Ellagic Acid Production Research Articles

Enhanced Production of Therapeutic Metabolites in Cork-Oak Somatic Embryos under Abiotic Stress Conditions

This study investigates the enhanced production of key therapeutic metabolites (ellagic acid, serotonin, and chlorogenic acid) in response to abiotic stress in in vitro cultures of Quercus suber somatic embryos. Findings indicate significant increases in metabolite levels under various stress conditions, highlighting the potential for commercial-scale production of these compounds, known for their antioxidant, anticancer, and anti-COVID-19 properties. Under osmotic/saline stress, ellagic acid production significantly increased, representing an 80% increase compared to control conditions. In embryos exposed to different stressors, serotonin accumulation showed a six-fold increase under osmotic/saline stress. Although the elicitors used did not increase chlorogenic acid levels, exploring alternative stress types may enhance its production. This research paves the way for sustainable, large-scale production of health-beneficial metabolites, addressing global health challenges and promoting resource sustainability.

Light-emitting diode-induced ellagic acid production in Drosera tokaiensis inhibits advanced glycation end-product formation

Drosera tokaiensis (DT) is an insectivorous plant that effectively inhibits advanced glycation end products (AGEs) generated from non-enzymatic reactions between reducing sugars and protein amino groups using phenolic compounds, including ellagic acid, myricitrin, and quercitrin. The accumulation of AGEs increases during the pathogenesis of non-communicable diseases, such as diabetes and nephropathy. Therefore, the extracts of Drosera species could have preventive effects against such diseases. The development of light-emitting diodes (LEDs) and their use in plant cultivation and tissue culture can enhance secondary metabolite production by controlling the wavelength of light in which plants are cultivated. We evaluated how LEDs affected the production of AGE-inhibitory compounds in extracts isolated from DT tissue cultures. DT was cultured for two months in different light wavelengths under unified photosynthetic photon flex density and the resulting extracts were used to examine changes in phenolic compound content and the inhibition of AGE formation. Cultivation in monochromatic light from LEDs had no significant effect on plant weight compared to conventional fluorescent cultivation, except for red LED light. Ellagic acid content increased regardless of LED coloration and the production and accumulation of myricitrin and quercitrin were inhibited. The extracts isolated from plantlets cultured under LEDs showed the same level of AGE inhibition as those cultured under fluorescent light. Therefore, for anti-glycation activity, DT can be grown stably and at a low cost using LED lights when compared to conventional culture methods.

Production and perspective approach of ellagic acid from mango pulp industrial waste by the Micrococcus luteus

AbstractEllagic acid (EA) has gained momentum recently due to its various properties like anti‐mutagenic, anti‐carcinogenic, anti‐oxidant, anti‐viral and many other benefits to human health. The present study focused on the microbial production of EA from mango pulp processing industrial waste as an alternate method for conventional chemical extraction. Our experiments demonstrated that the 100 μg/mL of EA was produced by Micrococcus luteus from 9% of mango pulp waste and the effect of several fermentation parameters of EA production with Pontecorvo medium has yielded 496.75 mg/g at pH 6.0, temperature 35°C, ammonium nitrate (nitrogen source), sucrose (carbon source), potassium chloride (mineral source) with 0.5% of inoculum after 48 h of incubation. EA synthesized was further confirmed with the standard EA in high performance liquid chromatography and Fourier transform infrared spectroscopy. Perspective approaches like drought resistant in plants, anti‐parasitic activity and anti‐cancer activities have been proven and they were docked against depression proteins (receptors), namely MAO‐A using Epic, LigPrep and Glide module of Schrödinger. EA was found to have an ability to bind with the candidate receptor proteins 2Z5X for depression. Microbial bioconversion of mango wastes from pulp processing industries which can reduce the environmental pollution and convert as value added product.

