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Analysis of streptokinase by validated liquid chromatography methods and correlation with an in vitro bioassay
Author(s) -
Cardoso Clóvis Dervil Appratto,
Perobelli Rafaela Ferreira,
Xavier Bruna,
Maldaner Fernanda Pavani Stamm,
da Silva Francielle Santos,
Dalmora Sérgio Luiz
Publication year - 2017
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.201600880
Subject(s) - chromatography , chemistry , sodium acetate , high performance liquid chromatography , streptokinase , size exclusion chromatography , calibration curve , reversed phase chromatography , sodium , sodium sulfate , detection limit , enzyme , biochemistry , psychology , organic chemistry , psychiatry , myocardial infarction
Reversed‐phase and size‐exclusion liquid chromatography methods were validated for the assessment of streptokinase. The reversed‐phase method was carried out on a Jupiter C 4 column (250 mm × 4.6 mm id) maintained at 25°C. The mobile phase consisted of 50 mM sodium sulfate solution pH 7.0 and methanol (90:10, v/v), run isocratically at a flow rate of 0.8 mL/min. The size‐exclusion method was carried out on a Protein KW 802.5 column (300 mm × 8.0 mm id), at 25°C. The mobile phase consisted of 40 mM sodium acetate solution pH 7.0, run isocratically at a flow rate of 1.0 mL/min. Retention times were 19.3 min, and 14.1 min, and calibration curves were linear over the concentration range of 0.25–250 μg/mL (25.75–25 750 IU/mL) ( r 2 = 0.9997) and 5–80 μg/mL (515–8240 IU/mL) ( r 2 = 0.9996), respectively, for reversed‐phase and size exclusion, with detection at 220 and 204 nm. Chromatographic methods were employed in conjunction with the in vitro bioassay for the content/potency assessment of Streptokinase, contributing to improve the quality control and ensure the efficacy of the biotherapeutic.