Does the Growth Differentiation Balance Hypothesis Explain Allocation to Secondary Metabolites in <Emphasis Type="Italic">Combretum apiculatum</Emphasis><Emphasis>,</Emphasis> an African Savanna Woody Species<Emphasis>?</Emphasis>

The growth differentiation balance hypothesis (GDBH) provides a framework that predicts a trade-off between costs of secondary metabolites (SMs) relative to the demand for photosynthate by growth. However, this hypothesis was developed using empirical evidence from plant species in northern boreal and temperate systems, leaving its applicability to species under different abiotic and biotic conditions questionable and generalizations problematic. The objective of this study was to investigate whether the GDBH explains allocation to SMs in the deciduous African savanna woody species C. apiculatum along a 6-point N gradient. The cornerstone prediction of the GDBH, i.e., the parabolic response in SMs along the N gradient, was not observed, with secondary metabolism showing compound-specific responses. Quercetin, myricetin, and kaempferol glycoside concentrations, all produced via the same pathway, responded differently across the N gradient. Flavonol glycoside, cinnamic acid, and quercetin glycoside concentrations decreased as N increased, which provides partial support for the carbon nutrient balance hypothesis. Simulated herbivory had no effect on photosynthesis, decreased foliar N and consequently increased C:N ratio, but did not induce an increase in SMs, with condensed tannins and flavonol glycosides being unaffected. Defoliated plants at low N concentration compensated for lost biomass, which suggests a tolerance response, but as predicted by the limiting resource model, plants at higher N concentration were evidently C limited and thus unable to compensate. Our results show that the GDBH does not explain allocation to SMs in C. apiculatum, and suggest that mechanistic explanations of plant allocation should consider the integrative defensive effect of changed SMs.     

Direct and Indirect Effects of Invasive Plants on Soil Chemistry and Ecosystem Function

Invasive plants have a multitude of impacts on plant communities through their direct and indirect effects on soil chemistry and ecosystem function. For example, plants modify the soil environment through root exudates that affect soil structure, and mobilize and/or chelate nutrients. The long-term impact of litter and root exudates can modify soil nutrient pools, and there is evidence that invasive plant species may alter nutrient cycles differently from native species. The effects of plants on ecosystem biogeochemistry may be caused by differences in leaf tissue nutrient stoichiometry or secondary metabolites, although evidence for the importance of allelochemicals in driving these processes is lacking. Some invasive species may gain a competitive advantage through the release of compounds or combinations of compounds that are unique to the invaded community—the “novel weapons hypothesis.” Invasive plants also can exert profound impact on plant communities indirectly through the herbicides used to control them. Glyphosate, the most widely used herbicide in the world, often is used to help control invasive weeds, and generally is considered to have minimal environmental impacts. Most studies show little to no effect of glyphosate and other herbicides on soil microbial communities. However, herbicide applications can reduce or promote rhizobium nodulation and mycorrhiza formation. Herbicide drift can affect the growth of non-target plants, and glyphosate and other herbicides can impact significantly the secondary chemistry of plants at sublethal doses. In summary, the literature indicates that invasive species can alter the biogeochemistry of ecosystems, that secondary metabolites released by invasive species may play important roles in soil chemistry as well as plant-plant and plant-microbe interactions, and that the herbicides used to control invasive species can impact plant chemistry and ecosystems in ways that have yet to be fully explored.     

Potential of Silver Nanoparticles in Overcoming the Intrinsic Resistance of Pseudomonas aeruginosa to Secondary Metabolites from Carnivorous Plants

Carnivorous plants are exemplary natural sources of secondary metabolites with biological activity. However, the therapeutic antimicrobial potential of these compounds is limited due to intrinsic resistance of selected bacterial pathogens, among which Pseudomonas aeruginosa represents an extreme example. The objective of the study was to overcome the intrinsic resistance of P. aeruginosa by combining silver nanoparticles (AgNPs) with secondary metabolites from selected carnivorous plant species. We employed the broth microdilution method, the checkerboard titration technique and comprehensive phytochemical analyses to define interactions between nanoparticles and active compounds from carnivorous plants. It has been confirmed that P. aeruginosa is resistant to a broad range of secondary metabolites from carnivorous plants, i.e., naphthoquinones, flavonoids, phenolic acids (MBC = 512 µg mL⁻¹) and only weakly sensitive to their mixtures, i.e., extracts and extracts’ fractions. However, it was shown that the antimicrobial activity of extracts and fractions with a significant level of naphthoquinone (plumbagin) was significantly enhanced by AgNPs. Our studies clearly demonstrated a crucial role of naphthoquinones in AgNPs and extract interaction, as well as depicted the potential of AgNPs to restore the bactericidal activity of naphthoquinones towards P. aeruginosa. Our findings indicate the significant potential of nanoparticles to modulate the activity of selected secondary metabolites and revisit their antimicrobial potential towards human pathogenic bacteria.     

