Overview of the applied quantification methods and corresponding reporter molecules. Left: total number of amine groups via qNMR (green), right: accessible number of amine groups via Ninhydrin assay (blue), SPDP assay (yellow) und 4-NB assay (violet).
Source: BAM, Biophotonics division
Nano- and microparticles of different sizes and materials are of increasing importance for a wide range of applications in the material and life sciences. Such materials are used as carriers for dye molecules in optical assays to detect disease relevant biomarkers and for drugs, as well as optical probes for bioimaging and sensing. This often involves the coupling of surface groups with linkers, sensor molecules, and analyte recognition moieties like (bio)ligands. Moreover, the surface chemistry of nanomaterials largely determines their interaction with their environment, biological systems, and thereby their possible toxicity.
To create optimal surface modifications and predict their interaction with their environment requires the knowledge of the chemical nature and number of the total and accessible surface groups. Therefore, a first bilateral comparison was performed by BAM and the National Research Council of Canada (NRC) to assess different analytical methods for the quantification of the total and accessible number of amine functionalities on silica nanoparticles of 20, 50, 80 and 120 nm diameter. Both laboratories used quantitative nuclear magnetic resonance (qNMR) for the quantification of the total amino groups and three different optical assays (Ninhydrin-, SPDP-, and 4-NB-assay) to determine the number of accessible amino groups. These methods had been developed and individually validated by both laboratories. The overall goal was to assess the comparability of the results across labs, to determine method-inherent limitations, and to identify best suited methods. Additionally, the ease of performance, robustness, and relative standard deviations were evaluated. This comparison presents a first step to the eventually required standardization of methods for surface group quantification.
Interlaboratory Comparison on the Quantification of Total and Accessible Amine Groups on Silica Nanoparticles with qNMR and Optical Assays
Filip Kunc, Nithiya Nirmalananthan-Budau, Bastian Rühle, Ying Sun, Linda J. Johnston, Ute Resch-Genger
published in Analytical Chemistry, Vol. 93, issue 46, pages 15271 - 15278, 2021
BAM Fachbereich Biophotonics