Cyclodextrin-based polymeric materials for the specific recovery of polyphenolic compounds through supramolecular host–guest interactions

While the specific recovery of valuable chemicals from waste streams represents an environmentally-friendly and potentially economically-relevant alternative to synthetic chemical productions, it remains a largely unmet challenge. This is partially explained by the complexity of designing sorption materials able to target one specific compound and able to function in complex matrices. In this work, a series of cyclodextrin-based polymers (CDPs) were designed to selectively extract phenolic compounds from a complex organic matrix that is olive oil mill wastewater (OMW). In order to endow these polymers with selective adsorption properties, several monomers and cross-linkers were screened and selected. The adsorption properties of the CDPs produced were first tested with selected phenolic compounds commonly found in OMW, namely syringic acid, p-coumaric acid, tyrosol and caffeic acid. The selected CDPs were subsequently tested for their ability to adsorb phenolic compounds directly from OMW, which is known to possess a high and complex organic content. It was demonstrated through high-performance liquid chromatography-mass spectroscopy analyses that efficient removal of phenolic compounds from OMW could be achieved but also that two compounds, namely tyrosol and hydroxytyrosol, could be selectively extracted from OMW.     

Multifunctional approaches to provide potential pharmacophores for the pharmacy shelf: Heracleum sphondylium L. subsp. ternatum (Velen.) Brummitt.

Heracleum sphondylium L. subsp. ternatum (Velen.) Brummitt. commonly known as “hogweed” is traditionally used to manage several human ailments. This investigation assessed, for the first time, the enzyme inhibitory properties, antioxidant activity, phytochemical profile, antimutagenic, and antimicrobial potential of the ethyl acetate, methanol, and water extracts of H. sphondylium. We also established the possible interactions of identified phenolic compounds with cholinesterases, amylase, glucosidase, and tyrosinase using in silico docking studies. Chlorogenic acid was found in high amounts in the methanol extract of H. sphondylium. The methanol extract was an effective inhibitor of acetylcholinesterase (1.70 mg galantamine equivalent (GALAE)/g extract) while the ethyl acetate extract showed pronounced inhibitory action against butyrylcholinesterase (1.77 mg GALAE/g extract). The extracts exhibited low inhibition against amylase (0.12-0.84 mmol acarbose equivalent (ACAE)/g extract) and a more pronounced inhibition against glucosidase (2.29–3.65 mmol ACAE/g extract). In silico results showed that rutin and quercetin (-70.4 and -72.2 Kcal/mol, for rutin and quercetin respectively) docked to the enzymatic cavity of acetylcholinesterase but these phenolic compounds showed less affinity with butyrylcholinesterase (-15.0 and -5.2 Kcal/mol, for rutin and quercetin respectively). The extracts did not induce any mutations on the bacterial strains, while they have excellent antimutagenic capacity against well-known mutagens (inhibition values 98%, 97% and 96%). The methanol extract (0.78 mg/ml) showed moderate antifungal activity while the ethyl acetate extract (0.78–3.12 mg/ml) showed weak to moderate antimicrobial activity. This study provides valuable baseline data which might serve for the development of future pharmacophores for the management of human ailments.     

Effect of Extraction Method on Total Flavonoid,Total Phenolic Content,Antioxidant and Anti-bacterial Activity of Zingiberis Officinale Rhizome

This research aims to compare between decocta and maceration method using water and ethanol as a solvent on total flavonoid, total phenolic content, antioxidant and antibacterial activity of Zingiber officinale rhyzome extract. The Rhyzome was dryed by oven method on 50 °C for 24 hours and 48 hours. Total flavonoid, total phenolic content and antioxidant activity were assay by spectrophotometric method. The antibacterial activity was determined by difussion agar method against Bacillus subtilis (Bs) and Escherichia colli InaCC B5 (Ec). Ethanol extract from dried rhyzome processed for 24 hours (R-24) indicated the best result for total flavonoid, total phenolic content and antioxidant activity. The yield of total flavonoid for R-24 was 286 μg/mL. The yield of total phenolic content for R-24 was 504 μg/mL and the yield of IC₅₀ antioxidant activity for R-24 was 5.753 μg/mL. The extract should antibacterial activity againts Bs with inhibition diameter of 10% and 20% of samples concentration are 10 and 11.5 mm respectively. Meanwhile, The inhibition diameter result against Ec for 10% and 20% of samples concentration are 7.5 and 8.5 mm respectively.     