Chemical Diversity and Defence Metabolism: How Plants Cope with Pathogens and Ozone Pollution

Chemical defences represent a main trait of the plant innate immune system. Besides regulating the relationship between plants and their ecosystems, phytochemicals are involved both in resistance against pathogens and in tolerance towards abiotic stresses, such as atmospheric pollution. Plant defence metabolites arise from the main secondary metabolic routes, the phenylpropanoid, the isoprenoid and the alkaloid pathways. In plants, antibiotic compounds can be both preformed (phytoanticipins) and inducible (phytoalexins), the former including saponins, cyanogenic glycosides and glucosinolates. Chronic exposure to tropospheric ozone (O₃) stimulates the carbon fluxes from the primary to the secondary metabolic pathways to a great extent, inducing a shift of the available resources in favour of the synthesis of secondary products. In some cases, the plant defence responses against pathogens and environmental pollutants may overlap, leading to the unspecific synthesis of similar molecules, such as phenylpropanoids. Exposure to ozone can also modify the pattern of biogenic volatile organic compounds (BVOC), emitted from plant in response to herbivore feeding, thus altering the tritrophic interaction among plant, phytophagy and their natural enemies. Finally, the synthesis of ethylene and polyamines can be regulated by ozone at level of S-adenosylmethionine (SAM), the biosynthetic precursor of both classes of hormones, which can, therefore, mutually inhibit their own biosynthesis with consequence on plant phenotype.     

Turnabout is fair play: Secondary roles for primary compounds

Chemically based resistance of plants to herbivorous insects is today essentially synonymous with allelochemically based resistance; the importance of plant secondary compounds in determining patterns of host-plant utilization has been established in a wide variety of insect-plant interactions. In contrast, primary metabolites, those involved in fundamental plant physiological processes, are rarely considered to be major determinants of host-plant resistance despite the fact that, as insect nutrients, they can have profound effects on behavior and physiology. The degree to which variation in plant primary metabolism results from the selective impact of herbivory may be greatly underestimated in that the biosynthetic and structural diversity of primary metabolites and the consequences of that diversity on herbivores are rarely taken into account in most studies of insect preference and performance. Qualitative and quantitative variation in the production of primary metabolites can result from herbivore selection pressure if production of primary metabolites is under genetic control and if plant fitness in the presence of herbivores is associated in a predictable way with genetically based primary metabolite variation. Variation in primary metabolism is likely to be particularly effective as a defense against highly oligophagous herbivores with limited mobility, especially those confined to structures containing allelochemicals that could neutralize the benefits associated with compensatory feeding.     

Genomics-Guided Efficient Identification of 2,5-Diketopiperazine Derivatives from Actinobacteria

Secondary metabolites derived from microorganism constitute an important part of natural products. Mining of the microbial genomes revealed a large number of uncharacterized biosynthetic gene clusters, indicating their greater potential to synthetize specialized or secondary metabolites (SMs) than identified by classic fermentation and isolation approaches. Various bioinformatics tools have been developed to analyze and identify such gene clusters, thus accelerating significantly the mining process. Heterologous expression of an individual biosynthetic gene cluster has been proven as an efficient way to activate the genes and identify the encoded metabolites that cannot be detected under normal laboratory cultivation conditions. Herein, we describe a concept of genomics-guided approach by performing genome mining and heterologous expression to uncover novel CDPS-derived DKPs and functionally characterize novel tailoring enzymes embedded in the biosynthetic pathways. Recent works focused on the identification of the nucleobase-related and dimeric DKPs are also presented.     

