Synthesis and antitumour activities of a novel class of dehydroabietylamine derivatives

Article abstract of DOI:10.1080/14786419.2011.648191

Structural modification is still a popular and important route in the forest chemical field for finding novel tricyclic diterpenes with more potential bioactivities and broad bioactive spectra. In this study, a series of dehydroabietylamine derivatives containing tricyclic diterpene structures were synthesised through oxidation in the 7th position of ring B and nitrification in the 12th position of ring C using dehydroabietylamine as the starting material. Structures of the synthesised compounds were confirmed by IR, ¹H-NMR, ¹³C-NMR, MS and HRMS. The cytotoxicities of these compounds against PC-3 (human prostate carcinoma cell line) and Hey-1B (human ovarian carcinoma cell line) cells by the MTT assay were investigated. The results showed that the presence of a nitro group at 12th position and a carbonyl group at 7th position resulted in an increase of cytotoxic activity. Our findings present more evidence, showing the relationship between the chemical structure and biological function.

Full text of DOI:10.1080/14786419.2011.648191

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Synthesis and antitumour activities of
a novel class of dehydroabietylamine
derivatives
Yong Chen a , Zhong-Xiang Lin b & Ai-Min Zhou c
a College of Science , Nanjing Forestry University , Nanjing
210037 , PR China
b College of Chemical Engineering , Nanjing Forestry University ,
Nanjing 210037 , PR China
c Department of Chemistry , Cleveland State University ,
Cleveland , OH 44115 , USA
Published online: 11 Jan 2012.
To cite this article: Yong Chen , Zhong-Xiang Lin & Ai-Min Zhou (2012) Synthesis and antitumour
activities of a novel class of dehydroabietylamine derivatives, Natural Product Research: Formerly
Natural Product Letters, 26:23, 2188-2195, DOI: 10.1080/14786419.2011.648191
To link to this article: http://dx.doi.org/10.1080/14786419.2011.648191
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Natural Product Research
Vol. 26, No. 23, December 2012, 2188–2195
Synthesis and antitumour activities of a novel class
of dehydroabietylamine derivatives
c
Yong Chena, Zhong-Xiang Linb and Ai-Min Zhou
*
aCollege of Science, Nanjing Forestry University, Nanjing 210037, PR China; bCollege of Chemical
Engineering, Nanjing Forestry University, Nanjing 210037, PR China; Department of Chemistry,
Cleveland State University, Cleveland, OH 44115, USA
c
(Received 28 March 2011; final version received 11 September 2011)
Structural modification is still a popular and important route in the forest
chemical field for finding novel tricyclic diterpenes with more potential
bioactivities and broad bioactive spectra. In this study, a series of dehydroabie-
tylamine derivatives containing tricyclic diterpene structures were synthesised
through oxidation in the 7th position of ring B and nitrification in the 12th
position of ring C using dehydroabietylamine as the starting material. Structures
of the synthesised compounds were confirmed by IR, 1H-NMR, 13C-NMR,
MS and HRMS. The cytotoxicities of these compounds against PC-3 (human
prostate carcinoma cell line) and Hey-1B (human ovarian carcinoma cell line)
cells by the MTT assay were investigated. The results showed that the presence
of a nitro group at 12th position and a carbonyl group at 7th position resulted
in an increase of cytotoxic activity. Our findings present more evidence, showing
the relationship between the chemical structure and biological function.
Keywords: dehydroabietylamine; antitumour; cytotoxicity
1. Introduction
Cancer remains the second leading cause of death in most countries and as a result, there is
a need for more effective compounds. Total synthesis and modification of the structures
of bioactive natural products are effective methods to find potential compounds with
anticancer activity. Natural tricyclic diterpenes are an important class of potential
anticancer drugs. Some tricyclic diterpenes from natural products have been synthesised or
modified, and their anticancer activities have been investigated (Gigante et al., 2003;
Grace, Jin, Wilson, & Coates, 2006; Son, Oh, Choi, Han, & Kwon, 2005). In particular,
functional group modification at C-7 and C-12 positions can be a way to prepare
compounds with potential biological interest (Gonzalez et al., 2010; Li, She, Zhang,
´
Wu, & Pan, 2003; Xiong, Wang, Pan, Sun, & Tu, 2006; Zhang & Lin, 2009).