Evaluation of different extraction approaches for the determination of phenolic compounds and their metabolites in plasma by nanoLC-ESI-TOF-MS

Sample preparation is an important step for the determination of phenolic compounds in biological samples. Different extraction methods have been tested to determine phenolic compounds and their metabolites in plasma by nano-liquid chromatography coupled to electrospray ionisation-time-of-flight mass spectrometry (nanoLC-ESI-TOF-MS). The sample treatment optimisation was performed using commercial foetal bovine serum spiked with representative phenolic standards, namely naringenin, luteolin, verbascoside, apigenin, rutin, syringic acid and catechin. Different protein-precipitation conditions were evaluated as well as enzymatic digestion with trypsin and solid-phase extraction using different phases such as C-18, ABN and ENV+, working at different pH values. The optimum extraction procedure consisted of a previous protein-precipitation step using HCl 200?mmol/L in methanol for 2.5?h at 50?°C followed by a solid-phase extraction using C-18 cartridges at pH?2.5. This procedure was finally applied to the plasma of rats overfed with a phenolic-rich Lippia citriodora extract. These samples were analysed by nanoLC-ESI-TOF-MS, enabling the identification of five compounds previously found in the administered L. citriodora extract and one metabolite.     

Antioxidant and antibacterial activity of silver nanoparticles biosynthesized using Chenopodium murale leaf extract

Silver is known for its antimicrobial effects and silver nanoparticles are gaining their importance due to their antimicrobial activities. The aims of the current study were to use plant extract for the biosynthesis of silver nanoparticles and to evaluate their antibacterial and antioxidant activity in vitro. The results indicated that silver nanoparticles (AgNPs) can be synthesized in a simple method using Chenopodium murale leaf extract. The TEM analysis showed that the sizes of the synthesized AgNps ranged from 30 to 50 nm. The essential oil of C. murale leaf extract was formed mainly of α-Terpinene, (Z)-Ascaridole and cis-Ascaridole. The total phenolic compounds and total flavonides were higher in AgNPs-containing plant extract compared to the plant extract. AgNPs-containing leaf extract showed a higher antioxidant and antimicrobial activity compared to C. murale leaf extract alone or silver nitrate. It could be concluded that C. murale leaf extract can be used effectively in the production of potential antioxidant and antimicrobial AgNPs for commercial application.     

Quick assessment of the economic value of olive mill waste water

Background

Olive biophenols are emerging as a valued class of natural products finding practical application in the food, pharmaceutical, beverage, cosmetic and nutraceutical industries due to their powerful biological activity which includes antioxidant and antimicrobial properties. Olive mill waste water (OMWW), a by-product in olive oil manufacturing, is rich in biophenols such as hydroxytyrosol and tyrosol. The amount of biophenols depends on the cultivar, the geographical area of cultivation, and the seasonal conditions. The goal of this study was to develop a straightforward method to assess the economic value of OMWW via quantification of hydroxytyrosol and tyrosol.

Results

The amount of hydroxytyrosol and tyrosol phenolic compounds in the OMWW from four different cultivars grown in four different regions of Sicily was analyzed using liquid–liquid and solid–liquid analytical protocols developed ad hoc. Results showed significant differences amongst the different cultivars and their geographical origin. In all samples, the concentration of hydroxytyrosol was generally from 2 to 10 times higher than that of tyrosol. In general, the liquid–liquid extraction protocol gave higher amounts of extracted biophenols. The cultivar Cerasuola had the highest amount of both hydroxytyrosol and tyrosol. The cultivar Nocellara Etnea had the lowest content of both biophenols.

