Design, synthesis and bioactivity of novel glycosylthiadiazole derivatives

Article abstract of DOI:10.3390/molecules19067832

A series of novel glycosylthiadiazole derivatives, namely 2-phenylamino-5- glycosyl-1,3,4-thiadiazoles, were designed and synthesized by condensation between sugar aldehydes A/B and substituted thiosemicarbazide C followed by oxidative cyclization by treating with manganese dioxide. The original fungicidal activities results showed that some title compounds exhibited excellent fungicidal activities against Sclerotinia sclerotiorum (Lib.) de Bary and Pyricularia oryzae Cav, especially compounds F-5 and G-8 which displayed better fungicidal activities than the commercial fungicide chlorothalonil. At the same time, the preliminary studies based on the Elson-Morgan method indicated that many compounds exhibited some inhibitory activity toward glucosamine-6- phosphate synthase (GlmS). The structure-activity relationships (SAR) are discussed in terms of the effects of the substituents on both the benzene and the sugar ring.

Full text of DOI:10.3390/molecules19067832

Molecules 2014, 19, 7832-7849; doi:10.3390/molecules19067832
OPEN ACCESS
molecules
ISSN 1420-3049
www.mdpi.com/journal/molecules
Article
Design, Synthesis and Bioactivity of Novel
Glycosylthiadiazole Derivatives
Guanghui Zong 1,†, Hanqing Zhao 1,2,†, Rui Jiang 1, Jianjun Zhang 1,*, Xiaomei Liang 1, Baoju Li 3,
Yanxia Shi 3,* and Daoquan Wang 1
1
Department of Applied Chemistry, China Agricultural University, Beijing 100193, China
2
Department of Fundamental Science, Beijing University of Agriculture, Beijing 102206, China
3
Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing 100081, China
†
The authors contributed equally to this paper.
* Authors to whom correspondence should be addressed; E-Mails: zhangjianjun@cau.edu.cn (J.Z.);
shiyanxia@caas.cn (Y.S.); Tel.: +86-10-6273-1115 (J.Z.); Fax: +86-10-6273-2219 (J.Z.).
Received: 21 April 2014; in revised form: 4 June 2014 / Accepted: 5 June 2014 /
Published: 11 June 2014
Abstract: A series of novel glycosylthiadiazole derivatives, namely 2-phenylamino-5-
glycosyl-1,3,4-thiadiazoles, were designed and synthesized by condensation between sugar
aldehydes A/B and substituted thiosemicarbazide C followed by oxidative cyclization by
treating with manganese dioxide. The original fungicidal activities results showed that
some title compounds exhibited excellent fungicidal activities against Sclerotinia
sclerotiorum (Lib.) de Bary and Pyricularia oryzae Cav, especially compounds F-5 and
G-8 which displayed better fungicidal activities than the commercial fungicide
chlorothalonil. At the same time, the preliminary studies based on the Elson-Morgan method
indicated that many compounds exhibited some inhibitory activity toward glucosamine-6-
phosphate synthase (GlmS). The structure-activity relationships (SAR) are discussed in
terms of the effects of the substituents on both the benzene and the sugar ring.
Keywords: 1,3,4-thiadiazole; carbohydrates; synthesis; fungicidal activity

Molecules 2014, 19
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1. Introduction
Carbohydrates play an important role in the field of pesticide investigation, and many natural
carbohydrate products used as pesticides have shown great vitality. Validamycin [1], validoxylamine A [2],
trehazolin [3], streptomycin [4] and kasugamycin [5] have already proven excellent activities against
pests and fungi, and most of them are considered to be non-toxic for mammals and have no adverse
effects on non-target organisms or on the environment. Many natural products containing glycosyl
residues in their structures, such as the derivatives of avermecin and spinosad, are well known green
pesticides and widely used in the control of many kinds of pests, taking the advantage of excellent
bioactivities, good environment compatibility and structure variability [6–9]. Encouraged by the
successes of the developed commercial pesticides based on natural carbohydrates, pesticide chemists
have paid considerable attention to the design, synthesis and activity evaluation of novel carbohydrate-
containing compounds as potential pesticides, finding that heterocyclic compounds modified by
carbohydrates exhibit excellent biological activities [10–13].
It is well established that 1,3,4-thiadiazole and their derivatives exhibit a broad spectrum of
biological activities not only in research on drugs with anticancer [14], antimicrobial [15],
antituberculosis [16], anticonvulsant [17], or anti-inflammatory activities [18,19] but also in pesticide
research such as antifungal [20,21], insecticidal [22], herbicidal [23] and also plant growth regulating
agents [24]. Consequently, studies on the synthesis and bioassays of 1,3,4-thiadiazole derivatives have
attracted increasing attention in the field of pesticide discovery. As pesticides, many of the 1,3,4-
thiadiazole derivatives showed high toxicity profiles and were taken off the market. Thus an interest in
developing novel bioactive agents with low toxicities and an acceptable impact on the environment is
increasing. One way to reach this goal is to modify the parent 1,3,4-thiadiazole structure.
In our previous studies [25–27], numerous 1,3,4-thiadiazole derivatives (I-III, Figure 1) were
designed and synthesized. Some of them exhibited good fungicidal activity against Rhizoctonia solani
Kühn, Verticillium dahlia Kleb. and Pyricularia oryzae Cav.
Figure 1. 1,3,4-Thiadiazole derivatives developed in our laboratory.

Molecules 2014, 19
7834
Considering the advantages of using carbohydrates in developing novel pesticides, we made some
efforts to investigate the antifungal activities of 1,3,4-thiadiazoles modified by carbohydrates, and a
series of thiadiazoline derivatives containing glucofuranose were synthesized in our laboratory [28].
The fungicidal activity results obtained showed that compound IV (Figure 1) exhibited excellent
fungicidal activities against Phytophtora parasitica Dast and Helminthosporium maydis Nisik & Miy.
Inspired by these promising results, we developed a great interest in searching for potential 1,3,4-
thiadiazole derivative pesticides containing furanoses. In the structure of compound IV, the 1,3,4-
thiadiazole moiety and the glucofuranose moiety were connected in a spirocyclic manner. In particular
the question of the kind of changes in the fungicidal activities that might happen if the two moieties
were connected directly to each other (through a single bond) caught our attention. Thus, a series of
xylose-based 1,3,4-thiadiazoles, were synthesized and evaluated, as mentioned in one of our former
Chinese patents [29]. At the same time, as there is some similarity between the target compounds and
the D-fructose-6-P, which is one of the substrates in the first committed step of the hexosamine
biosynthesis pathway [30] by glucosamine-6-phosphate synthase (GlmS; EC 2.6.1.16), their enzyme
inhibitory activities were evaluated, too. In this paper, we would like to report their synthesis (Schemes
1 and 2) and bioactivities in much greater details, and also their structure-activity relationship studies.
Furthermore, to investigate the effects of the protecting groups in the sugar ring on the activities of
compounds F and G, the deprotected compounds H, I and J were also synthesized and evaluated. We
report herein the preliminary results of the study.
2. Results and Discussion
2.1. Synthesis of the Title Compounds
The synthesis of the target compounds was outlined in Scheme 1. According to the known
methods [31–33], two furanosyl aldehydes (A and B) were prepared using D-glucose as the
starting material.
Scheme 1. Synthesis of the target compounds F/G.

