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AIM To establish an HPLC method for the simultaneous content determination of monocaffeyltartaric acid,forsythiaside A,chicoric acid,phillyrin,arctiin and harpagoside in Waigan Fenghan Granules.METHODS The analysis of 70%methanol extract of this drug was performed on a 30℃thermostatic Agilent Zorbax SB-C18 column(250 mm×4.6 mm,5 μm),with the mobile phase comprising of acetonitrile-0.1%phosphoric acid flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelengths were set at 278,330 nm.RESULTS Six constituents showed good linear relationships within their own ranges(r≥0.999 4),whose average recoveries were 96.80%-102.61%with the RSDs of 0.26%-1.93%.CONCLUSION This simple and accurate method can be used for the quality control of Xiao'er Qingyan Granules.
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Objective:Death causes and life reduction of malignant tumors in the residents of Zhuanqiao Town in Minhang District from 2013 to 2017 were analyzed to provide scientific evidence for the strategies on comprehensive prevention and control of cancer. Methods:The data of death causes of malignant tumors in the residents of Zhuanqiao Town were collected and analyzed. The mortality rate, annual percent change (APC), composition ratio, potential years of life lost (PYLL), potential years of life lost rate (PYLLR) and average years of life lost (AYLL) of the registered population were analyzed. Results:The standardized mortality rate of malignant tumors in the residents of Zhuanqiao Town from 2013 to 2017 was 128.05/105, and the rate was higher in males than that in females. The top four cancers regarding PYLL were lung cancer, liver cancer, stomach cancer and colorectal cancer, which were roughly the same order as the top four regarding the mortality. This indicates that these four cancers had a greater impact on residents. Lung cancer had a greater impact on female life expectancy. PYLL and SPYLL ranked the first in liver cancer in males and thus had a greater impact on the males. Breast cancer was one of the most important malignant tumors in causing early death of women. Conclusion:Malignant tumor has become an important public health problem endangering the health of residents. The focus of future work in the town remains to improve public awareness of carcinogenic risk factors, actively carry out health education, lifestyle intervention and early screening in order to reduce cancer risk, alleviate cancer burden and improve the life expectancy of residents.
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OBJECTIVE To design N dodecanol modified docetaxel(DTX) prodrug, prepare nanostructured lipid carrier(NLC) and investigate in vitro antitumor activity and in vivo pharmacodynamic. METHODS Nanostructured lipid carrier (DNLC) encapsulating n-dodecanol-modified DTX prodrug was prepared by ultrasonic method. The formulation was optimized by single-factor experiment and response surface optimization. The accumulated rates of DTX degraded from DNLC in different media was evaluated by high performance liquid chromatography (HPLC). The morphology of DNLC was observed by transmission electron microscopy (TEM). The particle size and PDI of DNLC were determined by Malvern particle size analyzer. The long-term stability of the preparations was investigated. In vitro cytotoxicity of DNLC was measured by MTT method. In vivo pharmacodynamics of DNLC were performed in 4T1 tumor xenograft balb/c mice using saline and DTX-Sol as control. RESULTS n-Dodecanol-modified DTX prodrug was synthesized and used to prepare DNLC. The optimal formulation was as following: mass ratio of emulsifier to co-emulsifier (Km) of 1∶3, solid-liquid lipid ratio of 1.43∶1, drug-lipid ratio of 1∶10, the emulsifier concentration of 60 mg•mL-1, the temperature of 70 ℃ and the stirring speed of 800 r•min-1. DNLC had a round appearance and a uniform spherical shape. And the particle size and PDI remained substantially stable within 30 d. The accumulated rates of DTX degraded from DNLC in PBS (pH 7.4), PBS (pH 7.4) containing 10 mmol•L-1 DTT and 10 mmol•L-1 H2O2 was (9.07±0.01)%, (21.52±0.35)% and (96.72±4.12)% at 24 h, respectively. After incubation of DTX-Sol and DNLC with 4T1 cells for 72 h, IC50 of DTX-Sol and DNLC were (1.2±0.2) and (13.2±4.3)nmol•L-1, respectively. The cytotoxicity of DTX-Sol group was stronger than that of DNLC group. At the end of the pharmacodynamics, the tumor volumes of the mice in saline, DTX-Sol and DNLC groups were (1 930.39±215.20), (1 013.64±138.65), and (765.16±177.43)mm3, respectively. And the change percentage of body weight in saline, DTX-Sol and DNLC groups were (-19.69±4.44)%, (-14.85±3.61)% and (-2.61±1.70)%. There were significant differences in tumor volume and body weight between the DNLC and DTX-Sol group (P<0.05). CONCLUSION The prepared DNLC shows good stability, redox sensitivity, obvious anti-tumor effect and lower toxicity. These RESULTS could provide a new experimental basis for the development of DTX prodrug loaded nano-drug delivery system.