Dehydroabietylamine (Figure 1) is a distinctive synthetic primary amine having a tricyclic
diterpene structure which is obtained as part of a mixture of amines prepared by the
hydrogenation of rosin acid nitrile. Dehydroabietylamine is widely used in the fields of
paper-making, medicine, pesticide and chemical industries (Rao, Song, & He, 2008;
Wilkerson, Galbraith, DeLucca, & Harris, 1993). Recently, antitumour and cytotoxic
*Corresponding author. Email: linzhongxiang36@gmail.com
ISSN 1478–6419 print/ISSN 1478–6427 online
ß 2012 Taylor & Francis
http://dx.doi.org/10.1080/14786419.2011.648191
http://www.tandfonline.com

Natural Product Research
2189
Figure 1. Reagents and conditions: (a) benzoyl chloride and substituted benzoyl chloride, toluene,
80ꢁC, 8 h, (b) CrO3, AcOH, 60ꢁC, 12 h and (c) Cu(NO3)2, Ac2O, 10ꢁC, 12 h.
activities of dehydroabietylamine derivatives are a focus of research in the forest chemical
field.
Pursuing our studies in the chemical transformation of dehydroabietylamine, we have
been looking at the possibility of preparing new compounds from dehydroabietylamine
through the functionalisation of rings B and C (Figure 1). In this communication, a series
of C-7 oxidised or C-12 nitrated abietane diterpenes were synthesised from dehydroabie-
tylamine, and their antitumour activities were evaluated against two cultured human-
tumour cell lines (PC-3 and Hey-1B) for investigating the influences of functional groups
modification on the bioactivities. Among these compounds, 3a, 4a, 4b and 4c showed good
cytotoxicities against PC-3 cancer cells (IC50 5.7–8.2 mg mLꢀ1) and Hey-1B cancer cells
(IC50 11.3–16.0 mg mLꢀ1). Our results have revealed that the presence of a nitro group at
12th position and a carbonyl group at 7th position resulted in an increase of cytotoxic
activity. These results may provide useful information for the design of anticancer drugs.
2. Results and discussion
2.1. Chemistry
To determine the effect of the type and the position of functional groups on bioactivity,
a series of novel tricyclic diterpenes with a carbonyl group in the 7th position on ring B or
a nitro group in the 12th position on ring C and benzamides with various substituents

2190
Y. Chen et al.
in the 18th position were synthesised using dehydroabietylamine as the starting material
(Figure 1).
2.1.1. Synthesis of analogues of N-benzoyl-dehydroabietylamine (2a)
Dehydroabietylamine (compound 1) was converted to compounds 2a–2c via acylation with
benzoyl chloride and substituted benzoyl chloride in toluene. The structures of compounds
2a–2c were confirmed by analyses of IR and 1H-NMR spectra. The presence of the
characteristic band of amides at 1644–1638 cmꢀ1 was observed in the IR spectra of 2a–2c.
The formation of amides was easily confirmed by their 1H-NMR spectrum due to a broad
one-proton singlet at ꢀ 6.10–6.24 ppm corresponding to the amides N–H. In contrast to
the standard spectra of dehydroabietylamine, the chemical shift value of H-18 in 2a–2c
changed from ꢀ 1.20–1.21 to ꢀ 3.31–3.53 ppm due to the deshielding effect caused by
formation of the amides.
2.1.2. Synthesis of analogues of N-benzoyl-dehydroabietylamine-7-one (3a)
The oxidation of an olefin to an ꢁ,ꢂ-unsaturated ketone has been utilised in the synthesis
and in the transformation of natural products. Oxidation with transition metals such as
complexes of copper and selenium compounds is the main method used in the laboratory.
In most cases, chromium (VI) has been the oxidant and varying results have been obtained
depending upon the specific chromium (VI) reagent used and the conditions employed.
