Quality parameters for the medicinal plant Drosera rotundifolia L.: A new approach with established techniques.

Monographs of the European Pharmacopoeia (Ph. Eur.) are the basis for quality control of medicinal plants and therefore important to ensure the consistency, quality, safety, and efficacy of phytopharmaceuticals. The traditional medicinal plant sundew (Drosera sp.) has disappeared from therapy due to nature conservation, but can now be cultivated sustainably on rewetted peatland. However, currently there is no valid Ph. Eur. monograph for the quality control of Droserae herba. In this study, sundew material from different species and sources was investigated with the aim of developing quality control methods based on the Ph. Eur. and defining a uniform quality standard for Droserae herba. It was possible to distinguish between sundew species of different quality, using macroscopic, microscopic, and chromatographic methods. Special emphasis was laid on the content of flavonoids and naphthoquinones as important quality parameters as their content differed between the sundew species. The differences in content and toxicity result in the recommendation that only round-leaved sundew (Drosera rotundifolia L.) should be used as a medicinal plant for the production of phytopharmaceuticals in the future.

Dramatic difference in rate of chromosome number evolution among sundew (Drosera L., Droseraceae) lineages.

Chromosome number change is a driver of speciation in eukaryotic organisms. Carnivorous sundews in the plant genus Drosera L. exhibit single chromosome number variation both among and within species, especially in the Australian Drosera subg. Ergaleium D.C., potentially linked to atypical centromeres that span much of the length of the chromosomes. We critically reviewed the literature on chromosome counts in Drosera, verified the taxonomy and quality of the original counts, and reconstructed dated phylogenies. We used the BiChrom model to test whether rates of single chromosome number increase and decrease, and chromosome number doubling differed between D. subg. Ergaleium and the other subgenera and between self-compatible and self-incompatible lineages. The best model for chromosome evolution among subgenera had equal rates of chromosome number doubling but higher rates of single chromosome number change in D. subg. Ergaleium than in the other subgenera. Contrary to expectation, self-incompatible lineages had a significantly higher rate of single chromosome loss than self-compatible lineages. We found no evidence for an association between differences in single chromosome number changes and diploidization after polyploidy or centromere type. This study presents an exemplar for critically examining published cytological data and rigorously testing factors that may impact the rates of chromosome number evolution.

Effects of Environmental Conditions on the Individual Architectures and Photosynthetic Performances of Three Species in Drosera.

The aim of this study was to determine the environmental conditions, individual architectures, and photosynthetic efficiencies of three sundew species: Drosera rotundifolia, D. anglica, and D. intermedia, found in well-preserved peatlands and sandy lake shores in NW Poland. Morphological traits and chlorophyll a fluorescence (Fv/Fm) were measured in 581 individuals of Drosera. D. anglica occupies the best-lit and warmest habitats, and also those that are the most heavily hydrated and the richest in organic matter; its rosettes are larger under conditions of higher pH, less organic matter, and less well-lit habitats. D. intermedia occupies substrates with the highest pH but the lowest conductivity, the poorest level of organic matter, and the least hydration. It is highly variable in terms of individual architecture. D. rotundifolia occupies habitats that are the most diverse, and that are often poorly lit, with the lowest pH but the highest conductivity. It is the least variable in terms of individual architecture. The value of the Fv/Fm ratio in Drosera is low (0.616 ± 0.137). The highest photosynthetic efficiency is achieved by D. rotundifolia (0.677 ± 0.111). It is significant for all substrates, indicating its high phenotypic plasticity. The other species have lower and similar Fv/Fm values (D. intermedia, 0.571 ± 0.118; D. anglica, 0.543 ± 0.154). Due to its very low photosynthetic efficiency, D. anglica avoids competition by occupying highly hydrated habitats. D. intermedia has adapted to the occupation of highly variable habitats in terms of hydration, while D. rotundifolia is primarily adapted to variable light conditions.

Expression and function of an S1-type nuclease in the digestive fluid of a sundew, Drosera adelae.

