Diversity Of Plant Secondary Metabolites Research Articles

Comparative Analysis of Polyphenolic Compounds in Different Amaranthus Species: Influence of Genotypes and Harvesting Year.

Amaranth is a nutritionally valuable crop, as it contains phenolic acids and flavonoids, yielding diverse plant secondary metabolites (PSMs) like phytosterol, tocopherols, and carotenoids. This study explored the variations in the contents of seventeen polyphenolic compounds within the leaves of one hundred twenty Amaranthus accessions representing nine Amaranthus species. The investigation entailed the analysis of phenolic content across nine Amaranthus species, specifically A. hypochondriacus, A. cruentus, A. caudatus, A. tricolor, A. dubius, A. blitum, A. crispus, A. hybridus, and A. viridis, utilizing ultra performance liquid chromatography with photodiode array detection (UPLC-PDA). The results revealed significant differences in polyphenolic compounds among accessions in which rutin content was predominant in all Amaranthus species in both 2018 and 2019. Among the nine Amaranthus species, the rutin content ranged from 95.72 ± 199.17 μg g-1 (A. dubius) to 1485.09 ± 679.51 μg g-1 (A. viridis) in 2018 and from 821.59 ± 709.95 μg g-1 (A. tricolor) to 3166.52 ± 1317.38 μg g-1 (A. hypochondriacus) in 2019. Correlation analysis revealed, significant positive correlations between rutin and kaempferol-3-O-β-rutinoside (r = 0.93), benzoic acid and ferulic acid (r = 0.76), and benzoic acid and kaempferol-3-O-β-rutinoside (r = 0.76), whereas gallic acid showed consistently negative correlations with each of the 16 phenolic compounds. Wide variations were identified among accessions and between plants grown in the two years. The nine species and one hundred twenty Amaranthus accessions were clustered into six groups based on their seventeen phenolic compounds in each year. These findings contribute to expanding our understanding of the phytochemical traits of accessions within nine Amaranthus species, which serve as valuable resources for Amaranthus component breeding and functional material development.

Chemical Composition of the Essential Oils of the Flowers Asphodelus aestivus Brot. Grown Wild in Jordan

Recent technological developments and methodological advances in GC-MS have become a common tool for investigating the quantity, quality, and chemical diversity of plant secondary metabolites. The flower parts of Asphodelus aestivus Brot. were studied, leading to the isolation and identification of various secondary metabolites, primarily essential oils: alcohol (26.9%); aldehyde (23%); alkanes, acetate derivatives, and aliphatic derivatives (19.2%); ketones (7.7%); and epoxides (3.8%). The principal oil components were 18.79% vetocitral C (trans), 17.27% hexadecyl acetate, 14.5% hexanal (2E), and 9.6% sabinene hydrate (trans). The identification of oil components was performed by matching their spectra with the mass spectra data bank.

Phytochemical diversity impacts herbivory in a tropical rainforest tree community.

Metabolomics provides an unprecedented window into diverse plant secondary metabolites that represent a potentially critical niche dimension in tropical forests underlying species coexistence. Here, we used untargeted metabolomics to evaluate chemical composition of 358 tree species and its relationship with phylogeny and variation in light environment, soil nutrients, and insect herbivore leaf damage in a tropical rainforest plot. We report no phylogenetic signal in most compound classes, indicating rapid diversification in tree metabolomes. We found that locally co-occurring species were more chemically dissimilar than random and that local chemical dispersion and metabolite diversity were associated with lower herbivory, especially that of specialist insect herbivores. Our results highlight the role of secondary metabolites in mediating plant-herbivore interactions and their potential to facilitate niche differentiation in a manner that contributes to species coexistence. Furthermore, our findings suggest that specialist herbivore pressure is an important mechanism promoting phytochemical diversity in tropical forests.