The conditions used most often have been chromium (VI) in acetic acid (Dauben, Lorber,
& Fullerton, 1969; Li & Si, 2003). Compounds 2a–2c were converted to 3a–3c via
oxidation with chromium trioxide in acetic acid. Compounds 3a–3c presented a typical
difference in their spectra from their parent compounds 2a–2c. Comparing the IR spectra
of compounds 2a and 3a, a new absorption appeared at 1685 cmꢀ1 for compound 3a,
indicating the presence of a keto group in the 7th position of ring B. In the MS spectrum of
compound 3a, the molecular ion peak was observed at m/z 404.3. The resonance of C-7
changed from ꢀ 30.53 to ꢀ 199.37 ppm by comparing the 13C-NMR spectra of 2a and 3a,
indicating the formation of carbonyl group. The IR, MS and 13C-NMR spectra of
compounds 3b and 3c had similar features.
2.1.3. Synthesis of analogues of N-benzoyl-12-nitrodehydroabietylamine (4a)
Nitration of aromatic rings is a classical organic synthesis reaction. In this study, nitration
of dehydroabietylamine derivatives was attempted using acetic anhydride–cupric nitrate
since this reagent gives low concentrations of nitronium ion in solution and is more
selective than a nitric acid–sulphuric acid mixture (Cambie & Franich, 1971). The presence
of the characteristic band of the nitro group at 1644–1638 cmꢀ1 was observed in the IR
spectra of 4a–4b. The presence of the nitro group in 4a was confirmed by the downfield
shift of H-11 and H-14 to 7.62 and 7.07 ppm, respectively, and by comparison with the
1H-NMR spectra of 2a (6.88 and 7.17 ppm, respectively) due to the deshielding effect
1
caused by formation of the nitro group. The H-NMR spectra of compounds 4b and 4c
had similar features.
2.2. Antitumour activities
The effects of the synthesised compounds (2a–2c, 3a–3c and 4a–4c) on the in vitro growth
of two human cancer cell lines PC-3 and Hey-1B were evaluated, and the results were
summarised as IC50 values in mg mLꢀ1 in Table 1.
Among these compounds, 2c, 3a, 4a, 4b and 4c showed potential activities against PC-3
and Hey-1B cancer cells. Apparently, the nitro groups at 12th and 25th positions play an

Natural Product Research
2191
Table 1. In vitro antitumour activities (IC50 mg mLꢀ1) of the
synthesised compounds.
Compounds
PC-3 IC50 (mg mLꢀ1
)
Hey-1B IC50 (mg mLꢀ1
)
2a
2b
2c
3a
3b
3c
4a
4b
4c
62.6
84.9
8.7
61.6
129.8
18.9
16.0
86.2
97.2
11.3
15.2
15.0
5.7
73.1
80.9
8.2
7.8
6.2
important role in cytotoxicity against cancer cells. Compounds 3a and 3b, with a carbonyl
group in the 7th position on ring B showed a remarkable increase in their cytotoxic
activities in comparison with their parent compounds 2a and 2b, suggesting the
importance of the carbonyl group in biological function.
3. Experimental
3.1. Chemistry
All chemicals and solvents were analytical grade, and solvents were purified by general
methods before being used. Melting points were determined with a XT6 melting point
1
apparatus and uncorrected. The H- and 13C-NMR spectra were recorded on a Bruker
DPX 300 spectrometer at 300 and 75 MHz, respectively. The chemical shifts are reported
in ppm relative to TMS. IR spectra were recorded on a Nicolet 360 FT–IR spectrometer.
Mass spectra were recorded on a Truace DSQ GC–MS. HRMS results were obtained
on an ABI MARINER–ESI–TOF. Thin layer chromatography was done using silica gel
GF254. Silica gel used for column chromatography was 100–200 mesh.
3.1.1. Synthesis of N-benzoyl-dehydroabietylamine (2a)
To a solution of dehydroabietylamine (1.43 g, 5 mmol) in toluene (50 mL) benzoyl chloride
(0.7 g, 5 mmol) was added. The mixture was stirred at room temperature for 1 h, and
heated at 80ꢁC with magnetic stirring for 8 h. The toluene was evaporated in vacuo.
