Synthesis and antitumor, antityrosinase, and antioxidant activities of xanthone

Article abstract of DOI:10.1080/10286020.2018.1454437

Ten substituted 1,3-dihydroxyxanthones were synthesized in one step. The yields ranged from 40 to 76%. Compounds 8–10 were first reported. Next, the compounds’ in vitro anti-proliferative activities against nine human cancer cell lines, antityrosinase, and antioxidant activities were evaluated. Compounds 1, 4, 6–7, and 9–10 exhibited enhanced cytotoxicity against certain cancer cells. Compounds 2, 8, 9, and 10 inhibited tyrosinase activity to a certain extent. In addition, compound 4 exhibited the best antioxidant activity, which was consistent with theoretical calculations. These results demonstrated that compounds 1–2, 4, and 6–10 were promising leads for further investigation.

Full text of DOI:10.1080/10286020.2018.1454437

Journal of Asian Natural Products Research
ISSN: 1028-6020 (Print) 1477-2213 (Online) Journal homepage: http://www.tandfonline.com/loi/ganp20
Synthesis and antitumor, antityrosinase, and
antioxidant activities of xanthone
Bei-Dou Zhou, Li-Lan Zeng, Yu-Gui Tong, Jia-Ying Fang, Zhi-Peng Ruan, Xiao-
Yun Zeng, Yuan-Yuan Fang, Gui-Fen Xu & Dong-Bao Hu
To cite this article: Bei-Dou Zhou, Li-Lan Zeng, Yu-Gui Tong, Jia-Ying Fang, Zhi-Peng Ruan,
Xiao-Yun Zeng, Yuan-Yuan Fang, Gui-Fen Xu & Dong-Bao Hu (2018): Synthesis and antitumor,
antityrosinase, and antioxidant activities of xanthone, Journal of Asian Natural Products Research,
DOI: 10.1080/10286020.2018.1454437
To link to this article: https://doi.org/10.1080/10286020.2018.1454437
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Journal of asian natural Products research, 2018
ꢁꢁpꢂ://ꢃꢄꢅ.ꢄꢆg/10.1080/10286020.2018.1454437
ꢀ
Synthesis and antitumor, antityrosinase, and antioxidant
activities of xanthone
Bei-Dou Zhou , Li-Lan Zeng , Yu-Gui Tong , Jia-Ying Fang , Zhi-Peng Ruana,b,
a,b
c
d
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f
Xiao-Yun Zeng , Yuan-Yuan Fang , Gui-Fen Xu and Dong-Bao Hu
ꢇ
b
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Pꢀꢇꢆmꢇꢈꢌꢍꢁꢅꢈꢇꢉ aꢋꢇꢉyꢂꢅꢂ ꢇꢋꢃ lꢇbꢄꢆꢇꢁꢄꢆy Mꢌꢃꢅꢈꢅꢋꢌ (Pꢍꢁꢅꢇꢋ uꢋꢅvꢌꢆꢂꢅꢁy), fꢍjꢅꢇꢋ Pꢆꢄvꢅꢋꢈꢌ uꢋꢅvꢌꢆꢂꢅꢁy, Pꢍꢁꢅꢇꢋ
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sꢀꢇꢋꢁꢄꢍ 515041, cꢀꢅꢋꢇ; sꢈꢀꢄꢄꢉ ꢄꢊ cꢀꢌmꢅꢈꢇꢉ Bꢅꢄꢉꢄgy ꢇꢋꢃ eꢋvꢅꢆꢄꢋmꢌꢋꢁ, Yꢍxꢅ nꢄꢆmꢇꢉ uꢋꢅvꢌꢆꢂꢅꢁy, Yꢍxꢅ 653100,
ꢊ
cꢀꢅꢋꢇ
ABSTRACT
ARTICLE HISTORY
Ten substituted 1,3-dihydroxyxanthones were synthesized in one
step. The yields ranged from 40 to 76%. Compounds 8–10 were first
reported. Next, the compounds’ in vitro anti-proliferative activities
against nine human cancer cell lines, antityrosinase, and antioxidant
activities were evaluated. Compounds 1, 4, 6–7, and 9–10 exhibited
enhanced cytotoxicity against certain cancer cells. Compounds 2, 8,
rꢌꢈꢌꢅvꢌꢃ 30 nꢄvꢌmbꢌꢆ 2017
aꢈꢈꢌpꢁꢌꢃ 14 Mꢇꢆꢈꢀ 2018
KEYWORDS
Xꢇꢋꢁꢀꢄꢋꢌ; ꢂyꢋꢁꢀꢌꢂꢅꢂ;
ꢇꢋꢁꢅꢁꢍmꢄꢆ ꢇꢈꢁꢅvꢅꢁy;
ꢇꢋꢁꢅꢁyꢆꢄꢂꢅꢋꢇꢂꢌ; ꢇꢋꢁꢅꢄxꢅꢃꢇꢁꢅꢄꢋ
9
, and 10 inhibited tyrosinase activity to a certain extent. In addition,
compound 4 exhibited the best antioxidant activity, which was
consistent with theoretical calculations. These results demonstrated
that compounds 1–2, 4, and 6–10 were promising leads for further
investigation.
1
. Introduction
Xanthone is a secondary metabolite isolated from certain plants and micro-organisms.
is type of compound has many important biological activities, such as antitumor [1],
antioxidant [2], antibacterial [3], antihypertensive [4], and antiplatelet aggregation activities
5]. e different activities are determined by the different substituents on xanthone’s core

