Polypodium vulgare L. (Wood Fern): In Vitro Cultures and the Production of Phytoecdysteroids

Messeguer, Joaquima; Melé, Enric; Reixach, Natàlia; Irurre-Santilari, Josep; Casas, Josefina

doi:10.1007/978-3-642-58833-4_17

Cited by 9 publications

(15 citation statements)

References 30 publications

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“…A second question of interest concerns the regulation of ecdysteroid production. It has been repeatedly observed that cell suspensions or callus cultures from ecdysteroid‐producing species produce at best very small amounts of ecdysteroids, while tissue and hairy‐root cultures produce much larger amounts [23,39–41]. This means that tissue organization is required for adequate productivity.…”

Section: Discussionmentioning

confidence: 99%

“…All cultures were incubated at 22 ± 2 °C under 16 h light (Sylvania cool white, 5 µ m ·s −1 ·m 2 ). Selected callus lines could be maintained in in vitro conditions by subculturing every 6 weeks [23].…”

Section: Methodsmentioning

confidence: 99%

“…We report here on the results of investigations on the biosynthetic steps of phytoecdysteroids using incorporation of labelled ecdysteroid and cholesterol derivatives by the above tissue cultures. Likewise, although in vitro calli cultures of P. vulgare do not produce ecdysteroids [23], they have the enzymatic capacity to carry out different reactions implied in the biosynthetic routes of these compounds. Taking advantage of the morphological characteristics of this plant material that facilitates the incorporation of labelled compounds, we also studied biotransformations of selected substrates related to ecdysteroid biosynthesis and previously shown to be efficiently converted by arthropod steroidogenic organs [24,25].…”

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Biotransformations of putative phytoecdysteroid biosynthetic precursors in tissue cultures of Polypodium vulgare

Reixach

Lafont

Camps

et al. 1999

European Journal of Biochemistry

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Incubation of calli and prothalli of Polypodium vulgare with different tritium-labelled ecdysteroids has led to modification of some previous assumptions about the biosynthesis of ecdysteroids in plants. Thus,-hydroxyecdysone was transformed efficiently in both tissues into 20-hydroxyecdysone (20E), but no 25-deoxyecdysteroids such as pterosterone and inokosterone were formed. Likewise, incubation of 2-deoxyecdysone (2dE) produced exclusively ecdysone (E) and 20E, indicating a high 2-hydroxylase activity in both tissues, despite calli not producing phytoecdysteroids. This 2-hydroxylation was also evident in the transformation of 2,22-dideoxyecdysone (2,22dE) into 22-deoxyecdysone (22dE). Different ecdysteroids that do not occur in P. vulgare were formed in the incubation of 3-dehydro-2,22,25-trideoxyecdysone (3D2,22,25dE) by 3a-reduction and 3b-reduction and 25-hydroxylation processes. The fact that 22,25-dideoxyecdysone and 22dE were the only 2-hydroxylated products formed in this case suggests that only compounds bearing a 3b-hydroxyl group are substrates for the 2-hydroxylase. Surprisingly, 22-hydroxylation was never observed with either 2,22dE or 3D2,22,25dE, raising the possibility that it could occur at an early step in the biosynthetic pathway. In this respect, labelled 22R-hydroxycholesterol was efficiently converted into E and 20E, whereas 22S-hydroxycholesterol was not transformed into ecdysteroids, because of its unsuitable configuration at C22. Finally, the conversion of 25-hydroxycholesterol into E and 20E was greatly enhanced after thermal treatment of prothalli which induces the release of previously stored ecdysteroids. Thus, P. vulgare prothalli and calli appear to be particularly suitable models for the study of ecdysteroid biosynthesis and its regulation in plants.Keywords: 20-hydroxyecdysone; fern; hydroxycholesterol derivatives; in vitro cultures; induction.Many plant species produce C 27 ±C 29 ecdysteroids of a wide structural diversity [1±5]. These analogues of insect molting hormone (20-hydroxyecdysone, 20E) are secondary metabolites which are expected to provide some protection to plants against non-adapted phytophagous insect species [6]. Although the presence of such compounds in plants was demonstrated more than 30 years ago, little has been learnt since about their biosynthetic pathway(s), and whether it operates in a sequence that is similar to or different from that in Arthropods is not known.In vivo and in vitro labelling experiments have established that these compounds are biosynthesized from mevalonate via C 27 , C 28 and C 29 phytosterols. Cholesterol has been proved to be the intermediate in the biosynthesis of C 27 phytoecdysteroids in Taxus baccata [7]. In biosynthetic studies with Ajuga reptans roots transformed with Agrobacterium rhizhogenes, 24-methylcholesterol and 24-ethylcholesterol have been proposed as intermediates for the biosynthesis of C 28 and C 29 ecdysteroids, respectively [8]. Adler & Grebenok [9] proposed lathosterol (5a-cholest-7-en-3b-ol) as the logical ...