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<p><b>OBJECTIVE</b>To construct a gene knock-out mutant of response regulator named RevS in Streptococcus suis serotype 2 virulent strain 05ZYH33, and to investigate the effects of its deletion on the biological characters of this pathogen and the pathogenesis to mice and piglets.</p><p><b>METHODS</b>Recombinant gene knock-out vector consisting of Spc(r) cassette was constructed and flanking was constructed consisting of Spc(r) cassette with flanking homology regions to the RevS genes while the isogenic RevS-deficient mutant was screened by allelic replacement. The effects of RevS deletion on the basic biological characters of 05ZYH33 including growth stability, colonial morphology, haemolysis, Gram staining, growth curve and protein expression were examined in vitro. The mice and piglets were infected with 10(8) CFU wild virulent and mutant isolates.</p><p><b>RESULTS</b>PCR analysis confirmed that the coding genes of RevS were replaced completely by Spc(r) cassette and the basic biological characters of 05ZYH33 did not undergo any apparent change. Balb/c mice infection assay indicated that RevS play a role in the pathogenesis of Streptococcus suis infections, while no remarkable difference was observed in the piglets' pathogenesis infection rates between mutant isolates deltaA05ZYH33 and wild-type isolates 05ZYH33.</p><p><b>CONCLUSION</b>The mutant of Streptococcus suis 05ZYH33 response regulator was successfully constructed, while the mutation did not obviously affect the bacterial biological characters, while the knock-out mutant of RevS was shown to be attenuated in pathogenesis to mice and piglets.</p>
Subject(s)
Animals , Mice , Bacterial Proteins , Genetics , Gene Knockout Techniques , Methods , Mice, Inbred BALB C , Models, Genetic , Polymerase Chain Reaction , Streptococcal Infections , Microbiology , Streptococcus suis , Genetics , VirulenceABSTRACT
<p><b>OBJECTIVE</b>To determine the prevalence of Streptococcus suis and major pathogenic serotypes in middle part of Jiangsu province.</p><p><b>METHODS</b>Tonsillar specimens from 303 slaughtered pigs aged 6 to 8 months were investigated for the presence of Streptococcus suis and major pathogenic serotypes by polymerase chain reaction (PCR) method. Bacteriological examination compared with molecular genetics identification for three Streptococcus suis isolates were also done.</p><p><b>RESULTS</b>The overall carrier rate of Streptococcus suis was up to 88.0%, with the percentages of serotype 1(14), 2(1/2), 7 and 9 were 9.6%, 8.5%, 11.3% and 29.5% respectively in 2005. While in 2006, the prevalence of Streptococcus suis was 82.5%, with capsular types 1 (14), 2 (1/2), 7 and 9 were accounted for 17.6%, 2.4%, 25.8% and 20.0% of all the specimens. All the three isolates belonged to Streptococcus suis serotype 2,named 2a, 2f and 14e, which exhibiting the virulent phenotype cps2+/gdh+/mrp-/lepf-/sly-/fbps+/orf2+/89k-, cps2+/lgdh+/mrp-/epf-/sly-/fbps-/orf2-/89k- and cps2+/gdh+/mrp-/epf-/sly-/fbps/orf2-/ respectively. These isolates were all susceptible to amoxicillin, ampicillin, penicillin and resistant to amikacin and tetraycline. Clinical signs were not noted in BALB/c mice and rabbit.</p><p><b>CONCLUSION</b>Prevalence of the Streptococcus suis among the healthy herds in the areas was very high, with various capsule types of Streptococcus suis involved in the same herds, and the virulent phenotype of these 3 isolates were very different from those prevalent Streptococcus suis serotype 2 virulent isolates frequently discovered from the epidemic areas.</p>
