[Optimization of processing technology of bark of Ilicis Rotundae Cortex based on quality by design concept].

Based on the concept of quality by design(QbD), this study optimized the processing technology of Ilicis Rotundae Cortex. According to the processing method and ingredient requirements of Ilicis Rotundae Cortex in the Chinese Pharmacopoeia, the content of syringin and pedunculoside, alcohol extract, fragmentation rate, and moisture content were taken as the critical quality attributes(CQAs). The soaking time, moistening time, and drying time were taken as critical process parameters(CPPs) by single factor tests. The weight coefficients of CQAs were determined by the analytic hierarchy process(AHP)-entropy weighting method, and the comprehensive score was calculated. With the comprehensive score as the response value, Box-Behnken design was employed to establish a mathematical model between CPPs and CQAs, and the design space for the processing of Ilicis Rotundae Cortex was built and verified. The results of ANOVA showed that the mathematical model had the P value below 0.05, the lack of fit greater than 0.05, adjusted R~2=0.910 5, and predicted R~2=0.831 0, which indicated that the proposed model had statistical significance and good prediction performance. Considering the factors in production, the best processing conditions of Ilicis Rotundae Cortex were decoction pieces of about 1 cm soaking for 1 h, moistening for 4 h, and drying at 60-70 ℃ in a blast drier for 2 h. The optimized processing technology of Ilicis Rotundae Cortex was stable and feasible, which can provide a reference for the standardized preparation and stable quality of Ilicis Rotundae Cortex.

Functional divergence of CYP76AKs shapes the chemodiversity of abietane-type diterpenoids in genus Salvia.

The genus Salvia L. (Lamiaceae) comprises myriad distinct medicinal herbs, with terpenoids as one of their major active chemical groups. Abietane-type diterpenoids (ATDs), such as tanshinones and carnosic acids, are specific to Salvia and exhibit taxonomic chemical diversity among lineages. To elucidate how ATD chemical diversity evolved, we carried out large-scale metabolic and phylogenetic analyses of 71 Salvia species, combined with enzyme function, ancestral sequence and chemical trait reconstruction, and comparative genomics experiments. This integrated approach showed that the lineage-wide ATD diversities in Salvia were induced by differences in the oxidation of the terpenoid skeleton at C-20, which was caused by the functional divergence of the cytochrome P450 subfamily CYP76AK. These findings present a unique pattern of chemical diversity in plants that was shaped by the loss of enzyme activity and associated catalytic pathways.

Antagonism of Rhizosphere Streptomyces yangpuensis CM253 against the Pathogenic Fungi Causing Corm Rot in Saffron (Crocus sativus L.).

Plant diseases lead to a significant decline in the output and quality of Chinese herbal medicines. Actinomycetes play a vital role in the rhizosphere ecosystem. This is especially true for Streptomyces, which have become a valuable biological control resource because of their advantages in producing various secondary metabolites with novel structures and remarkable biological activities. The purpose of this study was to isolate an effective antagonistic actinomycete against the pathogen of corm rot in saffron. An antagonistic actinomycete, CM253, was screened from the rhizosphere soil samples of Crocus sativus, by plate co-culture with four pathogenic fungi (Fusarium oxysporum, Fusarium solani, Penicillium citreosulfuratum, and Penicillium citrinum). CM253 inhibited the growth and development of F. oxysporum hyphae by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Furthermore, by analyzing the degrading enzyme, the growth-promoting performance, and the whole genome of strain CM253, it was identified as Streptomyces yangpuensis, which produces NH3, protease, glucanase, cellulase, IAA, and ACC deaminase. In addition, 24 secondary metabolite synthesis gene clusters were predicted in antiSMASH. We identified genes encoding 2,3-butanediol; methionine; isoprene (metH, mmuM, ispEFH, gcpE, idi, and ilvABCDEH); biofilm formation; and colonization (upp, rfbBC, efp, aftA, pssA, pilD, fliA, and dhaM). Above all, S. yangpuensis CM253 showed the potential for future development as a biocontrol agent.

Clinical Efficacy of Integrated Traditional Chinese and Western Medicine in the Treatment of Eczema: A Meta-Analysis.

