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.

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.

Mechanistic insights into the anti-tumor and anti-metastatic effects of Patrinia villosa aqueous extract in colon cancer via modulation of TGF-ß R1-smad2/3-E-cadherin and FAK-RhoA-cofilin pathways.

BACKGROUND: Patrinia villosa, a traditional medicinal herb commonly used for treating intestinal-related diseases, has been commonly prescribed by Chinese medicine practitioners as a key component herb to treat colon cancer, although its anti-tumor effect and mechanisms of action have not been fully elucidated. HYPOTHESIS/PURPOSE: This study aimed to investigate the anti-tumor and anti-metastatic effects of Patrinia villosa aqueous extract (PVW), and its underlying mechanisms. METHOD: The chemical profile of PVW was analysed by high-performance liquid chromatography with photodiode-array detection (HPLC-DAD) method. Cell-based functional assays MTT, BrdU, scratch, and transwell were conducted to evaluate the effects of PVW on human colon cancer HCT116 and murine colon26-luc cells, assessing cytotoxicity, cell proliferation, motility, and migration, respectively. Western blotting was performed to assess the effect of PVW on the expression of key intracellular signaling proteins. In vivo studies were conducted using zebrafish embryos and tumor-bearing mice to evaluate the anti-tumor, anti-angiogenesis, and anti-metastatic effects of PVW in colon cancer. RESULTS: Five chemical markers were identified and quantified in PVW. PVW exhibited significant cytotoxicity and anti-proliferative activity, as well as inhibitory effects on cell motility and migration in both HCT116 and colon 26-luc cancer cells via modulating protein expressions of TGF-ß R1, smad2/3, snail, E-cadherin, FAK, RhoA, and cofilin. PVW (0.01-0.1 mg/ml) could significantly decrease the length of subintestinal vessels of zebrafish embryos through decreasing mRNA expressions of FLT1, FLT4, KDRL, VEGFaa, VEGFc, and Tie1. PVW (> 0.05 mg/ml) also significantly suppressed colon cancer cells migration in the zebrafish embryos. Furthermore, oral administration of PVW (1.6 g/kg) significantly inhibited tumor growth by decreasing the expressions of tumor activation marker Ki-67 and CD 31 in tumor tissues of HCT116 tumor-bearing mice. PVW could also significantly inhibit lung metastasis in colon 26-luc tumor-bearing mice by modulating their tumor microenvironment, including immune cells populations (T cells and MDSCs), levels of cytokines (IL-2, IL-12, and IFN-γ), as well as increasing the relative abundance of gut microbiota. CONCLUSION: This study revealed for the first time the anti-tumor and anti-metastatic effects of PVW through regulation of TGF-ß-smad2/3-E-cadherin, and FAK-cofilin pathways in colon cancer. These findings provide scientific evidence to support the clinical use of P. villosa in patients with colon cancer.

New iridoids from Patrinia scabiosaefolia and their hypoglycemic effects by activating PI3K/Akt signaling pathway.

Growing in regions of Asia and North America, Patrinia scabiosaefolia is a wild vegetable and herb that has demonstrated health-promoting properties. Iridoids are one of the most bioactive phytochemicals in P. scabiosaefolia but the in-depth study is scarce. Herein we reported the separation and characterization of nine iridoids (compounds 1-9) from P. scabiosaefolia, and two compounds (2 and 6) were new. All the structures of the nine iridoids were characterized and confirmed with NMR (1D & 2D), HRMS, IR and UV. Compound 2 is a five-member ring iridoid, reminiscent of a broken C-1 and C-2 bond. Compound 6 has a typical monoene valerian iridoid, but the 5-deoxyglucose moiety at C-11 position is uncommon in this genus. The anti-diabetic evaluation of the isolated compounds revealed that compounds 1, 2, and 9 significantly increased the glucose absorption in 3 T3-L1 cells (P < 0.01). Further mechanism investigations have demonstrated that compound 1 promoted glucose uptake in dexamethasone-treated 3 T3-L1 adipocytes by activating PI3K/Akt signaling pathway. The expression of GLUT4 mRNA and protein was also upregulated. These results provide scientific references for the potential use of P. scabiosaefolia as a functional food to manage hyperglycemia.

[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.

