N-substituted hydroxyureas as urease inhibitors

Article abstract of DOI:10.1248/cpb.50.1280

In order to seek a urease inhibitor more potent than hydroxyurea (1), its alkyl- or phenyl-substituted derivatives were synthesized and evaluated for their effect on the jack bean urease. Of 16 compounds tested, m-methyl-(10) and m-methoxy-phenyl substituted hydroxyurea (13) showed the most potent inhibitory activities against the enzyme.

Full text of DOI:10.1248/cpb.50.1280

1280
Notes
Chem. Pharm. Bull. 50(9) 1280—1282 (2002)
Vol. 50, No. 9
N-Substituted Hydroxyureas as Urease Inhibitors
Shinichi UESATO, ,a Yuichiro HASHIMOTO,a Masaru NISHINO,a Yasuo NAGAOKA,a and
*
b
Hiroshi KUWAJIMA
a Department of Biotechnology, Faculty of Engineering, Kansai Univeristy; 3–3–35 Yamate-cho, Suita, Osaka 564–8680,
Japan: and b Faculty of Pharmaceutical Sciences, Kinki University; 3–4–1 Kowakae, Higashiosaka, Osaka 577–8502,
Japan. Received April 22, 2002; accepted June 10, 2002
In order to seek a urease inhibitor more potent than hydroxyurea (1), its alkyl- or phenyl-substituted deriva-
tives were synthesized and evaluated for their effect on the jack bean urease. Of 16 compounds tested, m-methyl-
(10) and m-methoxy-phenyl substituted hydroxyurea (13) showed the most potent inhibitory activities against the
enzyme.
Key words urease inhibitor; N-hydroxy-NЈ-(m-methylphenyl)urea; Helicobacter pylori; jack bean
t0: time interval (s) measured at zero molar sample concentration
Persons infected with Helicobacter pylori are exposed to a
high risk for stomach cancer. As eradication therapy, a com-
bination of antibiotics and a proton inhibitor has been
adopted. However, this therapy is unsatisfactory due to the
As a result, substitution with the lower alkyl groups at 1
led to a marked decrease in the inhibitory power [IC50 (mM):
Ͼ800 (2); Ͼ800 (3); 706 (4)] against the jack bean urease.
induction of drug-resistance. Thus, the urease activity of H.
However, N-phenyl-substituted derivative 5 inhibited the en-
pylori may be an attractive target for drug design of an anti-
zyme to the same extent [IC50 (mM): 19.3 mM] as did the lead
H. pylori agent. Hydroxyurea (1), an antineoplastic agent,1)
compound, hydroxyurea (1) [22.3 mM]. In the next experi-
ment, the o-, m- and p-substituted derivatives of 5 shown
below were thus synthesized in order to examine the effect of
substituents at the phenyl group: N-(fluorophenyl)-NЈ-hy-
has been known to inhibit various ureases from microorgan-
isms and vegetables, including H. pylori2,3) and jack bean.4)
In the present work, we have searched for a potent therapeu-
tic drug through the derivatization of 1 to N-substituted hy-
droxyureas (6—8), N-(o-methylphenyl)- (9),6) N-(m-methyl-
droxyureas.
