76 JOURNAL OF CHEMICAL RESEARCH 2016
s, 4H, piperazine, H-15, H-17), 3.02 (t, J = 5.4 Hz, 2H, H-5); 13C NMR
(DMSO-d6): δ 161.8, 158.0, 150.8, 149.2, 147.4, 145.7, 143.1, 137.5,
132.4, 126.5, 123.8, 122.7, 121.6, 120.9, 120.1, 110.5, 108.3, 105.4,
101.8, 63.5, 57.7, 55.6, 50.9, 42.8, 26.6; HRMS m/z [M – Cl]+ calcd for
[C28H28N5O4]+: 498.55; found: 498.30. Anal. calcd for C28H28ClN5O4:
C, 62.98; H, 5.29; N, 13.11; found: C, 62.85; H, 5.42; N, 13.02%.
12-[1-(2-Pyridyl)piperazin-1-ylmethyl]berberrubine (4d): Light
yellow solid; yield 61%; m.p. 213–215 °C (CHCl3); IR (KBr) cm–1:
3620 (OH), 3068 (C–H, Ar), 1652–1566 (C=C, Ar), 1269 (ArN);
1H NMR (CDCl3): δ 9.74 (s, 1H, –OH of berberine ring), 9.22 (s, 1H,
H-8), 8.05 (s, 1H, H-13), 7.27 (s, 1H, H-1), 7.20 (s, 1H, H-4), 6.92–6.56
(m, 4H, ArH, piperazine), 6.19 (s, 2H, –OCH2O–), 5.97 (s, 1H, H-11),
5.75 (t, J = 5.4 Hz, 2H, H-6), 3.91 (s, 2H, CH2, H-14 Mannich base),
3.85 (s, 3H, –OCH3), 3.67 (br s, 4H, piperazine, H-18, H-20), 3.20 (br
s, 4H, piperazine, H-15, H-17), 3.02 (t, J = 5.4 Hz, 2H, H-5); 13C NMR
(DMSO-d6): δ 159.8, 150.2, 149.9, 148.2, 147.7, 145.3, 143.4, 141.9, 137.8,
132.1, 126.6, 123.6, 122.5, 121.5, 120.7, 120.2, 114.3, 108.5, 107.6, 105.3,
101.3, 63.7, 57.6, 55.8, 50.5, 42.6, 26.1; HRMS m/z [M – Cl]+ calcd for
[C29H29N4O4]+: 497.56; found: 497.60. Anal. calcd for C29H29ClN4O4: C,
65.35; H, 5.48; N, 10.51; found: C, 65.47; H, 5.34; N, 10.62%.
6.03 (s, 1H, H-11), 5.86 (t, J = 5.3 Hz, 2H, H-6), 3.92 (s, 2H, CH2,
H-14 Mannich base), 3.88 (s, 3H, –OCH3), 3.68 (br s, 4H, piperadine),
3.04 (t, J = 5.3 Hz, 2H, H-5), 1.82 (br s, 6H, piperadine); 13C NMR
(DMSO-d6): δ 150.5, 149.3, 147.0, 145.8, 143.5, 137.2, 132.5, 126.1,
123.7, 122.6, 121.0, 120.6, 120.3, 108.6, 105.2, 101.6, 63.6, 57.8, 55.9,
52.4, 25.6, 26.5, 21.2; HRMS m/z [M – Cl]+ calcd for [C25H27N2O4]+:
419.49; found: 419.40. Anal. calcd for C25H27ClN2O4: C, 66.00; H, 5.98;
N, 6.16; found: C, 66.14; H, 6.04; N, 6.07%.
12-(Carbazol-1-ylmethyl)berberrubine (4i): Light yellow solid; yield
48%; m.p. 264–266 °C (CHCl3); IR (KBr) cm–1: 3614 (OH), 3013 (C–
1
H, Ar), 1648–1571 (C=C, Ar), 1283 (ArN); H NMR (CDCl3): δ 9.73
(s, 1H, –OH of berberine ring), 9.24 (s, 1H, H-8), 8.02 (s, 1H, H-13),
7.26 (s, 1H, H-1), 7.09 (s, 1H, H-4), 6.85–6.42 (m, 8H, ArH, carbazole),
6.22 (s, 2H, –OCH2O–), 6.05 (s, 1H, H-11), 5.73 (t, J = 5.5 Hz, 2H,
H-6), 4.16 (s, 2H, CH2, H-14 Mannich base), 3.71 (s, 3H, –OCH3), 3.14
(t, J = 5.4 Hz, 2H, H-5); 13C NMR (DMSO-d6): δ 150.9, 149.1, 147.2,
145.6, 143.3, 140.1 (2C), 137.1, 132.8, 128.6, 126.3, 125.4, 123.2, 122.4,
121.2, 120.9, 120.3, 119.2, 118.5, 110.7, 108.8, 105.1, 101.4, 63.0, 57.5,
55.3, 26.2; HRMS m/z [M – Cl]+ calcd for [C32H25N2O4]+: 501.55;
found: 501.60. Anal. calcd for C32H25ClN2O4: C, 71.57; H, 4.69; N,
5.22; found: C, 71.38; H, 4.58; N, 5.11%.