Therapeutic effect of Patrinia villosa on TNBS-induced ulcerative colitis via metabolism, vitamin D receptor and NF-κB signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Ulcerative colitis (UC) is a chronic inflammatory disease affecting the colon. Patrinia villosa Juss. (P.V) is an important traditional Chinese medicine widely used for more than 2000 years from ShenNongBenCaoJing, a famous ancient Chinese medicinal literary. P.V is often used in the treatment of UC, but the pathogenesis is not clear. AIM OF THE STUDY: This study was designed to analysis the metabolic pathways and relevant mechanisms of P.V on UC rats induced by TNBS. MATERIALS AND METHODS: The rat model of UC was established by 2,4,6-trinitrobenzene sulfonic acid (TNBS)/ethanol method. Three doses of P.V (21 g/kg, 43 g/kg, 64 g/kg) were administrated for 14 days. Disease activity index (DAI) scoring system and H&E staining were used to evaluate the efficacy. A method for simultaneous detection of 96 endogenous metabolic components was established by UPLC-MS. The method was used to detect the metabolites in serum and liver of rats with UC induced by TNBS. PLS-DA and Metaboanalyst were used to analyze the main metabolic pathways involved in the treatment of UC. The contents of IL-1ß, TNF-α and IL-6 in the colonic homogenate of rats were detected by enzyme-linked immunosorbent assay (ELISA). The expression levels of VDR, NF-κB, p-NF-κB, NLRP3 and caspase-1 in colon tissues of rats were detected by the method of Western blot. RESULTS: DAI scoring system and H&E staining indicated that P.V have the obvious therapeutic effect on UC induced by TNBS as a dosage-dependent manner. 36 potential biomarkers in serum and 26 potential biomarkers in liver were found in positive and negative ion mode of UPLC-MS, which significantly affected Glycine, serine and threonine metabolism, Glycerophospholipid metabolism, Purine metabolism, Histidine metabolism, Alanine, aspartate, and glutamate metabolism, Arginine and proline metabolism in serum, and significantly affected Purine metabolism, Lycine, serine and threonine metabolism, Glutathione metabolism, Glyoxylate and dicarboxylate metabolism in the liver. The contents of pro-inflammatory cytokines and receptors related to NF-κB signaling axis of model group were significantly higher than those of the control group, compared with the model group, their contents of the P.V group were significantly decreased (p < 0.01). Compared with the model group, the expression of NF-κB, p-NF-κB, NLRP3 and caspase-1 in colon tissues of the rats in P.V group were significantly decreased (p < 0.01). The expression of VDR in model group were significantly reduced compared to that in the control group, compared with the model group, the expression of VDR in P.V group were significantly increased (p < 0.01). CONCLUSION: P.V has an obvious therapeutic effect on UC induced by TNBS by regulating the energy metabolism, amino acid metabolism, purine metabolism, bile acid metabolism and lipid metabolism. P.V exerts anti-inflammatory effect by impacting bile acid levels, activating VDR, and inhibiting the overactivation of NF-κB signaling pathways.

A novel flavonoid with antioxidant activity from Patrinia villosa (Thunb.) Juss.

Patrinia villosa (Thunb.) Juss (P. villosa), a perennial herb, is widely used as a medicinal plant in Chinese folk. This study aims to isolate and identify the chemical constituents from P. villosa and evaluate their antioxidant activity. Normal silica column chromatography, ODS silica column chromatography, Sephadex LH-20 column chromatography and semi-preparative HPLC methods were used to obtain a new compound named 3-n-pentadecyl-4'-methoxyluteolin (1) and two known compounds including luteolin-7-O-ß-D-glucuronide methyl ester (2) and apigenin-7-O-ß-D-glucuronide methyl ester (3). The antioxidant activity of these compounds was determined by DPPH and ABTS methods and the IC50 values were calculated. The IC50 values of ABTS scavenging activity of 1, 2 and 3 were 12.99 ± 0.09 µM, 7.13 ± 0.07 µM and 5.15 ± 0.08 µM, respectively, and the IC50 values of DPPH scavenging activity of 1, 2 and 3 were 51.86 ± 0.41 µM, 23.95 ± 0.71 µM and 25.06 ± 0.65 µM, respectively. All the compounds exhibited good antioxidant activities in vitro.

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.

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.

The Herba Patriniae (Caprifoliaceae): A review on traditional uses, phytochemistry, pharmacology and quality control.

