Some anilides of 2-alkylthio- and 2-chloro-6-alkylthio-4-pyridinecarboxylic acids: Synthesis and photosynthesis-inhibiting activity

Article abstract of DOI:10.3390/60700603

Many compounds containing a -CONH- group display photosynthesis inhibiting activity. Based on this structural feature, a group of anilides of 2-alkylthio-(1b-4f) or 2-chloro-6-alkylthio-4-pyridinecarboxylic acids (5a-6c) was synthesised. The prepared compounds were tested for their inhibition of the oxygen evolution rate (OER) in spinach chloroplasts. A quasi-parabolic dependence between photosynthesis-inhibiting activity and the lipophilicity of the compounds was determined for 1b-4f as well as for 5a-6c. The inhibitory activity of compounds 1b-4f was higher than that of 5a-6c for comparable lipophilicity values.

Full text of DOI:10.3390/60700603

Molecules 2001, 6, 603-613
molecules
ISSN 1420-3049
http://www.mdpi.org
Some Anilides of 2-Alkylthio- and 2-Chloro-6-Alkylthio-4-
Pyridinecarboxylic Acids: Synthesis and Photosynthesis-
Inhibiting Activity
Miroslav Miletín1*, Martin Doležal1, Veronika Opletalová1, Jiří Hartl1 , Katarína Král’ová2, and
Miloš Macháček3
1 Department of Medicinal Chemistry and Drug Control, Charles University, Faculty of Pharmacy,
500 05 Hradec Králové, Czech Republic;
2 Institute of Chemistry, Faculty of Natural Sciences, Comenius University, Bratislava, Slovak
Republic
3 Department of Inorganic and Organic Chemistry, Charles University, Faculty of Pharmacy,
500 05 Hradec Králové, Czech Republic
* Author to whom correspondence should be addressed; e-mail miletin@faf.cuni.cz, tel. +420 49
5067387, fax +420 49 551 24 23.
Received: 16 February 2001; in revised form 28 May 2001 / Accepted: 1 June 2001 / Published: 30
June 2001
__________________________________________________________________________________
Abstract: Many compounds containing a -CONH- group display photosynthesis
inhibiting activity. Based on this structural feature, a group of anilides of 2-alkylthio-
(1b-4f) or 2-chloro-6-alkylthio-4-pyridinecarboxylic acids (5a-6c) was synthesised.
The prepared compounds were tested for their inhibition of the oxygen evolution rate
(OER) in spinach chloroplasts.
A
quasi-parabolic dependence between
photosynthesis-inhibiting activity and the lipophilicity of the compounds was
determined for 1b-4f as well as for 5a-6c. The inhibitory activity of compounds 1b-4f
was higher than that of 5a-6c for comparable lipophilicity values.
Keywords: 2-Alkylthio-4-pyridinecarboxylic acids; Anilides; Photosynthesis inhibition
__________________________________________________________________________________

Molecules 2001, 6
604
Introduction
Many herbicides acting as photosynthesis inhibitors possess in their molecules an >N-C(=X)- group
(X=O or N, not S) and a hydrophobic residue in close vicinity to this group. Shipman concluded that
the hydrophilic part of a herbicide binds electrostatically to the terminus of an α-helix at a highly
charged amino acid, whereas the hydrophobic part of the inhibitors extends into the hydrophobic part
of the membrane [1]. Recently, pronounced photosynthesis-inhibiting activity has been found for
alkoxy substituted phenylcarbamates [2,3] as well as for the local anesthetic of the anilide type –
trimecaine [4,5,6], i.e., for compounds with -CONH- groups in their molecules.
In order to bring together the structural features of the abovementioned compounds and our
previous results with 2-alkyl-4-pyridinecaboxylic acids anilides [7], some anilides of 2-alkylthio-4-
pyridinecarboxylic (1b-4f) and 2-chloro-6-alkylthio-4-pyridinecarboxylic acids (5a-6c) were
synthesised.
