Models for extradiol cleaving catechol dioxygenases: Syntheses, structures, and reactivities of iron(II)-monoanionic catecholate complexes

Article abstract of DOI:10.1021/ic001185d

Crystallographic and spectroscopic studies of extradiol cleaving catechol dioxygenases indicate that the enzyme-substrate complexes have both an iron(II) center and a monoanionic catecholate. Herein we report a series of iron(II)-monoanionic catecholate complexes, [(L)Fe^II(catH)](X) (1a, L = 6-Me₃-TPA (tris(6-methyl-2-pyridyl-methyl)amine), catH = CatH (1,2-catecholate monoanion); 1b, L = 6-Me₃-TPA, catH = DBCH (3,5-di-tert-butyl-1,2-catecholate monoanion); 1c, L = 6-Me₂-bpmcn (N, N′-dimethyl-N, N′-M-bis(6-methyl-2-pyridylmethyl)-trans-1,2-diaminocyclohexane), catH = CatH; 1d, L = 6-Me₂-bpmcn, catH = DBCH), that model such enzyme complexes. The crystal structure of [(6-Me₂-bpmcn)Fe^II(DBCH)]⁺ (1d) shows that the DBCH ligand binds to the iron asymmetrically as previously reported for 1b, with two distinct Fe-O bonds of 1.943(1) and 2.344(1) A. Complexes 1 react with O₂ or NO to afford blue-purple iron(III)-catecholate dianion complexes, [(L)Fe^III(cat)⁺ (2). Interestingly, crystallographically characterized 2d, isolated from either reaction, has the N-methyl groups in a syn configuration, in contrast to the anti configuration of the precursor complex, so epimerization of the bound ligand must occur in the course of isolating 2d. This notion is supported by the fact that the UV-vis and EPR properties of in situ generated 2d(anti) differ from those of isolated 2d(syn). While the conversion of 1 to 2 in the presence of O₂ occurs without an obvious intermediate, that in the presence of NO proceeds via a metastable S = 3/2 [(L)Fe(catH)(NO)]⁺ adduct 3, which can only be observed spectroscopically but not isolated. Intermediates 3a and 3b subsequently disproportionate to afford two distinct complexes, [(6-Me₃-TPA)Fe^III(cat)]⁺ (2a and 2b) and [(6-Me₃-TPA)Fe(NO)₂]⁺ (4) in comparable yield, while 3d converts to 2d in 90% yield. Complexes 2b and anti-2d react further with O₂ over a 24 h period and afford a high yield of cleavage products. Product analysis shows that the products mainly derive from intradiol cleavage but with a small extent of extradiol cleavage (89:3% for 2b and 78:12% for anti-2d). The small amounts of the extradiol cleavage products observed may be due to the dissociation of an α-methyl substituted pyridyl arm, generating a complex with a tridentate ligand. Surprisingly, syn-2d does not react with O₂ over the course of 4 days. These results suggest that there are a number of factors that influence the mode and rate of cleavage of catechols coordinated to iron centers.

Full text of DOI:10.1021/ic001185d

Inorg. Chem. 2001, 40, 3181-3190
3181
Models for Extradiol Cleaving Catechol Dioxygenases: Syntheses, Structures, and
Reactivities of Iron(II)-Monoanionic Catecholate Complexes
Du-Hwan Jo, Yu-Min Chiou, and Lawrence Que, Jr.*
Department of Chemistry and Center for Metals in Biocatalysis, University of Minnesota,
Minneapolis, Minnesota 55455
ReceiVed October 27, 2000
Crystallographic and spectroscopic studies of extradiol cleaving catechol dioxygenases indicate that the enzyme-
substrate complexes have both an iron(II) center and a monoanionic catecholate. Herein we report a series of
iron(II)-monoanionic catecholate complexes, [(L)FeII(catH)](X) (1a, L ) 6-Me3-TPA (tris(6-methyl-2-pyridyl-
methyl)amine), catH ) CatH (1,2-catecholate monoanion); 1b, L ) 6-Me3-TPA, catH ) DBCH (3,5-di-tert-
butyl-1,2-catecholate monoanion); 1c, L ) 6-Me2-bpmcn (N,N′-dimethyl-N,N′-bis(6-methyl-2-pyridylmethyl)-
trans-1,2-diaminocyclohexane), catH ) CatH; 1d, L ) 6-Me2-bpmcn, catH ) DBCH), that model such enzyme
complexes. The crystal structure of [(6-Me2-bpmcn)FeII(DBCH)]+ (1d) shows that the DBCH ligand binds to the
iron asymmetrically as previously reported for 1b, with two distinct Fe-O bonds of 1.943(1) and 2.344(1) Å.
Complexes 1 react with O2 or NO to afford blue-purple iron(III)-catecholate dianion complexes, [(L)FeIII(cat)]+
(2). Interestingly, crystallographically characterized 2d, isolated from either reaction, has the N-methyl groups in
a syn configuration, in contrast to the anti configuration of the precursor complex, so epimerization of the bound
ligand must occur in the course of isolating 2d. This notion is supported by the fact that the UV-vis and EPR
properties of in situ generated 2d(anti) differ from those of isolated 2d(syn). While the conversion of 1 to 2 in
the presence of O2 occurs without an obvious intermediate, that in the presence of NO proceeds via a metastable
S ) 3/2 [(L)Fe(catH)(NO)]+ adduct 3, which can only be observed spectroscopically but not isolated. Intermediates
3a and 3b subsequently disproportionate to afford two distinct complexes, [(6-Me3-TPA)FeIII(cat)]+ (2a and 2b)
and [(6-Me3-TPA)Fe(NO)2]+ (4) in comparable yield, while 3d converts to 2d in 90% yield. Complexes 2b and
anti-2d react further with O2 over a 24 h period and afford a high yield of cleavage products. Product analysis
shows that the products mainly derive from intradiol cleavage but with a small extent of extradiol cleavage (89:
3% for 2b and 78:12% for anti-2d). The small amounts of the extradiol cleavage products observed may be due
to the dissociation of an R-methyl substituted pyridyl arm, generating a complex with a tridentate ligand.
Surprisingly, syn-2d does not react with O2 over the course of 4 days. These results suggest that there are a
number of factors that influence the mode and rate of cleavage of catechols coordinated to iron centers.
Introduction
dihydroxybiphenyl 1,2-dioxygenase (1,2-BphC) and protocat-
echuate 4,5-dioxygenase (4,5-PCD) reveal that the iron active
The catechol dioxygenases are non-heme enzymes that
catalyze the oxidative cleavage of catechols in the final step
for the degradation of natural aromatic molecules into aliphatic
products.1 Cleavage of the C-C bond adjacent to the enediol
unit (extradiol) represents the more common oxidative cleavage
pathway in nature, utilizing enzymes that have Fe(II) or Mn(II)
in the active site. The crystal structures of these enzymes reveal
a square pyramidal geometry at the iron center that consists of
three endogenous ligands (two histidines and one glutamate)
in a facial array and two solvent molecules.2 In addition, the
structures of the enzyme-substrate (E‚S) complexes of 2,3-
site adopts a 5-coordinate structure with the three endogenous
ligands and a chelated catecholate.3 While the catecholate
coordinates in a bidentate fashion, it does so asymmetrically
with one Fe-Ocat bond 0.2 Å shorter than the other. A similar
bond asymmetry has also been observed in EXAFS studies of
the catechol 2,3-dioxygenase (2,3-CTD)-catechol complex4 and
ascribed to a monoanionic substrate. The remaining coordination
site is presumably reserved for O2, as suggested by the binding
of nitric oxide (NO) as a dioxygen surrogate,1a,5 which converts
the normally EPR-silent mononuclear high-spin Fe(II) center
3
(S ) 2) into an EPR active S )
/ species. This ternary
2
* To whom correspondence should be sent. Fax: (+1) 612-624-7029.
E-mail: que@chem.umn.edu.
(3) (a) Sugiyama, K.; Senda, T.; Narita, H.; Yamamoto, T.; Kimbara, K.;
Fukuda, M.; Yano, K.; Mitsui, Y. Proc. Jpn Acad., Ser. B 1995, 71,
32-35. (b) Senda, T.; Sugiyama, K.; Narita, H.; Yamamoto, T.;
Kimbara, K.; Fukuda, M.; Sato, M.; Yano, K.; Mitsui, Y. J. Mol. Biol.
1996, 255, 735-752. (c) Sugimoto, K.; Senda, T.; Aoshima, H.; Masai,
E.; Fukuda, M.; Mitsui, Y. Structure 1999, 7, 953-965.
(4) Shu, L.; Chiou, Y.-M.; Orville, A. M.; Miller, M. A.; Lipscomb, J.
D.; Que, L., Jr. Biochemistry 1995, 34, 6649-6659.
(5) (a) Arciero, D. M.; Lipscomb, J. D.; Huynh, B. H.; Kent, T. A.; Mu¨nck,
E. J. Biol. Chem. 1983, 258, 14981-14991. (b) Arciero, D. M.; Orville,
A. M.; Lipscomb, J. D. J. Biol. Chem. 1985, 260, 14035-14044. (c)
Arciero, D. M.; Lipscomb, J. D. J. Biol. Chem. 1986, 261, 2170-
2178.
