Adhesive injectable cellulose-based hydrogels with rapid self-healing and sustained drug release capability for promoting wound healing.

Injectable biocompatible hydrogels with multiple functions, including self-healing, adhesion, antibacterial activity, and suitable mechanical properties, are highly desirable for enhancing wound healing. In this study, a new class of multi-functional injectable self-healing cellulose-based hydrogels was synthesised using dynamic covalent acylhydrazone linkages for wound dressing. The carboxymethyl cellulose-graft-adipic dihydrazide (CMC-ADH)/4-Formylbenzoic acid-terminated poly(ethylene glycol) (PEG-FBA) (CMC-ADH/PEG-FBA) hydrogels have adjustable gelation time and excellent self-healing ability. In addition, drug release and in vitro antibacterial activities against Gram-positive and Gram-negative bacteria confirmed the sustained drug-release capacity of the hydrogels. Moreover, haemostasis and wound-healing effects were investigated using an in vivo haemorrhaging liver mouse model and a full-thickness skin defect model, and the results indicated that they not only promoted the wound-healing process but also presented excellent haemostatic effects. The CMC-ADH/PEG-FBA gels also exhibited good adhesion to irregular wounds and significantly enhanced angiogenic ability in vivo. This excellent wound-healing performance occurs because hydrogels can quickly stop bleeding, provide a moist and closed environment for the wound to prevent bacterial invasion, release ciprofloxacin (CIP), reduce inflammatory reactions, and promote wound tissue regeneration. In summary, the synthesised multi-functional gels are ideal candidates for treating haemorrhages and irregular wounds.

Stretchable and conductive cellulose hydrogel electrolytes for flexible and foldable solid-state supercapacitors.

In this study, the anti-freezing conductive hydrogel electrolytes with outstanding mechanical properties were synthesized by a facile and feasible method. The mechanical and anti-freezing properties of the synthesized polyacrylamide/lithium lhloride/water soluble cellulose acetate (PAM/LiCl/WSCA) hydrogels are significantly enhanced with the addition of WSCA and LiCl. The tensile strength and toughness of the gels were gradually increased to 341 KPa and 1.2 MJ/m3, respectively. The hydrogel electrolyte can remain soft and flexible at -80 °C, displaying certain elasticity and electrical conductivity. In addition, the super-capacitor assembled with PAM/LiCl/WSCA hydrogel as electrolyte showed excellent stability in capacitance retention after 500 times of folding cycles and 10,000 times of charge and discharge tests. The capacitor still maintains 64.64 % of its capacity at -40 °C. This facile strategy to fabricate anti-freezing conductive hydrogel electrolyte provides a new idea and way to the application of hydrogels as electrolytes in extreme cold environments.

New Delivery Route of Gambogic Acid Via Skin for Topical Targeted Therapy of Cutaneous Melanoma and Reduction of Systemic Toxicity.

Cutaneous melanoma is the deadliest form of skin cancer, and gambogic acid (GA) exhibits potent anti-melanoma activity. However, clinical application of GA via intravenous injection and oral administration is limited by systemic toxicity and rapid metabolism in the blood. Here, we developed a new, topical route of GA delivery for anti-melanoma activity and reduction of systemic toxicity. The results indicated that the barrier of the stratum corneum (SC) and low diffusion of GA in the hydrophilic viable skin (epidermis and dermis) limited the GA penetration through intact skin. The combination of azone (AZ) and propylene glycol (PG) showed obvious synergistic effects on skin penetration by GA via improving the permeability of the SC and greatly increasing the skin accumulation of GA, thereby forming a high drug concentration in the skin and achieving a topical targeted treatment of melanoma. In addition, GA (AZ-PG) achieved the same anti-melanoma effect via topical delivery as via intravenous injection. Intravenous injection and oral administration of GA induced remarkable pathological changes in various organs in mice, whereas GA was not toxic to various organs or to the skin via topical delivery. These findings indicated that topical administration of GA is an alternative route for melanoma treatment.

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.

Identification of Major Rhizobacterial Taxa Affected by a Glyphosate-Tolerant Soybean Line via Shotgun Metagenomic Approach.