Computational insights into the role of structurally diverse plant secondary metabolites as inhibitors against Imidazole Glycerol Phosphate Dehydratase of Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) is one of the major causes of death worldwide and there is a pressing need for the development of novel drug leads. The Imidazole Glycerol Phosphate Dehydratase (IGPD) of Mtb is one of the key enzymes in the histidine biosynthesis pathway and has been recognized as the potentially underexploited drug target for anti-tuberculosis treatment. In the present study, 6063 structurally diverse plant secondary metabolites (PSM) were screened for their efficiency in inhibiting the catalytic activity of IGPD through molecular docking. The top 150 PSMs with the lowest binding energy represent the chemical classes, including Tannins (34%), Flavonoid Glycosides (14%), Terpene Glycosides (10%), Steroid Lactones (9.3%), Flavonoids (6.6%), Steroidal Glycosides (4.6%), etc. Bismahanine, Ashwagandhanolide, and Daurisoline form stable IGPD-inhibitor complexes with binding free energies of −291.3 ± 16.5, −279.0 ± 25.0, and −279.8 ± 17.6 KJ/mol, respectively, as determined by molecular dynamics simulations. These PSM demonstrated strong H-bond interactions with the amino acid residues Ile279, Arg281, and Lys276 in the catalytic region of IGPD, as revealed by structural snapshots. On the basis of our findings, these three PSM could be considered as possible leads against IGPD and should be explored in vitro and in vivo. Communicated by Ramaswamy H. Sarma

REGULATION AND METABOLIC ENGINEERING OF THE GENERAL PHENYLPROPANOID PATHWAY IN RESPONSE TO STRESS IN PLANTS

The diversity of plant secondary metabolites is based on the phenylpropanoid pathway. As part of its functioning, phenylalanine is exposed to enzymes that convert this amino acid into phenolic compounds. The initial steps are catalyzed by phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), and 4-coumaroyl-CoA ligase (4CL), which are part of the general phenylpropanoid pathway. The products of the general phenylpropanoid pathway are associated with the growth and development of plants, responses to the external stimuli, signaling and protective functions. The study of the bio-chemical and molecular bases of the biosynthesis of phenylpropanoids is important, since their functioning underlies the understanding of the molecu-lar mechanisms of plant adaptation to external factors such as drought, salinity, lack of mineral nutrition components, and pathogens interaction. This review considers the relationship of the genes of the general phenylpropanoid pathway with the impact of pathogens and abiotic factors, as well as their genetic and metabolic engineering. PAL, C4H, and 4CH genes of many plant species usually represent genes families encoding several enzymes isoforms. Increased levels of expression correlate with increased production of phenylpropanoids, and enzymes activity varies with developmental stage, cell differentiation, and environmental exposure. Thus, PAL, C4H, and 4CH are one of the key enzymes involved in plant responses to stress. For example, PAL is involved in signaling in re-sponse to pathogens. It is also directly related to lignin biosynthesis, which strengthens cell walls and has antimicrobial activity. Genes of the general phenylpropanoid pathway often become objects of genetic and metabolic engineering. These manipulations can be aimed to increase the biosynthesis of flavonoids and other secondary metabolites, as well as to obtain genotypes that are resistant to biotic and abiotic factors

Plant scent and plant–insect interactions—Review and outlook from a macroevolutionary perspective

AbstractThe astonishing diversity of plants and insects and their entangled interactions are cornerstones in terrestrial ecosystems. Co‐occurring with species diversity is the diversity of plant secondary metabolites (PSMs). So far, their estimated number is more than 200 000 compounds, which are not directly involved in plant growth and development but play important roles in helping plants handle their environment including the mediation of plant–insect interactions. Here, we use plant volatile organic compounds (VOCs), a key olfactory communication channel that mediates plant–insect interactions, as a showcase of PSMs. In spite of the cumulative knowledge of the functional, ecological, and microevolutionary roles of VOCs, we still lack a macroevolutionary understanding of how they evolved with plant–insect interactions and contributed to species diversity throughout the long coevolutionary history of plants and insects. We first review the literature to summarize the current state‐of‐the‐art research on this topic. We then present various relevant types of phylogenetic methods suitable to answer macroevolutionary questions on plant VOCs and suggest future directions for employing phylogenetic approaches in studying plant VOCs and plant–insect interactions. Overall, we found that current studies in this field are still very limited in their macroevolutionary perspective. Nevertheless, with the fast‐growing development of metabolome analysis techniques and phylogenetic methods, it is becoming increasingly feasible to integrate the advances of these two areas. We highlight promising approaches to generate new testable hypotheses and gain a mechanistic understanding of the macroevolutionary roles of chemical communication in plant–insect interactions.

