- Email: [email protected]
The resonance Raman spectra of intermediate-spin ferrous porphyrin
Volume 63, number 3
CHEMlCAL
PHYSKS
LL?lTERS
1 June 1979
Teizo KITAGAWA and Junji TERAOKA Institute for Protein Research, Osurl-aUtir ersity, St&a. Osaka 565. Japmt Received 16 October
1978:in
fwl
form 3 April 1979
The resonance Raman and absorption spectra of unligatcd ferrous ocractb~ lporplwrin t&*fOEP)] and Its adduct tlith I”Velm or TtIF acre observed. The resommcc Ramaa spectrum of Te”(OEP) in CIi~Cl2 dispbq cd cbamctcristic features of iron porph>rio &ith three d, electron+. susgesting 3Ee 3~ the intermediate-spin (S = 1) sound st.tte_fe’*t.OIZP) in TIfF ‘i e\hibited the ferrous high spin feature like Fe2*(OEP) (_-McIm) in Cff=Ctz-
1. Introduction
The existence of intcr~nedi~te-span state (S = I) as a ground state of ferrous porphyrin had been predicted for a square pku~ar structure with a moderate @and field [I 1, and Collman et ai. [3] first isolated such ;Lcompound; tctr~phcnylporpi~~~~n~to iron
the aqueous solution of dithionite-reduced porphyrin in the presence of air [I I1 ; the absorpiion spectrum resembled titose of so-wiled “ohygensted“ ferrous porphyrin [ 121 snd oxygenated hemoglobin [ t 31 and the Rsman spectrum resembIed that of osyg.enated hemoglobin and chrtnged to ferrous high-spin type upon addition of 2-methyi imidnzotc Q-MeIm)_ To establish the absorption and Raman spectra of intermediate-spin ferrous porphyrin with more qualified preparation, i.e. in the absence of dithionite and contaminated water. we constructed a device to measure the absorption and Raman spectra of a hi$i!y evacuated system and successfutiy used it \\ith .I reactive intermediate-spin ferrota porphyrin_ Here we report the absorption and resonnnce Ramsn spectra of Fez” (OEP) with no asiaI iigmd and its adducr with 2-M&n or tetmhydrofuran (THF). proposing 3 Es ;LS a fikefy ground state.
3. Experimental Oct3eth~~porp~y~n~to iron (It) bis-pyrrdine complex [Fe”’ (OEP) @y&f (I). obtained by hydra&ehydrate reduction of F>3+ @XZP) Cl in pyridine (Py)_ was kindly provided by Dr_ H. Ogoshi md Mr.. H. Sugimoto of Kyoto University. The crystal of 1 wxi kept at 140°C for 2 h in 2 X 10M5 mm Hg in a Raman or visible absorption ceI1, yielding Fez’ (OEP) (2) [4]. 433
VoIume 63. number 3
1 June 1979
CHEiiICAL PHYSICS LJXJ-ERS
Since 2 is easily oxidized, the degassed solvent (THF, Py, or CH2CI2) was transferred to the ceII in the evacuated system_ To obtain Fez+ (CEP) (ZMeIm) (3), fresbiy recrystallized ZMeIm was put into the branched part of the cell, where it was kept at reduced pressure and room temperature during the heating of the cell. After 2 was formed in the ceII, 2-Meim was mixed with the solution of 2 without admitting air_ The Raman spectra were observed with the 488.0 nm excitation line of an argon ion laser (Spectra Physics. model 164) and a JEOL100D Raman spectrometer- The absorption spectra were measured with a Hitachi 124 spectrophotometer.
b.Py
3_ ResuIts and discussion The visible absorption spectra of 1,2, and 3 are shown in fig_ I_ I gave a typical spectrum of ferrous low-spin hemes (a). 2 in CH,CI, (b) and THF (c) gave spectra cfearly different from those of I and 3 (d)_ After observation of spectra b, c, and d, the solvent was rephced with Py in the evacuated system and reproduction of spectrum a was confirmed_ When evacuation was insufficient (<10d3 mm Hg), the reproduction of spectrum a was incomplete. indicating partial o.Gdation. The resonance Raman spectra of 1 in Py (A) and in CH$$ (B), 2 (C) and 3 (D) in CH+Ia are shown in fig. 2. Here again, reproduction of spectrum A was
I
1
Y)
1600
1
i
1500 I400 i;/ cm-1
9
1300
1&c
rig. 2- The resonance Raman spectra of Fe** (OEP) (PY)~ in pyridine (A) and in CH2C12 (B), Fez+ (OEP) in Cti~C.1~(C). and Fez* (OEP) (Z-MeIm) in CHzCtz (D)_ Excitation: 488.0 nm. The Raman lines marked by S me due to soWent or free 24feIm and those desigx&ed by b.Py indimte the resonzmeeenhanced Rzman Linesof bound pyridines.