[
structure. For example, the anti-proliferative activity of xanthone is substantially influenced
by the position and number of attached hydroxyl and prenyl groups [6]. Xanthones, such
CONTACT dꢄꢋg-Bꢇꢄ hꢍ
ꢀꢍꢃꢄꢋgbꢇꢄ@126.ꢈꢄm
tꢀꢌ ꢂꢍppꢉꢌmꢌꢋꢁꢇꢉ ꢃꢇꢁꢇ ꢊꢄꢆ ꢁꢀꢅꢂ ꢇꢆꢁꢅꢈꢉꢌ ꢈꢇꢋ bꢌ ꢇꢈꢈꢌꢂꢂꢌꢃ ꢇꢁ ꢀꢁꢁpꢂ://ꢃꢄꢅ.ꢄꢆg/10.1080/10286020.2018.1454437.
©
2018 iꢋꢊꢄꢆmꢇ uK lꢅmꢅꢁꢌꢃ, ꢁꢆꢇꢃꢅꢋg ꢇꢂ tꢇyꢉꢄꢆ & fꢆꢇꢋꢈꢅꢂ Gꢆꢄꢍp

2
B.ꢀD. ZHOU ET AL.
1
2
3
7
8
4
9
1
0
5
6
Scheme 1. syꢋꢁꢀꢌꢁꢅꢈ ꢆꢄꢍꢁꢌ ꢄꢊ ꢈꢄmpꢄꢍꢋꢃꢂ 1–10.
as 1,3-dihydroxy-6-methoxyxanthone, could be considered as new potential anticancer
agents for the selective treatment of MRP1-positive tumors [7].
1
,3-Dihydroxyxanthone (1) was of en used as a good lead compound for drug design.
A series of 1-hydroxyl-3-aminoalkoxy xanthone derivatives were synthesized, and most
of them exhibited effective cytotoxic activity against the four tested cancer cell lines [8].
1
,3,7-Trihydroxyxanthone (2) inhibited the growth of paclitaxel-resistant SMMC-7721
cells to a certain extent [9]. 1,3,8-Trihydroxyxanthone (3) displayed a certain degree of
antitumor activity against human HepG2 cells [6]. 1,3,5,6-Tetrahydroxyxanthone (4) exhib-
ited more potent antioxidant activity in a DPPH assay compared with ascorbic acid [10].
6
-Chloro-1,3-dihydroxyxanthone (5) was used as a lead compound for the treatment of
depression [11]. 7-Chloro-1,3-dihydroxyxanthone (6) was confirmed as the most active
agent against five tested cancer cell lines [12]. us, hydroxyl and chlorine are commonly
substituted in 1,3-dihydroxyxanthone. In addition to the two groups, nitro, trifluoromethyl,
and 2,4-difluorophenyl are also commonly used for structural modification. In particular,
fluorine-substituted group of en results in a significant improvement in the compound’s
biological activity.
e xanthone ring and phenolic hydroxyl group exhibit antitumor and antioxidant activ-
ities, respectively. Aromatics containing two metaphenolic hydroxyl groups may exhibit
antityrosinase activity. To further study substituted 1,3-dihydroxyxanthone, 10 compounds
were synthesized. Next, the compounds’ antitumor, antityrosinase, and antioxidant activi-
ties were determined. In addition, the antioxidant capacity was analyzed by computational
chemistry.
2
. Results and discussion
2
.1. Synthesis
Compounds 1–10 were synthesized by the corresponding salicylic acid and phloroglucinol
in phosphorus pentoxide-methanesulfonic acid (Eaton’s reagent) at 90 °C, according to
the reported method (Scheme 1) [13,14]. Better yields were found in this step (40–76%).

