Harnessing eugenol as a substrate for production of aromatic compounds with recombinant strains of Amycolatopsis sp. HR167

To harness eugenol as cheap substrate for the biotechnological production of aromatic compounds, the vanillyl alcohol oxidase gene (vaoA) from Penicillium simplicissimum CBS 170.90 was cloned in an expression vector suitable for Gram-positive bacteria and expressed in the vanillin-tolerant Gram-positive strain Amycolatopsis sp. HR167. Recombinant strains harboring hybrid plasmid pRLE6SKvaom exhibited a specific vanillyl alcohol oxidase activity of 1.1U/g protein. Moreover, this strain had gained the ability to grow on eugenol as sole carbon source. The intermediates coniferyl alcohol, coniferyl aldehyde, ferulic acid, guajacol, and vanillic acid were detected as excreted compounds during growth on eugenol, whereas vanillin could only be detected in trace amounts. Resting cells of Amycolatopsis sp. HR167 (pRLE6SKvaom) produced coniferyl alcohol from eugenol with a maximum conversion rate of about 2.3 mmol/h/l of culture, and a maximum coniferyl alcohol concentration of 4.7 g/1 was obtained after 16 h biotransformation without further optimization. Beside coniferyl alcohol, traces of coniferyl aldehyde and ferulic acid were also detected.     

Quantitative analysis of phenolics in selected crop species and biological activity of these compounds evaluated by sensitivity of Echinochloa crus-galli

The purpose of this study was to determine the content of selected phenolic compounds in white mustard, buckwheat, spring barley, oat and rye grown under field conditions. Moreover, the allelopathic efficiency of these compounds was evaluated by sensitivity of Echinochloa crus-galli. The aromatic acids: trans-cinnamic, salicylic, ferulic, chlorogenic, p-hydroxybenzoic, protocatechuic, p-coumaric and vanillic were separated from crop plants by TLC and determined spectrophotometrically. Differences in concentrations of analysed compounds were observed for most of the examined plant species. The highest concentration was noticed for cinnamic acid and ranged from 360 μg·g⁻¹ DW in rye to 2770 μg·g⁻¹ DW in spring barley. The relatively high concentration was noticed for ferulic acid (from 73.8 μg·g⁻¹ DW in buckwheat to 1046 μg·g⁻¹ DW in spring barley) and p-coumaric acid (from 50 μg·g⁻¹ DW in oat to 1499 μg·g⁻¹ DW in buckwheat). The observed differences in the phenolics content between two successive vegetation seasons can reflect the effect of abiotic and biotic environmental factors on the phenolics level in studied plants. In the greenhouse experiment the effect of particular compounds on the growth of Echinochloa crus-galli was also studied. It has been found that the examined phenolics, and especially trans-cinnamic acid and mixture of phenolic compounds, significantly inhibit the growth of Echinochloa crus-galli. The obtained results may contribute to the explanation of the biological activity of some phenolic compounds.     

Allelopathic Interactions Involving Phenolic Acids

A major concern regarding allelopathic interactions involving phenolic acids in no-till systems pertains to the fact that concentrations of individual phenolic acids recoverable from field soils are well below levels required for inhibition of germination and seedling growth in laboratory bioassays. Field soils contain a variety of phenolic acids as well as other toxic and nontoxic organic compounds that are available to interact with seeds and roots; whereas in laboratory bioassays, with few exceptions, single phenolic acids have been tested. Studies of mixtures of phenolic acids and other toxic (e.g., methionine) and nontoxic (e.g., glucose) organic compounds in laboratory bioassays indicate that the action of a single phenolic acid is not representative of the actions of such mixtures. Specifically, as the number of phenolic acids added to soil increased, concentrations of the individual phenolic acids required to bring about a growth inhibition declined. The addition of other organic compounds (e.g., glucose, methionine) to the soil also reduced the concentration of a phenolic acid (e.g., p-coumaric acid) required for growth inhibition. These results support the hypothesis that in the field mixtures of phenolic acids and other organic compounds can cause inhibitory effects even though the concentrations of individual compounds are well below their inhibitory levels.     