4OC
. . . ..!...... 500
600
700
X/nm Fis- t_ Absorption spectra af Fez+ (OEP) (Py?+ in pyridine (a). Fe*‘(OEP) in CH+Clz (b). Fe** (OEP) in THF (c) and Fe*‘(OEP) (XfeIm) in CH+& (d)- Soret region is measured for different prepsrarion \iith more dilute concentmtion_
confirmed after each measurement by replacing the solvent with i’y_ fn spectrum B, the resonance-enhanced Raman lines of bound pyridine [IO] (marked by b, Py) were prominent but disappeared in spectrum C, evidencing eIimination of bound pyridine- When 2hleim was added to 2, the resultant spectrum (D) had Raman characteristic features of ferrous high-spin hemoproteins [6,S] _Therefore, 2 is undoubtedly a planar ferrous porphyrin without any axial Iigand, and thus is an intermediate-spin ferrous porphyrin_ We emphasize that spectrum C in fig. 2 is remarkably close to the Raman spectrum of ferric low-spin iron porphy-
Volume 63, number 3
CHEMICAL
PHYSICS LETTERS
I June 1979
rin such as Fe3+ (OEP) (Im), [ 14]_ This similarity was also pointed out by Spiro and Burke for their intermediate-spin ferrous porphyrin [lo] _ It is we11established that the ITtype ferrous Iowspin hemoproteins
having three d, eiectrons give rise
to Rarnan spectra similar to those of ferric low-spin hemoproteins [7,_8,10], while the a type ferrous lowspin hemoproteins [S] having four d, electrons and feryi low-spin hemoprotein (Comp. II or horseradish peroxidase) [ 15,161 having two d, electrons show different Raman spectra. Therefore the Ramrtn results strongly suggests the presence of three d, electrons for the intermediate-spin ferrous state. There are three possible intermediate-spin states; 3B k’ 3A2g,and3E g_ The 3B,, state (d$d,,,d=z) is incompatible not only with the Raman spect&n but aiso with the MijssbauerJ2] and NMR data [5]. For the 3 A2e state (d&, d$ d;), the Raman band IV frequency 171 is expected to be higher than that of the ferric tow-spin motecules. as demonstmted by the feryl Iow-spin compound [ 15.16], due to Iess delocalization of electrons to the porphyrin rr* orbital and furthermore, the d=l orbikd is unlikely to be located lower than the 4, orbital_ Only the ‘E, state (d-z, d~d=~) is compatible with the MBssbauer [2].%hlR [5]-, and Raman data, although mixing of this state with the high-spin state through spin-orbit coupling cannot be ruled out- The empty dxz_.vT orbital is also consistent with the short Fe-N distance [2]. The spin state of the THF adduct of 2 is controversia1 [I ,4,17]. In fig_ 3 the resonance Raman spectrum of 2 in THF is compared with those of 3 (high spin) and 1 (low spin) in THF, although the latter is partly dissociated into 2. It is clear that the Raman spectrum of 2 in THF distinctly differs from that of 2 in Cl&Cl, _ suggesting complex formation of 2 with THF. Theresultant spectrum is close to that of 3 in THF which differs littIe from spectkm D in fig. 2_ Therefore, the THF adduct of Fe’* (OEP) is considered to take the ferrous high-spin state, in good agreement with magnetic susceptibiIity data (5. I pg at 25OC) [ i7]_
_kknowIedgement The authors wish to express their gratitude
to Dr.