Preparation of pure ellagic acid from waste pomegranate peel by supramolecular solvent based heat assisted extraction followed by liquid–liquid extraction and antisolvent precipitation

In this study, the applicability of supramolecular solvents (SUPRAS) produced by adding water to colloidal suspensions of four bioamphiphiles, namely n -hexanoic acid, n -heptanoic acid, n -octanoic acid, and n -nonanoic acid, in ethanol for the vortex assisted extraction of ellagic acid (EA) from waste pomegranate peel (WPP) was investigated, respectively. SUPRAS prepared by n -hexanoic acid/ethanol/water (30/30/40, w/w/w) and n -heptanoic acid/ethanol/water (10/30/60, w/w/w) showed better extraction efficiency in comparison with other SUPRAS. The influence of helper methods (vortex, ultrasound, and heat) and solid/solvent ratio on the extraction of EA using the above two SUPRAS was studied. SUPRAS-based heat assisted extraction was considered as the optimal one for the extraction of EA for WPP (SUPRAS was prepared by n -hexanoic acid/ethanol/water, 30/30/40, w/w/w). Other operation parameters, i.e. process temperature and extraction time, were also investigated. And then, the SUPRAS extract was further purified by liquid-liquid liquid–liquid extraction using water and ethyl acetate as extraction solvents successively. Finally, pure EA was recovered from the crude EA product by antisolvent precipitation using tetrahydrofuran as solvent and water as antisolvent. Under optimum conditions, the content of EA in the final product was 95.6% and the total yield was about 75%. • A feasible and low-cost strategy for pure ellagic acid production was developed. • Pomegranate peelwas environment-friendlyextracted peel was environment-friendly extracted with supramolecular solvents. • Efficient enrichment effect of LLE made antisolvent precipitation possible. • Tetrahydrofuran is better suited for the antisolvent precipitation of ellagic acid.

Ellagic acid production by solid-state fermentation (SSF) using pomegranate peels as a substrate: a review

Ellagic acid production by solid-state fermentation (SSF) using pomegranate peels as a substrate: a review

OPTIMIZATION OF POMEGRANATE PEEL EXTRACTS FOR THE BIOCONVERSION OF THE ELLAGITANNINS TO ELLAGIC ACID USING ASPERGILLUS NIGER, RHIZOPUS ORYZAE AND MIXED CULTURE

Objective: The present study is aimed at optimization of concentration of substrate (pomegranate peel extract) for the production of Ellagic acid using Aspergillus niger and Rhizopus oryzae. Methods: Test organisms were isolated and identified using standard microbiological techniques. Collected pomegranate peels were dried, grained and used as the substrate for solid-state fermentation. Prepared spore suspension of the test organisms was inoculated and incubated at room temperature. At regular intervals of 24 h, samples were drawn and subjected to analysis of reducing sugar, soluble proteins, hydrolysable tannins and production of extracellular tannase using standard methods. Results: The results of the present investigation demonstrated the optimum substrate concentration for the active conversion of ellagitannins to ellagic was 15g of A. niger, 20g for R. oryzae and 15g for mixed culture. Conclusion: From the current work, it was concluded that solid-state bioprocessing of fruit substrates and fruit wastes using fungi has shown to enrich phenolic antioxidants and improve phytochemical consistency.

Development of a bioprocess for production of ellagic acid from chestnut (Castanea sativa Mill.) waste by fermentation with Aspergillus spp.

Ellagic acid is a polyphenol found in plant-based foods with health benefits. In this study, valorization of chestnut shell (shell: inner pellicle and peel) to produce ellagic acid by fungal fermentation was investigated. Newly isolated black Aspergillus spp. (A. aculeatus ZGM6, A. japonicus ZGM4, A. niger ZDM2, A. tubingensis ZDM1) were screened to evaluate their potential for fermentation. LC-MS/MS and HPLC-PDA were carried out to identify ellagic acid and its derivatives in the fermentation medium. A. japonicus ZGM4 was found to yield the highest level of ellagic acid and used as the test microorganism to determine optimum fermentation conditions. Optimization studies were conducted by the Plackett-Burman Design followed by the Central Composite Design. Concentrations of lactose and yeast extract and fermentation time were found as effective factors on ellagic acid yield. Fermentation of chestnut shell under the optimized conditions (11.5 g/L lactose, 5.9 g/L yeast extract, 140 h) generated 6-fold increase in concentration of ellagic acid. Chestnut shell can be utilized as a natural source of ellagic acid that could be further enriched by fermentation.

Recent Advances in the Production and Applications of Ellagic Acid and Its Derivatives. A Review.

