Supercritical Carbon Dioxide Extraction of Flavonoids from Pomelo (Citrus grandis (L.) Osbeck) Peel and Their Antioxidant Activity

Supercritical carbon dioxide (SC-CO₂) extraction of flavonoids from pomelo (Citrus grandis (L.) Osbeck) peel and their antioxidant activity were investigated. Box-Behnken design combined with response surface methodology was employed to maximize the extraction yield of flavonoids. Correlation analysis of the mathematical-regression model indicated that a quadratic polynomial model could be used to optimize the SC-CO₂ extraction of flavonoids. The optimal conditions for obtaining the highest extraction yield of flavonoids from pomelo peel were a temperature of 80 °C, a pressure of 39 MPa and a static extraction time of 49 min in the presence of 85% ethanol as modifier. Under these conditions, the experimental yield was 2.37%, which matched positively with the value predicted by the model. Furthermore, flavonoids obtained by SC-CO₂ extraction showed a higher scavenging activity on hydroxyl, 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS) radicals than those obtained by conventional solvent extraction (CSE). Therefore, SC-CO₂ extraction can be considered as a suitable technique for the obtainment of flavonoids from pomelo peel.     

遂溪产沙田柚果皮中挥发油成分的GC-MS分析

采用水蒸汽蒸馏法提取广东遂溪产沙田柚果皮挥发油,应用气相色谱-质谱联用技术对其化学成分进行分析和鉴定。共分离出27个组分,鉴定了其中的19种化合物,占总挥发油含量的90.58%。其主要成分D-柠檬烯(37.18%)和β-月桂烯(26.93%),占总挥发油含量的64.11%。     

Adaptive Responses of Citrus grandis Leaves to Copper Toxicity Revealed by RNA-Seq and Physiology

Copper (Cu)-toxic effects on Citrus grandis growth and Cu uptake, as well as gene expression and physiological parameters in leaves were investigated. Using RNA-Seq, 715 upregulated and 573 downregulated genes were identified in leaves of C. grandis seedlings exposed to Cu-toxicity (LCGSEC). Cu-toxicity altered the expression of 52 genes related to cell wall metabolism, thus impairing cell wall metabolism and lowering leaf growth. Cu-toxicity downregulated the expression of photosynthetic electron transport-related genes, thus reducing CO₂ assimilation. Some genes involved in thermal energy dissipation, photorespiration, reactive oxygen species scavenging and cell redox homeostasis and some antioxidants (reduced glutathione, phytochelatins, metallothioneins, l-tryptophan and total phenolics) were upregulated in LCGSEC, but they could not protect LCGSEC from oxidative damage. Several adaptive responses might occur in LCGSEC. LCGSEC displayed both enhanced capacities to maintain homeostasis of Cu via reducing Cu uptake by leaves and preventing release of vacuolar Cu into the cytoplasm, and to improve internal detoxification of Cu by accumulating Cu chelators (lignin, reduced glutathione, phytochelatins, metallothioneins, l-tryptophan and total phenolics). The capacities to maintain both energy homeostasis and Ca homeostasis might be upregulated in LCGSEC. Cu-toxicity increased abscisates (auxins) level, thus stimulating stomatal closure and lowering water loss (enhancing water use efficiency and photosynthesis).     

Characterization of MicroRNAs Associated with Reproduction in the Brown Planthopper,Nilaparvata lugens

Insects have a robust capacity to produce offspring for propagation, and the reproductive events of female insects have been achieved at the molecular and physiological levels via regulatory gene pathways. However, the roles of MicroRNAs (miRNAs) in the reproductive development of the brown planthopper (BPH), Nilaparvata lugens, remain largely unexplored. To understand the roles of miRNAs in reproductive development, miRNAs were identified by Solexa sequencing in short-winged (SW) female adults of BPH. Small RNA libraries derived from three developmental phases (1 day, 3 days, and 5 days after emergence) were constructed and sequenced. We identified 905 miRNAs, including 263 known and 642 novel miRNAs. Among them, a total of 43 miRNAs were differentially expressed in the three developmental phases, and 14,568 putative targets for 43 differentially expressed miRNAs (DEMs) were predicted by TargetScan and miRanda. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the predicted miRNA targets illustrated the putative roles for these DEMs in reproduction. The progress events were annotated, including oogenesis, lipid biosynthetic process, and related pathways such as apoptosis, ABC transporters, and amino acid metabolism. Four highly abundant DEMs (miR-9a-5p, miR-34-5p, miR-275-3p, and miR-317-3p) were further screened, and miR-34-5p was confirmed to be involved in the regulation of reproduction. Overexpression of miR-34-5p via injecting its mimics reduced fecundity and decreased Vg expression. Moreover, target genes prediction for miR-34-5p showed they might be involved in 20E signaling cascades, apoptosis, and gonadal development, including hormone receptor 4 (HR4), caspase-1 (Cp-1), and spermatogenesis-associated protein 20 (SPATA20). These findings provide a valuable resource for future studies on the role of miRNAs in BPH reproductive development.     

