Metagenomics and untargeted metabolomics analyses to unravel the formation mechanism of characteristic metabolites in Cantonese soy sauce during different fermentation stages.

Cantonese soy sauce (CSS) is an important Chinese condiment due to its distinctive flavor. Microorganisms play a significant role in the flavor formation of CSS during fermentation. However, the correlation between microbes and flavor compounds as well as the potential fermentation mechanism remained poorly uncovered. Here we revealed the dynamic changes of microbial structure and characteristics metabolites as well as their correlation of CSS during the fermentation process. Metagenomics sequencing analysis showed that Tetragenococcus halophilus, Weissella confusa, Weissella paramesenteroides, Aspergillus oryzae, Lactiplantibacillus plantarum, Weissella cibaria were top six dominant species from day 0 to day 120. Sixty compounds were either positively or tentatively identified through untargeted metabolomics profile and they were 27 peptides, amino acids and derivatives, 8 carbohydrates and conjugates, 14 organic acids and derivatives, 5 amide compounds, 3 flavonoids and 3 nucleosides. Spearman correlation coefficient indicated that Tetragenococcus halophilus, Zygosaccharomyces rouxii, Pediococcus pentosaceus and Aspergillus oryzae were significantly related with the formation of taste amino acids and derivatives, peptides and functional substances. Additionally, the metabolisms of flavor amino acids including 13 main free amino acids were also profiled. These results provided valuable information for the production practice in the soy sauce industry.

The latest advances on soy sauce research in the past decade: Emphasis on the advances in China.

As an indispensable soybean-fermented condiment, soy sauce is extensively utilized in catering, daily cooking and food industry in East Asia and Southeast Asia and is becoming popular in the whole world. In the past decade, researchers began to pay great importance to the scientific research of soy sauce, which remarkably promoted the advances on fermentation strains, quality, safety, function and other aspects of soy sauce. Of them, the screening and reconstruction of Aspergillus oryzae with high-yield of salt and acid-tolerant proteases, mechanism of soy sauce flavor formation, improvement of soy sauce quality through the combination of novel physical processing technique and microbial/enzyme, separation and identification of soy sauce functional components are attracting more attention of researchers, and related achievements have been reported continually. Meanwhile, we pointed out the drawbacks of the above research and the future research directions based on published literature and our knowledge. We believe that this review can provide an insightful reference for international related researchers to understand the advances on soy sauce research.

Genome-Wide Comparison and Functional Characterization of HMGR Gene Family Associated with Shikonin Biosynthesis in Lithospermum erythrorhizon.

3-hydroxy-3-methylglutaryl-CoA reductase (HMGR), as the rate-limiting enzyme in the mevalonate pathway, is essential for the biosynthesis of shikonin in Lithospermum erythrorhizon. However, in the absence of sufficient data, the principles of a genome-wide in-depth evolutionary exploration of HMGR family members in plants, as well as key members related to shikonin biosynthesis, remain unidentified. In this study, 124 HMGRs were identified and characterized from 36 representative plants, including L. erythrorhizon. Vascular plants were found to have more HMGR family genes than nonvascular plants. The phylogenetic tree revealed that during lineage and species diversification, the HMGRs evolved independently and intronless LerHMGRs emerged from multi-intron HMGR in land plants. Among them, Pinus tabuliformis and L. erythrorhizon had the most HMGR gene duplications, with 11 LerHMGRs most likely expanded through WGD/segmental and tandem duplications. In seedling roots and M9 cultured cells/hairy roots, where shikonin biosynthesis occurs, LerHMGR1 and LerHMGR2 were expressed significantly more than other genes. The enzymatic activities of LerHMGR1 and LerHMGR2 further supported their roles in catalyzing the conversion of HMG-CoA to mevalonate. Our findings provide insight into the molecular evolutionary properties and function of the HMGR family in plants and a basis for the genetic improvement of efficiently produced secondary metabolites in L. erythrorhizon.

Characterization of the key aroma compounds in soy sauce by gas chromatography-mass spectrometry-olfactometry, headspace-gas chromatography-ion mobility spectrometry, odor activity value, and aroma recombination and omission analysis.