Volatile Organic Compounds as Mediators of Plant Communication and Adaptation to Climate Change.

Plant volatile organic compounds are the most abundant and structurally diverse plant secondary metabolites. They play a key role in plant lifespan via direct and indirect plant defenses, attracting pollinators, and mediating various interactions between plants and their environment. The ecological diversity and context-dependence of plant-plant communication driven by volatiles are crucial elements that influence plant performance in different habitats. Plant volatiles are also valued for their multiple applications in the food, flavor, pharmaceutical, and cosmetics industries. In the current review, we summarize recent advances that have elucidated the functions of plant volatile organic compounds as mediators of plant interaction at community and individual levels, highlighting the complexities of plant receiver feedback to various signals and cues. This review emphasizes volatile terpenoids, the most abundant class of plant volatile organic compounds, highlighting their role in plant adaptability to global climate change and stress-response pathways that are integral to plant growth and survival. Lastly, we identify research gaps and suggest future research directions. This article is protected by copyright. All rights reserved.

Potential lipolytic regulators derived from natural products as effective approaches to treat obesity.

Epidemic obesity is contributing to increases in the prevalence of obesity-related metabolic diseases and has, therefore, become an important public health problem. Adipose tissue is a vital energy storage organ that regulates whole-body energy metabolism. Triglyceride degradation in adipocytes is called lipolysis. It is closely tied to obesity and the metabolic disorders associated with it. Various natural products such as flavonoids, alkaloids, and terpenoids regulate lipolysis and can promote weight loss or improve obesity-related metabolic conditions. It is important to identify the specific secondary metabolites that are most effective at reducing weight and the health risks associated with obesity and lipolysis regulation. The aims of this review were to identify, categorize, and clarify the modes of action of a wide diversity of plant secondary metabolites that have demonstrated prophylactic and therapeutic efficacy against obesity by regulating lipolysis. The present review explores the regulatory mechanisms of lipolysis and summarizes the effects and modes of action of various natural products on this process. We propose that the discovery and development of natural product-based lipolysis regulators could diminish the risks associated with obesity and certain metabolic conditions.

Voluntary consumption of Lantana camara L. when browsing the heterogeneous vegetation of tropical forests: A goats’ perspective

Although Lantana camara L. (Verbenaceae) consumption has been considered a health hazard for many livestock species, its true feeding value for ruminants should not be disregarded. The present work describes the feeding behavior of 36 Criollo goats towards L. camara in the heterogeneous grazing/browsing context of the low deciduous forest (LDF). An observational methodology focusing on the bite-scale (frequency, size, height, and part of plant) was implemented for eight weeks. Almost all experimental goats (35 of 36) consumed freely and consistently L. camara, including bites (up to 30 cm) directed to leaves, stems, flowers, and fruits. Goats possess anatomical, physiological, and behavioral strategies that help them select a wide range of plant species, and the nutritional context could play an important role on their dietary decisions. Considering the heterogeneity present in the LDF, mixing plant species, avoiding large intake of specific feed resources, and maintaining a constant intake of diverse plant secondary metabolites seems to be the foundation of goat feeding behavior. In conclusion, voluntary L. camara consumption by Criollo goats in the LDF is constant, albeit small. Future works should focus on its phytochemical profile and relationships with the nutritional and health status of goats.