JOURNAL OF ASIAN NATURAL PRODUCTS RESEARCH
3
Table 1. tꢀꢌ ꢅꢋꢀꢅbꢅꢁꢄꢆy ꢇꢈꢁꢅvꢅꢁꢅꢌꢂ ꢄꢊ ꢈꢄmpꢄꢍꢋꢃꢂ 1–10 ꢇgꢇꢅꢋꢂꢁ fivꢌ ꢁꢍmꢄꢆ ꢈꢌꢉꢉ ꢉꢅꢋꢌꢂ.
Conc
Cell inhibition (%)
Compd
(μM)
HL-60
A-549
SMMC-7721
MCF-7
SW480
1
2
3
4
5
6
7
8
9
40
40
40
40
40
40
40
40
40
40
40
5
2.84 ± 1.ꢎ7
12.08 ± 3.05
−21.88 ± 5.3ꢎ
25.86 ± 1.75
4.20 ± 2.24
10.70 ± 0.81
3.33 ±3.51
−33.55 ± 0.44
51.43 ± 0.4ꢎ
20.16 ± 4.07
ꢎ4.76
2.ꢎ2 ± 1.5ꢎ
2.44 ± 1.63
25.85 ± 2.13
11.82 ± 0.54
ꢎ.20 ± 3.44
3.30 ± 3.77
0.15 ± 0.80
8.51 ± 1.52
65.57 ± 2.00
72.10 ± 2.8ꢎ
53.42
61.70 ± 0.73
3ꢎ.36 ± 1.30
3ꢎ.73 ± 0.8ꢎ
10.32 ± 0.55
2ꢎ.ꢎ1 ± 1.54
ꢎ8.7ꢎ ± 0.26
ꢎ0.17 ± 0.41
1ꢎ.20 ± 2.8ꢎ
57.72 ± 1.20
51.12 ± 0.67
61.ꢎ5
4.18 ± 1.78
12.55 ± 0.46
33.35 ± 1.33
2.34 ± 2.67
0.35 ± 0.21
ꢎ.76 ± 0.16
ꢎ.17 ± 1.63
10.73 ± 0.ꢎꢎ
31.82 ± 0.80
35.88 ± 0.27
52.ꢎ7
1.ꢎ5 ± 0.48
4.52 ± 0.23
21.52 ± 1.20
13.52 ± 0.45
7.32 ± 1.07
3.40 ± 0.2ꢎ
8.4ꢎ ± 0.44
0.00 ± 0.ꢎꢎ
14.ꢎ2 ± 2.10
28.38 ± 2.24
54.00
1
0
cꢅꢂpꢉꢇꢁꢅꢋ
tꢇxꢄꢉ
ꢎ6.65
64.78
88.23
5ꢎ.10
67.00
Table 2. tꢀꢌ ꢅꢋꢀꢅbꢅꢁꢄꢆy ꢇꢈꢁꢅvꢅꢁꢅꢌꢂ ꢄꢊ ꢈꢄmpꢄꢍꢋꢃꢂ 1–10 ꢇgꢇꢅꢋꢂꢁ ꢁꢀꢌ ꢄꢁꢀꢌꢆ ꢊꢄꢍꢆ ꢁꢍmꢄꢆ ꢈꢌꢉꢉ ꢉꢅꢋꢌꢂ.
Conc
Cell inhibition (%)
Compd
(μM)
HeLa
HepG2
Huh-7
MT-4
1
2
3
4
5
6
7
8
9
40
40
40
40
40
40
40
40
40
40
40
5
38.ꢎ4 ± 0.ꢎ0
23.38 ± 0.44
23.51 ± 2.78
0.00 ± 0.75
11.64 ± 2.18
63.85 ± 0.85
51.20 ± 0.37
ꢎ.ꢎ1 ± 2.48
43.07 ± 1.25
2ꢎ.37 ± 1.37
ꢎ6.30
5.80 ± 1.08
14.0ꢎ ± 1.05
26.75 ± 3.18
4.35 ± 1.24
2.85 ± 0.77
17.48 ± 1.1ꢎ
11.27 ± 1.18
1.64 ± 1.76
4.88 ± 1.61
34.2ꢎ ± 1.7ꢎ
80.6ꢎ
3.6ꢎ ± 1.78
4.63 ± 0.81
11.52 ± 3.3ꢎ
36.04 ± 1.38
2.47 ± 1.1ꢎ
5.57 ± 1.63
20.52 ± 1.65
33.ꢎ4 ± 2.45
21.3ꢎ ± 1.50
10.26 ± 1.ꢎꢎ
ꢎ0.17
1ꢎ.22 ± 3.08
25.16 ± 1.55
4.3ꢎ ± 4.07
ꢎ1.18 ± 0.42
17.02 ± 2.4ꢎ
28.11 ± 2.37
22.57 ± 1.8ꢎ
10.61 ± 1.70
60.74 ± 0.46
81.70 ± 1.50
ꢎꢎ.31
1
0
cꢅꢂpꢉꢇꢁꢅꢋ
tꢇxꢄꢉ
67.38
41.62
57.6ꢎ
ꢎ8.74
2
.2. Biological activity
2
.2.1. Antitumor activity
e in vitro cytotoxic activity of all compounds was assessed in nine human cancer cell
lines, including a promyelocytic leukemia cell line (HL-60), lung cancer cell line (A-549),
hepatocarcinoma cell line (SMMC-7721), breast cancer cell line (MCF-7), colon cancer cell
line (SW480), cervical carcinoma cell line (HeLa), hepatoma cell lines (HepG2 and Huh-7),
and T-cell leukemia cell line (MT-4) using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carbox-
ymethoxyphenyl)- 2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay [15]. Two
anticancer drugs, cisplatin and Taxol, were co-assayed as the positive control. e results
are listed in Tables 1 and 2. e inhibition rates of the compounds were greater than 50%,
and the IC50 values (concentration required to inhibit tumor cell proliferation by 50%)
relative to corresponding tumor cell lines were determined subsequently. ese results are
listed in Table 3.
Six compounds exhibited obvious antitumor activity. Compound 9 exhibited moderate
cytotoxicity against HL-60 cells with an inhibition rate of 51.4%. Compounds 9 and 10 dis-
played better cytotoxicity against A-549 cells with inhibition rates of 65.6 and 72.1%, respec-
tively. e IC50 values of compounds 9 and 10 against A-549 cells were reduced compared
with cisplatin, which is widely used in clinical practice. Compounds 1 and 9–10 exhibited

