Content/Potency Assessment of Botulinum Neurotoxin Type-A by Validated Liquid Chromatography Methods and Bioassays

Botulinum neurotoxin type-A (BoNTA) is one of the seven different serotypes (A to G) produced by Clostridium botulinum. A stability-indicating size-exclusion chromatography (SEC) method was developed and validated, and the specificity was confirmed by forced degradation study, interference of the ex...

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Bibliographic Details
Published in:Toxins 2019-01, Vol.11 (1), p.35
Main Authors: Xavier, Bruna, Perobelli, Rafaela Ferreira, Walter, Maurício Elesbão, da Silva, Francielle Santos, Dalmora, Sérgio Luiz
Format: Article
Language:English
Subjects:
Animals
Bioassays
Biocompatibility
Biological activity
Biological Assay
Biological products
Biopharmaceuticals
Biotechnology
botulinum neurotoxin type A
Botulinum toxin
Botulinum Toxins, Type A - analysis
Botulinum Toxins, Type A - toxicity
Cell culture
Cell Line
Cell Survival - drug effects
Chromatography
Chromatography, Gel
Chromatography, Liquid
Chromatography, Reverse-Phase
Cytotoxicity
Female
Humans
LD50 mouse bioassay
Lethal Dose 50
Liquid chromatography
Methods
Mice
Neurotoxins
Pharmaceuticals
Potassium
Proteins
Reproducibility of Results
reversed-phase chromatography
Serotypes
Size exclusion chromatography
Sodium
Standard deviation
Toxicity
Toxins
T−47D cell culture
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Summary:Botulinum neurotoxin type-A (BoNTA) is one of the seven different serotypes (A to G) produced by Clostridium botulinum. A stability-indicating size-exclusion chromatography (SEC) method was developed and validated, and the specificity was confirmed by forced degradation study, interference of the excipients, and peaks purity. The method was applied to assess the content and high-molecular-weight (HMW) forms of BoNTA in biopharmaceutical products, and the results were compared with those of the LD mouse bioassay, the T-47D cell culture assay, and the reversed-phase chromatography (RPC) method, giving mean values of 0.71% higher, 0.36% lower, and 0.87% higher, respectively. Aggregated forms showed significant effects on cytotoxicity, as well as a decrease in the bioactivity ( < 0.05). The employment of the proposed method in conjunction with the optimized analytical technologies for the analysis of the intact and altered forms of the biotechnology-derived medicines, in the correlation studies, enabled the demonstration of the capability of each one of the methods and allowed for great improvements, thereby assuring their safe and effective use.
ISSN:2072-6651
2072-6651
DOI:10.3390/toxins11010035