Genome Wide MeDIP-Seq Profiling of Wild and Cultivated Olives Trees Suggests DNA Methylation Fingerprint on the Sensory Quality of Olive Oil.

Secondary metabolites are particularly important to humans due to their pharmaceutical properties. Moreover, secondary metabolites are key compounds in climate change adaptation in long-living trees. Recently, it has been described that the domestication of Olea subspecies had no major selection signature on coding variants and was mainly related to changes in gene expression. In addition, the phenotypic plasticity in Olea subspecies was linked to the activation of transposable elements in the genes neighboring. Here, we investigated the imprint of DNA methylation in the unassigned fraction of the phenotypic plasticity of the Olea subspecies, using methylated DNA immuno-precipitation sequencing (MeDIP-seq) for a high-resolution genome-wide DNA methylation profiling of leaves and fruits during fruit development in wild and cultivated olives from Turkey. Notably, the methylation profiling showed a differential DNA methylation in secondary metabolism responsible for the sensory quality of olive oil. Here, we highlight for the first time the imprint of DNA methylation in modulating the activity of the Linoleate 9S lipoxygenase in the biosynthesis of volatile aromatic compounds. Unprecedently, the current study reveals the methylation status of the olive genome during fruit ripening.

EMS-Induced Mutagenesis of Clostridium carboxidivorans for Increased Atmospheric CO2 Reduction Efficiency and Solvent Production.

Clostridium carboxidivorans (P7) is one of the most important solvent-producing bacteria capable of fermenting syngas (CO, CO2, and H2) to produce chemical commodities when grown as an autotroph. This study aimed to develop ethyl methanesulfonate (EMS)-induced P7 mutants that were capable of growing in the presence of CO2 as a unique source of carbon with increased solvent formation and atmospheric CO2 reduction to limit global warming. Phenotypic analysis including growth and end product characterization of the P7 wild type (WT) demonstrated that this strain grew better at 25 °C than 37 °C when CO2 served as the only source of carbon. In the current study, 55 mutagenized P7-EMS mutants were developed by using 100 mM and 120 mM EMS. Interestingly, using a forward genetic approach, three out of the 55 P7-EMS mutants showed a significant increase in ethanol, butyrate, and butanol production. The three P7-EMS mutants presented on average a 4.68-fold increase in concentrations of ethanol when compared to the P7-WT. Butyric acid production from 3 P7-EMS mutants contained an average of a 3.85 fold increase over the levels observed in the P7-WT cultures under the same conditions (CO2 only). In addition, one P7-EMS mutant presented butanol production (0.23 ± 0.02 g/L), which was absent from the P7-WT under CO2 conditions. Most of the P7-EMS mutants showed stability of the obtained end product traits after three transfers. Most importantly, the amount of reduced atmospheric CO2 increased up to 8.72 times (0.21 g/Abs) for ethanol production and up to 8.73 times higher (0.16 g/Abs) for butyrate than the levels contained in the P7-WT. Additionally, to produce butanol, the P7-EMSIII-J mutant presented 0.082 g/Abs of CO2 reduction. This study demonstrated the feasibility and effectiveness of employing EMS mutagenesis in generating solvent-producing anaerobic bacteria mutants with improved and novel product formation and increased atmospheric CO2 reduction efficiency.

Effective identification of soybean candidate genes involved in resistance to soybean cyst nematode via direct whole genome re-sequencing of two segregating mutants.