Ellagitannins (ETs), characterized by their diversity and chemical complexity, belong to the class of hydrolysable tannins that, via hydrolysis under acidic or alkaline conditions, can yield ellagic acid (EA). They are mostly found as a part of extractives in angiosperms. As known antioxidants and chelators, EA and EA derivatives are drawing an increasing interest towards extensive technical and biomedical applications. The latter ones include possible antibacterial, antifungal, antiviral, anti-inflammatory, hepato- and cardioprotective, chemopreventive, neuroprotective, anti-diabetic, gastroprotective, antihyperlipidemic, and antidepressant-like activities, among others. EA’s synthesis and production challenges prompt further research on new methods and alternative sources. Conventional and prospective methods and raw materials for the production of EA and its derivatives are reviewed. Among the potential sources of EA, the residues and industrial streams of the pulp industry have been highlighted and considered as an alluring alternative in terms of commercial exploitation.

Enzymatic Biotransformation of Pomegranate Ellagitannins: Initial Approach to Reaction Conditions.

Background: Ellagitannase (Ellagitannin acyl hydrolase) is an inducible enzyme with great potential use in food industry since allows the ellagic acid release from ellagitannins.Objective: In this work, ellagitannase was produced by the fungus Aspergillus niger GH1 in solid state fermentation using polyurethane foam as solid support and pomegranate husk ellagitannins as sole carbon source and ellagitannase inducer and an initial approach to the enzymatic reaction conditions was reached.Materials and Methods: Ellagitannase was produced by Aspergillus niger GH1 in solid state fermentation and the ideal reaction conditions for ellagitannase activity based on ellagic acid quantification as ellagitannins biotransformation product by high performance liquid chromatographic are reported.Results:The enzyme ideal reaction conditions were substrate concentration of 1 mg.mL-1, 60 °C and pH 5.0, during 10 min of reaction. The kinetic enzyme constants (Vmax = 30.34 mM.mL-1.min-1 and Km = 1.48 x 103 mM) using punicalagin assubstrate were determined. Conclusion: The assay was completed in a short time and may find application in future studies of ellagic acid production.

BIOCONVERSION OF ELLAGITANNINS TO ELLAGIC ACID FROM POMEGRANATE PEELS BY SOLID-STATE FERMENTATION USING ASPERGILLUS NIGER AND RHIZOPUS ORYZAE

Ellagic acid an effective polyphenol antioxidant found in many fruits and vegetables. The antioxidant properties of ellagic acid have incited preliminary research into the impending health benefits of ellagic acid consumption. In the present research, an attempt is made to extract ellagic acid from pomegranate peels using solid state cultures of Aspergillus niger, Rhizopus oryzae and mixed cultures of both. The fungus has the potential to convert ellagitannins in pomegranate to ellagic acid as an intermediary metabolite. Test organisms were isolated from soil, identified and screened for production of tannase enzyme. Solid state fermentation of pomegranate husk (10% w/v) using A. niger, R. oryzae and mixed cultures was performed for the production of ellagic acid. Samples were collected daily from the fermentation broth and physico-chemical analyses like changes in the total tannin content and tannase enzyme assay were performed. After fermentation the production of ellagic acid in the medium was determined using HPLC. The amount of ellagic acid produced was 9.1mg (for A.niger), 79.1mg (for R. oryzae), 69.6mg (for mixed culture of both A. niger and R.oryzae).

Ellagic acid production using polyphenols from orange peel waste by submerged fermentation

Abstract Background Biotechnological processes are part of modern industry as well as stricter environmental requirements. The need to reduce production costs and pollution demands for alternatives that involve the integral use of agro-industrial waste to produce bioactive compounds. The citrus industry generates large amounts of wastes due to the destruction of the fruits by microorganisms and insects together with the large amounts of orange waste generated during the production of juice and for sale fresh. The aim of this study was used orange wastes rich in polyphenolic compounds can be used as source carbon of Aspergillus fumigatus MUM 1603 to generate high added value compounds, for example, ellagic acid and other molecules of polyphenolic origin through submerged fermentation system. Results The orange peel waste had a high concentration of polyphenols, 28% being condensed, 27% ellagitannins, 25% flavonoids and 20% gallotannins. The major polyphenolic compounds were catechin, EA and quercetin. The conditions, using an experimental design of central compounds, that allow the production of the maximum concentration of EA (18.68 mg/g) were found to be: temperature 30°C, inoculum 2 × 107 (spores/g) and orange peel polyphenols 6.2 (g/L). Conclusion The submerged fermentation process is an effective methodology for the biotransformation of molecules present in orange waste to obtain high value-added as ellagic acid that can be used as powerful antioxidants, antibacterial and other applications. How to cite: Sepulveda L, Laredo-Alcala E, Buenrostro-Figueroa JJ, et al. Ellagic acid production using polyphenols from orange peel waste by submerged fermentation. Electron J Biotechnol 2020;43. https://doi.org/10.1016/j.ejbt.2019.11.002 .