Results and Discussion
The synthesis of the anilides is shown in Schemes 1 and 2. The 2-alkylthio-4-cyanopyridines were
synthesised as described previously [8]. Subsequent treatment with ethanolic sodium hydroxide
solution afforded the corresponding acids 1-4. 2-chloro-6-alkylthio-4-pyridinecarboxylic acids 5 and 6
were obtained by a similar procedure from 2-chloro-6-alkylthio-4-carbamoylpyridines [9]. The
anilides were prepared from the acids by reaction of the corresponding acyl chlorides with substituted
anilines and aminophenols (Scheme 3). The melting points, yields, and elemental analyses for
compounds are given in Tables 2 and 4, and IR and 1H-NMR spectroscopic data in Tables 3 and 5.
N
Cl
N
SR
N
SR
ii,iii
i
CN
CN
COOH
1-4
iv,v
Reagents: i) Na+ -SR; ii) NaOH; iii) HCl;
iv) SOCl2; v) substituted aniline
N
SR
1b,d; 2b,f; 3a,b,d-f;
4a,b,e,f
Y
X1
X2
CONH
Scheme 1: Preparation of 2-alkylthio-4-pyridinecarboxylic acids and derived anilides.

Molecules 2001, 6
Cl
605
N
SR
Cl
N
SR
Cl
N
SR
i,ii
iii, iv
Y
X1
X2
CONH2
CONH
COOH
5,6
5a-c; 6a-c
Reagents: i) NaOH; ii) HCl; iii) SOCl2; iv) substituted aniline
Scheme 2: Preparation of 2-chloro 6-alkylthio-4-pyridinecarboxylic acids and derived anilides.
Cl
Cl
H2N
OH
H2N
H2N
HO
HO
a
b
c
CF3
Br
Br
H2N
H2N
Br
H2N
OH
CF3
d
e
f
Scheme 3: Aminophenols and anilines used.
The biological activity of anilides of 2-alkylthio-4-pyridinecarboxylic acids (1b-4f) and 2-chloro-6-
alkylthio-4-pyridinecarboxylic acids (5a-6c) with regards to inhibition of oxygen evolution rate in
spinach chloroplasts was investigated. The inhibitory activity of the compounds has been expressed by
IC50 values (Table 1). The IC50 values, i.e. molar concentrations of the compounds causing 50 %
activity decrease with respect to the untreated control, varied in the range of 6.0 - 69.1 µmol.dm-3 for
1b-4f and 14.2 - 32.5 µmol.dm-3, respectively, for 5a-6c.
A quasi-parabolic dependence of photosynthesis-inhibiting activity upon the lipophilicity (log P) of
the compounds was determined for 1b-4f as well as for 5a-6c (Table 1). The comparison of the
biological activity of compounds 1b-4f and 5a-6c having the same lipophilicity showed that the
introduction of a halogen substituent in the 6 position led to a partial decrease of the biological
activity. The previous study with anilides of 2-alkyl-4-pyridinecarboxylic acids showed that the site of
their inhibitory action is the intermediate Z+/D+ corresponding to the tyrosine radicals TyrZ and TyrD

Molecules 2001, 6
606
which are situated at 161th position in D1 and D2 proteins located on the donor side of photosystem
(PS) 2 [10]. The same site of action in the photosynthetic apparatus of spinach chloroplasts can also be
expected for the studied compounds 1b-4f and 5a-6c.
From the quasi-parabolic course of the dependence log (1/IC50) vs. log P it can be assumed that the
most active inhibitors are compounds with sufficiently high lipophilicity for securing their passage
through the lipidic parts of the biological membranes, but enabling also their sufficiently high
concentration in the aqueous phase. This is necessary for their interaction with the intermediates Z+/D+
situated at the lumenal side of photosynthetic membranes in D1 and D2 proteins [11].
Table 1. IC50 values for inhibition of oxygen evolution
rate in spinach chloroplasts and calculated logP
values of the compounds tested..
IC50 · 106
(mol.dm-3)
7.3
IC50 · 106
(mol.dm-3)
6.0
Compd.
calculated logP
Compd.
calculated logP
4.55 ± 0.44
4.90 ± 0.45
6.79 ± 0.54
4.24 ± 0.42
5.62 ± 0.44
6.66 ± 0.58
7.50 ± 0.54
3.75 ± 0.43
5.13 ± 0.46
7.01 ± 0.55
4.03 ± 0.44
5.41 ± 0.46
4.64 ± 0.45
4.57 ± 0.44
5.94 ± 0.46
5.17 ± 0.45
1b
2b
2f
3a
3b
3d
3f
4b
4f
8.0
13.9
12.7
4.8
11.3
69.1
10.3
35.2
32.5
14.2
16.2
19.7
18.3
14.2
5a
5b
5c
6a
6b
6c
4a
Acknowledgements
This study was supported by the Ministry of Education, Youth and Sports of the Czech Republic
(Research Plan No. 111600001), by the Grant Agency of Charles University (Grant No. 26/1998-
BCH), and by the Scientific Grant Agency VEGA of the Slovak Republic (No. 1/7262/20).