H_ Ogoshi and H_ Sugimoto of Kyoto University for their courtesy of providing FeZf (OEP) (Py)? _
1-e
1600
EC0
coo
‘300
.200
‘r.=
cse
0-u-t
J/Cl--I
Fig. 3. The resonmce Rnman specrm of Fe.‘+ (OEP), I-I?’ (OEP) (2DIeIm) and Fe2 (OEP) (P&jz in THT. T?w Ranan lines marhed b) b,Pv ar2 due to bound p: ridine-
References [ 1 j H. Koboqashi 2nd Y. k x~y3~~a. Bul!. Chem. Sot_ Japan IS (1972) 150(71 J-P. Col1m.m. J-L. Ziowi. N. Kim. G. Lang .mc! C A. Reed, J_ Am. Chem_ SOL 97 (1975) 2676_ [3] L.J. Radoncz\lcb,A. Bloom and J-L. Hoard, J. Am. Chem. Sot. 91(1972) 2073. [4j D. Dolphin, J.R. Snms, T-B. Tsin and K-L. N’ong. J. Xm. Chem. Sot. 9s (1976) 6970. [S] II. Gaff, GX LAiar rrnd C.X Reed. J. Am. Chem. SOC. 99 11977) 3641 161 T.G. Spiro and T-C. Strskas. j. Am. Chem. Sot- 96 (1971) 33& [7] T_ Kitsgwa. T_ Iizuka. M. %ito and Y. K?ogoLu. Chcm. Letters (1975) S-%9[S] T. Kitzgaws. I-_ K~ogoku, T_ Iizuka .md 11 I. S&o. J. Am. Chem. Sot. 9s (1976) 5169. [9] Y. Oaki. T. Kitngawa, Y. K>o~oku. C. Ik&imotoYutsudo, Y. Imai and R. &to_ Biochemistry. to be publisbed.
445
V5zme [IO]
63. number
3
CHEMICAL
PHYSICS LETTERS
T-G- Spiro and J-M- Burke, J_ Am_ Chem- Sot. 98 (1976) 5482 [I I] T- Kitagawa, Y. Ozaki. J. Teraoka, Y. Kyogoku and T. Yanumh. Biochim. Biophys. Acta 494 (1977) lOO_ [ 121 J-0. Alben, W-H. Fuchsman, CA_ Bcaudrcau and W_S. Caughey. Biochemistry 7 (1975) 625. [ 13 1 E Antonini and bt Bmnori. in: fiemoglobin and myo@bin in their reactions with li_gands(North-ffoIIand. Amstrrdam. 1971) p_ I9_
446
1 June 1979
[ I41 T. Kitamwa, Ii_ Ogoshi. E Watanabe and Z- Yoshida. Chem- Phys. Letters 30 (1975) 451; J. Phys. Chem. 79 (1975) 2629. [IS] R-H. Felton. A-Y. Ronuns, N-T. Yu and G-R_ Schonbaum. Biochim- Biophys. Acta 434 (1976) 82_ [ 161 G. Rakhit, T-G. Spiro and M. Uyedn, Biochem. BiophysRcs. Commun. 71 (1976) 803. [ 171 J-P. Colhmm and CA. Reed. J- Am. Chem- Sot- 95 (1973) 2048.
CHEMlCAL
PHYSKS
LL?lTERS
1 June 1979
Teizo KITAGAWA and Junji TERAOKA Institute for Protein Research, Osurl-aUtir ersity, St&a. Osaka 565. Japmt Received 16 October
1978:in
fwl
form 3 April 1979
The resonance Raman and absorption spectra of unligatcd ferrous ocractb~ lporplwrin t&*fOEP)] and Its adduct tlith I”Velm or TtIF acre observed. The resommcc Ramaa spectrum of Te”(OEP) in CIi~Cl2 dispbq cd cbamctcristic features of iron porph>rio &ith three d, electron+. susgesting 3Ee 3~ the intermediate-spin (S = 1) sound st.tte_fe’*t.OIZP) in TIfF ‘i e\hibited the ferrous high spin feature like Fe2*(OEP) (_-McIm) in Cff=Ctz-
1. Introduction
The existence of intcr~nedi~te-span state (S = I) as a ground state of ferrous porphyrin had been predicted for a square pku~ar structure with a moderate @and field [I 1, and Collman et ai. [3] first isolated such ;Lcompound; tctr~phcnylporpi~~~~n~to iron
the aqueous solution of dithionite-reduced porphyrin in the presence of air [I I1 ; the absorpiion spectrum resembled titose of so-wiled “ohygensted“ ferrous porphyrin [ 121 snd oxygenated hemoglobin [ t 31 and the Rsman spectrum resembIed that of osyg.enated hemoglobin and chrtnged to ferrous high-spin type upon addition of 2-methyi imidnzotc Q-MeIm)_ To establish the absorption and Raman spectra of intermediate-spin ferrous porphyrin with more qualified preparation, i.e. in the absence of dithionite and contaminated water. we constructed a device to measure the absorption and Raman spectra of a hi$i!y evacuated system and successfutiy used it \\ith .I reactive intermediate-spin ferrota porphyrin_ Here we report the absorption and resonnnce Ramsn spectra of Fez” (OEP) with no asiaI iigmd and its adducr with 2-M&n or tetmhydrofuran (THF). proposing 3 Es ;LS a fikefy ground state.