Most previous studies on volatile compounds in soy sauce were performed by gas chromatography-mass spectrometry (GC-MS). In this study, the volatile compounds of high-salt liquid-state fermentation soy sauce (HLFSS) were analyzed qualitatively and quantitatively by GC-MS and headspace-gas chromatography-ion mobility spectrometry (HS-GC-IMS). One hundred and seventy-four substances were detected using the two instruments, 87 by HS-GC-IMS and 127 by GC-MS. Aldehydes (26), ketones (28), esters (29), and alcohols (26) were the main compounds in HLFSS. In addition, ethyl pyruvate, (E)-2-pentenal and diethyl propanedioate were detected by HS-GC-IMS, which were previously not detected in HLFSS. Forty-eight aromatics including 34 key ones were identified by gas chromatography-olfactometry. Phenylacetaldehyde, methional, 2-methylbutanal, 1-octen-3-ol, ethyl acetate, 2-ethyl-4-hydroxy-5-methyl-3(2H)-furanone, 4-hydroxy-2,5-dimethyl-3(2H)-furanone and 4-ethyl guaiacol were identified as the main aroma compounds in HLFSS by aroma recombination and omission test. This study laid foundation for developing flavor assessment standards for soy sauce.

Applications of multi-omics techniques to unravel the fermentation process and the flavor formation mechanism in fermented foods.

Fermented foods are important components of the human diet. There is increasing awareness of abundant nutritional and functional properties present in fermented foods that arise from the transformation of substrates by microbial communities. Thus, it is significant to unravel the microbial communities and mechanisms of characteristic flavor formation occurring during fermentation. There has been rapid development of high-throughput and other omics technologies, such as metaproteomics and metabolomics, and as a result, there is growing recognition of the importance of integrating these approaches. The successful applications of multi-omics approaches and bioinformatics analyses have provided a solid foundation for exploring the fermentation process. Compared with single-omics, multi-omics analyses more accurately delineate microbial and molecular features, thus they are more apt to reveal the mechanisms of fermentation. This review introduces fermented foods and an overview of single-omics technologies - including metagenomics, metatranscriptomics, metaproteomics, and metabolomics. We also discuss integrated multi-omics and bioinformatic analyses and their role in recent research progress related to fermented foods, as well as summarize the main potential pathways involved in certain fermented foods. In the future, multilayered analyses of multi-omics data should be conducted to enable better understanding of flavor formation mechanisms in fermented foods.

Multiple Functions of MiRNAs in Brassica napus L.

The worldwide climate changes every year due to global warming, waterlogging, drought, salinity, pests, and pathogens, impeding crop productivity. Brassica napus is one of the most important oil crops in the world, and rapeseed oil is considered one of the most health-beneficial edible vegetable oils. Recently, miRNAs have been found and confirmed to control the expression of targets under disruptive environmental conditions. The mechanism is through the formation of the silencing complex that mediates post-transcriptional gene silencing, which pairs the target mRNA and target cleavage and/or translation inhibition. However, the functional role of miRNAs and targets in B. napus is still not clarified. This review focuses on the current knowledge of miRNAs concerning development regulation and biotic and abiotic stress responses in B. napus. Moreover, more strategies for miRNA manipulation in plants are discussed, along with future perspectives, and the enormous amount of transcriptome data available provides cues for miRNA functions in B. napus. Finally, the construction of the miRNA regulatory network can lead to the significant development of climate change-tolerant B. napus through miRNA manipulation.

Enhancing organic selenium content and antioxidant activities of soy sauce using nano-selenium during soybean soaking.

Nano-selenium has a greater potential than inorganic selenium in preventing selenium-deficiency diseases due to its higher safety. In this study, spherical nano-selenium particles (53.8 nm) were prepared using sodium selenite, ascorbic acid and chitosan. Selenium-enriched soy sauces were prepared by soaking soybean in nano-selenium and sodium selenite solutions (2-10 mg/L), respectively. Total selenium and organic selenium contents of soy sauces prepared by nano-selenium and sodium selenite were increased by 32-191-fold and 29-173-fold compared to the control (without selenium), and organic selenium accounted for over 90% of total selenium. Soy sauce prepared using 6 mg/L nano-selenium had the strongest antioxidant activities, which were 9.25-28.02% higher than the control. Nano-selenium (6 mg/L) markedly enhanced the koji's enzyme activities (9.76-33.59%), then the latter promoted the release of total phenolics (27.54%), total flavonoids (27.27%) and the formation of free amino acids (16.19%), Maillard reaction products (24.50%), finally the antioxidant activities of selenium-enriched soy sauce were enhanced.

