A novel synthesis of 3-(substituted)pyrimido[4,5-c]pyridazine-5,7(1H,6H)-diones

Article abstract of DOI:10.1016/j.tetlet.2010.01.007

An improved synthesis of 3-(substituted)pyrimido[4,5-c]pyridazine-5,7(1H,6H)-diones, a known subclass of 4-deazatoxoflavins, is reported. The approach involves treatment of 3-methyl-6-(1-methylhydrazinyl)uracil with representative phenyl and alkyl glyoxal

Full text of DOI:10.1016/j.tetlet.2010.01.007

Tetrahedron Letters 51 (2010) 1326–1328
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
A novel synthesis of 3-(substituted)pyrimido[4,5-c]pyridazine-5,7(1H,6H)-
diones
Anjanette J. Turbiak a, Jeff W. Kampf b, H. D. Hollis Showalter a,
*
a Department of Medicinal Chemistry, University of Michigan, Ann Arbor, MI 48109, USA
b Department of Chemistry, University of Michigan, Ann Arbor, MI 48109, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
An improved synthesis of 3-(substituted)pyrimido[4,5-c]pyridazine-5,7(1H,6H)-diones, a known subclass
of 4-deazatoxoflavins, is reported. The approach involves treatment of 3-methyl-6-(1-methylhydrazi-
nyl)uracil with representative phenyl and alkyl glyoxal monohydrates, which in turn are obtained by
selenium dioxide oxidation of the corresponding phenyl and alkyl methyl ketones. The first entry into
4-monosubstituted isomers is also reported.
Received 4 December 2009
Revised 23 December 2009
Accepted 5 January 2010
Available online 11 January 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Pyrimido[4,5-c]pyridazine-5,7(1H,6H)-
dione
4-Deazatoxoflavin
Phenylglyoxal
Cyclizations
6-(1-Methylhydrazinyl)uracil
1,6-Dimethylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-dione, also
known as 4-deazatoxoflavin, and its 3,4-disubstituted analogues
have been shown to serve as biomimetics of flavin and 5-deazaflavin
in their ability to oxidize amines to carbonyl compounds1 and to ab-
stract hydrogen equivalents from hydrogen donors under certain
conditions (Fig. 1).2 4-Deazatoxoflavin itself has also demonstrated
both inhibitory activity against Pseudomonas and DNA-binding
properties3 while not displaying the generalized cytotoxicity that
is characteristic of toxoflavin.4 There are scattered reports of
3,4-disubstituted3,5 and 3-mono-substituted2,6 analogues of 4-dea-
zatoxoflavin, but no reports of 4-monosubstituted analogues. The
described syntheses of 3-monosubstitued analogues generally
proceed in poor yields and via pathways that produce alternate
products. Our goal was to develop an improved synthesis that would
lead to clean products and possess generality for a wide range of
analogues for biological testing.
tone moieties, or as is often the case for 6-aminouracils,7 by nucleo-
philic attack by the 5-position of the uracil onto either glyoxal
carbonyl. Thus, we set out to evaluate a number of reaction
conditions to determine if regiochemical preferences for the uracil
and glyoxal reaction partners could be established. Initially, we trea-
ted 3-methyl-6-(1-methylhydrazinyl)uracil8 with phenylglyoxal
monohydrate in refluxing ethanol or water and obtained the result
shown in Scheme 1. Upon cooling a precipitate corresponding to
ꢀ85% of 3 was confirmed by 1H NMR and mass spectrometry analy-
sis, and a negative Tollens test for aldehyde. In addition to support-
ing spectral data, selective hydrazone formation onto the aldehyde
moiety under these conditions is well precedented from prior stud-
ies on glyoxal hydrazones of methylhydrazine.9,10 The ꢀ15% co-
crystallized impurity plus 5–10% additional material in the mother
liquor corresponded to one of two possible cyclization products 4
or 5 by spectral analysis with a distinctive 1H NMR methine proton
Our strategy is based on the known reaction of 6-(1-methylhydr-
azinyl)uracil with glyoxal and a-diketones to yield 3,4-unsubstitut-
ed- and 3,4-disubstituted- pyrimido[4,5-c]pyridazine-5,7(1H,6H)-
diones, respectively.3 Despite these prior examples, there are no
reports of reaction between this or related 6-(hydrazinyl)uracils
and unsymmetrical alkyl or aryl glyoxals. We imagined that reaction
could occur by several manifolds. One could proceed via initial
hydrazone formation onto either one or both of the aldehyde or ke-
O
O
N
4
5
6
3
6
X
X
3
NH
N
8
1
10
N
R
1
N
N
X
N
O
X= N; flavin
X= N; toxoflavin
X= CH; 4-deazatoxoflavin
X= CH; 5-deazaflavin
* Corresponding author. Tel.: +1 734 764 5504; fax: +1 734 647 8430.