The Potential of Plant Phenolics in Prevention and Therapy of Skin Disorders

Phenolic compounds constitute a group of secondary metabolites which have important functions in plants. Besides the beneficial effects on the plant host, phenolic metabolites (polyphenols) exhibit a series of biological properties that influence the human in a health-promoting manner. Evidence suggests that people can benefit from plant phenolics obtained either by the diet or through skin application, because they can alleviate symptoms and inhibit the development of various skin disorders. Due to their natural origin and low toxicity, phenolic compounds are a promising tool in eliminating the causes and effects of skin aging, skin diseases, and skin damage, including wounds and burns. Polyphenols also act protectively and help prevent or attenuate the progression of certain skin disorders, both embarrassing minor problems (e.g., wrinkles, acne) or serious, potentially life-threatening diseases such as cancer. This paper reviews the latest reports on the potential therapy of skin disorders through treatment with phenolic compounds, considering mostly a single specific compound or a combination of compounds in a plant extract.     

Chemical Ecology of Marine Angiosperms: Opportunities at the Interface of Marine and Terrestrial Systems

This review examines the state of the field for chemically mediated interactions involving marine angiosperms (seagrasses, mangroves, and salt marsh angiosperms). Small-scale interactions among these plants and their herbivores, pathogens, fouling organisms, and competitors are explored, as are community-level effects of plant secondary metabolites. At larger spatial scales, secondary metabolites from marine angiosperms function as reliable cues for larval settlement, molting, or habitat selection by fish and invertebrates, and can influence community structure and ecosystem function. Several recent studies illustrate the importance of chemical defenses from these plants that deter feeding by herbivores and infection by pathogens, but the extent to which allelopathic compounds kill or inhibit the growth of competitors is less clear. While some phenolic compounds such as ferulic acid and caffeic acid act as critical defenses against herbivores and pathogens, we find that a high total concentration of phenolic compounds within bulk plant tissues is not a strong predictor of defense. Residual chemical defenses prevent shredding or degradation of plant detritus by detritivores and microbes, delaying the time before plant matter can enter the microbial loop. Mangroves, marsh plants, and seagrasses remain plentiful sources of new natural products, but ecological functions are known for only a small proportion of these compounds. As new analytical techniques are incorporated into ecological studies, opportunities are emerging for chemical ecologists to test how subtle environmental cues affect the production and release of marine angiosperm chemical defenses or signaling molecules. Throughout this review, we point to areas for future study, highlighting opportunities for new directions in chemical ecology that will advance our understanding of ecological interactions in these valuable ecosystems.     

Comparison of Animal and Plant DNA-Adducts after Exposure to Benzo(a)pyrene

The occurrence of abnormal hypermodified nucleotides on the DNA upon xenobiotic exposure has long been considered as a characteristic of carcinogenesis and mutagenesis in animal cells. We have previously shown that DNA adducts could also be formed in plants exposed to xenobiotics in natural and controlled conditions. In this study we have compared the DNA adducts formed in different animal species and in different plants after benzo(a)pyrene (B(a)P) exposure.

The main DNA adduct in mice stomach and skin correspond to the 7,8-diol 9,10-epoxide B(a)P-guanine. In liver from rat, fish and Xenopus, this adduct is detected, but is not the major one. In plants analyzed, this adduct is never formed. Ten different adducts are detected in plants. This result indicates that the metabolic pathway leading to genotoxic metabolites is different with species. In conclusion, the result suggest that risk assessment for human and environment due to genotoxic compounds should be realized using multiple species assays.     

Sulfur-Containing Secondary Metabolites from Arabidopsis thaliana and other Brassicaceae with Function in Plant Immunity

Biosynthesis of antimicrobial secondary metabolites in response to microbial infection is one of the features of the plant immune system. Particular classes of plant secondary metabolites involved in plant defence are often produced only by species belonging to certain phylogenetic clades. Brassicaceae plants have evolved the ability to synthesise a wide range of sulfur-containing secondary metabolites, including glucosinolates and indole-type phytoalexins. A subset of these compounds is produced by the model plant Arabidopsis thaliana. Genetic tools available for this species enabled verification of immune functions of glucosinolates and camalexin (A. thaliana phytoalexin), as well as characterisation of their respective biosynthetic pathways. Current knowledge of the biosynthesis of Brassicaceae sulfur-containing metabolites suggests that the key event in the evolution of these compounds is the acquisition of biochemical mechanisms originating from detoxification pathways into secondary metabolite biosynthesis. Moreover, it is likely that glucosinolates and Brassicaceae phytoalexins, traditionally considered as separate groups of compounds, have a common evolutionary origin and are interconnected on the biosynthetic level. This suggests that the diversity of Brassicaceae sulfur-containing phytochemicals reflect phylogenetic clade-specific branches of an ancient biosynthetic pathway.     

