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Determining Lentinan Molecular Weight Using the BeSEC

2026-01-15Application Note

Abstract: Lentinan is a bioactive polysaccharide whose molecular weight strongly influences its immunomodulatory and functional properties. In this study, size-exclusion chromatography coupled with static light scattering and refractive index detection was employed to determine the absolute molecular weight and molecular weight distribution of lentinan samples, enabling clear differentiation among grades and supporting application-specific selection.

Keywords: Lentinan, Polysaccharide, Absolute molecular weight, Molecular weight distribution, Size-exclusion chromatography (SEC), Static Light scattering

Product	BeSEC
Industry	Pharmaceutical, Food Analysis
Sample	Lentinan (polysaccharide extracted from shiitake mushrooms)
Measurement Type	Absolute molecular weight and molecular weight distribution
Measurement Technology	Size-Exclusion Chromatography (SEC), Static Light Scattering

Introduction

Lentinan is a bioactive polysaccharide extracted from shiitake mushrooms, widely recognized for its immunomodulatory effects. It is commonly used as an adjuvant in cancer therapy to ease the side effects of chemotherapy and radiotherapy. Beyond oncology, lentinan exhibits antiviral and antioxidant activity and is frequently incorporated into health supplements to enhance immunity and slow aging. In food applications, it improves texture and stability while adding nutritional value.

Molecular weight plays a critical role in lentinan’s functionality. Higher molecular weight molecules tend to adopt more ordered structures and bind more effectively to immune cell receptors, resulting in stronger immunomodulatory effects. Lower molecular weight molecules dissolve and absorb more easily, but their biological activity may be shorter-lived.

Selecting the appropriate molecular weight is essential for optimizing performance in specific applications.

Experimental Section

This study utilized a Size Exclusion Chromatography (SEC) system equipped with refractive index (RI) and light scattering (LS) detectors. The light scattering detector is the BeSEC LS2 from Bettersize Instruments, with 90° and 7° angles. The BeSEC workstation combines light scattering with RI or UV signals to calculate molecular weight averages (Mn, Mw and Mz) and distributions.

System Configuration:

Detectors: Light Scattering (LS) + Refractive Index (RI)
Column: Shodex Ohpak LB-806M
Mobile phase: 0.05 M NaNO3 aqueous solution
Flow rate: 0.7 mL/min
Injection volume: 100 μL
Column temperature: 40 ℃
dn/dc: 0.129 mL/g

Sample Preparation:
Five lentinan samples were analyzed. Each powder was accurately weighed and dispersed in 0.05 M NaNO3, stirred until clear (1 to 3 mg/mL), filtered through a 0.22 μm PES syringe filter, and transferred into vials and placed in the autosampler for measurement.

Results and Discussion

Figure 1. Elution profiles of the multi-detector signals (top) and molecular weight distribution (bottom) for Sample A

Figure 2. Elution profiles of the multi-detector signals (top) and molecular weight distribution (bottom) for Sample B

Figure 3. Elution profiles of the multi-detector signals (top) and molecular weight distribution (bottom) for Sample C

Figure 4. Elution profiles of the multi-detector signals (top) and molecular weight distribution (bottom) for Sample D

Figure 5. Elution profiles of the multi-detector signals (top) and molecular weight distribution (bottom) for Sample E

Figures 1 to 5 show chromatograms of the five lentinan samples. The refractive index (RI) signal is shown in blue, the right-angle light scattering (RALS) signal in green, and the low-angle light scattering (LALS) signal in red. The molecular weight distribution, plotted as a purple line, represents molecular weight as a function of elution time.

Across all samples, the signals are clean with flat baselines, minimal noise, and strong signal-to-noise ratios. In Figure 1, the molecular weight profile decreases steadily with elution volume, consistent with SEC principles, larger species eluting first followed by smaller ones. Scattering peaks exhibit no tailing and the molecular weight curve remains stable at the end, indicating effective size-based separation.

Table 1. Molecular weight results of lentinan samples

No.	Mn (kDa)	Mw (kDa)	Mz (kDa)	Mw/Mn
Sample A	3559	4465	6128	1.25
Sample B	3218	6222	23741	1.37
Sample C	2903	4168	5676	1.13
Sample D	150.8	204.5	360.45	1.3
Sample E	29.2	37.1	47.5	1.27

Table 1 summarizes the molecular weight results for all five samples. Significant differences in molecular weight were observed, which directly impact solubility, viscosity, biological activity, and formulation behavior. Consequently, each sample may be better suited for different applications.

Conclusion

This study demonstrates the use of the BeSEC LS2 with light scattering detection to characterize the molecular weight of lentinan samples. The results reveal substantial variation in molecular weight among the five samples, providing critical insight for selecting the appropriate grade for pharmaceutical, nutraceutical, or food applications.

About the Authors

Bettersize-application-engineer-Zhibin-Guo

Zhibin Guo

Application Research Lab, Bettersize Instruments Ltd.

BeSEC

Advanced Light Scattering Detector

● For proteins: molecular weight, oligomer state, and aggregate characterization
● For polymers and polysaccharides: molecular weight distribution and size
● No column calibration required since Mw is independent of elution volume
● Low-angle detection enables accurate molecular weight without model assumptions or angle fitting
● Compatible with any GPC or SEC system

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