Novel procedures for olive leaves extracts processing: Selective isolation of oleuropein and elenolic acid.

Several processes have been developed in the past to selectively extract oleuropein and its aglycones from olive derived materials. In the present manuscript, we outline a novel approach for processing olive leaves aqueous extracts. This allowed first to select microwave irradiation as the methodology able to provide a large enrichment in oleuropein. Subsequently, the use of lamellar solids led to the selective and high yield concentration of the same. Adsorption on solids also largely contributed to the long term chemical stability of oleuropein. Finally, an eco-friendly, readily available, and reusable catalyst like H2SO4 supported on silica was applied for the hydrolysis of oleuropein into hydroxytyrosol and elenolic acid. This latter was in turn selectively isolated by an acid-base work-up providing its monoaldehydic dihydropyran form (7.8 % extractive yield), that was unequivocally characterized by GC-MS. The isolation of elenolic acid in pure form is described herein for the first time.

Solid-phase adsorption methodologies of naturally occurring anthraquinones: A review.

INTRODUCTION: Solid-phase extraction applied to plant matrices is nowadays a well-validated technique allowing to concentrate and purify different secondary metabolites. Several classes of phytochemicals have been selectively extracted by this methodology. During the last decade attention has been focused on biologically active anthraquinones from numerous sources like edible, healthy, and medicinal plants. OBJECTIVES: The aim of this review is to provide a detailed literature survey of the solid-phase adsorption methodologies for the extraction of natural anthraquinones reported so far and to discuss and propose future directions in this field of research. MATERIALS AND METHODS: Substructure search was performed in the SciFinder Scholar, PubMed, Medline, and Scopus databases. RESULTS: The first report about application of solid-phase adsorption for the purification of anthraquinones appeared in the literature in 2002. From this date, and in particular during recent years, the most notable examples included the use of chitin- and chitosan-based polymers, of molecularly imprinted polymers, of coated magnetic nanoparticles, of miniaturized matrix solid-phase dispersion, of functionalized resins, of differently structured lamellar solids, and finally of vortex-synchronized matrix solid-phase dispersion. CONCLUSIONS: The herein detailed solid-phase adsorption methodologies are powerful tools to selectively extract natural anthraquinones and/or provide anthraquinone-enriched phytopreparations. Nevertheless, many other important methods have been applied to synthetic anthraquinones (e.g., azo dyes). These could be conveniently employed also for natural anthranoids. Studies in this field are discussed in this review article.

Cognitive and biological effects of citrus phytochemicals in subjective cognitive decline: a 36-week, randomized, placebo-controlled trial.

BACKGROUND: Auraptene (AUR) and naringenin (NAR) are citrus-derived phytochemicals that influence several biological mechanisms associated with cognitive decline, including neuronal damage, oxidative stress and inflammation. Clinical evidence of the efficacy of a nutraceutical with the potential to enhance cognitive function in cohorts at risk of cognitive decline would be of great value from a preventive perspective. The primary aim of this study is to determine the cognitive effects of a 36-week treatment with citrus peel extract standardized in levels of AUR and NAR in older adults experiencing subjective cognitive decline (SCD). The secondary aim is to determine the effects of these phytochemicals on blood-based biomarkers indicative of neuronal damage, oxidative stress, and inflammation. METHODS: Eighty older persons with SCD will be recruited and randomly assigned to receive the active treatment (400 mg of citrus peel extract containing 0.1 mg of AUR and 3 mg of NAR) or the placebo at a 1:1 ratio for 36 weeks. The primary endpoint is a change in the Repeatable Battery for the Assessment of Neuropsychological Status score from baseline to weeks 18 and 36. Other cognitive outcomes will include changes in verbal and nonverbal memory, attention, executive and visuospatial functions. Blood samples will be collected from a consecutive subsample of 60 participants. The secondary endpoint is a change in interleukin-8 levels over the 36-week period. Other biological outcomes include changes in markers of neuronal damage, oxidative stress, and pro- and anti-inflammatory cytokines. CONCLUSION: This study will evaluate whether an intervention with citrus peel extract standardized in levels of AUR and NAR has cognitive and biological effects in older adults with SCD, facilitating the establishment of nutrition intervention in people at risk of cognitive decline. TRIAL REGISTRATION: The trial is registered with the United States National Library of Medicine at the National Institutes of Health Registry of Clinical Trials under the code NCT04744922 on February 9th, 2021 ( https://www. CLINICALTRIALS: gov/ct2/show/NCT04744922 ).

