Rynaxypyr: A new insecticidal anthranilic diamide that acts as a potent and selective ryanodine receptor activator

Article abstract of DOI:10.1016/j.bmcl.2007.09.012

Rynaxypyr is a highly potent and selective activator of insect ryanodine receptors with exceptional activity on a broad range of Lepidoptera. A strong correlation between insecticidal activity and ryanodine receptor activation is observed along with selective activity against insect over mammalian receptors. The synthesis and biological results are presented.

Full text of DOI:10.1016/j.bmcl.2007.09.012

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 17 (2007) 6274–6279
Rynaxypyre: A new insecticidal anthranilic diamide that acts
as a potent and selective ryanodine receptor activator
George P. Lahm,* Thomas M. Stevenson, Thomas P. Selby, John H. Freudenberger,
Daniel Cordova, Lindsey Flexner, Cheryl A. Bellin, Christine M. Dubas, Ben K. Smith,
Kenneth A. Hughes, J. Gary Hollingshaus, Christopher E. Clark and Eric A. Benner
DuPont Crop Protection, Stine-Haskell Research Center, 1094 Elkton Road, Newark, DE 19711, USA
Received 1 August 2007; revised 29 August 2007; accepted 4 September 2007
Available online 7 September 2007
Abstract—RynaxypyrTM is a highly potent and selective activator of insect ryanodine receptors with exceptional activity on a broad
range of Lepidoptera. A strong correlation between insecticidal activity and ryanodine receptor activation is observed along with
selective activity against insect over mammalian receptors. The synthesis and biological results are presented.
ꢀ 2007 Elsevier Ltd. All rights reserved.
Br
N
H
N
O
The ryanodine receptor (RyR) is a non-voltage-gated
calcium channel located in the sarcoplasmic reticulum
of muscle cells and endoplasmic reticulum of non-mus-
cle cells. RyRs regulate the release of intracellular cal-
cium stores critical for muscle contraction. Its name is
derived from the natural insecticide ryanodine, a plant
metabolite from Ryania speciosa, that has been found
to affect calcium release by locking channels in a par-
tially opened state.1 We recently reported the discovery
of a new class of insecticides, the anthranilic diamides,
which exhibit their action through activation of the
ryanodine receptor.2,3 In this paper we describe the dis-
covery of RynaxypyrTM (chlorantraniliprole, DPX-
E2Y45), a potent ryanodine receptor activator, as the
first new insecticide from this class (Fig. 1).4,5 Rynaxy-
pyrTM is characterized by its high levels of insecticidal
activity and low toxicity to mammals attributed to a
high selectivity for insect over mammalian ryanodine
receptors.6,7
Me
Cl
H
N
N
Cl
O
Me
N
Figure 1. RynaxypyrTM (chlorantraniliprole, DPX-E2Y45).
cium channel regulation and, in particular, the ryano-
dine receptor (RyR) represents a new biochemical
target for insect control and thus offers excellent promise
in integrated pest management strategies. RynaxypyrTM
binds to a receptor site distinct from that of ryanodine
and appears to be impacted by the channel’s state. Acti-
vation causes unregulated release of internal calcium
stores leading to Ca2+ depletion, feeding cessation, leth-
argy, muscle paralysis, and ultimately insect death.
Through cloning and expression of multiple insect RyRs
we have been able to provide genetic validation for the
mode of action.3,6
The ability of insects to rapidly develop resistance to
conventional pesticides poses a significant problem for
effective pest management. The discovery of control
agents that work by new biochemical mechanisms is
therefore of critical importance in crop protection. Cal-
We previously described the 3-chloropyridyl group of
the anthranilic diamides as an optimum substituent on
the pyrazole-5-carboxamide.2 Additionally, we found
the 6-methyl group of the anthranilamide to be one of
the more preferred substituents. Here we describe the
synthesis, insecticidal activity, and RyR activity of 1-
(3-chlororpyridyl)-pyrazole-6-methylanthranilic diamides
Keywords: RynaxypyrTM; Ryanodine; Receptor; Insecticide.