(1) (a) Lipscomb, J. D.; Orville, A. M. Metal Ions Biol. Syst. 1992, 28,
243-298. (b) Que, L., Jr. In Bioinorganic Catalysis, 2nd ed.; Reedijk,
J., Bouwman, E., Eds.; Marcel Dekker: New York, 1999; pp 269-
321. (c) Solomon, E. I.; Brunold, T. C.; Davis, M. I.; Kemsley, J. N.;
Lee, S.-K.; Lehnert, N.; Neese, F.; Skulan, A. J.; Yang, Y.-S.; Zhou,
J. Chem. ReV. 2000, 100, 235-349. (d) Kru¨ger, H.-J. In Biomimetic
Oxidations Catalyzed by Transition Metal Complexes; Meunier, B.,
Ed.; Imperial College Press: London, 2000; pp 363-413.
(2) (a) Han, S.; Eltis, L. D.; Timmis, K. N.; Muchmore, S. W.; Bolin, J.
T. Science 1995, 270, 976-980. (b) Kita, A.; Kita, S.-I.; Fusisawa,
I.; Inaka, K.; Ishida, T.; Horiiki, K.; Nozaki, M.; Miki, K. Structure
1999, 7, 25-34.
10.1021/ic001185d CCC: $20.00 © 2001 American Chemical Society
Published on Web 05/17/2001

3182 Inorganic Chemistry, Vol. 40, No. 13, 2001
Jo et al.
was then obtained by methylation of the secondary di-amine using
formic acid-formaldehyde. A mixture of the secondary amine (1.61
g, 5 mmol), formic acid (15 mL, 88%), and formaldehyde (15 mL,
37%) was refluxed at 90 °C with stirring for 3 days. The solution was
cooled in an ice bath and then brought to pH 12 with a saturated solution
of NaOH. The resulting solution was extracted with chloroform, dried
over Na2SO4, and evaporated to give a light yellow oil, trans-N,N′-
dimethyl-N,N′-bis(6-methyl-2-pyridylmethyl)-1,2-cyclohexadiamine.
enzyme-substrate-NO complex has been proposed to be a
model for the iron-dioxygen intermediate in the enzymatic
reaction mechanism.5b-c
To date, several synthetic Fe(III)-catecholate complexes
serve as functional models for extradiol cleaving dioxygenases,6
but there is only one mononuclear Fe(II)-catecholate complex
that is structurally characterized, [(6-Me3-TPA)FeII(DBCH)]-
(ClO4), a result reported in an earlier communication.7 In this
paper, we report the synthesis and characterization of a series
of mononuclear Fe(II)-catecholate complexes, [(L)FeII(catH)](X)
(L ) 6-Me3-TPA, 6-Me2-bpmcn; catH ) CatH, DBCH; X )
counterions)8 and their reactivities toward O2 and NO in order
to evaluate the plausibility of structural and reactivity features
postulated for the extradiol cleaving dioxygenases.
1
Yield: 1.41 g (80%). H NMR (CDCl3, ppm): 7.49(t, 2H, 4H-py),
7.41(d, 2H, 3H-py), 6.95(d, 2H, 5H-py), 3.90(d, 2H, AB, J ) 9.6 Hz;
NCH2), 3.75(d, 2H, AB, J ) 9.6 Hz; NCH2), 2.66(t, 2H, NCH), 2.53-
(s, 6H; py-CH3), 2.30(s, 6H; NCH3), 1.97(d, 2H), 1.76(d, 2H), 1.1∼1.4-
(m, 4H). 13C NMR (CDCl3, ppm): 160, 157, 136.5, 121, 119.7,
64.6(NCH), 60.3(NCH2), 36.6(pyCH3), 25.8, 25.6, 24.4(NCH3).
Metal Complexes. Caution. Perchlorate salts of metal complexes
with organic ligands are potentially explosive and should be handled
with great caution. NO is a poisonous gas and should only be used
with proper ventilation.
Experimental Section
[(6-Me3-TPA)FeII(CatH)](ClO4) (1a). A methanolic solution of
6-Me3-TPA (0.36 g, 1.0 mmol) was added to a methanol solution (30
mL) of Fe(ClO4)2‚6H2O (0.36 g, 1.0 mmol) and stirred for 10 min. To
this solution was slowly added a methanolic solution of CatH2 (0.11 g,
1.0 mmol) and triethylamine (0.10 g, 1.0 mmol). The resulting yellow
solution was stirred for 30 min under argon and the yellow micro-
crystalline product was filtered and dried under vacuum. Yield: 0.56
g (90%). Anal. Calcd. for 1a, C27H29ClFeN4O6: C, 54.33; H, 4.90; N,
9.39, Cl, 5.94. Found: C, 54.16; H, 4.70; N, 9.30; Cl, 6.16. UV-vis
[λmax, nm (ꢀ, M-1cm-1) in CH3CN]: 386 (2000).
Materials and Procedures. Reagents were purchased from com-
mercial sources and used as received unless otherwise noted. The
catechols (CatH2 and DBCH2) were purified by sublimation or
recrystallization from hexane. Preparation and handling of air-sensitive
materials were carried out under an argon atmosphere using standard
Schlenk techniques or in an anaerobic glovebox. All solvents were
distilled using standard methods under argon. Nitric oxide (99+%,
Matheson Gas Products, Inc.) was passed through a column of NaOH
pellets to prevent nitric acid contamination and the line purged with
argon gas for 30 min before using to ensure removal of any adventitious
O2.
[(6-Me3-TPA)FeII(DBCH)](ClO4) (1b). This compound was pre-
pared in a manner analogous to that described for 1a except that DBCH2
(0.22 g, 1.0 mmol) was used in place of CatH2.7 Yield: 0.62 g (82%).
Crystallographic quality crystals were obtained by vapor diffusion of
Et2O into an ethanol solution of 1b. Anal. Calcd. for 1b‚EtOH, C37H51-
ClFeN4O7: C, 58.85; H, 6.81; N, 7.42, Cl, 4.69. Found: C, 58.58; H,
6.81; N, 7.37; Cl, 4.93. UV-vis [λmax, nm (ꢀ, M-1cm-1) in CH3CN]:
395 (2200).
[(6-Me2-bpmcn)FeII(CatH)](CF3SO3) (1c). A CH2Cl2 solution of
6-Me2-bpmen (0.353 g, 1.0 mmol) was added to a CH2Cl2 solution
(30 mL) of Fe(CF3SO3)2 (0.354 g, 1.0 mmol) and stirred for 10 min.
To this solution was slowly added a CH2Cl2 solution of CatH2 (0.11 g,
1.0 mmol) and triethylamine (0.10 g, 1.0 mmol). The mixture was stirred
for 30 min and layered with diethyl ether to give greenish-yellow
crystals. Yield: 0.43 g (65%). Anal. Calcd. for 1c‚Et2O, C33H47F3-
FeN4O6S: C, 53.51; H, 6.40; N, 7.57, S, 4.33. Found: C, 53.35; H,
5.50; N, 7.73; S, 4.40. UV-vis [λmax, nm (ꢀ, M-1cm-1) in CH3CN]:
380 (900).
[(6-Me2-bpmcn)FeII(DBCH)](CF3SO3) (1d). This compound was
prepared in a manner analogous to that described for 1c except that
DBCH2 (0.22 g, 1.0 mmol) was used in place of CatH2. Crystallographic
quality crystals were obtained by vapor diffusion of diethyl ether into
a CH2Cl2 solution. Yield: 0.55 g (70%). Anal. Calcd. for 1d‚0.5CH2-
Cl2, C37.5H54ClF3FeN4O5S: C, 54.85; H, 6.63; N, 6.82, S, 3.90.
Found: C, 55.14; H, 6.62; N, 6.62; S, 3.90. UV-vis [λmax, nm (ꢀ,
M-1cm-1) in CH3CN]: 384 (1100).
6-Me3-TPA and 6-Me2-bpmcn. 6-Me3-TPA was synthesized ac-
cording to published procedures,9 while 6-Me2-bpmcn was prepared
by the following method. To an ethanolic solution (100 mL) of
6-methyl-pyridine 2-carboxaldehyde (4.8 g, 20 mmol) was added
racemic trans-1,2-diaminocyclohexane (2.28 g, 40 mmol), and the
mixture was stirred for 2 h at room temperature. The solution was
concentrated to give crystals of N,N′-bis(6-methyl-2-pyridylmethyl)-
1,2-cyclohexadiimine, which was filtered and washed with cold ethanol.
1
Yield: 6.08 g (90%). H NMR (CDCl3, ppm): 8.30(s, 2H, NdCH),
7.70(d, 2H, 3H-py), 7.52(t, 2H, 4H-py), 7.07(d, 2H, 5H-py), 3.51(t,
2H, NCH), 2.52(s, 6H, py-CH3), 1.81(m, 6H), 1.50(s, 2H). 13C NMR
(CDCl3, ppm): 161.6(N)CH), 157.6, 154.14, 136.5, 124, 118, 73.5-
(NCH), 32.6(py-CH3), 24.3, 24.2. The reduction of the di-imine was
then carried out using KBH4 in dry methanol. To the methanolic solution
(50 mL) of di-imine (3.2 g, 10 mmol) was slowly added excess KBH4
and the mixture was refluxed for 1 h. The solution was cooled and
then quenched with cold water (20 mL). Following chloroform
extraction (3 × 50 mL), the organic layer was dried with Na2SO4.