A novel and efficient concentration of pomegranate juice with enhanced antioxidant activity.

In the present study we investigated the performance of a panel of 13 solid sorbents comprising layered double hydroxides, zirconium phosphate-based materials, and phyllosilicates as heterogeneous supports for the concentration of pomegranate (Punica granatum L.) juice. Mg-containing clays exhibited an almost complete bleaching capacityof pomegranate juice and more interestingly provided blends with an increased antioxidant capacity (around 1.5-fold) respect to the parent juice when assayed for the Trolox equivalent antioxidant capacity (TEAC) coupled to ABTS decolorization test. Such an activity remained practically unaltered after 4 days during which the pomegranate concentrated preparations remained supported on clays. The approach investigated herein and used for the concentration of pomegranate juice and the discovery of the preservation for long periods of the antioxidant activities of pomegranate extracts when supported on solid sorbents have been reported herein for the first time in the literature to the best of our knowledge.

Pro-Osteogenic Properties of Violina pumpkin (Cucurbita moschata) Leaf Extracts: Data from In Vitro Human Primary Cell Cultures.

Traditional medicines rely mainly on use of plant extracts to mitigate or treat a wide range of disorders, including those that affect skeletal homeostasis. In this study, we investigated for the first time the potential pro-osteogenic effects of hexane, acetone and methanol extracts of the leaves of Cucurbita moschata, a very popular pumpkin cultivar in Western countries. We found that in Cucurbita moschata leaves, there are acetone-extractable substances-in particular, fatty acids such as 13-OH-9Z,11E,15E-octadecatrienoic acid (PU-13OH-FA), which is capable of both stimulating the function of human primary osteoblasts, which are responsible for bone formation, and inhibiting the differentiation of human osteoclasts, which are responsible for bone resorption. This dual effect was monitored by analyzing Runx2 expression, deposition of mineralized matrix, ALP activity, TRAP and actin ring staining respectively. This study suggests that bioactive chemicals from Cucurbita moschata leaves are potentially suitable as therapeutics for managing metabolic bone disorders such as osteoporosis and rheumatoid arthritis, and promoting tissue healing and functional recovery after bone fractures. The data we obtained increase knowledge on the biological activities of Cucurbita moschata, and in particular underline the potential benefits of consuming leaves which are a part of the plant currently little considered in the Western world.

Pre-concentration of active principles from different varieties of Camellia sinensis extracts by solid sorbents.

In this article we studied the phytochemical composition of leaves extracts of different varieties of Camellia sinensis(L.) Kuntze after treatment with 16 selected solid sorbents (namely hydrotalcites, magnesium oxide and hydroxide, zirconium phosphates, and phyllosilicates). The pre-concentration and selective adsorption of the main active principles of this food and medicinal plant [e.g. gallic acid, (-)-epicatechin, (-)-epicatechin gallate, and caffeine] were investigated. The quantities of phytochemicals adsorbed by solids were measured by HPLC analysis, coupled to photodiode array detection and calculated as the difference between the quantities in the parent untreated extracts and those recorded in the filtrates. Caffeine was selectively adsorbed by bentonite to a large extent, while for the remaining phytochemicals different patterns were recorded depending on the type of leaves extract. A comparison with pure chemicals revealed a strong effect of the phytocomplex composition on the adsorption yields. The methodology outlined herein may be useful to obtain tea extracts enriched in selective active principles also for industrial scopes.

7-Isopentenyloxycoumarin: What Is New across the Last Decade.