The worldwide commercial cultivation of transgenic crops, including glyphosate-tolerant (GT) soybeans, has increased widely during the past 20 years. However, it is accompanied with a growing concern about potential effects of transgenic crops on the soil microbial communities, especially on rhizosphere bacterial communities. Our previous study found that the GT soybean line NZL06-698 (N698) significantly affected rhizosphere bacteria, including some unidentified taxa, through 16S rRNA gene (16S rDNA) V4 region amplicon deep sequencing via Illumina MiSeq. In this study, we performed 16S rDNA V5-V7 region amplicon deep sequencing via Illumina MiSeq and shotgun metagenomic approaches to identify those major taxa. Results of these processes revealed that the species richness and evenness increased in the rhizosphere bacterial communities of N698, the beta diversity of the rhizosphere bacterial communities of N698 was affected, and that certain dominant bacterial phyla and genera were related to N698 compared with its control cultivar Mengdou12. Consistent with our previous findings, this study showed that N698 affects the rhizosphere bacterial communities. In specific, N698 negatively affects Rahnella, Janthinobacterium, Stenotrophomonas, Sphingomonas and Luteibacter while positively affecting Arthrobacter, Bradyrhizobium, Ramlibacter and Nitrospira.

Glyphosate application increased catabolic activity of gram-negative bacteria but impaired soil fungal community.

Glyphosate is a non-selective organophosphate herbicide that is widely used in agriculture, but its effects on soil microbial communities are highly variable and often contradictory, especially for high dose applications. We applied glyphosate at two rates: the recommended rate of 50 mg active ingredient kg-1 soil and 10-fold this rate to simulate multiple glyphosate applications during a growing season. After 6 months, we investigated the effects on the composition of soil microbial community, the catabolic activity and the genetic diversity of the bacterial community using phospholipid fatty acids (PLFAs), community level catabolic profiles (CLCPs), and 16S rRNA denaturing gradient gel electrophoresis (DGGE). Microbial biomass carbon (Cmic) was reduced by 45%, and the numbers of the cultivable bacteria and fungi were decreased by 84 and 63%, respectively, under the higher glyphosate application rate. According to the PLFA analysis, the fungal biomass was reduced by 29% under both application rates. However, the CLCPs showed that the catabolic activity of the gram-negative (G-) bacterial community was significantly increased under the high glyphosate application rate. Furthermore, the DGGE analysis indicated that the bacterial community in the soil that had received the high glyphosate application rate was dominated by G- bacteria. Real-time PCR results suggested that copies of the glyphosate tolerance gene (EPSPS) increased significantly in the treatment with the high glyphosate application rate. Our results indicated that fungi were impaired through glyphosate while G- bacteria played an important role in the tolerance of microbiota to glyphosate applications.

Impact of Glyphosate on the Rhizosphere Microbial Communities of An EPSPS-Transgenic Soybean Line ZUTS31 by Metagenome Sequencing.

BACKGROUND: The worldwide use of glyphosate has dramatically increased, but also has been raising concern over its impact on mineral nutrition, plant pathogen, and soil microbiota. To date, the bulk of previous studies still have shown different results on the effect of glyphosate application on soil rhizosphere microbial communities. OBJECTIVE: This study aimed to clarify whether glyphosate has impact on nitrogen-fixation, pathogen or disease suppression, and rhizosphere microbial community of a soybean EPSPS-transgenic line ZUTS31 in one growth season. METHOD: Comparative analysis of the soil rhizosphere microbial communities was performed by 16S rRNA gene amplicons sequencing and shotgun metagenome sequencing analysis between the soybean line ZUTS31 foliar sprayed with diluted glyphosate solution and those sprayed with water only in seed-filling stage. RESULTS: There were no significant differences of alpha diversity but with small and insignificant difference of beta diversity of soybean rhizosphere bacteria after glyphosate treatment. The significantly enriched Gene Ontology (GO) terms were cellular, metabolic, and single-organism of biological process together with binding, catalytic activity of molecular function. The hits and gene abundances of some functional genes being involved in Plant Growth-Promoting Traits (PGPT), especially most of nitrogen fixation genes, significantly decreased in the rhizosphere after glyphosate treatment. CONCLUSION: Our present study indicated that the formulation of glyphosate-isopropylamine salt did not significantly affect the alpha and beta diversity of the rhizobacterial community of the soybean line ZUTS31, whereas it significantly influenced some functional genes involved in PGPT in the rhizosphere during the single growth season.

