RESUMEN

Taraxacum officinale, a cosmopolitan perennial, is widely used as an edible, fodder, honey, and medicinal plant. Interestingly, detailed LC-QTOF-MS profiling of the flower extract revealed the presence of several flavonoid signals, most likely lacking literature confirmation. Targeted isolation, including extraction based on DoE planning, led to the isolation of 11 flavonoids, three of which were new (16, 18, and 33). Their structures were determined by the NMR technique. Both biflavones (16 and 18) were structured by two luteolin molecules, linked by a C-C bond through IB (C-6') and IIA (C-6″ or C-8″) rings. Novel flavonolignan (33) was composed of tricin and an unusual carboxyl function containing a lignan moiety. The content of 28 flavonoids (glycosides, aglycones, biflavones, and flavonolignans) was determined by the LC-MS/MS method in raw flowers and dandelion syrups and tincture. Multivariate analyses showed the separation of spring and autumn flowers and a high diversity of food products and helped to identify metabolites correlated with the samples.

Asunto(s)

RESUMEN

Herbal chemicals with a long history in medicine have attracted a lot of attention. Flavonolignans and flavonoids are considered as two classes of the above-mentioned compounds with different functional groups which exhibit several therapeutic capabilities such as antimicrobial, anti-inflammatory, antioxidant, antidiabetic, and anticancer activities. Based on the studies, high hydrophobic properties of the aforementioned compounds limit their bioavailability inside the human body and restrict their wide application. Nanoscale formulations such as solid lipid nanoparticles, liposomes, and other types of lipid-based delivery systems have been introduced to overcome the above-mentioned challenges. This approach allows the aforementioned hydrophobic therapeutic compounds to be encapsulated between hydrophobic structures, resulting in improving their bioavailability. The above-mentioned enhanced delivery system improves delivery to the targeted sites and reduces the daily required dosage. Lowering the required daily dose improves the performance of the drug by diminishing its side effects on non-targeted tissues. The present study aims to highlight the recent improvements in implementing lipid-based nanocarriers to deliver flavonolignans and flavonoids.

RESUMEN

Plant seeds, as those from milk thistle (Silybum marianum), are a valuable source of nonpolar and polar compounds with potentially interesting biological activity. The main nonpolar compounds are triglycerides, which are also the main components of all vegetable oils. In addition, specific polar compounds - flavonolignans, called silymarin, have been found in large amounts in milk thistle seeds extract. These flavonoids derivatives have different biological activity, for instance hepatoprotective effects. In order to extract and analyze both nonpolar (triglycerides) and polar compounds (flavonolignans) from milk thistle seeds through a sequential methodology, an on-line supercritical fluid extraction - supercritical fluid chromatography (SFE-SFC) method was developed. Different ways of transferring the extracts from SFE to SFC (i.e. direct on-column transfer and loop transfer) were compared, and particularly for their effect on chromatographic quality. In this respect, nonpolar and polar compounds caused different issues, especially as polar compounds required a significant portion of co-solvent in the extraction step, favoring early elution in the chromatographic column. First, on-line SFE-SFC was used for triglycerides analysis and allowed the comparison of transfer modes. Then, on-line kinetics were performed to measure defatting time before polar molecules extraction. Finally, the eventual benefit of loop transfer was also investigated for the analysis of flavonolignans, polar molecules whose analysis can be difficult by on-line SFE-SFC. The aim of this paper is to discuss the versatility of on-line SFE-SFC and how challenging the coupling can be, especially when both non-polar and polar molecules must be analyzed independently in a single sample.