BACKGROUND AND AIMS: Carnivorous plants trap and digest insects and similar-sized animals. Many studies have examined enzymes in the digestive fluids of these plants and have gradually unveiled the origins and gene expression of these enzymes. However, only a few attempts have been made at characterization of nucleases. This study aimed to reveal gene expression and the structural, functional and evolutionary characteristics of an S1-type nuclease (DAN1) in the digestive fluid of an Australian sundew, Drosera adelae, whose trap organ shows unique gene expression and related epigenetic regulation. METHODS: Organ-specificity in Dan1 expression was examined using glandular tentacles, laminas, roots and inflorescences, and real-time PCR. The methylation status of the Dan1 promoter in each organ was clarified by bisulphite sequencing. The structural characteristics of DAN1 were studied by a comparison of primary structures of S1-type nucleases of three carnivorous and seven non-carnivorous plants. DAN1 was prepared using a cell-free protein synthesis system. Requirements for metal ions, optimum pH and temperature, and substrate preference were examined using conventional methods. KEY RESULTS: Dan1 is exclusively expressed in the glandular tentacles and its promoter is almost completely unmethylated in all organs. This is in contrast to the S-like RNase gene da-I of Dr. adelae, which shows similar organ-specific expression, but is controlled by a promoter that is specifically unmethylated in the glandular tentacles. Comparison of amino acid sequences of S1-type nucleases identifies seven and three positions where amino acid residues are conserved only among the carnivorous plants and only among the non-carnivorous plants, respectively. DAN1 prefers a substrate RNA over DNA in the presence of Zn2+, Mn2+ or Ca2+ at an optimum pH of 4.0. CONCLUSIONS: Uptake of phosphates from prey is suggested to be the main function of DAN1, which is very different from the known functions of S1-type nucleases. Evolution has modified the structure and expression of Dan1 to specifically function in the digestive fluid.

Antibiofilm Activity of Sundew Species against Multidrug-Resistant Escherichia coli Strains.

Species of the genus Drosera, known for carnivorous plants, such as sundew, have been traditionally used for centuries as medicinal plants. Efficacy-determining compounds are naphthoquinones and flavonoids. Flavonoids possess a broad spectrum of bioactive properties, including biofilm inhibitory activity. Biofilms render antibiotics ineffective, contributing to the current rise in antimicrobial resistance. In this study, the biofilm inhibitory activity of two European sundew species (Drosera rotundifolia and Drosera intermedia) grown agriculturally in Germany and four commercial sundew products (declared as Drosera longifolia, Drosera sp. and Drosera planta trit.) against three multidrug-resistant Escherichia coli strains was tested. The aim of the study was to comparatively investigate the biofilm inhibitory potential of sundew species extracts grown locally in northern Germany and commercial sundew products. The minimum biofilm inhibitory concentration of the European sundew species was approx. 35 µg mL-1. In comparison, commercial sundew products ranged in concentration from 75 to 140 µg mL-1. Additionally, individual compounds isolated from European sundew were tested. Among these compounds, biofilm inhibitory activity was determined for four of the eight substances, with 2″-O-galloyl hyperoside standing out for its activity (38 µg mL-1). The whole plant extracts of Drosera rotundifolia and Drosera intermedia proved to be more effective than the commercial products and the single compounds in its biofilm inhibition activity against Escherichia coli strains. Sundew extracts may serve as a potential therapeutic approach for targeting biofilm production.

Effects of Extracts and Flavonoids from Drosera rotundifolia L. on Ciliary Beat Frequency and Murine Airway Smooth Muscle.

Extracts from Drosera rotundifolia are traditionally used to treat cough symptoms during a common cold. The present study aimed to investigate the impact of extracts from D. rotundifolia and active compounds on the respiratory tract. Tracheal slices of C57BL/6N mice were used ex vivo to examine effects on airway smooth muscle (ASM) and ciliary beat frequency (CBF). Phosphodiesterase (PDE) inhibition assays were carried out to test whether PDE1 or PDE4 are targeted by the active compounds. An ethanol-water extract, as well as an aqueous fraction of this extract, exerted antispasmodic properties against acetylcholine-induced contractions. In addition, contractions induced by 60 mM K+ were abrogated by the aqueous fraction. Effects on ASM could be attributed to the flavonoids quercetin, 2″-O-galloylhyperoside and hyperoside. Moreover, the Drosera extract and the aqueous fraction increased the CBF of murine tracheal slices. Quercetin and 2″-O-galloylhyperoside were identified as active compounds involved in the elevation of CBF. Both compounds inhibited PDE1A and PDE4D. The elevation of CBF was mimicked by the subtype-selective PDE inhibitor rolipram (PDE4) and by 8-methoxymethyl-IBMX. In summary, our study shows, for the first time, that a Drosera extract and its flavonoid compounds increase the CBF of murine airways while antispasmodic effects were transferred to ASM.