*
Corresponding author. Tel.: +1 302 366 5711; fax: +1 302 366
5738; e-mail: george.p.lahm@usa.dupont.com
0960-894X/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.09.012

G. P. Lahm et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6274–6279
6275
with varied 4-anthranilic and 3-pyrazole substituents,
leading to the discovery of RynaxypyrTM.
4. Metallation of 4 with lithium diisopropylamide fol-
lowed by addition of carbon dioxide affords the key
intermediate 3-bromopyrazole-5-carboxylic acid 5.
Schemes 1–3 illustrate the routes by which the com-
pounds of Table 1 were synthesized. Anthranilic dia-
mides 8 containing a 3-bromopyrazole (R3 = Br) could
be prepared as outlined in Scheme 1. A new method
for the preparation of the intermediate 3-bromopyrazole
3 was developed that takes advantage of the known
selective lithiation of N,N-dimethyl-sulfamoylpyrazole
1 at the 5-position.8 Thus, treatment of 1 with n-butylli-
thium followed by bromination with 1,2-dibromo-1,1,
2,2-tetrachloroethane affords 5-bromo-1-N,N-dim-
ethylsulfamoylpyrazole 2. Removal of the N,N-dim-
ethylsulfamoyl protecting group with TFA affords
the 3-bromopyrazole 3, which upon reaction with 2,3-
dichloropyridine provides 1-pyridyl-3-bromopyrazole
The benzoxazinone intermediates 7 could be prepared
by the coupling of the pyrazole carboxylic acids 5 with
anthranilic acids 6. A preferred method for direct con-
version of 5 to the benzoxazinone involves sequential
treatment with one equivalent of triethylamine and
methanesulfonyl chloride, followed by sequential addi-
tion of the anthranilic acid 6, triethylamine, and metha-
nesulfonyl chloride to provide benzoxazinone 7.
Treatment of 7 with amines, R2NH2, provided the
anthranilic diamides 8 which generally precipitated from
the reaction medium. Anthranilic diamides containing a
3-chloropyrazole (R3 = Cl) could be prepared in an
analogous fashion from 3-chloropyrazole prepared from
Br
Br
N
N
N
a
b
c
d
N
N
N
N
Br
Cl
N
H
SO2NMe2
SO2NMe2
N
1
2
3
4
Me
NH2
O
Br
N
Br
Br
N
R1
Me
O
N
N
N
Me
N
O
O
6
OH
Cl
N
f
NH
NH
Cl
Cl
N
O
OH
R1
e
N
N
O
R1
R2
5
7
8
Scheme 1. Reagents and conditions: (a) i—nBuLi, THF, À60 ꢁC; ii—BrCCl2CCl2Br, THF, À70 ꢁC, 89%; (b) TFA, 25 ꢁC, 78%; (c) 2,3-
dichloropyridine, K2CO3, DMF, 125 ꢁC, 64%; (d) i—LDA, THF, À78 ꢁC; ii—CO2 gas; iii—HCl, 87%; (e) i—5, MeSO2Cl, Et3N, MeCN; ii—6; iii—
Et3N, MeCN; iv—MeSO2Cl, 50–60%; (f) R2NH2, THF, 70–95%.
Cl
N
OH
N
Cl
N
NH2
HN
CO2Et
a
Cl
c
b
O
O
O
N
N
N
+
N
Cl
Cl
Cl
OH
CO2Et
OEt
OEt
N
N
N
9
10
11
12
13
d
OCF2H
OH
OCH2CF3
N
N
N
f
e
O
O
O
N
N
N
Cl
Cl
Cl
OH
OEt
OH
N
N
N
16
14
15
Scheme 2. Reagents and conditions: (a) NaOEt, EtOH, reflux, 55%; (b) POCl3, MeCN, 80 ꢁC, 91%; (c) i—MeCN, H2SO4, K2S2O8, reflux, 82%; ii—
aq NaOH, MeOH, 83%; (d) MeCN, H2SO4, K2S2O8, reflux, 82%; (e) i—CF3CH2I, K2CO3, DMF, 100 ꢁC, 44%; ii—aq NaOH, MeOH, 87%; (f) i—
CHClF2, K2CO3, DMF 42%; ii—aq NaOH, MeOH, 80%.