Experimental
General
Column chromatography was performed on silica gel (Silpearl, Kavalier Votice). Melting points
were determined on a Kofler block, and are uncorrected. IR spectra were recorded on a Nicolet Impact
1
400 spectrometer in chloroform. H-NMR spectra were determined for solutions in CDCl3 or DMSO
(substituted 4-pyridinecarboxylic acids) with a BS 587 (Tesla, Brno) 80 MHz apparatus or a Varian
Mercury - Vx BB 300 spectrometer operating at 300 MHz. Chemical shifts were recorded as δ values
in parts per million (ppm), and were indirectly referenced to tetramethylsilane via the solvent signal
(7.26 for 1H). Multiplicities are given together with the coupling constants (in Hz). Elemental analyses

Molecules 2001, 6
607
were performed on a EA 1110 CHNS-O CE INSTRUMENTS elemental analyser. Lipophilicity of the
compounds was computed using a program ACD/LogP version 1.0 (Advanced Chemistry
Development Inc., Toronto).
Synthesis of 2-alkylthio-4-cyanopyridines and 2-phenylmethylthio-4-cyanopyridine.
2-Chloro-4-cyanopyridine (10 mmol) and the appropriate thiol (10 mmol) were dissolved in 10 mL
of anhydrous N,N-dimethylformamide. To this solution sodium methoxide (10 mmol) in 5 mL
methanol was added dropwise with stirring under a nitrogen atmosphere at 20 °C. The stirring was
continued until TLC (6:1 hexane-ethyl acetate) indicated completion of the reaction. The reaction
mixture was concentrated in vacuo and after evaporation of the solvents, the 2-alkylthio-4-
cyanopyridines or 2-phenylmethylthio-4-cyanopyridine were distilled off from the oily product. The
boiling points corresponded with the previously described [8].
Synthesis of 2-chloro-6-alkylthio-4-carbamoylpyridines
2,6-Dichloro-4-carbamoylpyridine [13] (10 mmol) and the appropriate thiol (10 mmol) were
dissolved in anhydrous N,N-dimethylformamide (10 mL). To the stirred solution sodium methoxide
(10 mmol) in methanol (5 mL) was added dropwise. The reaction mixture was stirred at room
temperature until TLC (2:1 hexane-ethyl acetate) indicated the reaction was complete. The mixture
was then poured into cold water. The crude product was filtered off, purified by column
chromatography (2:1 hexane-ethyl acetate), and recrystallised from aqueous ethanol. The boiling
points and spectral data agreed with those previously described [9].
Synthesis of 2-alkylthio, 2-phenylmethylthio and 2-chloro-6-alkylthio-4-pyridinecarboxylic acids 1-6
The 2- or 2,6-substituted 4-cyano or carbamoylpyridine (10 mmol) in 10 mL of ethanol was mixed
with 25% aqueous sodium hydroxide (30 mmol) and refluxed until the evolution of the ammonia
ceased. The reaction mixture was then diluted with twice its volume of water and acidified with 10%
hydrochloric acid to pH 4-5. The crude product was collected, washed with water, and recrystallised
from aqueous ethanol. TLC for checking of the purity of final products was performed using hexane-
ethyl acetate-acetic acid (50:45:5) as the mobile phase. The yields, melting points, and elemental
analyses are given in Table 2, IR spectral data and 1H-NMR chemical shifts in Table 3.
Table 2. Analytical data of the prepared 2-alkylthio- and
2-chloro-6-alkylthio-4-pyridinecarboxylic acids.
X
N
SR
COOH

Molecules 2001, 6
608
Compd.