Conclusions

A quick method to assess the economic value of olive mill waste water via quantification of hydroxytyrosol and tyrosol in olive phenolic enriched extracts is now available.

     

Nutraceutical Potential of Leaf Hydro-Ethanolic Extract of the Edible Halophyte Crithmum maritimum L.

Aromatic halophytes represent an exceptional source of natural bioactive compounds for the food industry. Crithmum maritimum L., also known as sea fennel, is a halophyte plant colonizing cliffs and coastal dunes along Mediterranean and Atlantic coasts. It is well known to produce essential oils and polyphenols endowed with antioxidant and biological effects. The present work reports the phytochemical profile, as well as antioxidant, antimicrobial and antimutagenic properties of C. maritimum leaf hydro-alcoholic extract. From LC-ESI-MS analysis, eighteen phenolic compounds were depicted in sea fennel extract and the amount of total phenolic content exceeds 3% DW. Accordingly, C. maritimum extract showed strong antioxidant activities, as evidenced by in vitro (DPPH, ORAC, FRAP) and ex vivo (CAA-RBC and hemolysis) assays. An important antimicrobial activity against pathogenic strains was found as well as a strong capacity to inhibit Staphylococcus aureus (ATCC 35556) biofilm formation. Sea fennel extracts showed a significant decrease of mutagenesis induced by hydrogen peroxide (H₂O₂) and menadione (ME) in Saccharomyces cerevisiae D7 strain. In conclusion, our results show that C. maritimum is an exceptional source of bioactive components and exert beneficial effects against oxidative or mutagenic mechanisms, and pathogenic bacteria, making it a potential functional food.     

Characterization of Citrus nobilis Peel Methanolic Extract for Antioxidant,Antimicrobial, and Anti-Inflammatory Activity

Currently, the potential utilization of fruits and vegetable waste as a source of micronutrients and antioxidants has increased. The present study, therefore, aimed to determine the antimicrobial and anti-inflammatory activities of Citrus nobilis peel extract. A modified solvent evaporation technique was employed for peel extract preparation. For effective utilization of the natural product, quantitative analysis of phenolic compounds was carried out using liquid chromatography and mass spectroscopy technique. Phenolic and flavonoids were present in high amounts, while β-carotene and lycopene were present in vestigial amounts. The antimicrobial efficiency of peel extract was evaluated against four bacterial strains including Staphylococcus aureus (MTCC 3160), Klebsiella pneumoniae (MTCC 3384), Pseudomonas aeruginosa (MTCC 2295), and Salmonella typhimurium (MTCC 1254), and one fungal strain Candida albicans (MTCC 183), and zone of inhibition was comparable to the positive control streptomycin and amphotericin B, respectively. The extract of Citrus nobilis peels showed effective anti-inflammatory activity during human red blood cell membrane stabilization (HRBC) and albumin denaturation assay. The extracts also exhibited 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity ranging from 53.46 to 81.13%. Therefore, the obtained results suggest that Citrus nobilis peel could be used as an excellent source of polyphenols and transformed into value-added products.     