Molecules 2014, 19
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The substituted thiosemicarbazides C were synthesized from the corresponding substituted
arylamines as previously described [24,31]. The condensation between furanosyl aldehydes A or B and
substituted thiosemicarbazides C provided compounds D or E, respectively. Then the target
compounds F or G were prepared by treating compounds D or E with MnO2. Compound H was
obtained by deacetylation of compound F-8, and compounds I and J were obtained by
deisopropylidenation of the related compounds F-8 and G-7, as shown in Scheme 2.
Scheme 2. Synthesis of the sugar moiety-modified compounds.
N
N
N
N
N N
O
OR1
O
OR1
O
OR1
CF3COOH/H2O
MeONa
MeOH
R2
N
R2
N
R2
N
S
S
S
H
H
H
OH
O
O
C(Me)2
C(Me)2
O
O
OH
H: R1=H, R2
=
F-8: R1=Ac, R2=
Cl
F3C
I: R1=Ac, R2=
Cl
F3C
Cl
F3C
J: R1=Allyl, R2=
G-7: R1=Allyl, R2=
All the derivatives were synthesized according to the procedures described in Schemes 1 and 2 in
1
good overall yields of 65%–92%. The synthesized compounds were characterized by H-NMR, MS
and HRMS. Most of the 1H-NMR experiments of compounds F and G were conducted in CDCl3 as the
solvent. Nevertheless, the signal of NH was too weak in some cases, so we had to switch the solvent to
DMSO-d6. The physical data of the target compounds are given in Table 1.
Table 1. Physical Data of Compounds F/G.
Compd. R1
R2
Formula
Status
m.p./°C
Yield (%)
79
F-1
F-2
Ac
Ac
Ac
Ac
Ac
Ac
Ac
4-Br-C6H4-
C17H18BrN3O5S
C18H21N3O5S
C18H21N3O6S
C19H23N3O5S
C17H17Cl2N3O5S
C17H17Cl2N3O5S
C21H21N3O5S
White foamy solid
White foamy solid
yellow foamy solid
White foamy solid
White foamy solid
White foamy solid
White foamy solid
White foamy solid
White foamy solid
yellow foamy solid
White foamy solid
White foamy solid
White foamy solid
White foamy solid
White foamy solid
White foamy solid
yellow foamy solid
White foamy solid
White foamy solid
White foamy solid
204.1–204.7
187.9–188.1
177.7–178.7
133.7–134.3
147.7–148.5
61.8–62.9
4-CH3-C6H4-
4-CH3O-C6H4-
2,4-(CH3)2-C6H4-
3,4-Cl2-C6H4-
2,5-Cl2-C6H4-
1-Naphthyl-
82
F-3
73
F-4
81
F-5
83
F-6
85
F-7
57.0–58.1
65
F-8
Ac 4-Cl-3-CF3-C6H3- C18H17ClF3N3O5S
140.0–141.2
195.1–195.5
198.4–198.7
204.1–204.7
187.9–188.1
177.7–178.7
133.7–134.3
147.7–148.5
61.8–62.9
77
F-9
Ac
Ac
All
All
All
C6H4-
C17H19N3O5S
C17H18N4O7S
C18H20BrN3O4S
C19H23N3O4S
C19H23N3O5S
C20H25N3O4S
C18H19Cl2N3O4S
C18H19Cl2N3O4S
C22H23N3O4S
78
F-10
G-1
G-2
G-3
G-4
G-5
G-6
G-7
G-8
G-9
G-10
4-NO2-C6H4-
4-Br-C6H4-
73
68
4-CH3-C6H4-
4-CH3O-C6H4-
74
78
All 2,4-(CH3)2-C6H4-
76
All
All
All
3,4-Cl2-C6H4-
2,5-Cl2-C6H4-
1-Naphthyl-
81
76
57.0–58.1
68
All 4-Cl-3-CF3-C6H3- C19H19ClF3N3O4S
140.0–141.2
195.1–195.5
198.4–198.7
86
All
All
C6H4-
C18H21N3O4S
C18H20N4O6S
91
4-NO2-C6H4-
71

Molecules 2014, 19
7836
2.2. Fungicidal Activity of Compounds F/G against Six Fungus Species
Compounds F/G were evaluated in a series of in vitro fungicidal tests against six fungal species,
and compared with the commercial fungicide chlorothalonil. As shown in Table 2, the resulting data
revealed that most of the tested compounds displayed a certain degree of fungicidal activity against the
six species. Among them, the majority of the compounds showed better fungicidal activity against
S. sclerotiorum than the other five fungi. Among the 20 tested compounds, there were seven that
displayed an inhibition rate of 90% or more against S. sclerotiorum at a concentration of 50 µg/mL.
Therefore, the further activity evaluation of the compounds in our research was performed against
S. sclerotiorum.
Table 2. Fungicidal activity of compounds F/G against six fungus species (% control
at 50 µg/mL).
S.
P.
B.
P. oryzae
Cav.
29
P.
Compds No.
R. solani
sclerotiorum parasitica Dast cinerea
asparagi saecrdo
F-1
F-2
F-3
F-4
F-5
F-6
F-7
F-8
F-9
F-10
G-1
G-2
G-3
G-4
G-5
G-6
G-7
G-8
68
76
80
84
91
94
88
89
68
52
79
87
74
93
85
90
92
98
86
91
93
11
40
22
35
21
97
67
37
27
12
30
32
65
61
74
63
42
64
40
43
92
14
50
26
32
24
66
74
51
−11
6
72
79
50
88
84
77
85
80
64
81
84
60
65
70
69
67
77
62
53
63
59
58
77
80
62
56
63
68
59
55
55
100
−8
35
5
52
27
58
76
72
70
46
56
41
80
81
79
87
85
87
85
50
23
25
43
21
54
25
26
15
66
51
84
98
66
71
85
57
94
83
79
78
82
G-9
G-10
Chlorothalonil
2.3. Precise Fungicidal Activity of Compounds F/G Against S. sclerotiorum
The precise fungicidal activity of compounds F/G against S. sclerotiorum was evaluated, and the
data are shown in Table 3. For half of the compounds F/G, i.e., 10 out of the 20 tested compounds,
their EC50 values were <3 μg/mL. They thus exhibited good fungicidal activity against S. sclerotiorum.
Among them, compounds F-5, F-8, G-5, G-6 and G-8 (the EC50 values of which are 0.29, 1.50, 1.50,
1.62 and 0.46 μg/mL, respectively) exhibited excellent fungicidal activity and they are comparable
with the commercial fungicide chlorothalonil (with a EC50 value of 0.59 μg/mL).