Formula
R
X
C3H7
M. p. °C
Yield %
141-143
80
137-139
78
Calculated / Found
% C % H % N %S
M. w.
C9H11NO2S
197.3
C10H13NO2S
211.3
C11H15NO2S
225.3
C13H11NO2S
245.3
C9H10ClNO2S
231.7
C10H12ClNO2S
245.7
%Cl
54.80
54.55
56.85
56.61
58.64
58.48
-
5.62
5.79
6.20
6.41
6.71
6.89
-
7.10
6.95
6.63
6.46
6.22
6.05
-
16.25
16.08
15.17
14.93
14.23
14.02
-
-
-
-
-
1
2
3
4
5
6
iC4H9
C5H11
135-137
82
CH2C6H6
196-197 a)
76
C3H7
Cl
C4H9
Cl
119-120
77
93-95
75
46.66
46.45
48.88
48.67
4.35
4.21
4.92
4.85
6.05
6.19
5.70
5.82
13.84
13.65
13.05
12.87
15.30
15.55
14.43
14.65
a) ref. [12] M. p. 195-196°C
Table 3. IR and 1H-NMR spectroscopic data of the 2-alkylthio and
2-chloro-6-alkylthio-4-pyridinecarboxylic acids (DMSO).
Compd.
1
IR (cm-1)
δ 1H-NMR (ppm)
2975, 2935, 2890 (CH-aliph.) (CDCl3): 1.06 t, J=7.3 Hz, 3H, CH3; 1.76 m, 2H, CH2; 3.18 t,
2480 (COOH)
1725 (CO)
J= 7.3 Hz, 2H, SCH2; 7.56 m, 1H, H-5; 7.78 m, 1H, H-3; 8.59
m, 1H, H-6 a)
2960, 2930, 2870 (CH-aliph.) (CDCl3): 1.05 d, J=6.6 Hz, 6H, 2xCH3; 1.96 m, 1H, CH<; 3.11
2
3
5
6
2470 (COOH)
1725 (CO)
d, J= 6.9 Hz, 2H, SCH2; 7,53 d, J=4.95 Hz, 1H, H-5; 7.78 m,
1H, H-3; 8.59 d, J=4.95 Hz, 1H, H-6 a)
2970, 2925, 2860 (CH-aliph.) (CDCl3 ): 0.91 t, J=7 Hz, 3H, CH3; 1.3 - 1.5 m, 4H, 2xCH2; 1.73
2450 (COOH)
1730 (CO)
m, 2H, CH2; 3.19 t, J=7.3 Hz, 2H, SCH2; 7.56 d, J=4.95 Hz, 1H,
H-5; 7.79 m, 1H, H-3; 8.62 d, J=4.95 Hz, 1H, H-6 a)
2970, 2934, 2874 (CH-aliph.) (DMSO-d6): 1.05 t, J=7.2 Hz, 3H, CH3; 1.47-2.10 m, 2H, CH2;
2658 (COOH)
1705 (CO)
3.18 t, J=7.1 Hz, 2H, SCH2; 7.54 d, J=0.7 Hz, 1H, H-5; 7.67 d,
J=0.7 Hz, 1H, H-3; 11.65 s, 1H, COOH
2962, 2933, 2873 (CH-aliph.) (DMSO-d6): 0.96 t, J=6.2 Hz, 3H, CH3; 1.21-1.98 m, 4H, CH2;
2541 (COOH)
1707 (CO)
3.20 t, J=7.1 Hz, 2H, SCH2; 7.53 d, J=1.1 Hz, 1H, H-5; 7.66 d,
J=1.1 Hz, 1H, H-3; 11.62 s, 1H, COOH
a) A COOH signal was not observed in the 1H-NMR spectrum.

Molecules 2001, 6
609
General method for the synthesis of anilides of 2-alkylthio, 2-phenylmethylthio and 2-chloro-6-
alkylthio-4-pyridinecarboxylic acids (1b,d; 2b,f; 3a,b,d-f; 4a,b,e,f; 5a-c; 6a-c).