Asunto(s)

RESUMEN

Silybum marianum (SM) belongs to the family Asteraceae, which holds therapeutic significance in medicinal chemistry. Phytochemistry analysis revealed an abundance of active constituents, particularly silymarin, composed of polyphenols and flavonolignans. Silymarin is majorly found in leaves, seeds, and fruits and is comprised of seven flavonolignans. Silymarin derivatives, specifically silybin, were reported for their medicinal properties. This review summarizes the studies conducted to evaluate SM's pharmacological properties and proposed mechanisms. SM exhibited anticancer properties due to being capable of modifying the induction of apoptosis, inhibiting the STAT3 pathway, decreasing the transcription of various growth factors, impeding the growth of 4T1 cells and inducing cell cycle arrest in various types of cancers, i.e., skin cancer, liver cancer, breast cancer, ovarian cancer etc. Silymarin and its derivatives protect the liver and ameliorate various immune-mediated and autoimmune hepatic diseases. Moreover, antimicrobial, antidiabetic, cardioprotective, nephroprotective, and neuroprotective activities were also reported. Based on testified in vitro and in vivo studies, SM can serve as an alternative to cure various pathological ailments.

Asunto(s)

RESUMEN

A library of previously unknown halogenated derivatives of flavonolignans (silybins A and B, 2,3-dehydrosilybin, silychristin A, and 2,3-dehydrosilychristin A) was prepared. The effect of halogenation on the biological activity of flavonolignans was investigated. Halogenated derivatives had a significant effect on bacteria. All prepared derivatives inhibited the AI-2 type of bacterial communication (quorum sensing) at concentrations below 10 µM. All prepared compounds also inhibited the adhesion of bacteria (Staphyloccocus aureus and Pseudomonas aeruginosa) to the surface, preventing biofilm formation. These two effects indicate that the halogenated derivatives are promising antibacterial agents. Moreover, these derivatives acted synergistically with antibiotics and reduced the viability of antibiotic-resistant S. aureus. Some flavonolignans were able to reverse the resistant phenotype to a sensitive one, implying that they modulate antibiotic resistance.

Asunto(s)

RESUMEN

Two pairs of new enantiomeric flavonolignans, ±stachyols A and B (±1 and ± 2), along with two novel isoflavanelignans, stachyols C and D (3 and 4) were isolated from the roots of Indigofera stachyodes. Their chemical structures and absolute configurations were determined using nuclear magnetic resonance and comparison of experimental and theoretical electronic circular dichroism (ECD) spectra, as well as quantum chemical calculations. Of those compounds, 1 and 2 represented the first examples of flavonolignans with 5-deoxyflavonoids adduct phenylpropanoids. Moreover, 3 and 4 possess an unprecedented skeleton with isoflavanes adduct phenylpropanoids. The antioxidant activity was evaluated for all compounds in terms of ABTS+ and DPPH bioassays. Compounds 3 and 4 exhibited significant radical-scavenging activity in the ABTS+ assay, with IC50 values of 15.15 and 5.83 µM, respectively.

Asunto(s)

RESUMEN

Salinity is an important challenge around the world, effecting all physiological and biochemical processes of plants. It seems that seed priming can diminish the negative impacts of salinity. To study the effects of hydro-priming and inoculation with Piriformospora indica (Pi) and Azotobacter chroococcum (Az) on physio-biochemical traits, flavonolignans and fatty acids composition of milk thistle under saline conditions, a greenhouse experiment was carried out. Our results indicated that under salinity, seed priming, especially Pi, improved physio-biochemical properties in milk thistle. Under 120 mM NaCl, inoculation with Pi increased membrane stability index (MSI) and relative water content (RWC) (by 21.86 and 33.43%, respectively). However, peroxidase (POX) (5.57- and 5.68-fold in roots and leaves, respectively), superoxide dismutase (SOD) (4.74- and 4.44-fold in roots and leaves, respectively), catalase (CAT) (6.90- and 8.50-fold in roots and leaves, respectively) and ascorbate peroxidase (APX) (5.61- and 5.68-fold in roots and leaves, respectively) activities increased with increasing salinity. Contrary to salinity, hydro-priming with Az and Pi positively altered all these traits. The highest content of the osmolytes, adenosine triphosphate (ATP) content and rubisco activity were recorded in Pi treatments under 120 mM NaCl. Stearic acid (20.24%), oleic acid (21.06%) and palmitic acid (10.48%) increased, but oil content (3.81%), linolenic and linoleic acid content (22.21 and 15.07%, respectively) decreased under saline conditions. Inoculations of Pi positively altered all these traits. The present study indicated that seed priming with Pi under 120 mM NaCl resulted in maximum silychristin, taxidolin, silydianin, isosilybin, silybin and silymarin of milk thistle seeds.