Molecules 2014, 19
Table 3. Fungicidal Activity of Compounds F/G against S. sclerotiorum.
7837
Compds No.
Regression eq
Y = 4.93 + 0.57x
Y = 3.96 + 1.39x
Y = 4.77 + 0.58x
Y = 4.11 + 1.04x
Y = 5.31 + 0.58x
Y = 3.93 + 1.80x
Y = 2.04 + 2.18x
Y = 4.80 + 1.11x
Y = 4.51 + 0.72x
Y = 4.13 + 0.56x
Y = 3.84 + 1.29x
Y = 3.94 + 1.58x
Y = 4.75 + 0.63x
Y = 2.48 + 2.52x
Y = 4.83 + 0.96x
Y = 4.80 + 0.94x
Y = 4.68 + 1.06x
Y = 5.34 + 0.99x
Y = 4.32 + 1.26x
Y = 4.69 + 1.00x
5.19 + 0.84x
r
EC50 (μg/mL) EC90 (μg/mL)
F-1
F-2
F-3
F-4
F-5
F-6
F-7
F-8
F-9
F-10
G-1
G-2
G-3
G-4
G-5
G-6
G-7
G-8
0.9416
0.9880
0.9791
0.9717
0.9349
0.9758
0.9937
0.9543
0.9798
0.9765
0.9059
0.9993
0.9457
0.9859
0.9880
0.9871
0.9940
0.9961
0.9809
0.9534
0.9784
1.33
5.64
2.45
7.11
0.29
3.94
22.53
1.50
4.70
37.13
8.01
4.70
2.51
9.92
1.50
1.62
2.00
0.46
3.44
2.02
0.59
247.14
47.03
382.78
120.46
47.38
20.30
87.03
21.6
276.27
7558.45
79.59
30.32
273.14
31.89
32.10
37.28
32.30
8.91
G-9
G-10
Chlorothalonil
35.81
38.63
19.56
In general, the following structure-activity relationships (SAR) in compounds F/G were observed:
(1) for the two series F and G, on an overall level the latter (R2 = allyl) displayed a better fungicidal
activity than the former (R2 = Ac), i.e., there were six compounds in series G that exhibited better
fungicidal activity than their counterparts in series F; (2) for the F series, the fungicidal activity is
increased by improving the electron-withdrawing ability of substituents on the benzene ring, i.e.,
in compounds F-4 (R2 = 2,4-(CH3)2-C6H3-), F-2 (R2 = 4-CH3-C6H3-), F-9 (R2 = C6H3-), F-3
(R2 = 4-CH3O-C6H3-) and F-1 (R2 = 4-Br-C6H3-) with the EC50 values of 7.11, 5.64, 4.70, 2.45
and 1.33 μg/mL, respectively; (3) for the G series, the fungicidal activity is increased by improving
the electron-withdrawing ability of substituents on the benzene ring, too, i.e., compounds G-4
(R2 = 2,4-(CH3)2-C6H3-), G-2 (R2 = 4-CH3-C6H3-), G-9 (R2 = C6H3-), G-3 (R2 = 4-CH3O-C6H3-) and
G-6 (R2 = 2,5-Cl2-C6H3-) with the EC50 values of 9.92, 4.70, 3.44, 2.51 and 1.62 μg/mL, respectively;
(4) In both series F and G, the compounds with two electron-withdrawing groups in the benzene ring
have the best fungicidal activity in their own series, i.e., compounds F-5 (R2 = 3,4-Cl2-C6H3-) and G-8
(R2 = 4-Cl-3-CF3-C6H3-) with the EC50 values of 0.29 and 0.46 μg/mL, respectively. They were
slightly better than the commercial fungicide chlorothalonil (EC50 value = 0.59 μg/mL). Similarly,
compounds F-8 (R2 = 4-Cl-3-CF3-C6H3-) and G-5 (R2 = 3,4-Cl2-C6H3-), both with EC50 values of
1.50 μg/mL, also displayed excellent fungicidal activities. However, the compounds with two
electron-donating groups in the benzene ring displayed only moderate fungicidal activity, i.e.,
compounds F-4 (R2 = 2,4-Me2-C6H3-) and G-4 (R2 = 2,4-Me2-C6H3-) with the EC50 values of 7.11 and
9.92 μg/mL, respectively.

Molecules 2014, 19
7838
2.4. Effect of Structure Modifications on the Sugar Ring around Compound F/G
Having identified the relatively potent compounds of series F/G, such as F-8, G-8 and G-7, we next
focused our attention on investigating the effects of subtle structural changes in the sugar ring of
compounds F/G. To this end, compounds H, I and J were prepared and evaluated. The fungicidal
results against S. sclerotiorum are provided in Table 4. It is evident from the data (Tables 3 and 4) that,
among all the derivatives studied above, compounds with complete OH-protection in the sugar ring,
i.e., compounds F-8, G-8 and G-7, displayed the most promising results, with the EC50 values of 1.50,
0.46, 2.00 μg/mL respectively. Meanwhile, compound H without the OH-protection at the 3-position
in the sugar ring (and with an EC50 value of 4.61 μg/mL) has shown a slightly decreased fungicidal
activity. Compounds I and J without the OH-protection at both 1 and 2-position displayed a significant
decrease in their fungicidal activity, with EC50 values of 8.18, 18.91 μg/mL, respectively. The results
above demonstrate that appropriate protections of the hydroxyl groups in the sugar ring can make
positive contributions to the fungicidal activity against S. Sclerotiorum. Interestingly, however, the
enzyme inhibitory activity of compounds H, I and J is superior to that of compound G-8 (Table 5),
which may be associated with a better structural similarity between fructose 6-phosphate (Fru-6-P) and
compounds H, I and J.
Table 4. Effects of structural modifications in sugar ring of compounds F/G on the activity
against S. sclerotiorum.
OH
OAc
N
N
N
N
OAllyl
O
N
N
O
O
Cl
N
H
Cl
N
S
S
H
N
H
S
F3C
F3C
O
O
O
C(Me)2
C(Me)2
O
O
C(Me)2
F-8
O
H
G-7
OAllyl
O
OAc
OAllyl
O
N
N
N
N
N
N
O
Cl
N
Cl
N
H
N
H
S
S
S
H
F3C
OH
OH
F3C
O
C(Me)2
OH
O
OH
G-8
I
J
Compds No.
Regression eq
Y = 4.40 + 0.91x
Y = 4.27 + 0.80x
Y = 2.80 + 1.72x
r
EC50
4.61
EC90
H
I
0.8678
0.9963
0.9228
119.37
321.93
105.14
8.18
J
18.91
Table 5. Enzyme inhibition Rate of Compounds F and G at 0.35 mm.
Compd No.
F-1
Inhibition Rate (%)
Compd No.
G-1
Inhibition Rate (%)
13.2
15.3
17.7
18.5
18.3
19.1
17.9
12.0
10.8
16.1
15.2
25.8
17.4
28.1
F-2
G-2
F-3
G-3
F-4
G-4
F-5
G-5
F-6
G-6
F-7
G-7

Molecules 2014, 19
7839
Table 5. Cont.
Compd No.
Inhibition Rate (%)
Compd No.
Inhibition Rate (%)
F-8
F-9
F-10
H
18.1
14.7
13.3
29.4
35.8
G-8
G-9
G-10
J
26.5
20.3
24.2
36.2
I
2.5. Bioassay of Enzyme Inhibitory Activities [34–37]
Inhibitory activity of all the synthesized compounds towards Candida albicans GlcN-6-P synthase
was evaluated using the optimized Elson-Morgan method [38]. The absorption value of the solution
was measured at 585 nm, and then the concentration was counted by the specification curve which was
determined thanks to the relation between the absorption value and the concentration of glucosamine-
6-phosphate. Finally the enzyme inhibition rate was calculated according to Equation (1):
M 0 − M
I =
×100%
(1)
M 0
In Equation (1): I is the inhibition rate,
is the average concentration of glucosamine-6-phosphate
M 0
in the blank test, and
is the average concentration of glucosamine-6-phosphate in the presence of
M
target compounds. The inhibition rates were given in Table 5 at 0.35 mm.
The compounds of series F and G exhibited some enzyme inhibitory activities (Table 5).
Compounds G-5, G-7, G-8 and G-10 are more active against glucosamine-6-phosphate synthase than
the other compounds. On the whole, the enzyme inhibitory activity of G series of compounds is
superior to the F series. The enzyme inhibitory activity of compounds H, I and J is superior to that of
compound G-8 (Table 5), which may be associated with a better structural similarity between fructose
6-phosphate (Fru-6-P) and compounds H, I and J.
3. Experimental
3.1. General Methods
All starting materials and reagents were commercially available and used without further
purification except as indicated. 1H-NMR (300 MHz) and 13C-NMR (75 MHz) spectra was recorded in
CDCl3 or DMSO-d6 with a Bruker DPX300 spectrometer, using TMS as internal standard; Mass
spectra were obtained with Agilent 1100 series LC/MSD mass spectrometer. High-resolution mass
spectra (HRMS) was performed by the Peking University. Melting points were measured on a
Yanagimoto melting-point apparatus and are uncorrected.
3.2. Chemical Synthesis
General Procedure for the Syntheses of Title Compounds F, G, H, I and J. Substituted aldehydes A
and B were prepared from D-glucose as the starting material according to known methods [20–22].
Substituted thiosemicarbazides C were synthesized from amines as previously described [33,39].

