Phytochemical Study of Stachys iva Griseb. and In Vitro Evaluation of AhR Transcriptional Activity.

Genus Stachys L. consists of approximately 365 species, distributed mainly in temperate regions. Several members of this genus are widely used in the traditional medicine of different countries worldwide. In Greece, 54 Stachys taxa are found in parts of the mainland and/or insular country. The present study focused on the phytochemical investigation of Stachys iva Griseb. and the in vitro anti-inflammatory evaluation of the isolated compounds. In total, eighteen compounds were isolated and identified from the dichloromethane-methanol extract, belonging to iridoids, flavonoids, phenylethanoid glycosides, and phenolic acids. An in vitro approach assessed the aryl hydrocarbon receptor (AhR) modulatory effects of these compounds, revealing an AhR agonistic activity of the flavonoid aglycones apigenin and cirsimaritin in HepG2 and HT29 cell lines. The present study contributes to the evidence of the traditional uses of Stachys spp. and its bioactive constituents, justifying the ethnopharmacological use as an anti-inflammatory plant genus.

Sustainable Recovery of Phenolic Compounds from Distilled Rosemary By-Product Using Green Extraction Methods: Optimization, Comparison, and Antioxidant Activity.

Rosemary solid distillation waste (SWR), a by-product of the essential oil industry, represents an important source of phenolic antioxidants. Green technologies such as ultrasound-assisted extraction (UAE), microwave-assisted extraction (MAE), and accelerated solvent extraction (ASE) of phenolic compounds from SWR were optimized as valorization routes to maximize yield, rosmarinic acid (RMA), carnosol (CARO) and carnosic acid (CARA) contents. Response surface methodology was used in this context, with ethanol concentration (X1), extraction temperature (X2), and time (X3) being the independent variables. A second-order polynomial model was fitted to the data, and multiple regression analysis and analysis of variance were used to determine model fitness and optimal conditions. Ethanol concentration was the most influential extraction parameter, affecting phenolic compounds, while the influence of other parameters was moderate. The optimized conditions were as follows: X1: 67.4, 80.0, and 59.0%, X2: 70, 51, and 125 °C, and X3: 15, 10, and 7 min for MAE, UAE, and ASE, respectively. A comparison of optimized MAE, UAE, and ASE with conventional Soxhlet extraction techniques indicated that ASE provided a higher extraction yield and content of phenolic compounds. However, UAE represented the best process from an environmental point of view, allowing an improved extraction of phenolics from SWR with high energy efficiency and low energy costs.

Current status, technology, regulation and future perspectives of essential oils usage in the food and drink industry.

Nowadays, essential oils (EOs) have a wide use in many applications such as in food, cosmetics, pharmaceutical and animal feed products. Consumers' preferences concerning healthier and safer foodstuffs lead to an increased demand for natural products, in replacement of synthetic substances, used as preservatives, flavourings etc. EOs, besides being safe, are promising alternatives as natural food additives, and much research has been carried out on their antioxidant and antimicrobial activity. The initial purpose of this review is to discuss conventional and 'green' extraction techniques along with their basic mechanism for the isolation of EOs from aromatic plants. This review aims to provide a broad overview of the current knowledge about the chemical constitution of EOs while considering the existence of different chemotypes, since bioactivity is attributed to the chemical composition - qualitative and quantitative - of EOs. Although the food industry primarily uses EOs as flavourings, an overview on recent applications of EOs in food systems and active packaging is provided. EOs exhibit poor solubility in water, oxidation susceptibility, negative organoleptic effect and volatility, restricting their use. Encapsulation techniques have been proven to be one of the best approaches to preserve the biological activities of EOs and minimize their effects on food sensory qualities. Herein, different encapsulation techniques and their basic mechanism for loading EOs are discussed. EOs are highly accepted by consumers, who are often under the misconception that 'natural' means safe. This is, however, an oversimplification, and the possible toxicity of EOs should be taken into consideration. Thus, in the final section of the current review, the focus is on current EU legislation, safety assessment and sensory evaluation of EOs. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Bioactive Profile of Distilled Solid By-Products of Rosemary, Greek Sage and Spearmint as Affected by Distillation Methods.