E-mail address: showalh@umich.edu (H.D.H. Showalter).
Figure 1. Structure of toxoflavin, flavin and corresponding deaza analogues.
0040-4039/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2010.01.007

A. J. Turbiak et al. / Tetrahedron Letters 51 (2010) 1326–1328
1327
O
O
O
O
CH3
O
CH3
O
OH
OH
5
EtOH
N
N
H2N
N
N
CH3
N
H
N
N
H
Δ
1
CH3
1
2
3
H4
O
O
N
CH3
O
H3
CH3
O
N
N
N
N
N
N
N
CH3
CH3
4
5
Scheme 1. Reaction products from condensation of (hydrazinyl)uracil 1 with phenylglyoxal 2.
ate the 4-phenyl isomer 5 directly from hydrazone 3, various condi-
tions were examined to effect cyclodehydrative ring closure.
However, only starting hydrazone 3 was recovered (prolonged re-
flux in water, ethanol, or 1,2-dichloroethane), or decomposition
set in (refluxing 1,2-dichloroethane with protic or Lewis acid).
Upon isolation of 4, a variety of conditions were explored to
generate it selectively. Table 1 summarizes the results. As can be
readily seen, there were a number of conditions (entries d, e, f) that
yielded solely the 3-phenylpyrimidopyridazinedione (4). Interest-
ingly, however, the reaction conditions that afforded cyclized prod-
uct most rapidly (entry b, 2 equiv NaOAc, H2O, 1 h) consistently
yielded a mixture of both regioisomers, even when varying both
the rate and order of addition of the reactants. By careful chro-
matographic separation of 3- and 4-phenylpyrimidopyridazinedi-
one isomers, we determined that d(H3) = 8.35 ppm in DMSO-d6
for isomer 5. The pattern of d(H4) > d(H3) is observed for related
phenyl substituted compounds (Table 2).
Table 1
Reaction conditions and product distribution from the treatment of (hydrazinyl)uracil
(1) with phenylglyoxal monohydrate (2)
Entry
Reaction conditions
H2O or EtOH, , 24h
H2O, 2 equiv NaOAc,
H2O, , 2 h (2 equiv NaOAc
Product (yield)
a
b
c
D
3 (ꢀ50%) & 4 (ꢀ25%)
4 & 5; ꢀ1:1 (76%)
3 & trace 4
D
, 1 h
D
added after 30 min)
1,2-dichloroethane,
d
e
f
D
, 3 h
4 (52%)
4 (55%)
4 (34%)
3 & 4
1,2-dichloroethane, BF3ÁEt2O
p-TsOH, H2O, , 24 h
CH3OCH2CH2OH, , 24 h
D, 24 h
D
g
D
(H4) at d 8.72 ppm in DMSO-d6. Ultimately, we generated compound
4 exclusively (vide infra) and determinedits structure by single crys-
tal X-ray analysis. This showed the cyclized product to be the 3-phe-
nyl isomer 4, formed by uracil nucleophilic attack at the C-5 position
onto the aldehyde moiety of phenylglyoxal. In an attempt to gener-
Having established efficient conditions to the 3-phenylpyrimi-
dopyridazinedione (4), additional substituted phenyl and selected
Table 2
3- and 4-substituted pyrimido[4,5-c]pyridazine-5,7(1H,6H)-diones synthesized via condensation of 6-(hydrazinyl)uracil 1 with aryl and alkyl glyoxals (6)
Entry
R
Conditionsa
Product (yield)b
1a
1b
2a
2b
3a
3b
4a
4b
5a
5b
6a
6b
7a
7b
8a
8b
4-F-Ph
A
B
A
B
A
B
A
B
A
B
A
B
A
Bd
A
B
7a (56%)
7a:8a, 2:3 (55%)
7b (22%)
7b:8b, 1:2 (62%)
7c (85%)
7c:8c, 2:5 (43%)
7d (75%)
7d:8d, 1:1 (54%)
7e (42%)
7e:8e, 1:1 (51%)
7f (49%)
4-Cl-Ph
4-CF3-Ph
4-MeO-Ph
4-Cl-3-NO2-Ph
3-F-Ph
7f:8f, 1:1 (65%)
t-Bu
7g (19%)c
7g (60%)c
Me
7h (54%)
7h:8h, 3:1 (30%)
a
b
c
A: 1,2-dichloroethane,
Yields represent 1–2 runs for each condition and are not optimized.
