Search

BackgroundGinseng has been used in biomedicine to prevent and treat decreased physical and mental capacities. Total ginsenosides (TG) from ginseng root which have antitumor and immune-enhancing properties, are the principal active components of Panax ginseng, while the sulphation-modified TG derivative-3 (SMTG-d3) was expected to enhance the anticancer activity in conventional medicinal treatments.MethodsThe chlorosulphonic acid–pyridine technique, used for the sulfation modification of TG to improve their biological activity, and the infrared spectroscopic characteristics of TG and SMTG-d3 were investigated, and the effects of SMTG-d3 on immunocytes and cytokines relevant to tumor treatment were assessed. The MTT assay was used to assess the effect of TG and SMTG-d3 on the cytotoxicity and T-lymphocytic proliferation against mouse splenocytes. The LDH method was employed to evaluate NK activity induced by TG or SMTG-d3. The production levels of splenocytes-secreted IL-2 and IFN-γ and peritoneal macrophages-secreted TNF-α were determined using mouse ELISA kits.Results and discussionIt showed that the ideal conditions for the sulfation modification of TG: the volume ratio of chlorosulfonic acid to pyridine lower than 1:2.5; controlled amount of chlorosulfonic acid; and a yield of 51.5% SMTG-d3 (2 h, < 45°C). SMTG-d3 showed two characteristic absorption peaks at 1,230 cm−1 and 810 cm−1, indicating the formation of sulfuric acid esters and the presence of sulfuric acid groups. SMTG-d3 exhibited higher antitumor immunological activity than TG by promoting the proliferation of T lymphocytes and the production of IFN-γ and TNF-α, thus enhancing NK cell activity, and reducing cytotoxicity. The findings imply sulfated modification represents an effective method of enhancing the immunomodulatory activities of TG and could be used as the basis for developing new drug target compounds; SMTG-d3 can serve as an antitumor immunomodulator and can be considered an effective and prospective herbal formulation in clinical applications.

Abstract Background Starch is one of the most important renewable polysaccharides in nature for production of bio-ethanol. The starch saccharification step facilitates the depolymerization of starch to yield glucose for biofuels production. The filamentous fungus Aspergillus niger (A. niger) is the most used microbial cell factory for production of the commercial glucoamylase. However, the role of each component in glucoamylases cocktail of A. niger O1 for starch saccharification remains unclear except glucoamylase. Results In this study, we identified the key enzymes contributing to the starch saccharification process are glucoamylase, α-amylase and acid α-amylase out of 29 glycoside hydrolases from the 6-day fermentation products of A. niger O1. Through the synergistic study of the multienzymes for the starch saccharification in vitro, we found that increasing the amount of α-amylase by 5-10 times enhanced the efficiency of starch saccharification by 14.2-23.2%. Overexpression of acid α-amylase in strain O1 in vivo increased the total glucoamylase activity of O1 cultures by 15.0%. Conclusions Our study clarifies the synergistic effects among the components of glucoamylases cocktail, and provides an effective approach to optimize the profile of saccharifying enzymes of strain O1 for improving the total glucoamylase activity.

Dextrose equivalent of glucose from starch hydrolysis is a critical index for starch-hydrolysis industry. Improving glucose yield and decreasing the non]-fermentable sugars which caused by transglycosylation activity of the enzymes during the starch saccharification is an important direction. In this study, we identified two key α-glucosidases responsible for producing non-fermentable sugars in an industrial glucoamylase-producing strain Aspergillus niger O1. The results showed the transglycosylation product panose was decreased by more than 88.0% in agdA/agdB double knock-out strains than strain O1. Additionally, the B-P1 domain of agdB was found accountable as starch hydrolysis activity only, and B-P1 overexpression in ΔAΔB-21 significantly increased glucoamylase activity whereas keeping the glucoamylase cocktail low transglycosylation activity. The total amounts of the transglycosylation products isomaltose and panose were significantly decreased in final strain B-P1-3 by 40.7% and 44.5%, respectively. The application of engineered strains will decrease the cost and add the value of product for starch biorefinery.