酚酸类化感自毒物质对枸杞种子萌发的抑制作用

酚酸类化感自毒物质是导致农作物连作障碍的重要诱因之一。枸杞作为多年生木本植物,连作障碍突出,成因复杂。为揭示酚酸类物质对枸杞的化感自毒作用,研究了22种常见酚酸对枸杞种子萌发的抑制作用,并运用比较分子场分析方法(CoMFA)进行构效关系研究。结果发现,酚酸化合物对枸杞种子萌发作用的半抑制浓度IC₅₀范围为39.94—115.97 mg/L;CoMFA结果表明,运用立体场和静电场能较好解释该类化合物对枸杞种子萌发的化感自毒特征,苯环1/2/3/4位具有大体积、1位具有强负电性取代基的酚酸化合物,对枸杞种子萌发具有较强抑制作用。研究结果可为客观评价酚酸类化合物对枸杞的化感自毒作用提供基础数据支撑。     

Antibiotic potential of plant growth promoting rhizobacteria (PGPR) against Sclerotium rolfsii

High performance liquid chromatographic (HPLC) analysis of culture filtrates of plant growth promoting rhizobacteria (PGPR) and medium of inhibitory zone of interaction of Sclerotium rolfsii with PGPR, viz. Pseudomonas aeruginosa, Pseudomonas fluorescens 4, Pseudomonas fluorescens 4 (new) and Pseudomonas sp. varied from sample to sample. In all the culture filtrates of PGPRs, P. aeruginosa had nine phenolic acids in which ferulic acid (14.52 μg/ml) was maximum followed by other phenolic acids. However, the culture filtrates of P. fluorescens 4 had six phenolic acids with maximum ferulic acid (20.54 μg/ml) followed by indole acetic acid (IAA), caffeic, salicylic, o-coumeric acid and cinnamic acids. However, P. fluorescens 4 culture filtrate had seven phenolic acids in which salicylic acid was maximum (18.03 μg) followed by IAA, caffeic, vanillic, ferulic, o-coumeric and cinnamic acids. Pseudomonas sp. also showed eight phenolic acids where caffeic acid (2.75 μg) was maximum followed by trace amounts of ferulic, salicylic, IAA, vanillic, cinnamic, o-coumeric and tannic acids. The analysis of antibiosis zone of PGPRs showed fairly rich phenolic acids. A total of nine phenolic acids were detected in which caffeic acid was maximum (29.14 μg/g) followed by gallic (17.64 μg/g) and vanillic (3.52 μg/g) acids but others were in traces. In P. aeruginosa, antibiosis zone had seven phenolic acids where IAA was maximum (3.48 μg/g) followed by o-coumeric acid (2.08 μg/g), others were in traces. The medium of antibiosis zone of P. fluorescens 4 and P. fluorescens 4 new had eight phenolic acids in which IAA was maximum with other phenolic acids in traces.     

Batatasin‐III and the allelopathic capacity of Empetrum nigrum

Batatasin‐III (3,3‐dihydroxy‐5‐methoxybibenzyl) is a phenolic compound associated with the allelopathic effect of the evergreen dwarf shrub Empetrum nigrum, and has been referred to as the causal factor for the species being successful in dominating extensive ecosystems. Yet, only a few plant species have been tested for their response to batatasin‐III, and little is known about whether environmental factors modify this allelopathic effect. In this study, we tested the inhibitory effect of purified batatasin‐III through bioassays on 24 vascular plant species and, for certain species, we tested if this effect depended on growth substrate (mineral vs organic substrate), pH, and fertilization. Moreover, we tested if batatasin‐III predicted the allelopathic effect of E. nigrum by analyzing the inhibitory effect of E. nigrum leaves and humus in relation to their batatasin‐III content. Our results confirmed batatasin‐III as a stable compound capable of inhibiting germination and/or mean root elongation in all of the tested species, but this effect was modified by growth substrate. Surprisingly, the measured batatasin‐III content of E. nigrum leaves (mean value 19.7 ± 10.8 (SE) mg g^?1) and humus (mean value of 1 ± 1.5 (SE) μg g^?1) did not predict the inhibitory effect on mean root elongation. Although batatasin‐III was found to be phytotoxic to all the tested species, we conclude that this substance alone should not be used as a proxy for the allelopathic effect of E. nigrum.     