phenyl)- (10) and N-(p-methylphenyl)-NЈ-hydroxyurea (11),12)
Phenyl- or low alkyl-substituted hydroxyureas were first
synthesized as shown in Table 1. Thus, aniline was converted
to the isocyanate with triphosgene in the two-phase system of
N-(o-methoxyphenyl)- (12),6) N-(m-methoxylphenyl)- (13)13)
and N-(p-methoxylphenyl)-NЈ-hydroxyurea (14)12) were pre-
pared via the N-benzyloxy derivatives 6a—14a, respectively,
CH2Cl2 and aq. Na2CO3, which was treated with O-benzylhy-
in the same two-phase system as for 5. N-Hydroxy-NЈ-(o-ni-
droxylamine to give N-(benzyloxy)-NЈ-phenylurea (5a). This
trophenyl)- (15), N-hydroxy-NЈ-(m-nitrophenyl)- (16) and N-
compound was hydrogenated over 10% Pd–C to give the de-
hydroxy-NЈ-(p-nitrophenyl)-urea (17)12) were synthesized
sired N-hydroxy-NЈ-phenylurea5) (5). Further, N-ethyl-NЈ-hy-
using the procedure of Harmon, et al.,6) but with the addition
droxyurea6) (2), N-hydroxy-NЈ-propylurea6) (3) and N-butyl-
NЈ-hydroxyurea7) (4) were prepared by treating the corre-
Table 1. Synthesis of Substituted Hydroxyureas
sponding isocyanates with hydroxylamine hydrochloride in
aq. NaOH solution, respectively, according to the procedure
by Harmon, et al.6)
The inhibitory activities against jack bean urease were
measured as explained in a separate paper8): the control solu-
tion was prepared by dissolving Phenol Red and blank sol-
Yield (%)
vent (H2O, EtOH, or dimethyl sulfoxide (DMSO)) in 0.1 M
RϭH
C2H5
C3H7
C4H9
Ph
o-F-Ph
m-F-Ph
p-F-Ph
—
—
—
—
5a
6a
7a
8a
9a
10a
11a
12a
13a
14a
—
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
—
phosphate buffer (pH 7.7), whereas the sample solution was
made by dissolving Phenol Red and a test sample in a blank
solvent in 0.1 M phosphate buffer (pH 6.7). These control and
sample solutions, after addition of the urease solution, were
successively pre-incubated at 30 °C for 30 min, and diluted 6-
fold with the control buffer of pH 7.7 and the buffered urea
solution of pH 6.7, respectively, according to the colorimetric
timing method.9—11) The absorbance at 560 nm of the result-
ing control solution was adjusted to zero on the spectropho-
tometer, and the time interval for the absorbance of the sam-
ple solution to reach zero was measured at 560 nm. The per-
centage inhibition (%) of each sample was calculated based
on the following equation.8)
50.7a)
15.6a)
54.8a)
59.0b)
48.6b)
42.3b)
57.3b)
42.8b)
38.6b)
78.7b)
53.5b)
21.3b)
72.3b)
21.4a)
8.9a)
o-Me-Ph
m-Me-Ph
p-Me-Ph
o-MeO-Ph
m-MeO-Ph
p-MeO-Ph
o-NO2-Ph
m-NO2-Ph
p-NO2-Ph
—
—
11.8a)
percentage inhibition (%)ϭ[(tϪt0)/t]ϫ100
a) Based on the corresponding isocyanates. b) Based on the corresponding benzyl-
oxyureas.
t: time interval (s) measured at each molar sample concentration
To whom correspondence should be addressed. e-mail: uesato@ipcku.kansai-u.ac.jp
© 2002 Pharmaceutical Society of Japan

September 2002
1281
Table 2. Effects of Substitution at Phenylhydroxyureas on Jack Bean Ure-
ase (5: IC50ϭ19.3 mM)
N-(Benzyloxy)-NЈ-(o-fluorophenyl)urea (6a): Colorless plates (1010 mg,
53.3%). mp 101.8—102.3 °C (AcOEt). IR (KBr) cmϪ1: 3166, 3074, 2888,
1688, 1618, 1537, 1491, 1454, 1371, 1329, 1188, 1072, 984, 820, 752, 700.
1H-NMR (CD3OD) d: 4.87 (2H, s, –CH2–), 7.0—7.9 (9H, m, arom. H9). EI-
MS m/z: 260 (Mϩ).
IC50 (mM)
Substituent
ortho
meta
para
N-(o-Fluorophenyl)-NЈ-hydroxyurea (6): Colorless plates (320.7 mg). mp
140—142.8 °C (AcOEt). IR (KBr) cmϪ1: 3227, 2846, 1648, 1559, 1458,
1420, 1251, 1084, 752, 638. 1H-NMR (CD3OD) d: 7.03—7.17 (3H, m),
7.90—7.98 (1H, m). HR-EI-MS m/z: 170.0474 (Mϩ, Calcd for C7H7FN2O2:
170.0492).