7-Isopentenyloxycoumarin is among the most widespread naturally occurring prenyloxy umbelliferone derivatives. This secondary metabolite of mixed biosynthetic origin has been typically isolated from plants belonging to several genera of the Rutaceae and Apiaceae families, comprising widely used medicinal plants and in general plants with beneficial effects on human welfare, as well as edible fruits and vegetables. Although known for quite a long time (more than 50 years), only in the last two decades has this natural compound been revealed to exert powerful and promising pharmacological properties, such as active cancer chemopreventive, antibacterial, antiprotozoal, antifungal, anti-inflammatory, neuroprotective, and antioxidant properties, among the activities best outlined in the recent literature. The aim of this comprehensive miniature review article is to detail the novel natural sources and the effects described during the last decade for 7-isopentenyloxycoumarin and what has been reported on the mechanisms of action underlying the observed biological activities of this oxyprenylated secondary metabolite. In view of the herein described data, suggestions on how to address future research on the abovementioned natural product and structurally related derivatives in the best ways according to the authors will be also provided.

Solid phase adsorption of anthraquinones from plant extracts by lamellar solids.

Solid phase extraction is nowadays a well validated and powerful technique applicable to complex matrices like plant extracts and phytocomplexes. This process provides concentration and/or purification of selected secondary metabolites from these matrices for subsequent analysis and isolation. In this research article sixteen lamellar solids, comprising layered structures (hydrotalcites, zirconium phosphates, magnesium hydroxide), magnesium oxide, and the phyllosilicates talc and bentonite were investigated for their capacity and performance to selectively adsorb five naturally occurring and widespread anthraquinones (aloe, aloe-emodin, rhein, chrysophanol, and physcion) contained in three ethanolic extracts of well known plants with purgative effects (frangula, senna, and rhubarb). Ethanolic solutions of extracts from these species were vigorously magnetically stirred with fixed quantities of each solid support at room temperature for 1 h. Subsequent HPLC analysis, coupled to photodiode array detection, revealed that, among the solids tested, the hydrotalcite zinc aluminum oleate and magnesium aluminum azelate and magnesium oxide were largely the most effective to this concern allowing to recover anthraquinones (all or some) in good to excellent percentages. Another interesting result was the selective and total removal of rhein by some sorbents from senna and rhubarb extracts. Sorbents were also recyclable and could be re-used to accomplish additional steps without appreciable loss of adsorption capacity. The application of the title solid inorganic and mixed inorganic/organic supports for the selective adsorption and concentration in the solid phase of anthraquinones from commonly used laxative plant species is reported herein for the first time.

Solid phase adsorption of emodin on hydrotalcites and inorganic oxides: A preliminary study.

Solid phase extraction applied to plant matrices is nowadays a well validated technique allowing the concentration and purification of selected secondary metabolites for subsequent analysis. In this short communication we screened the efficiency of 16 selected solid supports including layered structures (hydrotalcites and zirconium phosphate), magnesium oxide and hydroxide, and finally the phyllosilicates talc and bentonite for the selective concentration of the anthraquinone emodin from raw solid extracts of Polygonum cuspidatum Siebold & Zucc. (sin. Reynoutria japonica Houtt.) (Polygonaceae), commonly known as "Japanese knotweed". An ethanolic solution of sample extract from this plant was vigorously mixed with fixed quantities of each solid support. Subsequent HPLC analysis, coupled to photodiode array detection, revealed that, among the solid supports assayed, the hydrotalcite zinc aluminum oleate and magnesium oxide were largely the most effective to this concern. Both were able to extract emodin from the raw extract in percentages of 81.5 % and 92.4 %, respectively. The application of the title supports for the extraction and concentration in the solid phase of anthraquinones from raw plant extracts have been reported herein for the first time.

Umbelliprenin as a novel component of the phytochemical pool from Artemisia spp.

Plants belonging to Artemisia spp. are known to biosynthesize a wide panel of 3,3-dimethylallyl- and sesquiterpenyl- substituted coumarins. In this short communication we applied a novel extraction methodology based on the use of subcritical butane under a counter-current mode to further characterize the presence of selected biologically active oxyprenylated phenylpropanoids, namely coumarins and ferulic acid derivatives, in extracts deriving from aerial parts of Artemisia vulgaris L. (commonly known as "common mugwort") (Asteraceae). In the mean time, we assessed the efficiency of the above mentioned extractive methodology with other routes like maceration and ultrasounds and microwaves-based methods using absolute EtOH as the solvents. UHPLC analysis coupled to UV/Vis detection revealed that, among the 5 pure chemical standard assayed, only umbelliprenin (7-farnesyloxycoumarin) was recorded, while boropinic acid, 4'-geranyloxyferulic acid, 7-isopentenyloxycoumarin, and auraptene were not detected. The best extractive yield (0.18 %) was obtained after extaction with subcritical butane. The presence of umbelliprenin in Artemisia plant species has been reported herein for the first time. This coumarin may represent the biosynthetic precursors of sesquiterpenyloxycoumarins with more complex structures typically found in this genus.