The resulting crude product was purified by recrystallisation from methanol. Compound
2a was obtained as white crystals (1.38 g, 71%), m.p. 162–164ꢁC. IR (KBr): 3433, 2925,
1638 and 823. 1H-NMR (ꢀH, CDCl3, 300 MHz): 1.01(3H, s, H-19), 1.20–1.24(9H, m, H-16,
17, 20), 1.32–1.84(7H, m, H-1ꢁ, 2, 3, 6), 1.95–2.02(1H, m, H-5), 2.31(1H, brd, J ¼ 12.7 Hz,
H-1ꢂ), 2.77–2.97(3H, m, H-7,15), 3.31–3.47(2H, m, H-18), 6.10(1H, s, 22-CONH),
6.88(1H, s, H-14), 6.99(1H, d, J ¼ 8.2 Hz, H-12), 7.17(1H, d, J ¼ 8.2 Hz, H-11),
7.39–7.51(3H, m, H-24, 25, 26) and 7.71–7.74(2H, m, H-23, 27). 13C-NMR (ꢀc, CDCl3,
75 MHz): 18.75(C-19), 18.88(C-2), 19.20(C-6), 24.07(C-16 or C-17), 25.55(C-20),
30.53(C-7), 33.50(C-15), 36.48(C-3), 37.64(C-4), 37.82(C-10), 38.44(C-1), 45.90(C-5),
50.42(C-18), 123.95(C-12), 124.3(C-11), 126.97(C-14 or C-23), 126.97(C-27), 128.61(C-24
or C-26), 131.39(C-25), 134.84(C-8 or C-22), 145.65(C-13), 147.12(C-9) and 167.79(C-21).
EI–MS m/z: 390.1[M þ H]þ, 339.0, 269.1, 187.2, 173.1, 133.9. HR–ESI–MS m/z: 390.2831
([M þ H]þ, C27H36NOþ; Calcd 390.2797).

2192
Y. Chen et al.
3.1.2. Synthesis of N-(o-chlorobenzoyl)-dehydroabietylamine (2b)
Compound 2b was similarly prepared according to the procedure of 2a. Yield 72%, white
1
crystals, m.p. 146–148ꢁC. IR (KBr): 3437, 2913, 1644 and 820. H-NMR (ꢀH, CDCl3,
300 MHz): 1.03(3H, s, H-19), 1.20–1.23(9H, m, H-16, 17, 20), 1.33–1.82(7H, m, H-1ꢁ, 2, 3,
6), 1.96–2.00(1H, m, H-5), 2.30(1H, brd, J ¼ 12.8 Hz, H-1ꢂ), 2.77–2.95(3H, m, H-7,15),
3.39–3.42(2H, m, H-18), 6.21(1H, s, 22-CONH), 6.88(1H, s, H-14), 6.99(1H, d, J ¼ 8.1 Hz,
H-12), 7.16(1H, d, J ¼ 8.1 Hz, H-11), 7.28–7.38(3H, m, H-25, 26, 27), 7.64–7.66(1H, m,
H-24). 13C-NMR (ꢀc, CDCl3, 75 MHz): 18.73(C-2 or C-19), 19.16(C-6), 24.07(C-16 or
C-17), 25.42(C-20), 30.34(C-7), 33.49(C-15), 36.41(C-3), 37.59(C-4), 37.63(C-10),
38.44(C-1), 45.81(C-5), 50.77(C-18), 123.90(C-12), 124.22(C-11), 126.96(C-14 or C-26),
127.12(C-27), 130.24(C-24), 131.21(C-22 or C-25), 134.76(C-8), 135.41(C-23),
145.63(C-13), 147.10(C-9) and 166.70(C-21). EI–MS m/z: 424.2[M þ H]þ, 401.0, 382.9,
330.3, 318.3, 302.3, 274.8, 271.0, 196.3, 151.9. HR–ESI–MS m/z: 424.2516 ([M þ H]þ,
C27H35ClNOþ; Calcd 424.2407).