Potential of Rhodococcus strains for biotechnological vanillin production from ferulic acid and eugenol

The potential of two Rhodococcus strains for biotechnological vanillin production from ferulic acid and eugenol was investigated. Genome sequence data of Rhodococcus sp. I24 suggested a coenzyme A-dependent, non-β-oxidative pathway for ferulic acid bioconversion, which involves feruloyl–CoA synthetase (Fcs), enoyl–CoA hydratase/aldolase (Ech), and vanillin dehydrogenase (Vdh). This pathway was proven for Rhodococcus opacus PD630 by physiological characterization of knockout mutants. However, expression and functional characterization of corresponding structural genes from I24 suggested that degradation of ferulic acid in this strain proceeds via a β-oxidative pathway. The vanillin precursor eugenol facilitated growth of I24 but not of PD630. Coniferyl aldehyde was an intermediate of eugenol degradation by I24. Since the genome sequence of I24 is devoid of eugenol hydroxylase homologous genes (ehyAB), eugenol bioconversion is most probably initiated by a new step in this bacterium. To establish eugenol bioconversion in PD630, the vanillyl alcohol oxidase gene (vaoA) from Penicillium simplicissimum CBS 170.90 was expressed in PD630 together with coniferyl alcohol dehydrogenase (calA) and coniferyl aldehyde dehydrogenase (calB) genes from Pseudomonas sp. HR199. The recombinant strain converted eugenol to ferulic acid. The obtained data suggest that genetically engineered strains of I24 and PD630 are suitable candidates for vanillin production from eugenol.     

Biological activity of metabolites of the herb <Emphasis Type="Italic">Kalanchoe diagremontania</Emphasis> (Hamet de la Bathie) Jacobs et Perr

In this study we investigated the hemolytic, antimicrobial, and phytoregulatory activity of various classes of lipids (triacylglycerols, free fatty acids (FFA), the glyceroglycolipids monogalactosyl diacylglycerol (MGDG), sulfoquinovosyl diacylglycerol (SQDG)), sterols, all of them were obtained from the medical herb Kalanchoe diagremontiana, and also pigments, phenolic compounds (FC), polysaccharides, and ethanol extract (EE) of the herbal. It was established that EE, FC, FFA, and sterols display pH-dependent membranothropic activity. FFA showed antimicrobial activity and stimulated growth of buckwheat stalk sprouts. K. diagremontiana glyceroglycolopids did not display expressed biological activity. Caroteniods displayed pH-independent membranothopic action and antibacterial activity. Chlorophylls displayed antimicrobial action, but did not influence erythrocytes and buckwheat sprouts. Polysaccharides acted against the microorganisms Safale S-04, Candida albicans, Fusarium oxysperum and buckwheat sprouts.     