A novel and efficient subcritical butane extraction method and UHPLC analysis of oxyprenylated phenylpropanoids from grapefruits peels.

Biologically active prenyoxyphenylpropanoids are well known to be biosynthesized by Citrus species, for which they have been found most abundantly in fruit peels. Although several extraction methodologies have been described, the development of novel and alternative extraction processes is a field of research of current interest. In this preliminary communication, we studied the performance of the subcritical butane promoted extraction of selected oxyprenylated phenylpropanoids from grapefruit peels under a counter-current mode using a handmade extraction apparatus coupled to UHPLC analysis. The application of such a method yielded 7-isopentenyloxycoumarin, auraptene, and boropinic acid in quantities higher than those recorded for other extraction methodologies like the ultrasound- and microwave-assisted macerations (0.234, 1.035, and 0.211 mg/g of dry extract respectively). The use of subcritical butane as the extraction solvent for oxyprenylated phenylpropanoids is reported herein for the first time and can be easily adopted for several other food matrices.

UHPLC-UV/Vis Quantitative Analysis of Hydroxylated and O-prenylated Coumarins in Pomegranate Seed Extracts.

A simple and rapid analytical UHPLC methodology with spectrophotometric (UV/Vis) detection, coupled with different extraction procedures, has been perfected to investigate the presence of biologically active O-prenylated umbelliferone derivatives, such as auraptene and umbelliprenin, in pomegranate (Punica granatum L.) seed extracts. Absolute ethanol was the most efficient extraction solvent in terms of yields, after a short ultrasound-assisted. The highest concentration values recorded under these experimental conditions were 1.99 µg/g of dry extract and 6.53 µg/g for auraptene and umbelliprenin, respectively. The parent metabolite umbelliferone was also detected (0.67 µg/g). The extraction and UHPLC analytical methodology set up in the present study proved to be an efficient, powerful, and versatile technique for the simultaneous qualitative analysis and quantification of oxyprenylated coumarins in pomegranate seed extracts. The characterization of such secondary metabolites in the mentioned phytopreparation represents, to the best of our knowledge, the first example in the literature.

HPLC Analysis and Skin Whitening Effects of Umbelliprenin-containing Extracts of Anethum Graveolens, Pimpinella Anisum, and Ferulago Campestris.

Umbelliprenin has recently been shown to have great potential as a skin whitening agent. Wishing to investigate the same effect in plant species known to biosynthesize this coumarin, three plants belonging to the Apiaceae family, namely Anethum graveolens L. (dill), Pimpinella anisum L. (anise), and Ferulago campestris (Besser) Grecescu (field ferula) were screened by HPLC analysis for their respective content of umbelliprenin in extracts obtained with different solvent mixtures and by maceration and ultrasound-assisted processes. EtOH was shown to be the best solvent, providing umbelliprenin yields ranging from 1.7% to 14.4% (with respect to the total amount of extract obtained). Extracts with the highest content of this farnesyloxycoumarin were then assayed as modulators of melanogenesis in cultured murine Melan A cells employing the same umbelliprenin obtained by chemical synthesis as the reference. A parallelism between the content of the coumarin and the recorded depigmenting effect (60% for the EtOH extract of F. campestris as the best value) was revealed for all plants extracts when applied at a dose of 100 µg/mL. Our results demonstrate that the same potential of umbelliprenin can be ascribed also to umbelliprenin-enriched plant extracts which reinforces enforce the widespread use of phyto-preparations for cosmetic purposes (e.g., A. graveolens).

An Example of a Novel Efficient Plant Extraction Technique: Electromagnetic Induction Heating.

