Maximizing Polysaccharides and Phycoerythrin in Porphyridium purpureum via the Addition of Exogenous Compounds: A Response-Surface-Methodology Approach.

Phycoerythrin and polysaccharides have significant commercial value in medicine, cosmetics, and food industries due to their excellent bioactive functions. To maximize the production of biomass, phycoerythrin, and polysaccharides in Porphyridium purpureum, culture media were supplemented with calcium gluconate (CG), magnesium gluconate (MG) and polypeptides (BT), and their optimal amounts were determined using the response surface methodology (RSM) based on three single-factor experiments. The optimal concentrations of CG, MG, and BT were determined to be 4, 12, and 2 g L-1, respectively. The RSM-based models indicated that biomass and phycoerythrin production were significantly affected only by MG and BT, respectively. However, polysaccharide production was significantly affected by the interactions between CG and BT and those between MG and BT, with no significant effect from BT alone. Using the optimized culture conditions, the maximum biomass (5.97 g L-1), phycoerythrin (102.95 mg L-1), and polysaccharide (1.42 g L-1) concentrations met and even surpassed the model-predicted maximums. After optimization, biomass, phycoerythrin, and polysaccharides concentrations increased by 132.3%, 27.97%, and 136.67%, respectively, compared to the control. Overall, this study establishes a strong foundation for the highly efficient production of phycoerythrin and polysaccharides using P. purpureum.

Intensifying the simultaneous removal of nitrogen and phosphorus of an integrated aerobic granular sludge-membrane bioreactor by Acinetobacter junii.

This work aims at intensifying the simultaneous removal of nitrogen and phosphorus of an integrated aerobic granular sludge (AGS) - membrane bioreactor (MBR) by Acinetobacter junii. After acclimation and enrichment in a sequencing batch reactor (SBR), Acinetobacter junii, a kind of denitrifying phosphate accumulating organism (DPAO), was successfully screened in the used SBR. Then it was verified to be capable of effectively enhancing the performance in the simultaneous removal of nitrogen and phosphorus of AGS-MBR. In the system, DPAO (Acinetobacter junii) mainly occurred in AGS, and the highest ratio even reached 22.8%, but its competitive advantages highly depend on the size of AGS. The presented results can cultivate AGS and enrich DPAO simultaneously to improve the removal of nitrogen and phosphorus of an AGS-MBR, which provide an environmentally friendly approach to upgrade traditional wastewater treatment processes.

Revealing of Intracellular Antioxidants in Dendrobium nobile by High Performance Liquid Chromatography-Tandem Mass Spectrometry.

The medicinal plant Dendrobium nobile is an important natural antioxidant resource. To reveal the antioxidants of D. nobile, high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was employed for metabolic analysis. The H2O2-induced oxidative damage was used in human embryonic kidney 293T (H293T) cells to assess intracellular antioxidant activities. Cells incubated with flower and fruit extracts showed better cell survival, lower levels of reactive oxygen species (ROS), and higher catalase and superoxide dismutase activities than those incubated with root, stem, and leaf extracts (p < 0.01). A total of 13 compounds were newly identified as intracellular antioxidants by association analysis, including coniferin, galactinol, trehalose, beta-D-lactose, trigonelline, nicotinamide-N-oxide, shikimic acid, 5'-deoxy-5'-(methylthio)adenosine, salicylic acid, isorhamnetin-3-O-neohespeidoside, methylhesperidin, 4-hydroxybenzoic acid, and cis-aconitic acid (R2 > 0.8, Log2FC > 1, distribution > 0.1%, and p < 0.01). They showed lower molecular weight and higher polarity, compared to previously identified in vitro antioxidants in D. nobile (p < 0.01). The credibility of HPLC-MS/MS relative quantification was verified by common methods. In conclusion, some saccharides and phenols with low molecular weight and high polarity helped protect H293T cells from oxidative damage by increasing the activities of intracellular antioxidant enzymes and reducing intracellular ROS levels. The results enriched the database of safe and effective intracellular antioxidants in medicinal plants.