Removal of chloroform yielded the oily N,N′-bis(6-methyl-2-pyridyl-
methyl)-1,2-cyclohexadiamine. Yield: 3.07 g (95%). 1H NMR (CDCl3,
ppm): 7.50(t, 2H, 4H-py), 7.21(d, 2H, 3H-py), 6.97(d, 2H, 5H-py),
4.01(d, 2H, AB, J ) 14 Hz; NCH2), 3.97(d, 2H, AB, J ) 14 Hz; NCH2),
2.51(s, 6H, py-CH3), 2.31(2H, NCH), 2.13(2H), 1.70(2H), 1.04-1.26-
(m, 4H). 13C NMR (CDCl3, ppm): 159.9, 157.5, 121.1, 119.0, 61.3-
(NCH2), 52.4(NCH), 31.6(py-CH3), 24.9, 24.4. 6-Me2-bpmcn ligand
[(6-Me3-TPA)FeIII(Cat)](BPh4) (2a). Complex 1a (60 mg, 0.1
mmol) in 20 mL methanol was reacted with NO, turning the yellow
solution brown in less than 1 min. Addition of NaBPh4 to the methanol
solution, followed by vapor diffusion of diethyl ether at room
temperature for 2∼3 days, resulted in the formation of two distinct
crystal forms in comparable yields, purple-blue needle crystals of 2a,
and brown prismatic crystals of 4 (see below). The crystals of 2a were
manually separated from the brown crystals of 4. Anal. Calcd. for 2a,
C51H48BFeN4O2: C, 75.10; H, 5.93; N, 6.87. Found: C, 74.85; H, 5.89;
N, 7.01. UV-vis [λmax, nm (ꢀ, M-1cm-1) in CH3CN]: 540 (2100),
940 (3200).
[(6-Me3-TPA)FeIII(DBC)](ClO4) (2b). Complex 1b (71 mg, 0.1
mmol) in 20 mL CH2Cl2 was reacted with NO to form a brown solution.
Diffusion of hexane into this solution at room temperature for 3-4
days resulted in the formation of two distinct crystal forms, blue crystals
of 2b and brown crystals of 4 (see below). The crystals of 2b were
manually separated from the brown crystals of 4. Anal. Calcd. for 2b,
(6) (a) Dei, A.; Gatteschi, D.; Pardi, L. Inorg. Chem. 1993, 32, 1389-
1395. (b) Ito, M.; Que, L., Jr. Angew. Chem., Int. Ed. Engl. 1997, 36,
1342-1344. (c) Ogihara, T.; Hikichi, S.; Akika, M.; Moro-oka, Y.
Inorg. Chem. 1998, 37, 2614-2615.
(7) Chiou, Y.-M.; Que, L., Jr. Inorg. Chem. 1995, 34, 3577-3578.
(8) Abbreviations: BF, benzoylformate; bpeen, trans-N,N′-diethyl-N,N′-
bis(2-pyridylmethyl)-1,2-ethylenediamine; BPG, N,N-bis(2-pyridyl-
methyl)glycine; bpmcn, trans-N,N′-dimethyl-N,N′-bis(2-pyridylmethyl)-
1,2-cyclohexadiamine; bpmen, trans-N,N′-dimethyl-N,N′-bis(2-pyridyl-
methyl)-1,2-ethylenediamine; cat, catecholate dianion (Cat or DBC);
catH, catecholate monoanion (DBCH or CatH); CatH, 1,2-catecholate
monoanion; DBCH, 3,5-di-tert-butyl-1,2-catecholate monoanion; 6-Me2-
bpmcn, trans-N,N′-dimethyl-N,N′-bis(6-methyl-2-pyridylmethyl)-1,2-
cyclohexadiamine; 6-Me3-TPA, tris(6-methyl-2-pyridylmethyl)amine;
NTA, N,N-bis(carboxymethyl)glycine; TPA, tris(2-pyridylmethyl)-
amine; Tpi-Pr2, hydrotris(3,5-di-iso-propyl-1-pyrazolyl)borate.
(9) Da Mota, M. M.; Rodgers, J.; Nelson, S. M. J. Chem. Soc. A 1969,
2036-2044.

Extradiol Cleaving Catechol Dioxygenases
Inorganic Chemistry, Vol. 40, No. 13, 2001 3183
C35H44ClFeN4O6: C, 59.37; H, 6.26; N, 7.91, Cl, 5.01. Found: C, 59.54;
H, 6.08; N, 7.76; Cl, 5.23. UV-vis [λmax, nm (ꢀ, M-1cm-1) in CH3-
CN]: 600 (2200), 1020 (3400).
[(6-Me2-bpmcn)FeIII(Cat)](CF3SO3) (2c). Complex 1c (78 mg, 0.1
mmol) was exposed to O2 in CH2Cl2 to form 2c. Diffusion of diethyl
ether into this solution at 4 °C for several months afforded microcrystals
of 2c. UV-vis [λmax, nm (ꢀ, M-1cm-1) in CH3CN]: 535 (2300), 874
(3000).
methods. All non-hydrogen atoms were refined with anisotropic thermal
parameters. Hydrogen atoms were placed at calculated positions and
refined as riding atoms. Final cycles of refinement converged with
discrepancy indices of R1 ) 0.0482, wR2 ) 0.0956 for 2a and R1 )
0.0539, wR2 ) 0.1290 for 4.
Physical Measurements. Elemental analyses (C, H, Cl, N, S) were
1
performed at M-H-W Laboratories (Phoenix, AZ). H- and 13C NMR
spectra were recorded on a Varian VXR-300 or IBM AC-300
spectrometer at room temperature. X-band EPR spectra were performed
at liquid helium temperature on a Varian E-109 spectrometer or a Bruker
E-500 spectrometer equipped with an Oxford ESR-10 cryostat. The
magnetic field was calibrated with a gaussmeter, and the microwave
frequency was measured with a counter. UV-visible spectra were
recorded on a Hewlett-Packard 8452 or 8453 diode array spectrometer.
Infrared spectra of KBr pellets were obtained on a Mattson Polaris
FT-IR spectrometer. Standard organic product analyses were performed
using a Hewlett-Packard HP-6890 Series gas chromatograph equipped
with a J & W Scientific DB-5 column and a flame ionization detector
or a reverse-phase isocratic HPLC (Waters 6000A isocratic system;
Kratos 769Z variable wavelength detector, 240 nm; Whatman Partisil
ODS-5 C18 column). Mass spectral analysis was carried out on a
Hewlett-Packard Model 5989B mass spectrometer.
[(6-Me2-bpmcn)FeIII(DBC)](CF3SO3) (2d). Complex 1d (78 mg,
0.1 mmol) was reacted with NO in 20 mL CH2Cl2. Diffusion of hexane
into this solution at room temperature for 3-4 days resulted in the
formation of purple-blue crystals of 2d in excellent yield (70 mg, 90%).
Anal. Calcd. for 2d, C37H52F3FeN4O5S: C, 57.14; H, 6.74, N; 7.20, S;
4.12. Found: C, 56.68; H, 6.77; N, 6.81; S, 3.88. UV-vis [λmax, nm
(ꢀ, M-1cm-1) in CH3CN]: 598 (2200), 950 (3100). Complex 2d was
also obtained by brief exposure of a solution of 1d (0.39 g, 0.5 mmol)
in 20 mL CH2Cl2 to O2, turning the solution blue in color. Addition of
diethyl ether to this solution and storage at 4 °C for 2 months afforded
blue crystals that had the same spectroscopic properties as isolated 2d
from the reaction of 1d with NO. Complex 2d was also obtained by
direct synthesis. To a CH3CN (20 mL) solution of Fe(ClO4)3‚10H2O
(0.267 g, 0.5 mmol) was added a CH3CN solution of 6-Me2-bpmcn
(0.177 g, 0.5 mmol). This solution was stirred for 30 min, then DBCH2
(0.11 g, 0.5 mmol) and triethylamine (0.20 g, 1 mmol) were added.
After stirring for 2 h, the reaction solution was evaporated to dryness.
The solid residue was dissolved in CH2Cl2, and this solution was layered
with diethyl ether; upon standing at 4 °C, diffraction quality crystals
of the perchlorate salt of 2d were obtained. The structure of the
perchlorate salt (unpublished data) matched that of the triflate salt
reported in this paper.
Oxygenation Studies. Reactivity studies were performed in CH3-
CN under an O2 atmosphere. In a typical stoichoimetric reaction,
complex 1 (0.05 mmol) was reacted with O2 in 20 mL of solvent. The
pale greenish-yellow color of the complex turned intense purple-blue
immediately, and the resulting solution gradually faded to yellow-brown
as the reaction proceeded. The solution was concentrated and extracted
with diethyl ether (3 × 5 mL). The solid residue was acidified with 3
N HCl to pH 3 to effect decomposition of the metal complexes and
extracted with diethyl ether (3 × 5 mL). The combined ether extract
was dried over anhydrous Na2SO4 and concentrated and the extract
was then submitted for GC, GC-MS (EI), and reverse-phase isocratic
HPLC separation (conditions are the same as previously reported10).