N-(Benzyloxy)-NЈ-(m-fluorophenyl)urea (7a): Colorless needles (762.8
mg, 40.3%). mp 113—114.5 °C (AcOEt). IR (KBr) cmϪ1: 3321, 3206, 2860,
1661, 1597, 1537, 1493, 1452, 1362, 1312, 1277, 1252, 1148, 1072, 973,
770. 1H-NMR (CD3OD) d: 4.56 (2H, s, –CH2–), 6.7—7.5, m, arom. H9). EI-
MS m/z: 260 (Mϩ).
–F
37.7
121.3
191.7
N.D.a)
35.7
12.7
48.0
152.8
340.0
533.3
–CH3
–OCH3
–NO2
16.2
N.D.a)
a) Not determined due to insolubility in the assay system.
of MeOH to improve the solubility of the starting isocyanates
in the reaction mixture. Table 2 illustrates the substitutional
and positional effects of F-, CH3-, CH3O- and NO2-groups at
the phenyl group. Introduction of these substituents at the o-
and p-position significantly decreased the anti-urease activ-
ity. This can be interpreted on the basis of: i) steric hindrance
by the ortho-substitution, which disturbs the hydroxamic acid
group from accessing the active site. ii) steric compression
due to bulky groups at the para-position, which hampers the
contact between the substituted phenyl group and a hy-
drophobic cavity around the active site. Substitution with F
did not significantly influence their activities, presumably
due to its small atomic radius. In contrast, m-methyl- and m-
methoxy-phenyl derivatives (10, 13) had a high potency, in-
hibiting the enzyme more strongly than 5, as well as 1,
though to a small degree (IC50 12.7 or 16.2 mM). We plan to
examine further the potency of these compounds as therapeu-
tic drugs.
N-(m-Fluorophenyl)-NЈ-hydroxyurea (7): Colorless columns (210.8 mg).
mp 103.4—105.7 °C (AcOEt). IR (KBr) cmϪ1: 3228, 2852, 1684, 1560,
1
1450, 1406, 1278, 1144, 763, 638. H-NMR (CD3OD) d: 6.65—6.78 (1H,
m), 7.22—7.26 (2H, m), 7.41—7.47 (1H, m). HR-EI-MS m/z: 170.0500
(Mϩ, Calcd for C7H7FN2O2: 170.0492).
N-(Benzyloxy)-NЈ-(p-fluorophenyl)urea (8a): Colorless fine crystals
(541.8 mg, 28.6%). mp 122.4—124.5 °C (AcOEt). IR (KBr) cmϪ1: 3312,
1
3223, 2926, 2860, 1659, 1524, 1412, 1211, 1074, 939, 831, 808, 698. H-
NMR (CD3OD) d: 4.54 (2H, s, –CH2–), 7.0—7.5 (9H, m, arom. H9). EI-MS
m/z: 260 (Mϩ).
N-(p-Fluorophenyl)-NЈ-hydroxyurea (8): Colorless fine crystals (202.7
mg). mp 135.6—137.1 °C (AcOEt). IR (KBr) cmϪ1: 3238, 1654, 1559,
1509, 1412, 1213, 1093, 830. 1H-NMR (CD3OD) d: 6.93—7.05 (2H, m),
7.29—7.50 (2H, m), 8.81. HR-EI-MS m/z: 170.0500 (Mϩ, Calcd for
C7H7FN2O2: 170.0492).
N-(Benzyloxy)-NЈ-(o-methylphenyl)urea (9a): Colorless needles (710 mg,
33.2%). mp 126.7—127.0 °C (AcOEt). IR (KBr) cmϪ1: 2862, 1686, 1533,
1
1458, 1252, 1074, 968, 916, 754, 702. H-NMR (CD3OD) d: 2.15 (3H, s,
CH3), 4.89 (2H, s, –CH2–), 7.0—7.5 (9H, m, arom. H9). EI-MS m/z: 256
(Mϩ).
