OBJECTIVE:To establish a method for the simultaneous content determination 5 anthraquinones of rhei radix-copti-dis rhizoma,and optimize its best mass ratio. METHODS:HPLC was performed on the column of Zorbax SB-C18 (250 mm × 4.6 mm,5 μm)with mobile phase of acetonitrile-0.05 mol/L potassium dihydrogen phosphate(48:52, V/V,adding sodium dodecyl sulfate 0.4 g per 100 mL,pH value was adjusted to 4.0);detection wavelength was 254 nm;flow rate was 1 mL/min;column tem-perature was 25℃;and injection volume was 10μL. When mass ratio of rhei radix-coptidis rhizoma was 1:1,1:2,2:1,2:3,3:2,the contents of aloe emodin,rhein,emodin,chrysophanol and emodin ether were investigated. RESULTS:The aloe emodin, rhein, emodin, chrysophanol and emodin ether showed good linear relationship with peak area in the range of 0.652-3.081, 0.704-3.422,1.280-6.197,0.633-0.324,1.326-5.954 μg(r≥0.9997);RSDs of precision,stability,reproducibility tests were low-er than 2.0%(n=6);and average recovery was 98.92%-100.37%(RSD≤2.26%,n=6). When mass ratio was 2:1,the total contents of 5 ingredients were the highest. CONCLUSIONS:The method has good precision,stability,reproducibility,and can be used for the quality verification of rhei radix-coptidis rhizoma under different compatibilities. The best mass ratio of rhei radix-copti-dis rhizoma is 2:1.

Cysticercosis,caused by the larval stage of the tapeworm Taenia pisiformis,known as Cysticercus pisiformis,is a parasitic ailment affecting lagomorphs,particularly domestic rabbits,posing a threat to the rabbit industry and the safety of rabbit meat products.This study aims to i-dentify the distribution of the TPO18 antigen in Cysticercus pisiformis and Taenia pisiformis and establish an indirect enzyme-linked immunosorbent assay(ELISA)for detecting antibodies against rabbit cysticercosis.The research involved the prokaryotic expression of the 18 kDa antigen of rabbit Cysticercus pisiformis and the isolation of soluble TPO18 protein post-purification.Immuni-zing rabbits with the TPO18 protein resulted in the production of polyclonal antibodies with a titer of up to 1∶51 200.Western blot analysis validated the antigenicity of the polyclonal antibodies a-gainst total proteins from rabbit Cysticercus pisiformis,Taenia pisiformis and the recombinant TPO18 protein.Immunohistochemistry revealed the distribution of the TPO18 antigen in rabbit Cysticercus pisiformis and Taenia pisiform is,indicating the effective reactivity of the polyclonal antibodies with total proteins from both parasites and the recombinant TPO18 protein.TPO18 an-tigen in rabbit Cysticercus pisiformis predominantly localized in the germinal layer and the paren-chyma,while in Taenia pisiformis,it was mainly present in the suckers,sucker peripheries,collec-ting duct upper cells,and parenchyma.An indirect ELISA based on the TPO18 antigen was devel-oped using the recombinant antigen,and its technical parameters were optimized.The optimized ELISA conditions included a serum dilution of 1∶100,antigen coating concentration of 5 mg/L,coating for 1 h at 37 ℃ followed by overnight incubation at 4 ℃,blocking with 1％BSA for 60 min at 37 ℃,serum reaction for 60 min,secondary antibody dilution at 1∶1 000,secondary antibody incubation for 60 min,substrate reacting for 15 min,with a cutoff value of 0.295.Sensitivity,speci-ficity,and repeatability tests of the ELISA demonstrated high sensitivity and specificity without cross-reactivity with positive sera of rabbit hemorrhagic disease virus,Hepatic coccidiosis,Eimer-ia stiedae,or Toxoplasma gondii.The intra-and inter-assay coefficients of variation were both less than 7％,indicating excellent repeatability.Application of this ELISA,compared to postmortem ex-amination,on 86 clinical serum samples showed a concordance rate of 97.7％.In conclusion,this study successfully established an indirect ELISA for detecting antibodies against rabbit Cysticercus pisiformis,presenting a novel monitoring approach for assessing rabbit infections with Cysticer-cus pisiformis.