Synthesis, structure analysis and cytotoxicity studies of novel unsymmetrically N,N′-substituted ureas from dehydroabietic acid

Article abstract of DOI:10.1248/cpb.56.1575

A series of novel unsymmetrically N,N′-substituted ureas were synthesized from dehydroabietic acid and their structures were characterized by IR, ¹H-NMR, ¹³C-NMR spectroscopy and single crystal X-ray diffraction. Three six-membered rings of urea 4c exhibited plane, half-chair and chair configurations, respectively. Their cytotoxicity activities against SMMC7721 liver cancer cells were evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide) method. The results showed that the title compounds exhibited highly effective cytotoxicity activities against SMMC7721 cells. Their IC₅₀ values are between 8.8 and 14.2 μmol/l. The change of N′ substituted groups resulted little difference to the cytotoxicity activities of ureas, which indicated that the cytotoxicity of this kind of ureas depend strongly on the tricyclic hydrophenanthrene structure.

Full text of DOI:10.1248/cpb.56.1575

November 2008
Chem. Pharm. Bull. 56(11) 1575—1578 (2008)
1575
Synthesis, Structure Analysis and Cytotoxicity Studies of Novel
Unsymmetrically N,Nꢀ-Substituted Ureas from Dehydroabietic Acid
a
Xiaoping RAO,*,a Zhanqian SONG,a Ling HE,b and Weihong JIA
a Institute of Chemical Industry of Forest Products, Chinese Academy of Forestry; Nanjing 210042, P. R. China: and
b Department of Pharmacology, China Pharmaceutical University; Nanjing 210009, P. R. China.
Received July 8, 2008; accepted August 12, 2008; published online August 13, 2008
A series of novel unsymmetrically N,Nꢀ-substituted ureas were synthesized from dehydroabietic acid and
their structures were characterized by IR, 1H-NMR, 13C-NMR spectroscopy and single crystal X-ray diffraction.
Three six-membered rings of urea 4c exhibited plane, half-chair and chair configurations, respectively. Their cy-
totoxicity activities against SMMC7721 liver cancer cells were evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-
2,5-diphenyltetrazolium bromide) method. The results showed that the title compounds exhibited highly effective
cytotoxicity activities against SMMC7721 cells. Their IC50 values are between 8.8 and 14.2 mmol/l. The change of
Nꢀ substituted groups resulted little difference to the cytotoxicity activities of ureas, which indicated that the cy-
totoxicity of this kind of ureas depend strongly on the tricyclic hydrophenanthrene structure.
Key words unsymmetrically N,Nꢀ-substituted urea; dehydroabietic acid; crystal structure; cytotoxicity activity
trum was recorded on a DPX-300 Bruker AVANCE 300 spectrometer
(CDCl3 as solvent).
Cancer is the primary cause of death in most of the coun-
tries and as a result there is a need for searching cancer-ef-
fective compounds.1) Natural products played an important
role in drugs discovery. They are still major source of new
antitumor drugs development, and many natural or natural
based antitumor drugs such as taxol and vinblastine were
found and clinically used in recent years.2,3)
Substituted ureas have been attracted great attention due to
a set of valuable properties allowing their use in industry,
agriculture, and medicine. Wide range of biological activities
made substituted ureas used as antitumor agents, insecti-
cides, plant growth regulators, gastroprotectives as well as
tranquillising and anticonvulsant agents. Moreover, ureas
substituted with amino acid groups have been shown to be
General Method for Preparation 4 Dehydroabietic acid (0.1 mol) dis-
solved in 30 ml CHCl3 was gradually added to a solution of 5 ml PCl3, the
reaction was performed for 3 h at 60—65 °C. Then cooled to room tempera-
ture and the mixture was filtrated and the solvent was distilled off under vac-
uum. The synthesized dehydroabietic chloride was slowly added to the
200 ml NH4OH solution, the mixture was quickly stirred for 2 h at room
temperature, and then the precipitates were filtered and crystallized from
acetone to give 2. To a 150 ml 10% NaOH solution, 12 ml Br2 and 100 ml
NaOH and 2 were added with ice cooling, the mixture was stirred for 4 h at
room temperature, then the mixture was extracted with ether three times, the
solution was dried with Na2SO4, the solvent was distilled off and 3 was ob-
tained. 0.1 mol amine was added to 20 ml CH2Cl2 solutions, the mixture was
stirred at 40 °C for 12 h, the solvents was distilled off under vacuum and
crystals were crystallized from acetone to give 4.