Background: Both traditional Chinese medicine (TCM) and Western medicine are widely applied in the treatment of eczema, but there are few reports on integrated TCM and Western medicine for eczema. Objective: This meta-analysis carried out the evaluation on the efficacy of integrated TCM and Western medicine in the treatment of eczema. Methods: PubMed, Web of Science, Embase, CNKI, and Wanfang databases were searched for all possible randomized controlled trials from 2000 to 2021. A meta-analysis of the included studies was also performed using Stata16 software. Results: A total of 16 studies including 1946 patients were included. Compared with the control group, the pooled results of the 16 studies showed that effective rate in the treatment group was higher (OR = 4.50, 95% CI: 3.16-6.40, P < 0.05), and the pooled data of 15 studies revealed that the cure rate was increased in the treatment group (OR = 2.60, 95% CI: 2.13-3.18, P < 0.05). Additionally, compared with the control group, pooled data of 11 studies demonstrated that lesion area after treatment was reduced in the treatment group (SMD = -1.91; 95% CI: -2.51, -1.31; P < 0.05), and pooled data of 9 studies showed that the degree of pruritus after treatment in the treatment group was lower (SMD = -1.69; 95% CI: -2.07, -1.30; P < 0.05). Conclusion: In comparison with Western medicine alone, integrated TCM and Western medicine are a more effective treatment for eczema, which can not only significantly improve the effective rate and cure rate but also reduce the lesion area and degree of pruritus.

Adsorption properties and influencing factors of Cu(II) on polystyrene and polyethylene terephthalate microplastics in seawater.

As an emerging contamination in the ocean, microplastics can act as effective vectors of pollutants, the ecological risks caused by the combined pollution of microplastics and other pollutants have attracted growing attention. In this work, Copper (Cu(II)) was chosen as the classic pollutant, polystyrene (PS) and polyethylene terephthalate (PET) pellets were used as the typical marine microplastics, the adsorption performance of Cu(II) on PS and PET beads was investigated by adsorption kinetics and isotherm experiments, and other influencing conditions, such as pH, salinity, coexisting heavy metals ions and aging treatment, were evaluated. The results indicated that the adsorption behavior of Cu(II) on PS and PET was spontaneous and endothermic in the simulated seawater environment, and the batch experimental data can be effectively described by pseudo-second-order model and Freundlich isothermal model. Besides, the adsorption capacity of microplastics for Cu(II) was the best at pH 7, the change of salinity had no obvious effect on the adsorption in the natural marine environment. Moreover, co-existence of lead (Pb(II)) exhibited evident impacts on Cu(II) sorption onto PS and PET, which confirmed the adsorption competition effect between them. Additionally, high temperature aging treatment of microplastics in different environments for different duration time could obviously affect the properties of microplastics. It was found that the microplastics after being exposed to high temperature environment in the air for 168 h showed relatively stronger adsorption amount for Cu(II). In summary, these findings suggested that electrostatic interaction and distributed diffusion mechanisms may be the main mechanisms of adsorption, while no new functional groups were generated after the adsorption, indicating that physisorption may dominate the adsorption performance of PS and PET pellets for Cu(II). This study provides supplementary insights into the role of microplastics as carriers of heavy metals in the marine environment.

First Report of Black Spot Caused by Penicillium citreosulfuratum on Saffron in Chongming Island, China.

