Production Of Α-tocopherol Research Articles

Elicitation and precursor induced approaches for the enhancement of α−tocopherol production using suspension cultures of Solanum lycopersicum

Elicitation and precursor feeding are the effective strategies for enhancing the synthesis of bioactive compounds in plant cell suspension cultures. The present study aimed to explore an efficient elicitation and precursor feeding protocol and its effect on inducing the accumulation of α-tocopherol in Solanum lycopersicum (tomato) suspension cell culture. The tomato cell suspension cultures were treated with different elicitors (Methyl Jasmonate, Salicylic acid and Yeast extract) and precursors (Homogentisic acid, Tyrosine, Hydroxypyruvic acid and Phytol) and the effect of α-tocopherol production was studied. Significant increase in the α-tocopherol was observed on day 5 upon methyl jasmonate treatment which represented 17.7 fold increase in comparison to the control. The treatment of precursor in combination viz., 150 μM Homogentisic acid + 150 μM Phytol showed the maximum enhancement of α-tocopherol up to 22 fold on day 10 compared to the untreated control. These results suggested that the suspension cultures combining with the optimal precursor feeding and elicitors enhanced the production of α-tocopherol in economically important tomato cell cultures.

Elicitor-Mediated Enhancement of α-Tocopherol in Cell Suspension Cultures of Nicotiana tabacum

Plant cell and tissue culture have been used as the alternative and potential renewable source for the production of valuable phytochemicals. Elicitation offers a reliable in vitro approach to produce or enhance potential phytochemicals. α-tocopherol, which is an isoform of vitamin E, is a potent fat-soluble phytochemical known in nature. The present study focused on enhancing the production of α-tocopherol in the cell suspension culture through an elicitation approach. Suspension cultures of Nicotiana tabacum were established from the leaf disk-derived callus. The cell suspension cultures were treated with different elicitors (methyl jasmonate, salicylic acid, and yeast extract) at the lag phase of the cell growth cycle. The effects of elicitors on cell cultures were determined in terms of biomass, and α-tocopherol enhancement was determined using high-performance liquid chromatography (HPLC). Different elicitors depending on the concentration exerted different effects on cell growth and α-tocopherol production. Methyl jasmonate treatment showed the significantly highest increase in α-tocopherol on the 6th day of treatment in tobacco suspension cultures. Methyl jasmonate at the concentration of 150 μM enhanced α-tocopherol content to 24-fold over the control. This study clearly shows that the elicitors had the potential to increase the accumulation of α-tocopherol considerably in tobacco cell cultures. The outcomes of this study could be of considerable importance to the nutraceutical and pharmaceutical industries.

Metabolic response induced by abiotic elicitors in Anacardium othonianum seedlings

Applying elicitors in plants growing under natural conditions change the metabolic content of volatile compounds extracted from their tissues. Here, we have investigated how abiotic elicitors influence the volatile composition of the ethyl acetate extract of Anacardium othonianum seedlings propagated in vitro and ex-vitro cultures. Abiotic factors such as salicylic acid, silver nitrate, light quality, and cultivation mode affected the composition of volatile compounds in ethyl acetate extract of A. othonianum leaves seedlings. GC/MS analysis demonstrated that α-tocopherol was the main metabolite in cultures added with chemical elicitors, reaching 22.48 ± 4.06% in the case of 30 µM salicylic acid. Higher concentrations of salicylic acid decreased the amount of α-tocopherol. The addition of AgNO3 had a linear effect on α-tocopherol. Thus, the production of this metabolite indicates that salicylic acid and AgNO3 elicitors positively affect α-tocopherol production at specific concentrations helping to understand how plant culture can be better manipulated.

Production of α-Tocopherol-Chitosan Nanoparticles by Membrane Emulsification.