4a: C20H30N2O, IR (cmꢁ1): 3463, 3376 (N–H); 2950 (–CH3, –CH2); 1658
potent human immunodeficiency virus (HIV)-1 protease in- (OꢂC–N); 816 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.27—6.88
hibitors.4—6)
(3H, CꢂCH–); 4.82 (1H, –NHCO); 4.34 (2H, CO–NH2–); 2.81 (1H,
–CH(Me)2); 2.90—1.68 (10H, –CH2–); 1.46 (1H, ꢃCH–); 1.28—1.19 (12H,
Dehydroabietic acid (DHA) is a natural occurring diter-
penic resin acid, which can be easily isolated from commer-
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 158.59 (1C, CꢂO); 146.88,
145.45, 134.69, 126.78, 124.27, 123.76 (6C, CꢂC); 47.15 (1C, C–N); 33.42
cial disproportionated rosin by crystallization of the 2-
aminoethanol salt.7) DHA and its derivatives exhibited wide
range of biological activities, such as antibacterial, antifun-
gal, gastroprotective, cytotoxic, antisecretory, antipepsin and
anti-penetrant activities.8,9) DHA was reported to have prop-
erties of enhancing the inhibitory activity of an anticancer
drug in cervical carcinoma cells, hepatocellular carcinoma
cells, or breast cancer cells.10) However, the cytotoxicity ac-
tivities of unsymmetrically N,Nꢀ-substituted ureas from DHA
have not yet been reported so far. Encouraged by these re-
search results, DHA was chosen as the raw material in
screening for new potential antitumor compounds.
(1C, C–C); 56.27, 30.21 (2C, CH–C); 38.09, 37.68, 37.59, 19.67, 18.81 (5C,
CH2–C); 25.11, 23.99, 21.03 (4C, CH3).
4b: C26H34N2O, IR (cmꢁ1): 3367, 3249 (N–H); 2950 (–CH3, –CH2); 1649
(OꢂC–N); 831 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.17—6.87
(3H, CꢂCH–); 5.80 (1H, –NHCO); 4.53 (1H, CO–NH–); 2.83 (1H,
–CH(Me)2); 2.91—1.70 (10H, –CH2–); 1.34 (1H, ꢃCH–); 1.26—1.18 (12H,
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 155.60 (1C, CꢂO); 146.88,
145.51, 139.49, 134.67, 128.88, 126.87, 124.32, 123.82, 122.46, 119.77
(12C, CꢂC); 47.15 (1C, C–N); 33.54 (1C, C–C); 56.73, 30.14 (2C, CH–C);
38.12, 37.71, 37.59, 19.70, 18.97 (5C, CH2–C); 25.11, 24.12, 20.95 (4C,
CH3).