Current Understanding of Modes of Action of Multicomponent Bioactive Phytochemicals: Potential for Nutraceuticals and Antimicrobials.

Plants produce a diversity of plant secondary metabolites (PSMs), which function as defense chemicals against herbivores and microorganisms but also as signal compounds. An individual plant produces and accumulates mixtures of PSMs with different structural features using different biosynthetic pathways. Almost all PSMs exert one or several biological activities that can be useful for nutrition and health. This review discusses the modes of action of PSMs alone and in combinations. In a mixture, most individual PSMs can modulate different molecular targets; they are thus multitarget drugs. In an extract with many multitarget chemicals, additive and synergistic effects occur. Experiments with the model system Caenorhabditis elegans show that polyphenols and carotenoids can function as powerful antioxidative and longevity-promoting PSMs. PSMs of food plants and spices often exhibit antioxidant, anti-inflammatory, and antimicrobial properties, which can be beneficial for health and the prevention of diseases. Some extracts from food plants and spices with bioactive PSMs have potential for nutraceuticals and antimicrobials.

Research progress on plant biosynthetic gene cluster

<p indent=0mm>Plants produce a wide variety of structurally and functionally diverse metabolites, many of which play an indispensable role in plant growth and development and in the response to abiotic and biotic stresses. In recent years, plant biosynthetic gene clustering has attracted extensive attention from scientists, and the rapid development of multi-omics technologies such as genome, transcriptome and metabolome makes it possible to efficiently mine plant metabolic gene clusters. Further, the studies of plant biosynthetic gene clusters will provide new insights to explore and utilize the diversity of plant secondary metabolites. Here, we not only systematically summarize the structural characteristics and regulatory models of plant biosynthetic gene clusters, but also review the application of new technologies in the discovery and identification of biosynthetic gene clusters.

In silico studies evidenced the role of structurally diverse plant secondary metabolites in reducing SARS-CoV-2 pathogenesis

Plants are endowed with a large pool of structurally diverse small molecules known as secondary metabolites. The present study aims to virtually screen these plant secondary metabolites (PSM) for their possible anti-SARS-CoV-2 properties targeting four proteins/ enzymes which govern viral pathogenesis. Results of molecular docking with 4,704 ligands against four target proteins, and data analysis revealed a unique pattern of structurally similar PSM interacting with the target proteins. Among the top-ranked PSM which recorded lower binding energy (BE), > 50% were triterpenoids which interacted strongly with viral spike protein—receptor binding domain, > 32% molecules which showed better interaction with the active site of human transmembrane serine protease were belongs to flavonoids and their glycosides, > 16% of flavonol glycosides and > 16% anthocyanidins recorded lower BE against active site of viral main protease and > 13% flavonol glycoside strongly interacted with active site of viral RNA-dependent RNA polymerase. The primary concern about these PSM is their bioavailability. However, several PSM recorded higher bioavailability score and found fulfilling most of the drug-likeness characters as per Lipinski's rule (Coagulin K, Kamalachalcone C, Ginkgetin, Isoginkgetin, 3,3′-Biplumbagin, Chrysophanein, Aromoline, etc.). Natural occurrence, bio-transformation, bioavailability of selected PSM and their interaction with the target site of selected proteins were discussed in detail. Present study provides a platform for researchers to explore the possible use of selected PSM to prevent/ cure the COVID-19 by subjecting them for thorough in vitro and in vivo evaluation for the capabilities to interfering with the process of viral host cell recognition, entry and replication.