Natural shikonin and acetyl-shikonin improve intestinal microbial and protein composition to alleviate colitis-associated colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) and inflammatory bowel disease (IBD) are the most common diseases of human digestive system. Nowadays, the influence of the inflammatory microenvironment on tumorigenesis has become a new direction, and the exploration of relative molecular mechanism will facilitate the discovery and identification of novel potential anti-cancer molecules. METHODS: Natural shikonin (SK) and acetyl-shikonin (acetyl-SK) was administered to azoxymethane (AOM)/dextran sodium sulphate (DSS)-induced colitis-associated colorectal cancer (CAC) mice model by gavage to investigate their therapeutic effects. Moreover, fresh feces and colon tissues were collected for determining the function of SK and acetyl-SK on the gut microbes and protein expression, respectively. RESULTS: Both SK and acetyl-SK decreased AOM/DSS-induced CAC, and regulated the intestinal flora structure in CAC mouse model. They, especially SK, improved species richness, evenness and diversity of intestinal flora, recovered the upregulated ratio of Firmicutes to Bacteroidota (F/B ratio) which symbolizes gut microbiota dysbiosis. SK and its derivative increased the beneficial bacteria g__norank_f__Muribaculaceae, Lactobacillus, Lachnospiraceae_NK4A136_Group, and reduced those harmful ones including Ileibacterium and Coriobacteriaceae UCG-002. Notably, AOM/DSS caused significant increase in the abundance of Ileibaterium valens and g__norank_f__norank_o__Clostridia_UCG-014, which were not previously reported in studies of colonic inflammation or cancer, and the disorder was reversed by 20 mg/kg of SK. In our current study, the action of SK and acetyl-SK is dose-dependent, and 20 mg/kg SK exhibited the most effective functions, even better than the positive drug mesalazine. Moreover, differential proteomics and ELISA results showed that SK could recover the increase of pro-inflammatory cytokines (including IL-1ß, IL-6 and TNF-α), the upregulation of pyruvate kinase isozyme type M2 (PKM2) and some other proteins (mainly concentrated in transcriptional mis-regulation in cancer and IL-17 signaling pathways), and the downregulation of Aldh1b1-Acc3-Maoa and Μgt2b34-Aldh1a1-Aldh1a7 involved in Wnt/ß-catenin signaling pathway. CONCLUSION: Our study identified SK and acetyl-SK, especially SK, as potential preventive agents for CAC through regulating both gut microbes and pathways involved in inflammation and cancer such as Wnt/ß-catenin signaling pathway.

Anti-microbial efficacy, mechanisms and druggability evaluation of the natural flavonoids.

AIMS: This study was conducted to evaluate 35 natural flavonoids for their in vitro susceptibility against E. coli (ATCC 25922), Ps. aeruginosa (ATCC 27853), B. subtilis (ATCC 530) and Staph. aureus (ATCC 6538) in search of a potential broad-spectrum antibiotic. METHODS AND RESULTS: Glabridin, a natural isoflavonoid isolated from Glycyrrhiza glabra L., was identified to be highly active with a MIC of 8-16 µg ml-1 against Staph. aureus, B. subtilis and E. coli. By the results of the docking simulation, we located the potential targets of glabridin as DNA gyrase and dihydrofolate reductase (DHFR). The subsequent DNA gyrase inhibition assays (glabridin: IC50  = 0.8516 µmol L-1 , ciprofloxacin: IC50  = 0.04697 µmol L-1 ), DHFR inhibition assays (glabridin: inhibition ratio = 29%, methotrexate: inhibition ratio = 45% under 100 µmol L-1 treatment) and TUNEL confirmed that glabridin acted as DNA gyrase inhibitor and DHFR mild inhibitor, exerting bactericidal activity by blocking bacterial nucleic acid synthesis. CCK-8 and in silico calculations were also conducted to verify the low cytotoxicity and acceptable druggability of glabridin. CONCLUSION: These findings suggest that glabridin represents the prototypical member of an exciting structural class of natural antimicrobial agents. SIGNIFICANCE AND IMPACT OF THE STUDY: This study reports a novel mechanism of bactericidal activity of glabridin against Staph. aureus.