4
B.ꢀD. ZHOU ET AL.
Table 3. ic50 vꢇꢉꢍꢌꢂ ꢄꢊ ꢈꢄmpꢄꢍꢋꢃꢂ 1, 4, 6–7, ꢇꢋꢃ 9–10 ꢇgꢇꢅꢋꢂꢁ ꢈꢄꢆꢆꢌꢂpꢄꢋꢃꢅꢋg ꢁꢍmꢄꢆ ꢈꢌꢉꢉ ꢉꢅꢋꢌꢂ.
IC50 (μM)
Compd
HL-60
A-549
SMMC-7721
HeLa
MT-4
1
4
6
7
9
–
–
–
–
–
–
–
–
2ꢎ.85 ± 0.56
–
–
–
–
13.73 ± 1.05
36.55 ± 0.6ꢎ
–
ꢎ.05 ± 0.40
–
6.14 ± 0.13
14.02 ± 0.37
27.16 ± 0.47
37.42 ± 1.37
26.26 ± 1.45
<0.008
–
38.12 ± 0.5ꢎ
–
6.ꢎ5 ± 0.38
<0.008
2ꢎ.ꢎ3 ± 0.47
24.8ꢎ ± 1.11
32.3ꢎ ± 0.54
<0.008
21.27 ± 1.48
18.55 ± 0.3ꢎ
0.52 ± 0.01
<0.008
1
0
–
cꢅꢂpꢉꢇꢁꢅꢋ
tꢇxꢄꢉ
5.77 ± 0.35
<0.008
nꢄꢁꢌ: “–”mꢌꢇꢋꢂ ꢋꢄꢁ bꢌꢅꢋg ꢃꢌꢁꢌꢆmꢅꢋꢇꢁꢌꢃ.
Table 4. aꢋꢁꢅꢁyꢆꢄꢂꢅꢋꢇꢂꢌ ꢇꢈꢁꢅvꢅꢁꢅꢌꢂ ꢄꢊ ꢈꢄmpꢄꢍꢋꢃꢂ 1–10.
Compd
Conc (g/ml)
Inhibition rate (%)
Compd
Conc (g/ml)
Inhibition rate (%)
kꢄjꢅꢈ ꢇꢈꢅꢃ
10
50
50
50
50
50
62.ꢎ2 ± 0.58
−15.21 ± 3.28
13.32 ± 1.72
1.15 ± 0.12
−17.34 ±1.38
5.40 ± 1.15
6
7
50
50
50
50
50
−0.58 ± 0.46
−3.33 ± 1.38
14.24 ± 2.64
11.25 ± 1.4ꢎ
1ꢎ.2ꢎ ± 0.12
1
2
3
4
5
8
9
10
moderate cytotoxicity against SMMC-7721 cells with inhibition rates ranging from 51.1 to
1.7%. Moreover, compounds 6 and 7 exhibited good cytotoxicity against SMMC-7721 cells
with inhibition rates of 98.8 and 90.2%, respectively. e IC values of compounds 6 and 7
6
5
0
against SMMC-7721 cells were also less than those for cisplatin. In particular, compound
exhibited strong cytotoxicity against SMMC-7721 cells (IC50 < 10 μM). Compounds 6
and 7 exhibited moderate cytotoxicity against HeLa cells with inhibition rates of 63.9 and
1.2%, respectively. In addition, compound 4 exhibited strong cytotoxicity against MT-4
cells with an inhibition rate of 91.2%.
6
5
2

.2.2. Antityrosinase activity
e results obtained from antityrosinase activity assay are presented in Table 4. All com-
pounds were compared with kojic acid. Compounds 2 and 8–10 inhibit tyrosinase activity
to a certain extent (>10%). Compound 5 exhibited minimal antityrosinase activity, whereas
the other compounds were inactive.
2
.2.3. Antioxidant activity
2
.2.3.1. DPPH radical scavenging activity. e antioxidant activity assay results are
presented in Table 5. All compounds were compared with trolox. Compound 4 exhibited
the best antioxidant activity, but its antioxidant activity was weaker than trolox. Its two
ortho-hydroxyl groups potentially played an important role in scavenging free radicals.
However, the other compounds did not exhibit antioxidant activity.
2
.2.3.2. Calculation of antioxidant activity. e antioxidant activity of compounds 1–10
was also studied using the density functional theory (DFT) B3LYP method with 6–311 + +
G (d, p) basis set (Table 6). Antioxidant activities were analyzed based on phenolic hydroxyl