4c: C28H38N2O, IR (cmꢁ1): 3367 (N–H); 2960 (–CH3, –CH2); 1654
(OꢂC–N); 831 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 5.87 (1H,
–NHCO); 4.46 (1H, CO–NH–); 2.78 (1H, –CH(Me)2); 2.29 (6H, –Ph-CH3);
2.87—1.71 (10H, –CH2–); 1.32 (1H, ꢃCH–); 1.28—1.19 (12H, –CH3). 13C-
NMR: (CDCl3, d/ppm, 300 MHz), 154.57 (1C, CꢂO); 145.82, 144.47,
135.79, 133.52, 130.33, 129.62, 125.73, 125.62, 123.74, 123.52, 123.25,
122.81 (12C, CꢂC); 46.28 (1C, C–N); 32.44 (1C, C–C); 55.62, 29.14 (2C,
In this report, we described the synthesis a series of un-
symmetrically N,Nꢀ-substituted ureas from DHA through 4
steps of reactions. The crystal structure of 4c was deter-
mined. The cytotoxicity activity of these novel ureas was CH–C); 37.07, 36.69, 32.44, 17.88, 16.80 (5C, CH2–C); 23.98, 23.02, 22.98,
19.83, 18.65 (6C, CH3).
studied, and their structure activity relationship was also in-
vestigated.
4d: C26H33FN2O, IR (cmꢁ1): 3367 (N–H); 2960 (–CH3, –CH2); 1639
(OꢂC–N); 831 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.21—6.87
(7H, CꢂCH–); 6.57 (1H, –NHCO); 4.74 (1H, CO–NH–); 2.78 (1H,
Experimental
Materials and Methods All chemicals purchased were of reagent grade –CH(Me)2); 2.94—1.76 (10H, –CH2–); 1.30 (1H, ꢃCH–); 1.27—1.19 (12H,
and used without further purification. Infrared spectrum was recorded as
KBr pellets on a Bio-Rad FTS-185 IR spectrophotometer. Melting points
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 155.19 (1C, CꢂO); 160.48,
146.72, 145.65, 135.02, 134.39, 126. 78, 124.31, 123.94, 122.29, 122.18,
115.75, 115.46 (12C, CꢂC); 47.44 (1C, C–N); 33.47 (1C, C–C); 56.90,
1
were determined by XT5 melting point apparatus. H- and 13C-NMR spec-
∗ To whom correspondence should be addressed. e-mail: rxping2001@163.com
© 2008 Pharmaceutical Society of Japan

1576
Vol. 56, No. 11
Table 1. Synthesis of Compounds
Table 2. Crystal Data and Structure Refinement for (4c)
Compound
R1
R2
mp (°C)
Empirical formula
Formula weight
Temperature (K)
Wavelength (Å)
Crystal system
Space group
a (Å)
C28H38N2O
418.60
293(2)
0.71073
Trigonal
P65
13.0880(19)
13.0880(19)
25.097(5)
90.00
90.00
120.00
3723.0(11)
1.117
0.067
4a
4b
4c
4d
4e
4f
H
H
H
H
H
H
H
H
Ph
195.8
234.6
227.8
152.1
226.2
245.5
208.8
2,5-(CH3)2Ph
p-FPh
o-FPh
2,6-F2Ph
3,5-F2Ph
b (Å)
c (Å)
4g
a (°)
4h
244.4
b (°)
g (°)
V (Å3)
4i
240.4
Density (calculated) (mg mꢁ3
)
)
4j
4k
H
H
iso-Pr
Bu
197.5
171.6
Absorption coefficient (mmꢁ1
F (0 0 0)
1362.0
Crystal size (mm)
q range for data collection (°)
Limiting indices
0.40ꢄ0.20ꢄ0.10
1.80 to 26.00
0ꢅhꢅ13, 0ꢅkꢅ13,
ꢁ30ꢅlꢅ30
30.14 (2C, CH–C); 38.13, 37.79, 35.82, 19.64, 18.95 (5C, CH2–C); 25.01,
24.01, 20.94 (4C, CH3).