α-tocopherol (α-T) has the highest biological activity with respect to the other components of vitamin E; however, conventional formulations of tocopherol often fail to provide satisfactory bioavailability due to its hydrophobic characteristics. In this work, α-tocopherol-loaded nanoparticles based on chitosan were produced by membrane emulsification (ME). A new derivative was obtained by the cross-linking reaction between α-T and chitosan (CH) to preserve its biological activity. ME was selected as a method for nanoparticle production because it is recognized as an innovative and sustainable technology for its uniform-particle production with tuned sizes and high encapsulation efficiency (EE%), and its ability to preserve the functional properties of bioactive ingredients operating in mild conditions. The reaction intermediates and the final product were characterized by 1HNMR, Fourier-transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC), while the morphological and dimensional properties of the nanoparticles were analyzed using electronic scanning microscopy (SEM) and dynamic light scattering (DLS). The results demonstrated that ME has high potential for the development of α-tocopherol-loaded nanoparticles with a high degree of uniformity (PDI lower than 0.2), an EE of almost 100% and good mechanical strength, resulting in good candidates for the production of functional nanostructured materials for drug delivery. In addition, the chemical bonding between chitosan and α-tocopherol allowed the preservation of the antioxidant properties of the bioactive molecule, as demonstrated by an enhanced antioxidant property and evaluated through in vitro tests, with respect to the starting materials.

FRET-Based Genetically Encoded Nanosensor for Real-Time Monitoring of the Flux of α-Tocopherol in Living Cells

Vitamin E plays an exemplary role in living organisms. α-Tocopherol is the most superior and active form of naturally occurring vitamin E that meets the requirements of human beings as it possesses the α-tocopherol transfer protein (α-TTP). α-Tocopherol deficiency can lead to severe anemia, certain cancers, several neurodegenerative and cardiovascular diseases, and most importantly male infertility. As a result of the depletion of its natural sources, researchers have tried to employ metabolic engineering to enhance α-tocopherol production to meet the human consumption demand. However, the metabolic engineering approach relies on the metabolic flux of a metabolite in its biosynthetic pathway. Analysis of the metabolic flux of a metabolite needs a method that can monitor the α-tocopherol level in living cells. This study was undertaken to construct a FRET (fluorescence resonance energy transfer)-based nanosensor for monitoring the α-tocopherol flux in prokaryotic and eukaryotic living cells. The human α-TTP was sandwiched between a pair of FRET fluorophores to construct the nanosensor, which was denoted as FLIP-α (the fluorescence indicator for α-tocopherol). FLIP-α showed excellence in monitoring the α-tocopherol flux with high specificity. The sensor was examined for its pH stability for physiological applications, where it shows no pH hindrance to its activity. The calculated affinity of this nanosensor was 100 μM. It monitored the real-time flux of α-tocopherol in bacterial and yeast cells, proving its biocompatibility in monitoring the α-tocopherol dynamics in living cells. Being noninvasive, FLIP-α provides high temporal and spatial resolutions, which holds an indispensable significance in bioimaging metabolic pathways that are highly compartmentalized.

Phytol, a Constituent of Chlorophyll, Induces Root-Knot Nematode Resistance in Arabidopsis via the Ethylene Signaling Pathway.

Root-knot nematodes (RKNs; Meloidogyne spp.) parasitize the roots or stems of a wide range of plant species, resulting in severe damage to the parasitized plant. The phytohormone ethylene (ET) plays an important role in signal transduction pathways leading to resistance against RKNs. However, little is currently known about the induction mechanisms of ET-dependent RKN resistance. Inoculation of Arabidopsis thaliana roots with RKNs decreased chlorophyll contents in aerial parts of the plant. We observed accumulation of phytol, a constituent of chlorophyll and a precursor of tocopherols, in RKN-parasitized roots. Application of sclareol, a diterpene that has been shown to induce ET-dependent RKN resistance, to the roots of Arabidopsis plants increased phytol contents in roots accompanied by a decrease in chlorophyll in aerial parts. Exogenously applied phytol inhibited RKN penetration of roots without exhibiting nematicidal activity. This phytol-induced inhibition of RKN penetration was attenuated in the ET-insensitive Arabidopsis mutant ein2-1. Exogenously applied phytol enhanced the production of α-tocopherol and expression of VTE5, a gene involved in tocopherol production, in Arabidopsis roots. α-Tocopherol exerted induction of RKN resistance similar to that of phytol and showed increased accumulation in roots inoculated with RKNs. Furthermore, the Arabidopsis vte5 mutant displayed no inhibition of RKN penetration in response to phytol. These results suggest that exogenously applied phytol induces EIN2-dependent RKN resistance, possibly via tocopherol production.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Essential role for phytol kinase and tocopherol in tolerance to combined light and temperature stress in tomato.