Molecules 2014, 19
7840
General Procedure for the Synthesis of Intermediate Compounds D/E. A solution of aldehyde A/B (5.5
mmol) and thiosemicarbazide C (5 mmol) in CH2Cl2 (100 mL) was heated to reflux for 6 h, at the end
of which time TLC (eluent: 2:1 petroleum ether-EtOAc) indicated that the reaction was complete.
The solvent was evaporated under diminished pressure at 40 °C to give a white solid, and the crude
product was used for next step directly without purification.
General Procedure for the Synthesis of Title Compounds F/G. To a stirred solution of compound D/E
(5.0 mmol) in CHCl3 (80 mL) was added MnO2 (10 g). The mixture was stirred for a further 1 h, at the
end of which time TLC (eluent: 2:1 petroleum ether-EtOAc) indicated that the reaction was complete.
After filtration, the filtrate was evaporated under reduced pressure to give a crude product, which was
purified on silica gel column chromatography with 4:1 petroleum ether-EtOAc as the eluent to give the
compounds F/G.
2-(4-Bromophenylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
thiadiazole (F-1). Yield: 79%. White solid, mp 239.1–240.2 °C. 1H-NMR (DMSO-d6): δ 10.61 (s, 1H,
NH), 7.62 (d, J = 8.9 Hz, 2H, ArH), 7.52 (d, J = 8.9 Hz, 2H, ArH), 6.07 (d, J = 3.7 Hz, 1H, H-1), 5.54
(d, J = 3.0 Hz, 1H, H-3), 5.28 (d, J = 3.0 Hz, 1H, H-4), 4.79 (d, J = 3.7 Hz, 1H, H-2), 2.02 (s, 3H,
13
CH3CO), 1.52, 1.31 (2s, 6H, Me2C); C-NMR (DMSO-d6) δ 168.82, 165.18, 154.23, 139.71, 131.74,
119.36, 113.27, 111.85, 104.36, 82.63, 76.51, 76.30, 26.33, 25.93, 20.61. ESI-MS m/z calcd.
for C17H17BrN3O5S (M-H) 454.0. Found: 454.0. HRMS for C17H19BrN3O5S [M+H]+ 456.0223.
Found: 456.0212.
2-(4-Tolylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-thiadiazole
1
(F-2). Yield: 82%. Pale-yellow solid, mp 227.6–228.0 °C. H-NMR (CDCl3): δ 10.42 (br, 1H, NH),
7.33-7.27 (m, 2H, ArH), 7.19 (m, 2H, ArH), 6.03 (d, J = 3.6 Hz, 1H, H-1), 5.72 (d, J = 3.1 Hz, 1H, H-3),
5.48 (d, J = 3.1 Hz, 1H, H-4), 4.65 (d, J = 3.7 Hz, 1H, H-2), 2.34 (s, 3H Ar-CH3), 2.00 (s, 3H, CH3CO),
13
1.59, 1.36 (2s, 6H, Me2C); C-NMR (CDCl3) δ 169.01, 138.05, 133.25, 130.13, 118.45, 112.86, 104.77,
83.30, 77.26, 76.92, 26.75, 26.22, 20.79. ESI-MS m/z calcd. for C18H21N3NaO5S (M+Na) 414.1. Found:
414.2. HRMS for C18H22N3O5S [M+H]+ 392.1275. Found: 392.1275.
2-(4-Methoxyphenylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
1
thiadiazols (F-3). Yield: 73%. Yellow solid, mp 186.7–188.0 °C. H-NMR (DMSO-d6): δ 10.28 (br,
1H, NH), 7.52 (d, J = 8.9 Hz, 2H, ArH), 6.93 (d, J = 8.9 Hz, 2H, ArH), 6.05 (d, J = 3.8 Hz, 1H, H-1),
5.50 (d, J = 3.0 Hz, 1H, H-3), 5.26 (d, J = 3.1 Hz, 1H, H-4), 4.76 (d, J = 3.8 Hz, 1H, H-2), 3.73 (s, 3H,
Ar-CH3O), 2.02 (s, 3H, CH3CO), 1.51, 1.30 (2s, 6H, Me2C); 13C-NMR (CDCl3) δ 169.08, 156.67,
133.90, 121.40, 114.97, 112.97, 104.83, 83.34, 77.29, 77.00, 55.61, 26.81, 26.29, 20.88. ESI-MS m/z
calcd. for C18H22N3O6S [M+H] 408.1. Found: 408.1. HRMS for C18H22N3O6S [M+H]+ 408.1224.
Found: 408.1218.
2-(2,4-Dimethylphenylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-
1
1,3,4-thiadiazols (F-4). Yield: 81%. White solid, mp 155.9–156.1 °C. H-NMR (CDCl3): δ 7.79 (br,
1H, NH), 7.29 (d, J = 8.0 Hz, 1H, ArH), 7.10–7.00 (m, 2H, ArH), 5.97 (d, J = 3.6 Hz, 1H, H-1), 5.65 (d,
J = 3.1 Hz, 1H, H-3), 5.43 (d, J = 3.1 Hz, 1H, H-4), 4.61 (d, J = 3.7 Hz, 1H, H-2), 2.32 (2s, 6H,

