Evaluating Water-Soluble Chitosan Molecular Weight with the BeSEC
2026-01-14Application Note
Abstract: Chitosan molecular weight plays a critical role in determining its functional performance across food, pharmaceutical, and environmental applications. In this study, size-exclusion chromatography coupled with static light scattering and refractive index detection was used to evaluate the absolute molecular weight and distribution of water-soluble chitosan, providing detailed insight into sample heterogeneity and polymer chain distribution.
Keywords: Chitosan, Absolute molecular weight, Polymer molecular weight distribution, Size-exclusion chromatography (SEC), Static Light scattering, Biopolymer characterization
| Product | BeSEC |
| Industry |
Pharmaceutical, Food Analysis |
| Sample | Water-soluble chitosan (polysaccharide) |
| Measurement Type | Molecular weight analysis |
| Measurement Technology |
Size-Exclusion Chromatography (SEC)
|
Introduction
Chitosan is obtained from chitin, a naturally abundant polysaccharide found in shrimp and crab shells and insect exoskeletons. Chemical deacetylation of chitin removes acetyl groups, producing chitosan.
Chitosan’s versatility drives its use across multiple industries:
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Food: Chitosan forms protective films to slow microbial growth and extends shelf life. It also acts as a thickener or emulsifier that improves flow and stability.
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Pharmaceuticals: Chitosan’s biocompatibility and biodegradability make it suitable for drug carriers, sustained-release systems, and wound dressings with hemostatic and antibacterial properties.
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Water Treatment: Chitosan functions as a flocculant for contaminant removal.
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Agriculture: Chitosan serves as a seed coating to enhance germination and disease resistance.
Molecular weight is a critical property influencing chitosan’s performance.
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Low molecular weight: dissolves easily, producing low viscosity solutions, and penetrates membranes more readily, often linked to stronger antimicrobial or antitumor activity.
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High molecular weight: creates viscous solutions, forms stable gels and scaffolds for tissue engineering, and offers higher adsorption capacity for metals and organics, albeit with slower kinetics.
Given these effects, accurate molecular weight evaluation is essential for selecting the proper grade for specific applications.
Experimental Section
This study employed a SEC system with refractive index (RI) and light scattering (LS) detectors. The light scattering detector equipped in the BeSEC LS2 from Bettersize Instruments includes 90° and 7° angles. The BeSEC workstation integrates light scattering with RI or UV signals to calculate molecular weight averages (Mn, Mw and Mz) and distributions.
System Configuration:
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Detector: Light Scattering (LS) + Refractive Index (RI)
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Column: Shodex Ohpak LB-806M
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Mobile phase: Water with 0.2M NaNO3
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Flow rate: 0.7 mL/min
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Injection volume: 100 μL
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Column temperature: 40 °C
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dn/dc: 0.129 mL/g
Sample Preparation:
A water-soluble chitosan sample was weighed and dispersed in 0.2 M NaNO3. After stirring to clarity (2–5 mg/mL), the solution was filtered through a 0.22 μm PES syringe filter and transferred to vials. The high ionic strength mobile phase minimizes electrostatic interactions between chitosan and the column.
Results and Discussion
Figure 1. Elution profiles of the multi-detector signals for Chitosan
Figure 2. Elution profile of the molecular weight for Chitosan
Figure 1 shows chromatograms collected using multiple detectors: refractive index (RI, blue), right-angle light scattering (RALS, green), and low-angle light scattering (LALS, red).
Figures 1 and 2 confirm clean signals with flat baselines, minimal noise, and strong signal-to-noise ratios. The molecular weight profile decreases gradually with elution volume, consistent with SEC principles, where larger species elute first, followed by smaller ones.
The light scattering peaks exhibit no tailing, and the molecular weight curve remains stable without upturn at the end, indicating effective size-based separation.
Table 1. Molecular weight results
| Sample | Mn (Da) | Mw (Da) | Mz (Da) |
| Chitosan |
71,254 |
181,663 | 3,266,638 |
Figure 3. Molecular weight distribution of Chitosan
Conclusion
This application note shows that the analyzed water-soluble chitosan sample possesses a relatively high molecular weight and broad distribution. The tailing peak in the molecular weight profile confirms the presence of a small fraction of larger chains. These findings provide a clear picture of sample heterogeneity and offer valuable guidance for selecting an appropriate chitosan grade for formulation or processing.
About the Authors
 |
Zhibin Guo |
BeSECAdvanced Light Scattering Detector
|
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