Natural production and quantification of ellagic acid in multiple plant parts of three Terminalia spp.

Ellagic acid (EA) is a natural phenol antioxidant, found in multiple fruits. Chemically, it is a dilactone of hexahydroxydiphenic acid. Owing to its immensely beneficial anti-proliferative, antioxidant, and anticancerous activities, it needs to be quantified in natural resources. An effective densitometric high-performance liquid chromatography (HPTLC) method was established for the simultaneous quantification of ellagic acid from different plant parts of Terminalia arjuna, T. bellerica, and T. chebula. The method was validated for intra-and inter-day precision, reproducibility, and specificity. The intraday precision and interday precision was 1.5% and 1.2% relative standard deviation (RSD), respectively. The reproducibility of standard EA was precise and the seven lanes provided similar banding pattern. The linearity curve for the EA produced a correlation coefficient (r) value of 0.98992, with regression equation Y=6330.441+935.150*X, and with a standard deviation value of 4.59%. For chromatogram development, toluene: ethyl acetate: formic acid (5: 5: 2.5) was optimized as mobile phase. With the exception of fruits of T. bellerica, all the plant parts of all three species contain a significant amount of EA; the highest EA content being (0.733 μg/μg dry weight) found in the roots of T. bellerica. In this study, the proposed HPTLC method for simultaneous quantification of EA was conducted in a simple, sensitive, specific, and precise manner that can be recommended for routine quality control and quantification of EA in plant materials.

Ellagic acid production by solid-state fermentation influenced by the inert solid supports

Ellagic acid and ellagitannins are defense phytochemicals and dietary polyphenols found in fruits and nuts and implicated with strong antioxidant, anticancer and antiatherosclerotic biological properties. Unfortunately, there are few reports on the bioconversion studies from ellagitannins to ellagic acid. The objective of the present work was to use of partially purified ellagitannins from pomegranate peels as inducers for production of ellagic acid by Aspergillus niger GH1 in solid-state fermentation on inert supports (perlite, polyurethane foam and nylon fiber). For total pomegranate polyphenols (TPP), two methodologies were compared. Higher recovery of TTP (70 g kg-1 peel) in a short time (2 h) were obtained by filtering through a muslin cloth followed by centrifugation at 3000 rpm for 15 min. Production of ellagic acid and ellagitannase enzyme were evaluated. Maximum ellagic acid production (231.22 mg g-1 TPP) at 24 h cultivation with an ellagitannase activity of 3213 U L-1 was obtained when using polyurethane foam as inert support. Polyurethane foam was the best solid support to produce high yields of ellagic acid production.

Solid state fermentation of pomegranate husk: Recovery of ellagic acid by SEC and identification of ellagitannins by HPLC/ESI/MS

Abstract 2, 3, 7, 8-Tetrahydroxy-chromeno [5, 4, 3-cde] chromene-5, 10-dione, commonly named ellagic acid was successfully separated by size-exclusion chromatography (SEC) from pomegranate husk ellagitannins purified by solid-state fermentation. Recovery of metabolites from fermented mash was carried out by the addition of ethanol and manual pressing. Culture extracts were loaded on a Sephadex LH-20 column, three fractions were separated, and second fraction was identified and evaluated by high performance liquid chromatography followed by electro spray ionization and mass spectrometry (HPLC/ESI/MS). It was possible to obtain a yield of 47 mg/g of pomegranate husk polyphenols. The analysis of HPLC/ESI/MS allowed to identify punicalagin (1082.97 m/z), punicalin (780.99 m/z) and ellagic acid (300.80 m/z). After the fermentation process, the main compound was ellagic acid. The present study describes a rapid and effective fermentation/chromatographic separation process for ellagic acid production.