Lipoxygenase (LOX) in Sweet and Hot Pepper (Capsicum annuum L.) Fruits during Ripening and under an Enriched Nitric Oxide (NO) Gas Atmosphere

Lipoxygenases (LOXs) catalyze the insertion of molecular oxygen into polyunsaturated fatty acids (PUFA) such as linoleic and linolenic acids, being the first step in the biosynthesis of a large group of biologically active fatty acid (FA)-derived metabolites collectively named oxylipins. LOXs are involved in multiple functions such as the biosynthesis of jasmonic acid (JA) and volatile molecules related to the aroma and flavor production of plant tissues, among others. Using sweet pepper (Capsicum annuum L.) plants as a model, LOX activity was assayed by non-denaturing polyacrylamide gel electrophoresis (PAGE) and specific in-gel activity staining. Thus, we identified a total of seven LOX isozymes (I to VII) distributed among the main plant organs (roots, stems, leaves, and fruits). Furthermore, we studied the FA profile and the LOX isozyme pattern in pepper fruits including a sweet variety (Melchor) and three autochthonous Spanish varieties that have different pungency levels (Piquillo, Padrón, and Alegría riojana). It was observed that the number of LOX isozymes increased as the capsaicin content increased in the fruits. On the other hand, a total of eight CaLOX genes were identified in sweet pepper fruits, and their expression was differentially regulated during ripening and by the treatment with nitric oxide (NO) gas. Finally, a deeper analysis of the LOX IV isoenzyme activity in the presence of nitrosocysteine (CysNO, a NO donor) suggests a regulatory mechanism via S-nitrosation. In summary, our data indicate that the different LOX isozymes are differentially regulated by the capsaicin content, fruit ripening, and NO.     

A Method for Assaying of Protein Kinase Activity In Vivo and Its Use in Studies of Signal Transduction in Strawberry Fruit Ripening

Strawberry (Fragaria × ananassa) fruit ripening is regulated by a complex of cellular signal transduction networks, in which protein kinases are key components. Here, we report a relatively simple method for assaying protein kinase activity in vivo and specifically its application to study the kinase, FaMPK6, signaling in strawberry fruit. Green fluorescent protein (GFP)-tagged FaMPK6 was transiently expressed in strawberry fruit and after stimuli were applied to the fruit it was precipitated using an anti-GFP antibody. The precipitated kinase activity was measured in vitro using ³²P-ATP and myelin basic protein (MBP) as substrates. We also report that FaMPK6 is not involved in the abscisic acid (ABA) signaling cascade, which is closely associated with FaMPK6 signaling in other plant species. However, methyl jasmonate (MeJA), low temperature, and high salt treatments were all found to activate FaMPK6. Transient manipulation of FaMPK6 expression was observed to cause significant changes in the expression patterns of 2749 genes, of which 264 were associated with MeJA signaling. The data also suggest a role for FaMPK6 in modulating cell wall metabolism during fruit ripening. Taken together, the presented method is powerful and its use will contribute to a profound exploration to the signaling mechanism of strawberry fruit ripening.     

Potential use of poly(lactic acid) nanofibers mats as Nano-sachets in postharvest of climacteric fruits and vegetables

This study is reported as the first demonstration of the use of solution blow spun mats in a Nano-sachet system containing a humidity absorber hydrogel and a paraffin wax tablet impregnated with potassium permanganate (KMnO₄) for exogenous ethylene (ex-C₂H₄) oxidation, in order to delay fruit ripening. The tomatoes quality during storage time was characterized by gas chromatography (GC) (ethylene concentration and CO₂ emission analysis—respiration rate), and colorimetry (green/red color parameter). Ex-C₂H₄ production over storage time was reduced for the Nano-sachet system when compared to the control system. Color changes, from 72 to 120 h, displayed fruit still predominantly green in the Nano-Sachet system (a* from −9.1 to −6.9), color transition green-red in the TNT-sachet (−7.5 to −0.3) and increased red color fruit in the control system (−6.1 to 3.7). This work suggests that nanofibrous mats can be used as a packaging system with the ability to protect, potentialize ethylene oxidation and extend climacteric fruits and vegetables quality in shelf life.     