By-products of essential oils (EOs) in the industry represent an exploitable material for natural and safe antioxidant production. One representative group of such by-products is distilled solid residues, whose composition is properly modulated by the distillation method applied for the recovery of EOs. Recently, in terms of Green Chemistry principles, conventional extraction and distillation processes are considered outdated and tend to be replaced by more environmentally friendly ones. In the present study, microwave-assisted hydro-distillation (MAHD) was employed as a novel and green method for the recovery of EOs from three aromatic plants (rosemary, Greek sage and spearmint). The method was compared to conventional ones, hydro-distillation (HD) and steam-distillation (SD), in terms of phytochemical composition of distilled solid residues, which was estimated by spectrophotometric and chromatographic methods. Total phenolic content (TPC), total flavonoid content (TFC) and antioxidant activity (ABTS, DPPH and FRAP) results highlighted the distilled solid residues as good sources of antioxidants. Moreover, higher antioxidant activity was achieved for MAHD extracts of solid residues in comparison to HD and SD extracts. A metabolomics approach was carried out on the methanolic extracts of solid residues obtained by different distillation methods using LC-MS analysis followed by multivariate data analysis. A total of 29 specialized metabolites were detected, and 26 of them were identified and quantified, presenting a similar phenolic profile among different treatments, whereas differences were observed among different species. Rosmarinic acid was the most abundant phenolic compound in all extracts, being higher in MAHD extracts. In rosemary and Greek sage extracts, carnosol and carnosic acid were quantified in significant amounts, while trimers and tetramers of caffeic acid (salvianolic acids isomers) were identified and quantified in spearmint extracts, being higher in MAHD extracts. The obtained results pointed out that MAHD extracts of distilled solid by-products could be a good source of bioactives with potential application in the food, pharmaceutical and cosmetic industries, contributing to the circular economy.

Development and Validation of an HPLC Method for the Analysis of Flowers of Wild-Growing Primula veris from Epirus, Greece.

An HPLC-PDA method was developed for the determination of the flavonoids in the flowers of Primula veris from Epirus, Greece. The aim was to investigate the chemical content of the over-harvested P. veris populations of Epirus and to develop and optimize an extraction protocol to allow fast, exhaustive, and repeatable extraction. Qualitative analysis revealed that the P. veris flowers from Epirus were particularly rich in flavonoids, especially flavonol triglycosides including derivatives of quercetin, isorhamnetin, and kaempferol. A phytochemical investigation of a 70% hydromethanolic extract from the flowers afforded a new flavonoid, namely, isorhamnetin-3-Ο-ß-glucopyranosyl-(1 → 2)-ß-glucopyranosyl-(1 → 6)-ß-glucopyranoside, which is also the main constituent of the flower extracts. Its structure elucidation was carried out by means of 1D and 2D NMR and mass spectrometry analyses. The HPLC-PDA method was developed and validated according to the International Council for Harmonisation guidelines. Since the main flavonol glycoside of the plant is not commercially available, rutin was used as a secondary standard and the response correction factor was determined. Finally, the overall method was validated for precision (% relative standard deviation ranging between 1.58 and 4.85) and accuracy at three concentration levels. The recovery ranged between 93.5 and 102.1% with relative standard deviation values < 5%, within the acceptable limits. The developed assay is fast and simple and will allow for the quality control of the herbal drug.

Anti-Ageing Potential of S. euboea Heldr. Phenolics.