Artemisia annua (AAH) is traditionally used as an anti-malarial, expectorant and antipyretic Chinese medicine. The aim of this study was to explore the therapeutic effect of Qinghao Powder (QHP) on chicken coccidiosis, evaluate the safe dosage of QHP, and provide test basis for clinical medication. High-performance liquid chromatography (HPLC) and thin-layer chromatography (TLC) were used to detect artemisinin in Qinghao Powder (QHP) for quality control. The level of artemisinin in QHP was 81.03 mg/g. A total of 210 chicks (14 days of age) were divided randomly into seven groups: three QHP treatments (0.15, 0.30, and 0.60 g/kg), a toltrazuril control (1.00 mL/L), a sulfachloropyrazine sodium control (SSC, 0.30 g/L), an E. tenella-infected control, and a healthy control group. All the groups were inoculated orally with 7 × 104E. tenella oocysts except for the healthy control group. After seven days of administration, compared with the infected control group, chicks which were administered QHP, SS, and toltrazuril showed less bloody feces, oocyst output, and cecal lesions, and the protection rates were improved. The maximum rBWG and ACI were found in the SS-medicated group, followed by the groups medicated with 0.60 and 0.30 g/kg QHP. Therefore, a 0.30 g/kg dose level of QHP in the feed was selected as the recommend dose (RD) in the target animal safety test, in which 80 broiler chicks (14 days of age) were randomly divided into four major groups (I-healthy control group; II-1× RD; III-3× RD; IV-6× RD), with each group subdivided into two subgroups (A and B) consisting of 10 chicks each. After 7-day (for sub-group A) or 14-day (for sub-group B) administration, compared with the healthy control, treatment-related changes in BWG, feed conversion ratio (FCR), relative organ weight (ROW) of the liver, WBC counts, and levels of RBC, HGB, ALT, AST, and TBIL were detected in the 3× and 6× RD groups. No differences were noted in necropsy for all doses, and histopathological examinations exhibited no QHP-associated signs of toxicity or abnormalities in the liver or kidney. The findings suggest that QHP at a dose of 0.30 g/kg feed would be appropriate for therapy and intermittent treatment of E. tenella-infected chicks, the dosage in clinical applications should be set according to the recommended dose to ensure animal safety.

The filamentous fungus Aspergillus niger is widely exploited by the fermentation industry for the production of enzymes, particularly glucoamylase. Although a variety of genetic techniques have been successfully used in wild-type A. niger, the transformation of industrially used strains with few conidia (e.g., A. niger N1) or that are even aconidial (e.g., A. niger O1) remains laborious. Herein, we developed genetic tools, including the protoplast-mediated transformation and Agrobacterium tumefaciens-mediated transformation of the A. niger strains N1 and O1 using green fluorescent protein as a reporter marker. Following the optimization of various factors for protoplast release from mycelium, the protoplast-mediated transformation efficiency reached 89.3% (25/28) for N1 and 82.1% (32/39) for O1. The A. tumefaciens-mediated transformation efficiency was 98.2% (55/56) for N1 and 43.8% (28/64) for O1. We also developed a marker-free CRISPR/Cas9 genome editing system using an AMA1-based plasmid to express the Cas9 protein and sgRNA. Out of 22 transformants, 9 albA deletion mutants were constructed in the A. niger N1 background using the protoplast-mediated transformation method and the marker-free CRISPR/Cas9 system developed here. The genome editing methods improved here will accelerate the elucidation of the mechanism of glucoamylase hyperproduction in these industrial fungi and will contribute to the use of efficient targeted mutation in other industrial strains of A. niger.