N-Hydroxy-NЈ-(o-methylphenyl)urea6) (9): Colorless needles (517.8 mg).
mp 233.1—235.3 °C (AcOEt). IR (KBr) cmϪ1: 3236, 2887, 2359, 2340,
1
Experimental
1647, 1612, 1458, 1354, 1288, 1248, 1082, 1042, 746. H-NMR (CD3OD)
All melting points (mp) were determined on a Yanagimoto MP-32 mi- d: 2.27 (3H, s, CH3), 7.0—7.5 (4H, m, arom. H4). HR-EI-MS m/z: 166.0727
cromelting point apparatus and are uncorrected. IR spectra were recorded on (Mϩ, Calcd for C8H10N2O2: 166.0742).
a Perkin-Elmer 1600 series FT-IR spectrometer, and low resolution (LR)-
N-(Benzyloxy)-NЈ-(m-methylphenyl)urea (10a): Colorless needles
1
FAB-MS on a JEOL JMS-HX 100 instrument. H-NMR (300 MHz) spectra (1012 mg, 47.5%). mp 96.3—97.2 °C (AcOEt). IR (KBr) cmϪ1: 2912, 2858,
were recorded on a Bruker AX-300 instrument, using tetramethylsilane 1611, 1541, 1489, 1420, 1362, 1292, 1175, 1088, 951, 905, 878, 824, 775,
(TMS) as a reference. High resolution (HR)- and LR-electron impact (EI)- 694, 627. 1H-NMR (CD3OD) d: 2.29 (3H, s, CH3), 4.86 (2H, s, –CH2–),
MS were measured with JEOL The Tandem MStation JMS-700. Chromatog- 6.8—7.5 (9H, m, arom. H9). EI-MS m/z: 256 (Mϩ).
raphy was carried out using silica gel 60 (Kanto Chemical, 70—230 mesh).
N-Hydroxy-NЈ-(m-methylphenyl)urea (10): Colorless needles (467.0 mg).
Analytical TLC and Preparative TLC were performed using Silica gel 60 mp 106.8—107.7 °C (AcOEt). IR (KBr) cmϪ1: 3383, 2883, 1647, 1555,
F254 (Merck, 0.25 mm) and Silica gel 60 F254 (Merck, 2 mm) glass plates, re- 1489, 1437, 1250, 1173, 1090, 891, 866, 777. HR-EI-MS m/z: 166.0749
spectively. Phenol Red solution (1 g/l phenol red in 47 vol% ethanol) was (Mϩ, Calcd for C8H10N2O2: 166.0742).
purchased from Kanto Chemical Co., Inc. (Tokyo, Japan). Urease (115
N-(Benzyloxy)-NЈ-(p-methylphenyl)urea (11a): Colorless needles (1488
units/mg derived from Jack bean) was obtained from Wako Pure Chemical mg, 64.5%). mp 106—106.8 °C (AcOEt). IR (KBr) cmϪ1: 3331, 3206, 3031,
Industries, Ltd. (Osaka, Japan). Hydroxyurea and isocyanates were pur- 2916, 2860, 1659, 1591, 1537, 1454, 1412, 1360, 1325, 1236, 1074, 945,
chased from Sigma-Aldrich Japan Co., Inc. (Tokyo).
814, 745, 694. 1H-NMR (CD3OD) d: 2.19 (3H, s, CH3), 4.80 (2H, s,
Preparation of N-Substituted Hydroxyureas (5—14) These com- –CH2–), 7.0—7.4 (9H, m, arom. H9). EI-MS m/z: 256 (Mϩ).
pounds were prepared via the corresponding N-(benzyloxy)ureas. In a typi-
N-Hydroxy-NЈ-(p-methylphenyl)urea12) (11): Colorless columns (760.7
cal experiment, a solution of Na2CO3 (14.56 mmol) in water (100 ml) was mg). mp 149—153.5 °C (AcOEt). IR (KBr) cmϪ1: 3240, 2918, 2540, 1896,
added to a solution of triphosgene (2.67 mmol) and non-substituted or sub- 1641, 1597, 1555, 1416, 1331, 1232, 1082, 1003, 870, 820, 704. HR-EI-MS
stituted aniline (7.28 mmol) in CH2Cl2 (100 ml), and the mixture was stirred m/z: 166.0727 (Mϩ, Calcd for C8H10N2O2: 166.0742).