In a changing environment, plants need to cope with the impact of rising temperatures together with high light intensity. Here, we used lipidomics in the tomato model system to identify lipophilic molecules that enhance tolerance to combined high-temperature and high-light stress. Among several hundred metabolites, the two most strongly up-regulated compounds were α-tocopherol and plastoquinone/plastoquinol. Both are well-known lipid antioxidants and contribute to the protection of photosystem II (PSII) against photodamage under environmental stress. To address the protective function of tocopherol, an RNAi line (vte5) with decreased expression of VTE5 and reduced levels of α-tocopherol was selected. VTE5 encodes phytol kinase, which acts in the biosynthetic pathway of tocopherols. vte5 suffered strong photoinhibition and photobleaching when exposed to combined high-light and high-temperature stress, but neither stress alone produced a visible phenotype. As vte5 had plastoquinone levels similar to those of the wild type under combined stress, the strong phenotype could be attributed to the lack of α-tocopherol. These findings suggest that VTE5 protects against combined high-light and high-temperature stress and does so by supporting α-tocopherol production.

Screening of microalgae and cyanobacteria strains for α-tocopherol content at different growth phases and the influence of nitrate reduction on α-tocopherol production

Tocopherols (vitamin E) are only synthesized by photosynthetic organisms and have wide applications in cosmetics and as dietary supplements in human nutrition and aquaculture. Tocopherols from microalgae and cyanobacteria are rarely investigated, and little is known about their contents. Therefore, 130 strains of cultured microalgae and cyanobacteria were analyzed for α-tocopherol content under various culture conditions. The growth phase had a significant effect on content of α-tocopherol. Maximal amounts were observed at the stationary growth phase. Reduction of nitrate concentration in media caused an increased production of α-tocopherol. The contents were significantly enhanced when the nitrate concentration was reduced to one fourth in culture media used. The content of α-tocopherol was found to reflect phylogenetic relationships at the level of classes, with classes of Rhodophyta and Cyanobacteria accumulating the lowest contents. Within each class, contents varied widely at the species level emphasizing the importance of extensive screening procedures for the identification of strains with high α-tocopherol contents.

Applicability of Euglena gracilis for biorefineries demonstrated by the production of α-tocopherol and paramylon followed by anaerobic digestion

In this study the use of Euglena gracilis biomass for α-tocopherol, paramylon and biogas production in a value-added chain was investigated. Therefore, we analyzed the dry cell weight and product concentrations at different growth phases during heterotrophic, photoheterotrophic and photoautotrophic cultivation in a low-cost minimal medium. Furthermore, the specific biogas yields for differently derived biomass with and without product recovery were investigated. We demonstrate that growth phase and cultivation mode not only have a significant impact on product formation, but also influence the yield of biogas obtained from anaerobic digestion of Euglena gracilis biomass. The maximum dry cell weight concentration ranged from 12.3±0.14gL−1 for heterotrophically to 3.4±0.02gL−1 for photoautotrophically grown Euglena gracilis cells. The heterotrophically grown biomass accumulated product concentrations of 5.3±0.12mgL−1 of α-tocopherol and 9.3±0.1gL−1 of paramylon or 805±10.9mL of biogasgvs−1 (per gram volatile solids). The results for photoautotrophically grown cells were 8.6±0.22mgL−1 of α-tocopherol and 0.78±0.01gL−1 of paramylon or 648±7.2mL of biogasgvs−1. For an energy-saving downstream procedure the extracting agent methanol does not have to be removed strictly. Samples with residual methanol showed a significantly increased biogas yield, because the solvent can be used as an additional substrate for methane production by archaebacteria.

