Optimizing the Extraction and Enrichment of Luteolin from Patrinia villosa and Its Anti-Pseudorabies Virus Activity.

Luteolin from Patrinia villosa exhibits strong antiviral activity. Here, the conditions for extracting and enriching luteolin from P. villosa were optimized. Response surface methodology was used to determine the optimal extraction parameters in terms of reflux time, solvent ratio, extraction temperature, material-to-liquid ratio, and number of extractions. Thereafter, a macroporous resin method was used to enrich luteolin from P. villosa. Finally, the following optimal extraction and enrichment conditions were established: an extraction time of 43.00 min, a methanol/hydrochloric acid solvent ratio of 13:1, an extraction temperature of 77.60 °C, a material/liquid ratio of 1:22, and a total of two extractions. NKA-9 was determined to be the most appropriate resin for enrichment. The ideal adsorption conditions were as follows: a pH of 5.0, a temperature of 25 °C, an initial luteolin concentration of 19.58 µg/mL, a sample loading volume of 2.9 BV, and a sample loading rate of 2 BV/h. The ideal desorption conditions were as follows: distilled water, 30% ethanol and 80% ethanol elution, and 5 BV at a flow rate of 2 BV/h. After optimization, the enrichment recovery rate was 80.06% and the luteolin content increased 3.8-fold. Additionally, the enriched product exhibited a significant inhibitory effect on PRV (Porcine pseudorabies virus) in vitro and in vivo, providing data for developing and applying luteolin from P. villosa.

Two Species Origins Comparison of Herba Patriniae Based on Their Ingredients Profile by UPLC-QTOF/MS/MS and Orthogonal Partial Least Squares Discriminant Analysis.

Herba Patriniae (HP) is widely used as a medicinal and edible material in China. Besides food value, HP attracts more attention due to its medicinal potential. Patrinia villosa Juss. (PV) and Patrinia scabiosaefolia Fisch. (PS) are the two species origins of HP. These two of HP show different effects on cell proliferation, migration, angiogenesis and anti-diabetic. As we have previously reported, PV and PS show significant differences on their anti-inflammatory ability in the same experimental model. Comparing the ingredient profiles of two different sources will not only facilitate the understanding of their medicinal effects, but also help the development and research of new activities. However, still now, there is no systematic and detailed study to compare the components of PV and PS. In present study, ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry was employed to achieve a high-throughput qualitative and thorough analysis of the chemical composition spectrum of HP. A total of 164 compounds were identified, among these compounds, 127 compounds were identified from PV, and 107 compounds were identified from PS. Most of the chemical components was discovered for the first time. Flavonoids, saponins, terpenoids and organic acids, as the main ingredients in PV and PS were 45.45 %vs 28.46 %, 12.61 % vs. 32.09 %, 14.33 % vs. 22.38 % and 14.58 % vs. 6.79 %, respectively. Flavonoids are the main components of PV, while PS is rich in saponins. PV and PS were classified into two groups by principal component analysis (PCA) and screened out the main molecular differences responsible by orthogonal partial least squares discriminant analysis (OPLS-DA). All the results will be a guide for the quality control, functional activity research, or better clinic use based on the ingredients profile between these two species. Besides, this first study on ingredients profile of two species origins will be beneficial for potential and best resources utilization of both PV and PS.

[Research progress on chemical constituents,pharmacological activities,and quality control of Patrinia villosa].

Patrinia villosa, regarding its functions in clearing heat and detoxification and eliminating carbuncles and pus, is widely used as a traditional medicinal herb that contains rich nutrition and substances such as various amino acids, vitamins, and soluble su-gar, and it is also an edible wild herb in Chinese folk tradition for 2 000 years. In 1973, Japanese scholars firstly separated three iridoids from Japanese P. villosa, and by 2021, chemical components such as flavonoids, iridoids, organic acids, triterpenoids, phenylpropanoids, and steroids have been found, which have multiple pharmacological effects, including antioxidant, antitumor, anti-diarrhea, antibacterial, sedative, and liver protection capabilities. Studies indicate that flavonoids, saponins, phenylpropanoids, and triterpenoids in P. villosa are vital substances for its pharmacological activities. However, the quality of this medicinal material cannot be controlled due to the unclear records in ancient books in the past dynasties and different drug use habits in different places, and thus its circulation is chaotic. At present, researchers have used flavonoids, organic acids, phenylpropanoids, triterpenoid saponins, and other compounds to conduct studies in this regard. Therefore, on the basis of the existing literature resources, we comprehensively summarize the chemical constituents, pharmacological activities, and quality control of P. villosa to further provide a reference for the safety and effectiveness of clinical drug use and lay a foundation for the follow-up experimental research.

