title: Evaluation of recombinant human parathyroid hormone by CZE method and its correlation with in vitro bioassay and LC methods.
abstract: A stability-indicating capillary zone electrophoresis (CZE) method was validated to assess the content/potency of the recombinant human parathyroid hormone (rhPTH 1-34), using ranitidine as internal standard (IS). A fused-silica capillary, (i.d. of 50µm; effective length of 40cm) was used at 25°C; the applied voltage was 20kV. The background electrolyte solution consisted of 50mmolL-1sodium dihydrogen phosphate solution at pH 3.0. Injections were performed using a pressure mode at 50 mbar for 45s, with detection by photodiode array (PDA) detector set at 200nm. Separation was obtained with a migration time of 5.3min, and was linear over the concentration range of 0.25-250µgmL-1(r2=0.9992). Specificity and stability-indicating capability were established in degradation studies, which also showed that there was no interference of the excipients. The accuracy was 100.28% with bias lower than 0.85%. Analyses of the same batches showed mean differences of the estimated content/potencies of 0.61%, 1.31% higher and 0.86% lower as compared to the validated reversed-phase and size exclusion liquid chromatography methods, and to the UMR-106 cell culture bioassay, respectively, with non-significant differences (p>0.05). Degraded forms were also subjected to the in vitro cytotoxicity test. The results obtained showed the capabilities of each one of the methods, and constitute an alternative strategy to monitor stability, improve the quality control and ensure the batch-to-batch consistency of bulk and finished biotechnology-derived medicine.
authors.affiliation: Postgraduate Program in Pharmaceutical Sciences, Federal University of Santa Maria, 97105-900 Santa Maria-RS, Brazil.
authors.forename: Fernanda Pavani Stamm
authors.initials: FP
authors.lastname: Maldaner
authors.affiliation: Postgraduate Program in Pharmaceutical Sciences, Federal University of Santa Maria, 97105-900 Santa Maria-RS, Brazil.
authors.forename: Rafaela Ferreira
authors.initials: RF
authors.lastname: Perobelli
authors.affiliation: Postgraduate Program in Pharmaceutical Sciences, Federal University of Santa Maria, 97105-900 Santa Maria-RS, Brazil.
authors.forename: Bruna
authors.initials: B
authors.lastname: Xavier
authors.affiliation: Department of Industrial Pharmacy, Federal University of Santa Maria, 97105-900 Santa Maria-RS, Brazil.
authors.forename: Gabriel Lunardi
authors.initials: GL
authors.lastname: Remuzzi
authors.affiliation: Postgraduate Program in Pharmaceutical Sciences, Federal University of Santa Maria, 97105-900 Santa Maria-RS, Brazil.
authors.forename: Maurício Elesbão
authors.initials: ME
authors.lastname: Walter
authors.affiliation: Department of Industrial Pharmacy, Federal University of Santa Maria, 97105-900 Santa Maria-RS, Brazil. Electronic address: sdalmora@terra.com.br.
authors.forename: Sérgio Luiz
authors.initials: SL
authors.lastname: Dalmora
mesh_terms: D000818:Animals; D001681:Biological Assay / Q000379:methods; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002850:Chromatography, Gel / Q000379:methods*; D056148:Chromatography, Reverse-Phase / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D007739:L Cells; D051379:Mice; D010281:Parathyroid Hormone / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D011899:Ranitidine / Q000378:metabolism / Q000592:standards; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism* / Q000494:pharmacology; D012015:Reference Standards; D015203:Reproducibility of Results
chemical_list: D010281:Parathyroid Hormone; D011994:Recombinant Proteins; D011899:Ranitidine
keywords: Biotechnology-derived medicine; Capillary zone electrophoresis; Liquid chromatography; Recombinant human parathyroid hormone; UMR-106 cell culture bioassay