A critical analysis of L-asparaginase activity quantification methods—colorimetric methods versus high-performance liquid chromatography

L-asparaginase or ASNase (L-asparagine aminohydrolase, E.C.3.5.1.1) is an enzyme clinically accepted as an antitumor agent to treat acute lymphoblastic leukemia (ALL) and lymphosarcoma through the depletion of L-asparagine (L-Asn) resulting in cytotoxicity to leukemic cells. ASNase is also important...

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Bibliographic Details
Published in:Analytical and bioanalytical chemistry 2018-11, Vol.410 (27), p.6985-6990
Main Authors: Magri, Agnes, Soler, Matheus F., Lopes, André M., Cilli, Eduardo M., Barber, Patrick S., Pessoa, Adalberto, Pereira, Jorge F. B.
Format: Article
Language:English
Subjects:
Acrylamide
Acute lymphoblastic leukemia
Analytical Chemistry
Anticancer properties
Asparaginase
Asparaginase - analysis
Asparaginase - metabolism
Asparagine
Asparagine - analogs & derivatives
Asparagine - metabolism
Aspartic acid
Aspartic Acid - metabolism
Biochemistry
Bioequivalence
Characterization and Evaluation of Materials
Chemical properties
Chemistry
Chemistry and Materials Science
Chromatography
Chromatography, High Pressure Liquid - methods
Colorimetry
Colorimetry - methods
Communication
Comparative analysis
Control methods
Correlation analysis
Cytotoxicity
Enzyme Assays - methods
Food industry
Food processing
Food processing industry
Food Science
High performance liquid chromatography
Humans
L-asparaginase
Laboratory Medicine
Leukemia
Liquid chromatography
Lymphatic leukemia
Lymphosarcoma
Methods
Monitoring/Environmental Analysis
Quality control
Substrates
Toxicity
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Summary:L-asparaginase or ASNase (L-asparagine aminohydrolase, E.C.3.5.1.1) is an enzyme clinically accepted as an antitumor agent to treat acute lymphoblastic leukemia (ALL) and lymphosarcoma through the depletion of L-asparagine (L-Asn) resulting in cytotoxicity to leukemic cells. ASNase is also important in the food industry, preventing acrylamide formation in processed foods. Several quantification techniques have been developed and used for the measurement of the ASNase activity, but standard pharmaceutical quality control methods were hardly reported, and in general, no official quality control guidelines were defined. To overcome this lack of information and to demonstrate the advantages and limitations, this work properly compares the traditional colorimetric methods (Nessler; L-aspartic acid β-hydroxamate (AHA); and indooxine) and the high-performance liquid chromatography (HPLC) method. A comparison of the methods using pure ASNase shows that the colorimetric methods both overestimate (Nessler) and underestimate (AHA and indooxine) the ASNase activity when compared to the values obtained with HPLC, considered the most precise method as this method monitors both substrate consumption and product formation, allowing for overall mass-balance. Correlation and critical analysis of each method relative to the HPLC method were carried out, resulting in a demonstration that it is crucial to select a proper method for the quantification of ASNase activity, allowing bioequivalence studies and individualized monitoring of different ASNase preparations. Graphical abstract ᅟ
ISSN:1618-2642
1618-2650
DOI:10.1007/s00216-018-1326-x