NMR Analysis of Extra Virgin Olive Oil of the Epirus Region of Greece with Emphasis on Selected Phenolic Compounds.

Extra virgin olive oil (EVOO) is recognized for its numerous health benefits, attributed to its rich phenolic components. NMR has emerged as a prevalent technique for precisely identifying these compounds. Among Mediterranean countries, Greece stands as the third-largest producer of olives, with the Epirus region notably advancing in olive cultivation, contributing significantly to the dynamic growth of the region. In this study, an NMR method was employed based on the acquisition of a 1H NMR spectrum along with multiple resonant suppression in order to increase the sensitivity. Using the above method, 198 samples of extra virgin olive oil, primarily sourced from the Epirus region, were analyzed, and both the qualitative and quantitative aspects of the phenolic compounds were obtained. In addition, we examined the effects of various factors such as variety, harvest month, and region origin on the phenolic compounds' concentration. The results revealed an average total phenolic content of 246 mg/kg, closely approaching the EU health claim limit of 250 mg/kg. Approximately 15% of the samples were confidently characterized as high-phenolic olive oil. The highest concentrations were observed in the Thesprotia samples, with several Lianolia varieties exceeding the total phenolic content of 400 mg/kg. Statistical tests demonstrated a significant influence of the olive variety and the month of fruit harvest on phenolic component concentration, followed by the region of origin. A very strong correlation was noted between the total phenolics content and the levels of oleocanthal and oleacein, with a correlation coefficient (r) of 0.924. Upon optimization of all factors affecting olive oil quality, the majority of the EVOOs from the Epirus region have the potential to be characterized as high in phenolic content.

Unlocking the potential of olive residues for functional purposes: update on human intervention trials with health and cosmetic products.

Olive mill waste (OMW) is a promising source of valuable compounds such as polyphenols, terpenes, sterols, and other bioactive compounds, which are of interest to the pharmaceuticals and cosmeceutical industries. This review examines the potential of OMW extracts for health and beauty applications based on evidence reports from human clinical trials. The results achieved to date indicate health-enhancing properties, but little is known about the underlying mechanisms of action, dose-response relationships, and long-term impacts. Therefore, while olive by-products, extracted using eco-friendly methods, present opportunities for the development of high-value health and cosmetic products, further studies are necessary to determine the full range of their effects and establish specific therapeutic strategies. © 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Effective separation of maslinic acid and oleanolic acid from olive pomace using high-speed shear off-line coupled with high-speed countercurrent chromatography and their antibacterial activity test.

In this work, a high-speed shear extraction off-line coupling high-speed countercurrent chromatography method was developed to separate maslinic acid and oleanolic acid from olive pomace. To improve extraction efficiency, the polar disparity between maslinic acid and oleanolic acid necessitated the concurrent utilization of both polar and non-polar solvents during high-speed shear extraction. Then, the high-speed shear extraction was directly feed to high-speed countercurrent chromatography for subsequently separation. A total of 250 min were needed to complete the extraction and separation process. This yielded two molecules from 3.3 g of defatted olive pomace: 7.2 mg of 93.8 % pure maslinic acid and 2.3 mg of 90.1 % pure oleanolic acid, both determined by HPLC at 210 nm. Furthermore, the compounds exhibited inhibitory activity against Escherichia coli and Staphylococcus aureus. At a concentration of 100 µg/mL, its efficacy in inhibiting hyaluronidase was comparable to that of the standard drug indomethacin. Compared with the conventional separation method, this coupled technique reduced the whole time due to the direct injection of sample extraction solution. This technique provides a useful approach for the separation of natural products with significant polarity differences.

Enhancing the quality and safety of Nocellara del Belice green table olives produced using the Castelvetrano method.

The Castelvetrano method is the most widely used among the various table olive processing styles in Sicily. After debittering, the product is stored at low temperatures to prevent the growth of undesirable microorganisms. In an effort to enhance the production process, yeast isolates underwent genotypic characterization and technological screening. The screening process identified two yeast strains Candida norvegica OC10 and Candida boidinii LC1, which can grow at low temperatures and tolerate high pH values (up to 10) and salinity [10% (w/v)]. During the monitoring period, the inoculated trials showed limited presence of spoilage/pathogenic microorganisms. Additionally, the yeasts limited oxidative phenomena and softening of the drupes. The organic compounds detected were higher in the inoculated trials than in the control, and cold storage induced aromatic decay, which was less pronounced in the trial inoculated with C. norvegica. Sensory analysis revealed that the inoculated trials scored higher in sweetness, hardness and crispness.

