I
and G. C. Voge13
lthaca College Ithaca, New York 14850
Bioinorganic Chemistry
Much of the current research activity in inorganic chemistry deals with synthetic analogs to biologically important molecules. Among the most important and most actively researched metal-containing compounds in biological systems is the class comprised of complexes between metal ions and porphyrin ligands. In this article we describe two experiments involving porphyrin materials which we use in our junior level laboratory. These experiments reflect recent developments in this research field and, at the same time, are readily understood and performed by the undergraduate student. During the first class meeting or two, we discuss the ubiquitous and essential nature of porphyrins and porphyrin-like material (1-4). The students' enthusiasm for these projects is enhanced once they recognize the connection between the materials they will he using and certain biologically active substances. Project I. The Thermodynamic Study of the Adduct Formation of a Zinc Porphyrln with a Neutral Lewis Base In a Non-Aqueous Solvent
The study of the adduct formation between metalloporphyrins and neutral Lewis bases in non-aqueous solvents is one area of porphyrin chemistry currently being pursued (1, 2). In particular, considerable work has been done on the interaction of zinc a,O,y,6-tetraphenylporphinewith nitrogen donors (2) and a limited study carried out on its interaction with oxygen, sulfur, and phosphorus donors (5). Approximately six weeks of this laboratory course can he spent studying the 1:l adduct formation between several phenyl substituted derivatives of zinc tetraphenylporphine (Fig. 1) and pyridine or imidazole in non-aqueous solvents such as benzene, o-dichlorobenzene, or toluene. The equilibrium studied can be represented by A B = A:B where A is the zinc porphyrin, B the Lewis base, and A:B the 1:l adduct. The difference in uv-visible spectra of the metalloporphyrin and its adduct provides a convenient way of studying the equilibrium by'the application of Beer's Law. Either the Soret region (400-500 nm) or the visible region (500-700 nm) is suitable for following the equilibrium. Figure 2 shows the visihle spectra of zinc tetra(4-fluorophenyl). porphine with varying amounts of pyridine. If the equilihrium constant is large enough to allow the molar extinction coefficient of the adduct, CAB, to he measured, it is easily shown that
Figure 1. rn and pPhenyi substituted derivatives of zinc n.S.y.6-tetraphenylporphine where X = Ci. Br. F. H. OCHs. CHJ have been used. See the table.
+
K=-- X 1
-X
1 [B]
X -
a-0.4
(1)
- UA
(1.46
where [B] is the equilibrium concentration of B, a is the ahsorbance of the equilihrium mixture of zinc porphyrin and the Lewis base, a A is the absorbance of the zinc porphyrin, and a A B is the absorbance of the adduct. If K is not large enough to allow CAB to be determined directly, eqn. (1)can be recast into the Rose-Drago equation
- Le
a-aA (2) [Alo - [Blo + [ A l o [ B 1 0 ~ me-m where [A10 and [B]o are the initial concentration of the zinc K-1-
LAB
'ACS-PRF undereraduate scholar for the summer of 1974.
High school r t u d c n t i participating in the Student Science Training Progmm spunsored bv \'SF during the summer of 1973. ,\uthors to whom rwre.ipondence s h d d b addreaied.
'
Figure 2. Visible spectra of zinc tetra(4-f1uoraphenyl)parphine with varying amounts of pyridine in the solvent benzene. A is the spectrum of zinc tetra(4fluoraphenyl)porphinewithout pyridine.
Vo/ume 53, Number 6.June 1976 / 387
porphyrin and the Lewis hase, respectively, and 6~ = a ~ l [A]o. Equation (2) provides an excellent graphical method for examining the precision of the data and for checking for any dependence of CAB on the Lewis hase concentration range employed (5, 6). All the systems studied in the lab experiment have equilihrium constants large enough for me to he measured directly.4 The equilihrium constants follow the order one would predict on the basis of the electron-donating or electron-withdrawing nature of the meta or para suhstituent. No special apparatus, such as a glove hag or dry box, was needed. The equilihrium does not appear to he sensitive to small traces of water. The work was performed on an open bench. However, all solvents and liquid reagents were stored over Linde 4A molecular sieves for 24 hr prior to use.