The isolation, identification, whole-genome sequencing of Clostridium butyricum LV1 and its effects on growth performance, immune response, and disease-resistance of Litopenaeus vannamei.

In this study, a strain of Clostridium butyricum was isolated from the intestine of Litopenaeus vannamei with the method of anaerobic microbial isolation and culture. Next, the probiotic properties of LV1 were evaluated with susceptibility tests, tolerance tests, and whole genome sequencing in vivo and in vitro, followed by the analysis of the effect of LV1 on the growth performance, immune response, and disease resistance of Litopenaeus vannamei. According to the results, the 16 S rDNA sequence of LV1 was 100% homolofgous to the reference sequence of Clostridium butyricum. Moreover, LV1 was resistant to several antibiotics including amikacin, streptomycin, and gentamicin and highly tolerated artificial gastric and artificial intestinal fluids. The whole genome of LV1 was 4625,068 bp in size and included 4336 coding genes. Among these genes, GO, KEGG, and COG databases exhibited the highest number of genes annotated to metabolic pathway classes and 105 genes annotated as glycoside hydrolases. Meanwhile, 176 virulence genes were predicted. The use of diets supplemented with 1.2 × 109 CFU/kg of LV1 live cells significantly increased the weight gain and specific growth rates of Litopenaeus vannamei and the activity of serum superoxide dismutase, glutathione peroxidase, acid phosphatase, and alkaline phosphatase (P < 0.05). Meanwhile, the use of these diets markedly improved the relative expression of intestinal immunity- and growth-related genes. In conclusion, LV1 has excellent probiotic properties. Specifically, the addition of 1.2 × 109 CFU/kg of LV1 live cells to the diet improved the growth performance, immune response, and disease-resistance of Litopenaeus vannamei.

As a novel anticancer candidate, ether extract of Dendrobium nobile overstimulates cellular protein biosynthesis to induce cell stress and autophagy.

Increasing data has confirmed the potential anticancer properties of Dendrobium, a traditional Chinese herb. However, most anticancer compositions from the plant of Dendrobium were usually extracted by high polar solvent, while weak polar compositions with excellent anticancer activity remained largely unexplored. In this study, the differences between ether extract and ethanol extract of Dendrobium nobile Lindl. on chemical components and anticancer activities were investigated, as well as the anticancer mechanisms among different extracts. The results demonstrated that the ether extract exhibited a stronger anticancer effect than ethanol extract, and its anticancer effect was mainly due to weak polar compounds rather than polysaccharides and alkaloids. Quantitative proteomics suggested that the ether extract significantly stimulated the over-expression of immature proteins, the endoplasmic reticulum stress and unfolded protein response were subsequently induced, the intracellular reactive oxygen species level was seriously elevated, and oxidative stress occurred in the meanwhile. Eventually, autophagy and apoptosis were activated to cause cell death. Our findings demonstrate that the ether extract of D. nobile is a potential candidate for anticancer drug development, and that future research on anticancer drugs derived from medicinal plants should also concentrate on weak polar compounds.

Quantitative Identification of Antioxidant Basis for Dendrobium Nobile Flower by High Performance Liquid Chromatography-Tandem Mass Spectrometry.

Dendrobium nobile is a beautiful orchid and a widely used medicinal plant. In vitro antioxidant assays suggested that D. noblie flower extracts showed significantly higher 2, 2'-azinobis-3-ethylbenzthiazoline-6-sulfonate (ABTS) and 1, 1-diphenyl-2-picrylhydrazyl (DPPH) scavenging rates and much more ferric-reducing power than those of root, stem, leaf and fruit. To better understand the antioxidant basis of D. nobile flower, high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) was used for metabolic identification and quantification. Finally, there were 72 metabolites among the total of 712 identified components showed significant association (coefficient >0.8, p < 0.05) with ABTS scavenging rates, DPPH scavenging rates, and ferric-reducing power. The three enriched classes of flower metabolites, including amino acids and their derivatives, organic acids and their derivatives, and flavonoids, formed the main antioxidant basis. The significantly accumulated rutin, astragalin, isomucronulatol-7-O-glucoside, quercetin 4'-O-glucoside, methylquercetin O-hexoside, caffeic acid, caffeic acid O-glucoside, and p-coumaric acid (Log2(fold change) >2, p < 0.01, distribution in flower >0.1%) made a key contribution to the higher antioxidant activities in flower. The relative quantification results of HPLC-MS/MS were verified by the common quantification methods. The antioxidant basis revealed of D. nobile flower will be helpful in the production of healthy or beauty products.