KEY MESSAGE: Three soybean candidate genes involved in resistance to soybean cyst nematode race 4 were identified via direct whole genome re-sequencing of two segregating mutants. The genes conferring resistance to soybean cyst nematode (SCN) race 4 (Hg type 1.2.3.5.7) in soybean (Glycine max L. Merr.) remains unknown. Next generation sequencing-based methods identify a wide range of targets, it is difficult to identify genes underlying traits. Use of the MutMap and QTL-seq methods to identify trait candidate genes needs backcrossing and is very time-consuming. Here we report a simple method to effectively identify candidate genes involved in resistance to SCN race 4. Two ethane methylsulfonate mutagenized mutants of soybean 'PI 437654', whose SCN race 4-infection phenotype altered, were selected. Six relevant whole genomes were re-sequenced, and then calling of genomic variants (SNPs and InDels) was conducted and compared to 'Williams 82'. The comparison eliminated many genomic variants from the mutant lines that overlapped two non-phenotypic but mutant progeny plants, wild-type PI 437654 and 'Zhonghuang 13'. Finally, only 27 mutations were found among 10 genes. Of these 10 genes, 3 genes, Glyma.09g054000, Glyma.16g065700 and Glyma.18g192200 were overlapped between two phenotypic mutant progeny plants. Therefore, the three genes may be the candidate genes involved in resistance of PI 437654 to soybean cyst nematode race 4. This method simplifies the effective identification of candidate genes.

Genome reorganization of the GmSHMT gene family in soybean showed a lack of functional redundancy in resistance to soybean cyst nematode.

In soybeans, eighteen members constitute the serine hydroxymethyltransferase (GmSHMT) gene family, of which the cytosolic-targeted GmSHMT08c member has been reported to mediate resistance to soybean cyst nematode (SCN). This work presents a comprehensive study of the SHMT gene family members, including synteny, phylogeny, subcellular localizations, haplotypes, protein homology modeling, mutational, and expression analyses. Phylogenetic analysis showed that SHMT genes are divided into four classes reflecting their subcellular distribution (cytosol, nucleus, mitochondrion, and chloroplast). Subcellular localization of selected GmSHMT members supports their in-silico predictions and phylogenetic distribution. Expression and functional analyses showed that GmSHMT genes display many overlapping, but some divergent responses during SCN infection. Furthermore, mutational analysis reveals that all isolated EMS mutants that lose their resistance to SCN carry missense and nonsense mutations at the GmSHMT08c, but none of the Gmshmt08c mutants carried mutations in the other GmSHMT genes. Haplotype clustering analysis using the whole genome resequencing data from a collection of 106 diverse soybean germplams (15X) was performed to identify allelic variants and haplotypes within the GmSHMT gene family. Interestingly, only the cytosolic-localized GmSHMT08c presented SNP clusters that were associated with SCN resistance, supporting our mutational analysis. Although eight GmSHMT members respond to the nematode infestation, functional and mutational analysis has shown the absence of functional redundancy in resistance to SCN. Structural analysis and protein homology modeling showed the presence of spontaneous mutations at important residues within the GmSHMT proteins, suggesting the presence of altered enzyme activities based on substrate affinities. Due to the accumulation of mutations during the evolution of the soybean genome, the other GmSHMT members have undergone neofunctionalization and subfunctionalization events.

Classification and experimental identification of plant long non-coding RNAs.

Our understanding of gene regulation is constantly evolving. It is now clear that the majority of cellular transcripts are non-coding RNAs. The spectrum of non-coding RNAs is diverse and includes short (<200 nt) and long non-coding RNAs (lncRNAs) (>200 nt). LncRNAs regulate gene expression through diverse mechanisms. In this review, we describe the emerging roles of lncRNA mediated plant gene regulation. We discuss the current classification of lncRNAs and their role in genome organization and gene regulation. We also highlight a subset of lncRNAs that are epigenetic regulators of plant gene expression. Lastly, we provide an overview of emerging techniques and databases that are employed for the identification and characterization of plant lncRNAs.

Combination of gold nanoparticles with low-LET irradiation: an approach to enhance DNA DSB induction in HT29 colorectal cancer stem-like cells.