4e: C26H33FN2O, IR (cmꢁ1): 3369 (N–H); 2958 (–CH3, –CH2); 1639
(OꢂC–N); 823 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.97—6.87
(7H, CꢂCH–); 6.58 (1H, –NHCO); 4.81 (1H, CO–NH–); 2.82 (1H,
–CH(Me)2); 2.91—1.48 (10H, –CH2–); 1.34 (1H, ꢃCH–); 1.24—1.21 (12H,
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 153.81 (1C, CꢂO); 154.47,
146.74, 145.63, 134.52, 127.59, 126.83, 124.45, 123.89, 122.94, 122.84,
121.81, 114.97 (12C, CꢂC); 47.46 (1C, C–N); 33.47 (1C, C–C); 57.31,
30.18 (2C, CH–C); 38.22, 37.72, 19.67, 18.95 (5C, CH2–C); 25.07, 24.01,
23.98, 21.01 (4C, CH3).
Reflections collected/unique
[Rintꢂ0.057]
5492/4852
Completeness to qꢂ25.99 (%)
Max. and min. transmission
Data/restraints/parameters
Goodness-of-fit on F2
Final R indices [Iꢃ2s(I)]
wR2ꢂ0.327
99.9
0.9736, 0.9933
4852/4/295
1.001
R1ꢂ0.105
R indices (all data)
R1ꢂ0.1044, wR2ꢂ0.1735
ꢁ5(6)
0.31, ꢁ0.25
4f: C26H32F2N2O, IR (cmꢁ1): 3353 (N–H); 2954 (–CH3, –CH2); 1645
(OꢂC–N); 823 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.33—6.74
(7H, CꢂCH–); 6.07 (1H, –NHCO); 4.51 (1H, CO–NH–); 2.83 (1H,
–CH(Me)2); 2.93—1.69 (10H, –CH2–); 1.32 (1H, ꢃCH–); 1.25—1.21 (12H,
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 154.57 (1C, CꢂO); 163.01,
159.78, 146.72, 145.62, 138.04, 134.45, 131.19, 126.79, 124.28, 123.90,
Absolute structure parameter
Largest diff. peak and hole (e·Aꢁ3
)
R1ꢂ∑||Fo|ꢁ|Fc||/∑|Fo|; wR2ꢂ∑[w(Fo2ꢁFc2)2/∑[w(Fo )2]1/2
.
2
116.38, 110.44 (12C, CꢂC); 47.31 (1C, C–N); 33.46 (1C, C–C); 57.12, 18.85 (5C, CH2–C); 25.07, 24.04, 23.98, 23.39, 23.32, 21.29 (6C, CH3).
30.13 (2C, CH–C); 38.17, 37.69, 19.66, 18.98 (5C, CH2–C); 25.02, 24.01,
23.98, 20.99 (4C, CH3).
4k: C25H39N2O, IR (cmꢁ1): 3326 (N–H); 2953 (–CH3, –CH2); 1633
(OꢂC–N); 819 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.14—6.84
(3H, CꢂCH–); 4.32 (1H, –NHCO); 3.78 (1H, –NHCO); 2.82 (1H,
4g: C26H32F2N2O, IR (cmꢁ1): 3362 (N–H); 2955 (–CH3, –CH2); 1628
(OꢂC–N); 826 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.28—6.86 –CH(Me)2); 2.87—1.46 (16H, –CH2–); 1.44 (1H, ꢃCH–); 1.45—1.08 (15H,
(6H, CꢂCH–); 6.65 (1H, –NHCO); 4.87 (1H, CO–NH–); 2.81 (1H,
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 157.27 (1C, CꢂO); 146.06,
–CH(Me)2); 2.93—1.68 (10H, –CH2–); 1.34 (1H, ꢃCH–); 1.31—1.19 (12H, 144.32, 133.75, 125.77, 123.26, 122.69 (6C, CꢂC); 46.26 (1C, C–N); 32.45
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 155.11 (1C, CꢂO); 146.79, (1C, C–C); 55.03, 31.55 (3C, CH–C); 38.56, 37.14, 36.90, 36.82, 29.25,
145.54, 139.24, 134.53, 129.02, 126.79, 124.29, 123.84, 122.93, 120.28
19.07, 18.83, 17.85 (8C, CH2–C); 24.11, 22.99, 20.28,12.84 (5C, CH3).