[Correlation analysis and breeding application of agronomic traits and quality indexes in Dendrobium nobile].

The present study aimed to explore the correlation between agronomic traits and quality indexes of Dendrobium nobile and its application value in agricultural breeding. The cultivated strains of D. nobile in Hejiang-Chishui producing areas were extensively collected,and the main agronomic traits and quality indexes were measured. The agronomic traits with significant correlation with quality indexes were screened out by the correlation analysis,and then the parental lines and self-bred F_1 generation plants were furtherverified. Among 96 lines of D. nobile,the content of soluble polysaccharides showed a significant negative correlation with dendrobine( P < 0. 01),and no significant correlation with agronomic traits in stems and leaves. The content of dendrobine exhibited a significant positive correlation with the stem width-thickness ratio( at the largest cross section; P < 0. 01),and no significant correlation with other agronomic traits. Regression analysis further verified the positive correlation between dendrobine content and stem width-thickness ratio( R2> 0. 9). Two lines,JC-10 and JC-35,with significant differences in stem width-thickness ratio were screened out( P <0. 05). The corresponding F1 generation plants by self-pollination both showed that the dendrobine content was higher with greater stem width-thickness ratio( P < 0. 01). The experimental results suggested that within a certain range,the dendrobine content was higher in D. nobile with flatter stem. Therefore,in the breeding of D. nobile,this specific trait could be used for screening plants with high content of quality indexes such as dendrobine.

Dendrobium liquor eliminates free radicals and suppresses cellular proteins expression disorder to protect cells from oxidant damage.

Dendrobium liquor obtained by soaking Dendrobium in Chinese liquor is considered as a health drink in China. Here, we found the pretreatment of extract of Dendrobium nobile Lindl. liquor (DNLE) attenuated the oxidative damage to cells caused by H2 O2 , while the abilities of DNLE of eliminating extracellular free radicals and promoting the activities of intracellular antioxidant enzymes were observed. Quantitative proteomics identified 375 differentially expressed proteins caused by H2 O2 treatment in 293T cells. However, only 12 differentially expressed proteins were found in DNLE-pretreated cells which under the same oxidative damage. This suggested that the pretreatment of DNLE could suppress the disorder of protein expressions caused by oxidative stress which could induce cell death. Besides, DNLE was helpful for avoiding the unfolded protein response (UPR) and cell cycle disorder caused by oxidative stress. Taken together, these results demonstrated that Dendrobium liquor could be a healthy herbal drink with antioxidant function. PRACTICAL APPLICATIONS: Dendrobium is used as an edible herb and a tonic food in traditional Chinese medicine. Dendrobium liquor obtained by soaking Dendrobium with Chinese liquor is also regarded as a nourishing health drink. However, there is rare research data on biological activity of Dendrobium liquor. Our current results demonstrated that the extract of Dendrobium nobile Lindl. liquor (DNLE) possessed the ability of eliminating free radicals in/out the human cells. More importantly, DNLE could help cells to resist the interference on cell life activities caused by oxidative stress. Since many evidences suggested that oxidative stress is linked to human disease and aging, and chemical antioxidant has some side effects on health, Dendrobium liquor can serve as a natural health drink with antioxidant function. Furthermore, the active ingredients in DNLE also possess the potential to be developed as natural antioxidant additive in food and cosmetics.

