Abstract
Scutellariae radix (S. radix) is one of the most important crude drugs used in Kampo medicines (KMs). A part of its pharmaceutical properties is due to flavone glycoside, baicalin (BI). A technique named eastern blotting was developed for the specific and easy identification of BI in the extracts of crude drugs and KMs using anti-BI monoclonal antibody (MAb). BI separated by silica gel thin-layer chromatography (TLC) transferred to a polyethersulfone (PES) membrane was treated with a NaIO4 solution and reacted with bovine serum albumin (BSA) preparing BI-BSA conjugate on the PES membrane. Anti-BI MAb was bound and then antibody labeled with peroxidase directed against anti-BI MAb. Finally, a substrate was added and then BI was detected. As little as 1 mg of BI was still detected on the PES membrane under immunostaining method. Various samples of S. radix and KMs which contain S. radix were qualitatively analyzed, and BI was visually detected by eastern blotting technique. Furthermore, this method was applied for the immunohistochemical study to investigate the distribution of BI in the fresh root of Scutellaria baicalensis using immunoblotting by transferred from fresh root to the PES membrane.
Highlights
Scutellaria baicalensis Georgi (Labiatae) is a well-known herb and widely found in East Asian from the north of China to east of Siberia
Anti-BI monoclonal antibody (MAb) was applied to the immunostaining technique for the visual detection of BI on the membrane
The specificity of anti-BI MAb was tested in a dot blot assay using BI and other glycosides such as flavonoids and terpenoids
Summary
Scutellaria baicalensis Georgi (Labiatae) is a well-known herb and widely found in East Asian from the north of China to east of Siberia. Immunoassay systems using monoclonal antibodies (MAbs) against bioactive compounds of low molecular weight have become important tools for the qualitative and quantitative analyses of active constituents from natural medicines and natural products [14]. In the investigation of staining technique for several low molecular weight compounds such as natural glycosides and phospholipids, the first successful immunostaining of natural glycosides on polyvinylidene difluoride (PVDF) membrane was reported on solasodine glycosides by our group following to the thin-layer chromatography blotting procedure of phospholipids [17,18]. As an extension of anti-BI MAb application, the present paper describes the development of eastern blotting technique for the specific analysis of BI in S. radix and KMs. immunohistochemical study to investigate the distribution of BI in the fresh root of S. baicalensis was demonstrated using immunoblotting by transferred from fresh root to the PES membrane
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