A simple and easy to handle extraction procedure based on the use of electromagnetic induction heating is described. To assess the potential, scopes, and limitations of this novel process, extraction and subsequent HPLC quantification of emodin from an hydroalcoholic extract of rhizome of Rheum palmatum (Chinese rhubarb) was selected as the reference experiment. Maceration at room temperature and by heating, ultrasound-assisted, and microwave-assisted extractions were also carried out for comparison. Results obtained with electromagnetic induction heating showed that this methodology performed largely better both in terms of time process and extraction yields.

Inhibition of HSV-2 infection by pure compounds from Thymus capitatus extract in vitro.

Thymus capitatus represents 1 of the 5 Tunisian species of the genus Thymus, which has long­standing use for flavouring and preserving several food products. Its constituents have been reported to endow antimicrobial properties, but little is known about their antiviral activities. The aim of this study was to examine the antiviral activity of pure compounds from the most bioactive inhibitory T. capitatus extract in vitro against herpes simplex virus Type 2 (HSV­2) infection and to identify their mechanism of action. Either the extracts or the essential oil exert inhibitory activity against HSV­2 infection, with the ethanolic extract showing the lowest EC50 value (2.3 µg/ml). Three pure compounds were then isolated from the ethanolic extract and investigated for their antiviral activity. ß­sitosterol showed the most favourable selectivity index and both cinnamaldehyde and carvacrol exerted moderate antiviral effect. Investigation of the mechanism of action revealed that all three compounds directly inactivated the infectivity of the virus particles. These findings suggest the use of T. capitatus ethanolic extract as source of anti­HSV­2 pure compounds and warrant further studies to evaluate their therapeutic potential.

Analysis of biologically active oxyprenylated phenylpropanoids in Tea tree oil using selective solid-phase extraction with UHPLC-PDA detection.

An efficient analytical strategy based on different extraction methods of biologically active naturally occurring oxyprenylated umbelliferone and ferulic acid derivatives 7-isopentenyloxycoumarin, auraptene, umbelliprenin, boropinic acid, and 4'-geranyloxyferulic acid and quantification by UHPLC with spectrophotometric (UV/Vis) detection from Tea tree oil is reported. Absorption of the pure oil on Al2O3 (Brockmann activity II) prior washing the resulting solid with MeOH and treatment of this latter with CH2Cl2 resulted the best extraction methodology in terms of yields of oxyprenylated secondary metabolites. Among the five O-prenylphenylpropanoids herein under investigation auraptene and umbelliprenin were never detected while 4'-geranyloxyferulic acid was the most abundant compound resulting from all the three extraction methods employed. The UHPLC analytical methodology set up in the present study resulted to be an effective and versatile technique for the simultaneous characterization and quantification of prenyloxyphenylpropanoids in Tea tree oil and applicable to other complex matrices from the plant kingdom.

Prenylated Coumarins of the Genus Citrus: An Overview of the 2006- 2016 Literature Data.

O-Prenyl coumarins (3,3-dimethylallyl, geranyl-, farnesyl- and related biosynthetic derivatives) represent a class of rarely occurring natural products. In the last two decades, such secondary metabolites have been found to possess promising and effective pharmacological properties, mainly in terms of anti-cancer and anti-inflammatory properties. To date, about 160 oxyprenylated coumarins have been extracted from plants, fungi, and bacteria. The genus Citrus has been demonstrated to be among the richest source of the title products. The aim of this comprehensive review is to make a survey of the so far reported literature citations about the main O-prenyl coumarins found in this genus from phytochemical and pharmacological point of views and for which no surveys of the in so far reported literature have been made.

A green deep eutectic solvent dispersive liquid-liquid micro-extraction (DES-DLLME) for the UHPLC-PDA determination of oxyprenylated phenylpropanoids in olive, soy, peanuts, corn, and sunflower oil.