Enzymatic Synthesis of Diacylglycerol-Enriched Oil by Two-Step Vacuum-Mediated Conversion of Fatty Acid Ethyl Ester and Fatty Acid From Soy Sauce By-Product Oil as Lipid-Lowering Functional Oil.

Soy sauce by-product oil (SSBO), a by-product of the soy sauce production process, is the lack of utilization due to an abundance of free fatty acid (FFA) and fatty acid ethyl ester (EE). The utilization of low-cost SSBO to produce value-added diacylglycerol (DAG)-enriched oil and its applications are promising for the sustainability of the oil industry. The objective of this study was to utilize SSBO containing a high content of EE and FFA as raw material to synthesize DAG-enriched oil and to evaluate its nutritional properties in fish. Based on different behaviors between the glycerolysis of EE and the esterification of FFA in one-pot enzymatic catalysis, a two-step vacuum-mediated conversion was developed for the maximum conversions of EE and FFA to DAG. After optimization, the maximum DAG yield (66.76%) and EE and FFA conversions (96 and 93%, respectively) were obtained under the following optimized conditions: lipase loading 3%, temperature 38°C, substrate molar ratio (glycerol/FFA and EE) 21:40, a vacuum combination of 566 mmHg within the initial 10 h and 47 mmHg from the 10th to 14th hour. Further nutritional study in fish suggested that the consumption of DAG-enriched oil was safe and served as a functional oil to lower lipid levels in serum and liver, decrease lipid accumulation and increase protein content in body and muscle tissues, and change fatty acid composition in muscle tissues. Overall, these findings were vital for the effective utilization of SSBO resources and the development of future applications for DAG-enriched oil as lipid-lowering functional oil in food.

Degradation Mechanism of Soybean Protein B3 Subunit Catalyzed by Prolyl Endopeptidase from Aspergillus niger during Soy Sauce Fermentation.

Soy sauce secondary precipitate formed due to the B3 subunit seriously affects soy sauce's appearance quality. In this study, a prolyl endopeptidase (APE) from Aspergillus niger, which could degrade approximately 50% of the B3 subunit and increase proline content by 24% in soy sauce, was isolated and identified. The results showed that APE was an acidic salt-tolerant serine protease (62 kDa), which was optimally active at 40 °C and pH 4.0, and retained more than 69% activity in 3 M NaCl solution over 10 days. As a potential substrate of APE, the B3 subunit contains 10 proline residues. High salinity could not damage the hydrogen bonds, salt bridges, and interior hydrophobicity of APE; thus, the spatial structures and activity of APE in 3 M NaCl solution were stable within 3 days and decreased thereafter. High salinity made the B3 subunit more rigid and lowered the catalytic activity of APE on the B3 subunit, hindering complete hydrolysis of the B3 subunit. This was the first report about the APE capable of degrading the B3 subunit and reducing the secondary precipitate of soy sauce, providing a new possibility to solve the secondary precipitate of soy sauce.

Deciphering the rhizobacterial assemblages under the influence of genetically engineered maize carrying mcry genes.