Molar Extinction Coefficients at 550 nma s X 10'
Mefsllop~rphvrin
a S ~ I ~ e nbenzene. t:
Outline of Experiments for Part I
I J Preparation of porphyrin 2) Preparation and purlfiration of zinc porphyrin 3) UV-vmhle rncetrn ot zinc oor~hvrinand its adduct
4) Bear's Law kudy of zinc pbrphyh 5) Determination of equilibrium constant
Phenyl suhstituted derivatives of tetraphenylporphine and the corresponding zinc complex were synthesized and purified by literature methods (7, 8). The literature methods are auite exolicit and the students should he encouraged to cbnsu~tthe original literature in planning their syntheses. A list of the reagents and equipment needed to prepare a convenient quantity of porphyrin per student follows. Figure 3. Tetrakis(4-N-mthylpyridy1)porphine (TMpyP) is water soluble aver an enended pH range.
A. Porphyrin
Reagents benzaldehyde (or derivative) pyrrole propionic acid methanol
15 ml(0.16 mole) 11ml (0.16 mole)
500 ml 100 ml
Equipment simple distillation apparatus 1round bottom flask (1000 ml) with 2 necks 1 dropping funnel 1condenser 1heating mantle 1magnetic stirrer 1Buchner funnel and filter flask B. Zinc Porphyrin
Reagents porphyrin 8 mmale (e.g., 2.5 g of tetraphenylporphine) zinc acetate 8 mmole (1.47 g) N,N'-dimethyiformamide Equipment 1round bottom flask (IWO ml) with 1 neck 1 condenser 1heating mantle 1magnetic stirrer 1 Buehner funnel and filter flask Pyridine was distilled from BaO and stored over Linde 4A molecular sieves. Imidazole was recrystallized from heuzene. Solvents such as benzene, toluene, and o-dichlorobenzene were dried over Linde 4A molecular sieves and used without further purification. Spectroscopic data were collected using either a Beckman DB, Perkin-Elmer 124, or a Cary 14. Suggested conditions for determination of equilibrium constants in the visible region are 388 / Journal of Chemical Education
[Zinc porphyrin]
-
3.5-4.0 x 10-6 M
[Imidazole]- 10-3-10-5 M [Pyridine] 10-2-10-4 M
The table lists the molar extinction coefficients of the phenyl suhstituted zinc porphyrins synthesized by our students. Project II. The Kinetics of Metal Ion Insertion into a Water Soluble Porphyrin P o r ~ h v r i n sand oorohvrin-like materials which are hio- . logicaily-active invariably contain a metal ion. Over the past dozen years, considerable effort has been made to determine the mechanism(s) by which metal ions insert into porphyrin molecules to form metalloporphyrins (2)
MZ++
-
5PH~
11-2
MP
+ i:q
~ +
p-2
However, manv of these studies have led to results which interpret because porphyrin aggregation are difficult andlor ionic streneth effects were overlooked (9). . . This experiment is concerned with the insertion of copper ions into the water soluble porphyrin, tetrakis(4-N-methylpyridy1)porphine (TMpyP) (Fig. 3). This porphyrin is water soluble over an extended D H ranee and has been shown to remain monomeric in solu&n even in the presence of inert electrolytes such as ~ o t a s s i u mnitrate (10). The Eourse of the-reaction is followed by the rate of conversion of the absorption spectrum from the metal-free
tb
-
'Listings of computer programs employing eqns. (I), (2), or (6) are available upon request.