vigorously at room temp. for 1.5 h. Then, a solution of O-benzylhydroxyl-
N-(Benzyloxy)-NЈ-(o-methoxyphenyl)urea (12a): Colorless columns (1136
amine (14.56 mmol) in MeOH (50 ml) was added to the mixture, which was mg, 57.3%). mp 89.0—90.2 °C (n-hexane–AcOEt). IR (KBr) cmϪ1: 3078,
stirred for another 15 h. The CH2Cl2 layer was separated, dried over MgSO4 2839, 1680, 1601, 1537, 1489, 1462, 1366, 1250, 1215, 1175, 1119, 986,
and concentrated in vacuo to give N-(benzyloxy)urea. This compound was 918, 810, 702, 665, 631. 1H-NMR (CD3OD) d: 3.87 (3H, s, OCH3), 4.86
dissolved in MeOH (60 ml) and hydrogenated over 10% Pd–C (90 mg) at (2H, s, –CH2–), 6.8—8.0 (9H, arom. H9). EI-MS m/z: 272 (Mϩ).
room temp. for 3 h. After removal of the catalyst, MeOH was evaporated in
vacuo, affording a desired substituted hydroxyurea.
N-Hydroxy-NЈ-(o-methoxyphenyl)urea6) (12): Powdery crystals (709.6
mg). mp 163.2—165.3 °C (AcOEt). IR (KBr) cmϪ1: 3215, 2841, 2340,
N-(Benzyloxy)-NЈ-phenylurea (5a): Colorless needles (1550 mg, 87.7%). 1643, 1595, 1547, 1462, 1427, 1259, 1215, 1177, 1124, 1082, 1047, 1003,
mp 106—107.6 °C (AcOEt). IR (KBr) cmϪ1: 3323, 3204, 2860, 1659, 1537,
1450, 1335, 1236, 1070, 949, 905, 789, 691. 1H-NMR (CD3OD) d: 4.56
(2H, s, –CH2–), 7.0—7.5 (10H, m, arom. H10). EI-MS m/z: 242 (Mϩ).
N-Hydroxy-NЈ-phenylurea5) (5): Colorless plates (574.2 mg). mp 138—
141.8 °C (AcOEt). IR (KBr) cmϪ1: 3238, 2895, 1636, 1543, 1449, 1230,
939, 876, 802, 764, 646. H-NMR (CD3OD) d: 3.93 (3H, s, OCH3), 6.8—
8.1 (4H, m, arom. H4). HR-EI-MS m/z: 182.0676 (Mϩ, Calcd for
C8H10N2O3: 182.0691).
1
N-(Benzyloxy)-NЈ-(m-methoxyphenyl)urea (13a): Colorless needles (1205
mg, 60.8%). mp 118.1—119.2 °C (n-hexane–AcOEt). IR (KBr) cmϪ1: 2837,
1074, 757, 690. 1H-NMR (DMSO-d6) d: 6.96 (1H, t, Jϭ7.3 Hz), 7.24 (2H, t, 1655, 1612, 1572, 1501, 1418, 1333, 1311, 1213, 1178, 1159, 1080, 1032,
Jϭ7.4 Hz), 7.60 (2H, d, Jϭ8.7 Hz), 8.71 (1H, s), 8.78 (1H, s), 8.91 (1H, d, 941, 891, 833, 789, 741, 702. 1H-NMR (CD3OD) d: 3.59 (3H, s, OCH3),
Jϭ0.7 Hz). HR-EI-MS m/z: 152.0582 (Mϩ, Calcd for C7H8N2O2: 152.0586).
4.86 (2H, s, –CH2–), 6.5—7.5 (9H, m, arom. H9). EI-MS m/z: 272 (Mϩ).