Genetically engineered (GE) maize has been thoroughly studied regarding its agro-environmental impact; however, its concerns for the soil environment remain. This work was aimed to decode rhizosphere microbe interactions and potential ecological hazards associated with GE maize. Rhizobacterial communities of field grown transgenic insect-resistant 2A5 maize carrying mcry1Ab and mcry2Ab genes were compared with control Z58 using PacBio sequencing platform. Also full-length 16S rDNA gene sequencing was used to verify the partial (V3-V4) sequencing results obtained in 2017. Measures of α-diversity displayed transgenic 2A5 to be significantly lower in species richness at the flowering stage; however, diversity remained undisturbed. ß-diversity was least affected by genetic modifications where similar community profiles were shared by transgenic 2A5 and control Z58. In addition, root exudation patterns were found to drive variations in bacterial assemblages based on developmental stages. For example, genus Massilia successfully colonized the rhizosphere at jointing stage, while Mucilaginobacter showed higher relative abundance in flowering stages of both 2A5 and Z58. These members are known to possess attributes related to plant growth. The impact of dual-transgene insertion on nifH gene abundance was also analyzed where no apparent significant difference in nifH gene copy number was observed. Our results confirmed that full-length 16S rDNA sequencing was sufficient to provide higher taxonomic resolution. Also, results of our 2-year field trials confirmed that there is no significant impact of mcry gene integration on belowground biomasses. Therefore, GE insect-resistant 2A5 maize carrying mcry1Ab and mcry2Ab genes can continue to benefit human populations by increasing crop productivity. In future, further research needs to be catalyzed to analyze the impact of Bt-insertion on microbial community structure across the years for ecosystem sustainability.

Differential microbial assemblages associated with shikonin-producing Borage species in two distinct soil types.

Shikonin and its derivatives are the main components of traditional Chinese medicine, Zicao. The pharmacological potential of shikonin and its derivatives have been extensively studied. Yet, less is known about the microbial assemblages associated with shikonin producing Borage plants. We studied microbial profiles of two Borage species, Echium plantagineum (EP) and Lithospermum erythrorhizon (LE), to identify the dynamics of microbial colonization pattern within three rhizo-compatments and two distinct soil types. Results of α and ß-diversity via PacBio sequencing revealed significantly higher microbial richness and diversity in the natural soil along with a decreasing microbial gradient across rhizosphere to endosphere. Our results displayed genotype and soil type-dependent fine-tuning of microbial profiles. The host plant was found to exert effects on the physical and chemical properties of soil, resulting in reproducibly different micro-biota. Analysis of differentially abundant microbial OTUs displayed Planctomycetes and Bacteroidetes to be specifically enriched in EP and LE rhizosphere while endosphere was mostly prevailed by Cyanobacteria. Network analysis to unfold co-existing microbial species displayed different types of positive and negative interactions within different communities. The data provided here will help to identify microbes associated with different rhizo-compartments of potential host plants. In the future, this might be helpful for manipulating the keystone microbes for ecosystem functioning.

Design, synthesis and biological evaluation of anilide (dicarboxylic acid) shikonin esters as antitumor agents through targeting PI3K/Akt/mTOR signaling pathway.

Triple-negative breast cancer (TNBC) has an unfavorable prognosis attribute to its low differentiation, rapid proliferation and high distant metastasis rate. PI3K/Akt/mTOR as an intracellular signaling pathway plays a key role in the cell proliferation, migration, invasion, metabolism and regeneration. In this work, we designed and synthesized a series of anilide (dicarboxylic acid) shikonin esters targeting PI3K/Akt/mTOR signaling pathway, and assessed their antitumor effects. Through three rounds of screening by computer-aided drug design method (CADD), we preliminarily obtained sixteen novel anilide (dicarboxylic acid) shikonin esters and identified them as excellent compounds. CCK-8 assay results demonstrated that compound M9 exhibited better antiproliferative activities against MDA-MB-231, A549 and HeLa cell lines than shikonin (SK), especially for MDA-MB-231 (M9: IC50 = 4.52 ± 0.28 µM; SK: IC50 = 7.62 ± 0.26 µM). Moreover, the antiproliferative activity of M9 was better than that of paclitaxel. Further pharmacological studies showed that M9 could induce apoptosis of MDA-MB-231 cells and arrest the cell cycle in G2/M phase. M9 also inhibited the migration of MDA-MB-231 cells by inhibiting Wnt/ß-catenin signaling pathway. In addition, western blot results showed that M9 could inhibit cell proliferation and migration by down-regulating PI3K/Akt/mTOR signaling pathway. Finally, a three-dimensional quantitative structure-activity relationship (3D-QSAR) model was also constructed to provide a basis for further development of shikonin derivatives as potential antitumor drugs through structure-activity relationship analysis. To sum up, M9 could be a potential candidate for TNBC therapy.