Quantitation of Multiple Sphingolipid Classes Using Normal and Reversed-Phase LC–ESI–MS/MS: Comparative Profiling of Two Cell Lines

Sphingolipids are an important class of compounds that regulate signal transduction and other vital cellular processes. Herein, we report sensitive normal and reversed phase LC–MS/MS methods for quantitation of multiple sphingolipid classes. In the normal-phase ESI/MS/MS method, a high content of organic solvents was utilized, which, although it included hexane, ethyl acetate, acetonitrile containing 2% methanol, 1–2% acetic acid, and 5 mM ammonium acetate, resulted in a very efficient electrospray ionization of the ceramides (Cers) and hexosylceramides (MHCers). Three normal-phase LC–MS/MS methods using segmented phases were developed to specifically target Cers, MHCers, or sphingomyelins (SMs). This segmentation scheme increases the number of data points acquired for a given analyte and enhances the sensitivity and specificity of the measurements. Nine separate reversed phase chromatography methods were developed for the three classes of compounds. These assays were used for comparing the levels of Cers, SMs, and MHCers from mouse embryonic fibroblast (pMEF) and human embryonic kidney (HEK293) cells. These findings were then compared with the reported data from RAW264.7 mouse macrophage cells, BHK21 hamster cells, and human plasma and serum samples. The analysis of cell lines, using both normal and reversed phase chromatography, revealed discrimination based on the type of chromatography chosen, while sphingolipid assays of samples containing different amounts of protein showed different results, even after normalizing for protein content. Also, LC/MS/MS profiles were provided for the classes and individual compounds so that they could be used as “molecular profiles” for class or individual sample analysis.     

Foraging in the Dark – Chemically Mediated Host Plant Location by Belowground Insect Herbivores

Root-feeding insects are key components in many terrestrial ecosystems. Like shoot-feeding insect herbivores, they exploit a range of chemical cues to locate host plants. Respiratory emissions of carbon dioxide (CO(2)) from the roots is widely reported as the main attractant, however, there is conflicting evidence about its exact role. CO(2) may act as a 'search trigger' causing insects to search more intensively for more host specific signals, or the plant may 'mask' CO(2) emissions with other root volatiles thus avoiding detection. At least 74 other compounds elicit behavioral responses in root-feeding insects, with the majority (>80?%) causing attraction. Low molecular weight compounds (e.g., alcohols, esters, and aldehydes) underpin attraction, whereas hydrocarbons tend to have repellent properties. A range of compounds act as phagostimulants (e.g., sugars) once insects feed on roots, whereas secondary metabolites often deter feeding. In contrast, some secondary metabolites usually regarded as plant defenses (e.g., dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA)), can be exploited by some root-feeding insects for host location. Insects share several host location cues with plant parasitic nematodes (CO(2), DIMBOA, glutamic acid), but some compounds (e.g., cucurbitacin A) repel nematodes while acting as phagostimulants to insects. Moreover, insect and nematode herbivory can induce exudation of compounds that may be mutually beneficial, suggesting potentially significant interactions between the two groups of herbivores. While a range of plant-derived chemicals can affect the behavior of root-feeding insects, little attempt has been made to exploit these in pest management, though this may become a more viable option with diminishing control options.     

Species-Specific Interactions between Plant Metabolites and Insect Juvenile Hormone Receptors

Because juvenile hormone (JH) controls insect development and its analogs are used as insecticides, juvenile hormone disruptors (JHDs) represent potential sources from which novel pesticides can be developed. Many plant species harbor JHD activity, which has previously been attributed plant secondary metabolites (i.e., diterpenes) that disrupt insect development by interfering with the JH-mediated heterodimer formation of insect juvenile receptor complexes. The results of the present study indicate that plant JHD activity is also concentrated in certain plant groups and families and that plant metabolites have insect group-specific activity. These findings suggest that reciprocal diversification has occurred between plants and insects through the evolution of the plant metabolites and JH receptors, respectively, and that plant metabolites could be developed into insect group-specific pesticides with limited effects on non-target species.     