Eco-friendly extraction of antibacterial compounds from enriched olive pomace: a design-of-experiments approach to sustainability.

The increasing interest in utilizing olive pomace bioactive molecules to advance functional elements and produce antioxidant and antimicrobial additives underscores the need for eco-friendly extraction and purification methods. This study aims to develop an eco-friendly extraction method to evaluate the effect of extraction parameters on the recovery of bioactive molecules from enriched olive pomace. The effects were identified based on total phenolic and flavonoid contents and antioxidant activity, employing a design of experimental methodology. The positive and the negative simultaneous effects showed that among the tested enrichments, those incorporating Nigella Sativa, dates, and coffee demonstrated superior results in terms of the measured responses. Furthermore, chromatographic analysis unveiled the existence of intriguing compounds such as hydroxytyrosol, tyrosol, and squalene in distinct proportions. Beyond this, our study delved into the structural composition of the enriched pomace through FTIR analysis, providing valuable insights into the functional groups and chemical bonds present. Concurrently, antimicrobial assays demonstrated the potent inhibitory effects of these enriched extracts against various microorganisms, underscoring their potential applications in food preservation and safety. These findings highlight enriched olive pomace as a valuable reservoir of bioactive molecules for food products since they can enhance their anti-oxidative activity and contribute to a sustainable circular economy model for olive oil industries.

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.

Olive (Olea europaea L.) Seed as New Source of Cholesterol-Lowering Bioactive Peptides: Elucidation of Their Mechanism of Action in HepG2 Cells and Their Trans-Epithelial Transport in Differentiated Caco-2 Cells.

The production of olive oil has important economic repercussions in Mediterranean countries but also a considerable impact on the environment. This production generates enormous quantities of waste and by-products, which can be exploited as new raw materials to obtain innovative ingredients and therefore make the olive production more sustainable. In a previous study, we decided to foster olive seeds by generating two protein hydrolysates using food-grade enzymes, alcalase (AH) and papain (PH). These hydrolysates have shown, both in vitro and at the cellular level, antioxidant and antidiabetic activities, being able to inhibit the activity of the DPP-IV enzyme and modulate the secretion of GLP-1. Given the multifunctional behavior of peptides, both hydrolysates displayed dual hypocholesterolemic activity, inhibiting the activity of HMGCoAR and impairing the PPI of PCSK9/LDLR, with an IC50 equal to 0.61 mg/mL and 0.31 mg/mL for AH and PH, respectively. Furthermore, both samples restored LDLR protein levels on the membrane of human hepatic HepG2 cells, increasing the uptake of LDL from the extracellular environment. Since intestinal bioavailability is a key component of bioactive peptides, the second objective of this work is to evaluate the capacity of AH and PH peptides to be transported by differentiated human intestinal Caco-2 cells. The peptides transported by intestinal cells have been analyzed using mass spectrometry analysis, identifying a mixture of stable peptides that may represent new ingredients with multifunctional qualities for the development of nutraceuticals and functional foods to delay the onset of metabolic syndrome, promoting the principles of environmental sustainability.

Microbial intervention improves pollutant removal and semi-liquid organo-mineral fertilizer production from olive mill wastewater sludge and rock phosphate.

Olive mill wastewater sludge (OMWS) represents a residual pollutant generated by the olive oil industry, often stored in exposed evaporation ponds, leading to contamination of nearby land and water resources. Despite its promising composition, the valorization of OMWS remains underexplored compared to olive mill wastewater (OMW). This study aims to identify potent native microbial species within OMWS suitable for bioremediation and its transformation into a high-value organic fertilizer. The microbial screening, based on assessing OMWS tolerance and phosphate solubilization properties in vitro, followed by a singular inoculation using a mixture of OMWS and rock phosphate (RP). Identification of FUN 06 (Galactomyces Geotrichum), a fungal species, employed as an inoculant in the treatment of sterile OMWS supplemented with RP. Results demonstrate that fungal inoculation notably diminished OMWS phytotoxicity while enhancing its physicochemical parameters, nutrient concentrations, and removal of toxic organic compounds by up to 90% compared to the control, and enhances plant growth, offering a sustainable approach to tackle environmental concerns. Additionally, metataxonomic analysis unveiled FUN 06's propensity to enhance the presence of microbial species engaged in pollutant degradation. However, higher RP dosage (10%) appeared to adversely affect bioprocess efficiency, suggesting a potential dose-related effect. Overall, FUN 06, isolated from OMWS evaporation ponds, shows promise for effective bioremediation and sustainable reuse. In fact, our results indicate that targeted microbial inoculation stands as an effective strategy for mitigating pollutants in OMWS, facilitating its conversion into a nutrient-rich organo-mineral fertilizer suitable for direct use, promoting its beneficial reuse in agriculture, thereby presenting a promising avenue for olive oil waste management.

