Zhongjun Ma

Physalins with anti-inflammatory activity are present in Physalis alkekengi var. franchetii and can function as Michael reaction acceptors

Michael reaction acceptors (MRAs) are a class of active molecules that are directly or indirectly involved in various cellular processes, including the regulation of many signaling pathways. In this study, the inducible nitric oxide synthase (iNOS) assay was used to demonstrate that the dichloromethane extract of Physalis alkekengi var. franchetii (DCEP) possesses anti-inflammatory activity that might be attributed to the modification of key cysteine residues in IKKβ by the MRAs in DCEP. To isolate these MRAs, glutathione (GSH) was employed, and a simple ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) screening method was developed to investigate the GSH conjugates with potential MRAs. Five physalins, including one new compound isophysalin A (2), together with four known steroidal compounds, physalin A (1), physalin O (3), physalin L (4) and physalin G (5), were isolated to evaluate the GSH conjugating abilities, and it was indicated that compounds 1, 2 and 3, which had a common α,β-unsaturated ketone moiety, exhibited conjugating abilities with GSH and also showed significant nitric oxide (NO) production inhibiting activities. The anti-inflammatory activities of compounds 1, 2 and 3 might be attributed to their targeting multiple cysteine residues on IKKβ; therefore, the alkylation of IKKβ by compound 1 was further studied by micrOTOF-MS. The result showed that six cysteine residues (C(59), C(179), C(299), C(370), C(412), and C(618)) were alkylated, which indicated that IKKβ is a potential target for the anti-inflammatory activity of physalin A.

Terpenoids from Tripterygium wilfordii

An abietane diterpenoid, triptobenzene Y and six sesquiterpene polyol esters, wilforsinines C-H, together with 14 known compounds, have been isolated from the roots of Tripterygium wilfordii. The structures of the compounds were elucidated on the basis of spectroscopic analyses. The quinone reductase (QR) induction assay indicated that two compounds showed moderate QR-inducing activities at concentrations of 25 μM and 50 μM, respectively.

Silver complexation and tandem mass spectrometry for differentiation of triterpenoid saponins from the roots of Pulsatilla chinensis (Bunge) Regel

For detection and differentiation of two types of triterpenoid saponins based on different aglycons of the lupane and oleanane skeleton from the roots of Pulsatilla chinensis (Bunge) Regel, the silver ion was introduced and electrospray ionization multi-stage tandem mass spectrometry was applied to analyze eleven triterpenoid saponin silver complexes. The quasi-molecular ion [M+Ag](+) was observed in the full-scan MS spectra of all the silver complexes. The MS(2) data of the [M+Ag](+) ion provided structural information on the sugar sequence of the oligosaccharide chains and the aglycon of the saponins. There are two patterns in the cleavage pathway of oleanane-type saponins. One is elimination of the sugar chain and subsequent loss of the carboxylic group which is the same as the cleavage of lupine-type saponins. The other is loss of the distinguishing ions at m/z 72 and 28 (C(2)H(4)) followed by loss of the carboxylic group. Diagnostic fragmentation pathways of the silver complexes of the saponins allow successful identification of the two types of saponins from the roots of Pulsatilla chinensis (Bunge) Regel.

Chemical constituents from the radix of Curcuma wenyujin.

Phytochemical study on the ethanol extract of the radixes of Curcuma wenyujin Y. H. Chen et C. Ling led to the isolation of three new compounds curcuminol F (1) curcuminol G (2) and wenyujinoside (3) and a known compound aurantiamide (4). Their structures were elucidated on the basis of extensive spectroscopic analysis.

Physalin A exerts anti-tumor activity in non-small cell lung cancer cell lines by suppressing JAK/STAT3 signaling

The signal transducers and activators of transcription 3 (STAT3) signaling pathway plays critical roles in the pathogenesis and progression of various human cancers, including non-small cell lung cancer (NSCLC). In this study, we aimed to evaluate the therapeutic potential of physalin A, a bioactive withanolide derived from Physalis alkekengi var. francheti used in traditional Chinese medicine, was evaluated in human NSCLC cells. Its and determined whether it effect oninhibited both constitutive and induced STAT3 activity, through repressing the phosphorylation levels of JAK2 and JAK3, resulting in anti-proliferation and pro-apoptotic effects on NSCLC cells was also determined, and. theThe antitumor effects of physalin A were also validated usingin an in vivo mouse xenograft models of NSCLC cells. Physalin A had anti-proliferative and pro-apoptotic effects in NSCLC cells with constitutively activated STAT3; it also suppressed both constitutive and induced STAT3 activity by modulating the phosphorylation of JAK2 and JAK3. Furthermore, physalin A abrogated the nuclear translocation and transcriptional activity of STAT3, thereby decreasing the expression levels of STAT3, its target genes, such as Bcl-2 and XIAP. Knockdown of STAT3 expression by small interfering RNA (siRNA) significantly enhanced the pro-apoptotic effects of physalin A in NSCLC cells. Moreover, physalin A significantly suppressed tumor xenograft growth. Thus, as an inhibitor of JAK2/3-STAT3 signaling, physalin A, has potent anti-tumor activities, which may facilitate the development of a therapeutic strategy for treating NSCLC.

