Microbial Models of Mammalian Metabolism<i>O</i>-Demethylations of Papaverine

Rosazza, John P. N.; Kammer, Marianne; Youel, Leisa; Smith, Robert V.; Erhardt, Paul; Truong, Duyen H.; Leslie, Steven W.

doi:10.3109/00498257709036245

Cited by 38 publications

(15 citation statements)

References 12 publications

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“…They are the only biotransformed compounds from papaverine in hepatic microsomes of guinea-pig. These papaverine biotransformation compounds can also be noted in microorganisms: a 6-demethyl derivative in Aspergillus a!liaceus and a 4'-demethyl in Cunninghamella echinulata (9). In this case, there seems to be a close parallel in the biotransformation of isoquinoline molecules by animals, plants and microorganisms.…”

Section: Biotransformed Compoundsmentioning

confidence: 88%

Papaverine Biotransformation bySilene albaCell Suspension

Verdeil¹,

Bister-Miel²,

Guignard³

et al. 1986

Planta Med

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Silene alba cell suspensions were supplied with papaverine hydrochloride. Seven biotransformation compounds were obtained of which four were alkaloidal. The two most important compounds are regioselective demethylation products: 6-monodemethylpapaverine and Inthe field of vegetal cell biotransformation of alkaloids, little work has been performed on isoquinoline molecules. The bioconversion of reticuline by calluses of Papaver somniferum L. was studied previously (1). In this work we study the papaverine biotransformation by Si/ene alba Miller E. H. L. Krause cell suspension. Because S. alba cells are non-alkaloid producers, the biotransformation products are easily isolated. MaterialsCell cultures of S. alba were grown in a Murashige and Skoog (2) liquid medium with sucrose (30 g.lj, adenine (7.4 106 M), 2,4-dichlorophenoxyacetic acid (5.l0_6 M) and kinetin (5. 10_6 M), as described earlier (3).The papaverine hydrochloride Codex was used as the substrate for these studies. The stability of a 1 g.l substrate solution was checked under experimental conditions after autoclaving in the culture medium; no new structures were demonstrated by thin layer chromatography (TLC). MethodsCell suspensions with papaverine hydrochloride were incubated at 23°C C on a rotary shaker at 180 rpm under illumination of 25 sE.m2.s' for 12 hours per day. After inoculation and seven days of growth, dry weight (DW) evaluations were performed. Cells samples were collected by reduced pressure sieving and dried at 60° C for 48 hours.The culture medium was first extracted. Residues of extractions are extracted again after hydrolysis with concentrated H2S04 to pH 0.9 for 36 hours alternating between room temperature and reflux conditions. Samples were periodically checked for products by TLC. Extractions were performed after alkalinization to pH 10 with concentrated NH4 OH using methylene chloride. The organic phase was shaken with HCI 0.1 N (pH 4.5). The aqueous phase was alkalized with concentrated NH4OH and extracted with methylene chloride, subsequently dried over anhydrous Na2SO4 and evaporated under reduced pressure. It had been previously determined that this method is capable of achieving a 96 % efficiency in removing papaverine hydrochloride. The cells were lyophilized and extracted with methylene chloride in a Soxhlet apparatus. Crude extract was cleaned and extracted as performed with the culture medium.Extracts were then separated by TLC and densitometry. TLC was performed on Merck silica gel 60 F 254 plates. Three solvent systems were used: chloroform/acetone (60/40, v/v); benzene/methanol (75/25, v/v); and acetone/methanol/ammonia 85/5/2, v/v/v). UV light (254 and 366 nm) and Dragendorff's reagent were used for the detection of papaverine and its metabolic alkaloid derivatives. Densitometry was performed using a Vernon type pH 16 photometer with a Mazda 1803 incandescent lamp. The Kiesel gel 60 F Merck 0.25 mm thick plates were developed in a chloroform/acetone (70/30, v/v) solvent system. Spots were made visible by immersing the...

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Section: Biotransformed Compoundsmentioning

confidence: 88%

Papaverine Biotransformation bySilene albaCell Suspension

Verdeil¹,

Bister-Miel²,

Guignard³

et al. 1986

Planta Med

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“…Other methoxylated compounds that are 0-demethylated by various species of Streptomyces include aconitine (2), anisole (27), 10,11-dimethoxyaporphine (25), glaucine (11), papaverine (24), and veratrate (31). Vanillate may be either demethylated or decarboxylated by S. setonii, depending on the original growth substrate (21,32).…”

Section: Discussionmentioning

confidence: 99%

Demethylation of Veratrole by Cytochrome P-450 in Streptomyces setonii

Sutherland

1986

Appl Environ Microbiol

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The actinomycete Streptomyces setonii 75Vi2 demethylates vanillic acid and guaiacol to protocatechuic acid and catechol, respectively, and then metabolizes the products by the ,-ketoadipate pathway. UV spectroscopy showed that this strain could also metabolize veratrole (1,2-dimethoxybenzene). When grown in veratrolecontaining media supplemented with 2,2'-dipyridyl to inhibit cleavage of the aromatic ring, S. setonii accumulated catechol, which was detected by both liquid chromatography and gas chromatography. Reduced cell extracts from veratrole-grown cultures, but not sodium succinate-grown cultures, produced a carbon monoxide difference spectrum with a peak at 450 nm that indicated the presence of soluble cytochrome P-450. Addition of veratrole or guaiacol to oxidized cell extracts from veratrole-grown cultures produced difference spectra that indicated that these compounds were substrates for cytochrome P-450. My results suggest that S. setonii produces a cytochrome P-450 that is involved in the demethylation of veratrole and guaiacol to catechol, which is then catabolized by the 1-ketoadipate pathway.