3.1.3. Synthesis of N-(p-nitrobenzoyl)-dehydroabietylamine (2c)
Compound 2c was similarly prepared according to the procedure of 2a. Yield 68%,
1
pale yellow crystals, m.p. 150–151ꢁC. IR (KBr): 3435, 2930, 1641 and 823. H-NMR
(ꢀH, CDCl3, 300 MHz): 1.05(3H, s, H-19), 1.23–1.25(9H, m, H-16, 17, 20), 1.33–1.82(7H,
m, H-1ꢁ, 2, 3, 6), 1.96–2.03(1H, m, H-5), 2.35(1H, brd, J ¼ 12.9 Hz, H-1ꢂ), 2.80–3.01(3H,
m, H-7, 15), 3.33–3.53(2H, m, H-18), 6.24(1H, s, 22-CONH), 6.92(1H, s, H-14),
7.01–7.03(1H, m, H-12), 7.19(1H, d, J ¼ 8.2 Hz, H-11), 7.91(2H, d, J ¼ 8.7 Hz, H-23,
27), 8.28(2H, d, J ¼ 7.8 Hz, H-24, 26). 13C-NMR (ꢀc, CDCl3, 75 MHz): 18.68(C-19),
18.84(C-2), 19.21(C-6), 24.02(C-16 or C-17), 25.45(C-20), 30.38(C-7), 33.46(C-15),
36.47(C-3), 37.59(C-4), 37.97(C-10), 38.36(C-1), 45.84(C-5), 50.78(C-18), 123.71(C-24 or
C-26), 124.00(C-12), 124.22(C-11), 126.98(C-14), 128.29(C-23 or C-27), 134.64(C-8),
140.55(C-22),145.73(C-13), 147.00(C-9), 149.40(C-25) and 166.12(C-21). EI–MS m/z:
435.3[M þ H]þ, 332.1, 302.9, 274.4, 270.5, 120.7. HR–ESI–MS m/z: 435.2625 ([M þ H]þ,
C27H35N2Oþ3 ; Calcd 435.2648).
3.1.4. Synthesis of N-benzoyl-dehydroabietylamine-7-one (3a)
A solution of CrO3 (1 g, 10 mmol) in 20 mL of acetic acid was added dropwise to a solution
of 2a (1.95 g, 5 mmol) in acetic acid. The mixture was stirred at room temperature for
30 min, and heated at 60ꢁC with magnetic stirring for 12 h. The mixture was then poured
into water and ice, extracted with chloroform. The combined organic layers were dried
over anhydrous sodium sulphate, and the solvent was evaporated under reduced pressure
(Li et al., 2003). The crude product was purified by silica gel column chromatography
using petroleum ether/acetone (3 : 1, v/v) as the eluent to give compound 3a (1.11 g, 55%)
as white needle-shaped crystals, m.p. 174–176ꢁC. IR (KBr): 3422, 2933, 1685, 1648 and
1
830. H-NMR (ꢀH, CDCl3, 300 MHz): 1.10(3H, s, H-19), 1.21–1.24(6H, m, H-16, 17),
1.28(3H, s, H-20), 1.35–1.89(5H, m, H-1ꢁ, 2, 3), 2.01–2.07(1H, m, H-5), 2.35(1H, brd,
J ¼ 12.5 Hz, H-1ꢂ), 2.67–2.80(2H, m, H-6), 2.84–2.97(1H, m, H-15), 3.18–3.55(2H, m, H-
18), 6.22(1H, s, 22-CONH), 7.27–7.49(5H, m, H-11, 12, 24, 25, 26), 7.70–7.73(2H, m, H-
23, 27), 7.83–7.84(1H, m, H-14). 13C-NMR (ꢀc, CDCl3, 75 MHz): 18.22(C-19), 18.67(C-2),
23.68(C-16 or C-17), 24.01(C-20), 33.41(C-15), 35.86(C-6), 36.11(C-3), 37.51(C-4),
37.76(C-10), 38.1(C-1), 44.42(C-5), 49.50(C-18), 123.59(C-11), 124.84(C-14), 127.06(C-23
or C-27), 128.40(C-24 or C-26), 130.4(C-12), 131.30(C-25), 132.69(C-8), 134.70(C-22),
146.58(C-13), 153.51(C-9), 168.09(C-21) and 199.37(C-7). EI–MS m/z: 404.3[M þ H]þ,

Natural Product Research
2193
390.4, 341.1, 300.8, 279.0, 244.0, 209.3, 191.9. HR–ESI–MS m/z: 404.2636 ([M þ H]þ,
C27H34NOþ2 ; Calcd 404.2590).
