Pyrene-4,5-oxide

Epoxide hydratase activity, with JH-benzo(a)pyrene 4,5-oxide as substrate, was assayed by the thin-layer chromatographic procedure of Jerina et al. (15). The protein content of microsomal and whole homogenate preparations was determined according to Lowry et al. (16), using bovine serum albumin as the standard, and microsomal cytochrome P-450 content was assayed by the method of Omura and Sato (17) on an Aminco DW-2A spectrophotometer. 

In a simulated atmosphere, direct epoxidation by ozone led to the formation of benzo[a]pyrene-4,5-oxide. Benzo [a] pyrene reacted with benzoyl peroxide to form the 6-benzoyloxy derivative (quoted, Nikolaou et al, 1984). It was reported that benzo [a] pyrene adsorbed on fly ash and alumina reacted with sulfur dioxide (10%) in air to form benzo[a]pyrene sulfonic acid (Nielsen et al., 1983). Benzo [a] pyrene coated on a quartz surface was subjected to ozone and natural sunlight for 4 and 2 h, respectively. The compounds 1,6-quinone, 3,6-quinone, and the 6,12-quinone of benzo[a]pyrene were formed in both instances (Rajagopalan et al., 1983). 

Epoxidation of aromatic hydrocarbons is an important method for the preparation of arene oxides. m-Chloroperbenzoic acid (MCPBA) is used in a two-phase system that involves treating the hydrocarbon with a large excess ( 10-fold) of MCPBA in methylene chloride-aqueous sodium bicarbonate at room temperature. The yields are moderate (10-60%). Because the arene oxides are sensitive to acids, the presence of sodium bicarbonate buffer is necessary. A number of K-region (see Section VII for a definition) epoxides like phenanthrene 9,10-oxide (1, 59%), 9,10-dimethylphenanthrene 9,10-oxide (2,40%), 9-phenylphenanthrene 9,10-epoxide (3,50%), pyrene 4,5-oxide (4, 14%), and chrysene 4,5-oxide (5,9%) have been prepared by this method.9... 

The cis- 1,2-glycols, obtainable from the parent aromatic hydrocarbon by osmium tetroxide hydroxylation, can be converted to the corresponding trans-1,2-glycols by oxidation-reduction, using a mixture of dimethyl sulfoxide, sulfur trioxide, and pyridine, followed by lithium aluminum hydride reduction. The trans- 1,2-glycols can be dehydrated to arene oxides using DMF-DMA as mentioned above. Benzo[a]pyrene 4,5-oxide (28) and 7,12-dimethylbenz[a]anthracene 5,6-oxide (30) have been prepared by this method in 68 and 80% yields, respectively.18... 

Armstrong et al.223 have shown that nonenzymatic trans addition of glutathione to synthetic ( + )-(4S,51 )- and (—)-(4R,5S)-benzo [a]pyrene 4,5-oxides (364 and 365) occurs at carbons 4 and 5 to give two diastereomeric pairs of positional isomers to almost equal extents. Correlations of the glutathione conjugates obtained from the 4,5-oxide derived from cytochrome P-450c-catalyzed oxidation of benzo [a] pyrene with those obtained from the syn-... 

The assay method described by Eaton and Stapleton (1989), measures the activities of both cytosolic glutathione 5-transferase and microsomal epoxide hydrolase toward benzo[a]pyrene-4,5-oxide as a substrate. These enzymes are important in the biotransformation of many epoxide xenobiotics, including potentially carcinogenic arene oxides. 

Enzyme assays were performed in 1.5 mL microcentrifuge tubes containing 165 /nL of incubation buffer (0.25 M Tris-HCl, 0.1 mM EDTA, pH 7.4), 50 fiL of 10 mM glutathione in incubation buffer, and 25 /nL of appropriately diluted enzyme. The reaction was initiated by addition of 10 /nL of benzo [a]pyrene-4,5-oxide in acetonitrile (0.875 mg/mL). The assay was stopped by addition of 0.25 mL of acetonitrile containing 300 /nM 2-methoxynaphthalene as an internal standard. The solution was kept overnight in the dark at 4°C, and then 500 /nL of distilled water was added. The mixture was centrifuged before analysis by HPLC. Production of both the glutathione and diol derivatives was linear with time and protein up to 15 minutes and 500 /ng/mL, respectively. 

Figure 18. Stereochemical course of the microsomal epoxide-hydrolase catalyzed hydration of (-) )- and (—)-benzo[a]pyrene 4,5-oxide. About 99% of the attack by water occurs at the (5S)-carbon of the (—)-(4R,5S)-oxide and about 85% at the (4S)-carbon of the (-l-)-(4S,5R)-oxide. ...

Data are relative to benzo[a]pyrene 4,5-oxide (2,300-2,700 histidine revertants per nmol in the two tester strains) which appears to be the most mutagenic K-region arene oxide known in this test system. 

Varying degrees of substrate enantioselectivity have been described for microsomal epoxide hydrolases. For example, for benzo(a]pyrene-4,5-oxide a 40-fold difference between the rates of hydration for the +) and (-) enantiomers was observed in vitro (Armstrong et al., 1980). Mono-substituted epoxides (such as 1,2-epoxyhexane and its geometrical isomer. 

Figure 10 Possible transition state for the reaction of an epoxide hydroiase-oxide-HjO complex, illustrating a general add-base catalysis mechanism for the hydrolysis of benzo[a]pyrene-4,5-oxide. (Taken from Armstrong, 1987.)...

R-configured carbon atom vide supra), microsomal epoxide hydrolase attacks at R- as well as S-configuxed carbon atoms. With some arene oxides, both enantiomers are attacked at the same carbon, irrespective of configuration (e.g., benzo[c]phenanthrene). Other arene oxides (e.g., benzo[a]-pyrene 4,5-oxide) undergo the primary reaction, predominantly at S-con-figured carbon (Yang, 1988). 

Aromatic amines Heterocychc amines. Cumene hydroperoxides Aflatoxin B1 epoxides Benzo [a] pyrene. 4,5. oxide TranS Stilbene oxide 4-nitrochinolone 1-oxide... 

Jennette, K.W., Jeffery, A.M., Blobstein, S.H., Beland, F.A., Harvey, R.G., and Weinstein, I.B. (1977) Nucleoside adducts from the in vitro reaction of benzo[o]pyrene-7,8-dihydrodiol-9,10-oxide or benzo[a]pyrene-4,5-oxide with nucleic acids. Biochemistry, 16, 932-938. 

Hydration Flounder, Sheepshead Styrene oxide, Benzo la] pyrene 4,5-oxide,... 

Fig. 31.27 Marker substrates- of EH. (1) cis-stilbene oxide (2) butadiene monoxide (3) benzo[a]pyrene-4,5-oxide (4) trans-stilbene oxide (5) 11,12-epoxyeicosatrienoic acid.

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