(12C, CꢂC); 47.45 (1C, C–N); 33.47 (1C, C–C); 56.84, 30.15 (2C, CH–C);
X-Ray Crystallography The crystal structure of 4c was determined by
38.13, 37.77, 37.70, 19.65, 18.95 (5C, CH2–C); 25.04, 24.03, 23.99, 20.88 X-ray single crystal diffraction. XRD data were collected on a Enraf-Nonius
(4C, CH3).
CAD-4 diffractometer equipped with MoKa (lꢂ0.07103) at 293 K. A sin-
4h: C25H38N2O, IR (cmꢁ1): 3358 (N–H); 2955 (–CH3, –CH2); 1626
gle crystal suitable for determination was mounted inside a glass fiber capil-
(OꢂC–N); 1529 (CꢂC); 832 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), lary. The structure of the title compound was solved by direct methods and
7.183—6.875 (3H, CꢂCH–); 4.203 (4H, CONCH2–); 3.682 (4H, OCH2–);
3.290 (4H, NCH2–); 3.237 (1H, –CH(Me)2); 2.829—1.725 (10H, –CH2–);
1.357 (1H, ꢃCH–); 1.278—1.222 (12H, –CH3). 13C-NMR: (CDCl3, d/ppm,
refined by full-matrix least squares on F2. All the hydrogen atoms were
added in their calculated positions and all the non-hydrogen atoms were re-
fined with anisotropic temperature factors. SHELXS97 were used to solve
300 MHz), 156.73 (1C, CꢂO); 146.93, 145.49, 134.45, 126.70, 124.14, the structure and SHELTL were used to refine the structure.11,12) The crystal-
123.76 (6C, CꢂC); 47.04 (1C, C–N); 33.39 (1C, C–C); 56.72, 30.12 (2C, lographic details are summarized in Table 2. The selected bond lengths and
CH–C); 47.04, 45.15, 44.53, 38.09, 37.79, 37.61, 19.72, 18.84 (10C, angles are shown in Table 3.
CH2–C); 23.90, 22.22, 21.37 (4C, CH3).
Pharmacology Test compounds 4a—c were evaluated in vitro, for cyto-
toxicity in SMMC7721 liver cancer cell lines at 1, 10, 15, 25 and 50 mmol/l
4i: C24H36N2O2, IR (cmꢁ1): 3358 (N–H); 2955 (–CH3, –CH2); 1626
(OꢂC–N); 832 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.18—6.87 concentrations using MTT assay, respectively.13)
(3H, CꢂCH–); 4.20 (4H, CONCH2–); 3.68 (4H, O–CH2–); 3.29 (4H,
NCH2–); 3.23 (1H, –CH(Me)2); 2.82—1.72 (10H, –CH2–); 1.35 (1H,
ꢃCH–); 1.28—1.22 (12H, –CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz),
Results and Discussion
Synthesis As shown in Chart 1, a series of unsymmetri-
cally N,Nꢀ-substituted ureas were synthesized from DHA
through 4 steps of reactions. Although high steric hindrance
156.71 (1C, CꢂO); 146.85, 145.64, 134.40, 126.78, 124.24, 123.89 (6C,
CꢂC); 47.41 (1C, C–N); 33.46 (1C, C–C); 57.02, 30.17 (2C, CH–C); 66.53,
44.24, 43.45, 38.19, 37.78, 19.73, 18.94 (9C, CH2–C); 24.92, 24.03, 21.19
(4C, CH3).