A mixture of the 2- or 2,6 substituted-4-pyridinecarboxylic acid (10 mmol) and thionyl chloride
(15 mmol) in 10 mL of dry benzene was refluxed for about 1 h. The excess of thionyl chloride was
removed by repeated evaporation of dry benzene solutions in vacuo. The resulting crude acyl chloride
dissolved in 10 mL of dry acetone was added dropwise to a stirred solution of substituted aniline or
aminophenol (10 mmol) in 10 mL of dry pyridine keeping the temperature at 10 °C. After addition of
the aniline or aminophenol was complete, stirring at 10 °C was continued for another 30 min. The low
temperature was essential in the case of aminophenols in order to avoid the partial esterification of acyl
chloride. The reaction mixture was poured into 40 mL of cold water. Crude anilide was collected and
recrystallised from aqueous ethanol. TLC was performed using hexane-ethyl acetate (50:50) as the
mobile phase. The yields, melting points and elemental analyses of the anilides are given in Table 4,
IR spectral data and 1 H-NMR chemical shifts in Table 5.
Table 4. Analytical data of the prepared anilides.
Z
N
R
Y
1
X
CONH
2
X
1
2
Formula
M. w.
C15H15ClN2O2S
322.8
C15H14Br2N2O2S
446.2
C16H17ClN2O2S
336.8
C18H16F6N2OS
422.4
C17H20N2O2S
316.4
C17H19ClN2O2S
350.9
C17H18Br2N2O2S
474.2
R
Y, Z
SC3H7
2´-OH, H
SC3H7
X
X
M. p. °C
Yield % % C
Calculated / Found
Compd.
1b
% H
4.68
4.52
3.16
3.23
5.09
5.07
3.82
3.72
6.37
6.45
5.46
5.39
3.83
3.81
5.05
4.93
% N % S % Cl(Br,F)
5´-Cl
H
3´-Br
5´-Br
5´-Cl
H
3´-CF3 164-165
5´-CF3
H
H
5´-Cl
H
3´-Br
5´-Br
4´-Br
H
161-163
57
55.81
55.96
40.38
40.41
57.05
57.11
51.18
51.31
64.53
64.48
58.19
58.23
43.06
43.15
53.83
53.97
8.68
8.49
6.28
6.22
8.32
8.27
6.63
6.48
8.85
8.71
7.98
7.88
5.91
5.77
7.39
7.23
9.93
10.06
7.19
7.12
9.52
9.56
7.59
7.75
10.13
10.28
9.14
9.19
6.76
6.67
8.45
8.31
10.98
10.76
35.82
35.71
10.53
10.45
26.99
26.78
-
152-153
65
1d
2b
2f
4´-OH, H
S-iC4H9
2´-OH, H
S-iC4H9
H, H
SC5H11
2´-OH, H
SC5H11
2´-OH, H
SC5H11
4´-OH, H
SC5H11
H, H
162-164
58
54
123-125
60
3a
3b
3d
3e
153-155
58
10.10
10.01
33.70
33.50
21.07
20.88
120-122
67
C17H19BrN2OS
379.3
94-95
52

Molecules 2001, 6
610
Table 4 (cont.)
C19H18F6N2OS
SC5H11
H, H
3´-CF3 122-124
52.29
52.16
67.84
67.57
61.54
61.65
57.15
57.31
55.26
55.38
55.81
55.68
50.43
50.33
50.43
50.29
57.05
56.91
51.76
51.58
51.76
51.63
4.16
4.21
4.79
4.97
4.08
3.86
3.79
3.71
3.09
3.01
4.68
4.51
3.95
3.91
3.95
3.86
5.09
5.02
4.34
4.31
4.34
4.23
6.42
6.36
8.33
8.09
7.55
7.41
7.02
6.87
6.14
6.03
8.68
8.79
7.84
7.72
7.84
7.95
8.32
8.48
7.55
7.68
7.55
7.71
7.35
7.21
9.53
9.75
8.66
8.72
8.03
8.14
7.02
7.18
9.93
9.72
8.97
8.85
8.97
8.81
9.52
9.39
8.63
8.41
8.63
8.38
26.12
25.95
-
3f
4a
4b
4e
4f
436.4
C19H16N2O2S
336.4
C19H15ClN2O2S
370.9
C19H15BrN2OS
339.3
C21H14F6N2OS
456.4
C15H15ClN2O2S
322.81
C15H14Cl2N2O2S
357.25
C15H14Cl2N2O2S
357.25
C16H17ClN2O2S
336.84
5´-CF3
H
56
149-150
60
SCH2C6H5
2´-OH, H
SCH2C6H5
2´-OH, H
SCH2C6H5
H, H
SCH2C6H5 3´-CF3 141-143
H, H
SC3H7
2´-OH, Cl
SC3H7
2´-OH, Cl
SC3H7
4´-OH, Cl
SC4H9
2´-OH, Cl
SC4H9
H
5´-Cl
H
194-196
78
9.56
9.38
4´-Br
H
108-109
55
20.01
19.85
24.98
24.75
10.98
11.21
19.85
20.07
19.85
20.03
10.53
10.79
19.10
19.31
19.10
19.33
5´-CF3
H
61
138-139
44
5a
5b
5c
6a
6b
6c
H
5´-Cl
H
152-153
53
3´-Cl
H
144-146
34
H
H
123-125
56
C16H16Cl2N2O2S
371.28
C16H16Cl2N2O2S
371.28
5´-Cl
H
158-160
59
2´-OH, Cl
SC4H9
4´-OH, Cl
3´-Cl
H
115-117
53
Table 5. IR and 1H-NMR spectroscopic data of the prepared anilides.
