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Objective:To evaluate the in vitro cross-neutralization of serum antibodies in human and mice immunized with inactivated SARS-CoV-2 vaccine against Delta and Beta variants. Methods:Human serum samples after a second and a third dose of inactivated SARS-CoV-2 vaccine and mouse serum samples after a two-dose vaccination were collected. The neutralizing antibodies in the samples against SARS-CoV-2 strains of prototype, Delta and Beta variants were detected using micro-neutralization assay in biosafety level Ⅲ laboratory. The seroconversion rates and geometric mean titers (GMTs) of antibodies were calculated.Results:The seroconversion rates of antibodies in human serum samples against different SARS-CoV-2 strains were all above 95%. After two-dose vaccination, the GMTs of neutralizing antibodies against the prototype, Delta and Beta strains were 109, 41 and 15, respectively. The GMTs decreased by 2.7 folds and 7.3 folds for the Delta and Beta variants as compared with the prototype strain. After the booster vaccination, the GMTs of neutralizing antibodies against the prototype, Delta and Beta strains were 446, 190 and 86, respectively. The GMTs of neutralizing antibodies against Delta and Beta variants decreased by 2.3 folds and 5.2 folds as compared with that against the prototype strain. The seroconversion rates of antibodies against different SARS-CoV-2 strains in mouse serum samples were all 100%. The GMTs of neutralizing antibodies against the prototype, Delta and Beta strains were 2 037, 862 and 408, respectively. The GMTs decreased by 2.4 folds and 5.0 folds for the Delta and Beta variants.Conclusions:Inactivated SARS-CoV-2 vaccine could induce a certain level of neutralizing antibodies against Delta and Beta variants in both human and mouse models. Moreover, a third dose of vaccine induced higher levels of neutralizing antibodies against Delta and Beta variants in human. This study provided valuable data for the clinical application and protective evaluation of the inactivated SARS-CoV-2 vaccine.
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A method based on QuEChERS coupled with ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was developed for the determination of seven kinds of fluorescent white agent residues in mushroom.After mixing with 10 mL of water, the sample was extracted with acidified acetonitrile, cleaned up by C18, primary secondary amine (PSA) and MgSO4.The separation of seven kinds of fluorescent white agents was performed on a C18 column with a gradient elution of acetonitrile and 0.1% acidified water as mobile phase.The target compounds were detected by electrospray ionization mass spectrometry in positive ion mode with multi reaction monitoring (MRM).Under the optimum conditions, the method had good linear relationship in the determination of the seven kinds of fluorescent white agents in a certain concentration range, with correlation coefficients greater than 0.991.Moreover, the limits of detection (S/N=3) were 0.05-0.4 μg/kg, the limits of quantitation (S/N=10) were 0.2-1.3 μg/kg, and the average recoveries for seven kinds of fluorescent white agent residues in msushroom were 70.1%-109.2%.In comparison with previous methods, the new procedure has characteristics of simple sample preparation and higher sensitivity.
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OBJECTIVE:To conduct the quality assessment of artificial Aquilaria sinensis by"cutting inducing technique", and provide reference for its scientific planting and harvest. METHODS:GC-MC and HPLC were adopted to detect the volatile in-gredients,characteristic spectrum,incense tetraol and alcohol-soluble extract contents in 3 batches of artificial A. sinensis(Num. 1, 2,3,respectively for 5,10,20 years)by"cutting inducing technique". RESULTS:The volatile ingredients of 3 batches of artifi-cial A. sinensis mainly consisted of aromatic compounds,sesquiterpene compounds,fatty acid compounds and chromone com-pounds. The characteristic spectrums of samples 2,3 were basically the same with the reference substance of A. sinensis. The in-cense tetraol contents of 3 batches of samples were 0.078%-0.254%,and alcohol-soluble extract contents were 12.4%-20.8%. The characteristic spectrum and the incense tetraol content of sample 1 were not conformed to the standards in Chinese Pharmacopoeia (2015 edition,part 1). CONCLUSIONS:Artificial A. sinensis by"cutting inducing technique"shows similar volatile ingredients to natural A. sinensis. The quality of artificial A. sinensis for more than 10 years is conformed to the standards in Chinese Pharmaco-poeia(2015 edition,part 1),which can be used as medicine,replacing the natural A. sinensis.
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OBJECTIVE:To conduct the quality assessment of artificial Aquilaria sinensis by"cutting inducing technique", and provide reference for its scientific planting and harvest. METHODS:GC-MC and HPLC were adopted to detect the volatile in-gredients,characteristic spectrum,incense tetraol and alcohol-soluble extract contents in 3 batches of artificial A. sinensis(Num. 1, 2,3,respectively for 5,10,20 years)by"cutting inducing technique". RESULTS:The volatile ingredients of 3 batches of artifi-cial A. sinensis mainly consisted of aromatic compounds,sesquiterpene compounds,fatty acid compounds and chromone com-pounds. The characteristic spectrums of samples 2,3 were basically the same with the reference substance of A. sinensis. The in-cense tetraol contents of 3 batches of samples were 0.078%-0.254%,and alcohol-soluble extract contents were 12.4%-20.8%. The characteristic spectrum and the incense tetraol content of sample 1 were not conformed to the standards in Chinese Pharmacopoeia (2015 edition,part 1). CONCLUSIONS:Artificial A. sinensis by"cutting inducing technique"shows similar volatile ingredients to natural A. sinensis. The quality of artificial A. sinensis for more than 10 years is conformed to the standards in Chinese Pharmaco-poeia(2015 edition,part 1),which can be used as medicine,replacing the natural A. sinensis.
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Objective To express virus-like particles(VLP) of enterovirus 71 (EV71) in Han-senula polymorpha.Methods The coding sequences of P1 and 3CD genes of EV71 were optimized accord-ing to codon usage bias of Hansenula polymorpha for achieving high expression , and then cloned into the ex-pression vector PMV of Hansenula polymorpha .The recombinant expression vector PMV-P1-3CD was trans-formed into Hansenula polymorpha AU 0501 .The transformants were stably cultured in selective medium (Yeast Nitrogen Base) and screened for strains with positive P1 and 3CD genes by PCR.Then an induced cultivation on the recombinant strains were performed in a medium supplemented with methanol to a final concentration of 1.0%and the expressed products were analyzed by SDS-PAGE and Western blot assays to select high expression strains .The high expression strains were cultured in 30 L fermentor and its fermenta-tion products were analyzed by electronic microscope after purification .Results EV71 recombinant expres-sion strains were successfully constructed .The results of SDS-PAGE showed that the expressed products had obvious VP3, VP1, VP0 protein bands with molecular weights of 26×103, 33×103 and 35×103, respective-ly, which were consistent with the expected molecular weight of the fusion proteins .Western blot demonstra-ted that the expressed products could be specifically recognized by the polyclonal antibody against EV 71-VP1 at 33 ×10 3 , indicating its high immunoreactivity .ELISA confirmed that the expression level of EV 71 fermen-tation products was reached to 200 mg/L.Electronic microscope analysis showed that the VLP of recombi-nant EV71 were 24-30 nm in diameter with normal structure .Conclusion The virus-like particles of human enterovirus 71 are successfully expressed in Hansenula polymorpha , which provides a foundation for the fur-ther development of EV 71 VLP vaccine .