Molecules 2014, 19
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Ar-CH3), 1.98 (s, 3H, CH3CO), 1.56, 1.34 (2s, 6H, Me2C); 13C-NMR (CDCl3) δ 172.26, 168.97,
152.98, 137.24, 136.09, 132.52, 132.10, 127.69, 123.01, 112.83, 104.71, 83.26, 77.28, 76.93, 26.75,
26.25, 20.93, 20.78, 17.89. ESI-MS m/z calcd. for C19H24N3O5S [M+H] 406.1. Found: 406.1. HRMS
for C19H24N3O5S [M+H]+ 406.1431. Found: 406.1422.
2-(3,4-Dichlorophenylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-
1,3,4-thiadiazole (F-5). Yield: 83%. White solid, mp 232.6–235.0 °C. 1H-NMR (DMSO-d6): δ 10.79 (s,
1H, NH), 8.07 (d, J = 2.5 Hz, 1H, ArH), 7.59 (d, J = 8.8 Hz, 1H, ArH), 7.49 (dd, J = 8.9, 2.5 Hz, 1H,
ArH), 6.08 (d, J = 3.7 Hz, 1H, H-1), 5.56 (d, J = 3.1 Hz, 1H, H-3), 5.30 (d, J = 3.1 Hz, 1H, H-4), 4.79
13
(d, J = 3.8 Hz, 1H, H-2), 2.02 (s, 3H, CH3CO), 1.52, 1.31 (2s, 6H, Me2C); C-NMR (DMSO-d6) δ
168.81, 164.87, 154.87, 140.24, 131.30, 130.72, 123.15, 118.55, 117.53, 111.88, 104.40, 82.63, 76.47,
76.29, 26.32, 25.91, 20.59. ESI-MS m/z calcd. for C17H18Cl2N3O5S [M+H] 446.0. Found: 446.1.
HRMS for C17H18Cl2N3O5S [M+H]+ 446.0339. Found: 446.0330.
2-(2,5-Dichlorophenylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-
1
1,3,4-thiadiazole (F-6). Yield: 85%. White solid, mp 119.6–120.6 °C. H-NMR (CDCl3): δ 8.22 (d,
J = 2.3 Hz, 1H, ArH), 7.69 (s, 1H, NH), 7.33 (d, J = 8.5 Hz, 1H, ArH), 7.00 (dd, J = 8.5, 2.4 Hz, 1H,
ArH), 6.05 (d, J = 3.6 Hz, 1H, H-1), 5.73 (d, J = 3.1 Hz, 1H, H-3), 5.50 (d, J = 3.1 Hz, 1H, H-4), 4.68
(d, J = 3.6 Hz, 1H, H-3), 2.04 (s, 3H, CH3CO), 1.59, 1.36 (2s, 6H, Me2C); 13C-NMR (CDCl3) δ 169.02,
165.30, 156.89, 137.41, 133.82, 130.28, 123.74, 120.59, 119.02, 113.09, 104.92, 83.30, 77.18, 77.04,
26.80, 26.27, 20.83. ESI-MS m/z calcd. for C17H18Cl2N3O5S [M+H] 446.0. Found: 446.1. HRMS for
C17H18Cl2N3O5S [M+H]+ 446.0339. Found: 446.0329.
2-(Naphthalen-1-ylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-
1
1,3,4-thiadiazole (F-7). Yield: 65%. Pale-yellow solid, mp 186.1–186.9 °C. H-NMR (DMSO-d6): δ
10.36 (s, 1H, NH), 8.25 (m, 1H, ArH), 8.13 (d, J = 7.5 Hz, 1H, ArH), 7.96 (m, 1H, ArH), 7.72 (d,
J = 8.2 Hz, 1H, ArH), 7.64–7.50 (m, 3H, ArH), 6.06 (d, J = 3.8 Hz, 1H, H-1), 5.54 (d, J = 3.1 Hz, 1H,
H-3), 5.29 (d, J = 3.1 Hz, 1H, H-4), 4.78 (d, J = 3.8 Hz, 1H, H-2), 2.01 (s, 3H, CH3CO), 1.52, 1.30 (2s,
6H, Me2C); 13C-NMR (75 MHz, DMSO-d6) δ 168.83, 167.94, 153.96, 136.33, 133.93, 128.33, 126.21,
126.01, 125.95, 125.90, 124.06, 121.99, 117.28, 111.79, 104.30, 82.61, 76.59, 76.28, 26.32, 25.92,
20.61. ESI-MS m/z calcd. for C21H22N3O5S [M+H] 428.1. Found: 428.1. HRMS for C21H22N3O5S
[M+H]+ 428.1275. Found: 428.1263.
2-(4-Chloro-3-(trifluoromethyl)phenylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-
1
2,3,4-triol-5S)-1,3,4-thiadiazole (F-8). Yield: 77%. Pale-yellow solid, mp 219.8–221.5 °C. H-NMR
(DMSO-d6): δ 10.95 (s, 1H, NH), 8.25 (d, J = 2.5 Hz, 1H, ArH), 7.86 (dd, J = 8.8, 2.5 Hz, 1H, ArH),
7.69 (d, J = 8.8 Hz, 1H, ArH), 6.09 (d, J = 3.7 Hz, 1H, H-1), 5.57 (d, J = 3.1 Hz, 1H, H-3), 5.30 (d,
J = 3.1 Hz, 1H, H-4), 4.80 (d, J = 3.7 Hz, 1H, H-2), 2.02 (s, 3H, CH3CO), 1.52, 1.31 (2s, 6H, Me2C);
13C-NMR (DMSO-d6) δ 168.84, 164.88, 155.12, 139.63, 132.26, 126.99 (q, J = 30.7 Hz), 124.53, 122.25
(d, J = 1.7 Hz), 122.11, 120.91, 116.03 (q, J = 5.5 Hz), 111.91, 104.43, 82.63, 76.39 (d, J = 11.6 Hz),
26.32, 25.92, 20.57. ESI-MS m/z calcd. for C18H18ClF3N3O5S [M+H] 480.1. Found: 480.1. HRMS for
C18H18ClF3N3O5S [M+H]+ 480.0602. Found: 480.0588.