Involvement of LeMDR, an ATP-binding cassette protein gene, in shikonin transport and biosynthesis in Lithospermum erythrorhizon.

BACKGROUND: Shikonin is a naphthoquinone secondary metabolite with important medicinal value and is found in Lithospermum erythrorhizon. Considering the limited knowledge on the membrane transport mechanism of shikonin, this study investigated such molecular mechanism. RESULTS: We successfully isolated an ATP-binding cassette protein gene, LeMDR, from L. erythrorhizon. LeMDR is predominantly expressed in L. erythrorhizon roots, where shikonin accumulated. Functional analysis of LeMDR by using the yeast cell expression system revealed that LeMDR is possibly involved in the shikonin efflux transport. The accumulation of shikonin is lower in yeast cells transformed with LeMDR-overexpressing vector than that with empty vector. The transgenic hairy roots of L. erythrorhizon overexpressing LeMDR (MDRO) significantly enhanced shikonin production, whereas the RNA interference of LeMDR (MDRi) displayed a reverse trend. Moreover, the mRNA expression level of LeMDR was up-regulated by treatment with shikonin and shikonin-positive regulators, methyl jasmonate and indole-3-acetic acid. There might be a relationship of mutual regulation between the expression level of LeMDR and shikonin biosynthesis. CONCLUSIONS: Our findings demonstrated the important role of LeMDR in transmembrane transport and biosynthesis of shikonin.

Antiviral activity of a synthesized shikonin ester against influenza A (H1N1) virus and insights into its mechanism.

This study aimed to examine the antiviral effects of shikonin ester ((R)-1-(5, 8-dihydroxy-1,4-dioxo-1,4-dihydronaphthalen-2-yl)-4-methylpent-3-en-1-yl3-(1H- indol-3-yl) propanoate (PMM-034) against influenza A (H1N1) virus. We investigated PMM-034 anti-H1N1 activity and its effect on caspase 3 gene expression during cellular apoptosis after influenza virus infection in vitro. Neuraminidase (NA) inhibition was assessed in comparison with oseltamivir in the influenza virus standard strains A/PR/8/34 to understand the viral mechanism. MDCK and A549 cells were used to investigate influenza viral infection and the structure-activity relationship between PMM-034 and NA was evaluated by pharmacophore-based docking modeling. The production of viral protein was tested by western blot. A/PR/8/34 induced cell inhibition but this was reduced by PMM-034 to 16µg/mL and this showed a selective index of 10mM. PMM-034 inhibited NA in a dose dependent manner, similar to oseltamivir inhibition. A sharp decrease in viral nucleocapsid protein mRNA was observed in infected cells after treatment with PMM-034. Apoptosis of infected A459 cells was inhibited by PMM-034 with decreased caspase 3 levels. ARG 118, ARG 152, ARG 371 and GLU 227 in the binding pocket of NA bound to PMM-034 in the docking model. Taken together, these results suggest PMM-034 shikonin ester blocked H1N1 infection and might be a potential anti-H1N1 drug.

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.

Identification of New Shikonin Derivatives as Antitumor Agents Targeting STAT3 SH2 Domain.

Signal transducer and activator of transcription 3 (STAT3) is hyper-activated in diversiform human tumors and has been validated as an attractive therapeutic target. Current research showed that a natural product, shikonin, along with its synthetic analogues, is able to inhibit the activity of STAT3 potently. The potential space of shikonin in developing novel anti-cancer agents encouraged us to carry out the investigation of the probable binding mode with STAT3. From this foundation, we have designed new types of STAT3 SH2 inhibitors. Combined simulations were performed to filter for the lead compound, which was then substituted, synthesized and evaluated by a variety of bioassays. Among the entities, PMM-172 exhibited the best anti-proliferative activity against MDA-MB-231 cells with IC50 value 1.98 ± 0.49 µM. Besides, it was identified to decrease luciferase activity, induce cell apoptosis and reduce mitochondrial transmembrane potential in MDA-MB-231 cells. Also, PMM-172 inhibited constitutive/inducible STAT3 activation without affecting STAT1 and STAT5 in MDA-MB-231 cells, and had no effect in non-tumorigenic MCF-10A cells. Moreover, PMM-172 suppressed STAT3 nuclear localization and STAT3 downstream target genes expression. Overall, these results indicate that the antitumor activity of PMM-172 is at least partially due to inhibition of STAT3 in breast cancer cells.