Time course study on accumulation of cell wall-bound phenolics and activities of defense enzymes in tomato roots in relation to <Emphasis Type="Italic">Fusarium</Emphasis> wilt

Major cell wall-bound phenolic compounds were detected and identified in roots of tomato at different stages of growth. Alkaline hydrolysis of the cell wall material of the root tissues yielded ferulic acid as the major bulk of the phenolic compounds. Other phenolic compounds identified were 4-hydroxybenzoic acid, vanillic acid, 4-hydroxybenzaldehyde, vanillin and 4-coumaric acid. All the six phenolic acids were higher in very early stage of plant growth. Ferulic acid, 4-hydroxybenzoic acid and 4-coumaric acid exhibited a decreasing trend up to 60 days and then the content of these phenolic acids increased somewhat steadily towards the later stage of growth. Total phenolics, phenylalanine ammonia-lyase (PAL) activity and peroxidase (POD) activity were in tandem match with the occurrence pattern of the phenolic acids. Ferulic acid showed highest antifungal activity against tomato wilt pathogen Fusarium oxysporum f. sp. lycopersici. The results of this study may be interpreted to seek an explanation for high susceptibility of tomato plants at flowering stage to Fusarium wilt. It may also be concluded that greater amounts of ferulic acid in combination with other phenolics and higher level of PAL and POD activities after 60 days of growth may have a role in imparting resistance against Fusarium wilt at a late stage of plant growth.     

ALLELOPATHY IN THE FIRST STAGES OF SECONDARY SUCCESSION ON THE PIEDMONT OF NEW JERSEY

Ambrosia artemisiifolia (ragweed) is a dominant species in the first year of old field succession but rarely persists for more than two years. Ragweed and Raphanus raphanistrum (wild radish), also an early invader, failed to become re-established in plots cleared of second stage perennial vegetation (dominated by Aster pilosus), despite the large number of seeds of these primary invaders present in the soil. Edaphic experiments revealed that this pattern of succession was not due to mineral or physical properties of the soil. Field soil from the second stage of succession inhibited the growth and germination of ragweed and wild radish while soil from the first stage had no effect. Inhibitory volatile materials from ragweed were not detected. However, root exudate of ragweed, and shoot extracts of ragweed and aster inhibited the germination and growth of early invaders of abandoned fields. These results indicate that the vegetational change from the first to the second stage of succession may be mediated at least partially by an allelopathic response. Chromatography and bioassay techniques revealed the inhibitory compounds to be phenolic acids, including caffeic and chlorogenic acids.     

Allelopathic Effects of Castanea henryi Aqueous Extracts on the Growth and Physiology of Brassica pekinensis and Zea mays

The present study investigated the allelopathic effects of aqueous extracts of Castanea henryi litter on the growth and physiological responses of Brassica pekinensis and Zea mays. Treatment with high concentrations of leaf extract (0.05 g/ml for B. pekinensis and 0.10 g/ml for Z. mays) significantly increased malonaldehyde content and reduced seed germination, seedling growth, chlorophyll content, and the activity levels of antioxidant enzymes. These effects generally increased with increasing extract concentration. However, in Z. mays, low extract concentrations actually promoted seed germination, shoot growth, chlorophyll content, and antioxidant enzyme activity. The allelopathic effects of the various C. henryi extracts decreased as follows: leaf extract > twig extract > shell extract. Eleven potential allelochemicals including rutin, quercetin, luteolin, procyanidin A2, kaempferol, allantoin, propionic acid, salicylic acid, jasmonic acid, methylmalonic acid, and gentisic acid were identified in the leaves of C. henryi which were linked to the strongest allelopathic effects. These findings suggest that the allelopathic effects of C. henryi differ depending on receptor plant species, and that leaves are the most allelopathic litter in C. henryi.     

Potential allelopathic effect of <Emphasis Type="Italic">Eucalyptus</Emphasis><Emphasis Type="Italic">grandis</Emphasis> across a range of plantation ages

