title: Granulocyte-macrophage colony stimulating factor: Evaluation of biopharmaceutical formulations by stability-indicating RP-LC method and bioassay.
abstract: The granulocyte-macrophage colony stimulating factor (GM-CSF) is a cytokine that regulates the proliferation and differentiation of hematopoietic cells and activates granulocytes and macrophages. A reversed-phase liquid chromatography (RP-LC) method was validated for the assessing of the stability of non-glycosylated recombinant rhGM-CSF (Molgramostim) in biopharmaceutical formulations. The RP-LC method was carried out on a Jupiter C(4) column (250 mm × 4.6 mm i.d.), maintained at 45 °C. The mobile phase A consisted of 0.1% TFA and the mobile phase B was acetonitrile with 0.1% TFA in acetonitrile, run at a flow rate of 1 mL/min, and using photodiode array (PDA) detection at 214 nm. Chromatographic separation was obtained with a retention time of 29.2 min, and was linear over the concentration range of 2-300 μg/mL (r(2) = 0.9992). Specificity was established in degradation studies. Moreover, the in vitro cytotoxicity test of the degraded products showed significant differences (p < 0.05). The method was applied to the assessment of rhGM-CSF and related proteins in biopharmaceutical dosage forms, and the results were correlated to those of a bioassay. It is concluded that the employment of RP-LC in conjunction with current methods allows a great improvement in monitoring stability, quality control and thereby assures the therapeutic efficacy.
authors.affiliation: Postgraduate Program in Pharmaceutical Sciences, Federal University of Santa Maria, RS, Brazil.
authors.forename: Diogo Paim
authors.initials: DP
authors.lastname: Leal
authors.forename: Ricardo Bizogne
authors.initials: RB
authors.lastname: Souto
authors.forename: Renato
authors.initials: R
authors.lastname: Schutkoski
authors.forename: Ana Cláudia
authors.initials: AC
authors.lastname: Bergamo
authors.forename: Sérgio Luiz
authors.initials: SL
authors.lastname: Dalmora
mesh_terms: D002853:Chromatography, Liquid; D004355:Drug Stability; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology*; D057230:Limit of Detection; D004364:Pharmaceutical Preparations; D015203:Reproducibility of Results
chemical_list: D004364:Pharmaceutical Preparations; D016178:Granulocyte-Macrophage Colony-Stimulating Factor