Elicitors, SA and MJ enhance carotenoids and tocopherol biosynthesis and expression of antioxidant related genes in Moringa oleifera Lam. leaves

Moringa oleifera Lam. leaves are rich source of carotenoids (provitamin A) and α-tocopherol (vitamin E), and there is a scope for their further enhancement, through elicitor mediation, thereby a great potential for addressing these vitamins deficiency. In the present study, we report the efficacy of foliar administration of biotic elicitors, carboxy-methyl chitosan and chitosan, and signaling molecules, methyl jasmonate (MJ) and salicylic acid (SA) for enhancement of major carotenoids and α-tocopherol. Highest α-tocopherol content of 49.7 mg/100 g FW was recorded upon foliar application of 0.1 mM SA after 24 h of treatment, which represented a 187.5 % increase in comparison to the untreated control. Similarly, a maximum of 52.6 mg/100 g FW lutein, and 21.8 mg/100 g FW β-carotene content were observed in leaves after 24 h of treatment with MJ, which represented a 54.0 and 20.3 % increase in comparison to the untreated control, respectively. Among the major genes of carotenoid biosynthetic pathway, the expression of lycopene β-cyclase (LCY-β) was maximum influenced after treatment with elicitors and signaling molecules, compared to phytoene synthase and phytoene desaturase, suggesting the LCY-β-mediated enhancement in the production of β-carotene in elicitor treated M. oleifera leaves. Enhanced production of α-tocopherol under respective elicitor treatment was further supported by 2.0–2.7 fold up-regulation of γ-tocopherol methyl transferase, compared to untreated control. This is the first report on elicitor-mediated enhanced production of tocopherol and carotenoids in foliage of economically important food plant.

Induction of a photomixotrophic plant cell culture of Helianthus annuus and optimization of culture conditions for improved α-tocopherol production

Tocopherols, collectively known as vitamin E, are lipophilic antioxidants, which are synthesized only by photosynthetic organisms. Due to their enormous potential to protect cells from oxidative damage, tocopherols are used, e.g., as nutraceuticals and additives in pharmaceuticals. The most biologically active form of vitamin E is α-tocopherol. Most tocopherols are currently produced via chemical synthesis. Nevertheless, this always results in a racemic mixture of different and less effective stereoisomers because the natural isomer has the highest biological activity. Therefore, tocopherols synthesized in natural sources are preferred for medical purposes. The annual sunflower (Helianthus annuus L.) is a well-known source for α-tocopherol. Within the presented work, sunflower callus and suspension cultures were established growing under photomixotrophic conditions to enhance α-tocopherol yield. The most efficient callus induction was achieved with sunflower stems cultivated on solid Murashige and Skoog medium supplemented with 30 g l(-1) sucrose, 0.5 mg l(-1) of the auxin 1-naphthalene acetic acid, and 0.5 mg l(-1) of the cytokinin 6-benzylaminopurine. Photomixotrophic sunflower suspension cultures were induced by transferring previously established callus into liquid medium. The effects of light intensity, sugar concentration, and culture age on growth rate and α-tocopherol synthesis rate were characterized. A considerable increase (max. 230%) of α-tocopherol production in the cells was obtained within the photomixotrophic cell culture compared to a heterotrophic cell culture. These results will be useful for improving α-tocopherol yields of plant in vitro cultures.