Two New Iridoid Glucosides from the Whole Plant of Patrinia scabiosifolia Link.

As a traditional Chinese medicine, Patrinia scabiosifolia Link has been used to treat various inflammatory-related diseases, and recent studies have shown that it possesses potent anti-inflammatory activity. Therefore, phytochemical investigation on whole plants of P. scabiosifolia were carried out, which led to the isolation of two new iridoid glucosides, patriniscabiosides A (1) and B (2), together with six known compounds (3-8). The structural elucidation of all compounds was performed by HRESIMS and extensive spectroscopic analyses including IR, 1D, 2D NMR, and electronic circular dichroism (ECD). All the isolated compounds were tested for their anti-inflammatory activity using the NF-κB-Dependent Reporter Gene Expression Assay, and compound 3 displayed anti-inflammatory activity through the inhibition of the NF-κB pathway, with an inhibitory rate of 73.44% at a concentration of 10 µM.

Integrated Network Pharmacology Analysis and In Vitro Validation Revealed the Potential Active Components and Underlying Mechanistic Pathways of Herba Patriniae in Colorectal Cancer.

Herba Patriniae (HP) are medicinal plants commonly used in colorectal cancer (CRC) patients. In this study, network pharmacology was used to predict the active components and key signaling pathways of HP in CRC. Patrinia heterophylla, one type of HP, was chosen for validation of the network pharmacology analysis. The phytochemical profile of Patrinia heterophylla water extract (PHW) was determined by UHPLC-MS. MTT, RT-PCR, and Western blot assays were performed to evaluate the bioactivities of PHW in colon cancer cells. Results showed that 15 potentially active components of HP interacted with 28 putative targets of CRC in the compound-target network, of which asperglaucide had the highest degree. Furthermore, the ErbB signaling pathway was identified as the pathway mediated by HP with the most potential against CRC. Both RT-PCR and Western blot results showed that PHW significantly downregulated the mRNA and protein levels of EGFR, PI3K, and AKT in HCT116 cells. Asperglaucide, present in PHW, exhibited an anti-migratory effect in HCT116 cells, suggesting that it could be an active component of PHW in CRC treatment. In conclusion, this study has provided the first scientific evidence to support the use of PHW in CRC and paved the way for further research into the underlying mechanisms of PHW against CRC.

Patrinia herba aqueous extract on the proliferation, apoptosis, invasion and migration of hepatocellular carcinoma cells.

This study aims to study the effect and mechanism of Patrinia herba aqueous extract on proliferation, apoptosis, invasion and migration of hepatocellular carcinoma cells. Hepatocellular carcinoma cells MHCC97-H were treated with 2.5, 5, 10 mg/mL P. herba aqueous extract, cell counting kit 8 (CCK-8), flow cytometry, plate cloning experiments, and Transwell measured cell survival, apoptosis, colony formation, invasion, and migration, respectively. Real-time quantitative PCR (qPCR) and western blot were used to detect long non-coding RNA (lncRNA) HTR2A-AS1 and expression of proteins P21, Caspase-3, E-cadherin and matrix metalloproteinase-2 (MMP-2), respectively. Transfected pcDNA3.1-HTR2A-AS1 in MHCC97-H cells, or transfected si-HTR2A-AS1 and treat with 10 mg/mL P. herba aqueous extract to evaluate their roles in cell proliferation, apoptosis, invasion, and migration. Different concentrations of P. herba aqueous extract significantly reduced the survival rate, colony formation, number of migrating cells, number of invading cells, and MMP-2 protein expression of MHCC97-H cells, and obviously increased the cell apoptosis rate, the expression levels of Caspase-3, E-cadherin protein and HTR2A-AS1 (P<0.05), which were all concentration-dependent. Overexpression of HTR2A-AS1 evidently decreased the survival rate, colony formation, number of migrating cells, number of invading cells, and MMP-2 protein levels in MHCC97-H cells, while remarkably enhanced the apoptosis rate of cells, P21, Caspase-3, and E-cadherin protein levels and HTR2A-AS1 expression level (P<0.05). Inhibition of HTR2A-AS1 greatly improved the survival rate, the number of clone formation, the number of migrating cells, the number of invading cells and the expression of MMP-2 protein of MHCC97-H cells treated with P. herba aqueous extract, dramatically reducing the cell apoptosis rate, P21, Caspase-3, E-cadherin protein levels and HTR2A-AS1 expression levels (P<0.05). P. herba aqueous extract may inhibit the proliferation, invasion and migration of hepatocellular carcinoma cells by up-regulating the expression of HTR2A-AS1 in hepatocellular carcinoma cells and induce apoptosis.