6276
G. P. Lahm et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6274–6279
Me
Me
Me
H
H
N
N
NH2
a
b
OH
c
N
O
O
CF3
CF3
CF3
O
17
18
19
R3
N
R3
R3
N
Me
H
Me
O
N
O
N
N
Me
d
e
O
N
O
Cl
N
N
+
NH
NH
O
Cl
Cl
N
CF3
Cl
O
CF3
N
N
O
O
CF3
R2
20
21
22
23
Scheme 3. Reagents and conditions: (a) Cl3CCH(OH)2, Na2SO4, (NH2OH)2–H2SO4, HCl, H2O, 35%; (b) H2SO4, H2O, 85%; (c) HOAc, H2O2, 75 ꢁC,
60%; (d) Et3N, CH3CN, 70 ꢁC, 23%; (e) R2NH2, THF, 40–50%.
Table 1. Insecticidal potency and Hv ryanodine receptor activity (Hv RyR EC50) of anthranilic diamides15
R3
N
O
N
Me
Cl
NH
NH
N
O
R1
R2
Entry
R1
R2
R3
Sfa
Pxa
Hva
Hv RyRa
EC50 nM (SEM)
LC50 (ppm)
LC50 (ppm)
LC50 (ppm)
D1
D2
Cl
Cl
Cl
Cl
Cl
Cl
Br
Br
I
Me
i-Pr
Me
i-Pr
Me
i-Pr
i-Pr
Me
Me
Me
Me
i-Pr
Me
i-Pr
Me
i-Pr
Me
i-Pr
CF3
CF3
0.02
0.03
0.02b
0.04b
0.03
0.05
0.03
0.18
0.26
0.13
0.53
0.39
0.68
0.30
0.21
0.14
0.11
0.03b
0.01c
0.01
0.02
0.04
0.03
0.09
0.01
0.06
0.10
0.07b
0.09
0.07
0.33
0.18
0.38
0.10
0.04
0.01
0.05
0.02b
0.04b
0.02c
0.07
0.05c
0.03
0.11
0.21
0.16
0.66
0.11
2.43
1.14
1.08
0.12
0.24
0.09
73 (4)
1834 (111)
52 (3)
D3
D4
Br
Br
458 (23)
69 (3)
D5
D6
Cl
Cl
270 (17)
1574 (97)
63 (4)
D7
D8
CF3
Br
D9
CF3
Br
205 (10)
118 (6)
NT
D10
D11
D12
D13
D14
D15
D16
D17
D18
I
CF3
CF3
Cl
Cl
Cl
Cl
Cl
Cl
CF3
CF3
3704 (80)
101 (9)
387 (9)
60 (3)
OCH3
OCH3
OCF2H
OCF2H
OCH2CF3
OCH2CF3
511 (34)
240 (4)
2734 (284)
Insect LC50s (ppm) are on fall armyworm (Sf, Spodoptera frugiperda), diamondback moth (Px, Plutella xylostella), and tobacco budworm (Hv,
Heliothis virescens).
a Mortality and RyR EC50 values were obtained for multiple test rates, each tested in replicate (n P 16 for LC50 values), (n P 3 for EC50 values).
LC50 and EC50 calculations were determined by Probit analysis using a maximum quasi-likelihood curve fitting algorithm.16 For each of the LC50
values listed, the range between the calculated LC50 and the corresponding lower or upper 90%-confidence interval value is less than 50% of the
calculated value unless otherwise noted.
b The range for the upper and/or lower 90%-confidence interval is less than 100% of the calculated value.
c The range for the upper and/or lower 90%-confidence interval is less than 300% of the calculated value.