Effect of olive leaf phytochemicals on the anti-acetylcholinesterase, anti-cyclooxygenase-2 and ferric reducing antioxidant capacity.

In this study, the phytochemical profile of fifty olive leaves (OL) extracts from Spain, Italy, Greece, Portugal, and Morocco was characterized and their anti-cholinergic, anti-inflammatory, and antioxidant activities were evaluated. Luteolin-7-O-glucoside, isoharmnentin, and apigenin were involved in the acetylcholinesterase (AChE) inhibitory activity, while oleuropein and hydroxytyrosol showed noteworthy potential. Secoiridoids contributed to the cyclooxygenase-2 inhibitory activity and antioxidant capacity. Compounds such as oleuropein, ligstroside and luteolin-7-O-glucoside, may exert an important role in the ferric reducing antioxidant capacity. It should be also highlighted the role of hydroxytyrosol, hydroxycoumarins, and verbascoside concerning the antioxidant activity. This research provides valuable insights and confirms that specific compounds within OL extracts contribute to distinct anti-cholinergic, anti-inflammatory, and anti-oxidative effects.

Ifanosine: Olea europaea L. and Hyphaene thebaica L. combination, from traditional utilization to rational formulation: Preclinical and clinical efficacy on hypertensives patients.

ETHNOPHARMACOLOGICAL RELEVANCE: Olea europaea L. and Hyphaene thebaica L. are commonly employed by traditional healers in Africa for treating and preventing hypertension, either individually or in a polyherbal preparation (Ifanosine). AIM OF THE STUDY: The primary aim was to assess the antihypertensive effects of Olea europaea L. leaves aqueous extract (OEL), Hyphaene thebaica L. mesocarp extract (HT), and the Ifanosine on isolated rat aorta rings. The secondary objective was to evaluate the clinical benefits of a new oral formulation of Ifanosine. MATERIALS AND METHODS: In vitro studies using an isometric transducer examined the antihypertensive effects of HT, OEL, and Ifanosine on rat aorta. Ussing chambers technic were employed to measure mucosal to serosal fluxes and total transepithelial electrical conductance (Gt) to assess the intestinal bioavailability of HT, OEL, and Ifanosine. HPLC was utilized to determine the phytochemical composition of OEL and HT extracts. Subchronic toxicity investigations involved two groups of rats, treated with either water (control) or Ifanosine at 5 g/kg for 28 days. Clinical benefits of the new Ifanosine formulation were evaluated in an observational study with 32 hypertensive patients receiving a fixed oral dose of 3.5 mg three times a day for 30 days. RESULTS: Aqueous extracts induced dose-dependent relaxation of rat aorta rings, with HT and OEL having higher IC50 values than Ifanosine (IC50 = 44.76 ± 1.35 ng/mL, 58.67 ± 1.02 ng/mL, and 29.46 ± 0.26 ng/mL, respectively). The pA2 values of OEL and HT were 1 and 0.6, respectively, while Ifanosine was 0.06. Intestinal bioavailability studies revealed better Prazosin bioavailability than plant extracts. Toxicological studies demonstrated the safety of Ifanosine, supported by histological examinations and biochemical parameters in rat blood. Biochemical analyses indicated flavonoids and phenolic acids as dominant active constituents. Clinical benefits in humans included reduced SBP, DBP, LDL-c, VLDL-c, and TAG, and increased HDL-c without overt adverse effects. CONCLUSION: This study validates the traditional use of OEL and HT for hypertension and advocates for alternative and combinatorial polyphytotherapy (ACP) to enhance traditional remedies.

Nutraceutical potential of olive pomace: insights from cell-based and clinical studies.