PURPOSE: High-linear energy transfer (high LET) irradiation has significant cytotoxic effects on different cancerous stem-like cells (CSLCs) such as colorectal CSLCs. A review of the literature has indicated that the presence of gold nanoparticles (GNPs) enables low-LET irradiation to produce highly non-homogeneous dose distributions like high-LET irradiation. The purpose of this study was to evaluate the radioresponsiveness of HT29 colorectal CSLCs under low-LET irradiation (X-ray) and in the presence of GNPs. METHODS: Radioresponsiveness was evaluated using the ϒ-H2AX foci formation assay, the clonogenic assay, the cell cycle progression assay and analyses of radiobiological parameters. RESULTS: In the presence of GNPs, the survival fraction of HT29 CSLCs was significantly reduced and caused significant changes in the radiobiological parameters after irradiation. In addition, ϒ-H2AX assay showed that in the presence of GNPs, the persistent DNA double-strand breaks were significantly increased in irradiated HT29 CSLCs. The relative biological effectiveness value of GNPs with X-rays was about 1.6 for HT-29 CSLCs at the 10% of cell survival fraction (D10 level) when compared to X-rays alone. CONCLUSION: Therefore, the combination of GNPs with X-ray irradiation has the potential to kill HT29 CSLCs greater than the X-ray alone, and may be considered as an alternative for high-LET irradiation.

Impact of wastewater effluent containing aged nanoparticles and other components on biological activities of the soil microbiome, Arabidopsis plants, and earthworms.

The amount of engineered nanomaterials (ENMs) in the environment has been increasing due to their industrial and commercial applications. Different types of metallic nanoparticles (NPs) have been detected in effluents from wastewater treatment plants (WWTPs). The effluents have been reclaimed for crop irrigation in many arid and semi-arid areas. Here, a soil micro-ecosystem was established including a microbiome, 4 Arabidopsis thaliana plants, and 3 Eisenia fetida earthworms, for a duration of 95 days. The impact of wastewater effluent (WE) containing aged NPs was studied. WE was taken from a local WWTP and exhibited the presence of Ti, Ag, and Zn up to 97.0 ± 9.4, 27.4 ± 3.9, and 4.1 ± 3.6 µg/L, respectively, as well as the presence of nanoscale particles (1-100 nm in diameter). The plants were irrigated with WE or deionized water (DIW). After 95 days, significantly higher concentrations of extractable Ti and Zn (439.2 ± 24.4 and 9.0 ± 0.5 mg/kg, respectively) were found in WE-irrigated soil than those in DIW-irrigated soil (161.2 ± 2.1 and 4.0 ± 0.1 mg/kg). The extractable Ag concentrations did not differ significantly between the WE- and DIW-irrigated soil. Although microbial biomass carbon and nitrogen were not significantly reduced, the population distribution of the microbial communities was shifted in WE-irrigated soil compared to the control. The abundance of cyanobacteria (Cyanophyta) was increased by 12.5% in the WE-irrigated soil as manifested mainly by an increase of Trichodesmium spp., and the abundance of unknown archaea was enhanced from 26.7% in the control to 40.5% in the WE-irrigated soil. The biomasses of A. thaliana and E. fetida were not significantly changed by WE exposure. However, A. thaliana had a noticeable shortened life cycle, and corrected total cell fluorescence was much higher in the roots of WE-irrigated plants compared to the control. These impacts on the soil micro-ecosystem may have resulted from the aged NPs and/or the metal ions released from these NPs, as well as other components in the WE. Taken together, these results should help inform the reuse of WE containing aged NPs and other components in sustainable agriculture.

Integration of sudden death syndrome resistance loci in the soybean genome.