Ecological Implications of Plant Secondary Metabolites - Phytochemical Diversity Can Enhance Agricultural Sustainability

Conventional agriculture production, although proficient in feeding an expanding human population, is having negative environmental impacts that are diminishing the sustainability of natural resources. Producers and consumers are increasingly interested in understanding how land management practices can enhance agricultural sustainability and improve human health. This perspective article offers a new approach to enhancing agricultural sustainability by growing crops and forages with diverse plant secondary metabolites (PSMs). Plants produce tens of thousands of PSMs to mediate interactions with soil, other plants, and animals. Plants use these metabolites to communicate with organisms in their environment, both above and belowground, and to modify the rhizosphere and influence chemical, physical, and biological attributes of soil. In pastures and rangelands, animal health benefits and production increases when animals ingest forages with different PSMs, which has implications for enhancing the biochemical richness of meat and dairy products for human consumption. A deeper understanding of PSMs, and their functional roles in agroecology, may help producers better manage their lands, reduce inputs, and minimize negative environmental impacts.

Upregulating the mevalonate pathway and repressing sterol synthesis in Saccharomyces cerevisiae enhances the production of triterpenes

Pentacyclic triterpenes are diverse plant secondary metabolites derived from the mevalonate (MVA) pathway. Many of these molecules are potentially valuable, particularly as pharmaceuticals, and research has focused on their production in simpler and more amenable heterologous systems such as the yeast Saccharomyces cerevisiae. We have developed a new heterologous platform for the production of pentacyclic triterpenes in S. cerevisiae based on a combinatorial engineering strategy involving the overexpression of MVA pathway genes, the knockout of negative regulators, and the suppression of a competing pathway. Accordingly, we overexpressed S. cerevisiae ERG13, encoding 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) synthase, and a truncated and deregulated variant of the rate-limiting enzyme HMG-CoA reductase 1 (tHMGR). In the same engineering step, we deleted the ROX1 gene, encoding a negative regulator of the MVA pathway and sterol biosynthesis, resulting in a push-and-pull strategy to enhance metabolic flux through the system. In a second step, we redirected this enhanced metabolic flux from late sterol biosynthesis to the production of 2,3-oxidosqualene, the direct precursor of pentacyclic triterpenes. In yeast cells transformed with a newly isolated sequence encoding lupeol synthase from the Russian dandelion (Taraxacum koksaghyz), we increased the yield of pentacyclic triterpenes by 127-fold and detected not only high levels of lupeol but also a second valuable pentacyclic triterpene product, β-amyrin.

Natural diversity of hydroxycinnamic acid derivatives, flavonoid glycosides, carotenoids and chlorophylls in leaves of six different amaranth species

Amaranth species are globally grown food crops. However, knowledge about the composition of their secondary metabolites is insufficient. Here, selected hydroxycinnamic acid derivatives, flavonoid glycosides, carotenoids and chlorophylls in the leaves of 14 genotypes from six different amaranth species were identified and quantified. For the first time, caffeic acid esters of isocitric and several aldaric acids were isolated and quantified in a leafy food matrix. High concentrations of hydroxycinnamic acid derivatives and chlorophylls, and moderate amounts of flavonoids and carotenoids were detected. A hierarchical clustering method of the metabolic profiles followed by Random Amplification of Polymorphic DNA (RAPD)-PCR fingerprinting was used to group the genotypes. Using this combined approach, three main groups of amaranth species were assigned. The information provided in this study increases the attractiveness of the amaranth genus as a food crop due to its strong diversity of plant secondary metabolites that are associated with numerous health-promoting benefits.

Phenolic compounds as functional ingredients and nutraceuticals: the case of Juice PLUS+