Allelopathy of the eucalypt has been considered as an important mechanism for the biodiversity reduction in the eucalypt plantation. To understand the allelopathic potential of the eucalypt (Eucalyptus grandis) roots and rhizosphere soil along a chronosequence (2, 4, 6, 8, 10 years), the germination and growth characteristics of three plant species (Raphanus sativus, Phaseolus aureus, and Lolium perenne) growing nearby or beneath the eucalypt plantations were measured. The results showed that aqueous extract of E. grandis root suppressed the germination and early seedling growth of the target plants. The younger E. grandis exhibited a comparatively stronger allelopathic potential. The highest dose root extracts from 4 years old E. grandis showed the strongest inhibitory effects on the germination rates of the target species, the inhibitory rates were about 48, 51.2, and 56.56% for R. sativus, P. aureus, and L. perenne, respectively. However, present biotests of rhizosphere soils from 6, 8, and 10-year-old plantations exhibited a remarkable stimulative effect on L. perenne, which indicated that the soil might neutralize or dilute allelopathic agents with the increase of plantation age. In addition, according to GC–MS analysis, more allelopathic potential compounds were found in the rhizosphere soil and roots of younger E. grandis plantation. Moreover, more allelochemicals were obtained from soil than from roots. The allelopathic compounds in roots and rhizosphere soil may play important roles in allelopathy of E. grandis plantation. More attention should be paid to the younger E. grandis plantations for the relative higher allelopathic effects.     

Allelopathic activity of different plant parts of <Emphasis Type="Italic">Peganum harmala</Emphasis> L. and identification of their growth inhibitors substances

This study was conducted to evaluate the inhibitory potential of P. harmala leaf, stem and root extract on germination and growth of Avena fatua L. and Convolvulus arvensis L., as well as identification of the phytotoxic substances responsible for this activity. According to our results, the degree of toxicity of different P. harmala plant parts can be arranged in the following order: leaves > stems > roots. The two test species differed in their sensitivity to P. harmala extracts. Inhibitory effect on shoot length and seedling dry weight was more pronounced in C. arvensis, whereas higher reduction in germination, root length and total chlorophyll content occurred in A. fatua. A significant amount of water-soluble phenolic acids were found in P. harmala plant extracts. Total phenolic acids content was higher in leaf extracts when compared to that of stem or root extracts. Seven phenolic acids including gallic acid, vanillic acid, 4-hydroxybenzoic acid, 3,4-dihydroxybenzoic acid, caffeic acid, syringic acid and ferulic acid were found in P. harmala leaf extracts. On the other hand, we identified four phenolic acids from stem (galllic acid, vanillic acid, 3,4-dihydroxybenzoic acid and caffeic acid) and root (galllic acid, 4-hydroxybenzoic acid, syringic acid and cinnamic acid) extracts. The greater number of growth inhibitors detected in the leaves might explain the stronger inhibitory activity. Overall, our results suggest that P. harmala might be used as a natural herbicide for weed control and consequently reduce dependence on synthetic herbicides.     

Growth strategy,phylogeny and stoichiometry determine the allelopathic potential of native and non‐native plants

Secondary compounds can contribute to the success of non‐native plant species if they reduce damage by native herbivores or inhibit the growth of native plant competitors. However, there is opposing evidence on whether the secondary compounds of non‐native plant species are stronger than those of natives. This may be explained by other factors, besides plant origin, that affect the potential of plant secondary compounds. We tested how plant origin, phylogeny, growth strategy and stoichiometry affected the allelopathic potential of 34 aquatic plants. The allelopathic potential was quantified using bioassays with the cyanobacterium Dolichospermum flos‐aquae. The allelopathic potential showed a strong phylogenetic signal, but was similar for native and non‐native species. Growth strategy was important, and emergent plants had twice the allelopathic potential as compared to submerged plants. Furthermore, the allelopathic potential was positively correlated to the foliar carbon‐to‐phosphorus (C:P) and total phenolic content. We conclude that eudicot plant species with an emergent growth strategy and high plant C:P ratio exhibit a high allelopathic potential. Unless non‐native plant species match this profile, they generally have a similar allelopathic potential as natives.     