3.1.5. Synthesis of N-(o-chlorobenzoyl)-dehydroabietylamine-7-one (3b)
Compound 3b was similarly prepared according to the procedure of 3a. Yield 50%,
1
white crystals, m.p. 163–164ꢁC. IR (KBr): 3442, 2930, 1684, 1644 and 840. H-NMR
(ꢀH, CDCl3, 300 MHz): 1.10(3H, s, H-19), 1.21–1.24(6H, m, H-16, 17), 1.27(3H, s, H-20),
1.41–1.91(5H, m, H-1ꢁ, 2, 3), 2.04–2.10(1H, m, H-5), 2.35(1H, brd, J ¼ 12.9 Hz, H-1ꢂ),
2.53–2.76(2H, m, H-6), 2.80–2.94(1H, m, H-15), 3.20–3.51(2H, m, H-18), 6.33(1H, s,
22-CONH), 7.23–7.44(5H, m, H-11, 12, 25, 26, 27), 7.56–7.59(1H, m, H-24), 7.80–7.81
(1H, m, H-14). 13C-NMR (ꢀc, CDCl3, 75 MHz): 18.24(C-19), 18.48(C-2), 23.70(C-20),
23.84(C-16 or C-17), 33.48(C-15), 35.91(C-6), 35.99(C-3), 37.52(C-4), 37.73(C-10),
37.80(C-1), 44.47(C-5), 49.86(C-18), 123.55(C-11), 124.83(C-14), 126.88(C-26 or C-27),
129.96(C-24), 130.47(C-12), 130.93(C-22), 132.54(C-8 or C-25), 135.67(C-23),
146.60(C-13), 153.32(C-9), 167.08(C-21) and 198.90(C-7). EI–MS m/z: 438.3[M þ H]þ,
436.3, 426.3, 422.3, 411.1, 383.4, 389.7. HR–ESI–MS m/z: 438.0242 ([M þ H]þ,
C27H33ClNOþ2 ; Calcd 438.2200).
3.1.6. Synthesis of N-(p-nitrobenzoyl)-dehydroabietylamine-7-one (3c)
Compound 3c was similarly prepared according to the procedure of 3a. Yield 52%,
pale yellow crystals, m.p. 204–205ꢁC. IR (KBr): 3389, 2925, 1662, 1523, 1345 and 833.
1H-NMR (ꢀH, CDCl3, 300 MHz): 1.13(3H, s, H-19), 1.20–1.31(9H, m, H-16, 17, 20),
1.41–1.88(5H, m, H-1ꢁ, 2, 3), 2.06–2.10(1H, m, H-5), 2.39(1H, brd, J ¼ 12.8 Hz, H-1ꢂ),
2.57–2.82(2H, m, H-6), 2.85–2.92(1H, m, H-15), 3.25–3.59(2H, m, H-18), 6.49(1H, s,
22-CONH), 7.31(1H, d, J ¼ 8.8 Hz, H-11), 7.41(1H, dd, J ¼ 8.0 Hz and 2.0 Hz, H-12),
7.78–7.79(1H, m, H-14), 7.91(2H, d, J ¼ 8.8 Hz, H-23, 27), 8.24(2H, d, J ¼ 9.2 Hz, H-24,
26). 13C-NMR (ꢀc, CDCl3, 75 MHz): 18.18(C-19), 18.81(C-2), 23.62(C-16 or C-17),
23.97(C-20), 33.35(C-15), 35.86(C-6), 36.10(C-3), 37.51(C-4), 37.81(C-10), 38.23(C-1),
44.24(C-5), 49.58(C-18), 123.58(C-24 or C-26), 123.72(C-11), 124.64(C-14), 128.32(C-23 or
C-27), 130.20(C-12), 132.97(C-8), 140.28(C-22), 146.67(C-13), 149.40(C-25), 153.55(C-9),
166.16(C-21) and 199.57(C-7). EI–MS m/z: 449.3[M þ H]þ, 431.3, 367.0, 294.1, 145.9,