Natural iridoids from Patrinia heterophylla showing anti-inflammatory activities in vitro and in vivo.

Inflammation, especially chronic inflammation, has been found to be closely related to the pathology of many diseases and the discovery of bioactive natural products to inhibit NO production is one of strategies to treat inflammation. In our continuous search for bioactive natural substances as potential anti-inflammatory agents, five new compounds (1-5) were extracted and purified from Patrinia heterophylla. The NMR and MS data analysis, along with electronic circular dichroism (ECD) calculations, led to the identification of these isolates, which were new iridoids. Using cell and zebrafish models, the in vitro and in vivo anti-inflammatory effects were conducted to evaluate the potency of anti-inflammation of these compounds. The preliminary mechanism was explored using molecular docking and Western blotting experiments.

Analysis of plasma migration components in Patrinia villosa (Thunb.) Juss. effective parts by UPLC-Q-TOF-MS.

Patrinia villosa (Thunb.) Juss. (PVJ) is described as pungent, bitter and slightly cold in Chinese medicine, and is associated with the large intestine, stomach and liver meridians. The preliminary experiments of our research team proved that PVJ total flavonoids have excellent inhibitory effects on liver cancer cells. The present experiment uses the UPLC-Q-TOF-MS technology and serum pharmacochemistry methods to analyze the chemical components in vitro and in vivo of PVJ antiliver tumors. A total of 14 chemical components were identified in the total flavonoids extract of PVJ, and it is mainly composed of flavonoids, flavonones, flavonols and phenolic acids. At the same time, seven prototypical components and seven metabolic components were detected in the drug-containing plasma. Hydrocaffeate and scutellarein are the phase I metabolites of caffeic acid and scutellarin, respectively. Sulfated apigenin, sulfated luteolin, sulfated kaempferol and methylated kaempferol are the II phase metabolites of apigenin, luteolin, kaempferol, respectively. The experiment provides a reference for the research and development of antitumor drug candidates, and provides a basis for revealing the bioactive components of PVJ and the antitumor mechanism.

Lignans, Monoterpenes and γ-Pyrone Derivatives from Patrinia scabiosifolia with Cytotoxic Activity against HCT-116 Cells.

One new dihydrobenzofuran neolignan, patrinianeolignan I, two new monoterpenes, 6,7-dehydrodissectol A and patriniaol A, and a new γ-pyrone derivative, hydroxymaltol 3-O-(6'-O-trans-caffeoyl)-ß-D-glucopyranoside, along with fifteen known lignans, eight known monoterpenes, and two known γ-pyrone derivatives, were isolated from the whole plant of Patrinia scabiosifolia. Their structures were elucidated by 1D- and 2D-NMR and HR-ESI-MS analysis. The absolute configuration of patrinianeolignan I was confirmed by circular dichroism (CD) spectrum. All compounds were evaluated in vitro for their cytotoxic activity against HCT-116 cells. The results showed that compounds patriniaol A and eudesmin exhibited moderate cytotoxicity against HCT-116 cells with IC50 values of 42.23 µM and 41.92 µM, respectively.

Anti-Melanogenic Effects of Ethanol Extracts of the Leaves and Roots of Patrinia villosa (Thunb.) Juss through Their Inhibition of CREB and Induction of ERK and Autophagy.