3. Experimental Oct3eth~~porp~y~n~to iron (It) bis-pyrrdine complex [Fe”’ (OEP) @y&f (I). obtained by hydra&ehydrate reduction of F>3+ @XZP) Cl in pyridine (Py)_ was kindly provided by Dr_ H. Ogoshi md Mr.. H. Sugimoto of Kyoto University. The crystal of 1 wxi kept at 140°C for 2 h in 2 X 10M5 mm Hg in a Raman or visible absorption ceI1, yielding Fez’ (OEP) (2) [4]. 433
VoIume 63. number 3
1 June 1979
CHEiiICAL PHYSICS LJXJ-ERS
Since 2 is easily oxidized, the degassed solvent (THF, Py, or CH2CI2) was transferred to the ceII in the evacuated system_ To obtain Fez+ (CEP) (ZMeIm) (3), fresbiy recrystallized ZMeIm was put into the branched part of the cell, where it was kept at reduced pressure and room temperature during the heating of the cell. After 2 was formed in the ceII, 2-Meim was mixed with the solution of 2 without admitting air_ The Raman spectra were observed with the 488.0 nm excitation line of an argon ion laser (Spectra Physics. model 164) and a JEOL100D Raman spectrometer- The absorption spectra were measured with a Hitachi 124 spectrophotometer.
b.Py
3_ ResuIts and discussion The visible absorption spectra of 1,2, and 3 are shown in fig_ I_ I gave a typical spectrum of ferrous low-spin hemes (a). 2 in CH,CI, (b) and THF (c) gave spectra cfearly different from those of I and 3 (d)_ After observation of spectra b, c, and d, the solvent was rephced with Py in the evacuated system and reproduction of spectrum a was confirmed_ When evacuation was insufficient (<10d3 mm Hg), the reproduction of spectrum a was incomplete. indicating partial o.Gdation. The resonance Raman spectra of 1 in Py (A) and in CH$$ (B), 2 (C) and 3 (D) in CH+Ia are shown in fig. 2. Here again, reproduction of spectrum A was
I
1
Y)
1600
1
i
1500 I400 i;/ cm-1
9
1300
1&c
rig. 2- The resonance Raman spectra of Fe** (OEP) (PY)~ in pyridine (A) and in CH2C12 (B), Fez+ (OEP) in Cti~C.1~(C). and Fez* (OEP) (Z-MeIm) in CHzCtz (D)_ Excitation: 488.0 nm. The Raman lines marked by S me due to soWent or free 24feIm and those desigx&ed by b.Py indimte the resonzmeeenhanced Rzman Linesof bound pyridines.
4OC
. . . ..!...... 500
600
700
X/nm Fis- t_ Absorption spectra af Fez+ (OEP) (Py?+ in pyridine (a). Fe*‘(OEP) in CH+Clz (b). Fe** (OEP) in THF (c) and Fe*‘(OEP) (XfeIm) in CH+& (d)- Soret region is measured for different prepsrarion \iith more dilute concentmtion_
confirmed after each measurement by replacing the solvent with i’y_ fn spectrum B, the resonance-enhanced Raman lines of bound pyridine [IO] (marked by b, Py) were prominent but disappeared in spectrum C, evidencing eIimination of bound pyridine- When 2hleim was added to 2, the resultant spectrum (D) had Raman characteristic features of ferrous high-spin hemoproteins [6,S] _Therefore, 2 is undoubtedly a planar ferrous porphyrin without any axial Iigand, and thus is an intermediate-spin ferrous porphyrin_ We emphasize that spectrum C in fig. 2 is remarkably close to the Raman spectrum of ferric low-spin iron porphy-
Volume 63, number 3
CHEMICAL
PHYSICS LETTERS
I June 1979
rin such as Fe3+ (OEP) (Im), [ 14]_ This similarity was also pointed out by Spiro and Burke for their intermediate-spin ferrous porphyrin [lo] _ It is we11established that the ITtype ferrous Iowspin hemoproteins
having three d, eiectrons give rise
to Rarnan spectra similar to those of ferric low-spin hemoproteins [7,_8,10], while the a type ferrous lowspin hemoproteins [S] having four d, electrons and feryi low-spin hemoprotein (Comp. II or horseradish peroxidase) [ 15,161 having two d, electrons show different Raman spectra. Therefore the Ramrtn results strongly suggests the presence of three d, electrons for the intermediate-spin ferrous state. There are three possible intermediate-spin states; 3B k’ 3A2g,and3E g_ The 3B,, state (d$d,,,d=z) is incompatible not only with the Raman spect&n but aiso with the MijssbauerJ2] and NMR data [5]. For the 3 A2e state (d&, d$ d;), the Raman band IV frequency 171 is expected to be higher than that of the ferric tow-spin motecules. as demonstmted by the feryl Iow-spin compound [ 15.16], due to Iess delocalization of electrons to the porphyrin rr* orbital and furthermore, the d=l orbikd is unlikely to be located lower than the 4, orbital_ Only the ‘E, state (d-z, d~d=~) is compatible with the MBssbauer [2].%hlR [5]-, and Raman data, although mixing of this state with the high-spin state through spin-orbit coupling cannot be ruled out- The empty dxz_.vT orbital is also consistent with the short Fe-N distance [2]. The spin state of the THF adduct of 2 is controversia1 [I ,4,17]. In fig_ 3 the resonance Raman spectrum of 2 in THF is compared with those of 3 (high spin) and 1 (low spin) in THF, although the latter is partly dissociated into 2. It is clear that the Raman spectrum of 2 in THF distinctly differs from that of 2 in Cl&Cl, _ suggesting complex formation of 2 with THF. Theresultant spectrum is close to that of 3 in THF which differs littIe from spectkm D in fig. 2_ Therefore, the THF adduct of Fe’* (OEP) is considered to take the ferrous high-spin state, in good agreement with magnetic susceptibiIity data (5. I pg at 25OC) [ i7]_
_kknowIedgement The authors wish to express their gratitude
to Dr.