Design, Synthesis, and Biological Evaluation of Chalcone-Containing Shikonin Derivatives as Inhibitors of Tubulin Polymerization.

The biological importance of microtubules in mitosis makes them an interesting target for the development of anticancer agents. In this study, a series of novel chalcone-containing shikonin derivatives was designed, synthesized, and evaluated for biological activities. Among them, derivative PMMB-259 [(R)-1-(5,8-dihydroxy-1,4-dioxo-1,4-dihydronaphthalen-2-yl)-4-methylpent-3-en-1-yl (E)-2-(4-(3-oxo-3-(3-(trifluoromethoxy)phenyl)prop-1-en-1-yl)phenoxy)acetate] was identified as a potent inhibitor of tubulin polymerization. Further investigation confirmed that PMMB-259 can induce MCF-7 cell apoptosis, reduce the mitochondrial transmembrane potential, and arrest the cell cycle at the G2 /M phase. Moreover, the morphological variation of treated cells was visualized by confocal microscopy. The results, along with docking simulations, further indicated that PMMB-259 can bind well to tubulin at the colchicine site. Overall, these studies may provide a new molecular scaffold for the further development of antitumor agents that target tubulin.

Design, synthesis, biological evaluation, and 3D-QSAR analysis of podophyllotoxin-dioxazole combination as tubulin targeting anticancer agents.

The advancement of cancer-fighting drugs has never been a simple linear process. Those drug design professionals begin to find inspiration from the nature after failing to find the ideal products by creative drug design and high-throughput screening. To obtain new molecules for inhibiting tubulin, podophyllotoxin was adopted as the leading compound and 1,3,4-oxadiazole was brought in to the C-4 site of podophyllotoxin in this research. A series of seventeen podophyllotoxin-derived esters have been achieved and then evaluated their antitumor activities against four different cancer cell lines: A549, MCF-7, HepG2, and HeLa. Among all the compounds, compound 7c showed the best antiproliferating properties with IC50  = 2.54 ± 0.82 µm against MCF-7 cancer cell line. It was obvious that the content of ROS grew significantly in MCF-7 in a way depending on the dosage. The time- and dose-dependent cell cycle assays revealed that compound 7c could apparently block cell cycle in the phase of G2/M along with the upregulation of cyclin A2 and CDK2 protein. According to further studies, confocal microscopy experiment has certified that compound 7c could restrain cancer from growing by blocking the polymerization of microtubule. Meanwhile, compound 7c could be ideally integrated with the colchicine site of tubulin. In future, it would be feasible to selectively design tubulin inhibitors with the help of 3D-QSAR. This means that it is hopeful to develop compound 7c as a potential agent against cancer due to its biological characteristics.

Impact of a Glyphosate-Tolerant Soybean Line on the Rhizobacteria, Revealed by Illumina MiSeq.

The global commercial cultivation of transgenic crops, including glyphosate-tolerant soybean, has increased widely in recent decades with potential impact on the environment. The bulk of previous studies showed different results on the effects of the release of transgenic plants on the soil microbial community, especially rhizosphere bacteria. In this study, comparative analyses of the bacterial communities in the rhizosphere soils and surrounding soils were performed between the glyphosate-tolerant soybean line NZL06-698 (or simply N698), containing a glyphosate-insensitive EPSPS gene, and its control cultivar Mengdou12 (or simply MD12), by a 16S ribosomal RNA gene (16S rDNA) amplicon sequencing-based Illumina MiSeq platform. No statistically significant difference was found in the overall alpha diversity of the rhizosphere bacterial communities, although the species richness and evenness of the bacteria increased in the rhizosphere of N698 compared with that of MD12. Some influence on phylogenetic diversity of the rhizosphere bacterial communities was found between N698 and MD12 by beta diversity analysis based on weighted UniFrac distance. Furthermore, the relative abundances of part rhizosphere bacterial phyla and genera, which included some nitrogen-fixing bacteria, were significantly different between N698 and MD12. Our present results indicate some impact of the glyphosate-tolerant soybean line N698 on the phylogenetic diversity of rhizosphere bacterial communities together with a significant difference in the relative abundances of part rhizosphere bacteria at different classification levels as compared with its control cultivar MD12, when a comparative analysis of surrounding soils between N698 and MD12 was used as a systematic contrast study.

