High-performance Countercurrent Chromatography to Access Rhodiola rosea Influenza Virus Inhibiting Constituents

Abstract

In a cytopathic effect inhibition assay, a standardized Rhodiola rosea root and rhizome extract, also known as roseroot extract (SHR-5), exerted distinct anti-influenza A virus activity against HK/68 (H3N2) (IC50 of 2.8 µg/mL) without being cytotoxic. For fast and efficient isolation and identification of the extractʼs bioactive constituents, a high-performance countercurrent chromatographic separation method was developed. It resulted in a three-stage gradient elution program using a mobile phase solvent system composed of ethyl acetate/n-butanol/water (1 : 4 : 5 → 2 : 3 : 5 → 3 : 2 : 5) in the reversed-phase mode. The elaborated high-performance countercurrent chromatographic method allowed for fractionation of the complex roseroot extract in a single chromatographic step in a way that only one additional orthogonal isolation/purification step per fraction yielded 12 isolated constituents. They cover a broad polarity range and belong to different structural classes, namely, the phenylethanoid tyrosol and its glucoside salidroside, the cinnamyl alcohol glycosides rosavin, rosarin, and rosin as well as gallic acid, the cyanogenic glucoside lotaustralin, the monoterpene glucosides rosiridin and kenposide A, and the flavonoids tricin, tricin-5-O-β-D-glucopyranoside, and rhodiosin. The most promising anti-influenza activities were determined for rhodiosin, tricin, and tricin-5-O-β-D-glucopyranoside with IC50 values of 7.9, 13, and 15 µM, respectively. The herein established high-performance countercurrent chromatographic protocol enables fast and scalable access to major as well as minor roseroot constituents. This is of particular relevance for extract standardization, quality control, and further in-depth pharmacological investigations of the metabolites of this popular traditional herbal remedy.

Antiviral Activity of the PropylamylatinTM Formula against the Novel Coronavirus SARS-CoV-2 In Vitro Using Direct Injection and Gas Assays in Virus Suspensions

“…Although numerous preclinical and clinical investigations have been conducted to identify effective drug candidates for the treatment and prevention of COVID-19, there are currently no antiviral agents approved for this serious disease. Natural products derived from plant, fungi, and marine sources have been, and are still, frequently used to treat viral infections, particularly those that cause respiratory illnesses such as rhinovirus and influenza virus [ 2 , 5 , 7 , 8 , 9 , 10 , 11 ]. Natural products are attractive therapies for viral as well as other microbial infections because they are often accessible in nature (making them low in cost), amenable to mass production, and have generally good safety profiles [ 9 ].…”

A supercritical fluid workflow for the quality assessment of herbal drugs and commercial preparations from Rhodiola rosea

“…Previously, seven constituents from R. rosea roots and rhizomes were isolated from a standardised 70% ethanolic extract (SHR‐5): p ‐tyrosol ( 1 ), rosin ( 2 ), rosiridin ( 3 ), salidroside ( 4 ), rosarin ( 5 ), rosavin ( 6 ), and tricin‐5‐ O ‐β‐ d ‐glucopyranoside ( 7 ) 28 . For isolation procedures, the dry powder of the SHR‐5 extract (batch no.…”

“…For isolation procedures, the dry powder of the SHR‐5 extract (batch no. 1521229, Voucher specimen JR‐20180904‐A1 deposited at the Department of Pharmacognosy, University of Vienna, Austria) was applied to high‐performance counter current chromatography gradient elution 28 . Isolated compounds were identified by the interpretation of one‐dimensional (1D) and two‐dimensional (2D) NMR and electrospray ionisation‐mass spectrometry (ESI‐MS) experiments.…”

“…Previously, seven constituents from R. rosea roots and rhizomes were isolated from a standardised 70% ethanolic extract (SHR‐5): p ‐tyrosol ( 1 ), rosin ( 2 ), rosiridin ( 3 ), salidroside ( 4 ), rosarin ( 5 ), rosavin ( 6 ), and tricin‐5‐ O ‐β‐ d ‐glucopyranoside ( 7 ). 28 For isolation procedures, the dry powder of the SHR‐5 extract (batch no. 1521229, Voucher specimen JR‐20180904‐A1 deposited at the Department of Pharmacognosy, University of Vienna, Austria) was applied to high‐performance counter current chromatography gradient elution.…”

“…rosin ( 2 ), rosarin ( 5 ), and rosavin ( 6 ), the monoterpene glucoside rosiridin ( 3 ) as well as the phenylethanoid p ‐tyrosol ( 1 ) and its glucoside salidroside ( 4 ) are well established marker compounds (Figure 1 ). Although tricin‐5‐ O ‐β‐ d ‐glucopyranoside ( 7 ) has not been considered in previous standardisation studies of Rhodiola , it was included in the present study for two reasons: (i) for its importance as anti‐influenza A virus active, 28 α‐amylase 29 and NO production inhibiting 30 compound and (ii) due to its polarity which increased the challenge for the development of SFx based protocols for extraction and separation. The established methods were validated and applied for the quantitation of 1 to 7 in 31 R. rosea samples (commercial products as well as herbal drugs).…”