Glyoxal from oxidation used directly in cyclization without purification.
D
, 1.5À24 h; B: H2O, 2 equiv NaOAc,
D, 0.2–1 h.
d
D
, 3 h.

1328
A. J. Turbiak et al. / Tetrahedron Letters 51 (2010) 1326–1328
alkyl glyoxal monohydrates were evaluated to explore the scope of
the new methodology. Non-commercial glyoxal monohydrates
were prepared by oxidation of the corresponding phenyl or alkyl
methyl ketone with selenium dioxide according to a standard liter-
ature procedure.11 Each of these was then treated with 6-(hydrazi-
nyl)uracil 1 under the following sets of conditions: (a) refluxing
1,2-dichloroethane, 1.5À24 h and (b) aqueous NaOAc (vide supra).
The results are shown in Table 2. In each case, product yields are
based on collected precipitate from the reaction mixture and were
not optimized. For each example shown, conducting the reaction in
1,2-dichloroethane yielded solely the 3-substituted pyrimidopyri-
dazinedione. For pyruvic aldehyde (Table 2, entry 8a), commercial
aqueous reagent was extracted into dichloromethane and the solu-
tion dried prior to use to effect a clean reaction to 7h. Similar bi-
phasic reaction with aqueous reagent resulted in significant co-
generation of hydrazone.
B. Cresswell, and Fred and Dee Lyons Graduate Fellowships. This
publication was also made possible by grant number GM007767
from NIGMS. Its contents are solely the responsibility of the
authors and do not necessarily represent the official views of
NIGMS.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.tetlet.2010.01.007.
References and notes
1. Nagamatsu, T.; Hashiguchi, Y.; Sakuma, Y.; Yoneda, F. Chem. Lett. 1982, 1309.
2. Yoneda, F.; Nakagawa, K.; Noguchi, M.; Higuchi, M. Chem. Pharm. Bull. 1981, 29,
379.
3. Billings, B. K.; Wagner, J. A.; Cook, P. D.; Castle, R. N. J. Heterocycl. Chem. 1975,
12, 1221.
4. Latuasan, H. E.; Berends, W. Biochim. Biophys. Acta 1954, 52, 502.
5. Sakuma, Y.; Nagamatsu, T.; Hashiguchi, Y.; Yoneda, F. Chem. Pharm. Bull. 1984,
32, 851.
6. Naya, S.; Shibayama, K.; Nitta, M. Heterocycles 2004, 63, 1393.
7. Migawa, M. T.; Hinkley, J. M.; Hoops, G. C.; Townsend, L. B. Synth. Commun.
1996, 26, 3317.
8. Daves, G. D., Jr.; Robins, R. K.; Cheng, C. C. J. Am. Chem. Soc. 1962, 84, 1724.
9. Kano, K.; Scarpetti, D.; Warner, J. C.; Anselme, J. P.; Springer, J. P.; Arison, B. H.
Can. J. Chem. 1986, 64, 2211.
10. Begtrup, M.; Nytoft, H. P. J. Chem. Soc., Perkin Trans. 1 1985, 81.
11. Fodor, G.; Kovacs, O. J. Amer. Chem. Soc. 1949, 71, 1045.
Typical experimental procedures. General procedure for the synthesis of phenyl/
alkyl glyoxal monohydrates (6) from phenyl/alkyl methyl ketones: The phenyl/
In contrast, reactions conducted in aqueous NaOAc yielded a
mixture of both regioisomers for all aryl examples examined. Un-
der these conditions both isomers were isolated in approximately
equal amounts in many cases (Table 1, entry b, Table 2, 4b, 5b,
6b), but interestingly, if one regioisomer was heavily favoured, it
was always the 4-substituted one (Table 2, entries 1b, 2b, 3b). Un-
der the same conditions, when the R group was t-butyl (Table 2,
entry 7b) only the 3-substituted isomer (7g) was formed and
was heavily favoured for methyl, reflecting an interesting inversion
of regioselectivity between aryl and alkyl glyoxals. To rule out the
possibility that product formation proceeds via a hydrazone inter-
mediate, compound 3 was refluxed for 16 h under the NaOAc con-
ditions. Only starting material was recovered.
alkyl methyl ketone (1.5 mmol) was dissolved in 3 mL p-dioxane and 120
lL of
H2O. SeO2 (3.0 mmol, 2 equiv) was added and the mixture was heated at reflux
for 24–72 h with consumption of starting material monitored by TLC. The
mixture was cooled to 25 °C and filtered through Celite, rinsing with EtOAc.