Characterization of Oxygenation Products. The intradiol cleavage
products, 3,5-di-tert-butyl-1-oxacyclohepta-3,5-diene-2,7-dione and 3,5-
di-tert-butyl-5-carboxymethyl-2-furanone, were identified by using GC-
[(6-Me3-TPA)Fe(NO)2](X) (4). Complex 4 was obtained from the
reaction of 1a or 1b with NO. As described earlier, these reactions
gave a mixture of products, 4 and 2a or 2b, which could be separated
manually. Anal. Calcd. for 4 (X ) ClO4), C21H24ClFeN6O6: C, 46.05;
H, 4.42; N, 15.34, Cl, 6.47. Found: C, 45.75; H, 4.65; N, 15.54; Cl,
6.37. Anal. Calcd. for 4 (X ) BPh4) C45H44BFeN6O2: C, 70.40; H,
5.78; N, 10.95. Found: C, 70.50; H, 5.86; N, 11.13. UV-vis [λmax
,
nm (ꢀ, M-1cm-1) in CH3CN]: 332 (sh, 1700), 374 (sh, 1300), 460 (sh,
360), 820 (200) nm. IR (cm-1, KBr pellet): ν(NO) 1801(s), 1726(s).
EPR (CH3CN at 5 K): g ) 2.02.
1
MS (EI) and H- and 13C NMR techniques. These data are consistent
with the published data,11 and authentic compounds were prepared by
treating 3,5-di-tert-butyl-o-benzoquinone with excess m-chloroperoxy-
benzoic acid at 0 °C.12 The two structural isomers of the extradiol
cleavage products, 3,5- and 4,6-di-tert-butyl-2-pyrone, were also
identified by using the same spectroscopic methods and compared with
the authentic compounds prepared according to the published proce-
dures.13
Crystallographic Studies. [(6-Me2-bpmcn)FeII(DBCH)](CF3SO3)
(1d) and [(6-Me2-bpmcn)FeIII(DBC)](CF3SO3) (2d). Yellow (1d) and
purple-blue (2d) crystals suitable for X-ray diffraction studies were
grown from CH2Cl2/diethyl ether and CH2Cl2/hexane solvent pairs,
respectively. The crystals were attached to glass fibers with heavy-
weight oil and mounted on a Siemens SMART system for data
collection at 173(2) K. An initial set of cell constants was calculated
from reflections harvested from three sets of 20 frames. These initial
sets of frames are oriented such that orthogonal wedges of reciprocal
space were surveyed. This produces orientation matrixes determined
from 204 reflections for 1d and 62 reflections for 2d. Final cell constants
are calculated from a set of 8192 (for 1d) and 3977 (for 2d) strong
reflections from actual data collection. Crystallographic data and
experimental conditions are summarized in Table 1. Both structures
were solved by direct methods. All non-hydrogen atoms were refined
anisotropically and hydrogen atoms were placed in ideal positions and
refined as riding atoms with relative isotropic displacement parameters.
The final full matrix least squares refinement coverged to R1 ) 0.0373,
wR2 ) 0.0909 for 1d and R1 ) 0.0563, wR2 ) 0.1213 for 2d.
[(6-Me3-TPA)FeIII(Cat)](BPh4) (2a) and [(6-Me3-TPA)Fe(NO)2]-
(ClO4) (4). Crystal data for a purple-blue crystal of 2a and a red-brown
prismatic crystal of 4 individually mounted on glass fibers with heavy-
weight oil were collected at 297 (2) and 177 (2) K, respectively.
Crystallographic data and experimental conditions are listed in Table
1. Measurements were carried out on an Enraf-Nonius CAD-4 diffrac-
tometer with graphite monochromated Mo KR radiation. Cell constants
were obtained from a least-squares refinements of the setting angles
for 50 reflections for 2a and for 46 reflections for 4. Lorentz and
polarization corrections were applied to the data and absorption
corrections based on ψ scans were carried out a maximum 2θ value of
52.0° for 2a and 56.0° for 4. Both structures were solved by direct
O2 Uptake. Manometric measurements of dioxygen uptake were
carried out using a similar apparatus as described previously.14 The
number of moles of O2 taken up was calculated with the ideal gas law.
The absolute accuracy of the reaction system was checked by using
Vaska’s complex, [IrCl(CO)(PPh3)2] (O2/Ir ) 1:1).15 For three replicate
runs using 0.2 mmol of 1b at 22 °C, the O2:Fe ratio was found to be
1.3(1).
Results and Discussion
The Fe(II)-catecholate complexes containing the tetradentate
ligands 6-Me3-TPA and 6-Me2-bpmcn (1) have been synthesized
to serve as models for the enzyme-substrate complexes of the
extradiol cleaving catechol dioxygenases. Treatment of FeIIX2
(X ) ClO4 or CF3SO3) with equimolar quantities of L, catechol
(10) Cox, D. D.; Que, L., Jr. J. Am. Chem. Soc. 1988, 110, 8085-8092.
(11) Jang, H. G.; Cox, D. D.; Que, L., Jr. J. Am. Chem. Soc. 1991, 113,
9200-9204.
(12) Demmin, T. R.; Rogic´, M. M. J. Org. Chem. 1980, 45, 1153-1156.
(13) Matsumoto, M.; Kuroda, K. J. Am. Chem. Soc. 1982, 104, 1433-
1434.
(14) (a) Karlin, K. D.; Cruse, R. W.; Gultneh, Y.; Farooq, A.; Hayes, J.
C.; Zubieta, J. J. Am. Chem. Soc. 1987, 109, 2668-2679. (b) Tolman,
W. B.; Liu, S.; Bentsen, J. G.; Lippard, S. J. J. Am. Chem. Soc. 1991,
113, 152-164.
(15) Vaska, L. Science 1963, 140, 809-810.

3184 Inorganic Chemistry, Vol. 40, No. 13, 2001
Table 1. Crystallographic Data of 1d, 2a, 2d, and 4
Jo et al.
1d‚0.5CH2Cl2
2a
empirical formula
formula weight (g/mol)
crystal habit, color
crystal system
space group
a (Å)
C
37.5H54ClF3FeN4O5S
C51H48BFeN4O2
815.59
prism, purple-blue
triclinic
P1h
9.696(4)
14.704(7)
15.315(13)
89.65(7)
85.33(7)
81.43(4)
821.21
block, yellow
monoclinic
P21/c
10.3726(5)
21.903(1)
17.6723(8)
90
97.121(1)
90
3984.0(3)
4
b (Å)
c (Å)
R, deg
â, deg
γ, deg
V (Å3)
2152(2)
2
1.259
Z
D
calc (g/cm3)
1.369
temperature (K)
θ range (deg)
hkl ranges
173(2)
297(2)
1.49 ∼ 27.49
-13, -28, -22 to 12, 27, 21
31610
8991 (Rint ) 0.0306)
Mo-KR, 0.71073
0.558
1.33 ∼ 25.98
0, -17, -18 to 11, 18, 18
8941
8415 (Rint ) 0.0359)
Mo-KR, 0.71073
0.396
no. of reflections collected
no. of unique reflections
radiation (Å)
absorption coefficient (mm-1
R1/wR2 (I > 2σ(I))a
R1/wR2 (all data)
)
0.0373/0.0909
0.0529/0.0965
0.0482/0.0956
0.0812/0.1094
2d
4
empirical formula
formula weight (g/mol)
crystal habit, color
crystal system
space group
a (Å)
C37H52F3FeN4O5S
777.74
needle, purple
monoclinic
P21/n
8.517(1)
50.379(6)
9.874(1)
C21H24ClFeN6O6
547.76
prism, red-brown
monoclinic
P21/n
11.934(4)
16.540(15)
12.408(4)
90
101.01(3)
90
b (Å)
c (Å)
R, deg
90
â, deg
105.938(3)
90
4073.8(9)
4
γ, deg
V (Å3)
2404(3)
4
1.513
Z
D
calc (g/cm3)
1.268
temperature (K)
θ range (deg)
hkl ranges
173(2)
177(2)
1.62 ∼ 27.51
-10, -59, -12 to 11, 65, 10
25658
9260 (Rint ) 0.0653)
Mo-KR, 0.71073
0.479
2.08 ∼ 28.0
-7, -21, -16 to 15, 21, 16
5800
5800 (Rint ) 0.033)
Mo-KR, 0.71073
0.788
no. of reflections collected
no. of unique reflections
radiation (Å)
absorption coefficient (mm-1
R1/wR2 (I > 2σ(I))a
R1/wR2 (all data)
)
0.0563/0.1213
0.1115/0.1380
0.0539/0.1290
0.0976/0.1512
a R1 ) ∑||Fo| - |Fc||/∑|Fo|. wR2 ) [∑[w(Fo - Fc2)2]/∑[w(Fo )2]]1/2, where w ) q/σ2(Fo ) + (a*P)2 + b*P.
2
2
2
Scheme 1
(catH2), and triethylamine under N2 affords pale green solutions
that yield [(L)FeII(catH)](X) (1a, L ) 6-Me3-TPA, catH ) CatH;
1b, L ) 6-Me3-TPA, catH ) DBCH; 1c, L ) 6-Me2-bpmcn,
catH ) CatH; 1d, L ) 6-Me2-bpmcn, catH ) DBCH) as pale
greenish yellow solids in 60∼90% yield. These Fe(II)-
catecholate complexes are very air-sensitive in solution and
exhibit a strong and broad νOH band at around 3300 cm-1 in
the solid state. The electronic spectra of these complexes show
only one intense absorption band near 390 nm, which gives
rise to their light yellow color. This feature likely arises from
an iron(II)-to-pyridine charge-transfer transition, as the absorp-
tion intensities of 1a and 1b are twice as large as those of 1c
and 1d due to the greater number of pendant pyridines. These
Fe(II)-catecholate complexes react with O2 to afford corre-
sponding Fe(III)-catecholate complexes (2) which in turn
undergo oxidative cleavage of the bound catecholate. In addition,
reactions of the Fe(II) complexes with NO give rise to transient
iron-nitrosyl intermediate adducts (3). Although these adducts
can be observed spectroscopically, they could not be isolated.