INDUCED PRODUCTION OF α-TOCOTRIENOL BY CO-SUPPRESSION OF TOCOPHEROL CYCLASE GENE IN VEGETABLE CROPS

This study involves the induced production of α-tocotrienol by the co-suppression of tocopherol cyclase gene. The Tocopherol/Tocotrienol Cyclase (TC) is a key enzyme involved in the biosynthesis of α-tocotrienol. The partial conserved cDNA of TC gene isolated from the oil palm (Elaeis guineensis) that shared greater than 75% sequence identity with TC sequences from other plant species was used to silence TC gene expression in Amaranthus sp. and Allium porrum leaves by co-suppression strategies. This was carried out using recombinant vector constructs designated as p4a11 and p4c9 containing TC driven by the maize ubiquitin promoter (Ubi1P) and the oil palm leaf-specific promoter (LHCB), respectively. In the Amaranthus sp. leaves infiltrated with p4a11, the α-tocopherol suppression was 85.0, 71.0 and 74.0% on day three, five and seven, respectively and complete suppression was observed in the leaves infiltrated with p4c9. Whereas, only 11.0, 7.0 and 29.0% suppression was detected in Allium porrum leaves infiltrated with p4a11 and 45.0, 4.0 and 57.0% of suppression was observed in Allium porrum infiltrated with p4c9 on day three, five and seven, respectively. Silencing of TC gene driven by LHCB (p4c9) causes the reduction of α-tocopherol production better than by Ubi1P (p4a11). The results have demonstrated that gene silencing through co-suppression is capable of producing α-tocotrienol. However, there is no correlation between the α-tocotrienol production and the α-tocopherol suppression. It has been suggested that the transgene mediated co-suppression mechanism had triggered the vitamin E biosynthesis metabolic pathway to shift towards producing α-tocotrienol.

Cloning and characterization of a g-tocopherol methyltransferase from Chinese cabbage

α-Tocopherol, with antioxidant properties, is synthesized by photosynthetic organisms and play important roles in human and animal nutrition. The final step of the α-tocopherol biosynthetic pathway is catalyzed by γ-tocopherol methyltransferase (γ-TMT). In the major oilseed crops, γ-tocopherol, the biosynthetic precursor of α-tocopherol, is the predominant form. The full-length cDNA of γ-TMT was obtained from Chinese cabbage ( Brassica. rapa , Pekinensis Group), named BoTMT, by reverse transcription polymerase chain reaction (RT-PCR). Sequence analysis indicates that, BoTMT gene consisted of the open reading frame of 1041 nucleotides encoding a protein of 39 kD polypeptide. A BoTMT whole cell system was developed for the production of α-tocopherol through the expression of BoTMT in Escherichia coli BL21 (DE3) strain. Keywords: γ-Tocopherol methyltransferase, Chinese cabbage, Perilla frustescens , tocopherol, high performance liquid chromatography (HPLC)

Methyltrioxorhenium-catalyzed oxidation of pseudocumene for vitamin E synthesis: A study of solvent and ligand effects

Vitamin E is an essential food component of major economical relevance with important antioxidant properties and biological activity. The oxidation of pseudocumene to trimethyl-1,4-benzoquinone would be a key transformation in an alternative industrial production of α-tocopherol that is important for the antioxidant activity of vitamin E. The methyltrioxorhenium (MTO)-catalyzed oxidation of pseudocumene has been revisited to offer a more environmentally friendly, economically beneficial and milder approach to this important industrial product. It has been observed that by choosing the solvent and Lewis base additives (as ligands of MTO), both yield and chemoselectivity are considerably improved, allowing milder reaction conditions compared to previously reported protocols.

Influence of growth regulators and elicitors on cell growth and α-tocopherol and pigment productions in cell cultures of Carthamus tinctorius L.