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2-(Phenylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-thiadiazole
(F-9). Yield: 78%. Pale-yellow solid, mp 213.7–214.1 °C. 1H-NMR (DMSO-d6): δ 10.63 (s, 1H, NH),
7.41-7.39 (m, 4H, ArH), 7.11 (m, 1H, ArH), 6.04 (d, J = 3.6 Hz, 1H, H-1), 5.74 (d, J = 3.1 Hz, 1H, H-3),
5.51 (d, J = 3.1 Hz, 1H, H-4), 4.66 (d, J = 3.7 Hz, 1H, H-2), 2.00 (s, 3H, CH3CO), 1.60, 1.36 (2s, 6H,
Me2C); 13C-NMR (DMSO-d6) δ 168.84, 165.55, 153.68, 140.43, 129.05, 121.97, 117.43, 111.82,
104.33, 82.62, 76.54, 76.30, 26.32, 25.93, 20.61. ESI-MS m/z calcd. for C17H20N3O5S [M+H] 378.1.
Found: 378.1. HRMS for C17H20N3O5S [M+H]+ 378.1118. Found: 378.1109.
2-(4-Nitrophenylamino)-5-(2R,3S-O-isopropylidene-4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
thiadiazole (F-10). Yield: 73%. Pale-yellow solid, mp 233.9–235.5 °C. 1H-NMR (DMSO-d6): δ 11.22
(s, 1H, NH), 8.27 (d, J = 8.7 Hz, 2H, ArH), 7.85 (d, J = 8.7 Hz, 2H, ArH), 6.10 (d, J = 3.4 Hz, 1H, H-1),
5.59 (d, J = 2.6 Hz, 1H, H-3), 5.32 (d, J = 2.6 Hz, 1H, H-4), 4.80 (d, J = 3.5 Hz, 1H, H-2), 2.02 (s, 3H,
13
CH3CO), 1.53, 1.31 (2s, 6H, Me2C); C-NMR (DMSO-d6) δ 168.62, 164.27, 155.82, 145.85, 140.67,
125.20, 116.75, 111.69, 104.22, 82.40, 76.22, 76.07, 26.10, 25.71, 20.36. ESI-MS m/z calcd. for
C17H17N4O7S (M-H) 421.1. Found: 421.0. HRMS for C17H19N4O7S [M+H]+ 423.0969. Found: 423.0957.
2-(4-Bromophenylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
1
thiadiazol (G-1). Yield: 68%. Pale-yellow solid, mp 204.1–204.7 °C. H-NMR (CDCl3): δ 10.50 (br,
1H, NH), 7.49-7.46 (m, 2H, ArH), 7.36-7.26 m, 2H, ArH), 6.04 (d, J = 3.6 Hz, 1H, H-1), 5.72 (m, 1H,
CH2=CHCH2), 5.63 (d, J = 3.1 Hz, 1H, H-3), 5.23-5.14 (m, 2H, CH2=CHCH2), 4.69 (d, J = 3.7 Hz, 1H,
H-2), 4.15 (d, J = 3.2 Hz, 1H, H-4), 4.04-3.86 (m, 2H, CH2=CHCH2), 1.58, 1.37 (2s, 6H, Me2C);
13C-NMR (DMSO-d6) δ 165.56, 155.14, 139.84, 134.03, 131.70, 119.28, 117.15, 113.08, 111.47,
111.39, 104.44, 81.82, 81.78, 77.57, 70.44, 26.52, 25.96. ESI-MS m/z calcd. for C18H19BrN3O4S (M-H)
452.0. Found: 451.9. HRMS for C18H21BrN3O4S [M+H]+ 454.0431. Found: 454.0415.
2-(4-Tolylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-thiadiazole
1
(G-2). Yield: 74%. Pale-yellow solid, mp 187.9–188.1 °C. H-NMR (CDCl3): δ 9.39 (br, 1H, NH),
7.29–7.26 (m, 2H, ArH), 7.18-7.16 (m, 2H, ArH), 6.02 (d, J = 3.7 Hz, 1H, H-1), 5.73 (m, 1H,
CH2=CHCH2), 5.62 (d, J = 3.1 Hz, 1H, H-3), 5.23-5.13 (m, 2H, CH2=CHCH2), 4.67 (d, J = 3.6 Hz, 1H,
H-2), 4.14 (d, J = 3.1 Hz, 1H, H-4), 4.02–3.85 (m, 2H, CH2=CHCH2), 2.33 (s, 3H, Ar-CH3), 1.56, 1.36
(2s, 6H, Me2C); 13C-NMR (DMSO-d6) δ 166.13, 154.21, 138.20, 134.08, 130.77, 129.40, 117.46,
117.11, 111.34, 104.40, 81.81, 77.62, 70.42, 26.53, 25.96, 20.28. ESI-MS m/z calcd. for C19H24N3O4S
[M+H] 390.1. Found: 390.1. HRMS for C19H24N3O4S [M+H]+ 390.1482. Found: 390.1468.
2-(4-Methoxyphenylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
1
thiadiazole (G-3). Yield: 78%. Pale-yellow solid, mp 177.7–178.7 °C. H-NMR (CDCl3): δ 9.70 (br,
1H, NH), 7.35–7.26 (m, 2H, ArH), 6.93–6.90 (m, 2H, ArH), 6.01 (d, J = 3.5 Hz, 1H, H-1), 5.72 (m, 1H,
CH2=CHCH2), 5.59 (d, J = 3.0 Hz, 1H, H-3), 5.22–5.13 (m, 2H, CH2=CHCH2), 4.66 (d, J = 3.6 Hz, 1H,
H-2), 4.13 (d, J = 3.0 Hz, 1H, H-4), 4.01–3.85 (m, 2H, CH2=CHCH2), 3.81 (s, 3H, Ar-CH3O), 1.56,
1.35 (2s, 6H, Me2C); 13C-NMR (DMSO-d6) δ 166.58, 154.53, 153.85, 134.12, 134.08, 119.21, 117.11,
114.26, 111.35, 104.39, 81.83, 77.65, 70.43, 55.19, 26.52, 25.96. ESI-MS m/z calcd. for C19H24N3O5S
[M+H] 406.1. Found: 406.2. HRMS for C19H24N3O5S [M+H]+ 406.1431. Found: 406.1417.

Molecules 2014, 19
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2-(2,4-Dimethylphenylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
thiadiazole (G-4). Yield: 76%. Pale-yellow solid, mp 133.7–134.3 °C. 1H-NMR (CDCl3): δ 9.41 (s, 1H,
NH), 7.58 (d, J = 9.0 Hz, 1H, ArH), 7.04–6.99 (m, 2H, ArH), 5.96 (d, J = 3.7 Hz, 1H, H-1), 5.82 (m,
1H, CH2=CHCH2), 5.53 (d, J = 3.1 Hz, 1H, H-3), 5.22–5.10 (m, 2H, CH2=CHCH2), 4.79 (d,
J = 3.7 Hz, 1H, H-2), 4.07 (d, J = 3.1 Hz, 1H, H-4), 4.12–3.87 (m, 2H, CH2=CHCH2), 2.25, 2.21 (2s,
6H, Ar-CH3), 1.46,1.29 (2s, 6H, Me2C); 13C-NMR (DMSO-d6) δ 168.89, 154.03, 136.75, 134.08,
133.48, 131.25, 129.92, 127.02, 122.20, 117.03, 111.29, 104.33, 81.83, 77.67, 70.40, 26.51, 25.95,
20.34, 17.72. ESI-MS m/z calcd. for C20H26N3O4S [M+H] 404.1. Found: 404.1. HRMS for
C20H26N3O4S [M+H]+ 404.1639. Found: 404.1624.
2-(3,4-Dichlorophenylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
1
thiadiazole (G-5). Yield: 81%. Pale-yellow solid, mp 147.7–148.5 °C. H-NMR (CDCl3): δ 10.77 (s,
1H, NH), 7.59 (s, 1H, ArH), 7.44 (m, 1H, ArH), 7.34 (m, 1H, ArH), 6.06 (d, J = 3.5 Hz, 1H, H-1), 5.74
(m, 1H, CH2=CHCH2), 5.65 (s, 1H, H-3), 5.24–5.15 (m, 2H, CH2=CHCH2), 4.71 (d, J = 3.5 Hz, 1H, H-2),
13
4.42–3.82 (m, 3H, H-4, CH2=CHCH2), 1.59, 1.38 (2s, 6H, Me2C); C-NMR (DMSO-d6) δ 165.25,
155.74, 140.37, 134.01, 131.25, 130.73, 122.95, 118.44, 117.48, 117.16, 111.41, 104.47, 81.79, 81.75,
77.50, 70.42, 26.51, 25.94. ESI-MS m/z calcd. for C18H20Cl2N3O4S [M+H] 444.0. Found: 444.0.
HRMS for C18H20Cl2N3O4S [M+H]+ 444.0546. Found: 444.0526.
2-(2,5-Dichlorophenylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
thiadiazole (G-6). Yield: 76%. Pale-yellow solid, mp 61.8–62.9 °C. 1H-NMR (CDCl3): δ 8.21 (m, 1H,
ArH), 7.73 (br, 1H, NH), 7.30 (m, 1H, ArH), 6.98 (m, 1H, ArH), 6.05 (d, J = 3.6 Hz, 1H, H-1), 5.74
(m, 1H, CH2=CHCH2), 5.63 (d, J = 3.2 Hz, 1H, H-3), 5.25–5.16 (m, 2H, CH2=CHCH2), 4.70 (d,
J = 3.6 Hz, 1H, H-2), 4.18 (d, J = 3.2 Hz, 1H, H-4), 4.05–3.87 (m, 2H, CH2=CHCH2), 1.57, 1.37 (2s,
13
6H, Me2C); C-NMR (DMSO-d6) δ 165.69, 157.21, 138.18, 134.05, 132.07, 130.79, 122.92, 120.35,
119.94, 117.16, 111.42, 104.50, 81.83, 77.66, 70.45, 26.52, 25.96. ESI-MS m/z calcd. for C18H20Cl2N3O4S
[M+H] 444.0. Found: 444.0. HRMS for C18H20Cl2N3O4S [M+H]+ 444.0546. Found: 444.0527.
2-(Naphthalen-1-ylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
thiadiazole (G-7). Yield: 68%. Pale-yellow solid, mp 57.0–58.1 °C. 1H-NMR (CDCl3): δ 10.28 (br, 1H,
NH), 8.23 (m, 1H, ArH), 8.12 (m, 1H, ArH), 7.95 (m, 1H, ArH), 7.00 (m, 1H, ArH), 7.58-7.49 (m, 3H,
ArH), 5.98 (d, J = 3.6 Hz, 1H, H-1), 5.78 (m, 1H, CH2=CHCH2), 5.37 (d, J = 3.1 Hz, 1H, H-3),
5.22–5.10 (m, 2H, CH2=CHCH2), 4.81 (d, J = 3.7 Hz, 1H, H-2), 4.14–3.64 (m, 3H, H-4, CH2=CHCH2),
13
1.48, 1.30 (2s, 6H, Me2C); C-NMR (DMSO-d6) δ 168.27, 154.99, 136.47, 134.05, 133.94, 128.30,
126.18, 126.03, 125.93, 125.83, 123.88, 122.04, 117.10, 111.36, 104.42, 81.85, 77.71, 70.44, 26.53,
25.97. ESI-MS m/z calcd. for C22H24N3O4S [M+H] 426.1. Found: 426.1. HRMS for C22H24N3O4S
[M+H]+ 426.1482. Found: 426.1462.
2-(4-Chloro-3-(trifluoromethyl)phenylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-
1
triol-5S)-1,3,4-thiadiazole (G-8). Yield: 86%. Pale-yellow solid, mp 140.0–141.2 °C. H-NMR
(CDCl3): δ 10.54 (s, 1H, NH), 7.81 (m, 1H, ArH), 7.63-7.48 (m, 2H, ArH), 6.05 (d, J = 3.5 Hz, 1H, H-1),
5.74 (m, 1H, CH2=CHCH2), 5.64 (d, J = 3.1 Hz, 1H, H-3), 5.24–5.15 (m, 2H, CH2=CHCH2), 4.71 (d,
J = 3.5 Hz, 1H, H-2), 4.17 (d, J = 3.0 Hz, 1H, H-4), 4.07–3.87 (m, 2H, CH2=CHCH2), 1.59, 1.38 (2s,