1 by chlorination with hexachloroethane. The intermedi-
ate 4-halo-6-methyl-anthranilic acids 6 (R1 = Cl, Br, I)
were prepared by halogenation of 6-methylanthranilic
acid with the corresponding N-halosuccinimide.5

G. P. Lahm et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6274–6279
6277
series of Lepidoptera under standard laboratory proce-
dures.14 Potency on fall armyworm (Spodoptera fru-
giperda, Sf), diamondback moth (Plutella xylostella,
Px), and tobacco budworm (Heliothis virescens, Hv)
was evaluated. Insecticidal activity is reported as an
LC50 in ppm, the lethal concentration required for
50% mortality. Also reported in Table 1 is the Hv
RyR EC50, a measure of receptor activity from a recom-
binant Spodopteran cell line that stably expresses the
H. virescens RyR. Sf9 cells are devoid of endogenous
RyRs, however expression of recombinant H. virescens
RyR confers sensitivity to anthranilic diamides and the
known RyR agent, caffeine.5b These recombinant cells
were plated into a 96-well plate and the compound-in-
duced calcium mobilization was monitored using a Flex-
StationTM plate reader as previously described.2 As
these cells are devoid of endogenous RyRs, the activity
reflects compound potency against the recombinant
H. virescens RyR. As a general rule all new compounds
showed high potency against the three species of Lepi-
doptera, with the greatest sensitivity to Px and the least
sensitivity to Hv. Compound D13, the least active com-
pound of Table 1, still proved substantially more active
than many insecticide standards. Structure–activity rela-
tionships for compounds D1–D6, where R1 was fixed as
Cl, and R3 was varied as chloro, bromo, and trifluorom-
ethyl, suggested the trend Br ꢀ CF3 > Cl, although these
differences were very small. In addition, the isopropyl
amides of compounds D1–D6 tended to show slightly
greater levels of insecticidal activity on Lepidoptera than
their corresponding methyl amides particularly on the
least susceptible Hv species. However, we observed the
opposite trend at the Heliothis ryanodine receptor. In
particular we noted that Hv RyR activity for each of
the methyl amides D1, D3, and D5 to be consistently
more potent than the corresponding isopropyl amides
D2, D4, and D6. While we cannot rule out decreased cell
permeability of the isopropyl amides as a possible cause
for the reduced receptor activity, one would expect a
similar reduction in insect toxicity and this is not
observed. We therefore believe the higher whole-organ-
ism toxicity for the isopropyl analogs may be the result
of improved bioavailability.
Chart 1. Differential receptor selectivity of RynaxypyrTM in insect and
mammalian cell lines expressing ryanodine receptors.
Preparation of the pyrazole-5-carboxylic acids created
several problems associated with practical aspects of
the lithiation on scale up. An alternate route for the
preparation of halopyrazole carboxylic acids (R3 is Cl
or Br) was developed and is outlined in Scheme 2 for
3-chloropyrazole-5-carboxylic acid 13.9 Reaction of
diethylmaleate 10 with 3-chloro-2-hydrazinopyridine 9
in the presence of sodium ethoxide afforded the pyrazo-
lone 11 in 55% yield. Subsequent treatment of 11 with
phosphoryl chloride in acetonitrile at 80 ꢁC afforded
the chloropyrazoline 12 in excellent yield. The bromop-
yrazoline analog of 12 could also be prepared with sim-
ilar results using phosphoryl bromide as the
bromination reagent. A variety of reagents and condi-
tions were explored for oxidation of 12 to the pyrazole
13, and potassium persulfate was found to be one of
the preferred oxidants.9
The fluoroalkoxy derivatives 15 and 16 were prepared in
two steps from pyrazolone 11. Persulfate oxidation of 11
provided pyrazolone 14 albeit in lower yields than the
chloro analog. Trifluoroethoxy pyrazole 15 was pre-
pared by alkylation with trifluoroethyliodide. The
difluoromethoxy analog 16 was prepared from 14 via
difluorocarbene addition generated from Freon 22 and
base. The pyrazole acids 13, 15, and 16 were used to pre-
pare anthranilic diamides by procedures analogous to
those of Scheme 1.