1282
Vol. 50, No. 9
N-Hydroxy-NЈ-(m-methoxyphenyl)urea13) (13): Powdery crystals (282.5 (DMSO-d6) d: 7.53 (1H, t, Jϭ8.2 Hz), 7.81 (1H, ddd, Jϭ0.9, 2.3, 8.2 Hz),
mg). mp 113.3—114.6 °C (AcOEt). IR (KBr) cmϪ1: 3371, 3214, 2839,
8.04 (1H, ddd, Jϭ0.9, 2.2, 8.2 Hz), 8.70 (1H, t, Jϭ2.2 Hz). HR-EI-MS m/z:
1456, 1435, 1296, 1202, 1163, 1092, 1053, 991, 887, 766, 689. 1H-NMR 197.0427 (Mϩ, Calcd for C7H7N3O4: 197.0437).
(CD3OD) d: 3.76 (3H, s, OCH3), 6.6—7.2 (4H, m, arom. H4). HR-EI-MS
N-Hydroxy-NЈ-(p-nitrophenyl)urea12) (17): Pale yellow needles (176.9
m/z: 182.0703 (Mϩ, Calcd for C8H10N2O3: 182.0691).
mg). mp 162.5—165 °C (CHCl3–MeOH). IR (KBr) cmϪ1: 3323, 2964, 2875,
1610, 1560, 1151, 1125, 1082, 824. 1H-NMR (DMSO-d6) d: 7.92 (2H, d,
Jϭ9.4 Hz), 8.16 (2H, d, Jϭ9.4 Hz), 9.13 (1H, s), 9.26 (1H, s), 9.50 (1H, s).
N-(Benzyloxy)-NЈ-(p-methoxyphenyl)urea (14a): Colorless needles
(1292.6 mg, 86.5%). mp 105.5—107 °C (AcOEt). IR (KBr) cmϪ1: 3344,
3211, 3067, 2862, 1649, 1518, 1420, 1323, 1269, 1232, 1178, 1069, 1030, HR-EI-MS m/z: 197.0427 (Mϩ, Calcd for C7H7N3O4: 197.0437).
1
947, 827, 804, 750, 654. H-NMR (CD3OD) d: 3.75 (3H, s, OCH3), 6.8—
7.5 (9H, m, arom. H9). EI-MS m/z: 272 (Mϩ).
Acknowledgments We are grateful to Dr. Masanori Morita (Research
N-Hydroxy-NЈ-(p-methoxyphenyl)urea12) (14): Colorless plates (625.3 Institute for Technology and Science, Kinki University) for measurement of
mg). mp 137.8—141.5 °C (AcOEt). IR (KBr) cmϪ1: 3441, 3320, 1654,
MS spectra. This research was financially supported by the Kansai Univer-
1597, 1551, 1511, 1240, 1025, 825. 1H-NMR (CD3OD) d: 3.70 (3H, s, sity Research Grants: Grant-in-Aid for Joint Research, 2000.
OCH3), 6.7—7.4 (4H, m, arom. H4). HR-EI-MS m/z: 182.0692 (Mϩ, Calcd
for C8H10N2O3: 182.0691).
References and Notes
Preparation of N-substituted Hydroxyureas (2—4, 15—17) In a typi-
cal experiment,14) a solution of NH2OH·HCl (7.61 mmol) in H2O (1.5 ml)
was added to a solution of NaOH (7.61 mmol) in H2O (1 ml). To this stirred
mixture was added isocyanate (7.61 mmol) over a period of 20 min under
ice-cooling. After stirring for another 45 min, the reaction mixture was fil-
tered to yield a precipitate.
1) Stearns B., Losee K. A., Bernstein J., J. Med. Chem., 6, 201 (1963).
2) Greig M. A., Neithercut W. D., Hossack M., McColl K. E., J. Clin.
Pathol., 44, 157—159 (1991).
3) Brzozowski T., Sliwowski Z., Majka J., Drozdowicz D., Konturek S. J.,
J. Physiol. Pharmacol., 47, 137—150 (1996).
4) Blakeley R. L., Hinds J. A., Kunze H. E., Webb E. C., Zerner B., Bio-
chemistry, 8, 1991—2000 (1969), the references cited herein.
