ABSTRACT
OBJECTIVE:To establish a method for the 5 main components (original syringin,chlorogenic acid,eleutheroside E,isofraxidin and quercetin-3-rhamnoside) in the fruits and roots of wild Acanthopanax senticosus. METHODS:UPLC was per-formed on the column of Waters ACQUITY UPLC HSS T3 with mobile phase of acetonitrile-0.3% phosphoric acid (gradient elu-tion)at a flow rate of 0.2 ml/min. Detection wavelength was 300 nm,column temperature was 30 ℃,and injection volume was 10μl. RESULTS:The linear range was 24.56-184.2 μg/ml for syringin(r=0.9993),18.454-138.405 μg/ml for chlorogenic acid(r=0.9993),8.416-63.12 μg/ml for eleutheroside E (r=0.9997),3.286-24.645 μg/ml for isofraxidin (r=0.9993) and 2.522-18.915μg/ml for quercetin-3-rhamnoside(r=0.9998);RSDs of precision,stability and reproducibility tests were lower than 1%;recover-ies were 99.14%-100.50%(RSD=0.48%,n=6)for syringing in the fruits of A. senticosus、99.03%-100.45%(RSD=0.50%,n=6) for chlorogenic acid in the fruits of A. senticosus、99.22%-100.44%(RSD=0.44%,n=6)for eleutheroside E in the fruits of A. sen-ticosus、99.80%-100.80%(RDS=0.44%,n=6)for isofraxidin in the fruits of A. senticosus、99.76%-101.10%(RSD=0.51%,n=6) for quercetin-3-rhamnoside in the fruits of A. senticosus;99.21%-101.20%(RSD=0.73%,n=6)for syringing in the root of A. senti-cosus、99.81%-101.20%(RSD=0.52%,n=6)for chlorogenic acid in the root of A. senticosus、100.00%-101.50%(RSD=0.62%, n=6)for eleutheroside E in the root of A. senticosus、99.22%-100.40%(RSD=0.47%,n=6)for isofraxidin in the root of A. senti-cosus. CONCLUSIONS:The method is simple and stable with good reproducibility,and can be used for the simultaneous determi-nation of original syringin,chlorogenic acid,eleutheroside E,isofraxidin and quercetin-3-rhamnoside in the fruits and root of wild A. senticosus.
ABSTRACT
OBJECTlVE To study the effect of ganoderma poIysaccharide( GLP)on zebrafish ( Danio rerio)survivaI,deveIopment and senescence. METHODS A p53-knockout zebrafish modeI was estabIished by injecting of morphoIino phosphorodiamidate(mO)into normaI zebrafish eggs in the singIe ceII deveIopment stage. Different concentrations of GLP 1,2 and 3 g·L-1 were used to treat both wiId type and p53-knockout zebrafish embryos 8 h post fertiIization(8 hpf)under 28℃ for 3 d. The survivaI and maIformation rates were caIcuIated after 72 hpf,and the effect of GLP on ceII senescence was evaI-uated by senescence associated β-gaIactosidase(SA-β-gaI)staining both in wiId type and p53-knockout zebrafish embryos. In addition,the differentiaI gene expression of cancer inhibitor gene(p53),teIomer-ase reverse transcriptase( TERT),murine doubIe minute2( mdm2)and p21 was examined by RT-PCR both in wiId type and p53-knockout zebrafish embryos after treatment with different concentrations of GLP. RESULTS GLP 1 and 2 g·L-1 had no effect on deveIopment of wiId type zebrafish embryos,but after GLP 3 g·L-1 treatment,most of zebrafish embryos dispIayed maIformation and the survivaI rate was onIy 48.6%. GLP 1,2 and 3 g·L-1 had no effect on survivaI and deveIopment of p53-knockout zebrafish embryos. After GLP 2 g·L-1 treatment for 3 d,the SA-β-gaI staining positive rate of wiId type zebrafish embryos was reduced compared with controI group(P﹤0.01),whiIe there was no significant change in p53-knockout zebrafish embryos. The resuIts of RT-PCR showed that GLP 2 g·L-1 treatment depressed p21 and p53 gene expression(P﹤0.05,P﹤0.01),and had no effect on mdm2 and TERTgene expres-sion of wiId type zebrafish embryos. For p53-knockout zebrafish embryos,the TERT,mdm2,p21 and p53 gene expression dispIayed no significant difference after GLP 2 g·L-1 treatment. CONCLUSlON GLP 2 g·L-1 may improve senescence of wiId type zebrafish embryo ceIIs. GLP ≥3 g·L-1 may Iead to death and abnormaI deveIopment of wiId type zebrafish embryos. The improvement of GLP on senescence of wiId type zebrafish embryo ceIIs might be mediated by its down-reguIation of p21 and p53 gene expression.