Abstract
A method for the quantitative determination of ganoderic acid A was constructed using the principle of indirect competitive enzyme-linked immunosorbent assay (ELISA), and this method was used to determine the ganoderic A contents of Ganoderma lucidum samples in the market. The conjugate of ganoderic acid A and bovine serum albumin was used for four rounds of immunization on test rabbits to obtain rabbit antiganoderic acid A antibody IgG. The enzyme-labeled plate was coated with the conjugate of ganoderic acid A and ovalbumin. The first stage reaction in the indirect competitive ELISA was that the conjugate of ganoderic acid A in the sample competed with the conjugate coated on the enzyme-labeled plate to bind rabbit antibodies. The second stage reaction was the combination of goat anti-rabbit IgG–horseradish peroxidase and rabbit antiganoderic acid A antibody IgG. The results of the determination of ganoderic acid A standard by this method showed that the coefficient of variation of repeated wells in the group was <5%, the detection limit of ganoderic acid A was 0.6 μg/L, and ganoderic acid A had a substantial dose-response relationship in the content range of 0.9–72.9 μg/L (R2 = 0.994). This method was used to measure the ganoderic A content of 12 varieties of G. lucidum in the market and showed the obvious differences in the ganoderic acid A contents of the different varieties. This method is simple, fast, and of great importance to the quality control of Ganoderma products.
Highlights
Ganoderma is rich in a variety of active ingredients, and its pharmacological effects are different
The detection methods of ganoderic acid A include visible light colorimetry [15], UV spectrophotometry [16, 17], high-performance liquid chromatography [18,19,20,21], and chromatography-mass spectrometry [22, 23]. e main drawbacks of these methods are the high cost of the highly specialized instrument, difficulty in popularizing the method, and long detection time. erefore, providing a simple and rapid detection method of ganoderic acid A is an urgent problem that needs to be solved. e purpose of this research was to use the principle of enzyme-linked immunosorbent assay (ELISA) to construct a method for the quantitative determination of ganoderic acid A and determine and analyze the ganoderic acid A contents of different G. lucidum varieties in the market
ELISA includes antigen coating, sample loading reaction, and substrate color development. e method used in this experiment was indirect competitive ELISA. e sample loading reaction involved two stages. e first stage is Journal of Food Quality the competition between the ganoderic acid A in the sample and the conjugate coated on the enzyme-labeled plate to bind rabbit antiganoderic acid A antibody IgG, and the second stage is the combination of goat anti-rabbit IgGhorseradish peroxidase (HRP) and rabbit antiganoderic acid A antibody IgG
Summary
Ganoderma is rich in a variety of active ingredients, and its pharmacological effects are different. Triterpene is one of the main active components of Ganoderma lucidum. Ganoderic acid A accounts for the largest proportion of G. lucidum triterpenes and is one of the Ganoderma triterpenoid markers [13]. Erefore, the development of a fast and simple method with a low detection limit for the determination of ganoderic acid A is of great importance for the quality control of Ganoderma products. E purpose of this research was to use the principle of enzyme-linked immunosorbent assay (ELISA) to construct a method for the quantitative determination of ganoderic acid A and determine and analyze the ganoderic acid A contents of different G. lucidum varieties in the market. ELISA includes antigen coating, sample loading reaction, and substrate color development. ELISA includes antigen coating, sample loading reaction, and substrate color development. e method used in this experiment was indirect competitive ELISA. e sample loading reaction involved two stages. e first stage is Journal of Food Quality the competition between the ganoderic acid A in the sample and the conjugate coated on the enzyme-labeled plate to bind rabbit antiganoderic acid A antibody IgG, and the second stage is the combination of goat anti-rabbit IgGhorseradish peroxidase (HRP) and rabbit antiganoderic acid A antibody IgG
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