Compd.
1b
IR (cm-1)
δ 1H-NMR (ppm)
2965, 2932, 2873 (DMSO-d6): 1.00 t, J=7, 3H, CH3; 1.66 m, 2H, CH2; 3.18 t, J=7, 2H, SCH2;
(CH aliph.),
1651 (CO)
6.93 d, J=8.5, 1H, H-3´; 7.12 dd, J=8.5, J=2.4, 1H, H-4´; 7.53 dd, J=5.1,
J=1.5, 1H, H-5; 7.73 qs, 2H, H-3 and H-6´; 8.60 d, J=5.1, 1H, H-6; 9.82 s,
1H, OH or NH; 10.08 s, 1H, NH or OH
2975, 2945, 2890 (DMSO-d6): 1.00 t, J=7, 3H, CH3; 1.75 m, 2H, CH2; 3.18 t, J=7, 2H, SCH2;
1d
2b
(CH aliph.),
1650 (CO)
7.52 d, J=5.1, 1H, H-5; 7.71 s, 1H, H-3; 8,00 s, 2H, H-2´and H-6´; 8.62 d,
J=5.1, 1H, H-6; 10.45 s, 1H, OH or NH; 10.51 s, 1H,NH or OH
2975, 2940, 2890 (DMSO-d6): 1.01 d, J=7, 6H, 2xCH3; 1.92 m, 1H, -CH<; 3.13 t, J=7.5, 2H,
(CH aliph.),
1655 (CO)
SCH2; 6.95 d, J=8.5, 1H, H-3´; 7.13 dd, J=8.5, J=2.5, 1H, H-4´; 7.52 dd, J=5,
J=1, 1H, H-5; 7.73 dd, J=1, J<1, 1H, H-3; 7.74 d, J=2.5, 1H, H-6´; 8.59 dd,
J=5, J<1, 1H, H-6
2962, 2929, 2869 (CDCl3) 1.05 d, J=6.4, 6H, 2xCH3; 1.94 m, 1H, CH; 3.14 d, J=6.7, 2H,
2f
(CH aliph.),
1662 (CO)
SCH2; 7.31 dd, J=5.2, J=1.5, 1H, H-5; 7.55 d, J=1.5, 1H H-3; 7.68 s, 1H, H-
4´; 8.17 s, 3H, H-2´, H-6´ and NH; 8.57 d, J=5.2, 1H, H-6

Molecules 2001, 6
611
Table 5 (cont.)