JOURNAL OF ASIAN NATURAL PRODUCTS RESEARCH
5
Table 5. sꢈꢆꢌꢌꢋꢅꢋg ꢆꢌꢂꢍꢉꢁꢂ ꢄꢊ dPPh ꢆꢇꢃꢅꢈꢇꢉ ꢂꢈꢇvꢌꢋgꢅꢋg ꢇꢈꢁꢅvꢅꢁꢅꢌꢂ ꢄꢊ ꢈꢄmpꢄꢍꢋꢃꢂ 1–10.
Compd
ꢆꢄꢉꢄx
1
2
3
4
5
Conc (g/ml)
Antioxidant rate (%)
Compd
Conc (g/ml)
Antioxidant rate (%)
ꢁ
25
50
50
50
50
50
ꢎ5.87 ± 0.26
1.82 ± 3.34
−0.42 ± 0.60
−0.060 ± 2.2ꢎ
47.13 ± 1.78
−1.26 ± 0.085
6
7
8
9
50
50
50
50
50
−2.10 ± 0.085
1.56 ± 1.53
1.ꢎ8 ± 0.26
0.06 ± 1.27
−2.56 ± 0.085
10
Table 6. eꢋꢌꢆgy ꢄꢊ ꢆꢇꢃꢅꢈꢇꢉ ꢇꢋꢃ Bde ꢄꢊ pꢀꢌꢋꢄꢉꢅꢈ ꢀyꢃꢆꢄxyꢉ ꢀyꢃꢆꢄgꢌꢋ ꢇꢁꢄmꢂ ꢅꢋ ꢄpꢁꢅmꢅzꢌꢃ ꢂꢁꢆꢍꢈꢁꢍꢆꢌ ꢄꢊ ꢈꢄm-
pꢄꢍꢋꢃꢂ 1–10.
Compd
Radical
Energy (hartree)
BDE (kJ/mol)
1
pꢇꢆꢌꢋꢁ
−801.320ꢎ686
−800.6700741
−800.66ꢎ4325
−876.5678327
−875.ꢎ176616
−875.ꢎ164876
−875.ꢎ2351ꢎ4
−876.5816776
−875.ꢎ105336
−875.ꢎ288505
−875.ꢎ16174
−ꢎ51.8126361
−ꢎ51.1622775
−ꢎ51.1612ꢎ87
−ꢎ51.1728141
−ꢎ51.1808811
−1260.ꢎ42ꢎ525
−1260.2ꢎ14480
−1260.2ꢎ07625
−1260.ꢎ4215ꢎ7
−1260.2ꢎ074ꢎ1
−1260.28ꢎꢎ141
−1005.8827418
−1005.22ꢎ7253
−1005.22ꢎ17ꢎ4
−1210.4286485
−120ꢎ.7747301
−120ꢎ.7725423
−1138.4686513
−1137.8167032
−1137.8155ꢎ84
−1230.ꢎ625517
−1230.3115557
−1230.3106038
0.00000000
3ꢎ1.740374
3ꢎ3.4313378
0.00000000
38ꢎ.833822ꢎ
3ꢎ2.ꢎ27ꢎ4ꢎ3
374.3ꢎ5345
0.00000000
445.1087818
3ꢎ6.8338225
430.2432707
0.00000000
3ꢎ0.327ꢎ871
3ꢎ2.ꢎ076556
362.5583352
341.2ꢎ7417ꢎ
0.00000000
3ꢎ3.3480546
3ꢎ5.1547187
0.00000000
3ꢎ3.1005773
3ꢎ5.3012548
0.00000000
3ꢎ7.332ꢎꢎ41
3ꢎ8.7717365
0.00000000
3ꢎꢎ.70ꢎꢎ8ꢎ5
405.4760283
0.00000000
3ꢎ4.5171811
3ꢎ7.428ꢎ27ꢎ
0.00000000
3ꢎ2.0078815
3ꢎ4.5166540
•
B1-o
B3-o
•
2
3
4
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
•
a7-o
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
•
a8-o
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
•
a5-o
•
a6-o
5
6
7
8
9
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
1
0
pꢇꢆꢌꢋꢁ
•
B1-o
B3-o
•
hydrogen atom bond dissociation energy (BDE). e theoretical results revealed that the
antioxidant activity was negatively correlated with the phenolic hydroxyl hydrogen bond
dissociation energy. Compound 4 exhibited the best antioxidant activity, given that A5-OH
and A6-OH formed an intramolecular hydrogen bond, and A6-OH exhibited the lowest
bond dissociation energy (341.2974179 kJ/mol). e computed results were consistent with
the experimental conclusions mentioned above.

6
B.ꢀD. ZHOU ET AL.
2
.3. Summarization
Ten xanthones were synthesized, and their antitumor, antityrosinase, and antioxidant activ-
ities were assessed. Compounds 1, 4, 6–7, and 9–10 displayed obvious antitumor activity.
Among these xanthones, compounds 6–7 and 9–10 exhibited the stronger cytotoxicity
against SMMC-7721 and A-549 cells compared with cisplatin, respectively. Compound 4
containing two ortho-hydroxyl groups had the better antioxidant activity. Compounds 2
and 8–10 inhibited tyrosinase to a certain extent. erefore, this type of compound could
be considered to be promising leads for the development of the more potent anticancer,
antityrosinase, and antioxidant agents. Further research will be conducted to investigate
the structure–activity relationships of this type of compound.
3
. Experimental
.1. Chemistry
Melting points were determined on a micro-melting point apparatus and are uncorrected
3
(
Shanghai Precision Scientific Instrument Co., Ltd, Shanghai, China). IR spectra were
1
13
recorded with KBr pellets (Bruker, Bremen, Germany). H and C NMR spectra were
1
determined in acetone-d and dimethyl sulfoxide-d at 500 MHz for H and 125 MHz for
6
6
1
3
C (Bruker, Fällanden, Switzerland). Low-resolution mass spectra were recorded on an
ABI mass spectrometer (American Applied Biosystems, Carlsbad, USA). High-resolution
mass (HRMS) data were recorded via a negative ion electron impact mass spectrometry
using a time-of-flight analyzer (Bruker Daltonics Inc, Boston, USA). e chemical reagents
purchased (Shanghai Aladdin Biochemical Technology Co., Ltd, Shanghai, China) were of
the highest commercially available purity and were used directly.
3
.2. General procedure for preparation of substituted 1,3-dihydroxy-9H-
xanthones
A mixture of phosphorus pentoxide (0.5 g) and methanesulfonic acid (10 ml) was heated at
1
10 °C until a clear solution was obtained. e solution was then lowered to 90 °C. A mixture
of phloroglucinol (0.2 g, 1.6 mmol) and the relevant substituted salicylic acid (1.6 mmol)
were added to this solution. Heating was continued for 20 min, and the reaction mixture
was poured into ice-water. e resulting solid was collected by filtration, washed with water,
dried, and purified by flash column chromatography to afford the target xanthone.
3
.2.1. 1,3-Dihydroxy-9H-xanthen-9-one (1)
Yellow solid; yield 48%; mp: 258–260 °C; IR (KBr) v 756, 829, 1159, 1223, 1290, 1467, 1570,
max
−
1 1
1
611, 1653, 3078, 3256 cm ; H NMR (500 MHz, CD COCD ): δ 6.27 (d, 1H, J = 2.2 Hz,
H-2), 6.44 (d, 1H, J = 2.2 Hz, H-4), 7.44–7.47 (m, 1H, H-7), 7.53 (d, 1H, J = 8.5 Hz, H-5),
.82–7.85 (m, 1H, H-6), 8.19 (dd, 1H, J = 1.6, 8.0 Hz, H-8), 9.88 (s, OH-3), 12.91 (s, OH-1);
C NMR (125 MHz, CD COCD ): δ 94.9, 99.0, 103.8, 118.5, 121.2, 125.1, 126.3, 136.2,
56.8, 158.8, 164.7, 166.5, 181.3; ESI-MS: m/z 227 [M − H] .
3 3
7
1
3
3
3
−
1