104.7. HR–ESI–MS m/z: 449.0154 ([M þ H]þ, C27H33N2O4þ; Calcd 449.2440).
3.1.7. Synthesis of N-benzoyl-12-nitrodehydroabietylamine (4a)
Powdered cupric nitrate trihydrate (2.42 g, 10 mmol) was added in portions to a stirred
solution of 2a (1.95 g, 5 mmol) in acetic anhydride (60 mL), and the blue solution was
stirred at 10ꢁC for 8 h. The mixture was then poured into water and ice. The pale yellow
precipitate was filtered off, washed with water, dried and recrystallised from methanol
to give 4a (1.52g, 70%) as pale yellow flakes (Cambie & Franich, 1971), m.p. 168–170ꢁC.
IR (KBr): 3424, 2931, 1648, 1524, 1349 and 825. 1H-NMR (ꢀH, CDCl3, 300 MHz):
1.00–1.02(3H, m, H-19), 1.19–1.27(9H, m, H-16, 17, 20), 1.35–1.81(7H, m, H-1ꢁ, 2, 3, 6),
1.99–2.14(1H, m, H-5), 2.29(1H, brd, J ¼ 12.3 Hz, H-1ꢂ), 2.70–3.03(3H, m, H-7, 15),
3.18–3.60(2H, m, H-18), 6.17(1H, s, 22-CONH), 7.07(1H, s, H-14), 7.39–7.52(3H, m,
H-24, 25, 26), 7.62(1H, s, H-11), 7.72–7.75(2H, m, H-23, 27). 13C-NMR (ꢀc, CDCl3,
75 MHz): 18.09(C-19), 18.40(C-2), 18.78(C-6), 23.63(C-16 or C-17), 25.28(C-20),
28.71(C-15), 30.22(C-7), 36.13(C-3), 37.68(C-4), 37.77(C-10), 38.32(C-1), 45.13(C-5),
50.23(C-18), 120.35(C-11), 126.85(C-23 or C-27), 127.99(C-14), 128.67(C-24 or C-26),
131.51(C-25), 134.82(C-22), 139.56(C-13), 141.05(C-8), 148.37(C-12), 149.06(C-9) and

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Y. Chen et al.
167.74(C-21). EI–MS m/z: 435.3[M þ H]þ, 432.1, 401.1, 294.1, 121.2, 86.8. HR–ESI–MS
m/z: 435.2759 ([M þ H]þ, C27H35N2O3þ; Calcd 435.2648).
3.1.8. Synthesis of N-(o-chlorobenzoyl)-12-nitrodehydroabietylamine (4b)
Compound 4b was similarly prepared according to the procedure of 4a. Yield 74%, pale
yellow flakes, m.p. 165–166ꢁC. IR (KBr): 3405, 2929, 1645, 1517, 1345 and 827. 1H-NMR
(ꢀH, CDCl3, 300 MHz): 1.03(3H, s, H-19), 1.19–1.27(9H, m, H-16, 17, 20), 1.35–1.86(7H,
m, H-1ꢁ, 2, 3, 6), 2.06–2.17(1H, m, H-5), 2.29(1H, brd, J ¼ 12.9 Hz, H-1ꢂ), 2.72–3.04(3H,
m, H-7, 15), 3.20–3.60(2H, m, H-18), 6.25(1H, s, 22-CONH), 7.08(1H, s, H-14), 7.28–
7.40(3H, m, H-25, 26, 27), 7.63–7.67(2H, m, H-11, 24). 13C-NMR (ꢀc, CDCl3, 75 MHz):
18.04(C-19), 18.40(C-2), 18.75(C-6), 23.61(C-16 or C-17), 25.12(C-20), 28.71(C-15),
30.05(C-7), 36.15(C-4), 37.63(C-3), 38.15(C-10), 38.33(C-1), 44.75(C-5), 50.50(C-18),
120.31(C-11), 127.12(C-26), 127.95(C-14 or C-27), 130.20(C-24), 131.24(C-22 or C-25),
135.35(C-23), 139.54(C-13), 141.17(C-8), 148.42(C-12), 149.11(C-9) and 166.80(C-21).