RESUMEN

Antibiotic resistance is currently a serious health problem. Since the discovery of new antibiotics no longer seems to be a sufficient tool in the fight against multidrug-resistant infections, adjuvant (combination) therapy is gaining in importance as well as reducing bacterial virulence. Silymarin is a complex of flavonoids and flavonolignans known for its broad spectrum of biological activities, including its ability to modulate drug resistance in cancer. This work aimed to test eleven, optically pure silymarin flavonolignans for their ability to reverse the multidrug resistance phenotype of Staphylococcus aureus and reduce its virulence. Silybin A, 2,3-dehydrosilybin B, and 2,3-dehydrosilybin AB completely reversed antibiotic resistance at concentrations of 20 µM or less. Both 2,3-dehydrosilybin B and AB decreased the antibiotic-induced gene expression of representative efflux pumps belonging to the major facilitator (MFS), multidrug and toxic compound extrusion (MATE), and ATP-binding cassette (ABC) families. 2,3-Dehydrosilybin B also inhibited ethidium bromide accumulation and efflux in a clinical isolate whose NorA and MdeA overproduction was induced by antibiotics. Most of the tested flavonolignans reduced cell-to-cell communication on a tetrahydrofuran-borate (autoinducer-2) basis, with isosilychristin leading the way followed by 2,3-dehydrosilybin A and AB, which halved communication at 10 µM. Anhydrosilychristin was the only compound that reduced communication based on acyl-homoserine lactone (autoinducer 1), with an IC50 of 4.8 µM. Except for isosilychristin and anhydrosilychristin, all of the flavonolignans inhibited S. aureus surface colonization, with 2,3-dehydrosilybin A being the most active (IC50 10.6 µM). In conclusion, the selected flavonolignans, particularly derivatives of 2,3-dehydrosilybin B, 2,3-dehydrosilybin AB, and silybin A are non-toxic modulators of S. aureus multidrug resistance and can decrease the virulence of the bacterium, which deserves further detailed research.

Asunto(s)

RESUMEN

CE method for the baseline separation of structurally similar flavonolignans silybin A, silybin B, isosilybin A, isosilybin B, silychristin, silydianin, and their precursor taxifolin in silymarin complex has been developed and validated. The optimized background electrolyte was 100 mmol/L boric acid (pH 9.0) containing 5 mmol/L heptakis(2,3,6-tri-O-methyl)-ß-CD and 10% (v/v) of methanol. The separation was carried out in an 80.5/72 cm (50 µm id) fused silica capillary at +25 kV with UV detection at 200 nm. Genistein (10 µg/mL) was used as internal standard. The resolution between the diastereomers of silybin and isosilybin was 1.73 and 2.59, respectively. The method was validated for each analyte in a concentration range of 2.5-50 µg/mL. The calibration curves were rectilinear with correlation coefficients ≥0.9972. The method was applied to determine flavonolignans in two dietary supplements containing Silybum marianum extract. The accuracy was evaluated by comparing the results of the CE analyses of the dietary supplements with those of the reference United States Pharmacopeial HPLC method. The unpaired t-test did not show a statistically significant difference between the results of both the proposed CE and the reference method (p > 0.05, n = 3).