KEY MESSAGE: Complexity and inconsistencies in resistance mapping publications of soybean sudden death syndrome (SDS) result in interpretation difficulty. This review integrates SDS mapping literature and proposes a new nomenclature system for reproducible SDS resistance loci. Soybean resistance to sudden death syndrome (SDS) is composed of foliar resistance to phytotoxins and root resistance to pathogen invasion. There are more than 80 quantitative trait loci (QTL) and dozens of single nucleotide polymorphisms (SNPs) associated with soybean resistance to SDS. The validity of these QTL and SNPs is questionable because of the complexity in phenotyping methodologies, the disease synergism between SDS and soybean cyst nematode (SCN), the variability from the interactions between soybean genotypes and environments, and the inconsistencies in the QTL nomenclature. This review organizes SDS mapping results and proposes the Rfv (resistance to Fusarium virguliforme) nomenclature based on supporting criteria described in the text. Among ten reproducible loci receiving our Rfv nomenclature, Rfv18-01 is mostly supported by field studies and it co-localizes to the SCN resistance locus rhg1. The possibility that Rfv18-01 is a pleiotropic resistance locus and the concern about Rfv18-01 being confounded with Rhg1 is discussed. On the other hand, Rfv06-01, Rfv06-02, Rfv09-01, Rfv13-01, and Rfv16-01 were identified both by screening soybean leaves against phytotoxic culture filtrates and by evaluating SDS severity in fields. Future phenotyping using leaf- and root-specific resistance screening methodologies may improve the precision of SDS resistance, and advanced genetic studies may further clarify the interactions among soybean genotypes, F. virguliforme, SCN, and environments. The review provides a summary of the SDS resistance literature and proposes a framework for communicating SDS resistance loci for future research considering molecular interactions and genetic breeding for soybean SDS resistance.

First draft genome assembly of the Argane tree ( Argania spinosa).

Background: The Argane tree ( Argania spinosa L. Skeels) is an endemic tree of southwestern Morocco that plays an important socioeconomic and ecologic role for a dense human population in an arid zone. Several studies confirmed the importance of this species as a food and feed source and as a resource for both pharmaceutical and cosmetic compounds. Unfortunately, the argane tree ecosystem is facing significant threats from environmental changes (global warming, over-population) and over-exploitation. Limited research has been conducted, however, on argane tree genetics and genomics, which hinders its conservation and genetic improvement. Methods: Here, we present a draft genome assembly of A. spinosa. A reliable reference genome of  A. spinosa was created using a hybrid  de novo assembly approach combining short and long sequencing reads. Results: In total, 144 Gb Illumina HiSeq reads and 7.2 Gb PacBio reads were produced and assembled. The final draft genome comprises 75 327 scaffolds totaling 671 Mb with an N50 of 49 916 kb. The draft assembly is close to the genome size estimated by k-mers distribution and covers 89% of complete and 4.3 % of partial Arabidopsis orthologous groups in BUSCO. Conclusion: The A. spinosa genome will be useful for assessing biodiversity leading to efficient conservation of this endangered endemic tree. Furthermore, the genome may enable genome-assisted cultivar breeding, and provide a better understanding of important metabolic pathways and their underlying genes for both cosmetic and pharmacological purposes.

Genome of wild olive and the evolution of oil biosynthesis.

Here we present the genome sequence and annotation of the wild olive tree (Olea europaea var. sylvestris), called oleaster, which is considered an ancestor of cultivated olive trees. More than 50,000 protein-coding genes were predicted, a majority of which could be anchored to 23 pseudochromosomes obtained through a newly constructed genetic map. The oleaster genome contains signatures of two Oleaceae lineage-specific paleopolyploidy events, dated at ∼28 and ∼59 Mya. These events contributed to the expansion and neofunctionalization of genes and gene families that play important roles in oil biosynthesis. The functional divergence of oil biosynthesis pathway genes, such as FAD2, SACPD, EAR, and ACPTE, following duplication, has been responsible for the differential accumulation of oleic and linoleic acids produced in olive compared with sesame, a closely related oil crop. Duplicated oleaster FAD2 genes are regulated by an siRNA derived from a transposable element-rich region, leading to suppressed levels of FAD2 gene expression. Additionally, neofunctionalization of members of the SACPD gene family has led to increased expression of SACPD2, 3, 5, and 7, consequently resulting in an increased desaturation of steric acid. Taken together, decreased FAD2 expression and increased SACPD expression likely explain the accumulation of exceptionally high levels of oleic acid in olive. The oleaster genome thus provides important insights into the evolution of oil biosynthesis and will be a valuable resource for oil crop genomics.