Phytochemical analysis of the triterpenoids with cytotoxicity and QR inducing properties from the total tea seed saponin of Camellia sinensis.

The tea seed triterpene saponin (TS) from Camellia sinensis was found to exhibit better antitumor activity in vivo in S180 implanted ICR mice and QR inducing activity for hepa lclc7 cells respectively compared with the total tea seed saponin (TTS), hydrolysate of the TTS and tea seed flavonoid glycosides (TF). By bioassay-guided isolation, the TS fraction was separated and seven major components were purified and identified as theasaponin E1 (1), theasaponin E2 (2), theasaponin C1 (3), assamsaponin C (4), theasaponin H1 (5), theasaponin A9 (6), and theasaponin A8 (7), among which compounds 4 and 5 were isolated from this genus for the first time. The antitumor bioassay of the isolated compounds showed that compounds 1, 2 and 3 exhibited potential activities against the human tumor cell lines K562 and HL60. Furthermore, compound 1 (the major constituent with a mass content of over 1%) showed significant QR inducing activity with an IR value of 4.2 at 4μg/ml. So it can be concluded that tea seed especially the compound 1 (theasaponin E1) could be used as an antitumor agent and a chemoprevention agent of cancer. The preliminary structure-activity relationship in the anti-tumor activity and QR inducing activity of tea saponins was discussed briefly.

Characterization of aromatase binding agents from the dichloromethane extract of Corydalis yanhusuo using ultrafiltration and liquid chromatography tandem mass spectrometry.

Aromatase represents an important target for the treatment of hormone-dependent breast cancer. In the present study, nine alkaloids from the dichloromethane extract of Corydalis yanhusuo were identified by liquid chromatography tandem mass spectrometry (LC-MS/MS) and tested for their aromatase binding activities using an ultrafiltration LC-MS method by investigating the differences of peak areas of compounds before and after incubations with aromatase. It was demonstrated that the quaternary protoberberine alkaloids and the tertiary protoberberine alkaloids exhibited potent aromatase binding activities. The quaternary ammonium group and the methyl group at C-13 position of tertiary protoberberine alkaloids might be necessary for the activity. The findings should provide guidance for the discovery of potential aromatase inhibitors from natural products.

Induction of quinone reductase (QR) by withanolides isolated from Physalis pubescens L. (Solanaceae).

In the present study, it was demonstrated that the dichloromethane extract of Physalis pubescens L. (DEPP) had weak potential quinone reductase (QR) inducing activity, but an UPLC-ESI-MS method with glutathione (GSH) as the substrate revealed that the DEPP had electrophiles (with an α,β-unsaturated ketone moiety). These electrophiles could induce quinone reductase (QR) activity, which might be attributed to the modification of the highly reactive cysteine residues in Keap1. Herein, four withanolides, including three new compounds physapubescin B (2), physapubescin C (3), physapubescin D (4), together with one known steroidal compound physapubescin (1) were isolated. Structures of these compounds were determined by spectroscopic analysis and that of physapubescin C (3) was confirmed by a combination of molecular modeling and quantum chemical DFT-GIAO calculations. Evaluation of the QR inducing activities of all withanolides indicated potent activities of compounds 1 and 2, which had a common α,β-unsaturated ketone moiety.

Unprecedent aminophysalin from Physalis angulata.

The 95% ethanol extract of the whole plant of Physalis angulata Linn. afforded one new skeletal physalin named aminophysalin A (1) and one new naturally occurring 5β-hydroxy-6a-chloro-5,6-dihydrophysalin B (2), together with five known physalins (3-7). Their structures were elucidated through MS, IR, NMR spectroscopy analyses and X-ray crystallography. Aminophysalin A (1) had an absolutely unusual structural feature in the chemistry of physalins with a nitrogen atom. Compounds 1-7 were evaluated for quinone reductase activities in hepa 1c1c7 cells. Physalin H (6) showed strong quinone reductase induction activity with IR (Induction ratio, QR induction activity) value of 3.74±0.02, using 4-bromoflavone as a positive control substance (2.17±0.01, 10 μg/mL), while compounds 1, 2, 3, 5 showed weak quinone reductase induction activity.

Immunoprecipitation coupled with HPLC-MS/MS to discover the aromatase ligands from Glycyrrhiza uralensis.

Affinity chromatography, applied to discover the enzyme inhibitors, needs special column with target protein and its carrier. Selection of stationary phase and mobile phase needs careful considerations due to the characteristics of proteins. In this study, a method immunoprecipitation (IP) coupled with HPLC-DAD-MS was developed to discover the aromatase ligands from Glycyrrhiza uralensis. An SB-C18 column was employed to separate target compounds without special consideration in mobile phase. Twenty-one compounds, including isolated compounds 4, 7, 8, 10, 11, 13, 15, 18-20, 23 and non-isolated compounds A-J, were found to have good affinity to aromatase by LC-MS. Seven of them (7, 15, 18, 19, 23, D, E) were detected to bind with aromatase in MCF-7 cells by IP coupled with HPLC-MS/MS. Bioassays disclosed aromatase inhibitory activities of the isolated compounds mentioned above, verifying the efficiency of IP coupled with HPLC-MS/MS as a method to screen aromatase ligands.