A-Hydroxyphenazine

B. a-Hydroxyphenazine (demethylalion). A solution of 4.2 g. (0.02 mole) of a-methoxyphenazine, from A above, in 125 ml. of 55% hydrobromic acid (Note 7) is placed in a 250-ml. round-bottomed flask fitted with a reflux condenser. The flask is immersed in an oil bath, and the solution is heated to 110-120° for 5 hours the evolved gases are absorbed with water in a trap. The reaction mixture is cooled to room temperature, diluted with about 125 ml. of water, almost neutralized with sodium hydro.xide (Note 8), and extracted six times with 30- to 40-ml. portions of ether. The combined ether extracts arc extracted with 25-ml. portions of 10% sodium hydroxide solution (Note 9) until no more purple sodium salt is remox ed from the ether. The aqueous extracts are combined, made acid to litmus with dilute acetic acid, and re-extracted four times with 50-ml. portions of ether. The combined ether extracts are dried over anhydrous sodium sulfate, and the ether is removed by distillation on a steam bath. The residue is recrystallized as follows It is dissolved in the least possible amount of hot alcohol, water... 

C. Pyocyanine alkylation). A solution of 2 g. (0.011 mole) of a-hydroxyphenazine in 13.4 g. (10 ml., 0.1 mole) of methyl sulfate (Note 10) is placed in a 250-ml. Erlenmeyer flask fitted with a calcium chloride drying tube and heated at 100° (oil bath) for 10 minutes. The solution is allowed to cool to room temperature, and about 7.5 ml. of dry ether is added. The dark brown solid is filtered on a 7-cm. Eiichner funnel and washed with about ISO ml. of dry ether in several portions (Note 11). 

The residue, without recrystallization, is pure enough for the preparation of a-hydroxyphenazine. 

Hydroxypentanal, 27, 43 a-Hydroxyphenazine, methylation to quaternary salt, 26, 87 2-(/3-Hydroxypropyl)-pyridine, 23, 83 2-(7-Hydroxypropyl)-pyridine, 23, 85 N-(7-Hydroxypropyl)-tetrahydroquin-oline, 26, 41... 

N-MethylaHon. The last step in a synthesis of pyocyanine (6), N-methylation of a-hydroxyphenazine (4), is carried out by heating 2 g. of (4) with 10 ml. of dimethyl sulfate at 100° for 10 min, cooling, and washing the dark brown metho-... 

Pyocyanine. 1 -Hydroxy-S-methylphenazini u m hydroxide inner salt. C,3HWNJ3 mol wt 210.23. C 74.27%, H 4.80%, N 13.32%, O 7.6]%. Produced by Pseudomonas aeruginosa (Ps. pyocyanea, Bacillus pyocyaneus). Isoln from broth cultures Wrede, Si rack. Z. Physiol Chem. 140, 1 (1924). Isoln from agar cultures Sohoental, Brit J, Exp Pathol. 22, 137 (1941). Synthesis by dimethyl sulfate meth-ylation of a -hydroxyphenazine and alkaline treatment of the resulting methosulfate Wrede, Strack, Z. Physio/, Chem. 181, 58 (1929) Ber. 62, 2051 (1929) Surrey, Org- Syn. 26, 86 (1946) coll, voL HI, 753 (1955). 

The same percentage yield was obtained with double the amounts specified above. The pyocyanine thus obtained can be stored in a vacuum desiccator in the dark for several weeks without appreciable decomposition. It slowly decomposes on longer standing to give the yellow a-hydroxyphenazine. 

Phenazines — The phenazines are biosynthesized by the shikimic acid pathway, through the intermediate chorismic acid. The process was studied using different strains of Pseudomonas species, the major producers of phenazines. The best-known phenazine, pyocyanine, seems to be produced from the intermediate phenazine-1-carboxylic acid (PCA). Although intensive biochemical studies were done, not all the details and the intermediates of conversion of chorismic acid to PCA are known. In the first step, PCA is N-methylated by a SAM-dependent methyltransferase. The second step is a hydroxylative decarboxylation catalyzed by a flavoprotein monooxygenase dependent on NADH. PCA is also the precursor of phenazine-1-carboxamide and 1-hydroxyphenazine from Pseudomonas species. - - ... 

Methanophenazine 239 functions as an electron carrier in the cytoplasmic membrane. The etherification of 2-hydroxyphenazine 237 with mesylate 238 was a key step in a total synthesis of 239 <00AC(E)2470>. 