Phytochemicals: Extraction, Isolation, and Identification of Bioactive Compounds from Plant Extracts.

There are concerns about using synthetic phenolic antioxidants such as butylated hydroxytoluene (BHT) and butylated hydroxyanisole (BHA) as food additives because of the reported negative effects on human health. Thus, a replacement of these synthetics by antioxidant extractions from various foods has been proposed. More than 8000 different phenolic compounds have been characterized; fruits and vegetables are the prime sources of natural antioxidants. In order to extract, measure, and identify bioactive compounds from a wide variety of fruits and vegetables, researchers use multiple techniques and methods. This review includes a brief description of a wide range of different assays. The antioxidant, antimicrobial, and anticancer properties of phenolic natural products from fruits and vegetables are also discussed.

Evaluation of the antimicrobial activities of ultrasonicated spinach leaf extracts using RAPD markers and electron microscopy.

Spinach (Spinacia oleracea L.) leaves represent an important dietary source of nutrients, antioxidants and antimicrobials. As such, spinach leaves play an important role in health and have been used in the treatment of human diseases since ancient times. Here, the aims were to optimize the extraction methods for recovering antimicrobial substances of spinach leaves, determine the minimum inhibitory concentrations (MICs) of the antimicrobial substances against Escherichia coli and Staphylococcus aureus and, finally, evaluate the effects of spinach leaves' antimicrobials on bacterial DNA using central composite face-centered methods. The effect of the extracts on both Gram-positive and Gram-negative bacterial models was examined by scanning electron microscopy (SEM) and random amplification of polymorphic (bacterial) DNA (RAPD). The optimal extraction conditions were at 45 °C, ultrasound power of 44% and an extraction time of 23 min. The spinach extracts exhibited antimicrobial activities against both bacteria with MICs in the 60-100 mg/ml range. Interestingly, SEM showed that the treated bacterial cells appear damaged with a reduction in cell number. RAPD analysis of genomic DNA showed that the number and sizes of amplicons were decreased by treatments. Based on these results, it was inferred that spinach leaf extracts exert bactericidal activities by both inducing mutations in DNA and causing cell wall disruptions.

Proteomic Profiling and the Predicted Interactome of Host Proteins in Compatible and Incompatible Interactions Between Soybean and Fusarium virguliforme.

Sudden death syndrome (SDS) is a complex of two diseases of soybean (Glycine max), caused by the soil borne pathogenic fungus Fusarium virguliforme. The root rot and leaf scorch diseases both result in significant yield losses worldwide. Partial SDS resistance has been demonstrated in multiple soybean cultivars. This study aimed to highlight proteomic changes in soybean roots by identifying proteins which are differentially expressed in near isogenic lines (NILs) contrasting at the Rhg1/Rfs2 locus for partial resistance or susceptibility to SDS. Two-dimensional gel electrophoresis resolved approximately 1000 spots on each gel; 12 spots with a significant (P < 0.05) difference in abundance of 1.5-fold or more were picked, trypsin-digested, and analyzed using quadruple time-of-flight tandem mass spectrometry. Several spots contained more than one protein, so that 18 distinct proteins were identified overall. A functional analysis performed to categorize the proteins depicted that the major pathways altered by fungal infection include disease resistance, stress tolerance, and metabolism. This is the first report which identifies proteins whose abundances are altered in response to fungal infection leading to SDS. The results provide valuable information about SDS resistance in soybean plants, and plant partial resistance responses in general. More importantly, several of the identified proteins could be good candidates for the development of SDS-resistant soybean plants.

Ultrasound Assisted Extraction of Phenolic Compounds from Peaches and Pumpkins.

