Moringa oleifera leaves have been widely used for the treatment of inflammation, diabetes, high blood pressure, and other diseases, due to being rich in polyphenols. The main objective of this work was to largely separate the main polyphenols from Moringa oleifera leaves using the technique of high-speed counter-current chromatography (HSCCC). The phenolic composition in Moringa oleifera leaves was first analyzed qualitatively and quantitatively by UPLC-Q-Exactive Orbitrap/MS and UPLC-QqQ/MS, respectively, indicating that quercetin and kaempferol derivatives, phenolic acid and apigenin are the main polyphenols in Moringa oleifera leaves, with quercetin and kaempferol derivatives predominating. Furthermore, the conditions of HSCCC for large-scale separation of polyphenols from Moringa oleifera leaves were optimized, which included the selection of the solvent system, flow rate and the sample load. Only by one-step HSCCC separation (within 120 min) under the optimized conditions, six quercetin and kaempferol derivatives, a phenolic acid and an apigenin could be individually isolated at a large scale (yield from 10% to 98%), each of which possessed high purity. Finally, the isolated polyphenols and phenolic extract from Moringa oleifera leaves (MLPE) were verified to have strong neuroprotective activities against H2O2-induced oxidative stress in PC-12 cells, suggesting that these compounds would contribute to the main beneficial effects of Moringa oleifera leaves.

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Characterization, Large-Scale HSCCC Separation and Neuroprotective Effects of Polyphenols from Moringa oleifera Leaves

Be restrained by the limited peak capacity, one dimension chromatography usually leads to an unsatisfactory separation with low purity of compounds in a complex mixture. To obtain more highly pure targets for standard reference and to discover new substances for structural elucidation, two-dimensional chromatography is more and more prevalent in many fields. As few metrics on assessment of preparative capability of two-dimensional chromatographic separations is reported, a methodology of in silico screening of various two-dimensional chromatographic separations with a minimal number of experiments was demonstrated in this work, which was based on three descriptors including the occupation rate of peaks and system homogeneity of a two-dimensional separation space, and the minimal distance of all nearest-neighbor distances of peaks. Combining the advantages of counter-current chromatography and liquid chromatography, we elaborated the methodology by employing off-line comprehensive two-dimensional counter-current chromatography with liquid chromatography to be in silico screened for separation of four saponins from Panax notoginseng at analytical scale to simulate the case of preparative scale transfer. The predictive results were presented by two-dimensional contour plots and verified by experiments. The result showed that the experimental results were in general accord with the predictive results. This article is protected by copyright. All rights reserved.

In Silico Screening of Off-line Comprehensive Two-Dimensional Counter-current Chromatography with Liquid Chromatography for Four Saponins Isolation.

Sustainable approaches for the study of alkaloids from plants using supercritical fluid-based processes

The separation regularity of the three-phase solvent system of counter-current chromatography based on polarity parameter modeling

The separation regularity of the three-phase solvent system of counter-current chromatography based on polarity parameter modeling.

Piper longum L. (Piperaceae) is famous for flavoring food and treatment purposes. N-Alkylamides (NAAs) are the most common secondary metabolites found in Piper species with broad spectrum of biological properties. Because of the similar structures and lower polarities, it is difficult to separate NAAs by traditional chromatographic methods. Herein, an efficient off-line two-dimensional coupled with gradient high-speed counter-current chromatography (HSCCC) method is reported for preparative separation of N-alkylamides from P. longum. The HSCCC solvent system with n-hexane/ethyl acetate/methanol/water (5:4:5:4, v/v) was used for the first dimensional separation. Two pure compounds i.e. piperlonguminine (1) and piperine (2) were successfully separated, coupled with an enriched trace ingredient. Furthermore, the enriched ingredient was used for the second dimensional separation with a gradient HSCCC mode by adjustment of the mobile phase with more enhanced elution ability. Eight minor compounds were successfully separated including retrofractamide A (3), retrofractamide C (4), dehydropipernonaline (5), pipernonaline (6), piperundecalidine (7), guineesine (8), brachystamide B (9), and (2E,4E,12Z)-N-isobutyl-2,4,12- octadecatrienamide (10). The structures of the isolated pure compounds were characterized by ESI-MS, 1H, and 13C NMR spectral analysis.

Efficient separation of N-Alkylamides from Piper longum L. using off-line two-dimensional coupled with gradient high-speed counter-current chromatography

The invention relates to the technical field of the separation and purification of effective constituents in traditional Chinese medicines, and specifically relates to a method of separating and purifying adenosine from trichosanthes kirilowii maxim. The method includes peeling the trichosanthes kirilowii maxim, and performing drying, crushing, distilled water reflux extraction, alcohol precipitation and supernatant concentration to prepare a crude trichosanthes kirilowii maxim product; performing pre-purification on the crude trichosanthes kirilowii maxim product through macroporous resin, and preparing a crude adenosine product after pre-purification; and using high speed countercurrent chromatography to separate and purify the crude adenosine product so as to obtain adenosine monomer ofwhich purity is higher than 98%, wherein a solvent system adopted by the high speed countercurrent chromatography is a tert-butyl methyl ether/n-butanol/acetonitrile/water system. The preparation method is low in cost, simple in operation and high in efficiency, can separate the adenosine monomer with high purity (>98%) from the trichosanthes kirilowii maxim in large batches and has good practical application values.

Li Ge

Papers

Efficient separation of N-Alkylamides from Piper longum L. using off-line two-dimensional coupled with gradient high-speed counter-current chromatography

Preparation method of high-purity adenosine in trichosanthes kirilowii maxim