5) Clifton G., Bryant S. R., Skinner C. G., J. Med. Chem., 13, 377—379
(1970).
6) Harmon R. E., Dabrowiak J. C., Brown D. J., Gupta S. K., Herbert M.,
Chitharanjan D., J. Med. Chem., 13, 577—579 (1970).
7) Ichimori K., Stuehr D. J., Atkinson R. N., King S. B., J. Med. Chem.,
42, 1842—1848 (1999).
N-Ethyl-NЈ-hydroxyurea6) (2): Colorless fine crystals (401.2 mg). mp
113—115 °C (AcOEt). IR (KBr) cmϪ1: 3396, 3238, 2981, 2879, 1684, 1654,
1
1458, 1153, 1070, 778, 724. H-NMR (CD3OD) d: 1.13 (3H, t, Jϭ7.2 Hz),
3.22 (2H, q, Jϭ7.2 Hz).
N-Hydroxy-NЈ-propylurea6) (3): Colorless fine crystals (140 mg). mp
116—118.3 °C (AcOEt). IR (KBr) cmϪ1: 3323, 2964, 2875, 1636, 1560,
1
1151, 1125, 1082, 824, 749. H-NMR (CD3OD) d: 0.92 (3H, t, Jϭ7.4 Hz),
1.47—1.59 (2H, m), 3.15 (2H, t, Jϭ7.0 Hz).
8) Ouan H.-J., Koyanagi J., Ohmori K., Uesato S., Tsuchido T., Saito S.,
Eur. J. Med. Chem., 37, in press (2002).
9) Van Slyke D. D., Archibald R. M., J. Biol. Chem., 154, 623—642
(1944).
N-Butyl-NЈ-hydroxyurea7) (4): Colorless fine crystals (550 mg). mp 122—
124 °C (AcOEt). IR (KBr) cmϪ1: 3328, 3159, 2959, 1560, 1430, 1317, 1145,
1082, 832. 1H-NMR (DMSO-d6) d: 0.87 (3H, t, Jϭ7.3 Hz), 1.20—1.32 (2H,
m), 1.34—1.44 (2H, m), 3.03 (2H, q, Jϭ6.7 Hz), 6.60 (1H, br t, Jϭ5.8 Hz), 10) Kobashi K., Hase J., Uehara K., Biochim. Biophys. Acta, 65, 380—383
8.18 (1H, s), 8.49 (1H, d, Jϭ1.0 Hz). (1962).
N-Hydroxy-NЈ-(o-nitrophenyl)urea (15): Pale yellow needles (320.9 mg). 11) Kobayashi K., Kumaki K., Hase J., Biochim. Biophys. Acta, 227,
mp 225—226 °C (CHCl3–MeOH). IR (KBr) cmϪ1: 3292, 1654, 1584, 1508,
1340, 1276, 1148, 743. H-NMR (DMSO-d6) d: 7.29 (1H, br t, Jϭ7.8 Hz), 12) Wilson B. D., Woodgate P. E., Henn R. W., FR Patent, 2,184,047,
7.71 (1H, br t, Jϭ7.9 Hz), 7.96 (1H, dd, Jϭ1.3, 8.4 Hz), 8.04 (1H, dd, Jϭ1.5,
8.3 Hz), 10.04 (1H, s). HR-EI-MS m/z: 197.0427 (Mϩ, Calcd for C7H7N3O4:
197.0437).
429—441 (1971).
1
1974.
13) Bodi T., Pinter I., Foldi Z., Molnar I., Racz L., HU Patent, 38,899,
1986.
N-Hydroxy-NЈ-(m-nitrophenyl)urea (16): Pale yellow fine crystals 14) For the preparation of NO2-substituted derivatives of 5, MeOH
(133.5 mg). mp 156—158.5 °C (CHCl3–MeOH). IR (KBr) cmϪ1: 3199,
2892, 1647, 1597, 1560, 1523, 1420, 1347, 1108, 892, 842. 1H-NMR
(2.5 ml) was added to the reaction mixture in order to promote the sol-
ubility of the isocyanates.