Molecules 2014, 19
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6H, Me2C); 13C-NMR (DMSO-d6) δ 165.26, 155.95, 139.76, 134.01, 132.21, 126.96 (q, J = 30.6 Hz),
124.53, 122.06 (d, J = 1.9 Hz), 122.01, 120.91, 117.13, 115.94 (q, J = 5.6 Hz), 111.44, 104.51, 81.81
(d, J = 2.1 Hz), 77.53, 70.45, 26.50, 25.93. ESI-MS m/z calcd. for C19H20ClF3N3O4S [M+H] 478.1.
Found: 478.2. HRMS for C19H20ClF3N3O4S [M+H]+ 478.0810. Found: 478.0802.
2-(Phenylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-thiadiazole
1
(G-9). Yield: 91%. Pale-yellow solid, mp 195.1–195.5 °C. H-NMR (CDCl3): δ 10.66 (s, 1H, NH),
7.46–7.35 (m, 4H, ArH), 7.09 (m, 1H, ArH), 6.04 (d, J = 3.6 Hz, 1H, H-1), 5.73 (m, 1H, CH2=CHCH2),
5.65 (d, J = 3.1 Hz, 1H, H-3), 5.23–5.13 (m, 2H, CH2=CHCH2), 4.69 (d, J = 3.6 Hz, 1H, H-2), 4.16 (d,
J = 3.1 Hz, 1H, H-4), 4.13–3.86 (m, 2H, CH2=CHCH2), 1.58, 1.37 (2s, 6H, Me2C); 13C-NMR (DMSO-d6)
δ 165.96, 154.62, 140.58, 134.06, 129.01, 121.80, 117.35, 117.11, 111.37, 104.43, 81.82, 77.62, 70.43,
26.52, 25.97. ESI-MS m/z calcd. for C18H21N3O4SNa (M+Na) 398.1. Found: 398.1. HRMS for
C18H22N3O4S [M+H]+ 376.1326. Found: 376.1323.
2-(4-Nitrophenylamino)-5-(2R,3S-O-isopropylidene-4S-O-allyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
thiadiazole (G-10). Yield: 71%. Pale-yellow solid, mp 198.4–198.7 °C. 1H-NMR (CDCl3): δ 11.75 (s,
1H, NH), 8.33–8.28 (m, 2H, ArH), 7.62–7.57 (m, 2H, ArH), 6.08 (d, J = 3.6 Hz, 1H, H-1), 5.80–5.69
(m, 2H, CH2=CHCH2, H-3), 5.25–5.15 (m, 2H, CH2=CHCH2), 4.74 (d, J = 3.7 Hz, 1H, H-2), 4.21 (d,
J = 3.2 Hz, 1H, H-4), 4.16-3.89 (m, 2H, CH2=CHCH2), 1.62, 1.40 (2s, 6H, Me2C); 13C-NMR (DMSO-d6)
δ 164.86, 156.82, 146.19, 140.78, 134.00, 125.32, 117.16, 116.84, 111.48, 104.55, 81.85, 81.79, 77.54,
70.49, 26.49, 25.91. ESI-MS m/z calcd. for C18H21N4O6S [M+H] 421.1. Found: 421.1. HRMS for
C18H21N4O6S [M+H]+ 421.1176. Found: 421.1173.
2-(4-Chloro-3-(trifluoromethyl)phenylamino)-5-(2R,3S-O-isopropylidene-4S-tetrahydrofuro-2,3,4-triol-
5S)-1,3,4-thiadiazole (H). To a solution of F-8 (0.48 g, 1.0 mmol) in MeOH (20 mL) was added
MeONa (0.05 g). The reaction mixture was stirred at rt for 0.5 h, at the end of which time TLC
(1:2 petroleum ether–EtOAc) indicated that the reaction was complete. Neutralization of the reaction
mixture with acidic ion exchange resin (Amberlite IR-120 (H+), Alfa Aesar, Tianjin, China) was
conducted, and the organic phase was concentrated under reduced pressure to give a crude product,
which could be purified by recrystallization from a mixture solvents of petroleum ether (10 mL) and
1
EtOAc (2 mL). Yield: 92%. White solid, mp 249.8–250.3 °C. H-NMR (DMSO-d6): δ 10.84 (s, 1H,
NH), 8.26 (d, J = 2.5 Hz, 1H, ArH), 7.86 (dd, J = 2.5, 8.8 Hz, 2H, ArH), 7.68 (d, J = 8.8 Hz, 1H, ArH),
6.01 (d, 1H, J = 3.6 Hz, H-1), 5.98 (d, J = 5.2 Hz, 1H, OH), 5.33 (d, J = 2.7 Hz, 1H, H-4), 4.60 (d,
J = 3.6 Hz, 1H, H-2), 4.68 (dd, J = 3.6 Hz, 5.2 Hz, 1H, H-3), 1.48, 1.29 (2s, 6H, Me2C);
13C-NMR (DMSO-d6) δ 165.13, 156.93, 139.88, 132.26, 127.00 (q, J = 30.7 Hz), 124.58, 121.97,
120.96, 115.93 (q, J = 5.6 Hz), 111.33, 104.52, 84.86, 78.49, 74.58, 26.66, 26.01. ESI-MS m/z calcd.
for C16H16ClF3N3O4S [M+H] 438.0. Found: 438.0. HRMS for C16H16ClF3N3O4S [M+H]+ 438.0497.
Found: 438.0478.
2-(4-Chloro-3-(trifluoromethyl)phenylamino)-5-(3S,4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-
thiadiazole (I). A solution of F-8 (0.48 g, 1.0 mmol) and CF3COOH (9 mL) and H2O (1 mL) was
stirred at rt for 12 h, at the end of which time TLC (1:2 petroleum ether-EtOAc) indicated that the
reaction was complete. The reaction mixture was neutralized with solid NaHCO3, and filtered through