Exploring the Degradation of Gallotannins Catalyzed by Tannase Produced by Aspergillus niger GH1 for Ellagic Acid Production in Submerged and Solid-State Fermentation.

Due to great interest on producing bioactive compounds for functional foods and biopharmaceuticals, it is important to explore the microbial degradation of potential sources of target biomolecules. Gallotannins are polyphenols present in nature, an example of them is tannic acid which is susceptible to enzymatic hydrolysis. This hydrolysis is performed by tannase or tannin acyl hydrolase, releasing in this way, biomolecules with high-added value. In the present study, chemical profiles obtained after fungal degradation of tannic acid under two bioprocesses (submerged fermentation (SmF) and solid state fermentation (SSF)) were determined. In both fermentation systems (SmF and SSF), Aspergillus niger GH1 strain and tannic acid as a sole carbon source and inducer were used (the presence of tannic acid promotes production of enzyme tannase). In case of SSF, polyurethane foam (PUF) was used like as support of fermentation; culture medium only was used in case of submerged fermentation. Fermentation processes were monitored during 72h; samples were taken kinetically every 8h; and all extracts obtained were partially purified to obtain polyphenolic fraction and then were analyzed by liquid chromatography-mass spectrometry (LC-MS). Molecules like gallic acid and n-galloyl glucose were identified as intermediates in degradation of tannic acid; during SSF was identified ellagic acid production. The results obtained in this study will contribute to biotechnological production of ellagic acid.

On-line monitoring of Aspergillus niger GH1 growth in a bioprocess for the production of ellagic acid and ellagitannase by solid-state fermentation

The present work describes the monitoring of CO2 production by Aspergillus niger GH1 in a bioprocess for the production of ellagitannase (EAH) and ellagic acid by solid state fermentation. Pomegranate ellagitannins, mainly punicalagin, were used as carbon source and EAH inducer. A second condition, using ellagitannins and maltose as growth promoting carbon source, was tested. The ellagic acid production was quantified and the EAH activity was assayed. The accumulated metabolites were identified by HPLC-ESI-MS/MS. Higher CO2 production (7.79mg/grams of dry material) was reached in media supplemented with maltose. Short-time lag phase (7.79h) and exponential phase (10.42h) were obtained using only ellagitannins, despite its lower CO2 production (3.79mg/grams of dry material). Without the use of maltose lower ellagic acid (11.85mg/L/h) and EAH (21.80U/L/h) productivities were reached. The use of maltose enhances the productivity of EA (33.18mg/L/h) and EAH (33.70U/L/h). Besides of punicalin and ellagic acid, two unknown compounds with mass weight of 702 and 290g/mol (ions 701 and 289m/z in negative mode, respectively) were identified and characterized by HPLC-ESI-MS/MS analysis.

Biotransformation of hydrolysable tannin to ellagic acid by tannase from Aspergillus awamori

Madhuca indica, locally known as mahua in India is a multipurpose tree species. Mahua, particularly bark contains a significant amount of hydrolysable tannin (17.31%) which can be utilized for ellagic acid production through biotransformation. In the present study, mahua bark utilized not only as a raw material for tannase production but also for ellagic acid a well-known therapeutic compound. After prior confirmation of hydrolysable tannin content in bark, it has been supplemented, as a substrate for tannase production through solid state fermentation of Aspergillus awamori. Tannase production, as well as biodegradation of the hydrolysable tannin reached a maximum at 72 h of incubation time. The optimum conditions for tannase production are solid to liquid ratio of 1:2, 35 °C, pH 5.5 and 72h incubation time which resulted 0.256 mg/mL of an extract of ellagic acid. Maximum tannase activity of 56.16 IU/gds at 35 °C and 72h of incubation time is recorded. It seems that tannase production and biotransformation of hydrolysable tannins using bark powder of mahua can be considered as an appropriate alternative to the existing procedures of ellagic acid production.