Saffron (Crocus sativus L.) is a world-famous source of dye and spices and an important medicinal plant, which is cultivated on a large scale on Chongming Island (31.62° N, 121.39° E) in Shanghai, China. In August 2020, a survey of saffron was conducted in this area, and black spots were observed on about 10% of plants. Characteristics of the disease were: The bottom of the corm was darkened in the groove and scattered black spots could be observed after peeling off the membranous scale leaves. The junction of lesions and healthy parts was light brown. As the lesions expanded, approximately 80% of the surface of the corm became dark brown to bluish gray. The inside of the corm was also necrotic. In order to isolate the pathogen, ten diseased corms with typical symptoms were selected. All corms were first treated with 75% ethanol for 30 s, 0.1% HgCl2 for 5 min, and then rinsed with sterile water 5 times. Next, tissue pieces (5 mm × 5 mm) at the margin of lesions were cut out and placed on the potato dextrose agar (PDA) medium. After incubating at 28°C for 5 days, fungi were separated and purified by using the hyphal-tip technique. A total of six pure cultures with different colony morphologies were obtained, of which only the isolate MF3 was present in ten diseased corms. The isolate MF3 was inoculated on the PDA and cultured at 28°C for 10 days and characteristics of the fungus colonies were: colonies sub-circular, reaching 28 to 30 mm diam, from above rough, dense, fluffy, blue-gray, with some white spots and central point raised, and outer margin form an irregular, narrow, white ring; from below, yellow with light-green. The hyphae were slender, with many septa. The conidiophores were typically smooth walled, short, and slender and either monoverticillate with very short stipes or as irregularly biverticillate. Phialides were ampulliform, 5 to 10 per metula, 5 to 8 × 2 to 3 µm. Conidia were smooth and globose, and ranged in diameter from 1.4 to 1.7 µm (n=50). Molecular identification of the fungus was made by PCR amplification of the internal transcribed spacer (ITS) region of rDNA and ß-tubulin gene using primers ITS1/ITS4 (White et al. 1990), TUB2Fd/TUB4Rd (Aveskamp et al. 2009) respectively. The ITS (MW881446) and ß-tubulin (MW911464) sequences of the fungus were similar to the ITS (MN592912) and ß-tubulin (KY469126) sequences of the epitype of Penicillium citreosulfuratum with 99.81% and 99.56% identity, respectively. According to the morphological and molecular characterization, the isolate MF3 was identified as P. citreosulfuratum (Visagie et al. 2016). For pathogenicity testing, the fungus was grown on PDA and incubated at 28°C for 5 days. Then mycelial plugs (5 mm diam.) were inoculated on the scalpel incision square wounds of surface-disinfected corms and mock-inoculated corms received only PDA plugs. Corms were placed in sterile plastic bottles and observed after culturing at 28°C for 21 days. Each treatment had three replicates and the experiment was repeated twice. The results showed that corms inoculated with P. citreosulfuratum developed diseased with similar symptoms as in the field. No disease symptoms were observed on control corms. Re-isolations were performed from inoculated corms, and all re-isolated fungi were confirmed as P. citreosulfuratum, verifying the fungus as the pathogen based on Koch's postulates. To our knowledge, this is the first report of the pathogen causing black spot disease of saffron. Although the disease is not fatal to saffron, to a certain extent it will cause a reduction in the production of the crop. In addition, this pathogen has not been reported to be pathogentic to other plant species.

Effects of BMP9 and pulsed electromagnetic fields on the proliferation and osteogenic differentiation of human periodontal ligament stem cells.

Periodontal ligament stem cells (PDLSCs) have been confirmed to have self-renewal capacity and multidifferentiation potential and are good candidates for periodontal tissue regeneration. Pulsed electromagnetic field (PEMF) has been demonstrated to promote osteogenesis in non-union fractures, partly by regulating mesenchymal stem cells or osteoblast activity. However, there is no report about the osteo-inductive effect of PEMF stimulation on human PDLSCs (hPDLSCs). Thus, we tested the hypothesis that PEMF biophysical stimulation alone has an influence on the proliferation and osteogenic differentiation of hPDLSCs. To detect the osteo-inductive potential of bone morphogenetic protein (BMP9), we transfected the STRO-1+ /CD146+ hPDLCSs with BMP9-expressing recombinant adenoviruses. We examined the proliferation and osteogenic differentiation of hPDLSCs treated with either PEMF (15 Hz, 1 h daily, different intensities), or BMP9, or both stimuli. Cell counting kit-8 (CCK-8) assay showed that PEMF of different intensities had no effect on the proliferation of hPDLSCs and did not enhance the proliferative capability of BMP9-transfected cells. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting showed that the combination of both PEMFs (1.8 or 2.4 mT) and BMP9 stimulation had a synergistic effect on early and intermediate osteogenic genes and protein expressions of runt-related transcription factor 2, alkaline phosphatase, osteopontin, and late mineralized extracellular matrix formation in hPDLSCs. Bioelectromagnetics. 38:63-77, 2017. © 2016 Wiley Periodicals, Inc.

[Purification technology of manninotirose in Rehmanniae Radix Praeparata by D-101 microporous adsorption resin].

This paper was aim to optimize the purification technology of Rehmanniae Radix Praeparata extract with macroporous adsorption resin. With the content of manninotriose as index, the absorptive flow and time were investigated, as well as kinds, amount, flow of eluent. D-101 type macroporous adsorption resin was the best choice for the purification of manninotriose. The optimized parameters were as follows: the content of manninotriose at 161.16-53.72 mg x g(-1), absorption time 240 min, eluting solvent of purified water, volume flow at 1.5 BV x h(-1), and eluant volume at 6 BV. D-101 type macroporous adsorption resin could significantly increase the purity of Rehmanniae Radix Praeparata extract with the advantage of high absorption, remove most part of impurity, and the effect of semi-works production was better.

Reporting patterns of adverse drug reactions over recent years in China: analysis from publications.

