and concentrated under reduced pressure to yield 1.5 L of crude extract. After washing with 3 L of petroleum ether (PE) to remove pigments，the crude extract was further partitioned with 4.5 L of ethyl acetate (AcOEt). The AcOEt layer was concentrated and dried in vaccum to obtain a dry residue (100 g). This dry residue was fractionated by column chromatography (CC) using a silica gel H-packed column (10 cm × 120 cm) with a gradient system of PE/AcOEt (98:2) to PE/AcOEt (0:100) to obtain 269 fractions Fr.Gp.1 (F1- F269) (one fraction per 0.5 L of eluant). Then, all fractions were combined according to thinlayer chromatography (TLC) analysis and afforded fractions A–M. Antioxidative activities of all prefractionational fractions were evaluated to sieve the further purified constituents [2]. Fr.H indicated strong antioxidative effect, so Fr.H (5.5 g) was further fractioned by CC (275 g, ф5 cm x 60 cm), eluted with a gradient of PE-AcOEt (96:4, 92:8, 88:12) to divided into frs.H1-H3. Fr.H1(2.8 g) was subjected to another CC fractionation (silica gel-H, ф2.5 cm x 60 cm), eluted with a gradient of PE-AcOEt (100:0, 98:2, 92:8) to afford frs.H1a-H1c. Fr.H1a (700 mg) was further re-crystallized by CHCl3/MeOH (50:50). Finally, 12 mg of MHBFC (Rf=0.59 with PE-EtOAc=4:1 as developer) was obtained by preparative liquid chromatography purification using an ODS-BP column (10 cm × 250 cm) at 3 mL/min with acetonitrile/0.2% acetic acid (85:15) as mobile phase, λ 238 nm, and tR 34.10-35.20 minute.
References
1.Jian, J., Lin, X., Huang, R., et al. (2009). The study on total flavonoids extraction of Yulangsan.Lishizhen Med. Materia Med. Res., 8: 1998-1999
2.Jian J., Li, Y., Jiang, W., et al. (2009).The effects of two chalcone monomers from Yulangsan on scavenging free radicals. Chin. J. Gerontology, 18: 2353-2354.
Table 1S1H- and 13C- NMR spectroscopic data of MHBFC (CDCl3, 500 MHz)
Position / δH / δC / Position / δH / δC1 / 141.8 / 10 / 122.0
2 / 7.57(overlapped) / 128.6 / 11 / 8.17(s) / 128.4
3 / 7.57(overlapped) / 130.6 / 12 / 117.0
4 / 7.57(overlapped) / 128.6 / 13 / 154.8
5 / 7.57(overlapped) / 130.6 / 14 / 8.2(d) / 145.7
6 / 7.57(overlapped) / 128.6 / 15 / 7.2(d) / 110.0
7 / 158.2 / 16 / 119.8
8 / 7.77(s) / 104.2 / 17 / 8.17(s) / 150.0
9 / 174.2 / 18 / 3.95(s) / 60.2
Table 2SBond lengths [nm] and angles [°] of MHBFC.