Patrinia villosa (Thunb.) Juss is a traditional herb commonly used in East Asia including Korea, Japan, and China. It has been administered to reduce and treat inflammation in Donguibogam, Korea. The mechanism for its anti-inflammatory effects has already been reported. In this study, we confirmed the efficacy of Patrinia villosa (Thunb.) Juss ethanol extract (Pv-EE) for inducing autophagy and investigate its anti-melanogenic properties. Melanin secretion and content were investigated using cells from the melanoma cell line B16F10. Pv-EE inhibited melanin in melanogenesis induced by α-melanocyte-stimulating hormone (α-MSH). The mechanism of inhibition of Pv-EE was confirmed by suppressing the mRNA of microphthalmia-associated transcription factor (MITF), decreasing the phosphorylation level of CREB, and increasing the phosphorylation of ERK. Finally, it was confirmed that Pv-EE induces autophagy through the autophagy markers LC3B and p62, and that the anti-melanogenic effect of Pv-EE is inhibited by the autophagy inhibitor 3-methyl adenine (3-MA). These results suggest that Pv-EE may be used as a skin protectant due to its anti-melanin properties including autophagy.

Chemical Composition of Patrinia scabiosifolia Flower Absolute and Its Migratory and Proliferative Activities in Human Keratinocytes.

Patrinia scabiosifolia (PS) has bioactivities such as antitumor and anti-inflammation effects. However, its effects on human skin physiological activities, such as skin regeneration and wound healing, remain unclear. In this study, we investigated the effects of absolute extracted from PS flower (PSF) on migration and proliferation of human dermal keratinocyte (HaCat). The yield of PSF absolute obtained by solvent extraction method was 0.105 % and its five constituents were found in GC/MS analysis. The PSF absolute induced the proliferation and migration of HaCats. The absolute increased the phosphorylation of serine/threonine-specific protein kinase (Akt) and extracellular signal-regulated kinase1/2 (Erk1/2) in HaCats. In addition, the absolute stimulated the outgrowth of collagen sprouting of HaCats. These results demonstrated, for the first time, that PSF absolute may have positive effects on skin regeneration and/or wound healing by inducing migration and proliferation of dermal keratinocytes via the Akt/Erk1/2 pathway. Therefore, PSF absolute may be a useful natural material for skin regeneration and/or wound healing.

Effects of saponins of patrinia villosa against invasion and metastasis in colorectal cancer cell through NF-κB signaling pathway and EMT.

BACKGROUND: Research has indicated that Herba Patriniae can suppress the growth of Several kinds of tumor cells in vitro and in vivo, thus displaying favorable antitumor activity. However, research regarding the effect of saponins of Patrinia villosa against CRC cell has not been reported. In the current study, We have revealed that the effects of saponins of patrinia villosa on colorectal cancer (CRC) cell invasion and epithelial-mesenchymal transition (EMT) as well as its underlying mechanism. METHODS: The CRC EMT model was induced through repeated TGF-ß1 stimulations on human CRC cell line SW480. Effects of saponins of patrinia villosa at various concentrations on CRC SW480 cell and EMT model cell proliferation were detected using MTT method, so as to select the optimal action concentration. Meanwhile, effects on SW480 cell and EMT model cell invasion were determined through Scratch assay and Transwell assay. Moreover, changes in expression of EMT-related proteins E-cadherin, N-cadherin and NF-ΚBp65 in each group were detected through Western blotting. RESULTS: Saponins of patrinia villosa at various concentrations could markedly inhibit the proliferation rate of CRC cell in an obvious concentration-dependent manner. Meanwhile, saponins of patrinia villosa at various concentrations could also remarkably suppress migration of cell developing EMT. In addition, the protein expression of E-cadherin and N-cadherin was down-regulated with the increase in saponins of patrinia villosa concentration, while that of NF-KBp65 was notably down-regulated. CONCLUSION: Saponins of patrinia villosa can act against tumor invasion and metastasis through inhibiting EMT in human CRC cell line, which may be achieved through down-regulating the NF-κB signaling pathway.

Iridoids from the Roots of Patrinia scabra and Their Inhibitory Potential on LPS-Induced Nitric Oxide Production.