JOURNAL OF ASIAN NATURAL PRODUCTS RESEARCH
7
3
.2.2. 1,3,7-Trihydroxy-9H-xanthen-9-one (2)
Yellow solid; yield 46%; mp: 206–209 °C; IR (KBr) v 536, 852, 976, 1074, 1168, 1279, 1335,
max
−1 1
1
476, 1578, 1614, 1652, 3047, 3395 cm ; H NMR (500 MHz, CD COCD ): δ 6.29 (d, 1H,
3 3
J = 2.1 Hz, H-2), 6.46 (d, 1H, J = 2.1 Hz, H-4), 7.66 (d, 1H, J = 9.1 Hz, H-5), 7.80 (dd, 1H,
1
3
J = 2.8, 9.1 Hz, H-6), 8.08 (d, 1H, J = 2.8 Hz, H-8), 9.99 (s, OH-Ar), 12.67 (s, OH-1);
C
NMR (125 MHz, CD COCD ): δ 95.1, 99.3, 103.5, 119.1, 120.7, 122.1, 130.6, 146.3, 155.1,
3
3
−
1
58.8, 164.6, 166.9, 180.4; ESI-MS: m/z 243 [M − H] .
3
.2.3. 1,3,8-Trihydroxy-9H-xanthen-9-one (3)
Yellow solid; yield 53%; mp: 223–226 °C; IR (KBr) v 737, 822, 1082, 1154, 1235, 1444,
max
−
1 1
1
573, 1605, 1652, 3377 cm ; H NMR (500 MHz, CD COCD ): δ 6.27 (d, 1H, J = 2.2 Hz,
3 3
H-2), 6.43 (d, 1H, J = 2.2 Hz, H-4), 7.35 (dd, 1H, J = 1.0, 8.4 Hz, H-7), 7.56 (dd, 1H, J = 1.0,
8
.4 Hz, H-5), 7.89 (t, 1H, J = 8.4 Hz, H-6), 9.92 (s, OH-Ar), 12.95 (s, OH-1); 13C NMR
125 MHz, CD COCD ): δ 94.5, 99.3, 104.3, 117.9, 120.6, 136.0, 145.2, 148.1, 151.3, 158.1,
(
3
3
−
1
64.9, 166.6, 180.3; ESI-MS: m/z 243 [M − H] .
3
.2.4. 1,3,5,6-Tetrahydroxy-9H-xanthen-9-one (4)
Rufous solid; yield 43%; mp: 130–133 °C; IR (KBr) v 522, 800, 966, 1180, 1265, 1364,
max
−
1 1
1
453, 1610, 1650, 1696, 3105, 3424 cm ; H NMR (500 MHz, CD COCD ): δ 6.27 (d, 1H,
3 3
J = 2.2 Hz, H-2), 6.50 (d, 1H, J = 2.2 Hz, H-4), 7.14 (d, 1H, J = 8.9 Hz, H-7), 7.98 (d, 1H,
1
3
J = 8.9 Hz, H-8), 12.89 (s, 1H, OH-1); C NMR (125 MHz, CD COCD ): δ 95.1, 99.3, 102.1,
3
3
−
1
05.8, 113.6, 114.5, 116.8, 125.4, 157.3, 158.3, 164.6, 166.2, 180.1; ESI-MS: m/z 259 [M − H] .
3
.2.5. 6-Chloro-1,3-dihydroxy-9H-xanthen-9-one (5)
Off-white solid; yield 41%; mp: 287–290 °C; IR (KBr) v 824, 1079, 1163, 1215, 1279,
max
−1 1
1
437, 1562, 1608, 1654, 3082, 3319 cm ; H NMR(CD SOCD 00 MHz): δ 6.21 (1H, d,
3 3,5
J = 2.1 Hz, H-2), 6.37 (1H, d, J = 2.1 Hz, H-4), 7.49 (1H, dd, J = 1.8, 8.5 Hz, H-7), 7.75 (1H,
d, J = 1.8 Hz, H-5), 8.09 (1H, d, J = 8.5 Hz, H-8), 11.3 (1H, s, OH-3), 12.6 (1H, s, OH-1);
1
3
C NMR(CD SOCD 5 MHz): δ 94.3, 98.5, 102.2, 117.7, 118.8, 125.0, 127.1, 139.9, 155.7,
3
3,12
−
1
57.3, 162.9, 166.1, 179.1; ESI-MS: m/z 261 [M − H] .
3
.2.6. 7-Chloro-1,3-dihydroxy-9H-xanthen-9-one (6)
Yellow solid; yield 51%; mp: 244–247 °C; IR (KBr) v 823, 1078, 1158, 1266, 1459, 1570,
max
−1 1
1
611, 1627, 2924, 3070, 3382 cm ; H NMR (500 MHz, CD COCD ): δ 6.30 (d, 1H,
J = 2.2 Hz, H-2), 6.46 (d, 1H, J = 2.2 Hz, H-4), 7.60 (d, 1H, J = 8.9 Hz, H-5), 7.84 (dd, 1H,
J = 2.7, 8.9 Hz, H-6), 8.11 (d, 1H, J = 2.7 Hz, H-8), 12.67 (s, 1H, OH-1); C NMR (125 MHz,
CD COCD ): δ 94.2, 98.5, 119.9, 120.3, 124.5, 129.2, 135.2, 136.4, 154.5, 157.9, 163.7, 166.4,
3 3
13
3
3
−
1
79.2; ESI-MS: m/z 261 [M − H] .
3
.2.7. 1,3-Dihydroxy-7-nitro-9H-xanthen-9-one (7)
Rufous solid; yield 40%; mp: 241–244 °C; IR (KBr) v 840, 969, 1183, 1348, 1584, 1622,
max
−1 1
1
651, 1704, 3101, 3440 cm ; H NMR (CD COCD 00 MHz): δ 6.34 (1H, d, J = 1.9 Hz,
3 3,5
H-2), 6.53 (1H, d, J = 1.9 Hz, H-4), 7.79 (1H, d, J = 9.2 Hz, H-5), 8.63 (1H, dd, J = 2.9,
.2 Hz, H-6), 8.94 (1H, d, J = 2.9 Hz, H-8), 10.2 (1H, s, OH-3), 12.5 (1H, s, OH-1); 13
NMR(CD COCD 5 MHz): δ 95.6, 99.9, 116.6, 116.8, 120.5, 122.4, 130.3, 144.4, 157.9,
9
C
3
3,12
−
1
60.0, 164.6, 167.3, 179.9; ESI-MS: m/z 272 [M − H] .