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“…Those previously documented include N-demethylation, deacetylation, and the formation of a group of related derivatives of dihydrovindoline ether (2a). In the hope of detecting new types of vindoline metabolites, we screened a variety of microorganisms from our own culture collection, many of which have demonstrated high capacity to metabolize alkaloids, steroids, and amino acid derivatives (14,10,15). Cultures obtained by soil enrichment procedures yielded no new vindoline metabolites.…”

Section: Methodsmentioning

confidence: 99%

“…They were incubated according to the twostage procedure previously described (1). All media were sterilized in an autoclave at 121°C for 15 Many types of microbial transformations of vindoline (la) are possible. Those previously documented include N-demethylation, deacetylation, and the formation of a group of related derivatives of dihydrovindoline ether (2a).…”

Section: Methodsmentioning

confidence: 99%

Microbial Transformations of Natural Antitumor Agents: O -Demethylation of Vindoline by Sepedonium chrysospermum

Nabih

Youel

et al. 1978

Antimicrob Agents Chemother

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Vindoline (la) was transformed to O-demethylvindoline (lb) by Sepedonium chrysospermum ATCC 13378 in 33% yield. Thin-layer and high-pressure liquid chromatographic methods distinguished lb from previously reported microbial metabolites of vindoline. The structural proof of lb was based on its mass spectral fragmentation pattern and on its chemical and nuclear magnetic resonance spectral properties.The Vinca alkaloids vincristine and vinblastine possess significant antitumor activity and are widely used clinically. They have resisted simple syntheses, and their structural complexity has hindered the preparation of other useful derivatives of the alkaloids by chemical methods. Microbial transformations are being examined for their potential to prepare potentially important Vinca alkaloid derivatives.Vindoline (la) is one of the monomeric Vinca alkaloids and one of the most abundant alkaloids in Vinca rosea L. Since it is more readily available than vincristine and vinblastine, it was used in an effort to identify new types of biotransformation products of this group of alkaloids. Previous reports document the production of a variety of metabolites of vindoline by streptomycetes. Known vindoline metabolites include ( Fig. 1): deacetylvindoline (Ic) and deacetyldihydrovindoline ether (2c) (9); dihydrovindoline ether (2a); 3-acetonyldihydrovindoline ether (2b); a ring contraction product known as 16-dehydroxy-14,15-epoxy-14-oxo-3-norvindoline (12); N-demethylvindoline (Id) (13); and a dimer consisting of two dihydrovindoline ether moieties linked through their 3 and 14 positions (11).This report describes selective cleavage of the methyl ether group of vindoline by Sependonium chrysospermum ATCC 13378. The reaction gives lb in 33% yields, and it is not complicated by the formation of other side products. MATERIALS AND METHODSMelting points were determined on a Thomas-Hoover apparatus and are uncorrected. Infrared spectra were determined on a Perkin-Elmer 267 grating infrared spectrophotometer. Nuclear magnetic resonance (NMR) spectra were determined on a VarianAssociates model T-60 spectrometer using tetramethylsilane as an internal standard. Low-resolution mass spectra were obtained on a Finnigan model 3200 spectrometer, and high-resolution mass spectral data were provided by the Department of Chemistry, Massachusetts Institute of Technology, Cambridge. UV spectra were obtained on a Beckman DK-2 instrument. Elemental analyses were determined by Integral Microanalytical Laboratories, Inc., Raleigh, N.C.Vindoline, deacetylvindoline, dihydrovindoline ether, N-demethylvindoline, 3-acetonyldihydrovindoline ether, and 16-dehydroxy-14,15-epoxy-14-oxo-3-norvindoline were obtained as authentic reference standards from Eli Lilly and Co., Indianapolis, Ind.The purity of each compound was verified by thinlayer chromatography (TLC), high-pressure liquid chromatography, and mass spectral analysis. Vindoline (7) dihydrovindoline ether and the dihydrovindoline ether dimer (11) were characterized by melting points and UV, NMR, and...

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Microbial Models of Mammalian MetabolismO-Demethylations of Papaverine

Cited by 38 publications

References 12 publications

Papaverine Biotransformation bySilene albaCell Suspension

Papaverine Biotransformation bySilene albaCell Suspension

Demethylation of Veratrole by Cytochrome P-450 in Streptomyces setonii

Microbial Transformations of Natural Antitumor Agents: O -Demethylation of Vindoline by Sepedonium chrysospermum

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