Asunto(s)

RESUMEN

OBJECTIVES: Plant-derived flavonolignans had been demonstrated to have various biological functions. They are an important class of natural products combined by a flavonoid unit and a phenylpropanoid unit. KEY FINDINGS: From the literature survey, 88 constituents from natural resources were identified. Different derivatives of flavonolignans were listed, fused phenylpropanoid unit with dioxane ring, or cyclic ether, or simple ether side chain, or lactone, and so on. Besides, the pharmacological effects of flavonolignans were summarized as well. It has a wide range of anti-tumour, antioxidant, anti-microorganic and anti-inflammatory effects. SUMMARY: This review had provided a full-scale profile of flavonolignans on its plant sources, phytochemistry and pharmacology, and also proposed some issues and perspectives which may be of concern in the future. It was greatly anticipated that the commercialization of the flavonolignans would lead to uplift the financial abilities of communities attending the growing of the flavonolignans and the relevant and potential production becoming an international herbal and pharmaceutical commodity.

Asunto(s)

RESUMEN

Silymarin is known for its hepatoprotective effects. Although there is solid evidence for its protective effects against Amanita phalloides intoxication, only inconclusive data are available for alcoholic liver damage. Since silymarin flavonolignans have metal-chelating activity, we hypothesized that silymarin may influence alcoholic liver damage by inhibiting zinc-containing alcohol dehydrogenase (ADH). Therefore, we tested the zinc-chelating activity of pure silymarin flavonolignans and their effect on yeast and equine ADH. The most active compounds were also tested on bovine glutamate dehydrogenase, an enzyme blocked by zinc ions. Of the six flavonolignans tested, only 2,3-dehydroderivatives (2,3-dehydrosilybin and 2,3-dehydrosilychristin) significantly chelated zinc ions. Their effect on yeast ADH was modest but stronger than that of the clinically used ADH inhibitor fomepizole. In contrast, fomepizole strongly blocked mammalian (equine) ADH. 2,3-Dehydrosilybin at low micromolar concentrations also partially inhibited this enzyme. These results were confirmed by in silico docking of active dehydroflavonolignans with equine ADH. Glutamate dehydrogenase activity was decreased by zinc ions in a concentration-dependent manner, and this inhibition was abolished by a standard zinc chelating agent. In contrast, 2,3-dehydroflavonolignans blocked the enzyme both in the absence and presence of zinc ions. Therefore, 2,3-dehydrosilybin might have a biologically relevant inhibitory effect on ADH and glutamate dehydrogenase.

Asunto(s)

RESUMEN

BACKGROUND: Flavonolignans like silybin, hydnocarpin, and siliandrin are a group of natural compounds combining the structural moieties of flavonoid and phenylpropanoid (lignan). Hydnocarpin and silandrin have been less explored because of their trace occurrence in nature. OBJECTIVE: The present study aimed at chemical conversion of silybin to hydnocarpin and siliandrin. Another objective was to synthesize a series of amide derivatives and biologically evaluate them with regard to their anti-cancer effects. METHODS: In order to selectively convert silybin to 23-iodo silybin, 23-iodo hydnocarpin D and 23- iodo isosilandrin, the ratio of Ph3P, imidazole and molecular iodine was meticulously adjusted. These three iodide compounds were converted into amide compounds by chemical transformation. MTT method was applied to evaluate their anti-cancer potency. The binding affinity to related proteins was calculated by molecular docking. RESULTS: A total of 45 new amido-derivatives were synthesized and structurally characterized by NMR and HRMS. Some of them showed moderate to good antiproliferative potency against cancer cells. The activity of compound 10j was further testified by colony formation assay and molecular docking. CONCLUSION: The synthesis of 23-iodo silybin, 23-iodo hydnocarpin D and 23-iodo isosilandrin from silybin was successfully accomplished by one simple iodination reaction. Some of the amide derivatives of sylibin/hydnocarpin D /silandrin exhibited a remarkable inhibitory effect of proliferation on cancer cells compared to silybin. These results would pave the way for further investigation on the derivatives of flavonolignans for the treatment of cancer.