8
B.ꢀD. ZHOU ET AL.
3
.2.8. 1,3-Dihydroxy-5,7-dinitro-9H-xanthen-9-one (8)
Rufous solid; yield 42%; mp: >338 °C; IR (KBr) v 807, 1074, 1158, 1262, 1348, 1463, 1540,
max
−1 1
1
615, 1642, 3087, 3422 cm ; H NMR (500 MHz, CD COCD ): δ 6.41 (d, 1H, J = 2.1 Hz,
3 3
H-2), 6.60 (d, 1H, J = 2.1 Hz, H-4), 9.17 (d, 1H, J = 2.8 Hz, H-8), 9.92 (d, 1H, J = 2.8 Hz,
1
3
H-6), 10.46 (s, OH-3), 12.19 (s, OH-1); C NMR (125 MHz, CD COCD ): δ 96.3, 100.8,
3
3
1
3
03.2, 123.9, 126.1, 126.2, 152.5, 154.3, 157.3, 157.7, 164.6, 168.0, 178.2; HR-ESI-MS: m/z
-
17.0049 [M - H] (Calcd for C H N O , 317.0051).
1
3
5
2
8
3
.2.9. 1,3-Dihydroxy-6-(trifluoromethyl)-9H-xanthen-9-one (9)
Yellow solid; yield 75%; mp: 228–331 °C; IR (KBr) vmax 817, 837, 906, 1065, 1077, 1134,
−
1
1
157, 1174, 1249, 1284, 1318, 1352, 1371, 1403, 1444, 1504, 1573, 1610, 1657, 3375 cm ;
1
H NMR(CD COCD 00 MHz): δ 6.26 (1H, d, J = 2.2 Hz, H-2), 6.41 (1H, d, J = 2.2 Hz,
3
3,5
H-4), 7.72 (1H, dd, J = 1.1, 8.3 Hz, H-7), 7.82 (1H, d, J = 1.1 Hz, H-5), 8.32 (1H, d, J = 8.3 Hz,
13
H-8), 10.0 (1H, s, OH-3), 12.6 (1H, s, OH-1); C NMR (CD COCD3,125 MHz): δ 95.1, 99.4,
3
1
03.9, 116.2, 121.1, 123.8, 125.3, 127.9, 132.9, 156.3, 158.7, 164.7, 167.0, 180.2; HR-ESI-MS:
-
m/z 295.0226 [M - H] (Calcd for C H F O , 295.0224).
1
4
6
3
4
3
.2.10. 7-(2,4-Difluorophenyl)-1,3-dihydroxy-9H-xanthen-9-one (10)
Yellow solid; yield 76%; mp: 295–298 °C; IR (KBr) vmax 809, 1078, 1103, 1175, 1283,
−
1
1
300, 1320, 1343, 1422, 1485, 1522, 1569, 1607, 1656, 2852, 2923, 2959, 3138, 3427 cm ;
1
H NMR(CD COCD 00 MHz): δ 6.29 (1H, d, J = 2.2 Hz, H-2), 6.48 (1H, d, J = 2.2 Hz,
3
3,5
H-4), 7.16–7.23 (2H, m, H-3 ’, H-5’), 7.66 (1H, d, J = 8.7 Hz, H-5), 7.68–7.78 (1H, m,
1
3
H-6’), 8.01–8.03 (1H, m, H-6), 8.32 (1H, t, J = 1.8 Hz, H-8), 12.8 (1H, s, OH-1); C NMR
(
CD COCD3,125 MHz): δ 95.0, 99.0, 99.2, 103.7, 105.0, 105.2, 105.4, 112.8, 112.9, 113.0,
3
1
13.2, 119.0, 126.3, 132.8, 136.6, 156.3, 164.7, 166.8, 180.9; HR-ESI-MS: m/z 339.0465
−
[
M−H] (Calcd for C H F O , 339.0474).
1
9
9
2
4
3
.3. Bioassay
.3.1. Cytotoxicity assays
e human tumor cell lines HL-60, A-549, SMMC-7721, MCF-7, SW-480, HeLa, HepG2,
3