Epigenetic regulation is well recognized for its importance in gene expression in organisms. DNA methylation, an important epigenetic mark, has received enormous attention in recent years as it's a key player in many biological processes. It remains unclear how DNA methylation contributes to gene transcription regulation in maize seeds. Here, we take advantage of recent technologies to examine the genome-wide association of DNA methylation with transcription of four types of DNA sequences, including protein-coding genes, pseudogenes, transposable elements, and repeats in maize embryo and endosperm, respectively.The methylation in CG, CHG and CHH contexts plays different roles in the control of gene expression. Methylation around the transcription start sites and transcription stop regions of protein-coding genes is negatively correlated, but in gene bodies positively correlated, to gene expression level. The upstream regions of protein-coding genes are enriched with 24-nt siRNAs and contain high levels of CHH methylation, which is correlated to gene expression level. The analysis of sequence content within CG, CHG, or CHH contexts reveals that only CHH methylation is affected by its local sequences, which is different from Arabidopsis.In summary, we conclude that methylation-regulated transcription varies with the types of DNA sequences, sequence contexts or parts of a specific gene in maize seeds and differs from that in other plant species. Our study helps people better understand from a genome-wide viewpoint that how transcriptional expression is controlled by DNA methylation, one of the important factors influencing transcription, and how the methylation is associated with small RNAs.

Interactions among metabolic pathways are important in plant biology. At present, not much is known about how folate metabolism affects other metabolic pathways in plants. Here we report a T-DNA insertion mutant (atdfb-3) of the plastidial folylpolyglutamate synthetase gene (AtDFB) was defective in seed reserves and skotomorphogenesis. Lower carbon (C) and higher nitrogen (N) content in the mutant seeds than that of the wild type were indicative of an altered C and N partitioning capacity. Higher levels of organic acids and sugars were detected in the mutant seeds compared with the wild type. Further analysis revealed that atdfb-3 seeds contained less total amino acids and individual Asn and Glu as well as NO3-. These results indicate significant changes in seed storage in the mutant. Defects in hypocotyl elongation were observed in atdfb-3 in darkness under sufficient NO3- conditions, and further enhanced under NO3- limited conditions. The strong expression of AtDFB in cotyledons and hypocotyl during early developmental stage was consistent with the mutant sensitivity to limited NO3- during a narrow developmental window. Exogenous 5-formyl-tetrahydrofolate completely restored the hypocotyl length in atdfb-3 seedlings with NO3- as the sole N source. Further study demonstrated that folate profiling and N metabolism were perturbed in atdfb-3 etiolated seedlings. The activity of enzymes involved in N reduction and assimilation was altered in atdfb-3. Taken together, these results indicate that AtDFB is required for seed reserves, hypocotyl elongation and N metabolism in darkness, providing novel insights into potential associations of folate metabolism with seed reserve accumulation, N metabolism and hypocotyl development in Arabidopsis.

Novel pleuromutilin derivatives designed based on the structure of valnemulin were synthesized and evaluated for their in vitro antibacterial activities. These pleuromutilin derivatives with substituted amino moiety exhibited excellent activities against methicillin-resistant Staphylococcus aureus, methicillin-resistant Staphylococcus epidermidis, Escherichia coli, and Streptococcus agalactiae. Compound 5b showed the highest antibacterial activities and even exceeded tiamulin. Moreover, the docking experiments provided information about the binding model between the synthesized compounds and peptidyl transferase center (PTC) of 23S rRNA.

The study aimed to investigate the effects of Hypericum perforatum extract on the endocrine immune network factors in the immunosuppressed rat. Wistar rats were used to establish an immunosuppression model. Immunosuppressed rats received Hypericum perforatum extract suspension for 7 days at doses of 600 mg/kg (containing hyperforin at 7.2mg/kg).Endocrine immune network factors including IL-1, DA, NE, and 5-HT were determined on day 8.The results indicated that Hypericum perforatum extract affects the endocrine immune network through decreasing endogenous IL-1,dopamine (DA), norepinephrine (NE), and 5- hydroxytryptamine (5-HT) levels. Hyperforin reverses immune suppression through enhancing DA levels to inhibit B cells while promoting T cell and macrophage functions to recover immune function. [ABSTRACT FROM AUTHOR]