Rapid flower closure of Drosera tokaiensis deters caterpillar herbivory.

Certain plants exhibit rapid movement in response to mechanical stimulation; however, the ecological functions of this behaviour are largely unknown. Here, we show that the rapid flower closure of Drosera tokaiensis (Droseraceae) in response to mechanical stimulation functions as a physical defence against a specialist herbivore Buckleria paludum (Pterophoridae) caterpillar. Following feeding damage on fruits, flowers, flower stalks and buds by B. paludum, D. tokaiensis closed its flowers nine times faster than during natural circadian closure. The extent of damage to ovules was significantly reduced when the flowers were able to close compared with the condition in which closure was physically inhibited by the application of a resin. Nonetheless, flower closure had no effect on the feeding damage to stamens and styles and promoted further damage to petals. Given that feeding on petals, stamens and styles had no significant effect on the number of mature seeds, rapid flower closure leading to the protection of ovules had an overall positive effect on the reproductive success of D. tokaiensis. Our study showed rapid plant movement as a novel case of induced physical defence against herbivory.

Leaf NPK stoichiometry, δ15 N, and apparent nutrient limitation of co-occurring carnivorous and noncarnivorous plants.

Previous meta-analyses suggested that carnivorous plants-despite access to N, P, and K from prey-have significantly lower leaf concentrations of these nutrients than noncarnivores. Those studies, however, largely compared carnivores in nutrient-poor habitats with noncarnivores in more nutrient-rich sites, so that the differences reported might reflect habitat differences as much as differences in nutrient-capture strategy. Here we examine three carnivorous and 12 noncarnivorous plants in the same nutrient-poor bog to compare their foliar nutrient concentrations, assess their patterns of nutrient limitation using leaf NPK stoichiometry, and estimate percentage N derived from prey by carnivores using a mixing model for stable N isotopes. We hypothesized that (1) carnivore leaf nutrient concentrations approach or exceed those of noncarnivores in the same nutrient-poor habitat; (2) species in different functional groups show different patterns of stoichiometry and apparent nutrient limitation; and (3) noncarnivores might show evidence of using other means of nutrient acquisition or conservation to reduce nutrient limitation. At Fallison Bog in northern Wisconsin, carnivorous plants (Drosera rotundifolia, Sarracenia purpurea, Utricularia macrorhiza) showed significantly lower leaf percentage C and N:P ratio, higher δ15 N, and no difference from noncarnivores in leaf N, P, K, and δ13 C. Sedges had significantly lower leaf percentage P, percentage C, and N:K ratio, and higher K:P ratio than nonsedges restricted to the Sphagnum mat, and may tap peat N via aerenchyma-facilitated peat oxidation (oxipeditrophy). Evergreen ericaceous shrubs exhibited significantly higher levels of percentage C and lower values of δ15 N than mat nonericads. Calla palustris-growing in the nutrient-rich moat at the bog's upland edge-had very high values of leaf N, K, δ15 N, and N:P ratio, suggesting that it may obtain nutrients from minerotrophic flows from the adjacent uplands and/or rapidly decaying peat. Stoichiometric analyses indicated that most species are N limited. A mixing model applied to δ15 N values for carnivores, noncarnivores, and insects produced an estimate of 50% of leaf N derived from prey for Utricularia, 42% for Sarracenia, and 41% for Drosera.

Microwave Radiation as an Inducer of Secondary Metabolite Production in Drosera rotundifolia In Vitro Plantlets.

In this study, low-intensity microwave irradiation (frequency of 2.45 GHz; 26.3 kW m-3 power density) for 0, 5, and 30 s was tested for the first time on Drosera rotundifolia in vitro plantlets to explore its effect on the production of highly valued phenolic compounds. Analysis of the extracts obtained from irradiated plantlets revealed time-dependent increases in the levels of photosynthetic pigments, particularly the carotenoids, whereas symptoms of growth decline were not observed. Similarly, the highest total antioxidant capacity and total phenolic and flavonoid contents were detected in 30-s-irradiated plantlets. High-performance liquid chromatography analysis revealed that the content of the bioactive phenolics 5-hydroxy-7-methylnaphthalene-1,4-dione (1), 2-(3,4-dihydroxyphenyl)-3,5,7-trihydroxychromen-4-one (2), and 2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-3-[(3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxychromen-4-one (3), which are considered to be responsible for the pharmacological properties of this species, was around twofold higher in 30-s-irradiated plantlets than in controls. Nevertheless, the accumulation of 5-hydroxy-2-methylnaphthalene-1,4-dione (4), which was present only in trace amounts in the plant roots, decreased by 30% upon microwave irradiation. The results indicate that microwave treatment acts as an effective inducer of the production of phenylpropanoid compounds, which opens up new opportunities for its use in biotechnological applications.