2958, 2931, 2859 (DMSO-d6): 0.88 dist.t, CH3; 1.38 m, 4H, 2xCH2; 1.66 m, 2H, CH2; 3.19 t,
3a
(CH aliph.),
1652 (CO)
2H, CH2; 6.96 m, 3H, arom.; 7.55 m, 2H, H-5 and 1H arom.; 7.74 s, 1H, H-
3; 8.59 d, J=5.1, 1H, H-6; 9.66 s, 1H, OH; 9.77 bs, 1H, NH
2975, 2940, 2870 (DMSO-d6): 0.88 dist. t, 3H, CH3; 1.36 m, 4H, (CH2)2; 1.63 m, 2H, CH2;
3b
(CH aliph.),
1640 (CO)
3.20 t, J=7, 2H, SCH2; 7.13 dd, J=8.5, J=2.5, 1H, H-4’; 7.53 dd, J=5, J=1.5,
1H, H-5; 7.72 dd, J=1.5, J=1, 1H, H-3; 7.75 d, J=2.5, 1H, H-6´; 8.61 dd, J=5,
J=1, 1H, H-6
2980, 2945, 2875 (DMSO-d6): 0.88 dist. t, 3H, CH3; 1.37 m, 4H, (CH2)2; 1.66 m, 2H, CH2;
3d
(CH aliph.),
1655 (CO)
3.19 t, J=7, 2H, SCH2; 7.52 d, J=5.1, 1H, H-5; 7.71 s, 1H, H-3; 8,00 s, 2H,
H-2´and H-6´; 8.61 d, J=5.1, 1H, H-6; 9.84 s, 1H, OH or NH; 10.44 s, 1H,
NH or OH
2952, 2926, 2852 (CDCl3) 0.91 dist. t, 3H, CH3; 1.38 m, 4H 2xCH2; 1.71 m, 2H, CH2;
3e
3f
(CH aliph.),
1658 (CO)
3.19 t, J=7, 2H, SCH2; 7.27 dd, J=5, J=1.5, 1H, H-5; 7.49 qs, 5H, H-3 and
C6H4; 8.01 bs, 1 H, NH; 8.52 d, J= 5, 1H, H-6
2958, 2930, 2858 (CDCl3) 0.91 dist. t, 3H, CH3; 1.38 m, 4H, 2xCH2; 1.71 m, 2H, CH2; 3.22 t,
(CH aliph.),
1669 (CO)
1660 (CO)
J=7, 2H, SCH2; 7.32 dd, J=5.2, J=1.5, 1H, H-5; 7.55 d, J=1.5, 1H, H-3; 7.68
s, 1H, H-4´; 8.17 qs, 3H, H-2´, H-6´and NH; 8.58 d, J=5.2, 1H, H-6
(CDCl3) 4.49 s, 2H, SCH2; 6.95 m, 1H, H-4´ or H-5´; 7.03 m, 1H, H-3´ or H-
6´; 7.16 m, 1H, H-5´ or H-4´; ca.7.25 m overlapped with the signal of
solvent, H-6´ or H-3´; 7.27-7.45 m, 5H, C6H5; 7.39 overlapping dd, J=5.2
Hz, J=1.6 Hz, 1H, H-5; 7.58 m, 1H, H-3; 7.75 bs, 1H, NH; 8.14 bs, 1H, OH;
8.63 d, J=5.2 Hz, 1H, H-6
4a
1655 (CO)
(CDCl3) 4.49 s, 2H, SCH2; 6.95 d, J=8.7 Hz, 1H, H-3´; 7.11 dd, J=8.7 Hz,
J=2.5, 1H, H-4´; 7.27-7.45 m, 5H, C6H5; 7.37 overlapping dd, J=5.2 Hz,
J=1.6 Hz, 1H, H-5; 7.55 m, 1H, H-6´; 7.56 m, 1H, H-3; 8.09 bs, 1H, NH;
8.64 dd, J=5.2 Hz, J=0.8 Hz,1H, H-6
4b
1653 (CO)
1668 (CO)
(CDCl3) 4.45 s, 2H, SCH2; 7.31 m, 6H, H-5 and C6H5; 7.45 qs, 5H, H-3 and
C6H4; 7.98 bs, 1 H, NH; 8.53 d, J= 5.2, 1H, H-6
(CDCl3) 4.46 s, 2H, SCH2; 7.36 m, 7H, H-3, H-5 and C6H5; 7.67 s, 1H, H-4´;
8.10 s, 2H, H-2´and H-6´; 8.24 bs, 1 H, NH; 8.57 d, J=5.2, 1H, H-6
4e
4f
2965, 2931, 2872 (DMSO-d6): 0.86-1.33 m, 3H, CH3; 1.50-2,02 m, 2H, CH2; 3.20 t,
5b
(CH aliph.),
1655 (CO)
J=7.2 Hz, 2H, CH2; 6.90 d, J=8.5 Hz, 1H, H-3´; 7.00-7.23 m, 1H,
NH; 7.10 dd, J1=2.2 Hz, J2=8.5 Hz, 1H, H-4´; 7.34 d, J=1.2 Hz, 1H,
H-5; 7.46 d, J=1.2 Hz, 1H, H-3; 7.69 d, J=2.2 Hz, 1H, H-6´; 8.18 s,
1H, -OH
2965, 2931, 2872 (DMSO-d6): 0.90-1.18 m, 3H, CH3; 1.46-2.04 m, 2H, CH2; 3.18 t,
5c
(CH aliph.),
1654 (CO)
J=7.1 Hz, CH2; 5.57 s, 1H, NH; 6.98 d, J=8.8 Hz, 1H, H-5´; 7.28 dd,
J1=2.4 Hz, J2=8.8 Hz, 1H, H-6´; 7.29 d, J=1.2 Hz, 1H, H-5; 7.41 d,
J=1.2 Hz, 1H, H-3; 7.73 d, J=2.4 Hz, 1H, H-2´; 7.83 s, 1H, OH

Molecules 2001, 6
612
Table 5 (cont.)