Biotransformation of Eugenol to Ferulic Acid by a Recombinant Strain of Ralstonia eutropha H16

The gene loci ehyAB, calA, and calB, encoding eugenol hydroxylase, coniferyl alcohol dehydrogenase, and coniferyl aldehyde dehydrogenase, respectively, which are involved in the first steps of eugenol catabolism in Pseudomonas sp. strain HR199, were amplified by PCR and combined to construct a catabolic gene cassette. This gene cassette was cloned in the newly designed broad-host-range vector pBBR1-JO2 (pBBR1-JO2ehyABcalAcalB) and transferred to Ralstonia eutropha H16. A recombinant strain of R. eutropha H16 harboring this plasmid expressed functionally active eugenol hydroxylase, coniferyl alcohol dehydrogenase, and coniferyl aldehyde dehydrogenase. Cells of R. eutropha H16(pBBR1-JO2ehyABcalAcalB) from the late-exponential growth phase were used as biocatalysts for the biotransformation of eugenol to ferulic acid. A maximum conversion rate of 2.9 mmol of eugenol per h per liter of culture was achieved with a yield of 93.8 mol% of ferulic acid from eugenol within 20 h, without further optimization.     

Infection with the fungal endophyte Epichloë festucae may alter the allelopathic potential of red fescue

Red fescue (Festuca rubra) is a perennial grass used as both forage and turfgrass. Asymptomatic plants of this species are systemically infected by the fungal endophyte Epichloë festucae, which has a beneficial effect on the infected plants. The aim of this study was to determine the effect of the endophyte Epichloë festucae on the allelopathic potential of F. rubra against four associated pasture species that are also considered as weeds in lawns, Trifolium pratense, Trifolium repens, Lotus corniculatus and Plantago lanceolata. Two experiments were designed to evaluate the allelopathic effect of extracts from the roots and leaves of endophyte‐infected (E+) and non‐infected (E?) plants on the germination and seedling growth of the four target species. Regardless of the endophyte status of the host plant, leaf extracts elicited a stronger reduction in germination and seedling growth than root extracts. Extracts from E+ plants reduced the speed of germination index of Trifolium spp. to a greater extent than those from E? plants. Radicle length of the target species was the parameter most affected by the presence of the endophyte in F. rubra. Root extracts from E+ plants had a greater inhibitory effect on the radicle growth of the target species than did root extracts from E? plants. A greater concentration in total phenolic compounds was found in the roots of E+ plants than of E?; however, this difference was not observed in the leaves. Thus, the allelopathic potential of F. rubra is altered in infected plants.     

Metabolism of phenolic compounds in <Emphasis Type="Italic">Vitis riparia</Emphasis> seeds during stratification and during germination under optimal and low temperature stress conditions

We studied the alterations in phenolic compounds in grape seeds during their stratification and germination under optimal conditions (+25 °C) and at low temperature (+10 °C). Biological materials in the study were seeds of Vitis riparia. Phenolic compounds were extracted from defatted seeds using 80 % methanol or 80 % acetone. The content of total phenolics was determined with the Folin-Ciocalteau reagent, while the content of tannins was determined by vanillin assay and the protein (BSA) precipitation method. The RP-HPLC method was used to determine phenolic compounds (phenolic acids, catechins) in the extracts. High amounts of tannins, catechins, gallic acid and lesser amounts of p-coumaric acid were found in the seeds. The content of total phenolics in acetone extracts was higher than that obtained using methanol. The amounts of phenolic acids and tannins found in V. riparia seeds after stratification were much lower. It may confirm a possible role of these compounds in dormancy of V. riparia seeds. After 72 h of low temperature treatment, inhibition of grape root growth and biochemical changes in seeds were detected. The chilling stimulated increased accumulation of some phenolic compounds (free gallic acid and catechins) in the seeds. These substances can protect plants against some abiotic stressors.     