EI–MS m/z: 469.3[M þ H]þ, 427.3, 395.1, 218.1, 155.9, 80.2. HR–ESI–MS m/z: 468.8725
([M þ H]þ, C27H34ClN2Oþ3 ; Calcd 469.2258).
3.1.9. Synthesis of N-(p-nitrobenzoyl)-12-nitrodehydroabietylamine (4c)
Compound 4c was similarly prepared according to the procedure of 4a. Yield 69%, pale
yellow flakes, m.p. 178–180ꢁC. IR (KBr): 3435, 2928, 1661, 1523, 1348 and 829. 1H-NMR
(ꢀH, CDCl3, 300 MHz): 1.02–1.03(3H, m, H-19), 1.17–1.26(9H, m, H-16, 17, 20), 1.31–
1.87(7H, m, H-1ꢁ, 2, 3, 6), 1.99–2.11(1H, m, H-5), 2.31(1H, brd, J ¼ 12.6 Hz, H-1ꢂ),
2.63–3.05(3H, m, H-7, 15), 3.24–3.59(2H, m, H-18), 6.26(1H, s, 22-CONH), 7.16(1H, s, H-
14), 7.61(1H, s, H-11), 7.88–7.90(2H, d, J ¼ 9.0 Hz, H-23, 27), 8.24–8.27(2H, d, J ¼ 8.4 Hz,
H-24, 26). 13C-NMR (ꢀc, CDCl3, 75 MHz): 18.10(C-19), 18.36(C-2), 18.72(C-6),
23.65(C-16 or C-17), 25.25(C-20), 28.71(C-15), 30.13(C-7), 36.12(C-3), 37.67(C-4),
37.91(C-10), 38.27(C-1), 45.26(C-5), 50.58(C-18), 120.29(C-11), 123.79(C-24 or C-26),
126.31(C-14), 128.15(C-23 or C-27), 139.62(C-13), 140.42(C-22), 140.98(C-8), 148.3(C-12),
148.98(C-9), 149.46(C-25) and 165.98(C-21). EI–MS m/z: 478.3[M ꢀ H]þ, 436.4, 338.5,
227.4, 156.9. HR–ESI–MS m/z: 480.2605 ([M þ H]þ, C27H34N3O5þ; Calcd 480.2498).
3.2. Biological assays
The biological assays were performed as described by Cui, Fan, Huang, Liu, and Zhou
(2009). Hey-1B and PC-3 cells (5000 cells per well) were seeded in 96-well plates. One
day after seeding, the cells were treated with the compounds (2a–2c, 3a–3c and 4a–4c)
at various concentrations and incubated for 36 h. After removing the media, each well
was refilled with 100 uL fresh media and 10 uL MTT reagent. The cells were incubated
under the same conditions for 5 h. About 100 uL of MTT detergent was added to each
well, and plates were incubated in the dark overnight at room temperature. The OD value
was measured at 490 nm by a LD 400C Luminescence Detector.
4. Conclusions
We have prepared a series of dehydroabietylamine derivatives with a carbonyl group in the
7th position on ring B or a nitro group in the 12th position on ring C and various
benzamides substituted in the 18th position. The cytotoxicities of the synthesised
compounds against PC-3 and Hey-1B cells were investigated.
The results have demonstrated that the presence of a nitro group in the 12th position
on ring C or a carbonyl group in the 7th position on ring B were very important
in determining the biological activity of these compounds. Our findings provide new

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evidence, showing the relationship between chemical structure and biological function.
The information obtained from these studies may be useful for the design of novel
chemotherapeutic drugs for cancer (Cui et al., 2009). These results confirm that
dehydroabietylamine-type diterpenes do have interesting antitumour properties and
encourage us to research targets recognised for these diterpenes in PC-3 and Hey-1B cells;
furthermore, these results also encourage us to synthesise additional dehydroabietylamine
derivatives with the aim of obtaining compounds with more potent biological activity
(Gonzalez et al., 2010).
´
Acknowledgements
The generous financial support of the Natural Science Foundation of China (30871989 and
31170536) was gratefully acknowledged. This project was funded by the Priority Academic Program
Development of Jiangsu Higher Education Institutions.
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