The complete biodegradation pathway of ellagitannins by Aspergillus niger in solid‐state fermentation

Our research group has found preliminary evidences of the fungal biodegradation pathway of ellagitannins, revealing first the existence of an enzyme responsible for ellagitannins degradation, which hydrolyzes pomegranate ellagitannins and it was called ellagitannase or elagitannin acyl hydrolase. However, it is necessary to generate new and clear information in order to understand the ellagitannin degradation mechanisms. This work describes the distinctive and unique features of ellagitannin metabolism in fungi. In this study, hydrolysis of pomegranate ellagitannins by Aspergillus niger GH1 was studied by solid-state culture using polyurethane foam as support and pomegranate ellagitannins as substrate. The experiment was performed during 36 h. Results showed that ellagitannin biodegradation started after 6 h of fermentation, reaching the maximal biodegradation value at 18 h. It was observed that ellagitannase activity appeared after 6 h of culture, then, the enzymatic activity was maintained up to 24 h of culture reaching 390.15 U/L, after this period the enzymatic activity decreased. Electrophoretic band for ellagitannase was observed at 18 h. A band obtained using non-denaturing electrophoresis was identified as ellagitannase, then, a tandem analysis to reveal the ellagitannase activity was performed using Petri plate with pomegranate ellagitannins. The extracts were analyzed by HPLC/MS to evaluate ellagitannins degradation. Punicalin, gallagic acid, and ellagic acid were obtained from punicalagin. HPLC/MS analysis identified the gallagic acid as an intermediate molecule and immediate precursor of ellagic acid. The potential application of catabolic metabolism of ellagitannin hydrolysis for ellagic acid production is outlined.

Simultaneous extraction and biotransformation process to obtain high bioactivity phenolic compounds from Brazilian citrus residues.

Recent studies have pointed to a reduction in the incidence of some cancers, diabetes, and neuro-degenerative diseases as a result of human health benefits from flavanones. Currently, flavanones are obtained by chemical synthesis or extraction from plants, and these processes are only produced in the glycosylated form. An interesting environmentally friendly alternative that deserves attention regarding phenolic compound production is the simultaneous extraction and biotransformation of these molecules. Orange juice consumption has become a worldwide dietary habit and Brazil is the largest producer of orange juice in the world. Approximately half of the citrus fruit is discarded after the juice is processed, thus generating large amounts of residues (peel and pectinolytic material). Hence, finding an environmentally clean technique to extract natural products and bioactive compounds from different plant materials has presented a challenging task over the last decades. The aim of this study was to obtain phenolics from Brazilian citrus residues with high bioactivity, using simultaneous extraction (cellulase and pectinase) and biotransformation (tannase) by enzymatic process. The highest hesperetin, naringenin and ellagic acid production in the experiment were 120, 80, and 11,250 µg g(-1), respectively, at 5.0 U mL(-1) of cellulase and 7.0 U mL(-1) of tannase at 40°C and 200 rpm. Also, the development of this process generated an increase of 77% in the total antioxidant capacity. These results suggest that the bioprocess obtained innovative results where the simultaneous enzymatic and biotransformatic extracted flavanones from agro-industrial residues was achieved without the use of organic solvents. The methodology can therefore be considered a green technology.

Optimization of Ellagitannase Production by Aspergillus niger GH1 by Solid-State Fermentation

Ellagic acid is one of the most bioactive antioxidants with important applications in pharmaceutical, cosmetic, and food industries. However, there are few biotechnological processes developed for its production, because it requires precursors (ellagitannins) and the corresponding biocatalyst (ellagitannase). The aim of this study was to optimize the culture conditions for ellagitannase production by Aspergillus niger in solid-state fermentation (SSF). The bioprocess was carried out into a column bioreactor packed with polyurethane foam impregnated with an ellagitannins solution as carbon source. Four strains of Aspergillus niger (PSH, GH1, HT4, and HC2) were evaluated for ellagitannase production. The study was performed in two experimental steps. A Plackett–Burman design was used to determine the influencing parameters on ellagitannase production. Ellagitannins concentration, KCl, and MgSO4 were determined to be the most significant parameters. Box–Behnken design was used to define the interaction of the selected parameters. The highest enzyme value was obtained by A. niger PSH at concentrations of 7.5 g/L ellagitannins, 3.04 g/L KCl, and 0.76 g/L MgSO4. The methodology followed here allowed increasing the ellagitannase activity 10 times over other researcher results (938.8 U/g ellagitannins). These results are significantly higher than those reported previously and represent an important contribution for the establishment of a new bioprocess for ellagic acid and ellagitannase production.