Instead, 3a and 3b decompose to afford two distinct species:
[(6-Me3-TPA)FeIII(cat)]+ (2a and 2b) and [(6-Me3-TPA)Fe-
(NO)2]+ (4) as final products, while 3d yields only the
corresponding Fe(III)-catecholate complex (2d). Scheme 2
summarizes the reactions of complexes 1 with O2 and NO. The
crystal structures of 1d, 2a, 2d, and 4 were solved and are
described below.

Extradiol Cleaving Catechol Dioxygenases
Inorganic Chemistry, Vol. 40, No. 13, 2001 3185
Scheme 2
Scheme 3
amine ligand (N2), while the longer Fe1-O1 bond is trans to
a pyridine nitrogen (N3). The 0.40 Å difference in Fe-Ocat
bonds and the presence of one counterion per molecule unit
strongly suggest that the catecholate ligand is a monoanion.
Indeed the difference in Fe-Ocat bond lengths for 1d is 0.09 Å
larger than the 0.31 Å difference found for 1b.7 Deprotonation
of the O2 oxygen is consistent with model studies,10 showing
that the O2 oxygen is the more basic ligand and affords a
stronger interaction with the iron center. This deprotonation
occurs despite potential steric interactions between the 3-tert-
butyl group of the DBCH ligand and one of the R-methyl groups
of the 6-Me2-bpmcn ligand, so electronic factors appear more
important than steric factors in determining which catechol
oxygen becomes deprotonated. Furthermore the O1 oxygen is
only 2.65 Å away from an oxygen atom of the counterion
CF3SO3-, suggesting a hydrogen bonding interaction which
would stabilize the monoanionic form of the catecholate. The
steric interactions among the R-methyl groups of 6-Me2-bpmcn
and the 3-tert-butyl group of DBCH ligand give rise to bond
angles much larger than those that are associated with an ideal
octahedron (O2-Fe1-N4 107.42(5)°, N3-Fe1-N4 111.89(5)°).
Table 2. Selected Bond Lengths (Å) for 1b, 1d, 2a, and 2d
1ba
1d
2a
2d
Fe(1)-O(1)
Fe(1)-O(2)
Fe(1)-N(1)
2.263(8) 2.3443(13) 1.896(2) 1.926(2)
1.953(8) 1.9434(11) 1.972(2) 1.928(2)
2.21(1)
2.2505(14) 2.151(3) 2.193(3)
Fe(1)-N(2) [N(11)]# 2.30(1)#
2.2405(14) 2.230(3)# 2.226(2)
Fe(1)-N(3) [N(21)]# 2.172(9)# 2.2415(14) 2.248(2)# 2.217(2)
Fe(1)-N(4) [N(31)]# 2.31(1)#
2.215(2)
1.396(2)
1.339(2)
1.401(2)
2.202(3)# 2.269(3)
1.343(3) 1.345(3)
1.342(3) 1.348(3)
1.391(4) 1.402(4)
C(1)-O(1)
C(2)-O(2)
C(1)-C(2)
1.40(1)
1.35(1)
1.40(2)
The Fe-N bonds of 6-Me2-bpmcn have lengths of 2.22-
2.25 Å, typical of R-methylpyridines coordinated to high-spin
Fe(II) ions.16 However, the coordination configuration of the
6-Me2-bpmcn ligand differs from that of related ligands in
mononuclear17 and dinuclear iron complexes18 such as [(L)-
FeII(CH3CN)2]2+, [(L)FeIII(µ-O)(µ-OH)FeIII(L)]3+, and [(L)-
FeIII(µ-O)(µ-O2CR)FeIII(L)]3+ (L ) bpeen, bpmcn, bpmen; R
) -H, -CH3). In all these complexes, the tetradentate ligand
coordinates to the metal center in a cis-R configuration, i.e.,
the two pyridyl groups are trans to each other as shown in
Scheme 3. However, the 6-Me2-bpmcn ligand in 1d coordinates
to the iron(II) center in a cis-â configuration, i.e., the two pyridyl
groups are cis to each other. This distinct configuration may
arise from the introduction of the R-methyl groups of 6-Me2-
bpmcn.
a Ref 7.
[(6-Me3-TPA)FeIII(Cat)](BPh4) (2a) and [(6-Me2-bpmcn)-
FeIII(DBC)](CF3SO3) (2d). The crystal structures of 2a and 2d
are shown in Figure 2 and selected bond lengths are given in
Table 2. The structure of 2a is comparable to those of previously
Figure 1. ORTEP plot of [(6-Me2-bpmcn)Fe(DBCH)](CF3SO3) (1d)
showing 50% probability thermal ellipsoids and the labeling scheme
for selected atoms. The dashed line shows the hydrogen bonding
interaction (2.65 Å) between HO1 on the DBCH ligand and O3 on
CF3SO3 anion. All other hydrogen atoms are omitted for clarity.
(16) (a) Zang, Y.; Kim, J.; Dong, Y.; Wilkinson, E. C.; Appelman, E. H.;
Que, L., Jr. J. Am. Chem. Soc. 1997, 119, 4197-4205. (b) Constable,
E. C.; Baum, G.; Bill, E.; Dyson, R.; van Eldik, R.; Fenske, D.; Kaderli,
S.; Morris, D.; Neubrand, A.; Neuburger, M.; Smith, D. R.; Wieghardt,
K.; Zehnder, M.; Zuberbu¨hler, A. D. Chem. Eur. J. 1999, 5, 498-
508.
Structural Characterization. [(6-Me2-bpmcn)FeII(DBCH)]-
(CF3SO3) (1d). The crystal structure of 1d is shown in Figure
1 with selected bond lengths listed in Table 2. This structure is
comparable with that previously reported for [(6-Me3-TPA)-
FeII(DBCH)](ClO4) (1b).7 The iron center has a distorted
octahedral environment with a tetradentate ligand and a bidentate
catecholate ligand. The catecholate ligand binds to the iron
asymmetrically with two distinct Fe-O bond lengths of 1.943(1)
and 2.344(1) Å. The shorter Fe1-O2 bond is trans to the weaker
(17) (a) Chen, K.; Que, L., Jr. Chem. Commun. 1999, 1375-1376. (b) Jo,
D.-H. Unpublished observations.
(18) (a) Arulsamy, N.; Hodgson, D. J.; Glerup, J. Inorg. Chim. Acta 1993,
209, 61-69. (b) Glerup, J.; Goodson, P. A.; Hazell, A.; Hazell, R.;
Hodgson, D. J.; McKenzie, C. J.; Michelsen, K.; Rychlewska, U.;
Toftlund, H. Inorg. Chem. 1994, 33, 4105-4111. (c) Fenton, R. R.;
Stephens, F. S.; Vagg, R. S.; Williams, P. A. Inorg. Chim. Acta 1995,
236, 109-115. (d) Glerup, J.; Michensen, K.; Arulsamy, N.; Hodgson,
D. J. Inorg. Chim. Acta 1998, 274, 155-166. (e) Zheng, H.; Zang,
Y.; Dong, Y.; Young, V. G., Jr.; Que, L., Jr. J. Am. Chem. Soc. 1999,
121, 2226-2235.

3186 Inorganic Chemistry, Vol. 40, No. 13, 2001
Jo et al.
Figure 2. ORTEP plots of [(6-Me3-TPA)Fe(Cat)]+ (2a) and [(6-Me2-bpmcn)Fe(DBC)]+ (2d) showing 30% and 50% probability thermal ellipsoids,
respectively, and the labeling scheme for selected atoms. Hydrogen atoms are omitted for clarity.
reported Fe(III)-catecholate complexes with tetradentate tri-
podal ligands.10,11,19 The catecholate oxygens, N1 and N31,
define an approximate plane of symmetry. As previously
observed for Fe(6-Me3-TPA) complexes,16a,18e,20 the in-plane
Fe1-N1 and Fe1-N31 bonds are shorter than the two out-of-
plane Fe-N bonds, due to steric interactions among the
R-methyl groups on the pyridine rings. For the same reason,
the average Fe-Npy bond length of 2a (2.23 Å) is 0.1 Å longer
than that of [(TPA)FeIII(DBC)](BPh4) (2.13 Å). The catecholate
ligand chelates to the iron asymmetrically with Fe-O bond
lengths of 1.896(2) and 1.972(2) Å. This asymmetry (∆rFe-O
) 0.076 Å) in 2a is not as pronounced as in [(NTA)FeIII(DBC)]
(0.092 Å)19a or [(BPG)FeIII(DBC)] (0.1 Å)10 but is larger than
in [(TPA)FeIII(DBC)]+ (0.019 Å).11 Also, due to the steric
congestion introduced by 6-Me3-TPA, the bond angles related
to O1 are larger than others (O1-Fe1-N11 100.27(10)°, O1-
Figure 3. ORTEP plot of [(6-Me3-TPA)Fe(NO)2]+ (4) showing 50%
probability thermal ellipsoids and the labeling scheme for selected
atoms. Hydrogen atoms are omitted for clarity.