Quantitative assessment of secondary metabolites and cancer cell inhibiting activity by high performance liquid chromatography fingerprinting in Dendrobium nobile.

Dendrobium nobile is an important medicinal food beneficial for human health, well known for polysaccharides and dendrobine. For fast, accurate, and comprehensive comparison of its quality, high performance liquid chromatography (HPLC) fingerprinting method was constructed. Firstly, spring frost stressed D. nobile herb was observed for assessment. Decreased leaf thickness, chlorophyll, and drying rate, and increased free-proline indicated heavy damages on growth. But, the content of polysaccharides increased significantly in during-frost (DF), and dropped significantly in after-frost (AF). The content of dendrobine accumulated significantly in AF. Then, low similarity among HPLC fingerprints of before-frost (BF), DF, and AF, and 75.82% of significantly variant peaks indicated the changing of much more components. Especially, some less-polar components increased significantly in DF, but not in AF. Moreover, the highest suppression rates (SRs) to A549 lung cancer cells were up to 33.08% in DF, but only 15.63% and 12.12% in BF and AF. After association analysis, eleven less-polar components were found to be significantly and positively correlated to SRs under relatively high concentration. The result shows that frost stress not only causes damages to plant growth, but also promotes the accumulation of some health-beneficial bioactive metabolites. HPLC based fingerprinting method shows good applicability on quality evaluation and bioactivity correlation analysis of complexed agricultural products.

[Quantitative proteomics of inhibitory mechanism of Dendrobium denneanum ether extract on lung cancer cells].

Dendrobium denneanum have been used for a long time as rare medicinal herbs in traditional Chinese medicine. Our previous works found that ether extract of D. denneanum had higher anticancer activities than alcohol or water extract,thus with better development prospects. Quantitative proteomics based on SILAC technique was used to investigate the anticancer mechanism of D. denneanum on lung tumor cell line A549,and 4 855 proteins were detected in A549 cells. Quantitative proteomics experiments found that 193 proteins of A549 cells were up-regulated,and 44 proteins were down-regulated by ether extract of D. denneanum. Those proteins are associated with synthesis,transport and metabolism of biological macromolecules,chaperone,DNA repair,oxidoreductase,cell adhesion,cell cycle,apoptosis and autophagy. Through the function analysis of differentially expressed proteins,it was inferred that ether extract of D. denneanum caused cell protein metabolism disorder,endoplasmic reticulum stress response,abnormal self-repair mechanism of cells,damage of cell adhesion and proliferation; besides,it caused a dramatic increase in ROS level in A549 cells,and upset the balance of intracellular oxidation reduction system. Affected by the above factors,lung cancer cells initiated apoptosis and autophagy,which accelerated cell death. This research explains the anticancer mechanism of D. denneanum from the perspective of quantitative proteomics,and lays a foundation for future research and development of new anticancer drugs based on ether extract of D. denneanum.

Ligand fishing of anti-neurodegenerative components from Lonicera japonica using magnetic nanoparticles immobilised with monoamine oxidase B.

In this work, monoamine oxidase B was immobilised onto magnetic nanoparticles to prepare a new type of affinity solid-phase extraction adsorbent, which was used to extract the possible anti-neurodegenerative components from the Lonicera japonica flower extracts. Coupled with high-performance liquid chromatography with mass spectrometry, two monoamine oxidase B ligands were fished-out and identified as isochlorogenic acid A and isochlorogenic acid C, which were found to be inhibitors of the enzyme for the first time, with similar half maximal inhibitory concentration values of 29.05 ± 0.49 and 29.77 ± 1.03 µM, respectively. Furthermore, equilibrium-dialysis dissociation assay of enzyme-inhibitor complex showed that both compounds have reversible binding patterns to monoamine oxidase B, and kinetic analysis demonstrated that they were mixed-type inhibitors for monoamine oxidase B, with Ki and Kis values of 9.55 and 37.24 µM for isochlorogenic acid A, 9.53 and 35.50 µM for isochlorogenic acid C, respectively. The results indicated that isochlorogenic acid A and isochlorogenic acid C were the major active components responsible for the anti-degenerative activity of the flowers of L. japonica, while magnetic nanoparticles immobilised monoamine oxidase B could serve as an efficient solid-phase extraction adsorbent to specifically extract monoamine oxidase B inhibitors from complex herbal extracts.