H_ Ogoshi and H_ Sugimoto of Kyoto University for their courtesy of providing FeZf (OEP) (Py)? _
1-e
1600
EC0
coo
‘300
.200
‘r.=
cse
0-u-t
J/Cl--I
Fig. 3. The resonmce Rnman specrm of Fe.‘+ (OEP), I-I?’ (OEP) (2DIeIm) and Fe2 (OEP) (P&jz in THT. T?w Ranan lines marhed b) b,Pv ar2 due to bound p: ridine-
References [ 1 j H. Koboqashi 2nd Y. k x~y3~~a. Bul!. Chem. Sot_ Japan IS (1972) 150(71 J-P. Col1m.m. J-L. Ziowi. N. Kim. G. Lang .mc! C A. Reed, J_ Am. Chem_ SOL 97 (1975) 2676_ [3] L.J. Radoncz\lcb,A. Bloom and J-L. Hoard, J. Am. Chem. Sot. 91(1972) 2073. [4j D. Dolphin, J.R. Snms, T-B. Tsin and K-L. N’ong. J. Xm. Chem. Sot. 9s (1976) 6970. [S] II. Gaff, GX LAiar rrnd C.X Reed. J. Am. Chem. SOC. 99 11977) 3641 161 T.G. Spiro and T-C. Strskas. j. Am. Chem. Sot- 96 (1971) 33& [7] T_ Kitsgwa. T_ Iizuka. M. %ito and Y. K?ogoLu. Chcm. Letters (1975) S-%9[S] T. Kitzgaws. I-_ K~ogoku, T_ Iizuka .md 11 I. S&o. J. Am. Chem. Sot. 9s (1976) 5169. [9] Y. Oaki. T. Kitngawa, Y. K>o~oku. C. Ik&imotoYutsudo, Y. Imai and R. &to_ Biochemistry. to be publisbed.
445
V5zme [IO]
63. number
3
CHEMICAL
PHYSICS LETTERS
T-G- Spiro and J-M- Burke, J_ Am_ Chem- Sot. 98 (1976) 5482 [I I] T- Kitagawa, Y. Ozaki. J. Teraoka, Y. Kyogoku and T. Yanumh. Biochim. Biophys. Acta 494 (1977) lOO_ [ 121 J-0. Alben, W-H. Fuchsman, CA_ Bcaudrcau and W_S. Caughey. Biochemistry 7 (1975) 625. [ 13 1 E Antonini and bt Bmnori. in: fiemoglobin and myo@bin in their reactions with li_gands(North-ffoIIand. Amstrrdam. 1971) p_ I9_
446
1 June 1979
[ I41 T. Kitamwa, Ii_ Ogoshi. E Watanabe and Z- Yoshida. Chem- Phys. Letters 30 (1975) 451; J. Phys. Chem. 79 (1975) 2629. [IS] R-H. Felton. A-Y. Ronuns, N-T. Yu and G-R_ Schonbaum. Biochim- Biophys. Acta 434 (1976) 82_ [ 161 G. Rakhit, T-G. Spiro and M. Uyedn, Biochem. BiophysRcs. Commun. 71 (1976) 803. [ 171 J-P. Colhmm and CA. Reed. J- Am. Chem- Sot- 95 (1973) 2048.