Huh-7, and MT-4 were used in the cytotoxic assay. ese cell lines were obtained from
ATCC (Manassas, VA, USA). Cells were cultured in RMPI-1640 or DMEM medium
(
Biological industries, Kibbutz Beit-Haemek, Israel) supplemented with 10% fetal bovine
serum (Biological Industries) at 37 °C in a humidified atmosphere with 5% CO . e cyto-
2
toxicity assay was evaluated using MTS (Promega, Madison, WI, USA) assay. Briefly, cells
were seeded into each well of a 96-well cell culture plate. Af er 12 h of incubation at 37 °C,
the test compound (40 μM) was added. Af er incubation for 48 h, cells were subjected to
the MTS assay. Compounds with a growth inhibition rate of 50% were further evaluated
at concentrations of 0.064, 0.32, 1.6, 8, and 40 μM in triplicate. Cisplatin and paclitaxel
(
Sigma, St. Louis, MO, USA) served as positive controls. e IC value of each compound
50
was calculated using the method of Reed and Muench [16]. e results were expressed as
average ± standard deviation.

JOURNAL OF ASIAN NATURAL PRODUCTS RESEARCH
9
3
.3.2. Mushroom tyrosinase in vitro inhibition
Sample or kojic acid standard (final concentration, 10 g/ml) was mixed with L-Dopa (final
concentration, 1.25 mM) in the 96-well microplate. Each test article was assayed in triplicate.

e reaction was initiated by adding tyrosinase (final concentration, 25 U/ml). e plate
was incubated at room temperature on shaker for 5 min. Plates were read at 490 nm. e
results were expressed as average ± standard deviation. e effect of tyrosinase inhibition
was calculated using the following equation:
Tyrosinase inhibition (% )
ꢀ
ꢁ
=
1 − Mean OD490nmof Sample/Mean OD490nmof Untreated Control × 100%
3
.3.3. Antioxidant activity
3
.3.3.1. In vitro antioxidant capacity. Sample or the trolox standard (final concentration,
2
5 g/ml) was added to 96-well microplates. Each test article was assayed in triplicate. e
reaction was initiated by adding DPPH solution (final concentration, 100 μM) at 30 °C for
1

h. e plate was read at 515 nm. e results were expressed as average ± standard deviation.
e antioxidant effect was calculated using the following equation:
Antioxidant inhibition (%)
ꢀ
ꢁ
=
1 − Mean OD515nm of sample∕Mean OD515nm of untreated control ×100%
3
.3.3.2. Calculated methods
All DFT calculations were performed at 298 K in the gas phase with Gaussian 09. e
geometry was further optimized at the B3LYP/6–311++G (d, p) level. Vibrational analysis
was also performed at the same level. No imaginary frequencies were noted, confirming
the considered conformers as real minima.
Acknowledgments

e authors gratefully thank the Project of Yunnan Provincial Department of Education (No.
016ZZX200), the Education Scientific Research Project for Middle-Age and Young Teachers of
Fujian Province (No. JA15453), the Science and Technology Planning Project of Putian City (No.
2
2
014S02(3)), and the Scientific Research Projects of Putian University (Nos. 2,014,053 and 2,015,016)
for supporting this program.
Disclosure statement
No potential conflict of interest was reported by the authors.
Funding

is work was supported by the Project of Yunnan Provincial Department of Education [grant num-
ber 2016ZZX200], the Education Scientific Research Project for Middle-Age and Young Teachers of
Fujian Province [grant number JA15453], the Science and Technology Planning Project of Putian
City [grant number 2014S02(3)], and the Scientific Research Projects of Putian University [grant
number 2014053 and 2015016] for supporting this program.

10
B.ꢀD. ZHOU ET AL.
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