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Biotransformations of putative phytoecdysteroid biosynthetic precursors in tissue cultures of Polypodium vulgare

Reixach

Lafont

Camps

et al. 1999

European Journal of Biochemistry

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“…PEs are natural products that can be found in several tissues of some plant species [2]. We reported the presence of PEs in P. vulgare , mainly in sporophytes, rhizomes and protallus tissues [10].…”

Section: Discussionmentioning

confidence: 99%

“…Polypodium vulgare is a fern with a worldwide distribution that usually grows in dark and humid areas, and produces PE, the highest content being in rhizomes (up to 0.4% dry weight). Different culture lines of prothalli and calli from this plant have been obtained and their PE content has been described by our group [5–10]. Among those models, a calli line with undetectable levels of PE, but that is able to transform E into 20E, was selected for this study.…”

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The first cytochrome P450 in ferns

2005

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The fern Polypodium vulgare is a phytoecdysteroid (PE)‐producing plant. Cultures of P. vulgare prothalus produce PE, whereas prothalus‐derived callus cultures do not. However, this callus line can transform topically applied ecdysone (E) to 20‐hydroxyecdysone (20E), which is the last step in the biosynthetic pathway of the main plant PE. This hydroxylation is catalysed by a cytochrome P450 enzyme. E treatment of the callus line results in an increased amount of P450, showing a linear correspondence between the amount of P450 and in vivo E 20‐hydroxylation activity, estimated by measuring the bioconversion of E to 20E. This activity can be inhibited by molecules that bind to the P450‐heme group. E shows a P450‐substrate‐binding spectrum with microsomes that overexpress the P450 protein. Finally, a P450 protein was purified from E‐treated calli, this being the first P450 to be described in the pterydophyte group.

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Ecdysterone: Possible sources of origin in urine

Kwiatkowska

Grucza²,

Chajewska³

et al. 2022

Drug Testing and Analysis

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Ecdysterone (crustecdysone; beta‐ecdysone; 20‐hydroxyecdysone) is a naturally occurring steroid hormone belonging to the ecdysteroid class. The presented study investigated the possible concentration range of ecdysterone in urine after consumption of various preparations of spinach, drinking tea (made from Rhapoiticum Carthamoides) and topical use of a cream containing Cyanotis arachnoides. It is very important to establish reference ranges reflecting concentrations compatible with dietary habits and common uses of care products. The data obtained in the research may be used in the interpretation of results of routine analyses. In addition, elimination time and observed concentrations provided by the studies conducted by the Polish Anti‐Doping Laboratory can be used by WADA. In the case of spinach, peak elimination occurred within the first few hours, followed by a rapid decline. As for the other plants, instead of clear peak concentrations, gradual elimination was observed. Individual differences were observed between volunteers depending on route of administration. Differences in ecdysterone elimination following ingestion of spinach‐based and other plant products were observed too. The highest observed ecdysterone concentration was related to the paste consumption, and it was 691 ng/ml. Finally, our findings were compared with the data collected for the samples routinely tested as part of the monitoring program. During 2.5 years, the presence of ecdysterone was confirmed in as many as 507 samples out of 11 191 total samples tested. The concentration range was very wide, from 1 ng/ml (which is the LOD for this method in the Polish Anti‐Doping Laboratory) to over 2000 ng/ml.

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Polypodium vulgare L. (Wood Fern): In Vitro Cultures and the Production of Phytoecdysteroids

Cited by 9 publications

References 30 publications

Biotransformations of putative phytoecdysteroid biosynthetic precursors in tissue cultures of Polypodium vulgare

Biotransformations of putative phytoecdysteroid biosynthetic precursors in tissue cultures of Polypodium vulgare

The first cytochrome P450 in ferns

Ecdysterone: Possible sources of origin in urine

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