Big effects from small changes: possible ways to explore nature's chemical diversity

Fungi or bacteria that produce secondary metabolites often have the potential to bring up various compounds from a single strain. The molecular basis for this well-known observation was confirmed in the last few years by several sequencing projects of different microorganisms. Besides well-known examples about induction of a selected biosynthesis (for example, by high- or low-phosphate cultivation media), no overview about the potential in this field for finding natural products was given. We have investigated the systematic alteration of easily accessible cultivation parameters (for example, media composition, aeration, culture vessel, addition of enzyme inhibitors) in order to increase the number of secondary metabolites available from one microbial source. We termed this way of revealing nature's chemical diversity the 'OSMAC (One Strain-Many Compounds) approach' and by using it we were able to isolate up to 20 different metabolites in yields up to 2.6 g L(-1) from a single organism. These compounds cover nearly all major natural product families, and in some cases the high production titer opens new possibilities for semisynthetic methods to enhance even more the chemical diversity of selected compounds. The OSMAC approach offers a good alternative to industrial high-throughput screening that focuses on the active principle in a distinct bioassay. In consequence, the detection of additional compounds that might be of interest as lead structures in further bioassays is impossible and clearly demonstrates the deficiency of the industrial procedure. Furthermore, our approach seems to be a useful tool to detect those metabolites that are postulated to be the final products of an amazing number of typical secondary metabolite gene clusters identified in several microorganisms. If one assumes a (more or less) defined reservoir of genetic possibilities for several biosynthetic pathways in one strain that is used for a highly flexible production of secondary metabolites depending on the environment, the OSMAC approach might give more insight into the role of secondary metabolism in the microbial community or during the evolution of life itself.     

Larval Beetles Form a Defense from Recycled Host-Plant Chemicals Discharged as Fecal Wastes

Larvae of the leaf-feeding beetles Neolema sexpunctata and Lema trilinea carry feces on their backs that form shields. We used the generalist predatory ant, Formica subsericea, in a bioassay to determine whether shields were a physical barrier or functioned as a chemical defense. Fecal shields protected both species against ant attack. Larvae of both species reared on lettuce produced fecal shields that failed to deter ants. Commelina communis, N. sexpunctata's host, lacks noxious secondary compounds but is rich in phytol and fatty acids, metabolites of which become incorporated into the fecal defense. In contrast, the host plant of L. trilinea, Solanum dulcamara, contains steroidal glycoalkaloids and saponins, whose partially deglycosylated metabolites, together with fatty acids, appear in Lema feces. Both beetle species make modifications to host-derived precursors before incorporating the metabolites into shields. Synthetic chemicals identified as shield metabolites were deterrent when applied to baits. This study provides experimental evidence that herbivorous beetles form a chemical defense by the elimination of both primary and secondary host-derived compounds. The use of host-derived compounds in waste-based defenses may be a more widely employed strategy than was hitherto recognized, especially in instances where host plants lack elaborate secondary compounds.     

Arbuscular Mycorrhizal Fungi and Plant Growth-Promoting Pseudomonads Increases Anthocyanin Concentration in Strawberry Fruits (Fragaria x ananassa var. Selva) in Conditions of Reduced Fertilization

Anthocyanins are a group of common phenolic compounds in plants. They are mainly detected in flowers and fruits, are believed to play different important roles such as in the attraction of animals and seed dispersal, and also in the increase of the antioxidant response in tissues directly or indirectly affected by biotic or abiotic stress factors. As a major group of secondary metabolites in plants commonly consumed as food, they are of importance in both the food industry and human nutrition. It is known that arbuscular mycorrhizal (AM) fungi can influence the plant secondary metabolic pathways such as the synthesis of essential oils in aromatic plants, of secondary metabolites in roots, and increase flavonoid concentration. Plant Growth-Promoting Bacteria (PGPB) are able to increase plant growth, improving plant nutrition and supporting plant development under natural or stressed conditions. Various studies confirmed that a number of bacterial species living on and inside the root system are beneficial for plant growth, yield and crop quality. In this work it is shown that inoculation with AM fungi and/or with selected and tested Pseudomonas strains, under conditions of reduced fertilization, increases anthocyanin concentration in the fruits of strawberry.     

Improved Release and Metabolism of Flavonoids by Steered Fermentation Processes: A Review

This paper provides an overview on steered fermentation processes to release phenolic compounds from plant-based matrices, as well as on their potential application to convert phenolic compounds into unique metabolites. The ability of fermentation to improve the yield and to change the profile of phenolic compounds is mainly due to the release of bound phenolic compounds, as a consequence of the degradation of the cell wall structure by microbial enzymes produced during fermentation. Moreover, the microbial metabolism of phenolic compounds results in a large array of new metabolites through different bioconversion pathways such as glycosylation, deglycosylation, ring cleavage, methylation, glucuronidation and sulfate conjugation, depending on the microbial strains and substrates used. A whole range of metabolites is produced, however metabolic pathways related to the formation and bioactivities, and often quantification of the metabolites are highly underinvestigated. This strategy could have potential to produce extracts with a high-added value from plant-based matrices.     