Replacement of the chlorine substituent at R1 with the
groups Br, I, and CF3 (compounds D7–D12) indicated
sensitivity to substituents at R1. While the Br analog
D7 was of equal potency to the Cl analog D2, the methyl
amide D8 and both sets of I and CF3 analogs D9–D12
were somewhat less active.
Anthranilic diamides containing a CF3 substituent at R1
(i.e. 23) were prepared by the procedure shown in
Scheme 3. Isatin 19 was prepared in two steps from 2-
methyl-4-trifluoromethyl aniline by first reaction with
chloral hydrate and hydroxylamine to yield 18 followed
by cyclization with sulfuric acid.10,11 Oxidation of 19 to
the isatoic anhydride 20 was accomplished with hydro-
gen peroxide in acetic acid.12 The isatoic anhydride 20
could be coupled directly with pyrazole acid chlorides
such as 21 to yield the benzoxazinones 22 in modest
yield.13 Reaction of 22 with amines afforded the anthra-
nilic diamides 23.
Alkoxy and haloalkoxy derivatives at R3 (D13–D18)
followed the structure–activity trend on Hv of OCH2C-
F3 > OCHF2 > OCH3. In particular, we observed lower
activity for the methoxy derivatives D13–D14 in spite
of their strong activity at the Heliothis ryanodine
receptor for both the methyl and isopropyl amides.
This trend was observed for other methoxy derivatives
and may be the result of metabolic detoxification via
demethylation. In support of this, we observed a
>30- to 50-fold decrease in insect toxicity for the hy-
droxy (desmethyl) derivatives. Additionally, the re-
duced Lepidopteran activity of the difluoromethoxy
Insecticidal activity of the anthranilic diamides is sum-
marized in Table 1. Compounds were tested against a

6278
G. P. Lahm et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6274–6279
4. Lahm, G. P.; Selby, T. P.; Stevenson, T. M. PCT Int.
Appl. WO 03/015519, 2003; Chem. Abstr. 2003, 138,
200332.
analog D15 was a bit of a surprise owing to the strong
activity of the analogous i-Pr amide D16, and the
exceptional activity at the Heliothis ryanodine receptor.
Also, consistent with the trend for D1–D6 we observed
greater activity for the methyl amides of D13–D18 at
the Hv RyR.
5. Presented in part at the 11th IUPAC International
Congress on the Chemistry of Crop Protection, Kobe,
Japan August 2006. Proceedings: (a) Lahm, G. P.;
Stevenson, T. M.; Selby, T. P.; Freudenberger, J. F.;
Dubas, C. M.; Smith, B. K.; Cordova, D.; Flexner, L.;
Clark, C. E.; Bellin, C. A.; Hollingshaus, J. G. In Pesticide
Chemistry: Crop Protection, Public Health and Environ-
mental Safety; Ohkawa, H., Miyagawa, H., Lee, P. W.,
Eds.; Wiley-VCH: Weinheim, 2007; pp 111–120; (b)
Cordova, D.; Benner, E. A.; Sacher, M. D.; Rauh, J. J.;
Sopa, J. S.; Lahm, G. P.; Selby, T. P.; Stevenson, T. M.;
Flexner, L.; Caspar, T.; Ragghianti, J. J.; Gutteridge, S.;
Rhoades, D. F.; Wu, L.; Smith, R. M.; Tao, T. In
Pesticide Chemistry: Crop Protection, Public Health and
Environmental Safety; Ohkawa, H., Miyagawa, H., Lee, P.
W., Eds.; Wiley-VCH: Weinheim, 2007; pp 121–126.
6. Caspar, T.; Cordova, D.; Gutteridge, S.; Rauh, J. J.;
Smith, R. M.; Wu, L.; Tao, Y. PCT Intl. Appl. WO 04/
027042, 2004; Chem. Abstr. 2004, 140, 282468.