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Celite. The filtrate was evaporated under reduced pressure to give a crude product, which was purified
on silica gel column chromatography with 1:1 petroleum ether-EtOAc as the eluent to give the
1
compound. Yield: 79%. Pale-yellow solid, mp 115.6–116.5 °C. H-NMR (DMSO-d6): δ 10.87 (d,
J = 12.2 Hz, 1H, NH), 8.26 (d, J = 2.6 Hz, 1H, ArH), 7.83 (dd, J = 2.6, 8.8 Hz, 2H, ArH), 7.68 (d,
J = 8.8 Hz, 1H, ArH), 7.05 (br, 1H, OH), 5.89 (br, 1H, OH), 5.55–5.52 (m, 1H, H-1), 5.40–5.26 (m, 1H,
H-3), 5.19–5.17 (m, 1H, H-4), 4.13–4.00 (m, 1H, H-2), 1.92 (s, 3H, OCH3). ESI-MS m/z calcd. for
C15H12ClF3N3O5S (M-H) 438.0. Found: 438.0. HRMS for C15H14ClF3N3O5S [M+H]+ 440.0289.
Found: 440.0272.
2-(Naphthalen-1-ylamino)-5-(3S,4S-O-acetyl-tetrahydrofuro-2,3,4-triol-5S)-1,3,4-thiadiazole
(J).
Deisopropylidenation of G-7 (0.52 g, 1.2 mmol) was accomplished by following the same procedure
1
employed for the preparation of compound J. Yield: 73%. Yellow solid, mp 57.2–59.7 °C. H-NMR
(DMSO-d6): δ 10.19 (s, 1H, NH), 8.24–8.22 (m, 1H, ArH), 8.09 (d, J = 7.4 Hz, 1H, ArH), 7.97–7.94
(m, 1H, ArH), 7.70 (d, J = 8.1 Hz, 1H, ArH), 7.58–7.49 (m, 3H, ArH), 6.62 (d, J = 5.5 Hz, 0.5H), 6.51
(d, J = 7.3 Hz, 0.5H), 5.77–5.70 (m, 1H, CH2=CHCH2), 5.57 (d, J = 4.3 Hz, 0.5H), 5.43–5.28 (m, 2H),
5.20–5.05 (m, 2.5H), 4.01–3.87 (m, 4H). ESI-MS m/z calcd. for C19H20N3O4S [M+H] 386.1. Found:
386.0. HRMS for C19H20N3O4S [M+H]+ 386.1169. Found: 386.1169.
3.3. Fungicidal Assays
Each of the test compounds was dissolved in DMSO (10 mL). Fungicidal activities of compounds F,
G, H, I and J against Sclerotinia sclerotiorum (Lib.) de Bary, Phytophthora parasitica Dast, Botrytis
cinerea Pers., Rhizoctonia solani Kühn., Pyricularia oryzae Cav. and Phoma asparagi Saecrdo were
evaluated using the mycelium growth rate test as previously reported [40].
Inhibition rates of compounds F and G against S. sclerotiorum, P. Parasitica Dast, B. cinerea, R
solani, P. oryzae Cav and P. asparagi Saecrdo at 50 μg/mL were determined first and the results are
given in Table 2. The inhibition rate of compounds F, G, H, I and J against S. sclerotiorum were
further determined at the concentrations of 50, 20, 10, 5 and 2 μg/mL, respectively. Then EC50 and
EC90 values were estimated using logit analysis [41]. The commercial fungicide chlorothalonil was
used as a control in the above bioassay.
3.4. Enzyme Inhibitory Activities Bioassay
Inhibitory activities of all the synthesized compounds towards Candida albicans GlcN-6-P synthase
were evaluated using the mycelium growth rate test as previously reported [38]. Three replicates were
performed. Absorbance at λ = 585 nm was measured and GlcN-6-P concentration in the sample was
read from the standard curve (solutions of glucosamine-HCl (0.1–1 mM) were assayed simultaneously, to
obtain a standard line from the plot of extinction against concentration of glucosamine). In each
experiment, two control samples, one without enzyme and one without substrates, were assayed in the
same way.

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4. Conclusions
In summary, a series of novel glycosylthiadiazole derivatives were synthesized, and their
bioactivities were evaluated. The bioassays showed that they had the inhibitory activities against
glucosamine-6-phosphate synthase, and at the same time, the results from the Chinese Academy of
Agricultural Science have shown that most of the tested compounds have good fungicidal activities
against S. sclerotiorum. Among all the novel compounds tested, compounds F-5 and G-8 displayed
better fungicidal activities than the commercial fungicide chlorothalonil. The SAR of the designed
compounds was studied: compounds with two electron-withdrawing substituents in the benzene ring
have better fungicidal activities than those with two electron-donating substituents. The compounds
with the protecting groups in the sugar ring have less inhibitory activities against Glms than those
without protecting groups, but displayed better fungicidal activities against S. sclerotiorum, and the
advantage depends on the number and type of the protection groups. Further studies are in progress.
Supplementary Materials
Supplementary materials can be accessed at: http://www.mdpi.com/1420-3049/19/6/7832/s1.
Acknowledgments
We acknowledge financial support of this investigation by NSFC of China (21172257), the
National High Technology Research and Development Programme of China (2011AA10A206), the
National Basic Research Program of China (2010CB126105, 2011BAE06B02-01, 2012BAK25B03-01)
and Key laboratory of Horticultural Crops Genetic Improvenment, Ministry of Agriculture in China.
Author Contributions
Main text paragraph Guanghui Zong was in charge of the synthesis experiments, Hanqing Zhao
was in charge of bioassay, they wrote the manuscript together; Rui Jiang provided help in the synthesis
1
experiments and bioassay experiments; H and 13C-NMR spectra were tested by Xiaomei Liang;
Jianjun Zhang provided guidance and suggestions for all the experiments and he also provided proper
suggestions when wrote and revised the manuscript; Daoquan Wang provided guidance and
suggestions in the synthesis experiments. Yanxia Shi and Baoju Li provided guidance and suggestions
in the bioassay experiments.
Conflicts of Interest
The authors declare no conflict of interest.
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Sample Availability: Samples of the compounds C-5, C-6, C-8, F-8 and G-8 are available from
the authors.
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