An activity-guided fractionation procedure of the 70% aqueous EtOH extract from the roots of Patrinia scabra led to the isolation and characterization of five new iridoids, patriscabrins A-E (1-5), along with 13 known compounds. The structures of 1-5 were determined by interpretation of spectroscopic data, particularly by 1D and 2D NMR, ECD, and VCD studies. Thereafter, isolates were evaluated for their inhibitory effects on lipopolysaccharide-induced nitric oxide production in RAW 264.7 cells. Of these, the new iridoids 2 and 5 and the known lignan patrineolignan B (6) exhibited IC50 values of 14.7 to 17.8 µM.

[Evolution and transition of source of Patrinia Herba and Patrinia Radix].

Both Patrinia Herba and Patrinia Radix are traditional Chinese herbal medicines. The herbal source and medicinal part of them are confusing in the herbal medicine market of China. To explore the evolution and transition of the herbal source and medicinal part of Patrinia Herba and Patrinia Radix, this paper systematically summarizes the record of the herbal source and medicinal part of them in ancient classics of herbal medicine in China. According to the findings, before Ming Dynasty, Patrinia Herba originated from the radix of the plants with yellow flowers of Patrinia. In Ming and Qing Dynasty, Patrinia Herba originates from the whole plant (including the radix)of the plant with white flowers of Patrinia. In Ming Dynasty, Patrinia Radix, stemming from the radix of the plants with yellow flowers of Patrinia, started to be used as a traditional Chinese herbal medicine, which had the same herbal source with that of Patrinia Herba before Ming Dynasty. Therefore, Patrinia Herba and Patrinia Radix can be seen as the same traditional Chinese herbal medicine, and the genuine of Patrinia Herba should be the radix and the whole herba of P. scabiosaefolia and P. heterophylla.

A new sesquiterpene lactone glycoside and a new quinic acid methyl ester from Patrinia villosa.

A new sesquiterpene lactone glycoside (1) and a new quinic acid methyl ester (2) were isolated from Patrinia villosa, together with another two known compounds chlorogenic acid n-butyl ester (3), 3, 4-di-O-caffeoylquinic acid methyl ester (4). Their structures were established using 1D/2D-NMR spectroscopy, mass spectrometry, and comparing with spectroscopic data reported in the literature.

New flavonoid from Patrinia villosa.

CONTEXT: Patrinia villosa (Thunb.) Juss (Valerianaceae) is an important ancient herbal medicine widely used for inflammation, wound healing, and abdominal pain. But little is known of the phytochemical constituents of this herbal plant. OBJECTIVE: The objective of this study is to isolate and identify the bioactive components from P. villosa. MATERIALS AND METHODS: A 70% EtOH extract of P. villosa was subjected to normal-phase silica, ODS silica gel column chromatography, and semi-preparative HPLC chromatography after partitioned successively with light petroleum, dichloromethane and n-BuOH. Chemical structures of the compounds were elucidated by spectroscopic methods including UV, 1D-NMR, 2D-NMR, HR-ESI-MS, and CD spectra. The cytotoxic activity of the new component was determined with the SMMC-7721 cell line using the MTT method after incubation for 48 h. RESULTS: A new flavonoid named patriniaflavanone A (1) along with four known compounds was isolated from P. villosa. The four known compounds were identified as luteolin 7-O-glucuronide-6″-methyl ester (2), p-hydroxyphenylacetic acid methyl ester (3), trans-caffeic acid (4), and trans-caffeic acid methylate (5) by comparison of their spectral data with the reported data. The IC50 value of patriniaflavanone A (1) on SMMC-7721 was 61.27 µM. DISCUSSION AND CONCLUSION: This is the first report on the isolation and identification of patriniaflavanone A (1), and compounds 2-5 were isolated for the first time from the title plant. Patriniaflavanone A (1) exhibited moderate cytotoxic activity.

A systematic approach for authentication of medicinal Patrinia species using an integration of morphological, chemical and molecular methods.