Transgenic studies reveal the positive role of LeEIL-1 in regulating shikonin biosynthesis in Lithospermum erythrorhizon hairy roots.

BACKGROUND: The phytohormone ethylene (ET) is a key signaling molecule for inducing the biosynthesis of shikonin and its derivatives, which are secondary metabolites in Lithospermum erythrorhizon. Although ETHYLENE INSENSITIVE3 (EIN3)/EIN3-like proteins (EILs) are crucial transcription factors in ET signal transduction pathway, the possible function of EIN3/EIL1 in shikonin biosynthesis remains unknown. In this study, by targeting LeEIL-1 (L. erythrorhizon EIN3-like protein gene 1) at the expression level, we revealed the positive regulatory effect of LeEIL-1 on shikonin formation. RESULTS: The mRNA level of LeEIL-1 was significantly up-regulated and down-regulated in the LeEIL-1-overexpressing hairy root lines and LeEIL-1-RNAi hairy root lines, respectively. Specifically, LeEIL-1 overexpression resulted in increased transcript levels of the downstream gene of ET signal transduction pathway (LeERF-1) and a subset of genes for shikonin formation, excretion and/or transportation (LePAL, LeC4H-2, Le4CL-1, HMGR, LePGT-1, LeDI-2, and LePS-2), which was consistent with the enhanced shikonin contents in the LeEIL-1-overexpressing hairy root lines. Conversely, LeEIL-1-RNAi dramatically repressed the expression of the above genes and significantly reduced shikonin production. CONCLUSIONS: The results revealed that LeEIL-1 is a positive regulator of the biosynthesis of shikonin and its derivatives in L. erythrorhizon hairy roots. Our findings gave new insights into the molecular regulatory mechanism of ET in shikonin biosynthesis. LeEIL-1 could be a crucial target gene for the genetic engineering of shikonin biosynthesis.

Transgenic analysis reveals LeACS-1 as a positive regulator of ethylene-induced shikonin biosynthesis in Lithospermum erythrorhizon hairy roots.

The phytohormone ethylene (ET) is a crucial signaling molecule that induces the biosynthesis of shikonin and its derivatives in Lithospermum erythrorhizon shoot cultures. However, the molecular mechanism and the positive regulators involved in this physiological process are largely unknown. In this study, the function of LeACS-1, a key gene encoding the 1-aminocyclopropane-1-carboxylic acid synthase for ET biosynthesis in L. erythrorhizon hairy roots, was characterized by using overexpression and RNA interference (RNAi) strategies. The results showed that overexpression of LeACS-1 significantly increased endogenous ET concentration and shikonin production, consistent with the up-regulated genes involved in ET biosynthesis and transduction, as well as the genes related to shikonin biosynthesis. Conversely, RNAi of LeACS-1 effectively decreased endogenous ET concentration and shikonin production and down-regulated the expression level of above genes. Correlation analysis showed a significant positive linear relationship between ET concentration and shikonin production. All these results suggest that LeACS-1 acts as a positive regulator of ethylene-induced shikonin biosynthesis in L. erythrorhizon hairy roots. Our work not only gives new insights into the understanding of the relationship between ET and shikonin biosynthesis, but also provides an efficient genetic engineering target gene for secondary metabolite production in non-model plant L. erythrorhizon.

A potent anticancer agent of shikonin derivative targeting tubulin.

In this study, a shikonin ester derivative, compound , was selected to evaluate its anticancer activities and we found that compound exhibited better antitubulin activities against the human HepG2 cell line with an IC50 value of 1.097 µM. Furthermore, the inhibition of tubulin polymerization results indicated that compound demonstrated the most potent antitubulin activity (IC50 = 13.88), which was compared with shikonin and colchicine as positive controls (IC50 = 25.28 µM and 22.56 µM), respectively. Compound was simulated to have good binding site with tubulin and arrested the cell cycle at G2/M phase, which also induces apoptosis in HepG2 cells, in which P53 and members of Bcl-2 protein family were both involved in the progress of apoptosis revealed by western blot. Confocal microscopy observations revealed compound targeted tubulin and altered its polymerization by interfering with microtubule organization. Based on these results, compound functions as a potent anticancer agent targeting tubulin.