The filtrate was concentrated, dissolved in CH2Cl2 and purified by SiO2 flash
chromatography (elution with 0–20% EtOAc in hexanes). Representative
procedure for the synthesis of 3-substituted pyrimido[4,5-c]pyridazine-
5,7(1H,6H)-diones. 1,6-Dimethyl-3-phenylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-
The 1H NMR spectra for compounds 4a2 and 7h6 are identical to
those described in earlier syntheses, and thus confirm previous
structural assignments.
In conclusion, we describe novel and efficient procedures to
pyrimido[4,5-c]pyridazine-5,7(1H,6H)-diones. The data argue for
a reaction manifold that proceeds by selective nucleophilic attack
by the 5-position of the uracil onto the glyoxal aldehyde moiety
in nonpolar solvents, whereas for hydroxylic solvents there is a
preference for hydrazone formation. Under mildly basic conditions
(e.g., aqueous NaOAc), selective C-5 attack is also observed, but
now proceeds onto both glyoxal carbonyls with the ratio of
3- and 4-substituted products dependent on electronic and steric
factors. In no case is there evidence for ring closure via an interme-
diate hydrazone. Thus, our methodology allows for the selective
incorporation of an aryl or alkyl substituent at the 3-position, rep-
resenting a direct access to this subclass of pyrimidopyridazinedi-
ones. We also report the first entry into 4-monosubstituted
isomers and are working to develop conditions that provide this
isomer selectively.
dione (4): To
a
refluxing suspension of 6-(hydrazinyl)uracil 18 (85 mg,
0.5 mmol) in 2 mL of 1,2-dichloroethane was added phenylglyoxal
monohydrate 2 (84 mg, 0.55 mmol). Additional 2 (10 mg) was added after
3 h and the solution was heated for 3 h more. The orange mixture was
concentrated to a solid residue that was triturated in hot EtOH. The solids were
collected, washed with EtOH and dried to leave 70 mg (52%) of pure 4 as yellow
fibres; mp 260–262 °C (lit2 mp 250 °C): Rf 0.37 (SiO2; 95:5 CH2Cl2: MeOH); 1
H
NMR (CDCl3) d 3.50 (s, 3H), 4.32 (s, 3H), 7.55–7.58 (m, 3H), 7.93–7.95 (m, 2H),
8.69 (s, 1H); 1H NMR (DMSO-d6) d 3.26 (s, 3H), 4.17 (s, 3H), 7.55–7.57 (m, 3H),
8.08 (d, J = 3.4 Hz, 2H), 8.72 (s, 1H); 13C NMR (DMSO-d6) d 28.25, 43.94, 123.65,
126.79, 128.81, 129.67, 130.85, 133.45, 133.58, 147.65, 155.09, 160.86; MS m/z
269.1 (M+H), 291.1 (M+Na). Representative procedure for synthesis of mixture of
3- and 4-substituted pyrimido[4,5-c]pyridazine-5,7(1H,6H)-diones. 1,6-Dimethyl-
3-phenylpyrimido[4,5-c]pyridazine-5,7(1H,6H)-dione (4) and 4-phenyl isomer (5):
To a refluxing mixture of 6-(hydrazinyl)uracil 18 (85 mg, 0.5 mmol), NaOAc
(82 mg, 1 mmol), and 4 mL of distilled H2O was added phenylglyoxal
monohydrate (84 mg, 0.55 mmol) in
a single charge. Yellow precipitate
began to form immediately. After heating for 1 h, the supension was
maintained at 25 °C for 1 h. The solids were collected, washed with H2O, and
dried to leave 102 mg (76%) of ꢀ1:1 mixture of 4:5 by 1H NMR. TLC (SiO2; 97:3
CH2Cl2/MeOH);) showed two partially overlapping spots, Rf 0.24. A small
sample of the mixture was purified by preparative SiO2 thin layer
chromatography, eluting 2 Â with 98:2 CH2Cl2/MeOH), to provide pure 5;
mp >205 °C (dec): 1H NMR (CDCl3) d 3.38 (s, 3H), 4.25 (s, 3H), 7.42–7.44 (m,
3H), 7.54–7.57 (m, 2H), 8.10 (s, 1H); 1H NMR (DMSO-d6) d 3.14 (s, 3H), 4.07 (s,
3H), 7.48–7.50 (m, 5H), 8.35 (s, 1H); MS m/z 269.1 (M+H), 291.1 (M+Na).
Acknowledgements
A.J.T. gratefully acknowledges the financial support of the
American Chemical Society Division of Medicinal Chemistry, Sheila