Four common Patrinia species, including P. heterophylla, P. monandra, P. scabiosifolia and P. villosa, have been documented as herbal medicines with various clinical applications, such as anti-cancer, anti-diarrhea and sedative. However, the authentication of medicinal Patrinia species poses a problem, particularly with the processed herbal materials. This study aimed to systematically authenticate the four medicinal Patrinia species in the market using morphological and chemical characterization, as well as DNA markers. We found the species identity authenticated by traditional morphologies were in good agreement with both chemical and molecular results. The four species showed species-specific patterns in chromatographic profiles with distinct chemical markers. We also revealed the power of complete chloroplast genomes in species authentication. The sequences of targeted loci, namely atpB, petA, rpl2-rpl23 and psaI-ycf4, contained informative nucleotides for the species differentiation. Our results also facilitate authentication of medicinal Patrinia species using new DNA barcoding markers. To the best of our knowledge, this is the first report on the application of morphology, chemical fingerprinting, complete chloroplast genomes and species-specific Insertion-Deletions (InDels) in differentiating Patrinia species. This study reported on the power of a systematic, multidisciplinary approach in authenticating medicinal Patrinia species.

Hepatoprotective activity of Patrinia scabiosaefolia Fisch and quality evaluation based on UPLC fingerprint and multi-component analysis.

To establish a quality evaluation method for Patrinia scabiosaefolia Fisch (PS), as well as to study the anti-inflammatory and hepatoprotective effects of the aqueous extract of Patrinia scabiosaefolia Fisch (APS). We used ultra performance liquid chromatography (UPLC) to establish fingerprint and content determination method for PS. The alcoholic liver injury model was prepared by feeding Lieber-DeCarli alcohol liquid feed to mice. We determined the levels of ALT, AST, TC, TG in serum, as well as GSH, MDA in the liver. The mRNA relative expression levels of TNF-α, IL-6, IL-1ß, INOS and COX-2 were detected by qRT-PCR, and liver tissues were taken for pathological examination. The fingerprints of 16 batches of PS were established, and 3 component peaks were identified, which were chlorogenic acid (CA), isochlorogenic acid A (ICAA) and isochlorogenic acid C (ICAC). The similarity of the 6 common peaks was between 0.924 and 1.000. A mice model of alcoholic liver injury was successfully made by mixing alcohol liquid feed. The levels of ALT, AST, TC and TG in serum and MDA, TNF-α, IL-1ß, LL-6, COX-2 and INOS mRNA in liver were effectively reduced in the drug administration group. The levels of GSH in mouse liver tissue were increased in the drug administration group. The method has good repeatability, stability and feasibility, and it meets the requirements for Quality evaluation. APS exhibits a protective effect against alcoholic liver injury (ALI) in mice.

Glycosides from the aerial parts of Patrinia villosa.

An investigation of the Korean medicinal plant Patrinia villosa (THUNB.) JUSS. (Valerianaceae) led to the isolation of two new flavonoid glycosides, patrivilosides 1 (1) and 2 (2), a new iridoid glycoside, patrinovalerosidate (3), and two new saponins, patrinovilosides A (4) and B (5), along with six known compounds including three flavonoid glycosides and three iridoid glycosides. The structures of the new compounds were elucidated based on analysis of their one dimensional (1D)- and 2D-NMR spectra along with their mass spectrometric data and the results of acid hydrolysis.

Facile synthesis of the naturally cytotoxic triterpenoid saponin Patrinia-glycoside B-II and its conformer.

The first chemical synthesis of the natural triterpenoid saponin Patrinia-glycoside B-II, namely oleanolic acid 3-O-α-L-rhamnopyranosyl-(1→2)-[ß-D-gluco-pyranosyl-(1→3)]-α-L-arabinopyranoside, has been accomplished in a linear 11-step sequence 11 with 9.4% overall yield. The abnormal 1C4 conformation of the arabinose residue was found to occur via conformational fluctuation during preparation of the intermediates. Molecular mechanism and quantum chemistry calculations showed that Patrinia-glycoside B-II and its conformer 1 cannot interconvert under normal conditions. Preliminary structure-activity relationships studies indicated that the 4C1 chair conformation of the arabinose residue in the unique α-L-rhamnopyranosyl-(1→2)-α-L-arabinopyranosyl disaccharide moiety is one of the chief positive factors responsible for its cytotoxic activity against tumors.