Dynamic calcium signals mediate the feeding response of the carnivorous sundew plant.

Some of the most spectacular examples of botanical carnivory-in which predator plants catch and digest animals presumably to supplement the nutrient-poor soils in which they grow-occur within the Droseraceae family. For example, sundews of the genus Drosera have evolved leaf movements and enzyme secretion to facilitate prey digestion. The molecular underpinnings of this behavior remain largely unknown; however, evidence suggests that prey-induced electrical impulses are correlated with movement and production of the defense hormone jasmonic acid (JA), which may alter gene expression. In noncarnivorous plants, JA is linked to electrical activity via changes in cytoplasmic Ca2+. Here, we find that dynamic Ca2+ changes also occur in sundew (Drosera spatulata) leaves responding to prey-associated mechanical and chemical stimuli. Furthermore, inhibition of these Ca2+ changes reduced expression of JA target genes and leaf movements following chemical feeding. Our results are consistent with the presence of a conserved Ca2+-dependent JA signaling pathway in the sundew feeding response and provide further credence to the defensive origin of plant carnivory.

A novel approach for reliable qualitative and quantitative prey spectra identification of carnivorous plants combining DNA metabarcoding and macro photography.

Prey spectra (the number and composition of captured arthropods) represent a crucial aspect of carnivorous plant ecology, yet remain poorly studied. Traditional morphology-based approaches for prey identification are time-intensive, require specialists with considerable knowledge of arthropod taxonomy, and are hampered by high numbers of unidentifiable (i.e., heavily digested) prey items. We examined prey spectra of three species of closely-related annual Drosera (Droseraceae, sundews) from tropical northern Australia using a novel DNA metabarcoding approach with in-situ macro photography as a plausibility control and to facilitate prey quantity estimations. This new method facilitated accurate analyses of carnivorous plant prey spectra (even of heavily digested prey lacking characteristic morphological features) at a taxonomic resolution and level of completeness far exceeding morphology-based methods and approaching the 100% mark at arthropod order level. Although the three studied species exhibited significant differences in detected prey spectra, little prey specialisation was observed and habitat or plant population density variations were likely the main drivers of prey spectra dissimilarity.

Multifaceted phytogenic silver nanoparticles by an insectivorous plant Drosera spatulata Labill var. bakoensis and its potential therapeutic applications.

The current investigation highlights the green synthesis of silver nanoparticles (AgNPs) by the insectivorous plant Drosera spatulata Labill var. bakoensis, which is the first of its kind. The biosynthesized nanoparticles revealed a UV visible surface plasmon resonance (SPR) band at 427 nm. The natural phytoconstituents which reduce the monovalent silver were identified by FTIR. The particle size of the Ds-AgNPs was detected by the Nanoparticle size analyzer confirms that the average size of nanoparticles was around 23 ± 2 nm. Ds-AgNPs exhibit high stability because of its high negative zeta potential (- 34.1 mV). AFM studies also revealed that the Ds-AgNPs were spherical in shape and average size ranges from 10 to 20 ± 5 nm. TEM analysis also revealed that the average size of Ds-AgNPs was also around 21 ± 4 nm and the shape is roughly spherical and well dispersed. The crystal nature of Ds-AgNPs was detected as a face-centered cube by the XRD analysis. Furthermore, studies on antibacterial and antifungal activities manifested outstanding antimicrobial activities of Ds-AgNPs compared with standard antibiotic Amoxyclav. In addition, demonstration of superior free radical scavenging efficacy coupled with potential in vitro cytotoxic significance on Human colon cancer cell lines (HT-29) suggests that the Ds-AgNPs attain excellent multifunctional therapeutic applications.

Drosera-Inspired Dual-Actuating Double-Layer Hydrogel Actuator.