The present study examined the effects of plant growth hormones, incubation period, biotic (Trametes versicolor, Mucor sp., Penicillium notatum, Rhizopus stolonifer, and Fusarium oxysporum) and abiotic (NaCl, MgSO(4), FeSO(4), ZnSO(4), and FeCl(3)) elicitors on cell growth and α-tocopherol and pigment (red and yellow) productions in Carthamus tinctorius cell cultures. The cell growth and α-tocopherol and pigment contents improved significantly on Murashige and Skoog (MS) liquid medium containing 50.0 μM α-naphthalene acetic acid (NAA) and 2.5 μM 6-Benzyladenine (BA) at 28 days of incubation period. Incorporation of T. versicolor (50 mg l(-1)) significantly enhanced the production of α-tocopherol (12.7-fold) and red pigment (4.24-fold). Similarly, supplementation of 30 mg l(-1) T. versicolor (7.54-fold) and 70 mg l(-1) Mucor sp. (7.40-fold) significantly increased the production of yellow pigment. Among abiotic elicitors, NaCl (50-70 mg l(-1)) and MgSO(4) (10-30 mg l(-1)) significantly improved production of α-tocopherol (1.24-fold) and red pigment (20-fold), whereas yellow pigment content increased considerably by all the abiotic elicitor treatments. Taken together, the present study reports improved productions of α-tocopherol and the pigment as a stress response of safflower cell cultures exposed to these elicitors.

Supplemental vitamin E and selenium effects on egg production, egg quality, and egg deposition of α-tocopherol and selenium

SUMMARY Diets were fed to laying hens for 12 wk during hot summer weather (34 to 35°C high temperatures daily) to investigate the effects of feeding higher levels of the dietary antioxidants dl -α-tocopherol and selenium (from 2 sources, inorganic or organic) on egg production and egg quality. High basal levels of selenium were present in the corn-soybean meal diets (0.25 ppm), resulting in selenium treatment levels of 0.55 or 0.75 ppm; supplemented α-tocopherol treatments were 50, 100, or 150 IU/kg of diet. Increasing dietary selenium had a positive effect on egg production and egg mass (g of egg/d) as well as stored egg vitelline membrane strength. Vitamin E supplementation did not affect egg production except during wk 7 of the trial, during a particularly hot period in which hens fed the higher levels of vitamin E (100 or 150 ppm) did not have a decline in egg production. Yolk vitelline membrane strength improved with vitamin E supplementation in fresh eggs and eggs stored for 2 wk. Vitamin E supplementation also affected egg pH, improving albumen pH by decreasing pH of freshly laid eggs. Yolk α-tocopherol increased linearly with vitamin E supplementation. Yolk selenium content also increased with dietary Se supplementation and was deposited more efficiently when feeding the organic source (Sel-Plex) compared with the inorganic source of selenium. In summary, vitamin E and selenium can be supplemented to a laying hen ration to improve the vitelline membrane strength of fresh and aged eggs while also increasing the levels of these nutrients in the egg yolk.

Increased synthesis of α-tocopherol, paramylon and tyrosine by Euglena gracilis under conditions of high biomass production

To analyse the production of different metabolites by dark-grown Euglena gracilis under conditions found to render high cell growth. The combination of glutamate (5 g l(-1) ), malate (2 g l(-1) ) and ethanol (10 ml l(-1) ) (GM + EtOH); glutamate (7·15 g l(-1) ) and ethanol (10 ml l(-1) ); or malate (8·16 g l(-1) ), glucose (10·6 g l(-1) ) and NH(4) Cl (1·8 g l(-1) ) as carbon and nitrogen sources, promoted an increase of 5·6, 3·7 and 2·6-fold, respectively, in biomass concentration in comparison with glutamate and malate (GM). In turn, the production of α-tocopherol after 120 h identified by LC-MS was 3·7 ± 0·2, 2·4 ± 0·1 and 2 ± 0·1 mg [g dry weight (DW)](-1) , respectively, while in the control medium (GM) it was 0·72 ± 0·1 mg (g DW)(-1) . For paramylon synthesis, the addition of EtOH or glucose induced a higher production. Amino acids were assayed by RP-HPLC; Tyr a tocopherol precursor and Ala an amino acid with antioxidant activity were the amino acids synthesized at higher concentration. Dark-grown E. gracilis Z is a suitable source for the generation of the biotechnologically relevant metabolites tyrosine, α-tocopherol and paramylon. By combining different carbon and nitrogen sources and inducing a tolerable stress to the cell by adding ethanol, it was possible to increase the production of biomass, paramylon, α-tocopherol and some amino acids. The concentrations of α-tocopherol achieved in this study are higher than others reported previously for Euglena, plant and algal systems. This work helps to understand the effect of different carbon sources on the synthesis of bio-molecules by E. gracilis and can be used as a basis for future works to improve the production of different metabolites of biotechnological importance by this organism.