Fe1-N21 106.92(10)°, O1-Fe1-N31 107.46(10)°).
Complex 2d also has a distorted octahedral geometry with
average Fe-N and Fe-O bond lengths of 2.226(3) and 1.927(2)
Å, respectively, typical of high-spin Fe(III) complexes.16 Unlike
in 2a, the Fe-Ocat bond lengths (∆rFe-O ) 0.002 Å) are nearly
identical. The larger bond angles, O2-Fe1-N3 110.01(9)° and
N3-Fe1-N4 97.78(9)°, result from steric congestion among
R-methyl groups of 6-Me2-bpmcn and the 3-tert-butyl group
of the DBCH ligand, as in 1d. Surprisingly, although the 6-Me2-
bpmcn ligands in both 1d and 2d have the same cis-â topology,
their conformations differ. While the two N-methyl groups in
1d are anti to each other, those on 2d are syn. The epimerization
of the 6-Me2-bpmcn ligand may result from steric interactions
that develop upon oxidation of the metal center.
Table 3. Selected Bond Lengths (Å) and Bond Angles (deg) for 4
Fe(1)-N(1w)
Fe(1)-N(1)
Fe(1)-N(21)
N(2w)-O(2w)
1.699(3)
2.160(3)
2.207(3)
1.165(4)
Fe(1)-N(2w)
Fe(1)-N(11)
N(1w)-O(1w)
1.690(3)
2.251(3)
1.168(4)
N(1w)-Fe(1)-N(2w) 110.4(2) N(1w)-Fe(1)-N(1) 125.8(2)
N(1w)-Fe(1)-N(11) 99.7(2) N(1w)-Fe(1)-N(21) 97.62(14)
N(2w)-Fe(1)-N(1)
N(2w)-Fe(1)-N(21)
N(1)-Fe(1)-N(21)
123.8(2) N(2w)-Fe(1)-N(11) 97.3(2)
96.4(2) N(1)-Fe(1)-N(11)
75.60(12)
76.97(11) N(11)-Fe(1)-N(21) 152.52(12)
Fe(1)-N(2w)-O(2w) 162.4(4) Fe(1)-N(1w)-(O1w) 159.7(4)
[(6-Me3-TPA)Fe(NO)2](ClO4) (4). The crystal structure of
4 is shown in Figure 3 and selected bond lengths and angles
are listed in Table 3. Complex 4 has a trigonal bipyramidal iron
center with two equatorial nitrosyl groups. The 6-Me3-TPA
ligand binds to the iron as a meridional tridentate ligand with
one of the pyridine arms remaining unbound and just dangling
away from the iron. As observed in other structures of Fe(6-
Me3-TPA) complexes,16a,20-21 the axial Fe-Npy bonds are
somewhat elongated (2.251(3) and 2.207(3) Å) relative to the
equatorial Fe-Nam bond (2.160(3) Å) to minimize steric
interactions between the R-methyl substituents on the pyridines.
The Fe(NO)2 unit in the complex is nearly planar and the
two NO ligands are almost equivalent crystallographically. The
Fe(NO)2 unit of 4 is characterized by shorter Fe-N and N-O
bond lengths (average 1.695(3) and 1.167(4) Å, respectively)
(19) (a) Que, L., Jr.; Kolanczyk, R. C.; White, L. S. J. Am. Chem. Soc.
1987, 109, 5373-5380. (b) Cox, D. D.; Benkovic, S. J.; Bloom, L.
M.; Bradley, F. C.; Nelson, M. J.; Que, L., Jr.; Wallick, D. E. J. Am.
Chem. Soc. 1988, 110, 2026-2032. (c) Koch, W. O.; Kru¨ger, H.-J.
Angew. Chem., Int. Ed. Engl. 1995, 34, 2671-2674. (d) Mialane, P.;
Anxolabe´he`re-Mallart, E.; Blondin, G.; Nivorojkine, A.; Guilhem, J.;
Tchertanova, L.; Cesario, M.; Ravi, N.; Bominaar, E.; Girerd, J.-J.;
Mu¨nck, E. Inorg. Chim. Acta 1997, 263, 367-378. (e) Duda, M.;
Pascaly, M.; Krebs, B. Chem. Commun. 1997, 835-836. (f) Mialane,
P.; Tchertanov, L.; Banse, F.; Sainton, J.; Girerd, J.-J. Inorg. Chem.
2000, 39, 2440-2444.
(20) (a) Chiou, Y.-M.; Que, L., Jr. J. Am. Chem. Soc. 1992, 114, 7567-
7568. (b) Zang, Y.; Elgren, T. E.; Dong, Y.; Que, L., Jr. J. Am. Chem.
Soc. 1993, 115, 811-813.
(21) (a) Zang, Y.; Jang, H. G.; Chiou, Y.-M.; Hendrich, M. P.; Que, L., Jr.
Inorg. Chim. Acta 1993, 213, 41-48. (b) Chiou, Y.-M.; Que, L., Jr.
J. Am. Chem. Soc. 1995, 117, 3999-4013.

Extradiol Cleaving Catechol Dioxygenases
Inorganic Chemistry, Vol. 40, No. 13, 2001 3187
Table 4. Comparison of Properties of Fe(II)- and
Fe(III)-Catecholate Complexes
complex
λ
max (ꢀ)a
EPR (g)
E/D
kobs (s-1
)
1a
386 (2000)
395 (2200)
380 (900)
384 (1100)
12b
-
1b
1c
1d
2a
12b
1.1 × 10-2 d
1.7 × 10-2 d
2.2 × 10-2 d
EPR-silent
EPR-silent
540 (2100), 940 (3200) 7.3, 4.5, 1.9b 0.06
6.0
0
2b
600 (2200), 1020 (3400) 7.5, 4.5, 1.9b 0.065 7.8 × 10-5 e
6.0
0
2c (in situ) 535 (2100), 950 (3100) 8.5, 5.6, 3.1b 0.13
6.3, 5.6, 2.0 0.02
2c (isolated) 535 (2300), 874 (3000) 4.3b
0.33
8.4, 5.5, 3.0 0.13
6.3, 5.5, 2.0 0.02
2d (in situ) 590 (2000), 1030 (3500) 8.4, 3.2c
6.8, 5.2, 2.0 0.03
2d (isolated) 598 (2200), 950 (3100) 9.3, 4.8, 4.1c 0.25 <10-6 e
8.4, 5.5, 3.0 0.13
0.13 2.7 × 10-5 e
Figure 4. Absorption spectral changes for the reaction of 1d with O2
in CH3CN at 22 °C. (a) Unreacted 1d, (b) in situ formed 2d, and (c)
isolated 2d.
a Obtained in CH3CN at 22 °C. b Obtained in CH3CN at 5 K.
c Obtained in CH2Cl2/toluene (2:1) at 5 K. d Rate constants for the
formation of Fe(III)-catecholate complexes under 1 atm O2 at 22 °C.
e Rate constants for the decomposition of Fe(III)-catecholate complexes
under 1 atm O2 at 22 °C.
than those of iron-mononitrosyl complexes.22-24 The short Fe-
NO distances indicate substantial covalent character between
the iron and the nitrosyl ligands. The N1w-Fe1-N2w angle
of 110.4(2)° is larger than the O1w-Fe1-O2w angle of
95.0(2)°, indicating that the iron-dinitrosyl group adopts an
“attracto” conformation. The “attracto” conformation refers to
the fact that the two NO ligands bend toward each other, usually
observed for the first-low (3d) transition metal complexes
containing good π-acceptor ligands.22a Complex 4 represents a
species with an {Fe(NO)2}9 electronic configuration, thereby
increasing the small number of structurally characterized {Fe-
(NO)2}9 complexes with nitrogen- and/or oxygen-donor
ligands.25
Reactions of Fe(II)-Catecholate Complexes with O2. The
light yellow solutions of the Fe(II)-catecholate complexes (1)
react immediately with O2 to afford purple-blue intermediate
species (2), the color of which arises from intense catecholate-
to-Fe(III) LMCT bands (Figure 4, Table 4), as previously found
for other [(L)FeIII(catecholate)] (L ) tetradentate ligands)
complexes.11,19 EPR spectra of 1a and 1b in CH3CN show a
strong and broad signal at g ) 12 (Figure 5a), which is
tentatively assigned to an integer-spin resonance arising from
the S ) 2 high-spin iron(II) center.26 These features are replaced,
upon exposure of 1a and 1b to O2, by signals arising from high-
spin iron(III) centers. Complexes 2a and 2b exhibit nearly axial
EPR spectra, consisting of a major component with E/D )
0.06-0.065 and a minor component with E/D near 0 (Figure
5c, Table 4). Complex 2b further reacts with O2 to afford
oxidative cleavage products (see below).
Figure 5. X-band EPR spectra in CH3CN at 5 K of (a) 1a, (b) iron-
nitrosyl intermediate 3a (E/D ) 0.006), (c) 2a (E/D ) 0.06), and (d)
4. Instrumental conditions: microwave power, 0.2 mW; microwave
frequency, 9.63 GHz; modulation frequency, 100 kHz; modulation
amplitude, 1 mT; gain, 60.