2958, 2931, 2872 (DMSO-d6): 0.78-1.10 m, 3H, CH3; 1.5-1.95 m, 4H, CH2CH2; 3.18 t,
6a
(CH aliph.),
1646 (CO)
J=7.0 Hz, 2H, CH2; 6.70-7.26 m, 3H, H-4´, H-5´, H-6´; 7.32 d, J=1.0
Hz, 1H, H-5; 7.43 d, J=1,2 Hz, 1H, H-3; 7.58 d, J=7.8 Hz, 1H, H-3´;
7.72 s, 1H, NH; 8.47 s, 1H, OH
2959, 2931, 2872 (DMSO-d6): 0.73-1.32 m, 3H, CH3; 1.50-2.02 m, 4H, CH2CH2; 3.22 t,
6b
(CH aliph.),
1656 (CO)
J=7.2 Hz, 2H, CH2; 6.90 d, J=8.3 Hz, 1H, H-3´; 7.00-7.23 m, 1H,
NH; 7.10 dd overlapped by another signal, J1=2.2 Hz, J2=8.5 Hz, 1H,
H-4´; 7.35 d, J=1.2 Hz, 1H, H-5; 7.45 d, J=1.2 Hz, 1H, H-3; 7.69 d,
J=2.2 Hz, 1H, H-6´; 8.14 s, 1H, OH
2959, 2930, 2872 (DMSO-d6): 0.96 t, J=6.4 Hz, 3H, CH3; 1.14-1.93 m, 4H, CH2CH2;
6c
(CH aliph.),
1649 (CO)
3.19 t, J=7.1 Hz, CH2; 5.62 s, H, NH; 7.27 dd ovelapped by d, J1=2.4
Hz, J2=8.8 Hz, 2x1 H, H-5´, H-6´; 7.29 d, J=1.2 Hz, 1H, H-5; 7.40 d,
J=1.2 Hz, 1H, H-3; 7.73 d, J=2.4 Hz, 1H, H-2´; 7.91 s, 1H, OH
Biological assays
Chloroplasts were prepared from locally purchased spinach according to the procedure described by
Walker [14]. The oxygen evolution rate (OER) in spinach chloroplasts was determined
spectrophotometrically (Specord UV VIS Zeiss Jena, Germany) by the Hill reaction. The
measurements were carried out in phosphate buffer (20 mmol, pH = 7.2) containing sucrose (0.4
mol.dm-3), MgCl2 (5 mmol.dm-3) and NaCl (15 mmol.dm-3) using 2,6-dichlorophenol-indophenol as
electron acceptor.
Chlorophyll content in the samples was 30 mg.dm-3 and the samples were irradiated (~ 100 W.m-2)
from 10cm distance with a halogen lamp (250 W) using a water filter to prevent warming of the
o
samples (suspension temperature 22 C). The compounds were dissolved in dimethyl sulfoxide
(DMSO) because of their limited water solubility. The applied DMSO concentration (up to 5 %) did
not affect OER.
References
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Molecules 2001, 6
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Sample availability: Supporting samples of the following compounds are available at MDPI: 5, 1b, 2f,
3b, 3e, 3f, 5a-c, 6a-c.
© 2001 by MDPI (http://www.mdpi.org). Reproduction is permitted for noncommercial purposes