Olive oil production yields a substantial volume of by-products, constituting up to 80% of the processed fruits. The olive pomace by-product represents a residue of significant interest due to the diverse bioactive compounds identified in it. However, a thorough characterization and elucidation of the biological activities of olive pomace are imperative to redirect its application for functional food, nutraceutical, and pharmaceutical purposes both for animals and humans. In this review, we examine data from experimental models, including immortalized human vascular endothelial cells, human corneal and conjunctival epithelial cells, human colorectal adenocarcinoma cells, non-tumorigenic human hepatoma cells, and murine macrophages alongside clinical trials. These studies aim to validate the safety, nutritional value, and pharmacological effects of olive pomace. In vitro studies suggest that biophenols extracted from olive pomace possess antioxidant, anti-inflammatory, and antiproliferative properties that could be beneficial in mitigating cardiovascular disorders, particularly atherosclerosis, hepatosteatosis, and dry-eye disease. Protective effects against dry-eye disease were confirmed in a mouse model assay. Olive pomace used in the feed for fish and poultry has demonstrated the ability to enhance animals' immunity and improve nutritional quality of meat and eggs. Human clinical trials are scarce and have revealed minimal biological changes following the consumption of olive pomace-enriched foods. However, alterations in certain biomarkers tentatively suggest cardioprotective properties. The review underscores the value of olive pomace while addressing potential drawbacks and future perspectives, with a specific focus on the need for further investigation into the animal feed and human nutritional properties of olive pomace. © 2024 Society of Chemical Industry.

Chemical Data and Relationships for a Scoring Algorithm of Extra Virgin Olive Oil's Nutritional Value.

Extra virgin olive oil (EVOO) is a valuable product and is highly appreciated by consumers for its great nutritional value. However, to date, there has been a lack of uniform systems capable of ranking the nutritional value of EVOO based on its chemical composition in terms of macro- and micronutrients (including phenolic compounds and tocopherols). The aim of this study was to propose a scoring algorithm to rank the nutritional value of EVOO samples, considering their chemical composition in macro- and micronutrients and their sensitivity to oxidation phenomena. Data from more than 1000 EVOO samples were used to assess the variability of the data, considering the selected negative parameters (free acidity, peroxide value, spectrophotometric indices) and positive components (composition in tocopherols via HPLC-DAD, phenolic compounds via HPLC-DAD, and fatty acids via GC-MS) so as to ensure the universal validity of the scoring algorithm. The dataset included samples from the main producing countries worldwide, in addition to Australia, across several production years; data were selected to represent different production realities. A mathematical model was set up for each chemical component, resulting in six variable values. By combining these values with a dimensionless constant value, the algorithm for computing the nutritional value score (NVS) was defined. It allows the nutritional value of an oil to be ranked on a scale of 0 to 100 based on its chemical composition. The algorithm was then successfully tested using chemical data from about 300 EVOO samples obtained from laboratories from different Italian regions. The proposed NVS is a simple and objective tool for scoring the nutritional value of an EVOO, easy to understand for both producers and consumers.

New olive-pomace fertilizer tested with a 2-tiers approach: Biomarkers on Eisenia fetida, physiochemical effects on Solanum lycopersicum and Olea europaea.

Every year, the olive oil industry generates a substantial amount of pomace, a semi-solid residue made up of skin, pulp, pit, and kernel fragments. Rather than being disposed of, the pomace can be dried and transported to an extraction facility where pomace oil can be extracted. Utilizing its high thermal capacity, the extracted pomace can be used as a supplementary fuel in the drying process, resulting in the production of ashes. In this study, the effect of pomace waste applied to the soil was investigated by testing two mixtures with different proportions of de-oiled pomace flour and kernel ash (50:50 and 70:30, respectively) in powder and pellet form. We used a dual approach, evaluating the effects of the mixtures on both soil communities and plant physiology and productivity, to assess the actual usability of the fertilizer in agriculture. The biomarker approach was valuable in assessing the sublethal effects of the two mixtures in powder form in soil. After 30 days of exposure, the bioindicator organism Eisena fetida showed lipid peroxidation, glutathione S-transferase and lactate dehydrogenase levels similar to the control, while lysozyme activity was reduced in all treatments. The powder mixture was lethal to the tomato plants, while there was no evidence of any damage to the olive trees. During 60 days of monitoring, both mixtures in pellet form showed a slight increase in physiological parameters, suggesting a benefit to the photosynthetic system. The improved carbon assimilation in tomato plants treated with the mixtures results in increased plant productivity, both in terms of number and weight of fruits, while maintaining the antioxidant content. This study paves the way for the use of the pomace mixture as a soil improver, thus increasing the value of this waste product.

Olive leaf protein: Extraction optimization, in vitro digestibility, structural and techno-functional properties.

