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  • Author or Editor: Zhengtao Wang x
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A simple and accurate high-performance thin-layer chromatography (HPTLC)-bioautographic method was developed for the quantitative analysis of magnolol and honokiol in the herbal medicine Magnoliae officinalis Cortex. The samples were separated on a silica gel HPTLC plate with a mixture solution of toluene-methanol (10:1, v/v) as the mobile phase. Spots were visualized by dipping in 2,2-diphenyl-1-picrylhydrazyl radical (DPPH*) reagent and measured at a wavelength of 550 nm in a reflection mode, scanning after derivatization for 40 min. The method had excellent linearity (r 2 = 0.9939 for magnolol and r 2 = 0.9989 for honokiol, respectively) in the concentration range of 0.16–0.97 mg spot−1 for both analytes. The recoveries were 94.5–105.9% for magnolol and 86.6–103.4% for honokiol, respectively. The established HPTLC-bioautographic method was evaluated comprehensively in quantitative and antioxidant activity analysis of magnolol and honokiol in Magnoliae officinalis Cortex and various plants.

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In traditional Chinese medicine, plants of the genus Peganum have been used to treat cough, hypertension, diabetes, asthma, jaundice, lumbago, and many other ailments. In this study, seeds of the plants of genus Peganum, including P. harmala Linn., P. multisectum (Maxim) Bobr, P. nigellastrum Bunge, and Peganum variety were collected from different provinces in China. A simple, rapid, and effective thin-layer chromatographic (TLC) fingerprint combined with bioautographic technique has been established for the identification of acetylcholinesterase (AChE) inhibitors from these seeds. The methanol extracts of seeds were separated on silica gel plates with ethyl acetate-methanol-ammonia 10:1.5:0.5 (ν/ν) as mobile phase, and then the plates were inspected under UV 366 nm and visualized by spraying with both Dragendorff’s and vanillin-sulfuric acid reagents as well as by bioautographic assay. Moreover, the limits on AChE inhibitive activity of harmine and harmaline were found to be 0.01 μg, in comparison to that of galanthamine of also 0.01 μg. The TLC fingerprints combined with the bioautographic method could distinguish the seeds of the different species of genus Peganum investigated. Moreover, harmine and harmaline displayed similar AChE inhibition compared to galanthamine.

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Summary

A thin-layer chromatography (TLC)–bioautography–mass spectrometry (MS) method coupled with a liquid chromatography–MS-controlled autopurification system was developed and applied for screening and isolating natural dipeptidyl peptidase IV (DPP IV) inhibitor from plant extracts. An unknown constituent with potential DPP IV inhibitory activity from bulbs of Fritillaria cirrhosa was discovered using TLC–bioautography, followed by using a TLC interface with mass spectrometry to obtain m/z of the target compound, and the purification of the compound was directly achieved with a mass-directed autopurification system for 40 runs of injection. Finally, 2.1 mg of the compound was obtained and identified as peimisine by 1H and 13C nuclear magnetic resonance (NMR) spectroscopy. The half maximal inhibitory concentration (IC50) of DPP IV inhibitory activity was determined at 80.5 µm comparing to 58.0 µm of the standard DPP IV inhibitor diprotin A by a spectrophotometric method.

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