Study on Quality Control Markers in Commercial Ganoderma lucidum Products
Date Issued
2015
Date
2015
Author(s)
KAO, YA-LING
Abstract
Ganoderma lucidum, a medicinal fungi, has been used in Traditional Chinese Medicine (TCM) for its health promoting constituents, including polysaccharides, proteins, triterpenes, sterols, and fatty acids. However, G. lucidum contains various and complex bioactive compounds. It was difficult to distinguish the bioactivity of commercial G. lucidum products from specific component. Most products were claimed on the basis of their total sugar or crude polysaccharides contents, but immunomodulatory protein (LZ-8) and (1,3)-β-D-glucan, were not shown in the nutrition facts. In this study, we decided to examine whether the contents of total sugar or (1,3)-β-D-glucan could be used as the quality and functional indicators, and what would be the appropriate concentrates for the evaluation in product activities. First, we determined content of total sugar and content of (1,3)-β-D-glucan in commercial products with phenol-sulfuric acid method and fluorometric method. Our results showed that there were significant differences on the contents of total sugar and (1,3)-β-D-glucan in the eight commercial products. Samples with the same contends of total sugar or (1,3)-β-D-glucan had different capability to induce secreation of TNF-α and IL-6 in murine macrophages and RAW 264.7. Therefore, the quantity of total sugar and (1,3)-β-D-glucan might not be an appropriate functional indicators. Our results also revealed that the approach of diluting the commercial product solutions which were cultured with murine macrophages to 30 to 250 mg commercial product in 100 mL PBS could be used to distinguish bioactivities of commercial products. Further, the eight commercial products could not increase production of cytokine (IFN-γ, IL-2, and IL-4) by murine splenocytes. In addition, SDS-PAGE and Western blotting analysis of these samples showed that commercial G. lucidum products could rarely include immunomodulatory protein LZ-8. In the third part, mycelia from G. lucidum were lysed by ultrasonic reactor and analyzed for the content of LZ-8 by HPLC. LZ-8 peak have been observed as lysis solution filtered through glass fiber membranes and added 100 μg/mL standard LZ-8 to the solution. As shown above, we assumed that the component which interfered the HPLC analysis of LZ-8 should be polysaccharide. Sample solution was further treated with exo-(1,3)-β-D-glucanase and endo-(1,3)-β-D-glucanase in order to remove the potented interfering substance, but no LZ-8 peak have been observed. As a result, it was inferred that the interfering substance might be polysaccharides but not (1,3)-β-D-glucans. In conclusion, the content of total sugar, (1,3)-β-D-glucan, and LZ-8 could not be the functional indicators. The appropriate dosages of sample cultured with murine macrophages were 30 to 250 mg of commercial product in 100 mL PBS and could be used to assess product activities. In the third part, we inferred that polysaccharids might be the main factor to interfer the HPLC analysis of the free form of LZ-8.
Subjects
(1,3)-β-D-glucan
Immunomodulation
G. lucidum mycelia
Type
thesis
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