7. Cordova, D.; Benner, E. A.; Sacher, M. D.; Rauh, J. J.;
Sopa, J. S.; Lahm, G. P.; Selby, T. P.; Stevenson, T. M.;
Flexner, L.; Gutteridge, S.; Rhoades, D. F.; Wu, L.;
Smith, R. M.; Tao, Y. In Synthesis and Chemistry of
Agrochemicals VII; Lyga, J. W., Theodoridis, G., Eds.;
ACS: Washington, DC, 2007; pp 223–234.
8. (a) Effenberger, F.; Roos, M.; Ahmad, R.; Krebs, A.
Chem. Berl. 1991, 124, 1639; (b) Dudfield, C. T.; Ham-
ilton, E. K.; Osbourn, C. E. Synlett 1990, 277.
RynaxypyrTM possesses low acute mammalian toxicity
with an acute oral LD50 of >5000 mg/kg in rats, and lit-
tle to no toxicity in 90-day studies, at dosing as high as
1500 mg/kg/day. In addition, no developmental toxicity
is observed in rats or rabbits with doses as high as
1000 mg/kg/day. Comparative studies of ryanodine
receptor activation between insect and mammalian cell
lines were conducted to determine if differential receptor
selectivity is a contributing factor to the low mammalian
toxicity. Unlike insects, mammals express three RyR
isoforms. RyR1 and RyR2 are predominately found in
skeletal and cardiac muscle, respectively, whereas
RyR3 is heterogeneously distributed throughout various
tissues. As the data in Chart 1 shows, RynaxypyrTM is
ꢀ300-fold less potent against RyRs in the mouse myo-
blast cell line, C2C12, than in insect RyRs from Dro-
sophila melanogaster and H. virescens. This mouse cell
line has been shown to predominately express RyR1.17
Increased selectivity is observed with the rat cell line,
PC12, which predominately expresses RyR2.17 In this
cell line RynaxypyrTM has an EC50 value >100 lM
(ꢀ25% activation at 100 lM, >2000-fold selectivity).
Of the mammalian cell lines tested, the human cell line,
IMR32, was the least sensitive to RynaxypyrTM, with no
receptor activation observed at 100 lM. This cell line
has been shown to express functional RyRs however it
is unclear which isoforms are being expressed.18 Overall,
RynaxypyrTM exhibits strong differential selectivity for
insect over mammalian RyRs. Therefore, it is likely that
such selectivity contributes significantly to the observed
low mammalian toxicity.
9. Freudenberger, J. H.; Lahm, G. P.; Selby, T. P.; Steven-
son, T. M.; PCT Int. Appl., WO 03/016283, 2003; Chem.
Abstr. 2003, 138, 205053.
10. Da Silva, J. F. M.; Garden, S. J.; Pinto, A. C. J. Braz.
Chem. Soc. 2001, 12, 273.
11. Demerson, C. A.; Humber, L. G.; Philipp, A. H.; Martel,
R. R. J. Med. Chem. 1976, 19, 391.
12. Reissenweber, G.; Mangold, D. Angew. Chem. 1980, 92,
196.
13. (a) Tsubota, M.; Hamashima, M. Heterocycles 1984, 21,
706; (b) Coppola, G. M. J. Heterocycl. Chem. 1999, 36,
563.
From a number of strong candidates identified in Table
1, RynaxypyrTM was selected for development based on
the combination of insecticidal potency and low mam-
malian toxicity. In numerous field trials we observed
exceptional activity across a wide range of pests and
with great consistency. We believe RyanxypyrTM will
be a valuable addition to the chemistry of crop protec-
tion based on the combination of an extremely favorable
toxicology profile, the new mode of action and out-
standing insecticidal properties.