Drosera is a small insectivorous plant whose antennae can fold up, encircle, and prey. The rapid movement of the antennae is achieved by the synergistic effect of a double-layer structure with the antennae contracts on the front and expands on the back. In this work, a drosera-inspired dual-actuating double-layer hydrogel actuator is proposed, in which the temperature-responsive poly(N, N-diethyl acrylamide) (PDEAAm) layer acts as the main actuation layer and a moisture-responsive poly(acrylamide) (PAAm) layer acts as the auxiliary actuation layer. In a water environment with low temperature, both the PAAm and PDEAAm layers absorb water and expand with a swelling property. When the temperature exceeds the lower critical solution temperature of PDEAAm, the PDEAAm layer undergoes a hydrophilic-hydrophobic transition and shrinks rapidly. Therefore, the synergistic effect of the double-layer hydrogel enables the double-layer hydrogel to achieve a large bending angle at high temperature. In addition, when designing and fabricating shape-patterned double-layer hydrogels, complex shape changes can be achieved. Due to the physical and chemical properties, the actuator can be used to grab, transport, and release objects. This drosera-inspired double-layer hydrogel actuator has high practical value, which may provide new insights for the design and manufacture of artificial intelligence materials.

Field-Grown and In Vitro Propagated Round-Leaved Sundew (Drosera rotundifolia L.) Show Differences in Metabolic Profiles and Biological Activities.

Drosera rotundifolia L. is a carnivorous plant used in traditional medicine for its therapeutic properties. Because of its small size, its collection in nature is laborious and different cultivation methods have been studied to ensure availability. However, only a few studies exist where the lab-grown sundew tissue and field-grown sundew would have been compared in their functionality or metabolic profiles. In this study, the antioxidant and antiviral activities of lab-grown and field-grown sundew extracts and their metabolic profiles are examined. The effect of drying methods on the chromatographic profile of the extracts is also shown. Antioxidant activity was significantly higher (5-6 times) in field-grown sundew but antiviral activity against enterovirus strains coxsackievirus A9 and B3 was similar in higher extract concentrations (cell viability ca. 90%). Metabolic profiles showed that the majority of the identified compounds were the same but field-grown sundew contained higher numbers and amounts of secondary metabolites. Freeze-drying, herbal dryer, and oven or room temperature drying of the extract significantly decreased the metabolite content from -72% up to -100%. Freezing was the best option to preserve the metabolic composition of the sundew extract. In conclusion, when accurately handled, the lab-grown sundew possesses promising antiviral properties, but the secondary metabolite content needs to be higher for it to be considered as a good alternative for the field-grown sundew.

Gland cell responses to feeding in Drosera capensis, a carnivorous plant.

Glands of Drosera absorb and transport nutrients from captured prey, but the mechanism and dynamics remain unclear. In this study, we offered animal proteins in the form of fluorescent albumin (FITC-BSA) and observed the reactions of the glands by live cell imaging and fluorescence microscopy. The ultrastructure of these highly dynamic processes was also assessed in high-pressure frozen and freeze substituted (HPF-FS) cells. HPF-FS yielded excellent preservation of the cytoplasm of all cell types, although the cytosol looked different in gland cells as compared to endodermoid and stalk cells. Especially prominent were the ER and its contacts with the plasma membrane, plasmodesmata, and other organelles as well as continuities between organelles. Also distinct were actin microfilaments in association with ER and organelles. Application of FITC-BSA to glands caused the formation of fluorescent endosomes that pinched off the plasma membrane. Endosomes fused to larger aggregates, and accumulated in the bulk cytoplasm around the nucleus. They did not fuse with the cell sap vacuole but remained for at least three days; in addition, fluorescent vesicles also proceeded through endodermoid and transfer cells to the epidermal and parenchymal cells of the tentacle stalk.

Metabolomic analysis reveals reliance on secondary plant metabolites to facilitate carnivory in the Cape sundew, Drosera capensis.

BACKGROUND AND AIMS: Secondary metabolites are integral to multiple key plant processes (growth regulation, pollinator attraction and interactions with conspecifics, competitors and symbionts) yet their role in plant adaptation remains an underexplored area of research. Carnivorous plants use secondary metabolites to acquire nutrients from prey, but the extent of the role of secondary metabolites in plant carnivory is not known. We aimed to determine the extent of the role of secondary metabolites in facilitating carnivory of the Cape sundew, Drosera capensis. METHODS: We conducted metabolomic analysis of 72 plants in a time-series experiment before and after simulated prey capture. We used ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) and the retention time index to identify compounds in the leaf trap tissue that changed up to 72 h following simulated prey capture. We identified associated metabolic pathways, and cross-compared these compounds with metabolites previously known to be involved in carnivorous plants across taxa. KEY RESULTS: For the first time in a carnivorous plant, we have profiled the whole-leaf metabolome response to prey capture. Reliance on secondary plant metabolites was higher than previously thought - 2383 out of 3257 compounds in fed leaves had statistically significant concentration changes in comparison with unfed controls. Of these, ~34 compounds are also associated with carnivory in other species; 11 are unique to Nepenthales. At least 20 compounds had 10-fold changes in concentration, 12 of which had 30-fold changes and are typically associated with defence or attraction in non-carnivorous plants. CONCLUSIONS: Secondary plant metabolites are utilized in plant carnivory to an extent greater than previously thought - we found a whole-metabolome response to prey capture. Plant carnivory, at the metabolic level, likely evolved from at least two distinct functions: attraction and defence. Findings of this study support the hypothesis that secondary metabolites play an important role in plant diversification and adaptation to new environments.