Olive Pomace Phenolic Compounds Stability and Safety Evaluation: From Raw Material to Future Ophthalmic Applications

Nowadays, increasing interest in olive pomace (OP) valorization aims to improve olive’s industry sustainability. Interestingly, several studies propose a high-value application for OP extracts containing its main phenolic compounds, hydroxytyrosol and oleuropein, as therapy for ocular surface diseases. In this work, the stability and accessibility of OP total phenolic and flavonoid content, main representative compounds, and antioxidant activity were assessed under different pretreatment conditions. Among them, lyophilization and supercritical CO₂ extraction were found to increase significantly most responses measured in the produced extracts. Two selected extracts (CONV and OPT3) were obtained by different techniques (conventional and pressurized liquid extraction); Their aqueous solutions were characterized by HPLC-DAD-MS/MS. Additionally, their safety and stability were evaluated according to EMA requirements towards their approval as ophthalmic products: their genotoxic effect on ocular surface cells and their 6-months storage stability at 4 different temperature/moisture conditions (CPMP/ICH/2736/99), together with pure hydroxytyrosol and oleuropein solutions. The concentration of hydroxytyrosol and oleuropein in pure or extract solutions was tracked, and possible degradation products were putatively identified by HPLC-DAD-MS/MS. Hydroxytyrosol and oleuropein had different stability as standard or extract solutions, with oleuropein also showing different degradation profile. All compounds/extracts were safe for ophthalmic use at the concentrations tested.     

Chemoselective and efficient carbomethoxylation of the alcoholic chain of phenols by dimethyl carbonate (DMC)

The efficiency of dimethyl carbonate (DMC) as chemoselective carbomethoxylating agent of the alcoholic chain of phenols has been investigated. In the presence of 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) or sulfuric acid as catalysts, new carbomethoxylated phenolic compounds were obtained in quantitative yields. A new efficient derivatization of the aliphatic alcoholic chain of the precious natural hydroxytyrosol is described, which increases the lipophilicity of the hydroxytyrosol. The antioxidant activity of this new carboxymethylated hydroxytyrosol 8 has been investigated using DPPH radical scavenging test. The results showed that this new compound has an antioxidant activity similar to hydroxytyrosol.     

LC-MS/MS based metabolite profiling and lipase enzyme inhibitory activity of Kaempferia angustifolia Rosc. with different extracting solvents

Kaempferia angustifolia also known as kunci pepet in Indonesia, has been widely used as a traditional medicine to treat cold, cough, stomachache, diarrhea, fever, and dysentery, also used as a slimming agent. The level of biological activity depends on the composition and concentration of bioactive compounds present in the plants. In addition, extraction solvents affects the composition and concentration of bioactive compounds. Therefore, this study aimed at identifying the metabolite profile of K. angustifolia and to evaluate the inhibitory potential of their various solvent extracts towards lipase enzyme. Extracts were prepared using water and different concentration of ethanol (30–99%) and then analyzed their metabolite profile using LC-MS/MS. Lipase inhibitory activity was assessed using in vitro enzymatic inhibition assay. In this study, profile of K. angustifolia was shown to be rich in terpenoids (monoterpenoids, sesquiterpenoids, diterpenoids), and phenolics (carboxylic acid and phenolic acid). Most of the identified compounds were detected in ethanol extract of K. angustifolia. The ethanol extract at 100 μg/mL displayed 59.82% inhibitory activity towards lipase and was found to have the highest inhibitory activity compared to the other extracts. A partial least square-discriminant analysis (PLS-DA) was performed for clustering the extracts based on the peak area of 53 putatively identified compounds. Based on the result obtained, 50% ethanol extract is the best extract that gives the highest inhibition results and 15 metabolites were identified, mainly from the carboxylic acid and terpenoid groups.     

Chemical Composition and Antioxidant,Anti-Inflammatory,and Enzyme Inhibitory Activities of an Endemic Species from Southern Algeria: Warionia saharae