Genome-wide researches and applications on Dendrobium.

MAIN CONCLUSION: This review summarizes current knowledge of chromosome characterization, genetic mapping, genomic sequencing, quality formation, floral transition, propagation, and identification in Dendrobium. The widely distributed Dendrobium has been studied for a long history, due to its important economic values in both medicine and ornamental. In recent years, some species of Dendrobium and other orchids had been reported on genomic sequences, using the next-generation sequencing technology. And the chloroplast genomes of many Dendrobium species were also revealed. The chromosomes of most Dendrobium species belong to mini-chromosomes, and showed 2n = 38. Only a few of genetic studies were reported in Dendrobium. After revealing of genomic sequences, the techniques of transcriptomics, proteomics and metabolomics could be employed on Dendrobium easily. Some other molecular biological techniques, such as gene cloning, gene editing, genetic transformation and molecular marker developing, had also been applied on the basic research of Dendrobium, successively. As medicinal plants, insights into the biosynthesis of some medicinal components were the most important. As ornamental plants, regulation of flower related characteristics was the most important. More, knowledge of growth and development, environmental interaction, evolutionary analysis, breeding of new cultivars, propagation, and identification of species and herbs were also required for commercial usage. All of these studies were improved using genomic sequences and related technologies. To answer some key scientific issues in Dendrobium, quality formation, flowering, self-incompatibility and seed germination would be the focus of future research. And genome related technologies and studies would be helpful.

Rapid quantification of aloin A and B in aloe plants and aloe-containing beverages, and pharmaceutical preparations by microchip capillary electrophoresis with laser induced fluorescence detection.

A microchip capillary electrophoresis coupled with laser induced fluorescence detection method for the fast determination of aloin was developed and comprehensively applied for the quantification of aloin A and B present in seven aloe plant species, 42 aloin-containing crude drugs, ten aloe pharmaceutical preparations, and four aloe gel-containing functional foods. The excitation and emission wavelengths for detection of both aloins were set at 473 and 520 nm, respectively. Sample analysis on a 35 mm length of glass microchip channel was completed within 40 s. An interference study indicated that the other main anthraquinones present in the samples did not interrupt with the target aloins detection, demonstrating the good selectivity of this method. It is demonstrated that this method is fast, facile, and specific for determination of aloin A and B from matrix samples which can be applied to the quality control of a wide varieties of aloe species and aloe-derived products.

ROS-Mediated 15-Hydroxyprostaglandin Dehydrogenase Degradation via Cysteine Oxidation Promotes NAD+-Mediated Epithelial-Mesenchymal Transition.

Nicotinamide adenine dinucleotide (NAD) levels decrease with aging as a result of aging-associated CD38 upregulation. Here, we established a cell model with decreased cellular NAD levels by overexpressing CD38 or treating cells with FK866, an inhibitor of nicotinamide phosphoribosyltransferase. We revealed that decreased NAD triggered reactive oxygen species (ROS)-mediated degradation of 15-hydroxyprostaglandin dehydrogenase (15-PGDH), which drove cells to undergo epithelial-mesenchymal transition (EMT). Moreover, we showed that oxidation of the Cys44 residue to sulfonic acid in 15-PGDH led to its degradation via non-canonical ubiquitination-proteasome and autophagy pathways. Mutation of Cys44 to alanine abolished ROS-induced 15-PGDH degradation. We demonstrated that 15-PGDH silencing promoted EMT, whereas supplementation with NAD precursors increased NAD and 15-PGDH stability, and reversed the EMT process. Taken together, these results suggest that declining NAD levels contribute to age-dependent increases in cancer incidence, and repletion of NAD precursors is beneficial for increasing 15-PGDH expression.