2c generated from the reaction of 1c and O2 are red-shifted from
those of 2c isolated from the same solution that was left standing
in the refrigerator for several months (Figure S1b and c in
Supporting Information). Similarly, the LMCT bands of 2d
generated in situ are also red-shifted relative to those of isolated
2d (Figure 4 b and c), obtained upon long-term standing of in
situ 2d solutions generated either from the reaction of 1d with
a limited amount of O2 or with NO (see below). Thus there
appear to be two isomers of 2c and 2d, one that is generated
directly in the reaction of 1c or 1d and O2 and the other obtained
upon long-term crystallization.
EPR studies support the notion of two forms of 2c and 2d
(Table 4). Exposure of EPR-silent 1d to O2 immediately affords
2d with EPR features associated with high-spin iron(III) S )
5/2 species having E/D values of 0.03 and 0.13 (Figure 6c). In
contrast, isolated 2d has EPR features from an E/D ) 0.13
species and a more rhombic E/D ) 0.25 species (Figure 6d).
Even crystals of 2d exhibit signals from both E/D ) 0.13 and
The spectral changes associated with the reaction of 1c and
1d with O2 are more complex. The visible spectral changes for
1c and 1d are displayed in Figures S1 (Supporting Information)
and 4, respectively. Interestingly, the LMCT bands of in situ
(22) (a) Enemark, J. H.; Feltham, R. D. Coord. Chem. ReV. 1974, 13, 339-
406. (b) McCleverty, J. A. Chem. ReV. 1979, 79, 53-76. (c) Richter-
Addo, G. B.; Legzdins, P. Metal Nitrosyls; Oxford University Press:
New York, 1992.
(23) (a) Chiou, Y.-M.; Que, L., Jr. Inorg. Chem. 1995, 34, 3270-3278.
(b) Ray, M.; Golombek, A. P.; Hendrich, M. P.; Yap, G. P. A.; Liable-
Sands, L. M.; Rheingold, A. L.; Borovik, A. S. Inorg. Chem. 1999,
38, 3110-3115.
(24) Pohl, K.; Wieghardt, K.; Nuber, B.; Weiss, J. J. Chem. Soc., Dalton
Trans. 1987, 187-192.
(25) (a) Chong, K. S.; Rettig, S. J.; Storr, A.; Trotter, J. Can. J. Chem.
1979, 57, 3119-3125. (b) Chong, K. S.; Rettig, S. J.; Storr, A.; Trotter,
J. Can. J. Chem. 1979, 57, 3113-3118. (c) Kisko, J. L.; Hascall, T.;
Parkin, G. J. Am. Chem. Soc. 1998, 120, 10561-10562.
(26) Hendrich, M. P.; Debrunner, P. Biophys. J. 1989, 56, 489-506.

3188 Inorganic Chemistry, Vol. 40, No. 13, 2001
Jo et al.
Scheme 4
reactions consist of an initial fast formation of the blue
chromophore, followed by a much slower first-order decay of
the color (Figure 4 and Table 4). While the rates of formation
are quite comparable for the three complexes studied, the rates
of decomposition differ significantly. Not surprisingly, 2a and
2c are much more stable than 2b and 2d due to the absence of
electron-donating tert-butyl groups. Complex 2b and in situ
generated 2d react with O2 with rates of 3-8 × 10-5 s-1, which
are 3 orders of magnitude slower than that for [(TPA)FeIII-
(DBC)]+ (4.8 × 10-2 s-1 in DMF).11 Although the metal centers
in 2b and 2d have greater Lewis acidity, as indicated by their
lower energy LMCT bands, we speculate that the steric
hindrance introduced by 6-methyl substituents on the pyridines
increases the barrier to the reaction. As noted earlier, isolated
2d does not react with O2 after 4 days, so the configuration of
the tetradentate ligand also exerts a significant effect on the
reactivity of the catecholate complex toward O2. Given that the
decay of in situ generated 2d in the presence of O2 is first order
in 2d, it would appear that the isomerization of in situ generated
2d to isolated 2d must be significantly slower than the reaction
of in situ generated 2d with O2 to afford oxidative cleavage
products.
Figure 6. X-band EPR spectral changes observed in the reactions of
1d with NO and O2 at 5 K. (a) 1d in CH3CN, (b) iron-nitrosyl
intermediate 3d in CH3CN (E/D ) 0.01), (c) in situ formed 2d (E/D )
0.03, 0.13) from the reaction of 1d and O2 in CH2Cl2/toluene (2:1), (d)
isolated 2d (E/D ) 0.13, 0.25) in CH2Cl2/toluene (2:1) from the reaction
of 1d with NO. Instrumental conditions: microwave power, 0.2 mW;
microwave frequency, 9.63 GHz; modulation frequency, 100 kHz;
modulation amplitude, 1 mT; gain, 60.
0.26 species, as does 2d obtained by direct synthesis. There is
also a corresponding difference in the EPR properties of in situ
generated 2c and isolated 2c (Figure S2 in Supporting Material).
It is not unusual for high-spin iron(III) complexes to show E/D
heterogeneity, which may arise from the sensitivity of the S )
5/2 zero field splitting parameters to different conformations of
the cyclohexane ring.27 Despite the complexity of the EPR
spectra, it is clear that in situ generated 2c and 2d are distinct
from their isolated counterparts.
Most importantly, the reactivities of two forms of 2d toward
O2 differ significantly. While the in situ generated complex
reacts further with O2 to afford about 90% yield of oxidative
cleavage products (see below), isolated 2d does not react with
O2 at all over a period of 4 days. Thus there must be a significant
difference between the in situ generated and isolated forms of
2d.
We propose the difference to arise from the two distinct ligand
configurations found in the crystal structures of 1d and 2d, cis-
â, anti for 1d and cis-â, syn for 2d. This structural difference
reflects the epimerization of one N-CH3 group in the conversion
of 1d to 2d. However, this change entails breaking both an Fe-
Nam and an Fe-Npy bond followed by rebinding with the
inversion of the amine nitrogen and would be expected to have
a large activation barrier. The in situ generated 2c and 2d are
thus the kinetically controlled products, which retain the cis-â,
anti configuration of the precursor complexes; they then must
isomerize upon standing to form the thermodynamically more
favored products having the cis-â, syn configuration observed
in the crystal structure of 2d.
As reported previously,7 an O2 uptake study for 1b showed
that 1.3(1) molecules of O2 were consumed per molecule of
1b. This result is consistent with 1 molecule of O2 oxidizing 4
molecules of 1b in the initial fast step (i.e. 0.25 O2 per 1b with
O2 being reduced to H2O), and another molecule of O2 cleaving
the bound DBC on 2b in the slower step. The reaction of 2b
with O2 gives rise to mainly intradiol cleavage of the bound
DBC in 89% yield. The major products observed are 3,5-di-
tert-butyl-1-oxacyclohepta-3,5-diene-2,7-dione (A, 53%) and
3,5-di-tert-butyl-5-(carboxymethyl)-2-furanone (B, 36%), shown
in Scheme 4. However, extradiol cleavage is also observed,
accounting for 3% of the products. Both 4,6-di-tert-butyl-2-
pyrone (C) and 3,5-di-tert-butyl-2-pyrone (D) are found.
Similarly, the reaction of 2d with O2 results in 78(62:16)%
intradiol cleavage and 12(4:8)% extradiol cleavage. Previous
work has demonstrated that [(L)FeIII(DBC)] (L ) tetradentate
tripodal ligands) complexes afford exclusively intradiol cleavage
products.10,11,19 To date, extradiol cleavage has been observed
only for complexes with facial tridentate ligands such as R3-
Kinetic studies of the reaction of complexes 1 with O2 were
undertaken by monitoring the intensity changes of the LMCT
bands of 2 under pseudo-first-order conditions in CH3CN at 22
°C ([O2] ) 8.1 mM in CH3CN under 1 atm pressure).28 The
TACN and Tpi-Pr2 6,29
Thus, while it is expected that 2b and
.
2d yield predominantly intradiol cleavage products, it is
surprising that there is a small amount of extradiol cleavage as
well. Such cleavage may derive from complexes wherein one
of the pyridyl arms has dissociated from the metal center. The
steric congestion around the metal center engendered by the
(27) Simaan, J.; Poussereau, S.; Blondin, G.; Girerd, J.-J.; Defaye, D.;
Philouze, C.; Guilhem, J.; Tchertanov, L. Inorg. Chim. Acta 2000,
299, 221-230.
(28) Sawyer, D. T. Oxygen Chemistry; Oxford University Press: New York,
1991; p 21.
(29) Jo, D.-H.; Que, L., Jr. Angew. Chem., Int. Ed. 2000, 39, 4284-4287.

Extradiol Cleaving Catechol Dioxygenases
Inorganic Chemistry, Vol. 40, No. 13, 2001 3189
Table 5. Comparison of Physicochemical Properties of Iron-Nitrosyl Adducts with {Fe(NO)1,2}x (x ) 7, 9) Configurations
complexa
{Fe(NO)2}9
Fe-NO (Å) FeN-O (Å)
(Fe-N-O) (deg)
[Fe-(NO)2] (deg)
[Fe-(NO)2] (deg) ν(NO) (cm-1
) reference
4
1.690(3)
1.699(3)
1.688(3)
1.694(3)
1.694(2)
1.168(4)
1.165(4)
1.164(3)
1.172(3)
1.159(2)
159.7(4)
162.4(4)
166.8(3)
161.7(3)
164.3(2)
110.4(2)
112.57(13)
110.6(1)
108.1(2)
95.0
99.8
97.9
91.0
1801, 1726 this work
1811, 1726 25c
[(PhTpt-Bu)Fe(NO)2]
[(Pz3,5-me2)Fe(NO)2]2
[(GaPzO)Fe(NO)2]
{Fe(NO)}7
b
1802, 1780 25b
1750, 1710
1724, 1671 25a
1.706(4)
1.700(4)
1.158(5)
1.148(5)
157.9(4)
159.3(5)
[(TPA)Fe(BF)(NO)]+
1.72(2)
1.78
1.738(5)
1.15(2)
1.10
1.142(7)
159(2)
156(5)
155.5(10)
n.a.
n.a.
n.a.
n.a.
n.a.
n.a.