The Dynamics of Carbon–Nutrient Balance: Effects of Cottonwood Acclimation to Short- and Long-Term Shade on Beetle Feeding Preferences

The carbon–nutrient balance hypothesis (CNBH) predicts that shading should increase leaf palatability to herbivores by decreasing concentrations of carbon (C) -based chemical defenses and increasing nitrogen (N). We measured cottonwood (Populus deltoids) growth, leaf chemistry, and beetle (Plagiodera versicolora) feeding preferences on saplings grown in either continuous high (HH) or low (LL) light, and saplings switched from high to low (HL) or low to high (LH) light for nine days. As expected, based on the CNBH, shading increased total N and decreased total phenol glycoside (C-based secondary metabolites) concentrations in plants from all shade treatments (LL, HL, and LH), relative to HH plants, with plant growth and gross leaf chemistry being affected by initial and final light regime. In contrast, while specific phenol glycoside concentrations were affected by the initial and final light regime, they also showed an initial × final light interaction. Beetles tended to prefer LL to HH plants. Beetles unexpectedly preferred HH to either HL or LH switched plants, most likely because high concentrations of a specific phenol glycoside – salicin – occurred in both switched treatments and inhibited beetle feeding. Plant chemical allocation during light acclimation led to unpredictable changes in specific C-based compounds, even though plant growth and gross chemistry conformed to expectations for shading effects and the CNBH. The response of this herbivore to altered concentrations of a specific compound confounded predictions based on average dynamics of suites of chemicals. Our findings may help explain why relationships between light availability and herbivory in field studies, where light varies on many time scales, can differ from those predicted by the CNBH. Understanding both dynamic plant chemical responses to altered resource availability and controls over allocation to specific compounds would likely enhance future predictability of specific environment-plant-herbivore interactions.     

Plant Secondary Metabolites as Rodent Repellents: a Systematic Review

The vast number of plant secondary metabolites (PSMs) produced by higher plants has generated many efforts to exploit their potential for pest control. We performed a systematic literature search to retrieve relevant publications, and we evaluated these according to PSM groups to derive information about the potential for developing plant-derived rodent repellents. We screened a total of 54 publications where different compounds or plants were tested regarding rodent behavior/metabolism. In the search for widely applicable products, we recommend multi-species systematic screening of PSMs, especially from the essential oil and terpenoid group, as laboratory experiments have uniformly shown the strongest effects across species. Other groups of compounds might be more suitable for the management of species-specific or sex-specific issues, as the effects of some compounds on particular rodent target species or sex might not be present in non-target species or in both sexes. Although plant metabolites have potential as a tool for ecologically-based rodent management, this review demonstrates inconsistent success across laboratory, enclosure, and field studies, which ultimately has lead to a small number of currently registered PSM-based rodent repellents.     

In Vitro Metabolism of Six C-Glycosidic Flavonoids from Passiflora incarnata L.

Several medical plants, such as Passiflora incarnata L., contain C-glycosylated flavonoids, which may contribute to their efficacy. Information regarding the bioavailability and metabolism of these compounds is essential, but not sufficiently available. Therefore, the metabolism of the C-glycosylated flavones orientin, isoorientin, schaftoside, isoschaftoside, vitexin, and isovitexin was investigated using the Caco-2 cell line as an in vitro intestinal and epithelial metabolism model. Isovitexin, orientin, and isoorientin showed broad ranges of phase I and II metabolites containing hydroxylated, methoxylated, and sulfated compounds, whereas schaftoside, isoschaftoside, and vitexin underwent poor metabolism. All metabolites were identified via UHPLC-MS or UHPLC-MS/MS using compound libraries containing all conceivable metabolites. Some structures were confirmed via UHPLC-MS experiments with reference compounds after a cleavage reaction using glucuronidase and sulfatase. Of particular interest is the observed cleavage of the C–C bonds between sugar and aglycone residues in isovitexin, orientin, and isoorientin, resulting in unexpected glucuronidated or sulfated luteolin and apigenin derivatives. These findings indicate that C-glycosidic flavones can be highly metabolized in the intestine. In particular, flavonoids with ortho-dihydroxy groups showed sulfated metabolites. The identified glucuronidated or sulfated aglycones demonstrate that enzymes expressed by Caco-2 cells are able to potentially cleave C–C bonds in vitro.