Olive leaf, as an important by-product of olive farming, is generated from the pruning and harvesting of olive trees and represents >10 % of the total olive weight. The present study was conducted to evaluate the composition, functional and structural characterizations, as well as the in vitro digestibility of olive leaf proteins isolated from ultrasonic-assisted extraction, comparing to classical and industrial techniques. The ultrasound-assisted extraction of olive leaf protein was optimized by the simultaneous maximization of the yield and purity of protein using a Box-Behnken design (BBD) of response surface methodology (RSM). The results indicated that the optimal extraction conditions were as follows: pH of 10.99, temperature of 40.48 °C, sonication time of 47.25 min, and solvent/solid ratio of 24.08 mL/g. Under these conditions, the extraction yield and protein content were 11.67 and 51.2 %, respectively, which were significantly higher than those obtained by the conventional techniques. Regarding the functionality of protein, extraction technique had significant impacts on the structural and functional properties of proteins. In general, ultrasound assisted extraction had higher solubility, and better foaming and thermal properties and in vitro digestibility but lower emulsifying stability and fluid binding capacity compared to conventional ones. Ultrasound-assisted alkaline extraction has great potential to produce edible olive leaf protein with modified functional properties that can be used for various aims in the food applications.

Bioethanol lignin-rich residue from olive stones for electrospun nanostructures development and castor oil structuring.

This work describes the chemical and structural characterization of a lignin-rich residue from the bioethanol production of olive stones and its use for nanostructures development by electrospinning and castor oil structuring. The olive stones were treated by sequential acid/steam explosion pretreatment, further pre-saccharification using a hydrolytic enzyme, and simultaneous saccharification and fermentation (PSSF). The chemical composition of olive stone lignin-rich residue (OSL) was evaluated by standard analytical methods, showing a high lignin content (81.3 %). Moreover, the structural properties were determined by Fourier-transform infrared spectroscopy, nuclear magnetic resonance, and size exclusion chromatography. OSL showed a predominance of ß-ß' resinol, followed by ß-O-4' alkyl aryl ethers and ß-5' phenylcoumaran substructures, high molecular weight, and low S/G ratio. Subsequently, electrospun nanostructures were obtained from solutions containing 20 wt% OSL and cellulose triacetate with variable weight ratios in N, N-Dimethylformamide/Acetone blends and characterized by scanning electron microscopy. Their morphologies were highly dependent on the rheological properties of polymeric solutions. Gel-like dispersions can be obtained by dispersing the electrospun OSL/CT bead nanofibers and uniform nanofiber mats in castor oil. The rheological properties were influenced by the membrane concentration and the OSL:CT weight ratio, as well as the morphology of the electrospun nanostructures.

Bioassay-guided detection, identification and assessment of antibacterial and anti-inflammatory compounds from olive tree flower extracts by high-performance thin-layer chromatography linked to spectroscopy.

Olive trees are one of the most widely cultivated fruit trees in the world. The chemical compositions and biological activities of olive tree fruit and leaves have been extensively researched for their nutritional and health-promoting properties. In contrast, limited data have been reported on olive flowers. The present study aimed to analyse bioactive compounds in olive flower extracts and the effect of fermentation-assisted extraction on phenolic content and antioxidant activity. High-performance thin-layer chromatography (HPTLC) hyphenated with the bioassay-guided detection and spectroscopic identification of bioactive compounds was used for the analysis. Enzymatic and bacterial in situ bioassays were used to detect COX-1 enzyme inhibition and antibacterial activity. Multiple zones of antibacterial activity and one zone of COX-1 inhibition were detected in both, non-fermented and fermented, extracts. A newly developed HPTLC-based experimental protocol was used to measure the high-maximal inhibitory concentrations (IC50) for the assessment of the relative potency of the extracts in inhibiting COX-1 enzyme and antibacterial activity. Strong antibacterial activities detected in zones 4 and 7 were significantly higher in comparison to ampicillin, as confirmed by low IC50 values (IC50 = 57-58 µg in zone 4 and IC50 = 157-167 µg in zone 7) compared to the ampicillin IC50 value (IC50 = 495 µg). The COX-1 inhibition by the extract (IC50 = 76-98 µg) was also strong compared to that of salicylic acid (IC50 = 557 µg). By comparing the locations of the bands to coeluted standards, compounds from detected bioactive bands were tentatively identified. The eluates from bioactive HPTLC zones were further analysed by FTIR NMR, and LC-MS spectroscopy. Multiple zones of antibacterial activity were associated with the presence of triterpenoid acids, while COX-1 inhibition was related to the presence of long-chain fatty acids.