4j: C23H36N2O, IR (cmꢁ1): 3356 (N–H); 2961 (–CH3, –CH2); 1633 of tricycle hydrophethane structure to the carboxylic acid (1),
(OꢂC–N); 826 (Ar-H); 1H-NMR: (CDCl3, d/ppm, 300 MHz), 7.15—6.85
the reaction activity was greatly improved by conversion of
the acid group to chloride intermediate. Previous study re-
vealed that PCl3 is the best chloride reagent. (2) can be ob-
tained from chloride in the presence of NH4OH at room tem-
perature. Through Hoffman degrading reaction, isocyanate
(3H, CꢂCH–); 4.30 (1H, –NHCO); 3.06 (1H, –NHCO); 2.79 (1H,
–CH(Me)2); 2.86—1.67 (10H, –CH2–); 1.43 (2H, ꢃCH–); 1.31—0.88 (18H,
–CH3). 13C-NMR: (CDCl3, d/ppm, 300 MHz), 157.40 (1C, CꢂO); 147.06,
145.35, 134.73, 126.77, 124.30, 123.71 (6C, CꢂC); 47.23 (1C, C–N); 33.46
(1C, C–C); 56.15, 41.48, 30.31 (3C, CH–C); 38.15, 37.94, 37.85, 19.81,

November 2008
1577
(3) can be obtained. Isocyanate can be converted to unsym- spectively. The data of inhibition ratios are listed in Table 4.
metrically N,Nꢀ-substituted ureas (4) with substituted amine. As can be concluded from the results, the inhibition ratios of
Structure Analysis Full crystallographic details of 4c all compounds increased with the increment of concentra-
have been deposited at the Cambridge Crystallographic Data tions, they reached very high activity at the concentration of
Center and allocated the deposition number CCDC 683542. 25 mM, the inhibition ratios reached about 66—82%, espe-
White crystals of 4c suitable for X-ray analysis were ob- cially compounds 4a, 4d, 4g and 4h exhibited higher activi-
tained by solvent evaporation under room temperature. Its ties than others, the inhibition ratios are near 80%.
single-crystal structure was determined by X-ray crystallog-
Detailed bioassays were carried out for compounds 4a, 4d,
raphy. As shown in Fig. 1, the molecular structure of 4c con- 4g and 4h at sample concentrations of 1.0, 10, 15, 25, 50 mM,
tains four crystallographically unique six-membered rings. respectively. The inhibition ratios and IC50 values of these
Their torsion angles show ring B (C6, C7, C10—C13) and C compounds are listed in Table 5. The compounds exhibited
(C10, C11, C14—C17) exhibits a chair and half-chair config- high activity at 50 mM, the inhibition ratios reached between
uration, respectively, they form trans ring junction with two 90% and 99.1%, IC50 values are between 8.8 and 14.2. Com-
methyl groups (C18, C19) in axis positions. However, the six pound 4a exhibited highest activity against SMMC7721 liver
atoms in the other two six-membered rings are coplanar. cancer cells with IC50 value of 8.8 mM. The title compound is
From the view of the crystal packing (Fig. 2), it is found that
weak inter-molecular p ···p stacking interactions exist in the
structure.