Effects of reactive oxygen species on α-tocopherol production in mitochondria and chloroplasts of Euglena gracilis

The effects of reactive oxygen species (ROS) on α-tocopherol production in mitochondria and chloroplasts of Euglena gracilis were investigated. Addition of an organic carbon source to the medium resulted in increased mitochondrial activity, intracellular O2 - concentration and α-tocopherol productivity in E. gracilis W14ZUL (a chloroplast deficient mutant). α-Tocopherol productivity of the wild-type strain (with both mitochondria and chloroplast) was higher than that of the W14ZUL strain. In the case of the wild strain, the O2 − generated in chloroplasts was efficiently scavenged by the α-tocopherol synthesized inside the chloroplast. In photoheterotrophic culture (with an organic carbon source), there was a positive correlation between α-tocopherol production and O2 − generation. Addition of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (an inhibitor of photosynthesis) resulted in increased O2 − generation and α-tocopherol productivity. These results indicate that the ROS generated in mitochondria and chloroplasts play important roles in α-tocopherol production by E. gracilis. The presence of chloroplasts and generation of intracellular ROS are important for efficient production of α-tocopherol.

Effect of mixed organic substrate on α-tocopherol production by Euglena gracilis in photoheterotrophic culture

Effects of organic carbon sources on cell growth and alpha-tocopherol productivity in wild and chloroplast-deficient W14ZUL strains of Euglena gracilis under photoheterotrophic culture were investigated. In both strains, the increase in cell growth was particularly high when glucose was added as the sole organic carbon source. On the other hand, alpha-tocopherol production per dry cell weight was enhanced by adding ethanol. Ethanol addition also increased the chlorophyll concentration in wild strain and mitochondria activity in W14ZUL strain. For effective alpha-tocopherol production, the effects of mixture of glucose and ethanol were investigated. The results showed that, when a mixture of glucose (6 g/l) and ethanol (4 g/l) was used, alpha-tocopherol productivity per culture broth was 3.89 x 10(-2) mg l(-1) h(-1), which was higher than the value obtained without addition of organic carbon source (0.92 x 10(-2) mg l(-1) h(-1)). In addition, under fed-batch cultivation using an internally illuminated photobioreactor, the alpha-tocopherol production per culture broth was 23.43 mg/l, giving a productivity of 16.27 x 10(-2) mg l(-1) h(-1).

HPLC screening of natural vitamin E from mediterranean plant biofactories—a basic tool for pilot-scale bioreactors production of α-tocopherol

The study was performed in order to investigate a simple, efficient, reliable and rapid method of extracting and quantifying natural vitamin E for pressurized liquid extraction (PLE) as well as high-performance liquid chromatography (HPLC) analysis. Lyophilized Corylus avellana L. nut samples were powdered by high-speed milling with Waring blender for 40 s. alpha-Tocopherol was extracted from the nut tissue powder using dehydrated hexane fortified with 0.01% butylated hydroxytoluene (BHT, co-antioxidant). The rate of alpha-tocopherol accumulation showed differences among nut samples collected in different areas of Italy. Sarda Piccola nut biofactory contained higher amount (81.17 microg/g d.w) of alpha-tocopherol than other-local eleven Italian cultivar nuts. These results provide insight into the biofactory basis for alpha-tocopherol accumulation in hazelnut and give the suitable cultivar tissues to establish pilot-scale bioreactors production of natural bioactive vitamin E.