Asunto(s)

RESUMEN

The aim of this study was to investigate secondary metabolite production in Silybum marianum L. cell suspension cultures obtained from seeds treated with gamma rays (200 and 600 Gy) and 0.05% colchicine. The effects of these treatments on callus induction, growth, viability, and silymarin production were studied, along with the changes in the transcriptome and DNA sequence of chalcone synthase (CHS) genes. The effect of gamma radiation (200 and 600 Gy) on silymarin production in S. marianum dry seeds was also studied using HPLC-UV. All three treatments induced high callus biomass production from leaf segments. The viability of the cell suspension cultures was over 90%. The flavonolignan content measured in the extracellular culture medium of the S. marianum cell suspension was highest after treatment with 600 Gy, followed by 0.05% colchicine, and finally, 200 Gy, after a growth period of 12 days. In general, an increased expression of CHS1, CHS2, and CHS3 genes, accompanied by an increase of silymarin content, was observed in response to all the studied treatments, although the effect was greatest on CHS2 expression. Bioinformatics analysis confirmed that the three CHS2 clones exhibited the highest genetic variation, both in relation to each other and to the CHS1 and CHS3 clones. Based on the results, S. marianum plants obtained from seeds previously exposed to 600 and 200 Gy as well as colchicine constitute a renewable resource with the potential to obtain large amounts of silymarin.

Asunto(s)

RESUMEN

2,3-Dehydrosilybin (DHS) was previously shown to chelate and reduce both copper and iron ions. In this study, similar experiments with 2,3-dehydrosilychristin (DHSCH) showed that this congener of DHS also chelates and reduces both metals. Statistical analysis pointed to some differences between both compounds: in general, DHS appeared to be a more potent iron and copper chelator, and a copper reducing agent under acidic conditions, while DHSCH was a more potent copper reducing agent under neutral conditions. In the next step, both DHS and DHSCH were tested for metal-based Fenton chemistry in vitro using HPLC with coulometric detection. Neither of these compounds were able to block the iron-based Fenton reaction and, in addition, they mostly intensified hydroxyl radical production. In the copper-based Fenton reaction, the effect of DHSCH was again prooxidant or neutral, while the effect of DHS was profoundly condition-dependent. DHS was even able to attenuate the reaction under some conditions. Interestingly, both compounds were strongly protective against the copper-triggered lysis of red blood cells, with DHSCH being more potent. The results from this study indicated that, notwithstanding the prooxidative effects of both dehydroflavonolignans, their in vivo effect could be protective.

RESUMEN

The present study aimed to investigate the enzymatic potential of Silybum marianum leaves to bioconvert phenolic acids produced in S. marianum callus into silymarin derivatives as chemopreventive agent. Here we demonstrate that despite the fact that leaves of S. marianum did not accumulate silymarin themselves, expanding leaves had the full capacity to convert di-caffeoylquinic acid to silymarin complex. This was proven by HPLC separations coupled with electrospray ionization mass spectrometry (ESI-MS) analysis. Soaking the leaf discs with S. marianum callus extract for different times revealed that silymarin derivatives had been formed at high yield after 16 h. Bioconverted products displayed the same retention time and the same mass spectra (MS or MS/MS) as standard silymarin. Bioconversion was achieved only when using leaves of a specific age, as both very young and old leaves failed to produce silymarin from callus extract. Only medium leaves had the metabolic capacity to convert callus components into silymarin. The results revealed higher activities of enzymes of the phenylpropanoid pathway in medium leaves than in young and old leaves. It is concluded that cotyledon-derived callus efficiently produces compounds that can be bio-converted to flavonolignans in leaves tissue of S. marianum.