1794
1776
1690
23a
30
24
[(EDTA)Fe(NO)]c
[(Me3TACN)Fe(N3)2(NO)]
a PhTpt-Bu, phenyl tris(3-tert-butylpyrazolyl)borate; Pz3,5-me2, 3,5-dimethylpyrazolyl anion; GaPzO, dimethyl(N,N-dimethyl ethanolamino)(3,5-
dimethyl-pyrazolyl)gallate[N(2),N(3),O]; BF, benzoylformate; EDTA, ethylenediaminetetraacetic acid; Me3TACN, 1,4,7-trimethyl-1,4,7-triazacy-
clononane. b Average values. c Determined by using GNXAS analysis.
Scheme 5
1d with NO initially affords a 2d product with cis-â, anti ligand
configuration, as indicated by its UV-vis spectrum (λmax ) 590
and 1030 nm); this species then epimerizes upon long term
standing to the thermodynamically more favored cis-â, syn
configuration (λmax ) 598 and 950 nm).
Complex 4 exhibits a UV-vis spectrum with absorption
shoulders at 332 (1700) and 374 (1300) nm and weak, broad
bands at 460 (360) and 820 (200) nm. It has an IR spectrum
with two intense nitrosyl stretching vibrations at 1801 and 1726
cm-1, consistent with the presence of an Fe(NO)2 moiety.31 Its
X-band EPR spectrum in a frozen CH3CN solution (Figure 5d)
shows only one intense isotropic signal at g ) 2.02, which
clearly indicates the presence of an S ) 1/2 species. The complex
can thus be assigned as an {Fe(NO)2}9 system and is related to
other iron-dinitrosyl complexes, which exhibit an isotropic g
) 2.03 signal.32 The formation of 4 is a result of the
disproportionation of 3a and 3b, resulting in the transfer of NO
and one electron from one iron center to another. The expected
free catechol byproduct was found in the mother liquor.
introduction of the 6-methyl substituent and resulting longer
Fe-Npy bonds may promote the dissociation. This would
generate a site for O2 attack at the metal center and result in
extradiol cleavage, following a mechanism previously proposed
for complexes of tridentate ligands.6b,29
Summary and Perspective
As part of our efforts to model extradiol cleaving catechol
dioxygenases, we have prepared a series of Fe(II)-catecholate
complexes (1) as structural mimics for the enzyme-substrate
complexes. The bound catecholate is monoanionic but acts as
a bidentate ligand. The fact that one oxygen is negatively
charged while the other is neutral results in a coordination mode
where the Fe-Ocat bonds differ in length by as much as 0.4 Å.
A similar coordination mode has thus been proposed for the
catecholate substrate bound to catechol 2,3-dioxygenase, on the
basis of the appearance of a short (1.93 Å) Fe-O bond upon
substrate binding in the EXAFS analysis of this complex.4 This
notion is supported by more recent crystallographic data on the
enzymes.3 However the iron centers of the model complexes
are six-coordinate, while those of the enzyme-substrate com-
plexes are five-coordinate and thus have one available site for
O2 binding. Nevertheless the model complexes can react with
Reactions of Fe(II)-Catecholate Complexes with NO.
Scheme 5 summarizes the reactions of complexes 1 with NO.
When NO is bubbled through a solution of 1 (CH3OH for 1a
and CH2Cl2 for 1b, 1d) at 22 °C under anaerobic conditions,
the initial light yellow color darkens to brown and then changes
to purple-blue. These brown intermediate solutions (3) exhibit
UV-vis absorption features near 400, 500, and 700 nm, typical
of synthetic S ) 3/2 Fe-NO complexes (Table 6).23,30 Interme-
diates 3 also show typical axial S ) 3/2 EPR signals, with g )
4.0, 3.9, 2.0 for 3a (E/D ) 0.006) and 4.1, 4.0, 2.0 for 3d (E/D
) 0.01) (Figures 5b and 6b), accounting for >95% of the total
spin in the samples. Thus the nitrosyl adducts of 1 can be
formulated as [(L)Fe(catH)(NO)]+ (3) with an {Fe(NO)}7
electronic configuration. Unfortunately, none of the complexes
3 could be isolated due to their instability. Complexes 3a and
3b convert to two distinct products in comparable yields: deep-
blue crystals of [(6-Me3-TPA)FeIII(cat)]+ (2a and 2b) and
brown-red crystals of [(6-Me3-TPA)Fe(NO)2]+ (4), while 3d
yields only the corresponding Fe(III)-catecholate complex (2d)
in 90% yield (and presumably an equivalent of HNO). Similar
to that observed for the reaction of 1d with O2, the reaction of
(31) (a) Beck, W.; Melnikoff, A.; Stahl, R. Chem. Ber. 1966, 99, 3721-
3727. (b) Poletti, A.; Foffani, A.; Cataliotti, R. Spectrochim. Acta 1970,
26A, 1063-1069.
(32) (a) Bulter, A. R.; Glidewell, C.; Li, M.-H. AdV. Inorg. Chem. 1988,
32, 335-393. (b) Vanin, A. F. FEBS Lett. 1991, 289, 1-3. (c)
Kennedy, M. C.; Gan, T.; Antholine, W. E.; Petering, D. H. Biochem.
Biophys. Res. Commun. 1993, 196, 632-635. (d) Butler, A. R.; Flitney,
F. W.; Williams, D. L. H. Trends Pharmacol. Sci. 1995, 16, 18-22.
(e) Noguchi, T.; Honda, J.; Nagamune, T.; Sasabe, H.; Inoue, Y.; Endo,
I. FEBS Lett. 1995, 358, 9-12. (f) Muller, B.; Kleschyov, A. L.;
Stoclet, J. C. Br. J. Pharm. 1996, 119, 1281-1285.
(30) Brown, C. A.; Pavlosky, M. A.; Westre, T. E.; Zhang, Y.; Hedman,
B.; Hodgson, K. O.; Solomon, E. I. J. Am. Chem. Soc. 1995, 117,
715-732.

3190 Inorganic Chemistry, Vol. 40, No. 13, 2001
Jo et al.
Table 6. UV-vis Spectral Features of Iron-Nitrosyl Adducts with {Fe(NO)}7 Configurationa
complex
λ
max (ꢀ)
reference
this work
3a
3d
360 (2600), 500 (720), 718 (440)
372 (2300)
388 (1700)
360 (2400), 550 (250), 750 (150)
372 (2500)
390 (2450)
358 (1900), 492 (720), 690 (240)
372 (1600)
this work
23a
[(6-Me3-TPA)Fe(BF)(NO)]+
388 (1200)
350 (1100), 520 (250), 654 (160)
420 (730)
[(TPA)Fe(BF)(NO)]+
23a
30
[(EDTA)Fe(NO)]
340 (1300), 430 (900), 640 (200)
a Obtained in CH3CN at 22 °C.
3
O2 and NO. With NO, S ) /2 {Fe(NO)}7 adducts are formed,
complexes to date are thus imperfect. Future efforts will be
directed at obtaining five-coordinate iron(II)-catecholate
complexes that react with O2 and afford extradiol cleavage
directly.
as for the enzymes,5 although the biomimetic adducts are
susceptible to decomposition. The weaker Fe-OHcat bond is
presumably easily broken and NO replaces it. On the other hand,
exposure of complexes 1 to O2 results in their immediate one-
electron oxidation to the corresponding iron(III)-catecholate
dianion complex, a reactivity pattern distinct from that of the
enzymes.
These iron(III)-catecholate complexes further react with O2
to afford oxidative cleavage products. Like previously reported
iron(III)-catecholate complexes of tetradentate ligands, the
major products derive from intradiol cleavage.10,11,19 However,
unlike the latter complexes, some extradiol cleavage is observed.
Biomimetic extradiol cleavage has thus far been observed only
for five-coordinate iron(III)-catecholate complexes.6,29 So we
speculate that, due to steric congestion around the iron center,
there is a subpopulation of complexes with one pyridyl arm
dissociated that gives rise to the minor products. Our model
Acknowledgment. This work was supported by the National
Institutes of Health (Grant GM-33162). D.-H.J. is grateful to
the Korean Science and Engineering Foundation (KOSEF) for
a postdoctoral fellowship. We thank Prof. Doyle Britton, Dr.
Victor G. Young, Jr., and Dr. Maren Pink of the University of
Minnesota X-ray Crystallographic Laboratory for determining
the crystal structures of 1d, 2a, 2d, and 4.
Supporting Information Available: Crystallographic data for 1d,
2a, 2d, and 4 (CIF format) and Figures S1 and S2 showing UV-vis
and EPR spectral changes for 2c (PDF format). These materials are
available free of charge via the Internet at http://pubs.acs.org.
IC001185D