Warionia saharae Benth. & Coss. (Asteraceae) is an endemic species of North Africa naturally grown in the southwest of the Algerian Sahara. In the present study, this species’ hydromethanolic leaf extract was investigated for its phenolic profile characterized by ultra-high-performance liquid chromatography coupled with a diode array detector and an electrospray mass spectrometer (UHPLC-DAD-ESI/MS). Additionally, the chemical composition of W. saharae was analyzed by gas chromatography–mass spectrometry, and its antioxidant potential was assessed through five in vitro tests: DPPH^● scavenging activity, ABTS^●+ scavenging assay, galvinoxyl scavenging activity, ferric reducing power (FRP), and cupric reducing antioxidant capacity. The UHPLC-DAD-ESI/MS analysis allowed the detection and quantification of 22 compounds, with taxifolin as the dominant compound. The GC–MS analysis allowed the identification of 37 compounds, and the antioxidant activity data indicate that W. saharae extract has a very high capacity to capture radicals due to its richness in compounds with antioxidant capacity. The extract also showed potent α-glucosidase inhibition as well as a good anti-inflammatory activity. However, weak anti-α-amylase and anticholinesterase activities were recorded. Moreover, an in silico docking study was performed to highlight possible interactions between three significant compounds identified in W. saharae extract and α-glucosidase enzyme.     

Sambucus Nigra Extracts–Natural Antioxidants and Antimicrobial Compounds

Due to the health-promoting properties of elderberry fruits, which result from their rich chemical composition, this raw material is widely used in herbal medicine and the food industry. The aim of the study was to demonstrate the antibacterial activity of the elderberry fruit extracts. The research showed that the content of phenolic acids and flavonoids in the extracts determined their antibacterial activity. The research showed that the content of phenolic acids and flavonoids in the extracts determined their antibacterial activity. The following phenolic acids were predominant: chlorogenic acid, sinapic acid, and t-cinnamic acid. Their average content was, respectively, 139.09, 72.84, 51.29 mg/g extract. Rutin and quercetin (their average content was 1105.39 and 306.6 mg/g extract, respectively) were the dominant flavonoids. The research showed that the elderberry polyphenol extracts exhibited activity against selected strains of bacteria within the concentration range of 0.5–0.05%. The following bacteria were the most sensitive to the extracts: Micrococcus luteus, Proteus mirabilis, Pseudomonas fragii, and Escherichia coli. Of the compounds under analysis, apigenin, kaempferol and ferulic, protocatechuic, and p-coumarin acids had the greatest influence on the high antibacterial activity of elderberry extracts. The results of the microbiological and chemical analyses of the composition of the extracts were analyzed statistically to indicate the bioactive compounds of the greatest antimicrobial significance.     

Comparison of different sample preparation treatments for the analysis of wine phenolic compounds in human plasma by reversed phase high-performance liquid chromatography

Phenolic compounds are common constituents of wine. Due to their healthy properties the analysis in human fluids is interesting within bioavailability evaluation. They have been reported not to be stable in human plasma, particularly at room temperature. Most sample treatments have been reported for a single compound. Our aim in this paper is to study sample handling control conditions and improve phenolic stability in human plasma samples. We tested various sample treatments to determine whether they could be used for analysing a set of phenolic compounds usually present in wines.The compounds studied were six phenolic acids, five flavonoids, trans-resveratrol and tyrosol. The effect of the following factors was explored: temperature, pH, the addition of antioxidants and the addition of anticoagulants.The results suggest that the plasma samples should be kept at temperatures below −20 °C before analysis and that 1% ascorbic acid plus 10 μl/ml o-phosphoric acid should be added. Anticoagulants (heparin or EDTA) do not play a significant role in the stability of polyphenolic compounds.The recovery values of a number of sample treatments (solid phase extraction, extraction with methanol, deproteinization, inhibition of enzymatic plasma activity) were compared. The recovery values for most phenolic compounds were better if the enzymatic plasma activity was inhibited and acidified ethanol was used for deproteinization.     