Identification and comparative analysis of microRNAs in barnyardgrass (Echinochloa crus‐galli) in response to rice allelopathy

Rice allelopathy is a hot topic in the field of allelopathy, and behaviour of donor allelopathic rice has been well documented. However, few study addresses response of receiver barnyardgrass (BYG). We found that expression of miRNAs relevant to plant hormone signal transduction, nucleotide excision repair and the peroxisome proliferator‐activated receptor and p53 signalling pathways was enhanced in BYG co‐cultured with the allelopathic rice cultivar PI312777, the expression levels of these miRNAs in BYG plants were positively correlated with allelopathic potential of the co‐cultured rice varieties. Treatment of BYG plants with rice‐produced phenolic acids also increased miRNA expression in BYG, while treatment with rice‐produced terpenoids had no obvious effect on miRNA expression. In the hydroponic system, the largest number of Myxococcus sp. was found in the growth medium containing rice with the highest allelopathic potential. The addition of phenolic acids in the hydroponic medium also increased the number of Myxococcus sp. More interestingly, inoculation with Myxococcus xanthus significantly increased miRNA expression in the treated BYG. Jointed treatments of ferulic acid and M. xanthus led to strongest growth inhibition of BYG. The results suggest that there exist involvement of Myxococcus sp. and mediation of miRNA expression in rice allelopathy against BYG.     

Silybum marianum cell cultures stably transformed with Vitis vinifera stilbene synthase accumulate t‐resveratrol in the extracellular medium after elicitation with methyl jasmonate or methylated β‐cyclodextrins

The growing demand for t‐resveratrol for industrial uses has generated considerable interest in its production. Heterologous resveratrol production in plant cell suspensions, apart from requiring the introduction of only one or two genes, has the advantage of high biomass yield and a short cultivation time, and thus could be an option for large‐scale production. Silybum marianum is the source of the flavonolignan silymarin. Phenylpropanoid synthesis in cultures of this species can be activated by elicitation with methyl jasmonate and methylated β‐cyclodextrins, with products of the pathway (coniferyl alcohol and some isomers of the silymarin complex) being released into the medium. Given that stilbene synthase shares the same key precursors involved in flavonoid and /or monolignol biosynthesis, we explored the potential of metabolically engineered S. marianum cultures for t‐resveratrol production. Cell suspensions were stably transformed with Vitis vinifera stilbene synthase 3 and the expression of the transgene led to extracellular t‐resveratrol accumulation at the level of milligrams per litre under elicitation. Resveratrol synthesis occurred at the expense of coniferyl alcohol. Production of silymarin was less affected in the transgenic cultures, since the flavonoid pathway is limiting for its synthesis, due to the preferred supply of precursors for the monolignol branch. The fact that the expressed STS gene took excessively produced precursors of non‐bioactive compounds (coniferyl alcohol), while keeping the metabolic flow for target secondary compounds (i.e. silymarin) unaltered, opens a way to extend the applications of plant cell cultures for the simultaneous production of both constitutive and foreign valuable metabolites.     

Metabolic Regulation of Floral Scent in Petunia axillaris Lines: Biosynthetic Relationship between Dihydroconiferyl Acetate and iso-Eugenol

Aromatic scent-related compounds in flowers were comprehensively analyzed by high-performance liquid chromatography (HPLC) based on their absorption spectra to understand regulation of metabolism leading to floral scent diversity in Petunia axillaris lines. An unknown compound occurring at similar levels to scent compounds in some plant lines was identified to be dihydroconiferyl acetate. Based on the structure, dihydroconiferyl acetate is likely to be a biosynthetically closely related compound to aromatic scent compounds, especially iso-eugenol. Similar time-course changes of the concentrations suggest that the metabolism of dihydroconiferyl acetate is underlaid by the similar regulation to aromatic scent compounds. Dihydroconiferyl acetate and iso-eugenol occurred almost exclusively, implying that metabolism of the common precursors to each compound is selectively regulated in these plants. The branching of the biosynthetic pathway into dihydroconiferyl acetate and iso-eugenol is probably one of regulatory steps leading to scent diversity in P. axillaris lines.