The ultrasound-assisted extraction (UAE) method was used to optimize the extraction of phenolic compounds from pumpkins and peaches. The response surface methodology (RSM) was used to study the effects of three independent variables each with three treatments. They included extraction temperatures (30, 40 and 50°C), ultrasonic power levels (30, 50 and 70%) and extraction times (10, 20 and 30 min). The optimal conditions for extractions of total phenolics from pumpkins were inferred to be a temperature of 41.45°C, a power of 44.60% and a time of 25.67 min. However, an extraction temperature of 40.99°C, power of 56.01% and time of 25.71 min was optimal for recovery of free radical scavenging activity (measured by 1, 1-diphenyl-2-picrylhydrazyl (DPPH) reduction). The optimal conditions for peach extracts were an extraction temperature of 41.53°C, power of 43.99% and time of 27.86 min for total phenolics. However, an extraction temperature of 41.60°C, power of 44.88% and time of 27.49 min was optimal for free radical scavenging activity (judged by from DPPH reduction). Further, the UAE processes were significantly better than solvent extractions without ultrasound. By electron microscopy it was concluded that ultrasonic processing caused damage in cells for all treated samples (pumpkin, peach). However, the FTIR spectra did not show any significant changes in chemical structures caused by either ultrasonic processing or solvent extraction.

Effects of plant-derived anti-leukemic drugs on individualized leukemic cell population profiles in Egyptian patients.

Leukemias are a group of cancer types that originate from blood-forming tissues. In this disease, an abnormally large number of immature white blood cells is produced by the bone marrow. The relationship between treatments with plant-derived drugs and leukemia-associated immunophenotypes (LAIPs) of clinically isolated leukemia cells has yet to be established. The aim of the present study was to develop a preliminary clinical prognostic map for commonly expressed LAIPs in patients clinically diagnosed with leukemia, as well as to assess the potential involvement of LAIPs in the response rate to 10 natural products of plant origin. An increased expression of LAIPs, including CD4, CD14, CD33 and CD34, was considered a surrogate marker of the desired response of leukemia cells to treatment with plant-derived drugs. By contrast, the increased expression of the LAIPs, MPO and DR, was associated with poor prognostic outcomes following treatment with the plant-derived drugs. The results showed that 5 of the 10 plant-derived drugs tested induced the expression of several desirable LAIPs biomarkers. These findings clearly highlight the potential treatment efficacy of certain plant-derived drugs against leukemic cell types.

Domestication footprints anchor genomic regions of agronomic importance in soybeans.

Present-day soybeans consist of elite cultivars and landraces (Glycine max, fully domesticated (FD)), annual wild type (Glycine soja, nondomesticated (ND)), and semi-wild type (semi-domesticated (SD)). FD soybean originated in China, although the details of its domestication history remain obscure. More than 500 diverse soybean accessions were sequenced using specific-locus amplified fragment sequencing (SLAF-seq) to address fundamental questions regarding soybean domestication. In total, 64,141 single nucleotide polymorphisms (SNPs) with minor allele frequencies (MAFs) > 0.05 were found among the 512 tested accessions. The results indicated that the SD group is not a hybrid between the FD and ND groups. The initial domestication region was pinpointed to central China (demarcated by the Great Wall to the north and the Qinling Mountains to the south). A total of 800 highly differentiated genetic regions and > 140 selective sweeps were identified, and these were three- and twofold more likely, respectively, to encompass a known quantitative trait locus (QTL) than the rest of the soybean genome. Forty-three potential quantitative trait nucleotides (QTNs; including 15 distinct traits) were identified by genome-wide association mapping. The results of the present study should be beneficial for soybean improvement and provide insight into the genetic architecture of traits of agronomic importance.

Simultaneous extraction, optimization, and analysis of flavonoids and polyphenols from peach and pumpkin extracts using a TLC-densitometric method.