12-[1-(2-Furoyl)piperazin-1-ylmethy]berberrubine (4e): Light
yellow solid; yield 53%; m.p. 247–249 °C (CHCl3); IR (KBr) cm–1:
3625 (C–H, Ar), 3059 (C–H, Ar), 1713 (C=O), 1649–1570 (C=C,
Evaluation of antioxidant capacity by DPPH and ABTS•+ assay
The free radical scavenging activities of the berberine derivatives 4a–i,
as IC50 values in μM, were determined as described previously by us17,18
via assays using 1,1-diphenyl-2-picrylhydrazyl free radicals (DPPH)
and 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) radical
cations (ABTS•+).
1
Ar), 1285 (ArN); H NMR (CDCl3): δ 9.75 (s, 1H, –OH of berberine
ring), 9.32 (s, 1H, H-8), 7.97 (s, 1H, H-13), 7.30 (s, 1H, H-1), 7.19 (s, 1H,
H-4), 6.79–6.48 (m, 5H, ArH, piperazine), 6.25 (s, 2H, –OCH2O–),
6.08 (s, 1H, H-11), 5.83 (t, J = 5.5 Hz, 2H, H-6), 4.15 (s, 2H, CH2, H-14
Mannich base), 3.81 (s, 3H, –OCH3), 3.62 (br s, 4H, piperazine, H-18,
H-20), 3.13 (br s, 4H, piperazine, H-15, H-17), 3.03 (t, J = 5.3 Hz, 2H,
H-5); 13C NMR (DMSO-d6): δ 171.6, 150.1, 149.7, 147.5, 145.1, 143.2,
137.6, 132.2, 131.9, 130.5, 128.4, 126.8, 123.0, 122.8, 121.7, 121.2,
120.5, 120.1, 108.1, 105.7, 101.5, 63.3, 57.9, 55.2, 50.7, 42.1, 37.1, 26.4;
HRMS m/z [M – Cl]+ calcd for [C30H30N3O5]+: 512.58; found: 512.50.
Anal. calcd for C30H30ClN3O5: C, 65.75; H, 5.52; N, 7.67; found: C,
65.59; H, 5.61; N, 7.78%.
12-(1-Benzhydrylpiperazin-1-ylmethyl)berberrubine (4f): Light
yellow solid; yield 55%; m.p. 275–277 °C (CHCl3); IR (KBr) cm–1:
3619 (OH), 3019 (C–H, Ar), 1655–1579 (C=C, Ar), 1273 (ArN);
1H NMR (CDCl3): δ 9.82 (s, 1H, –OH of berberine ring), 9.20 (s, 1H,
H-8), 8.03 (s, 1H, H-13), 7.21 (s, 1H, H-1), 7.02 (s, 1H, H-4), 6.93–6.49
(m, 10H, ArH, piperazine), 6.35 (s, 2H, –OCH2O–), 6.11 (s, 1H, H-11),
5.74 (t, J = 5.5 Hz, 2H, H-6), 5.22 (s, 1H, –CH), 3.94 (s, 2H, CH2, H-14
Mannich base), 3.76 (s, 3H, –OCH3), 3.65 (br s, 4H, piperazine, H-18,
H-20), 3.16 (br s, 4H, piperazine, H-15, H-17), 3.10 (t, J = 5.5 Hz, 2H,
H-5); 13C NMR (DMSO-d6): δ 150.3, 149.5, 147.9, 145.0, 143.9, 141.6,
137.3, 132.3, 131.7, 130.1, 129.3, 128.3, 127.7, 126.0, 123.1, 122.2, 121.9,
120.8, 120.0, 108.2, 105.9, 101.7, 80.4, 63.9, 57.2, 55.5, 50.2, 42.3, 26.0;
HRMS m/z [M – Cl]+ calcd for [C37H36N3O44]+: 586.70; found: 586.60.