14. A representative testing protocol for Spodoptera frugiperda
is described. Experimental compounds were formulated at
500 ppm using a solution containing 10% acetone, 90%
water and 300 ppm X-77ꢂ Spreader (Loveland Industries,
Inc.). Serial dilutions were made to obtain concentrations of
0.01, 0.03, 1, 3, 10, 30, 100 and 500 ppm. The formulated
compounds were sprayed to run-off on 4-week old soybean
plants. Once the plants had dried, a leaf (or trifoliate) was
excised from the treated plant. The leaves were cut into 24
pieces and placed singly into a 5.5 cm by 3.5 cm cell of a 16-
well plastic tray (Mullinix Packages, Inc.). Each cell also
contained a 2.5 square of moistened chromatography paper
(Whatman No. 3MM) to prevent desiccation. One third
instar larvae of Spodoptera frugiperda was placed into each
cell. A total of 16 insects were tested per rate. Larval
mortality was assessed at 96 hours post-infestation. Percent
References and notes
1. Coronado, R.; Morrissette, J.; Sukhareva, M.; Vaughan,
D. M. Am. J. Physiol. 1994, 266, C1485.
2. Lahm, G. P.; Selby, T. P.; Freudenberger, J. H.; Steven-
son, T. M.; Myers, B. J.; Seburyamo, G.; Smith, B. K.;
Flexner, L.; Clark, C. E.; Cordova, D. Bioorg. Med. Chem.
Lett. 2005, 15, 4898.
3. Cordova, D.; Benner, E. A.; Sacher, M. D.; Rauh, J. J.;
Sopa, J. S.; Lahm, G. P.; Selby, T. P.; Stevenson, T. M.;
Flexner, L.; Gutteridge, S.; Rhoades, D. F.; Wu, L.;
Smith, R. M.; Tao, Y. Pestic. Biochem. Physiol. 2006, 84,
196.
control
was
evaluated,
[(#dead/total
number
insects) · 100], and LC50’s calculated.
15. All new compounds gave satisfactory spectral data con-
sistent with their structures. Synthesis of RynaxypyrTM
(D3): To a solution of 2-[3-bromo-1-(3-chloro-2-pyridi-
nyl)-1H-pyrazol-5-yl]-6-chloro-8-methyl-4H-3,1-benzoxa-
zin-4-one (5.5 g, 12.2 mmol) in tetrahydrofuran (100 mL)
was added methylamine (2 M in THF, 15.0 mL,
30.0 mmol) dropwise at room temperature. The reaction
mixture was stirred for 10 min and concentrated. The

G. P. Lahm et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6274–6279
6279
residual solid was purified by chromatography on silica gel
by elution with methylene chloride followed by ethyl
acetate and concentrated to a white solid. Trituration of
the solid with ether/hexane afforded 4.46 g of a white solid.
Yield 76%; mp 239–240 ꢁC; IR (Nujol) mmax 3388, 3262,
128.52, 131.61, 131.83, 132.22, 136.69, 139.47, 139.91,
140.05, 147.77, 149.09, 156.22, 166.81; HRMS (APESI,
M+) C18H14Cl2BrN5O2: m/z calcd 500.0868, m/z found
500.0851 (M+).
16. McCullagh, P.; Nelder, J. A. In Generalized Linear
Models, second ed.; Chapman and Hall: London, 1989;
pp 1–511.
17. Bennett, D. L.; Cheek, T. R.; Berridge, M. J.; DeSmedt,
H.; Parys, J. B.; Missiaen, L.; Bootman, M. D. J. Biol.
Chem. 1996, 271, 6356.
1
1661, 1638, 1578, 1529, 1503, 1463, 1416, 1377 cmÀ1; H
NMR (400 MHz CDCl3) d 1.25 (s, 3 H), 2.17 (s, 3H), 2.94
(d, 2H, J = 4.8 Hz), 6.20 (q, 1H, J = 4.8 Hz), 7.12 (s, 1H),
7.20 (d, 1H, J = 2.4 Hz), 7.23 (d, 1H, J = 2.4 Hz), 7.37 (dd,
1H, J = 4.7, 8.0 Hz), 7.84 (dd, 1H, 1.7, 8.0 Hz), 8.46 (dd,
1H, J = 1.7, 4.7 Hz), 10.08 (s, 1H); 13C NMR (100 MHz,
DMSO) d 18.35, 26.77, 111.34, 126.03, 127.26, 127.48,
18. IMR32 cells challenged with caffeine exhibit a dose-
dependent release of internal Ca2+ stores.