Cytotoxicity Activity In order to investigate whether the
synthesized unsymmetrically N,Nꢀ-substituted ureas inhibit
the growth of tumor cells, SMMC7721 liver cancer cells
were selected as tumor agent. All compounds were subjected
to a preliminary screening for cytotoxicity activity at sample
concentrations of 1.0, 10 and 25 mM using the MTT assay, re-
Table 3. Selected Bond Lengths and Angles for (4c)
Bond length [Å]
O–C20
1.269(9)
N1–C20
N1–C17
N2–C20
N2–C21
1.323(10)
1.464(10)
1.365(10)
1.402(11)
Fig. 1. ORTEP Diagram of Compound 4c with H Atoms Represented by
Small Spheres of Arbitrary Radius
Bond angles [°]
C20–N1–C17
C20–N2–C21
C6–C5–C4
C8–C13–C12
N1–C17–C16
N1–C17–C11
C16–C17–C11
N1–C17–C19
O–C20–N1
127.3(7)
127.9(7)
120.3(11)
115.2(8)
109.7(7)
108.4(6)
109.5(6)
103.3(7)
124.2(8)
120.0(7)
115.7(7)
119.9(9)
119.3(9)
121.8(10)
O–C20–N2
N1–C20–N2
C22–C21–N2
N2–C21–C26
C21–C22–C27
Torison angles [°]
C5–C6–C7–C10
C20–N1–C17–C16
C15–C16–C17–N1
C17–N1–C20–O
C17–N1–C20–N2
C21–N2–C20–O
C21–N2–C20–N1
N2–C21–C26–C25
ꢁ178.3(10)
ꢁ54.7(11)
169.7(7)
ꢁ7.6(15)
176.3(7)
6.5(14)
ꢁ177.2(8)
173.2(8)
Fig. 2. The Packing Diagram of Compound 4c in the Unit Cell
Chart 1. Synthetic Scheme of Unsymmetrically N,Nꢀ-Substituted Ureas

1578
Vol. 56, No. 11
Table 4. Inhibition Ratios of Compounds against SMMC7721 Liver Can-
cer Cells at Different Concentration
nism as TBIDOM.
Conclusions
Concentration (mM)
In summary, we have synthesized a series of unsymmetri-
cally N,Nꢀ-substituted ureas from dehydroabietic acid and
their structures were identified by IR, 1H-NMR spectroscopy
and single crystal X-ray diffraction. The tricyclic hy-
drophenanthrene structure of 4c exhibited classic configura-
tion. Three six-membered rings of molecule 4c exhibited
plane, half-chair and chair configurations, respectively. This
kind of compounds will be a new class of antitumor agents
because of its strong indication of cytotoxicity activities and
natural based properties. The change of Nꢀ substituted
groups resulted little difference to the cytotoxicity activities
of ureas, which indicated that the cytotoxicity of this kind of
ureas depend strongly on the tricyclic hydrophenanthrene
structure.
Compounds
1.0
10
25
4a
4b
4c
4d
4e
4f
4g
4h
4i
3.4
0
0
2.5
0
0
3.4
3.4
0
0
0
34.7
16.9
19.5
17.8
0
81.3
74.6
73.7
77.1
68.6
69.5
76.3
78.8
66.9
67.8
70.3
0
20.3
25.4
0
0
0
4j
4k
Table 5. Inhibition Ratios of Compounds against SMMC7721 Liver Can-
cer Cells at Different Concentration
Acknowledgement This research was financially supported by grants
from Forestry Commonwealth Industry Special Foundation of China (No.
200704008) and National Natural Science Foundation of China (No.
30771690).
Concentration (mM)
Compounds
IC50
1.0
10
15
25
50
References
4a
4d
4g
4h
3.4
2.5
3.4
3.4
34.7
17.8
20.3
25.4
61.1
50.5
58.4
59.2
81.3
77.1
76.3
78.8
99.1
90.4
92.3
98.1
8.8
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According to the structure and cytotoxicity activity of
these compounds, change of the substituted groups of R1 and
R2 can little improve the cytotoxicity activity, which indicat-
ing that the cytotoxicity activity of this kind of ureas depend
strongly on the tricyclic hydrophenanthrene structure.
Possible Mechanism In our previous study,14) we have
explored the possible mechanisms of dehydroabietic de-
rivaties (TBIDOM) on SMMC-7721 liver cancer cells. Pre-
treatment of SMMC-7721 cells with TBIDOM significantly
induced a decrease of Bcl-2 protein expression and an in-
crease of caspase-3 activity and Bax protein expression. Its
mechanism may have been related to the decrease in the ex-
pression of anti-apoptotic protein, Bcl-2, accompanied by a
drop in the mitochondrial membrane potential and the activa-
tion of caspase-3, that led to apoptotic body formation and fi-
nally apoptosis. We think the ureas possess the same mecha-