Asunto(s)

RESUMEN

Silybum marianum (L.) Gaertn. (Asteraceae), commonly known as milk thistle, is a botanical natural product used to self-treat multiple diseases such as Type 2 diabetes mellitus and nonalcoholic steatohepatitis (NASH). An extract from milk thistle seeds (achenes), termed silymarin, is comprised primarily of several flavonolignans. Systemic concentrations of these flavonolignans can influence the potential biologic effects of silymarin and the risk for pharmacokinetic silymarin-drug interactions. The aims of this research were to determine the roles of organic anion transporting polypeptides (OATPs/Oatps) in silymarin flavonolignan disposition and in pharmacokinetic silymarin-drug interactions. The seven major flavonolignans from silymarin were determined to be substrates for OATP1B1, OATP1B3, and OATP2B1. Sprague Dawley rats were fed either a control diet or a NASH-inducing diet and administered pitavastatin (OATP/Oatp probe substrate), followed by silymarin via oral gavage. Decreased protein expression of Oatp1b2 and Oatp1a4 in NASH animals increased flavonolignan area under the plasma concentration-time curve (AUC) and maximum plasma concentration. The combination of silymarin inhibition of Oatps and NASH-associated decrease in Oatp expression caused an additive increase in plasma pitavastatin AUC in the animals. These data indicate that OATPs/Oatps contribute to flavonolignan cellular uptake and mediate the interaction between silymarin and NASH on pitavastatin systemic exposure.

Asunto(s)

RESUMEN

In the last few decades, targeting cancer by the use of dietary phytochemicals has gained enormous attention. The plausible reason and believe or mind set behind this fact is attributed to either lesser or no side effects of natural compounds as compared to the modern chemotherapeutics, or due to their conventional use as dietary components by mankind for thousands of years. Silibinin is a naturally derived polyphenol (a flavonolignans), possess following biochemical features; molecular formula C25H22O10, Molar mass: 482.44 g/mol, Boiling point 793 °C, with strikingly high antioxidant and anti-tumorigenic properties. The anti-cancer properties of Silibinin are determined by a variety of cellular pathways which include induction of apoptosis, cell cycle arrest, inhibition of angiogenesis and metastasis. In addition, Silibinin controls modulation of the expression of aberrant miRNAs, inflammatory response, and synergism with existing anti-cancer drugs. Therefore, modulation of a vast array of cellular responses and homeostatic aspects makes Silibinin an attractive chemotherapeutic agent. However, like other polyphenols, the major hurdle to declare Silibinin a translational chemotherapeutic agent, is its lesser bioavailability. After summarizing the chemistry and metabolic aspects of Silibinin, this extensive review focuses on functional aspects governed by Silibinin in chemoprevention with an ultimate goal of summarizing the evidence supporting the chemopreventive potential of Silibinin and clinical trials that are currently ongoing, at a single platform.

Asunto(s)

RESUMEN

Silybum marianum (L.) Gaertn. is an important medicinal plant belonging to Mediterranean flora. The medicinal properties of the species are mainly due to silymarin, a combination of different flavonolignans contained in the fruit. As for silymarin, so far a wide variability of possible S. marianum chemotypes has been described. In the present study the flavonolignan profile of 40 different S. marianum wild accessions was analysed at both population and single plant level, further extending the analysis to progenies derived from crosses between parental lines with different chemotypes. The results of this work indicate that S. marianum wild populations can be composed either of individuals with the same chemotype, or heterogeneous mixtures of individuals characterized by different chemotypes. Only three chemotypes (A, B and C) have been identified among Italian wild populations. Based on data collected we furthermore propose that chemotype C is the result of the hybridization between A and B chemotypes. If assessed at single plant level, chemotypes are extremely stable therefore evidencing a strong genetic control of silymarin biosynthetic pathway. Chemotypes A and B are present in all the analysed regions and no clear correlation between chemotypes and geographic features has been found. In conclusion, this work provides a general procedure for the characterization of different and stable chemotypes, for a deeper understanding of silymarin biosynthetic pathway, and in order to implement S. marianum breeding programmes aiming to improve silymarin quality.