Subject(s)
Animals , Mice , Amikacin , Therapeutic Uses , Amoxicillin , Therapeutic Uses , Ampicillin , Therapeutic Uses , China , Epidemiology , Mice, Inbred BALB C , Molecular Epidemiology , Methods , Penicillins , Therapeutic Uses , Polymerase Chain Reaction , Streptococcal Infections , Drug Therapy , Epidemiology , Microbiology , Streptococcus suis , Classification , Genetics , Virulence , Tetracycline , Therapeutic Uses , VirulenceABSTRACT
<p><b>OBJECTIVE</b>To clone and express Streptococcus suis serotype 2 (S. suis 2) sly gene for constructing an foundation on identification of S. suis 2 protective antigen.</p><p><b>METHODS</b>The sly gene was amplified from S. suis 2 clinical isolate strain 05ZYH33 genome DNA by PCR. The gene fragment was inserted into the expression vector pET-30b(+) to build pET30b-sly. When recombinant vector pET30b-sly was identified by restriction enzyme cutting and DNA sequencing as a correct one, subsequently it was transformed to E. coli Rosetta for expression under IPTG induction. The obtained fusion protein was purified by Ni-NTA affinity chromatography. The immunologic and hemolysis activity of the purified protein was proved through Western blot and hemolysis assay respectively.</p><p><b>RESULTS</b>The PCR product was around 1500 bp. The gene segment inserted into the recombinant vector was proven to be completely identical with the sly gene sequence in the total genome sequence of S. suis 2. The target protein expressed was up to 30% of the total somatic protein under IPTG induction. The protein purity reached above 80% after purification. The protein could be recognized by human serum infected with S. suis 2 and could dissolve swine erythrocytes with the Hemolytic titer as 256.</p><p><b>CONCLUSION</b>The expression vector pET30b-sly was successfully constructed. The target protein could be over-expressed in E. coli and possessed its biological activity after purification.</p>
Subject(s)
Animals , Humans , Bacterial Proteins , Genetics , Metabolism , Pharmacology , Blotting, Western , Chromatography, Affinity , Hemolysis , Recombinant Proteins , Genetics , Metabolism , Pharmacology , Streptococcus suis , Genetics , Metabolism , SwineABSTRACT
<p><b>OBJECTIVE</b>To rapidly and sensitively detect the four virulence-associated factors of Streptococcus suis, a multiplex PCR was developed.</p><p><b>METHODS</b>In the process of this reaction, four distinct DNA targets were amplified. One target was based on the serotype 2 (and 1/2) specific cps gene and the others were based on Streptococcus suis mrp, epf (epf*) and sly gene, encoding the MRP, EF(EF*) and Sly proteins of Streptococcus suis. 72 isolates, which including 48 strains of Streptococcus suis and 24 strains of negative control, and 49 clinical specimens were detected by the multiplex PCR assay.</p><p><b>RESULTS</b>All PCR products were detected by electrophoresis on 1.2% agarose gels. With the 48 Streptococcus suis strains, the positive detection rates of cps2+, mrp+, epf+, epf*+ and sly+ were 16/48, 14/48, 12/48, 3/48 and 26/48,respectively. The results were confirmed by bacteriological examination. There were no specific amplification products including 49 clinical specimens and 24 negative control strains.</p><p><b>CONCLUSION</b>The results demonstrated that multiplex PCR was a highly specific and sensitive diagnostic tool for the detection of virulence-associated factors of streptococcus suis.</p>