A green dispersive liquid-liquid microextraction (DLLME) using deep eutectic solvent (DES) as the extracting solvent has been developed and applied for the simultaneous quantification of ferulic acid, umbelliferone, boropinic acid, 7-isopentenyloxycoumarin, 4'-geranyloxyferulic acid (GOFA), and auraptene in some vegetable oils using ultra high performance liquid chromatography (UHPLC) with photodiode array detection (PDA). All parameters in the extraction step, including selection and loading of both extracting and dispersing solvents, amount of both extractant and disperser solvent were investigated and optimized. PhAA/TMG DES achieved higher recovery and enrichment factor compared to other DESs. The validated method showed good linearity with correlation coefficients, r2>0.9990 for all the analytes. Furthermore, this is the first time that eco-friendly solvents are used for the extraction of oxyprenylated phenylpropanoids and the corresponding extract analyzed with ultra high performance liquid chromatography with photodiode array detection.

The interaction of auraptene and other oxyprenylated phenylpropanoids with glucose transporter type 4.

BACKGROUND: Glucose transporter 4 (GLUT4) is firmly established to play a pivotal role in glucose metabolism and in particular in modulating the insulin-stimulated glucose transport in several tissues, such as skeletal muscle and adipose tissue. Stimulation of GLUT4 by insulin results in its translocation to the plasma membrane, activation of several kinases, and finally in a large glucose influx into cells. PURPOSE: In this study we investigated the modulating properties of four biologically active oxyprenylated ferulic acid and umbelliferone derivatives and of their unprenylated parent compounds on GLUT-4 mediated glucose uptake and translocation. METHODS: Oxyprenylated phenylpropanoids have been synthesized in high yields and purity by already reported methodologies. All the synthesized chemicals were tested for their capacity to modulate GLUT4 mediated glucose uptake and GLUT4 translocation in L6 rat skeletal myoblasts in the concentration range 0.1 - 10 µM. Insulin (0.1 µM) was used as positive control. Western blot analysis was employed to assess if GLUT4 translocation occurred prior to increase of glucose uptake. Statistical analyses were carried out by the Dunnett multiple comparison test. RESULTS: 4'-Geranyloxyferulic acid (GOFA), 7-isopentenyloxycoumarin, and auraptene (7-geranyloxycoumarin) increased glucose uptake in a concentration-dependent manner, and significant increases were observed at 0.1 µM for GOFA, and 10 µM for 7-isopentenyloxycoumarin, and auraptene. These products also were able to significantly promote the translocation of GLUT4 to the plasma membrane of L6 myotubes. After treatment with compounds for 15 min, the incorporated amounts of GOFA, 7-isopentenyloxucoumarin, and auraptene were 0.15, 0.32, and 1.77 nmols/60-mm culture dish, respectively. A sample of raw Italian propolis, found to be rich in GOFA and auraptene, was also seen to mimic insulin-effect in the concentration range 0.01 - 1.0 mg/ml. CONCLUSIONS: Among the compounds assayed, auraptene showed to possess potentialities to be a potent activator of both translocation of GLUT4 and glucose influx into skeletal muscle cells with the highest bioavailability among effective compounds. Its capacity to modulate sugar metabolism, coupled to its presence in edible Citrus fruits, can be regarded as an additional reason to account for the already known stimulating properties of some vegetable (e.g. bitter orange).

Quantification of biologically active O-prenylated and unprenylated phenylpropanoids in dill (Anethum graveolens), anise (Pimpinella anisum), and wild celery (Angelica archangelica).

An analytical strategy based on different extraction methodologies and HPLC with spectrophotometric (UV-vis) detection has been developed to investigate the presence of and to quantitate biologically active selected unprenylated and O-prenylated phenylpropanoids, namely umbelliferone, 4'-geranyloxyferulic acid, 7-isopentenyloxycoumarin, auraptene, and umbelliprenin in dill (Anethum graveolens L.), anise (Pimpinella anisum L.), and wild celery (Angelica archangelica L.). Absolute ethanol or 7:3 water/ethanol mixtures were seen to be the most powerful extraction solvents to perform "classic" maceration or ultrasound-assisted one in terms of yields in secondary metabolites. For anethum and anise, umbelliprenine was found to be the most abundant prenyloxy secondary metabolite, while in wild celery 4'-geranyloxyferulic acid recorded the highest concentration. Our experimental approach demonstrated to be efficient for the simultaneous identification and quantitation of the above mentioned prenyloxyphenylpropanoids in the title plant species, that is reported herein for the first time in the literature.