Differential relieving effects of shikonin and its derivatives on inflammation and mucosal barrier damage caused by ulcerative colitis.

BACKGROUND: Ulcerative colitis (UC) is one of the most challenging human diseases. Natural shikonin (SK) and its derivatives (with have higher accumulation) isolated from the root of Lithospermum erythrorhizon have numerous beneficial effects, such as wound healing and anti-inflammatory activities. Some researchers have reported that hydroxynaphthoquinone mixture (HM) and SK attenuate the acute UC induced by dextran sulfate sodium (DSS). However, no existing study has systemically investigated the effectiveness of SK and other hydroxynaphthoquinone natural derivative monomers on UC. METHODS: In this study, mice were treated with SK and its derivatives (25 mg/kg) and mesalazine (200 mg/kg) after DSS administration daily for one week. Disease progression was monitored daily by observing the changes in clinical signs and body weight. RESULTS: Intragastric administration natural single naphthoquinone attenuated the malignant symptoms induced by DSS. SK or its derivatives remarkably suppressed the serum levels of pro-inflammatory cytokines while increasing the inflammatory cytokine interleukin (IL)-10 . Additionally, both SK and alkanin restrained the activities of cyclooxygenase-2 (COX-2), myeloperoxidase (MPO) and inducible nitric oxide synthase (iNOS) in serum and colonic tissues. SK and its derivatives inhibited the activation of nucleotide binding oligomerization domain-like receptors (NLRP3) inflammasome and NF-κB signaling pathway, thereby relieving the DSS-induced disruption of epithelial tight junction (TJ) in colonic tissues. CONCLUSIONS: Our findings shed more lights on the pharmacological efficacy of SK and its derivatives in UC against inflammation and mucosal barrier damage.

Establishment of the hairy root culture of Echium plantagineum L. and its shikonin production.

Echium plantagineum L. (Boraginaceae) is an invasive species in Australia and contains medicinal shikonins in its roots. In this study, the hairy root lines of E. plantagineum were established using Agrobacterium rhizogenes strain ATCC15834 and confirmed by the amplification of the rolB gene. Results showed significant difference in shikonin production between the hairy root lines in the 1/2B5 and M9 media. The biomass of the lines in the 1/2B5 medium was fivefold of that in the M9 medium. However, the components of detected shikonins were similar in these two liquid media. By contrast, different accumulation profiles appeared in the hairy root lines. HPLC analysis revealed the presence of nine possible related compounds, including shikonins, and acetylshikonin was the most abundant shikonin derivative. The content of acetylshikonin in the 1/2B5 medium (36.25 mg/L on average) was twofold of that in the M9 medium. Our results showed that the hairy root cultures of E. plantagineum can be used in enhancing the production of potential pharmaceutical compounds, such as acetylshikonin.

Design, synthesis and biological evaluation of benzoylacrylic acid shikonin ester derivatives as irreversible dual inhibitors of tubulin and EGFR.

In this study, a series of shikonin derivatives combined with benzoylacrylic had been designed and synthesized, which showed an inhibitory effect on both tubulin and the epidermal growth factor receptor (EGFR). In vitro EGFR and cell growth inhibition assay demonstrated that compound PMMB-317 exhibited the most potent anti-EGFR (IC50 = 22.7 nM) and anti-proliferation activity (IC50 = 4.37 µM) against A549 cell line, which was comparable to that of Afatinib (EGFR, IC50 = 15.4 nM; A549, IC50 = 6.32 µM). Our results on mechanism research suggested that, PMMB-317 could induce the apoptosis of A549 cells in a dose- and time-dependent manner, along with decrease in mitochondrial membrane potential (MMP), production of ROS and alterations in apoptosis-related protein levels. Also, PMMB-317 could arrest cell cycle at G2/M phase to induce cell apoptosis, and inhibit the EGFR activity through blocking the signal transduction downstream of the mitogen-activated protein MAPK pathway and the anti-apoptotic kinase AKT pathway; typically, such results were comparable to those of afatinib. In addition, PMMB-317 could suppress A549 cell migration through the Wnt/ß-catenin signaling pathway in a dose-dependent manner. Additionally, molecular docking simulation revealed that, PMMB-317 could simultaneously combine with EGFR protein (5HG8) and tubulin (1SA0) through various forces. Moreover, 3D-QSAR study was also carried out, which could optimize our compound through the structure-activity relationship analysis. Furthermore, the in vitro and in vivo results had collectively confirmed that PMMB-317 might serve as a promising lead compound to further develop the potential therapeutic anticancer agents.