Bond Bond length Bond Bond length Bond Bond angle
C(1)-C(6) 1.392(2) C(18)-H(18A) 0.9800 C(11)-C(10)-C(9) 116.80(12)
C(1)-C(2) 1.395(2) C(18)-H(18B) 0.9800 C(12)-C(11)-C(10) 122.56(13)
C(1)-C(7) 1.4845(19) C(18)-H(18C) 0.9800 C(12)-C(11)-H(11) 118.7
C(2)-C(3) 1.383(2) O(1)-H(1) 0.8400 C(10)-C(11)-H(11) 118.7
C(2)-H(2) 0.9500 Bond Bond angle C(11)-C(12)-C(13) 116.21(13)
C(3)-C(4) 1.389(2) C(6)-C(1)-C(2) 118.99(13) C(11)-C(12)-H(12) 121.9
C(3)-H(3) 0.9500 C(6)-C(1)-C(7) 119.40(13) C(13)-C(12)-H(12) 121.9
C(4)-C(5) 1.382(2) C(2)-C(1)-C(7) 121.60(12) O(3)-C(13)-C(12) 126.00(12)
C(4)-H(4) 0.9500 C(3)-C(2)-C(1) 120.53(13) O(3)-C(13)-C(16) 110.09(13)
C(5)-C(6) 1.387(2) C(3)-C(2)-H(2) 119.7 C(12)-C(13)-C(16) 123.91(13)
C(5)-H(5) 0.9500 C(1)-C(2)-H(2) 119.7 C(15)-C(14)-O(3) 112.71(13)
C(6)-H(6) 0.9500 C(2)-C(3)-C(4) 120.11(15) C(15)-C(14)-H(14) 123.6
C(7)-O(1) 1.3035(17) C(2)-C(3)-H(3) 119.9 O(3)-C(14)-H(14) 123.6
C(7)-C(8) 1.3833(19) C(4)-C(3)-H(3) 119.9 C(14)-C(15)-C(16) 105.78(13)
C(8)-C(9) 1.4195(19) C(5)-C(4)-C(3) 119.66(14) C(14)-C(15)-H(15) 127.1
C(8)-H(8) 0.9500 C(5)-C(4)-H(4) 120.2 C(16)-C(15)-H(15) 127.1
C(9)-O(2) 1.2776(17) C(3)-C(4)-H(4) 120.2 C(13)-C(16)-C(17) 118.85(13)
C(9)-C(10) 1.4838(19) C(4)-C(5)-C(6) 120.43(14) C(13)-C(16)-C(15) 105.93(12)
C(10)-C(17) 1.3987(19) C(4)-C(5)-H(5) 119.8 C(17)-C(16)-C(15) 135.21(13)
C(10)-C(11) 1.4105(18) C(6)-C(5)-H(5) 119.8 O(4)-C(17)-C(16) 118.54(12)
C(11)-C(12) 1.380(2) C(5)-C(6)-C(1) 120.26(14) O(4)-C(17)-C(10) 122.42(12)
C(11)-H(11) 0.9500 C(5)-C(6)-H(6) 119.9 C(16)-C(17)-C(10) 119.03(12)
C(12)-C(13) 1.381(2) C(1)-C(6)-H(6) 119.9 O(4)-C(18)-H(18A) 109.5
C(12)-H(12) 0.9500 O(1)-C(7)-C(8) 121.65(13) O(4)-C(18)-H(18B) 109.5
C(13)-O(3) 1.3671(17) O(1)-C(7)-C(1) 115.55(12) H(18A)-C(18)-H(18B) 109.5
C(13)-C(16) 1.3949(19) C(8)-C(7)-C(1) 122.80(12) O(4)-C(18)-H(18C) 109.5
C(14)-C(15) 1.338(2) C(7)-C(8)-C(9) 120.37(13) H(18A)-C(18)-H(18C) 109.5
C(14)-O(3) 1.3779(19) C(7)-C(8)-H(8) 119.8 H(18B)-C(18)-H(18C) 109.5
C(14)-H(14) 0.9500 C(9)-C(8)-H(8) 119.8 C(7)-O(1)-H(1) 109.5
C(15)-C(16) 1.437(2) O(2)-C(9)-C(8) 120.69(13) C(13)-O(3)-C(14) 105.49(11)
C(15)-H(15) 0.9500 O(2)-C(9)-C(10) 117.02(12) C(17)-O(4)-C(18) 114.70(11) 1.4219(19) 0.9800
C(16)-C(17) 1.3972(19) C(8)-C(9)-C(10) 122.21(12)
C(17)-O(4) 1.3784(16) C(17)-C(10)-C(11)119.42(13)
C(18)-O(4) 1.4219(19) C(17)-C(10)-C(9) 123.74(12)
1
Figure 1S The extraction and isolation processes of MHBFC.
Figure 2SThe chemical structure of MHBFC.
Figure 3SMolecular stereo configuration and crystal packing of MHBFC.