The result of the retrosynthetic analysis of rac-lO is 2-hydroxyphenazine (9) and the terpenoid unit rac-23, which may be linked by ether formation [29]. The rac-23 component can be dissected into the alkyl halide rac-24 and the (E)-vinyl halide 25. A Pd(0)-catalyzed sp -sp coupling reaction is meant to ensure both the reaction of rac-24 and 25 and the ( )-geometry of the C-6, C-7 double bond. Following Negishi, 25 is accessible via carboalumination from alkyne 27, which might be traced back to (E,E)-farnesyl acetone (28). The idea was to produce 9 in accordance with one of the methods reported in the literature, and to obtain rac-24 in a few steps from symmetrical 3-methyl-pentane-1,5-diol (26) by selective functionalization of either of the two hydroxyl groups. 

To further elucidate the function of methanophenazine (10) as electron carrier its redox potential was determined [45]. As Scheme 2 reveals, the redox potential of lO/dihydro-10 must range somewhere between the values for F420H2/F420 ( 0 =-360 mV) [46] and H2/2H ( =-420 mV), and the redox potential of the heterodisulfide CoB-S-S-CoM (22). Whilst the values of F420H2/F420 and H2/2H" have long been known, the redox potential of 22 remained an unknown quantity. 2-Hydroxyphenazine (9) had proved to be a satisfactory model sub-... 

That the second option can also be successfully used has recently been revealed by our synthesis of 2-methoxyphenazine (117) [90]. The reduction of o-bromo-o -nitrodiphenylamine 132 accessible via intermolecular Pd-catalyzed JV-arylation provides the o-amino-o -bromodiphenylamine 133, which can then be cyclized to give 117 in a Pd-catalyzed intramolecular AT-arylation by employing Pd2(dba)3 as the Pd complex and 134 as the phosphine ligand. It should be noted that the outcome of both the intermolecular and the intramolecular AT-arylations heavily depends on the appropriate choice of the Pd complex as well as the phosphine. Ether cleavage leads to 2-hydroxyphenazine (9). 

Rapid and sensitive HPLC methods were developed for the detection of an antimicrobial growth promoter and its main metabolites containing quinoxaline-2-carboxylic acid <2005MI1495>. The major phenazine pigments of Pseudomonas aeruginosa such as 1-hydroxyphenazine and phenazine-l-carboxylic acid <1997JCH(A)(771)99>, and 2-aminophenazine as an impurity in a bactericide <1999MI632>, were also analyzed by HPLC methods. 

Using ether-treated cells of P. aureofaciens, dicarbonyl-14C2 phenazme-l,6-dicarboxylic acid las (121)1 was found to be an efficient and specific precursor for phenazine-1-carboxylic acid (123), and also for 2-hydroxyphenazine-l-carboxylic acid (124). The rate of growth of the organism appeared to be important, because an incorporation was also recorded of the labelled (121) into (123), albeit at a lower level, with cultures that had been grown rapidly. The position of phenazine-1,6-dicarboxylic acid (121) as a universal intermediate in the biosynthesis of phenazines now seems secure. The previously reported failure of dimethyl phenazine-1,6-dicarboxylate (122) to act as a precursor of phenazines cf. Vol. 10, p. 28 Vol. 9, p. 29) has been confirmed with ether-treated cells of P. aureofaciens. Efficient hydrolysis of (125) to (123) did, however, occur.101... 

When phenolate anions are used as substrates, phenazine dioxides are produced. The same product—2-hydroxyphenazine dioxide (173 R1 = H, R2 = OH)—is formed, whether phenol, resorcinol, hydroquinone, or benzoquinone is used,9,390,392 illustrating the variability of the relationship between the oxidation levels of the substrates and products. Benzofuroxan, a trialkylphosphine, and a quinone produce blue-violet phos-phonium betaine derivatives (e.g., 173 R1 = PR3+, R2 = 0-).393 The ether 173 (R1 = H, R2 = O-alkyl) is produced from a hydroquinone monoether.390 1-Hydroxyphenazine dioxide (173 R1 = OH, R2 = H) can be prepared from benzofuroxan, cyclohexane- 1,2-dione, and a base, followed by oxidation of the mixture of mono- and di-iV-oxide formed.394 /J-Naphthol provides benzo[a] phenazine dioxide (174).9,390... 

It was of interest to see whether the betaine (171) would serve as a precursor for 2-hydroxyphenazine-l-carboxylic acid (177) and 2-hydroxyphenazine, which are produced by Ps. aureofaciens along with 1-carboxyphenazine (170). ° 2-Hydroxyphenazine-l-carboxylic acid could, it was thought, arise by opening of the epoxide ring of (172) in an alternative manner to the one which would give pyocyanin. In the event, however, only 1-carboxyphenazine served as a precursor for (177) and 2-hydroxyphenazine the respective levels of incorporation are in accord with the sequence (170)— (177)— 2-hydroxyphenazine. As 1-... 

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