A new amine isolated from Urtica thunbergiana Siebold & Zucc.

One new amine 2-dimethyl-Penidilamine (1), together with seventeen known compounds (2-18) were isolated from the 95% ethanol extract of Urtica thunbergiana Siebold & Zucc. Their structures were characterised by extensive spectroscopic analysis including NMR, mass spectra and single X-ray crystallography. Among them, compound 1 is a new compound, and compounds 3, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 17 and 18 were isolated from Urtica thunbergiana Siebold & Zucc for the first time. Among them, compound 1, 10, 15, 17 and 18 exhibited significant α-glucosidase inhibitory activity with an IC50 value of 65.12 µM, 7.42 µM, 26.24 µM, 71.31 µM and 72.55 µM, respectively. Our study provided the scientific report for the medicinal value of Urtica thunbergiana Siebold & Zucc.

Large-scale preparative isolation of bergenin standard substance from Saxifraga atrata using polyamide coupled with MCI GEL® CHP20P as stationary phases in medium pressure chromatography.

In this study, polyamide and MCI GEL® CHP20P were employed as stationary phases in medium pressure chromatography (MPC) for the efficient preparative separation of bergenin from Saxifraga atrata. Ethanol-water, methanol-water, and acetonitrile-water mobile phases all showed good enrichment capacity for bergenin fraction when polyamide was used as a stationary phase. After 5 cycles of polyamide MPC using acetonitrile/water, 1.2 g of bergenin fraction was isolated from 180 g Saxifraga atrata herb. Further purification of this fraction was conducted using MCI GEL® CHP20P styrene-divinylbenzene beads. The bergenin fraction was separated into two fractions, and after three runs of MPC, 714.2 mg of bergenin with purity above 99% was obtained. The results demonstrate that the combination of polyamide and styrene-divinylbenzene MPC can be utilized for preparative isolation of compounds from natural products with high yield and purity.

Targeted isolation of 1,1-diphenyl-2-picrylhydrazyl inhibitors from Saxifraga atrata using medium- and high- pressure liquid chromatography combined with online high performance liquid chromatography-1,1-diphenyl-2- picrylhydrazyl detection.

Traditional Tibetan medicine (TTM) is a valuable source of novel therapeutic lead molecules inspired by natural products (NPs). The health benefits of Saxifraga atrata are well documented in TTM, but reports on its chemical composition are limited, most likely due to the complicated purification process. Herein, target separation and identification of 4 main radical scavenging compounds from the methanolic extract of S. atrata was were performed using medium- and high-pressure liquid chromatography coupled with online HPLC-DPPH detection. The sample was pretreated using medium pressure liquid chromatography with MCI GELⓇ CHP20P styrene-divinylbenzene beads as a stationary phase, yielding 1.4 g of the target DPPH inhibitors (Fr4, 11.9% recovery). The compounds were further purified and isolated using HPLC on RP-C18 (ReproSil-Pur C18 AQ) followed by HILIC (Click XIon) column separation, resulting in 2.8 mg of fraction Fr4-1-1, 6.8 mg of fraction Fr4-2, 244.9 mg of the Fr4-3-1 sample, and 38.3 mg of Fr4-4-1. The structure and purity of the target compounds were determined, and four compounds (ethyl gallate, 11-O-galloylbergenin, rutin and isoquercitrin) were isolated with >95% purity. The developed methodology is efficient for targeted isolation of high-purity radical scavengers from NP extracts and could be used for rapid identification and isolation of DPPH inhibitors from various NPs.

An efficient strategy based on liquid-liquid extraction and pH-zone-refining counter-current chromatography for selective enrichment, separation, and purification of alkaloids and organic Acids from natural products.

This study presents an efficient strategy based on liquid-liquid extraction and pH-zone-refining counter-current chromatography for selective enrichment, separation, and purification of alkaloids and organic acids from natural products. First, an acid or base modified two-phase solvent system with maximum or minimum partition coefficient was developed for the liquid-liquid extraction of the crude extract. As a result, alkaloids or organic acids could be selectively enriched in the upper or lower phase. Then pH-zone-refining counter-current chromatography was employed to separate and purify the selectively enriched alkaloids or organic acids efficiently. The selective enrichment and separation of five bufadienolide from toad venom of Bufo marinus was used as an example to show the advantage of this strategy. As a result, 759 mg of selectively enriched bufadienolide was obtained from 2 g of crude extract and the total content of five targets was increased from 14.64 to 83%. A total of 31 mg of marinobufagin-3-adipoyl-l-arginine, 42 mg of telocinobufagin-3-pimeloyl-l-arginine, 51 mg of telocinobufagin-3-suberoyl-l-arginine, 132 mg of marinobufagin-3-suberoyl-l-arginine, and 57 mg of bufalin-3-suberoyl-l-arginine were all simultaneously separated from 500 mg of selectively enriched sample, with the purity of 92.4, 97.5, 90.3, 92.1, and 92.8%, respectively.

Preparative isolation of arylbutanoid-type phenol [(-)-rhododendrin] with peak tailing on conventional C18 column using middle chromatogram isolated gel column coupled with reversed-phase liquid chromatography.

Reversed-phase liquid chromatography coupled with middle chromatogram isolated gel column was employed for the efficient preparative separation of the arylbutanoid-type phenol [(-)-rhododendrin] from Saxifraga tangutica. Universal C18 (XTerra C18) and XCharge C18 columns were compared for (-)-rhododendrin fraction analysis and preparation. Although tailing and overloading occurred on the XTerra C18 column, the positively charged reversed-phase C18 column (XCharge C18) overcame these drawbacks, allowing for favorable separation resolution, even when loading at a on a preparative scale (3.69 mg per injection). The general separation process was as follows. First, 365.0 mg of crude (-)-rhododendrin was enriched from 165 g Saxifraga tangutica extract via a middle chromatogram isolated gel column. Second, separation was performed on an XTerra C18 preparative column, from which 73.8 mg of the target fraction was easily obtained. Finally, the 24.0 mg tailing peak of (-)-rhododendrin on XTerra C18 column was selectively purified on the XCharge C18 analytical column. These results demonstrate that the tailing nonalkaloid peaks can be effectively used for preparative isolation on XCharge C18 columns.

Preparative isolation of highly polar free radical inhibitor from Floccularia luteovirens using hydrophilic interaction chromatography directed by on-line HPLC-DPPH assay.

Wild edible macro fungi Floccularia luteovirens proved to be a valuable source for the identification of novel lead molecules with therapeutic potential. Nevertheless, the chemical constituents of Floccularia luteovirens are rarely reported due to absence of efficient purification methods. In this study, a hydrophilic interaction chromatography directed by on-line HPLC-DPPH assay has been developed and successfully applied for the isolation of free radical inhibitor from the methanolic extract of Floccularia luteovirens. Using a hydrophilic interaction chromatographic column coupled with the HPLC-DPPH assay for screening the potential radical scavengers, the mid-pressure hydrophilic interaction chromatography (HILIC) proved to be more efficient in the pretreatment stage, yielding the fraction rich in free radical scavengers in good yield (5.9% recovery from 130.0 g of fresh F. luteovirens). From highly potent fraction, the target compound was isolated using the Click XION preparative chromatography with 17.2% recovery. The isolated compound was L-(+)-ergothioneine, where the purity (>95%) and antioxidant activity of were confirmed by chromatography and HPLC-DPPH assay, while the structure of this compound was elucidated from HR ESI-MS and NMR data. This method proved to be very efficient for the recognition and isolation of highly polar free radical inhibitors from fungi extracts, and is also applicable for the purification of highly polar compounds from other sources.