Identification of new shikonin derivatives as STAT3 inhibitors.

The signal transducer and activator of transcription 3 is a constitutively activated oncogenic protein in various human tumors and represents a valid target for anticancer drug design. In this study, we have achieved a new type of STAT3 inhibitors based on structural modifications on shikonin scaffold, guided by computational modelling. By tests, PMMB-187 exhibited a more outstanding profile than shikonin on a small panel of human breast cancer cells, especially for the MDA-MB-231 cells. For the cellular mechanisms research, PMMB-187 was found to induce cell apoptosis in MDA-MB-231 cells, associated with the reduction of mitochondrial membrane potential, production of ROS and alteration of the levels of apoptosis-related proteins. Furthermore, PMMB-187 inhibited constitutive/inducible STAT3 activation, transcriptional activity, nuclear translocation and downstream target genes expression in STAT3-dependent breast cancer cells MDA-MB-231. Besides, no obvious inhibitory effect on activation of STAT1 and STAT5 was observed with PMMB-187 treatment. Most notably, the in vivo studies further revealed that PMMB-187 could dramatically suppress the MDA-MB-231 cells xenografted tumor growth. The in vitro and in vivo results collectively suggest that PMMB-187 may serve as a promising lead compound for the further development of potential therapeutic anti-neoplastic agents.

Transcriptome analysis explores genes related to shikonin biosynthesis in Lithospermeae plants and provides insights into Boraginales' evolutionary history.

Shikonin and its derivatives extracted from Lithospermeae plants' red roots have current applications in food and pharmaceutical industries. Previous studies have cloned some genes related to shikonin biosynthesis. However, most genes related to shikonin biosynthesis remain unclear, because the lack of the genome/transcriptome of the Lithospermeae plants. Therefore, in order to provide a new understanding of shikonin biosynthesis, we obtained transcriptome data and unigenes expression profiles in three shikonin-producing Lithospermeae plants, i.e., Lithospermum erythrorhizon, Arnebia euchroma and Echium plantagineum. As a result, two unigenes (i.e., G10H and 12OPR) that are involved in "shikonin downstream biosynthesis" and "methyl jasmonate biosynthesis" were deemed to relate to shikonin biosynthesis in this study. Furthermore, we conducted a Lamiids phylogenetic model and identified orthologous unigenes under positive selection in above three Lithospermeae plants. The results indicated Boraginales was more relative to Solanales/Gentianales than to Lamiales.

Effect of Traditional Chinese Herbal Medicine with Antiquorum Sensing Activity on Pseudomonas aeruginosa.

Traditional Chinese herbal medicines (TCHMs) were tested for their ability of antiquorum sensing. Water extracts of Rhubarb, Fructus gardeniae, and Andrographis paniculata show antiquorumsensing activity when using Chromobacterium violaceum CV12472 as reporter; the sub-MIC concentrations of these TCHMs were tested against AHL-dependent phenotypic expressions of PAO1. Results showed significant reduction in pyocyanin pigment, protease, elastase production, and biofilm formation in PAO1 without inhibiting the bacterial growth, revealing that the QSI by the extracts is not related to static or killing effects on the bacteria. The results indicate a potential modulation of bacterial cell-cell communication, P. aeruginosa biofilm, and virulence factors by traditional Chinese herbal medicine. This study introduces not only a new mode of action for traditional Chinese herbal medicines, but also a potential new therapeutic direction for the treatment of bacterial infections, which have QSI activity and might be important in reducing virulence and pathogenicity of pathogenic bacteria.