Presence of virgin olive oil phenolic metabolites in human low density lipoprotein fraction: determination by high-performance liquid chromatography-electrospray ionization tandem mass spectrometry

The biological benefits of olive oil in preventing the oxidation of low density lipoprotein (LDL) would seem to be linked to its high monounsaturated fatty acid contents, but also to its respective phenolic compounds contents. One prerequisite to assess the in vivo physiological significance of phenolic compounds is to determine their presence in human LDL following the ingestion of virgin olive oil.In this work, olive oil phenolic metabolites were identified using high-performance liquid chromatography in tandem with electrospray mass spectrometry (HPLC-ESI-MS/MS) detection, after solid phase extraction (SPE). Quantitative methods were developed in carrying out linearity, precision, sensitivity and recovery tests. The results from two methods of LDL separation were compared and shorter LDL isolation procedure showed a better recovery for antioxidants compounds in LDL. The metabolites identified in LDL were: hydroxytyrosol monoglucuronide, hydroxytyrosol monosulfate, tyrosol glucuronide, tyrosol sulfate and homovanillic acid sulfate. The fact that olive oil phenolic metabolites are able to bind LDL strengthens claims that these compounds act as in vivo antioxidants.     

Detection and quantification of phenolic compounds in olive oil by high resolution H nuclear magnetic resonance spectroscopy

High resolution ¹H NMR spectroscopy has been employed as a versatile and rapid method to analyze the polar fraction of extra virgin olive oils containing various classes of phenolic compounds. The strategy for identification of phenolic compounds is based on the NMR chemical shifts of a large number of model compounds assigned by using two-dimensional (2D) NMR spectroscopy. Furthermore, 2D NMR was applied to phenolic extracts in an attempt to discover additional phenolic compounds. The ¹H NMR methodology was successful in detecting simple phenols, such as p-coumaric acid, vanillic acid, homovanillyl alcohol, vanillin, free tyrosol, and free hydroxytyrosol, the flavonols apigenin and luteolin, the lignans (+) pinoresinol, (+) 1-acetoxypinoresinol and syringaresinol, two isomers of the aldehydic form of oleuropein and ligstroside, the dialdehydic form of oleuropein and ligstroside lacking a carboxymethyl group, and finally total hydroxytyrosol and total tyrosol reflecting the total amounts of free and esterified hydroxytyrol and tyrosol, respectively. The absolute amount of each phenolic constituent was determined in the polar fraction by using anhydrous 1,3,5-triazine as an internal standard.     

Bleacing of olive mill wastewater by clay in the presence of hydrogen peroxide

Treatment of olive mill wastewater (OMW) with clayey soils in the presence of hydrogen peroxide (H₂O₂) allows the elimination of phenolic compounds responsible for the black-brownish color of this industrial effluent. The aim of this research was to define optimal physicochemical parameters for the bleaching of OMW with clay in the presence of hydrogen peroxide. Two clayey soil powders were tested (A and B) and the results obtained indicate that high bleaching could be reached after 24 hours exposure of OMW to 7 % (W/V) clay material A in the presence of 0.5 % (V/V) hydrogen peroxide. Under these conditions, the bleaching led to about 87 % decrease of polyphenols (PF) and a 66 % decrease of the Chemical Oxygen Demand (COD). The structure of clay and its concentration in iron salts have an effective adsorbent and catalytic effect on the removal of the majority of polyphenols.     

Total phenolic and flavonoid contents,antioxidant, antidiabetic and antiplasmodial activities of Garcinia forbesii King: A correlation study