Background: Maize is a major staple food crop globally and contains various concentrations of vitamins. Folates are essential water-soluble B-vitamins that play an important role as one-carbon (C1) donors and acceptors in organisms. To gain an understanding of folate metabolism in maize, we performed an intensive in silico analysis to screen for genes involved in folate metabolism using publicly available databases, followed by examination of the transcript expression patterns and profiling of the folate derivatives in the kernels of two maize inbred lines. Results: A total of 36 candidate genes corresponding to 16 folate metabolism-related enzymes were identified. The maize genome contains all the enzymes required for folate and C1 metabolism, characterized by highly conserved functional domains across all the other species investigated. Phylogenetic analysis revealed that these enzymes in maize are conserved throughout evolution and have a high level of similarity with those in sorghum and millet. The LC-MS analyses of two maize inbred lines demonstrated that 5-methyltetrahydrofolate was the major form of folate derivative in young seeds, while 5-formyltetrahydrofolate in mature seeds. Most of the genes involved in folate and C1 metabolism exhibited similar transcriptional expression patterns between these two maize lines, with the highest transcript abundance detected on day after pollination (DAP) 6 and the decreased transcript abundance on DAP 12 and 18. Compared with the seeds on DAP 30, 5-methyltetrahydrofolate was decreased and 5-formyltetrahydrofolate was increased sharply in the mature dry seeds. Conclusions: The enzymes involved in folate and C1 metabolism are conserved between maize and other plant species. Folate and C1 metabolism is active in young developing maize seeds at transcriptional levels. [ABSTRACT FROM AUTHOR]

Investigations into the biochemical processes and regulatory mechanisms of nitrogen (N) utilization can aid in understanding how N is used efficiently in plants. This report describes a deficiency in N utilization in an Arabidopsis (Arabidopsis thaliana) transfer DNA insertion mutant of the mitochondrial folylpolyglutamate synthetase gene DFC, which catalyzes the conjugation of glutamate residues to the tetrahydrofolate during folate synthesis. The mutant seedlings displayed several metabolic changes that are typical of plant responses to low-N stress, including increased levels of starch and anthocyanin synthesis as well as decreased levels of soluble protein and free amino acid, as compared with those in wild-type seedlings when external N was sufficient. More striking changes were observed when dfc seedlings were grown under N-limited conditions, including shorter primary roots, fewer lateral roots, higher levels of glycine and carbon-N ratios, and lower N content than those in wild-type seedlings. Gene expression studies in mutant seedlings revealed altered transcript levels of several genes involved in folate biosynthesis and N metabolism. The biochemical and metabolic changes also suggested that N assimilation is drastically perturbed due to a loss of DFC function. The observation that elevated CO2 partly rescued the dfc phenotypes suggests that the alterations in N metabolism in dfc may be mainly due to a defect in photorespiration. These results indicate that DFC is required for N utilization in Arabidopsis and provide new insight into a potential interaction between folate and N metabolism. [ABSTRACT FROM AUTHOR]

The herbal Dichroa febrifuga Lour is llie dried roots of Saxifragaceae plaiil species Dichroa febrifuga Lour llial are endowed with llie efficacies of inducing vomiting, insecticide, anli-malaria and relieving heat. It has been broadly applied for treating malaria in folk. We reviewed the advances in pharmacological activity of effective component febrifugine in Dichroa febrifuga Lour and its derivatives, and found that Dichroa febrifuga Lour has been documented effective in antitumor, anti-coccidium, anti-scar, anti-malaria, and the treatment of diabetes mel-litus and osteoporosis as well. On the basis of this, we proposed to develop safe and efficient derivatives continuously by structural modification and perform immune study and their antitumor activity. [ABSTRACT FROM AUTHOR]