Olive oil and its derivatives for promoting performance, health, and struggling thermal stress effects on broilers.

Olive oil (OL) production is the most significant agro-industrial business and has a high impact on the economy of numerous Mediterranean countries. However, OL extraction results in massive amounts of byproducts, including a solid residue (olive cake or olive pomace) and an aqueous stage (olive mill wastewater), which have serious environmental effects due to their hazardous nature and excessive organic content. Despite these byproducts causing environmental pollution, they can be applied for animal feeding. According to the literature, OL or its derivatives have been used to promote broiler performance, feed utilization, and health status in broilers as growth promoters or protein sources. Furthermore, using OL and its derivatives could improve heat resistance in stressed broilers via struggling thermal stress effects. In this framework, we highlighted the use of OL and its byproducts in broiler feeding to promote performance and health status. Additionally, the role of these byproducts and OL in combating thermal stress is investigated for sustainable strategy and promoting circular economy in broiler industry.

Identification of Phenolic Compounds Extracted from OMW Using LC-MS.

The extraction of olive oil produces annually huge quantities of Olive Mill Wastewater (OMW) that are considered as a source of pollution due to their high concentration in organic matter. This study aims to valorize Olive mill wastewater and investigates the effect of the extraction method and solvents on the contents and profiling of phenolic compounds and their antioxidant potential. It was revealed that the liquid-liquid method using ethyl acetate is the most effective followed by the maceration using chloroform/methanol (1:1), their polyphenol contents are respectively at 1.17 g GAE/L of OMW and 1.07 g GAE/L of OMW. In addition, the antioxidant activity was studied using ABTS test. It has shown that the methanolic extract has the best antioxidant activity at 15.75 mg/L. Moreover, we noticed a negative correlation between the phenolic compounds' concentration and their antioxidant activity which indicates that the phenolic profile may not be the same in the different extracts that's why a primary identification of the phenolic profile using UHPLC-MS was monitored and the results showed different chromatographic profiles between the samples.

Oleuropein in olive leaf, branch, and stem extracts: stability and biological activity in human cervical carcinoma and melanoma cells.

Olive leaves as a main byproduct of olive oil and fruit industry are a valuable source of phytochemicals such as polyphenols, with multiple biomedical effects. Apart from leaves, olive branches and stems make up a significant amount of olive waste. It is well known that the drying process and long-term storage affect the stability and concentration of polyphenols present in raw materials. For that matter, two different means of storing olive waste, at room temperature and +4 °C, were compared by determining the content of the polyphenol oleuropein (OLE) in olive leaf, branch, and stem extracts (LE, BE, and SE) by HPLC-DAD method. Total phenols (TPC), o-diphenols (o-DPC), and total flavonoids (TFC) content in extracts were assessed by UV-Vis measurements. LE prepared from leaves stored at +4 °C had the highest OLE content, 30.7 mg g-1 of dry extract (DE). SE from stems stored at +4 °C was the richest in TPC and TFC (193 mg GAE/g DE and 82.9 mg CE/g DE, respectively), due to the higher purity of the extract. The biological activity of extracts was determined on cervical cancer (HeLa), melanoma (A375), metastatic melanoma (A375M) tumor cell lines, and on spontaneously immortalized cell line of keratinocytes (HaCaT), using the MTT assay. The data show that all extracts had a similar dose-dependent effect on cell viability in HeLa cells, while the effect of LE on melanoma A375 and A375M, and HaCaT cells was cell-line dependent.

Optimization of phenolic compounds extraction from olive mill wastewater using response surface methodology.

Large amounts of pomace are produced during the extraction of olive oil. This is because only 21% of an olive's weight is made up of oil, while 79% includes water, bark and pulp. This project extracted total phenolic compounds (TPCs) from olive mill wastewater as efficiently as possible. The TPCs were analyzed and the total antioxidant activity (TAA) was evaluated by spectrophotometry. Data were evaluated using the response surface method (RSM). The largest TAA and TPC were discovered in extracting using 80% ethanol at 25 °C, and 2.5 bar with pH = 4. The highest amount of TPCs was 11.614 mg of gallic acid per 100 mL, and a value of 71.06% was reported for TAA. The results of the quadratic model showed that R2 is equal to 0.937, because it has a larger coefficient and the pH factor had the least effect. The temperature factor had the greatest impact on the extraction of TPC and TAA, and the mutual temperature and pH impacts affected the extraction positively. As a result, it can be concluded that the RSM was a useful tool for assessing the ideal circumstances for phenolic component extraction.