In recent years, the use of Sideritis species as bioactive agents is increasing exponentially. The present study aimed to investigate the chemical constituents, as well as the anti-ageing potential of the cultivated Sideritis euboea Heldr. The chemical fingerprinting of the ethyl acetate residue of this plant was studied using 1D and 2D-NMR spectra. Isomeric compounds belonging to acylated flavone derivatives and phenylethanoid glycosides were detected in the early stage of the experimental process through 2D-NMR techniques. Overall, thirty-three known compounds were isolated and identified. Some of them are reported for the first time not only in S. euboea, but also in genus Sideritis L. The anti-ageing effect of the ethyl acetate residue and the isolated specialized products was assessed as anti-hyaluronidase activity. In silico docking simulation revealed the interactions of the isolated compounds with hyaluronidase. Furthermore, the in vitro study on the inhibition of hyaluronidase unveiled the potent inhibitory properties of ethyl acetate residue and apigenin 7-O-ß-d-glucopyranoside. Though, the isomers of apigenin 7-O-p-coumaroyl-glucosides and also the 4'-methyl-hypolaetin 7-O-[6'''-O-acetyl-ß-d-allopyranosyl]-(1→2)-ß-d-glucopyranoside exerted moderate hyaluronidase inhibition. This research represents the first study to report on the anti-hyaluronidase activity of Sideritis species, confirming its anti-inflammatory, cytotoxic and anti-ageing effects and its importance as an agent for cosmetic formulations as also anticancer potential.

NMR analysis of cultivated Sideritis euboea Heldr.

INTRODUCTION: Over the last decades, the demand of medicinal plants has vastly increased. The health trend and social changes are leading to a growing demand for herbal products, as medicines and in particular as food supplements. Various cultivations of medicinal plants have been developed in order not only to respond on this enormous market, but also to offer high-quality products. Cultivated plants have attracted great scientific interest for their rich and high standard chemical compositions. OBJECTIVE: Our study focuses on the phytochemical profile of cultivated Sideritis euboea Heldr., a shrub with a great commercial value. The whole isolation course of all extracts, fractions and sub-fractions was continuously monitored and traced down using an NMR strategy, which permitted their detail characterisation and enabled us to choose the more abundant extract. METHODS: Aerial parts of cultivated S. euboea were extracted with solvents of increasing polarity and their secondary metabolites were identified by NMR analyses. RESULTS: So far, using various analytical methods, 16 secondary metabolites have been isolated from the methanol extract; two iridoids, seven flavones, five phenylethanoid glucosides and two phenolic acids. In addition to the previous investigation of the wild population, the present study revealed that its chemical profile is similar and even richer in phenolic compounds. CONCLUSION: The metabolic profile of cultivated S. euboea was studied through 1 H-NMR spectra; the structures of the isolated secondary metabolites were assigned based on their one-dimensional and two-dimensional NMR spectra. The NMR strategy allowed us to detect even the minor compounds in the extracts.

NMR-Based Chemical Profiling, Isolation and Evaluation of the Cytotoxic Potential of the Diterpenoid Siderol from Cultivated Sideritis euboea Heldr.

Diterpenes are characteristic compounds from the genus Sideritis L., possessing an array of biological activities. Siderol is the main constituent of the ent-kaurene diterpenes in Sideritis species. In order to isolate the specific compound and evaluate for the first time its cytotoxic activity, we explored the dichloromethane extract of cultivated Sideritis euboea Heldr. To track the specific natural bioactive agent, we applied NMR spectroscopy to the crude plant extract, since NMR can serve as a powerful and rapid tool both to navigate the targeted isolation process of bioactive constituents, and to also reveal the identity of bioactive components. Along these lines, from the rapid 1D 1H NMR spectrum of the total crude plant extract, we were able to determine the characteristic proton NMR signals of siderol. Furthermore, with the same NMR spectrum, we were able to categorize several secondary metabolites into chemical groups as a control of the isolation process. Therefore, this non-polar extract was explored, for the first time, revealing eleven compounds-one fatty acid ester; 2-(p-hydroxyphenyl)ethylstearate (1), three phytosterols; ß-sitosterol (2), stigmasterol (3), and campesterol (4); one triterpenoid; ursolic acid (5), four diterpenoids; siderol (6), eubol (7), eubotriol (8), 7-epicandicandiol (9) and two flavonoids; xanthomicrol (10) and penduletin (11). The main isolated constituent was siderol. The antiproliferative potential of siderol was evaluated, using the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) assay, on three human cancer cell lines DLD1, HeLa, and A549, where the IC50 values were estimated at 26.4 ± 3.7, 44.7 ± 7.2, and 46.0 ± 4.9 µΜ, respectively. The most potent activity was recorded in the human colon cancer cell line DLD1, where siderol exhibited the lowest IC50. Our study unveiled the beneficial potential of siderol as a remarkable cytotoxic agent and the significant contribution of NMR spectroscopy towards the isolation and identification of this potent anticancer agent.