Anal. calcd for C37H36ClN3O4: C, 71.43; H, 5.83; N, 6.75; found: C,
71.52; H, 5.93; N, 6.59%.
FRAP assay
The FRAP assay of the compounds was performed using the modified
method described by Benzie and Strain.19 FRAP analysis measured the
in vitro antioxidant activity of all the newly synthesised compounds
by the reduction of a colourless Fe3+ tripyridyltriazine complex into
a blue-coloured Fe2+–tripyridyltriazine complex. The antioxidant
potentials of the compounds were estimated as their power to reduce
the 2,4,6-tripyridyl-s-triazine (TPTZ)–Fe(III) complex to TPTZ–
Fe(II) complex, which is simple, fast, and reproducible. The stock
solutions included 300 mM acetate buffer (3.1 g C2H3NaO2·3H2O and
16 mL C2H4O2), pH 3.6, 10 mM TPTZ solution in 40 mM hydrochloric
acid and 20 mM ferric chloride hexahydrate solution. The fresh
working solution was prepared by mixing acetate buffer (25 mL),
TPTZ (2.5 mL) and ferric chloride hexahydrate solution (2.5 mL).
The temperature of the solution was raised to 37 °C before use and
the solution was allowed to react with the FRAP solution (300 μL) in
a volume ratio of 10:1:1 respectively. Methanol (1 mL) was used as a
blank, test samples were dissolved in methanol and ascorbic acid as
standard was dissolved in water. The reaction was allowed to run for
30 min. The coloured product (ferrous tripyridyl triazine complex) was
monitored at λ = 593 nm. The experiments were performed in triplicate
and the mean was calculated for each compound.
12-(Morpholinomethyl)berberrubine (4g): Light yellow solid; yield
72%; m.p 219–221 °C (CHCl3); IR (KBr) cm–1: 3612 (OH), 3071 (C–H,
In vitro anticancer bioassay
1
Cancerous growth inhibitory efficacies (expressed as IC50 values in
μM) were assessed using cervical cancer cell lines HeLa and CaSki
and their cytotoxicities (expressed as CC50 values in μM) towards
normal cell lines were evaluated using Madin–Darby canine kidney
(MDCK) cell lines as described previously by us.17,18
Ar), 1641–1585 (C=C, Ar), 1263 (ArN); H NMR (CDCl3): δ 9.86 (s,
1H, –OH of berberine ring), 9.27 (s, 1H, H-8), 7.94 (s, 1H, H-13), 7.32
(s, 1H, H-1), 7.18 (s, 1H, H-4), 6.23 (s, 2H, –OCH2O–), 6.15 (s, 1H,
H-11), 5.71 (t, J = 5.4 Hz, 2H, H-6), 4.11 (s, 2H, CH2, H-14 Mannich
base), 3.82 (s, 3H, –OCH3), 3.72 (br s, 4H, morpholine, H-18, H-20),
3.19 (br s, 4H, morpholine, H-15, H-17), 2.98 (t, J = 5.5 Hz, 2H, H-5);
13C NMR (DMSO-d6): δ 150.4, 149.8, 147.6, 145.4, 143.6, 137.4, 132.9,
126.7, 123.5, 122.3, 121.4, 120.4, 120.2, 108.4, 105.5, 101.9, 66.1, 63.4,
57.0, 55.1, 52.8, 26.3; HRMS m/z [M – Cl]+ calcd for [C24H25N2O5]+:
421.47; found: 421.30; Anal. calcd for C24H25ClN2O5: C, 63.09; H, 5.51;
N, 6.13; found: C, 62.98; H, 5.37; N, 6.02%.
This work was supported by the KU Research Professor
Programme of Konkuk University, Seoul, South Korea.
Received 24 November 2015; accepted 21 December 2015
Published online: 13 January 2016
12-(Piperidin-1-ylmethyl-berberrubine (4h): Light yellow solid;
yield 66%; m.p. 206–208 °C (CHCl3); IR (KBr) cm–1: 3633 (OH), 3023
1
(C–H, Ar), 1663–1562 (C=C, Ar), 1266 (ArN); H NMR (CDCl3):
References
δ 9.96 (s, 1H, –OH of berberine ring), 9.19 (s, 1H, H-8), 7.95 (s, 1H,
H-13), 7.24 (s, 1H, H-1), 7.05 (s, 1H, H-4), 6.29 (s, 2H, -OCH2O–),
1
2