The protective effects of diets rich in fruits and vegetables are not only due to fibre, vitamins and minerals, but also to a diversity of plant secondary metabolites, in particular phenolic compounds and flavonoids. These compounds are now widely used as food ingredients and supplements, with the final aim of making the health benefits associated with their intake more widely and easily available. The bioavailability of these compounds after dietary intake has been a topic of increasing research in recent years, and could be one of the crucial issues when trying to develop commercial products containing polyphenols. Moreover, it is now well recognised that following ingestion, phenolic compounds undergo extensive metabolism during their passage through the gastrointestinal tract and, with very few exceptions, only metabolites and catabolites of the parent compounds enter the circulatory system. Metabolism occurs initially in the lumen of the small intestine with cleavage of sugar moieties after which the released aglycone undergoes glucuronidation, sulfation and/or methylation [1]. After entry of these metabolites into the circulatory system, phase II metabolism may also occur in the liver and other organs [1]. However, a large proportion of ingested polyphenols are not absorbed in the small intestine [1] and so pass to the large intestine, where substantial structural modifications are mediated by the local colonic microbiota [1]. The resultant low‐molecular‐weight compounds, may be absorbed and circulate in the body prior to excretion in urine in amounts that greatly exceed those of metabolites absorbed in the small intestine, whose concentration in plasma rarely exceeds the nmol/l levels [1]. Here we evaluated the composition and the absorption kinetics of a set commercially available encapsulated nutraceuticals, namely Juice PLUS+ Vineyard (a berry blend, JBB), Juice PLUS+ Fruit Blend (JFB) and Juice PLUS+ Vegetable Blend (JVB), kindly supplied by Juice PLUS+ company. The analysis of the three supplements by uHPLC–MSn allowed to tentatively identify 119 (poly)phenolic compounds. The main microbial/human metabolites were then tracked in a short pharmacokinetic study involving 20 healthy volunteers, demonstrating that a diversity of compounds circulate in vivo after Juice PLUS+ capsules intake, becoming potentially responsible of the health related effects which have been previously reported in a plethora of intervention studies [examples 2–4].Support or Funding InformationPart of this work has been supported by “Green Beat – Institute of Nutrient Research and Sport Nutrition”

Identification of a Novel L-rhamnose Uptake Transporter in the Filamentous Fungus Aspergillus niger.

The study of plant biomass utilization by fungi is a research field of great interest due to its many implications in ecology, agriculture and biotechnology. Most of the efforts done to increase the understanding of the use of plant cell walls by fungi have been focused on the degradation of cellulose and hemicellulose, and transport and metabolism of their constituent monosaccharides. Pectin is another important constituent of plant cell walls, but has received less attention. In relation to the uptake of pectic building blocks, fungal transporters for the uptake of galacturonic acid recently have been reported in Aspergillus niger and Neurospora crassa. However, not a single L-rhamnose (6-deoxy-L-mannose) transporter has been identified yet in fungi or in other eukaryotic organisms. L-rhamnose is a deoxy-sugar present in plant cell wall pectic polysaccharides (mainly rhamnogalacturonan I and rhamnogalacturonan II), but is also found in diverse plant secondary metabolites (e.g. anthocyanins, flavonoids and triterpenoids), in the green seaweed sulfated polysaccharide ulvan, and in glycan structures from viruses and bacteria. Here, a comparative plasmalemma proteomic analysis was used to identify candidate L-rhamnose transporters in A. niger. Further analysis was focused on protein ID 1119135 (RhtA) (JGI A. niger ATCC 1015 genome database). RhtA was classified as a Family 7 Fucose: H+ Symporter (FHS) within the Major Facilitator Superfamily. Family 7 currently includes exclusively bacterial transporters able to use different sugars. Strong indications for its role in L-rhamnose transport were obtained by functional complementation of the Saccharomyces cerevisiae EBY.VW.4000 strain in growth studies with a range of potential substrates. Biochemical analysis using L-[3H(G)]-rhamnose confirmed that RhtA is a L-rhamnose transporter. The RhtA gene is located in tandem with a hypothetical alpha-L-rhamnosidase gene (rhaB). Transcriptional analysis of rhtA and rhaB confirmed that both genes have a coordinated expression, being strongly and specifically induced by L-rhamnose, and controlled by RhaR, a transcriptional regulator involved in the release and catabolism of the methyl-pentose. RhtA is the first eukaryotic L-rhamnose transporter identified and functionally validated to date.

Phylogeny Explains Variation in The Root Chemistry of Eucalyptus Species.