PURPOSE: The goal of this study was to clarify the reporting patterns of self-reported adverse drug reactions (ADRs) in China. METHODS: A variety of sources were searched, including the official website of China FDA, the national center for ADR monitoring center, publications from PubMed, and so on. We retrieved the relevant information and made descriptive and comparative analysis from the year 2009 to 2013. RESULTS: The ADR reporting numbers were 638,996, 692,904, 852,799, 1,200,000 and 1,317,000 from 2009 to 2013, respectively. Healthcare professionals contributed significantly, and their proportion always exceeded 80% before 2012. The average report per million inhabitants has increased from 479 to 983 from 2009 to 2013. However, the proportion of new or serious report was always below 25%. The reports mainly concern anti-infective agents and traditional Chinese medicine (TCM), especially TCM injection. The proportion of ADR reports in geriatric patients has increased for 4 consecutive years. CONCLUSIONS: ADR report numbers and reporting rates in China are on the rise. However, the proportion of new or serious reports as well as the proportion of reports contributed by consumers and pharmaceutical companies are still quite low. More attention should be paid to the elderly, anti-infective agents and TCM, especially TCM injections.

[Data validation methods and discussion on Chinese materia medica resource survey].

From the beginning of the fourth national survey of the Chinese materia medica resources, there were 22 provinces have conducted pilots. The survey teams have reported immense data, it put forward the very high request to the database system construction. In order to ensure the quality, it is necessary to check and validate the data in database system. Data validation is important methods to ensure the validity, integrity and accuracy of census data. This paper comprehensively introduce the data validation system of the fourth national survey of the Chinese materia medica resources database system, and further improve the design idea and programs of data validation. The purpose of this study is to promote the survey work smoothly.

Unraveling the genetic basis of seed tocopherol content and composition in rapeseed (Brassica napus L.).

BACKGROUND: Tocopherols are important antioxidants in vegetable oils; when present as vitamin E, tocopherols are an essential nutrient for humans and livestock. Rapeseed (Brassica napus L, AACC, 2 n = 38) is one of the most important oil crops and a major source of tocopherols. Although the tocopherol biosynthetic pathway has been well elucidated in the model photosynthetic organisms Arabidopsis thaliana and Synechocystis sp. PCC6803, knowledge about the genetic basis of tocopherol biosynthesis in seeds of rapeseed is scant. This project was carried out to dissect the genetic basis of seed tocopherol content and composition in rapeseed through quantitative trait loci (QTL) detection, genome-wide association analysis, and homologous gene mapping. METHODOLOGY/PRINCIPAL FINDINGS: We used a segregating Tapidor × Ningyou7 doubled haploid (TNDH) population, its reconstructed F(2) (RC-F(2)) population, and a panel of 142 rapeseed accessions (association panel). Genetic effects mainly contributed to phenotypic variations in tocopherol content and composition; environmental effects were also identified. Thirty-three unique QTL were detected for tocopherol content and composition in TNDH and RC-F(2) populations. Of these, seven QTL co-localized with candidate sequences associated with tocopherol biosynthesis through in silico and linkage mapping. Several near-isogenic lines carrying introgressions from the parent with higher tocopherol content showed highly increased tocopherol content compared with the recurrent parent. Genome-wide association analysis was performed with 142 B. napus accessions. Sixty-one loci were significantly associated with tocopherol content and composition, 11 of which were localized within the confidence intervals of tocopherol QTL. CONCLUSIONS/SIGNIFICANCE: This joint QTL, candidate gene, and association mapping study sheds light on the genetic basis of seed tocopherol biosynthesis in rapeseed. The sequences presented here may be used for marker-assisted selection of oilseed rape lines with superior tocopherol content and composition.

Codon optimization enhances secretory expression of Pseudomonas aeruginosa exotoxin A in E. coli.

Pseudomonas aeruginosa exotoxin A (PEA) is a number of family of bacterial ADP-ribosylating toxins and possesses strong immunogenicity. The detoxified exotoxin A, as a potent vaccine adjuvant and vaccine carrier protein, has been extensively used in human and animal vaccinations. However, the expression level of PEA gene in Escherichia coli is relative low which is likely due to the presence of rare codon and high levels of GC content. In order to enhance PEA gene expression, we optimized PEA gene using E. coli preferred codons and expressed it in E. coli BL21 (DE3) by using pET-20b(+) secretory expression vector. Our results showed that codon optimization significantly reduced GC content and enhanced PEA gene expression (70% increase compared with that of the wild-type). Moreover, the codon-optimized PEA possessed biological activity and had the similar toxic effects on mouse L292 cells compared with the wild-type PEA gene. Codon optimization will not only improve PEA gene expression but also benefit further modification of PEA gene using nucleotide-mediated site-directed mutagenesis. A large number of purified PEA proteins will provide the necessary conditions for further PEA functional research and application.