BACKGROUND: The use of medicinal plants has been reported throughout human history. In the fight against illnesses, medicinal plants represent the primary health care system for 60 % of the world's population. Flavonoids are polyphenolic compounds with active anti-microbial properties; they are produced in plants as pigments. Quercetin, myricetin, and rutin are among the most well-known and prevalent flavonoids in plants, with an antioxidant activity capable of decreasing the oxidation of low density lipoproteins [LDLs]. To date, this research is the first of its kind to employ a coupled thin-layer chromatography (TLC) and a densitometric quantification method with a Box-Behnken design (BBD) response surface methodology (RSM) for optimization of ultrasonic-assisted extraction and determination of rutin and quercetin from peach and ellagic acid and myricetin from pumpkin fruits. RESULTS: The effect of process variables (extraction temperature (°C), extraction power (%) and extraction time (min)) on ultrasound-assisted extraction (UAE) were examined by using BBD and RSM. TLC followed by Quantity-One™ (BioRad) image analysis as a simple and rapid method was used for identification and quantification of the compounds in complex mixtures. The results were consistent under optimal conditions among the experimental values and their predicted values. A mass spectrometry (MALDI-TOF MS) technique was also used to confirm the identity of the natural products in the TLC spots resolved. CONCLUSION: The results show that the coupled TLC-densitometric methods & BBD can be a very powerful approach to qualitative and quantitative analysis of; rutin and quercetin from peach extracts; and ellagic acid and myricetin contents from pumpkin extracts.

Employing response surface methodology for the optimization of ultrasound assisted extraction of lutein and ß-carotene from spinach.

The extraction of lutein and ß-carotene from spinach (Spinacia oleracea L.) leaves is important to the dietary supplement industry. A Box-Behnken design and response surface methodology (RSM) were used to investigate the effect of process variables on the ultrasound-assisted extraction (UAE) of lutein and ß-carotene from spinach. Three independent variables, extraction temperature (°C), extraction power (%) and extraction time (min) were studied. Thin-layer chromatography (TLC) followed by UV visualization and densitometry was used as a simple and rapid method for both identification and quantification of lutein and ß-carotene during UAE. Methanol extracts of leaves from spinach and authentic standards of lutein and ß-carotene were separated by normal-phase TLC with ethyl acetate-acetone (5:4 (v/v)) as the mobile phase. In this study, the combination of TLC, densitometry, and Box-Behnken with RSM methods were effective for the quantitative analysis of lutein and ß-carotene from spinach extracts. The resulting quadratic polynomial models for optimizing lutein and ß-carotene from spinach had high coefficients of determination of 0.96 and 0.94, respectively. The optimal UAE settings for output of lutein and ß-carotene simultaneously from spinach extracts were an extraction temperature of 40 °C, extraction power of 40% (28 W/cm3) and extraction time of 16 min. The identity and purity of each TLC spot was measured using time-of-flight mass spectrometry. Therefore, UAE assisted extraction of carotenes from spinach can provide a source of lutein and ß-carotene for the dietary supplement industry.

Effects of ultrasonic treatments on the polyphenol and antioxidant content of spinach extracts.

The objective was to test ultrasound treatments on spinach leaves during extraction, and conventional extraction was used as a control. The effects of different combinations of the ultrasonic water bath factors tested on phenolic compound yields included frequency (37 and 80 kHz), exposure time (5, 10, 15, 20, 25, and 30 min), temperature (30, 40, and 50°C), and ultrasonic power (30%, 50%, and 70%). The best conditions for extraction yields were ultrasonic frequency of 37 kHz, extraction time of 30 min, reaction temperature of 40°C, and ultrasonic power of 50%. The mean yield (mg/100g), total phenol (mg gallic acid/g DW), flavonoids (mg/g DW), % DPPH free-radical scavenging activity, and % ferric reducing antioxidant power were all high (64.88 ± 21.84, 33.96 ± 11.30, 27.37 ± 11.85, 64.18 ± 16.69 and 70.25 ± 9.68). Treatments were significantly different. The interaction among the ultrasonic parameters was significant. Temperature and power had significant effects on all other dependent variables.