Plants are dependent on their root systems for survival, and thus are defended from belowground enemies by a range of strategies, including plant secondary metabolites (PSMs). These compounds vary among species, and an understanding of this variation may provide generality in predicting the susceptibility of forest trees to belowground enemies and the quality of their organic matter input to soil. Here, we investigated phylogenetic patterns in the root chemistry of species within the genus Eucalyptus. Given the known diversity of PSMs in eucalypt foliage, we hypothesized that (i) the range and concentrations of PSMs and carbohydrates in roots vary among Eucalyptus species, and (ii) that phylogenetic relationships explain a significant component of this variation. To test for interspecific variation in root chemistry and the influence of tree phylogeny, we grew 24 Eucalyptus species representing two subgenera (Eucalyptus and Symphyomyrtus) in a common garden for two years. Fine root samples were collected from each species and analyzed for total phenolics, condensed tannins, carbohydrates, terpenes, and formylated phloroglucinol compounds. Compounds displaying significant interspecific variation were mapped onto a molecular phylogeny and tested for phylogenetic signal. Although all targeted groups of compounds were present, we found that phenolics dominated root defenses and that all phenolic traits displayed significant interspecific variation. Further, these compounds displayed a significant phylogenetic signal. Overall, our results suggest that within these representatives of genus Eucalyptus, more closely related species have more similar root chemistry, which may influence their susceptibility to belowground enemies and soil organic matter accrual.

RNA interference targeting CYP76AH1 in hairy roots of Salvia miltiorrhiza reveals its key role in the biosynthetic pathway of tanshinones

Plant cytochrome P450s (CYPs) are well known as the largest family of enzymes that contribute to both primary metabolism and the chemical diversity of plant secondary metabolites. It is important to elucidate the in vivo role of CYPs in secondary metabolism, in order to apply them in the production of valuable metabolites in medicinal plants via metabolic engineering. CYP76AH1 has been suggested to catalyze the conversion of the carbon skeleton miltiradiene into the intermediate ferruginol, which is involved in the biosynthesis of tanshinones, the chief bioactive ingredients of Salvia miltiorrhiza. However, its role in planta remains to be elucidated. In this work, we constructed a CYP76AH1 RNAi system for hairy roots. Metabolic analysis of RNAi-AH1 hairy root lines showed a significantly increased accumulation of miltiradiene compared to the control lines. At the same time, the concentration of ferruginol decreased revealing the in vivo catalytic activity of CYP76AH1. The content of tanshinones decreased significantly after silencing of CYP76AH1, which verified its key role in the biosynthesis of tanshinones, and indicated that it could be used as a target for metabolic engineering.

Development of a rapid antimicrobial screening method for natural products using genetic programming

The importance of natural products in modern drug discovery is undisputed [1], not least thanks to the enormous structural diversity of plant secondary metabolites. However, the identification of compounds with a desired activity within this vast abundance remains a challenge. High-throughput screening is a successful approach, where simple tests gauge in a short time a compound's potential for further development. Our group employs screening techniques to search for novel antimicrobial agents from natural sources. In its simplest (and most straightforward) way, antimicrobial activity is detected by measuring the absorbance of a bacterial suspension after 24h incubation at a single wavelength. While this method is well-known and standardized [2], it is unspecific and fairly prone to assay interferences. With the goal to facilitate the screening process and thus increase its throughput, we have developed an assay that allows the antimicrobial activity screening of natural products against three microorganisms (Staphylococcus aureus, Escherichia coli, Candida albicans) simultaneously, as opposed to one at a time. In an attempt to get more comprehensive information, we mined the VIS and IR spectra of the bacterial/fungal suspension using genetic programming. Genetic programming is a machine learning algorithm, which follows the principal of biological evolution to develop (or evolve) a mathematical formula that, when applied to the spectra, yields a conjecture about present bacteria or fungi. This method has previously been used to successfully type single bacterial strains with the aid of IR spectroscopy [3]. Here, we present the development of this assay, with focus on the underlying mechanism, optimization process, the potential and pitfalls of genetic programming in bioactivity screening of natural products.