Factors influencing the production of stilbenes by the knotweed, Reynoutria x bohemica

Abstract

Background: Japanese knotweed, Reynoutria japonica, is known for its high growth rate, even on adverse substrates, and for containing organic substances that are beneficial to human health. Its hybrid, Reynoutria x bohemica, was described in the Czech Republic in 1983 and has been widespread ever since. We examined whether Reynoutria x bohemica as a medicinal plant providing stilbenes and emodin, can be cultivated in spoil bank substrates and hence in the coalmine spoil banks changed into arable fields. We designed a pot experiment and a field experiment to assess the effects of various factors on the growth efficiency of Reynoutria x bohemica on clayish substrates and on the production of stilbenes and emodin in this plant.

Results: In the pot experiment, plants were grown on different substrates that varied in organic matter and nutrient content, namely the content of nitrogen and phosphorus. Nitrogen was also introduced into the substrates by melilot, a leguminous plant with nitrogen-fixing rhizobia. Melilot served as a donor of mycorrhizal fungi to knotweed, which did not form any mycorrhiza when grown alone. As expected, the production of knotweed biomass was highest on high-nutrient substrates, namely compost. However, the concentration of the organic constituents studied was higher in plants grown on clayish low-nutrient substrates in the presence of melilot. The content of resveratrol including that of its derivatives, resveratrolosid, piceatannol, piceid and astringin, was significantly higher in the presence of melilot on clay, loess and clayCS. Nitrogen supplied to knotweed by melilot was correlated with the ratio of resveratrol to resveratrol glucosides, indicating that knotweed bestowed some of its glucose production upon covering part of the energy demanded for nitrogen fixation by melilot's rhizobia, and that there is an exchange of organic substances between these two plant species. The three-year field experiment confirmed the ability of Reynoutria x bohemica to grow on vast coalmine spoil banks. The production of this species reached 2.6 t of dry mass per hectare.

Conclusions: Relationships between nitrogen, phosphorus, emodin, and belowground knotweed biomass belong to the most interesting results of this study. Compared with melilot absence, its presence increased the number of significant relationships by introducing those of resveratrol and its derivatives, and phosphorus and nitrogen. Knotweed phosphorus was predominantly taken up from the substrate and was negatively correlated with the content of resveratrol and resveratrol derivatives, while knotweed nitrogen was mainly supplied by melilot rhizobia and was positively correlated with the content of resveratrol and resveratrol derivatives.

Figure 1

Aboveground biomass (d.w.) of Reynoutria × bohemica grown in pots with various substrates based on miocene clay from coalmine spoil banks with (black columns) and without (open columns) Melilotus alba (significant differences are indicated by asterisks) in 2006 (a - left) and 2007 (b - right). ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd. Equal letters indicate non-significant differences between the substrates; lower case - without melilot, upper case - with melilot.

Figure 2

Belowground biomass (d.w.) of Reynoutria × bohemica grown in pots with various substrates based on miocene clay from coalmine spoil banks with (black columns) and without (open columns) Melilotus alba (significant differences are indicated by asterisks) in 2007. ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd. Equal letters indicate non-significant differences between the substrates; lower case - without melilot, upper case - with melilot.

Figure 3

Resveratrol content in Reynoutria × bohemica roots and rhizomes grown in pots with various substrates based on miocene clay from coalmine spoil banks with (black columns) and without (open columns) Melilotus alba (significant differences are indicated by asterisks) in 2006 (a - left) and 2007 (b - right). ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd. Equal letters indicate non-significant differences between the substrates; lower case - without melilot, upper case - with melilot.

Figure 4

Resveratrol contained in all its derivatives was measured in Reynoutria × bohemica roots and rhizomes grown in pots with various substrates based on miocene clay from coalmine spoil banks with (black columns) and without (open columns) Melilotus alba (significant differences are indicated by asterisks) in 2006 (a - left) and 2007 (b - right). ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd. Equal letters indicate non-significant differences between the substrates; lower case - without melilot, upper case - with melilot.

Figure 5

Emodin content in Reynoutria × bohemica roots and rhizomes grown in pots with various substrates based on miocene clay from coalmine spoil banks with (black columns) and without (open columns) Melilotus alba (significant differences are indicated by asterisks) in 2006 (a - left) and 2007 (b - right). ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd. Equal letters indicate non-significant differences between the substrates; lower case - without melilot, upper case - with melilot.

Figure 6

Nitrogen content in Reynoutria × bohemica roots and rhizomes grown in pots with various substrates based on miocene clay from coalmine spoil banks with (black columns) and without (open columns) Melilotus alba (significant differences are indicated by asterisks) in 2006 (a - left) and 2007 (b - right). ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd. Equal letters indicate non-significant differences between the substrates; lower case - without melilot, upper case - with melilot.

Figure 7

Phosphorus content in Reynoutria × bohemica roots and rhizomes grown in pots with various substrates based on miocene clay from coalmine spoil banks with (black columns) and without (open columns) Melilotus alba (significant differences are indicated by asterisks) in 2006 (a - left) and 2007 (b - right). ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd. Equal letters indicate non-significant differences between the substrates; lower case - without melilot, upper case - with melilot.

Figure 8

Mycorrhizal colonization F% of Reynoutria × bohemica roots grown with melilot (a - left) and aboveground biomass of Melilotus alba (b - right), in pots with various substrates based on miocene clay from coalmine spoil banks in 2006 and 2007. ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd. Equal letters within the same year indicate non-significant differences between the substrates.

Figure 9

Aboveground (black columns) and belowground (open columns) biomass (d.w.) of Reynoutria × bohemica grown in a spoil bank changed into arable field, from April 2006 (planted) to September 2008. Means ± S.E. indicated.

Figure 10

Stilbenes (resveratrol and resveratrol in its derivatives) in belowground biomass of R. × bohemica grown in a spoil bank changed into arable field from April 2006 (planted) to September 2008. Means ± S.E. indicated.

Figure 11

Diagram of relationships between root and rhizome Reynoutria × bohemica characteristics (resveratrol, stilbenes, emodin, phosphorus and nitrogen contents and biomass) measured in 2007, melilot biomass measured in preceding year and phosphorus and nitrogen contents in the substrate, all in the absence and presence of melilot, respectively. Correlation coefficients are shown in cases of relationships that were significant at P ≤ 0.05.

Figure 12

Relationship between the ratio of resveratrol contained in its aglycons (resveratrol and piceatannol) to its glucosides (astringines, piceid, resveratrolosid), and differences in nitrogen concentration in belowground biomass of Reynoutria × bohemica grown with Melilotus alba (measured values) and without melilot (average per substrate). ClayC = clay enriched with slow-release biofertilizer Conavit; ClayCS = clay enriched with Conavit and arbuscular-mycorrhizal product Symbivit, both produced by Symbiom Ltd.

Figure 13

A typical HPLC record of the stilbenes and emodin measured in knotweed rhizomes.