porphyrin to the metalloporphyrin, CUP. Therefore, an important starting place is the determination of the spectrum of TMpyP in the free base form ( P H z ) , ~the diacid form (PHd2+,p H O), and as the metalloporphyrin. From these spectra a convenient wavelength (or wavelengths) may he chosen to study the reaction. We have used 544 nm in our CUP + experiments. For the reaction Cu2+ ZqPHq Zqq[H+],the rate law may he written as
-
-
+
where P represents the metal-free porphyrin in its various acidified forms, pH2, pH3+, and PHd2+. The rate constant, k,, is ionic strength and p H dependent. ,O All of the studies are conducted such that [ C U ~ + ] ~>> [PIt=o and, therefore, the ahove expression may he rewritten as
where kobn= ~ , [ C U ~ + ] ~ (5) We, therefore, have a pseudo-nth-order reaction and from a plot of log(a, - a ) versus time, i t may be shown that n = 1. Experiments are conducted a t fixed p H (no buffer) and fixed ionic strength but varying copper(I1) concentration to determine rn (see suggested conditions under Experimental Section). In our experiments, as in those of Hamhright and coworkers ( 9 ) ,i t is found that m = 1. Experiments conducted a t varying ionic strength or p H lead to a determination of the deoendence of hr on these quantities. For the p H dependence, a good fit wiih the data is obtained bv assumine that the onlv attackine form of the porphyrin is P H ~
-
cu2+
+
PH,
k,,
CUP
+
2 ~ '
Outline of Experiments for Part 11 1) Synthesis of porphyrin and CuTMpyP 2 ) Absorption spectra of TMpyP and CuTMpyP 3) Preparation of stock solutions of Cu(NOd2, KNOB, and HN03. Determination of concentration of Cu2+stock. 4) Determination of rate constant
Tetrakis(4-N-methylpyridyl)porphinemay he synthesized starting with either commercially availahle or synthesized tetrakis(4-pyridy1)porphine (10, 12) or purchased from Stream Chemicals, Danvers, Massachusetts. Suggested conditions for determination of rate constants are Determination of m: pH = 2.0, rr = constant chosen value (0.2, 0.7, 1.0.2.0). At a rr = 2.0 M we have used [Cu2+]o= 5.0 X lo-', 1.0 X 1 0 V , 5.0 X 1.0 X lo-', 2.0 X 10@. At lower ionic strength, the low end of the copper concentration range leads to rates which are inconveniently slow. Determination of influence of pH: p = constant, pH = 1.2, 1.5, 1.8, 2.0, 2.2, 2.5, 2.8; For rr = 2.0 M , we used [Cu2+]o= 1.0 X (high p H ) - 1.0 X 1 0 V (low pH). Many variations of the experiments we have outlined above are possible. In particular, one could study the rate of insertion of zinc ion into tetraphenylporphine in a nonaqueous environment and fdlow ;hi+ wi;h the study of the adduct formation of zinc tetmphmylporphine ahme A n other possihility would he to studv the rate of inserrion of zinc ion into tetrakis~4-N-merhyIp~idyl~p~,rphine and to studv adduct formation of ZnT\lnvP with imidaaole or pyridine in aqueous solution. The role of catalysts in the copper insertion reaction might be studied; almost any buffer having metal complexing tendency will catalyze the insertion (as, for example, acetate). If the apparatus is available, the temperature dependencies of both the thermodynamic and kinetic constants could he investigated. A copy of the detailed handout which is given to our students is available upon request.
..
Literature Clted I11 Falk. J. E., "Porphyrina and Metalloporphyins." Elecvier Publishing Co.. Amster-
The expression which results from this mechanism is
dam. 1964. (21 Hambright. P., C o o 4 Chem. Re".. 6.247 11971) and private mrrapndence. (31 Martin,D. L.,andHuheey, J. E..J CHEM. EDUC.49.177 119721. (4) Hughes, M . N.,"The Inorganic Chemistry of Biological Processes." John WilW and Sons, New York. 1972. (51 Vogoi, Glenn C., and Searby, Lynn A.,lnorg Chsm.. 12,936 (1973). (6) Rose. N,J..andDrago. R. S.. J Amer. Chem. Soc.. 81,6138 11959). (7) Adler, A. D.. et d.,J. Or& Chem.. 32.475 (18681. (8) Adler, A. D.. etal., J. Inorg. Nucl Chem., 32.2443 (1970). I91 Baker, H.. Hambright, P.,and Wagnar,L.. J. Amer Chsm. Soc., 95.5942 (19731. (101 Ps.ternaek,R. F.,etal., J. Amer C h z m Sm.. 94,4511 119721. (111 Pastexrack, R. F., Sutin, N., and Turner, D. H., J. Amer Cham. S o c . 38. WW (121
The data may he analyzed to obtain values of ho, K.1, and Kaz or, if computer programs4 are not availahle, values of K,I and K.2 at varying ionic strengths can he found in the literature (9, 11).
(147EI ,.- .-,. ong go. F ~ . . ~ i n a r e l l~.G.,and~im,~. i, B., J H e m o w l . c h m , 6,927 11969).
51n using these shorthand forms for writing the porphyrin species, the charges at the periphery of the molecule (ion) are frequently neglected for convenience.
Volume 53,Number 6.June 1976 / 389