Asunto(s)

RESUMEN

Dandelion (Taraxacum officinale L.) roots, leaves, and flowers have a long history of use in traditional medicine. Compared to the above organs, dandelion fruits are the least known and used. Hence, the present paper was aimed at the phytochemical analysis of T. officinale fruit extract and estimating its antiradical, antiplatelet, and antioxidant properties related to hemostasis. Methanolic extract of fruits (E1), enriched with polyphenols (188 mg gallic acid equivalents (GAE)/g), was successfully separated into cinnamic acids (E2; 448 mg GAE/g) and flavonoids (E3; 377 mg GAE/g) extracts. Flavonoid extract was further divided into four fractions characterized by individual content: A (luteolin fraction; 880 mg GAE/g), B (philonotisflavone fraction; 516 mg GAE/g), C (flavonolignans fraction; 384 mg GAE/g), and D (flavone aglycones fraction; 632 mg GAE/g). High DPPH radical scavenging activity was evaluated for fractions A and B (A > B > Trolox), medium for extracts (Trolox > E3 > E2 > E1), and low for fractions C and D. No simple correlation between polyphenol content and antiradical activity was observed, indicating a significant influence of qualitative factor, including higher anti-oxidative effect of flavonoids with B-ring catechol system compared to hydroxycinnamic acids. No cytotoxic effect on platelets was observed for any dandelion preparation tested. In experiments on plasma and platelets, using several different parameters (lipid peroxidation, protein carbonylation, oxidation of thiols, and platelet adhesion), the highest antioxidant and antiplatelet potential was demonstrated by three fruit preparations-hydroxycinnamic acids extract (E2), flavonoid extract (E3), and luteolin fraction (A). The results of this paper provide new information on dandelion metabolites, as well as their biological potential and possible use concerning cardiovascular diseases.

Asunto(s)

RESUMEN

Mature fruits (i.e., achenes) of milk thistle (Silybum marianum (L.) Gaertn., Asteraceae) accumulate high amounts of silymarin (SILM), a complex mixture of bioactive flavonolignans deriving from taxifolin. Their biological activities in relation with human health promotion and disease prevention are well described. However, the conditions of their biosynthesis in planta are still obscure. To fill this gap, fruit development stages were first precisely defined to study the accumulation kinetics of SILM constituents during fruit ripening. The accumulation profiles of the SILM components during fruit maturation were determined using the LC-MS analysis of these defined developmental phases. The kinetics of phenylalanine ammonia-lyase (PAL), chalcone synthase (CHS) and peroxidase (POX) activities suggest in situ biosynthesis of SILM from l-Phenylalanine during fruit maturation rather than a transport of precursors to the achene. In particular, in contrast to laccase activity, POX activity was associated with the accumulation of silymarin, thus indicating a possible preferential involvement of peroxidase(s) in the oxidative coupling step leading to flavonolignans. Reference genes have been identified, selected and validated to allow accurate gene expression profiling of candidate biosynthetic genes (PAL, CAD, CHS, F3H, F3'H and POX) related to SILM accumulation. Gene expression profiles were correlated with SILM accumulation kinetic and preferential location in pericarp during S. marianum fruit maturation, reaching maximum biosynthesis when desiccation occurs, thus reinforcing the hypothesis of an in situ biosynthesis. This observation led us to consider the involvement of abscisic acid (ABA), a key phytohormone in the control of fruit ripening process. ABA accumulation timing and location during milk thistle fruit ripening appeared in line with a potential regulation of the SLIM accumulation. A possible transcriptional regulation of SILM biosynthesis by ABA was supported by the presence of ABA-responsive cis-acting elements in the promoter regions of the SILM biosynthetic genes studied. These results pave the way for a better understanding of the biosynthetic regulation of SILM during the maturation of S. marianum fruit and offer important insights to better control the production of these medicinally important compounds.

Asunto(s)