Garcinia forbesii King belongs to Clusiaceae is a source of secondary metabolites especially xanthones with various biological activities. G. forbesii King is also known for its empirical use for malaria and diabetes. This study investigated the total phenolic and flavonoid contents, in vitro antioxidant, antidiabetic and antiplasmodial activities of four extracts attained from the stem bark of G. forbesii King. The total phenolic and flavonoid contents were determined by spectrophotometric methods and antioxidant activity was evaluated by DPPH, ABTS, FRAP assays. In vitro antidiabetic activity was assessed by α-glucosidase and α-amylase assays and antiplasmodial activity was studied against chloroquine sensitive Plasmodium falciparum strain 3D7. The highest value of total phenolic (187.37 ± 0.06 mg GAE/g) and flavonoid (35.97 ± 0.02 mg QE/g) contents were recorded in n-hexane and methanolic extracts. n-Hexane extract showed the highest DPPH activity with IC₅₀ of 8.12 ± 0.02 μg/mL. Ethyl acetate extract exhibited better scavenging ability for ABTS with IC₅₀ of 3.88 ± 0.04 μg/mL. The FRAP assay showed better activity in methanol extract with an inhibition value of 73.68 ± 3.66 µM Fe²⁺/g. The strong inhibition against α-glucosidase and α-amylase were displayed by dichloromethane extract with IC₅₀ of 35.13 ± 2.01 μg/mL and 4.83 ± 0.20 μg/mL. n-Hexane and methanol extracts showed significant antiplasmodial activity with IC₅₀ of 0.23 ± 0.01 μg/mL and 0.73 ± 0.01 μg/mL, respectively. The correlation analysis indicated a positive relationship of total phenolic and flavonoid contents with antiplasmodial activity. The results revealed that n-hexane and methanol extracts could be used as a potential natural antiplasmodial, while dichloromethane extract is a promising natural antidiabetic.     

Phenolic Profile,Antioxidant Capacity and Antimicrobial Activity of Nettle Leaves Extracts Obtained by Advanced Extraction Techniques

Nettle is a widely known plant whose high biological activity and beneficial medicinal effects are attributed to various bioactive compounds, among which polyphenols play an important role. In order to isolate polyphenols and preserve their properties, advanced extraction techniques have been applied to overcome the drawbacks of conventional ones. Therefore, microwave-assisted extraction (MAE) has been optimized for the isolation of nettle leaves polyphenols and it was compared to pressurized liquid extraction (PLE) and conventional heat-reflux extraction (CE). The obtained extracts were analyzed for their individual phenolic profile by UPLC MS² and for their antioxidant capacity by ORAC assay. MAE proved to be the more specific technique for the isolation of individual phenolic compounds, while PLE produced extracts with higher amount of total phenols and higher antioxidant capacity. Both techniques were more effective compared to CE. PLE nettle extract showed antimicrobial activity against bacteria, especially against Gram-negative Pseudomonas fragi ATCC 4973 and Campylobacter jejuni NCTC 11168 strains. This suggests that PLE is suitable for obtaining a nettle extract with antioxidant and antimicrobial potential, which as such has great potential for use as a value-added ingredient in the food and pharmaceutical industry.     

Phenylalanine Increases the Production of Antioxidant Phenolic Acids in Ginkgo biloba Cell Cultures

The aims of this study were to evaluate the antioxidant properties, to investigate the content of major secondary metabolites in Ginkgo biloba cell cultures, and to determine the change in the production of phenolic acids by adding phenylalanine to the culture medium. Three in vitro methods, which depend on different mechanisms, were used for assessing the antioxidant activity of the extract: 1,1-diphenyl-2-picrylhydrazil (DPPH), reducing power and Fe²⁺ chelating activity assays. The extract showed moderate activity both in the DPPH and in the reducing power assays (IC₅₀ = 1.966 ± 0.058 mg/mL; ASE/mL = 16.31 ± 1.20); instead, it was found to possess good chelating properties reaching approximately 70% activity at the highest tested dose. The total phenolic, total flavonoid, and condensed tannin content of G. biloba cell culture extract was spectrophotometrically determined. The phenolic acid content was investigated by RP-HPLC, and the major metabolites—protocatechuic and p-hydroxybenzoic acids—were isolated and investigated by ¹H NMR. The results showed that phenylalanine added to G. biloba cell cultures at concentrations of 100, 150, and 200 mg/150 mL increased the production of phenolic acids. Cultures that were grown for 3 weeks and collected after 4 days of phenylalanine supplementation at high concentration showed maximal content of phenolic acids (73.76 mg/100 g DW).