Self-Nanoemulsifying Drug Delivery Systems (SNEDDS) Containing Rice Bran Oil for Enhanced Fenofibrate Oral Delivery: In Vitro Digestion, Ex Vivo Permeability, and In Vivo Bioavailability Studies.

Lipid-based drug delivery systems (LbDDS), such as self-nanoemulsifying drug delivery systems (SNEDDS), constitute a prominent formulation approach for enhancing the aqueous solubility and oral bioavailability of poorly water-soluble compounds. Utilization of biorefinery wastes, such as oil from rice bran, may prove advantageous to both improving drug solubilization and absorption and to achieving sustainable agri-food waste valorization. Here, we assessed the effect of four SNEDDS compositions differing in the oil (rice bran oil and corn oil) and surfactant type (Kolliphor RH40 and EL) on the oral bioavailability of fenofibrate, a BCS class II compound. Prior to the in vivo oral administration of the SNEDDS in rats, drug solubilization was tested in vitro using the static digestion model, followed by the ex vivo permeability study of the predigested SNEDDS using the non-everted gut sac model. No significant variation was observed in the solubilization capacity within the different SNEDDS formulations. On the other hand, the ex vivo permeability data of the predigested SNEDDS correlated well with the in vivo bioavailability data designating the superiority of rice bran oil with Kolliphor EL as the surfactant, to enhance the oral absorption of fenofibrate. Results indicated that valorization of agro-industrial waste such as rice bran oil may prove useful in enhancing the oral performance of LbDDS in the case of fenofibrate, while at the same time maximizing the use of agricultural by-products via the creation of new sustainable value chains in the pharmaceutical field.

Development and Characterization of a Self-Nanoemulsifying Drug Delivery System Comprised of Rice Bran Oil for Poorly Soluble Drugs.

Poor aqueous solubility and low bioavailability are limiting factors in the oral delivery of lipophilic drugs. In a formulation approach to overcome these limitations, rice bran (RB) oil was evaluated as drug carrier in the development of self-nanoemulsifying drug delivery systems (SNEDDS). The performance of RB in formulations incorporating Kolliphor RH40 or Kolliphor EL as surfactants and Transcutol HP as cosolvent was compared to a common oil vehicle, corn oil (CO). Serial dilutions of the preconcentrates were performed in various media [distilled water and simulated intestinal fluids mimicking fasted state (FaSSIF) and fed state (FeSSIF)] and at different dilution ratios to simulate the in vivo droplets' behavior. The developed SNEDDS were assessed by means of phase separation, droplet size, polydispersity index, and ζ-potential. Complex ternary diagrams were constructed to identify compositions exhibiting monophasic behavior, droplet size < 100 nm, and polydispersity index (PDI) < 0.25. Multifactor analysis and response surface areas intended to determine the factors significantly affecting droplet size. The oil capacity to accommodate lipophilic drugs was assessed via fluorescence spectroscopy based on the solvatochromic behavior of Nile Red. Solubility studies were performed to prepare fenofibrate- and itraconazole-loaded SNEDDS and assess their droplet size, whereas dissolution experiments were conducted in simulated intestinal fluids. Caco-2 cell viability studies confirmed the safety of the SNEDDS formulations at 1:100 and 1:1000 dilutions after cell exposure in culture for 4 h. The obtained results showed similar performance between RB and CO supporting the potential of RB as oil vehicle for the effective oral delivery of lipophilic compounds.