Detection of the pandemic H1N1/2009 influenza A virus by a highly sensitive quantitative real-time reverse-transcription polymerase chain reaction assay.

A quantitative real time reverse-transcription polymerase chain reaction (qRT-PCR) assay with specific primers recommended by the World Health Organization (WHO) has been widely used successfully for detection and monitoring of the pandemic H1N1/2009 influenza A virus. In this study, we report the design and characterization of a novel set of primers to be used in a qRT-PCR assay for detecting the pandemic H1N1/2009 virus. The newly designed primers target three regions that are highly conserved among the hemagglutinin (HA) genes of the pandemic H1N1/2009 viruses and are different from those targeted by the WHO-recommended primers. The qRT-PCR assays with the newly designed primers are highly specific, and as specific as the WHO-recommended primers for detecting pandemic H1N1/2009 viruses and other influenza viruses including influenza B viruses and influenza A viruses of human, swine, and raccoon dog origin. Furthermore, the qRT-PCR assays with the newly designed primers appeared to be at least 10-fold more sensitive than those with the WHO-recommended primers as the detection limits of the assays with our primers and the WHO-recommended primers were 2.5 and 25 copies of target RNA per reaction, respectively. When tested with 83 clinical samples, 32 were detected to be positive using the qRT-PCR assays with our designed primers, while only 25 were positive by the assays with the WHO-recommended primers. These results suggest that the qRT-PCR system with the newly designed primers represent a highly sensitive assay for diagnosis of the pandemic H1N1/2009 virus infection.

[Simultaneous determination of 6 antibiotics and metronidazole in acne removal products bt high performance liquid chromatography].

An analytical method for the simultaneous determination of 6 antibiotics (minocycline, oxytetracycline, tetracycline hydrochloride, chlorotetracycline hydrochloride, doxycycline hydrochloride and chloramphenicol) and metronidazole in acne removal products of cosmetic was established using high performance liquid chromatography (HPLC). The drugs in the sample were extracted with methanol. The separation was performed on an Agilent ZORBAX SB-C18 column (250 mm x 4.6 mm, 5 microm) at 20 degrees C with methanol, acetonitrile and 0.002 mol/L oxalic acid solution as mobile phases with gradient elution at a flow rate of 0.8 mL/min. The detection was performed by a diode array detector (DAD) at 268 nm. The injection volume was 10 microL. The quantification was performed by external standard method. The calibration curves showed good linearity within the range of 1 - 30 mg/L with the correlation coefficients no less than 0.997 0. The detection limits were in the range of 1.1 - 1.2 microg/g. The recoveries were between 91.9% and 107.7% in three spiked levels of 5, 10 and 20 mg/L with the relative standard deviations (RSDs) of 0.13% - 1.74%. The method was used in the analysis of acne removal products, and metronidazole was found in 15% of the total test samples. The method is rapid, sensitive, accurate, effective in separation, and can be used in the determination of the six antibiotics and metronidazole in acne removal products.

RfaB, a galactosyltransferase, contributes to the resistance to detergent and the virulence of Salmonella enterica serovar Enteritidis.

In this study, a deletion mutant of rfaB (DeltarfaB) was observed to be susceptible to sodium dodecyl sulfate and less tolerant to bile salts. In addition, pre-incubation in 10% bile salts increased bacterial tolerance to 30% bile salts. We also showed that the DeltarfaB mutant invaded HeLa cells less than the wild type and resulted in a lower ratio of intracellular bacteria. Competitive infection of mice showed that the DeltarfaB mutant was defective in the colonization of host organs and was cleared more quickly in fecal shedding. Transforming of a plasmid containing a wild-type allele of rfaB (pRB3-rfaB) partially rescued the defect of the DeltarfaB mutant. The results suggest that RfaB, which is responsible to add the glycosyl residue to the core lipopolysaccharide, contributes to the tolerance to detergent and the virulence of Salmonella enterica serovar Enteritidis.