Analysis of Expressed Sequence Tags from Chinese Bayberry Fruit (Myrica rubra Sieb. and Zucc.) at Different Ripening Stages and Their Association with Fruit Quality Development

A total of 2000 EST sequences were produced from cDNA libraries generated from Chinese bayberry fruit (Myrica rubra Sieb. and Zucc. cv. “Biqi”) at four different ripening stages. After cluster and assembly analysis of the datasets by UniProt, 395 unigenes were identified, and their presumed functions were assigned to 14 putative cellular roles. Furthermore, a sequence BLAST was done for the top ten highly expressed genes in the ESTs, and genes associated with disease/defense and anthocyanin accumulation were analyzed. Gene-encoding elements associated with ethylene biosynthesis and signal transductions, in addition to other senescence-regulating proteins, as well as those associated with quality formation during fruit ripening, were also identified. Their possible roles were subsequently discussed.     

Removal of Cu(II) and Zn(II) Using Lignocellulosic Fiber Derived from Citrus reticulata (Kinnow) Waste Biomass

Abstract

The biosorption of Cu(II) and Zn(II) using dried untreated and pretreated Citrus reticulata waste biomass were evaluated. The Cu(II) and Zn(II) sorption were found to be dependent on the solution pH, the biosorbent dose, the biosorbent particle size, the shaking speed, the temperature, the initial metal ions (800 mg/L), and the contact time. Twenty-eight physical and chemical pretreatments of Citrus reticulata waste biomass were evaluated for the sorption of Cu(II) and Zn(II) from aqueous solutions. The results indicated that biomass pretreated with sulphuric acid and EDTA had maximum Cu(II) and Zn(II) uptake capacity of 87.14 mg/g and 86.4 mg/g respectively. Moreover, the Langmuir isotherm model fitted well than the Freundlich model with R ² > 0.95 for both metal ions. The sorption of Cu(II) and Zn(II) occurred rapidly in the first 120 min and the equilibrium was reached in 240 min. FTIR and SEM studies were also carried out to investigate functional groups present in the biomass and the surface morphological changes of biomass.     

柚子皮生物吸附剂化学改性及其对Pb~(2+)吸附性能的研究

以废弃物柚子皮作为原材料,分别制备了未改性的柚子皮吸附剂、碱改性柚子皮吸附剂和疏基乙酸改性柚子皮吸附剂。比较它们对铅离子的吸附效率,并通过比较各种因素对吸附剂吸附铅离子效果的影响,得出最佳的吸附条件:温度为45℃、pH为4的巯基乙酸改性柚子皮吸附能力最佳。     

Triacylglycerol and Glycerophospholipid Identification and Accumulation During Ripening of Pistacia lentiscus L. (Lentisc) Fruit

Triacylglycerol (TAG) and phospholipid (PL) compositions of vegetable oils are considered a marker of quality and are often used in industry to control the purity of the oils and to detect adulteration. In this study, the TAG and PL composition of developing fruit of Pistacia lentiscus were investigated for the first time. The total TAG content was found to increase rapidly during fruit ripening from 105 to 966 mg/100 g of oil respectively between the 35th and the 175th day after fructification (DAF). During this period, 16 different molecular species of TAG were identified and quantified. POO was the major TAG from the second stage of maturation. Only four classes of PL were identified in the P. lentiscus oil: the phosphatidic acid (PA), the phosphatidylethanolamine (PE), the phosphatidylglycerol (PG) and the phosphatidylinositol (PI). The mass spectra obtained showed the presence of nine molecular species of PA, five species of PE and seven molecular species for each PG and PI classes. The total phospholipid content decreased rapidly during fruit ripening, from 45.5 % at the 15th DAF to 6.88 % at the 175th DAF.     

FAAH Inhibition Restores Early Life Stress-Induced Alterations in PFC microRNAs Associated with Depressive-Like Behavior in Male and Female Rats

Early life stress (ELS) increases predisposition to depression. We compared the effects of treatment with the fatty acid amide hydrolase (FAAH) inhibitor URB597, and the selective serotonin reuptake inhibitor paroxetine, on ELS-induced depressive-like behavior and the expression of microRNAs (miRs) associated with depression in the medial prefrontal cortex (mPFC), hippocampal CA1 area, lateral habenula and dorsal raphe in rats. We also examined the mRNA expression of serotonergic (htr1a and slc6a4) and endocannabinoid (cnr1, cnr2 and faah) targets in the mPFC following ELS and pharmacological treatment. Adult males and females exposed to the ‘Limited Bedding and Nesting’ ELS paradigm demonstrated a depressive-like phenotype and late-adolescence URB597 treatment, but not paroxetine, reversed this phenotype. In the mPFC, ELS downregulated miR-16 in males and miR-135a in females and URB597 treatment restored this effect. In ELS females, the increase in cnr2 and decrease in faah mRNAs in the mPFC were reversed by URB597 treatment. We show for the first time that URB597 reversed ELS-induced mPFC downregulation in specific miRs and stress-related behaviors, suggesting a novel mechanism for the beneficial effects of FAAH inhibition. The differential effects of ELS and URB597 on males and females highlight the importance of developing sex-specific treatment approaches.     