Phytochemical Profiling and Biological Activity of the Australian Carnivorous Plant, Drosera magna.

Phytochemical profiling was undertaken on the crude extracts of Drosera magna to determine the nature of the chemical constituents present. In total, three new flavonol diglycosides (1-3), one new flavan-3-ol glycoside (4), and 12 previously reported compounds of the flavonol (5, 9), flavan-3-ol (15), flavanone (8), 1,4-napthoquinone (6, 7, 13, 14), 2,3-dehydroxynapthalene-1,4-dione (10-12), and phenolic acid (16) structure classes were isolated and identified. Compounds 1-9, 13, 17, and 18 were assessed for antimicrobial activity, with compounds 6, 7, 8, and 9 showing significant activity. Compounds 1, 2, and 6 were also evaluated for anthelmintic activity against larval forms of Hemonchus contortus, with compound 6 being active.

Low-dose Drosera rotundifolia induces gene expression changes in 16HBE human bronchial epithelial cells.

Drosera rotundifolia has been traditionally used for the treatment of respiratory diseases in phytotherapy and homeopathy. The mechanisms of action recognized so far are linked to the known effects of specific components, such as flavonoids, but are not completely understood. In this study, the biological functions of D. rotundifolia were explored in vitro following the treatment of bronchial epithelial cells, which are the potential targets of the pharmacological effects of the herbal medicine. To do so, the whole plant ethanolic extract was 1000-fold diluted in water (D. rotundifolia 3×) and added to a 16HBE human cell line culture for 3 h or 6 h. The effects on gene expression of the treatments and corresponding controls were then investigated by RNA sequencing. The differentially expressed genes were validated through RT-qPCR, and the enriched biological functions involved in the effects of treatment were investigated. D. rotundifolia 3× did not impair cell viability and was shown to be a stimulant of cell functions by regulating the expression of dozens of genes after 3 h, and the effects were amplified after 6 h of treatment. The main differentially expressed genes encoded ligands of epithelial growth factor receptor, proteins involved in xenobiotic detoxification and cytokines, suggesting that D. rotundifolia 3× could stimulate self-repair systems, which are impaired in airway diseases. Furthermore, D. rotundifolia 3× acts on a complex and multifaceted set of genes and may potentially affect different layers of the bronchial mucosa.

Organ-specific expression and epigenetic traits of genes encoding digestive enzymes in the lance-leaf sundew (Drosera adelae).

Over the last two decades, extensive studies have been performed at the molecular level to understand the evolution of carnivorous plants. As fruits, the repertoire of protein components in the digestive fluids of several carnivorous plants have gradually become clear. However, the quantitative aspects of these proteins and the expression mechanisms of the genes that encode them are still poorly understood. In this study, using the Australian sundew Drosera adelae, we identified and quantified the digestive fluid proteins. We examined the expression and methylation status of the genes corresponding to major hydrolytic enzymes in various organs; these included thaumatin-like protein, S-like RNase, cysteine protease, class I chitinase, ß-1, 3-glucanase, and hevein-like protein. The genes encoding these proteins were exclusively expressed in the glandular tentacles. Furthermore, the promoters of the ß-1, 3-glucanase and cysteine protease genes were demethylated only in the glandular tentacles, similar to the previously reported case of the S-like RNase gene da-I. This phenomenon correlated with high expression of the DNA demethylase DEMETER in the glandular tentacles, strongly suggesting that it performs glandular tentacle-specific demethylation of the genes. The current study strengthens and generalizes the relevance of epigenetics to trap organ-specific gene expression in D. adelae. We also suggest similarities between the trap organs of carnivorous plants and the roots of non-carnivorous plants.