Dysregulation of Muscle-Specific MicroRNAs as Common Pathogenic Feature Associated with Muscle Atrophy in ALS,SMA and SBMA: Evidence from Animal Models and Human Patients

Motor neuron diseases (MNDs) are neurodegenerative disorders characterized by upper and/or lower MN loss. MNDs include amyotrophic lateral sclerosis (ALS), spinal muscular atrophy (SMA), and spinal and bulbar muscular atrophy (SBMA). Despite variability in onset, progression, and genetics, they share a common skeletal muscle involvement, suggesting that it could be a primary site for MND pathogenesis. Due to the key role of muscle-specific microRNAs (myomiRs) in skeletal muscle development, by real-time PCR we investigated the expression of miR-206, miR-133a, miR-133b, and miR-1, and their target genes, in G93A-SOD1 ALS, Δ7SMA, and KI-SBMA mouse muscle during disease progression. Further, we analyzed their expression in serum of SOD1-mutated ALS, SMA, and SBMA patients, to demonstrate myomiR role as noninvasive biomarkers. Our data showed a dysregulation of myomiRs and their targets, in ALS, SMA, and SBMA mice, revealing a common pathogenic feature associated with muscle impairment. A similar myomiR signature was observed in patients’ sera. In particular, an up-regulation of miR-206 was identified in both mouse muscle and serum of human patients. Our overall findings highlight the role of myomiRs as promising biomarkers in ALS, SMA, and SBMA. Further investigations are needed to explore the potential of myomiRs as therapeutic targets for MND treatment.     

Furanocoumarin content and phototoxicity of rough lemon (Citrus jambhiri) foliage exposed to enhanced ultraviolet-B (UVB) irradiation

Rooted cuttings ofC. jambhiri were grown under enhanced levels of UVB radiation for 95 days. Bacterial phototoxicity and furanocoumarin content were determined in extracts made from various tissues from the aboveground biomass. Young, newly expanded leaves contained significantly higher concentrations of furanocoumarins than older leaves and stems. Additionally, the proportional concentration of psoralen was higher in young leaves than in old leaves. While treatment with UVB did not result in a change in the overall level of furanocoumarins, it did cause an increase in the ratio of psoralen to bergapten. Bacterial phototoxicity paralleled the distribution of furanocoumarin content among tissue types; analysis of covariance suggested that the phototoxic properties of the extracts could be accounted for on the basis of furanocoumarin content alone.     

Changes in the Content of Organic Acids and Expression Analysis of Citric Acid Accumulation-Related Genes during Fruit Development of Yellow (Passiflora edulis f. flavicarpa) and Purple (Passiflora edulis f. edulis) Passion Fruits

Organic acids are key components that determine the taste and flavor of fruits and play a vital role in maintaining fruit quality and nutritive value. In this study, the fruits of two cultivars of passion fruit Yellow (Passiflora edulis f. flavicarpa) and purple (Passiflora edulis f. edulis) were harvested at five different developmental stages (i.e., fruitlet, green, veraison, near-mature and mature stage) from an orchard located in subtropical region of Fujian Province, China. The contents of six organic acids were quantified using ultra-performance liquid chromatography (UPLC), activities of citric acid related enzymes were determined, and expression levels of genes involved in citric acid metabolism were measured by quantitative real-time PCR (qRT-PCR). The results revealed that citric acid was the predominant organic acid in both cultivars during fruit development. The highest citric acid contents were observed in both cultivars at green stage, which were reduced with fruit maturity. Correlation analysis showed that citrate synthase (CS), cytosolic aconitase (Cyt-ACO) and cytosolic isocitrate dehydrogenase (Cyt-IDH) may be involved in regulating citric acid biosynthesis. Meanwhile, the PeCS2, PeACO4, PeACO5 and PeIDH1 genes may play an important role in regulating the accumulation of citric acid. This study provides new insights for future elucidation of key mechanisms regulating organic acid biosynthesis in passion fruit.