ETHNOPHARMACOLOGICAL RELEVANCE: Herba Patriniae has been used for thousands of years in China as a traditional Chinese medicine with heat-clearing and detoxicating effects. It is applied widly for the treatment of rheumatoid arthritis, diarrhea, acute hepatitis, pelvic inflammatory disease and ulcerative colitis in clinic. Two species, namely Patrinia scabiosaefolia Fisch. (PS) and Patrinia villosa Juss. (PV) from the Caprifoliaceae family, are considered as Herba Patriniae in the pharmaceutical industry. AIM OF THE REVIEW: This paper aims to comprehensively outline the traditional uses, botanical description, phytochemistry, pharmacology, toxicology, quality control, pharmacokinetics and patents of Herba Patriniae, and elaborate the same/different characteristics between PS and PV. MATERIALS AND METHODS: Detailed information of Herba Patriniae was collected from various online databases (Pubmed, Web of Science, Google Schola, China National Knowledge Infrastructure Database, National Intellectual Property Administration, PRC National Medical Products Administration), and those published resources (M.Sc. Thesis and books). RESULTS: A total of 233 compounds have been identified in Herba Patriniae, including triterpenoid saponins, flavonoids, organic acids, iridoids, and volatiles. A very distinct difference was observed, that PS is rich in triterpenoid saponins and volatiles, while PV contains more flavonoids. Two source species of Herba Patriniae gave similar pharmacological effects on anti-cancer, anti-inflammatory, antioxidant, antimicrobial, sedative and hypnotic effects. But there were no reports were on antipruritic, proangiogenic and anti-diarrheal effects for PS, and no studies on anti-diabetic effects for PV. Generally, Herba Patriniae showed non-toxic in the clinical dose, but mild side effects, such as temporary leukopenia, dizziness and nausea, could be found when large and excessive dosage is used. A variety of compounds have been quantified for the quality control of PS and PV. The variety, growth environment, growth time, and harvest time not only affected the contents but also the pharmacological activities of the bioactive compounds. In the past year, patents for compositions containing PV and PS have been filed, mainly involving human health, hygiene, agriculture, and animal husbandry. Unfortunately, the research on pharmacokinetics is insufficient. Only the prototype components and metabolites were repored after intragastric administration of total flavonoids extract from PV in rats. CONCLUSION: Herba Patriniae has displayed a significant medicinal value in clinic, but the differences in phytochemistry, pharmacological effects and the content of compounds have been found between two official recorded species. About side effects and pharmacokinetic characteristics, the differences between two species have not been well studied. For a better clinical use of Herba Patriniae, it is urgent to establish systematic pharmacology, quality control, pharmacokinetics, and clinical researches on the same/different characteristics between PS and PV.

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.

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.

Patriscabrin F from the roots of Patrinia scabra attenuates LPS-induced inflammation by downregulating NF-κB, AP-1, IRF3, and STAT1/3 activation in RAW 264.7 macrophages.

BACKGROUND: The roots of Partrinia scabra have been used as a medicinal herb in Asia. We previously reported that the inhibitory effect of patriscabrin F on lipopolysaccharide (LPS)-induced nitric oxide (NO) production was the most potent than that of other isolated iridoids from the roots of P. scabra. PURPOSE: We investigated the anti-inflammatory activity of patriscabrin F as an active compound of P. scabra and related signaling cascade in LPS-activated macrophages. METHOD: The anti-inflammatory activities of patriscabrin F were determined according to its inhibitory effects on NO, prostaglandin E2 (PGE2), and pro-inflammatory cytokines. The molecular mechanisms were revealed by analyzing nuclear factor-κB (NF-κB), activator protein-1 (AP-1), interferon regulatory factor 3 (IRF3), and Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway. RESULTS: Patriscabrin F inhibited the LPS-induced production of NO, PGE2, tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß, and IL-6 in both bone-marrow derived macrophages (BMDMs) and RAW 264.7 macrophages. Patriscabrin F downregulated LPS-induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), TNF-α, IL-1ß, and IL-6 at the transcriptional level. Patriscabrin F suppressed LPS-induced NF-κB activation by decreasing p65 nuclear translocation, inhibitory κBα (IκBα) phosphorylation, and IκB kinase (IKK)α/ß phosphorylation. Patriscabrin F attenuated LPS-induced AP-1 activity by inhibiting c-Fos phosphorylation. Patriscabrin F suppressed the LPS-induced phosphorylation of IRF3, JAK1/JAK2, and STAT1/STAT3. CONCLUSION: Taken together, our findings suggest patriscabrin F may exhibit anti-inflammatory properties via the inhibition of NF-κB, AP-1, IRF3, and JAK-STAT activation in LPS-induced macrophages.

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.

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.

A Cytotoxic Natural Product from Patrinia villosa Juss.

BACKGROUND AND PURPOSE: Patrinia villosa Juss is an important Chinese herbal medicine widely used for thousands of years, but few reports on the ingredients of the herb have been presented. In this study, we aim to isolate the bioactive compound from the plant. MATERIAL AND METHODS: The air-dried leaves of P. villosa (15kg) were extracted three times with 70% EtOH under reflux. The condensed extract was suspended in H2O and partitioned with light petroleum, dichloromethane and n-BuOH. The dichloromethane portion was then subjected to normal-phase silica gel column chromatography, ODS silica gel column chromatography and semi-preparative HPLC to yield compound 1. Cytotoxicities of 1 were assayed on HepG2, A549 and A2780 cell lines. The mechanism of apoptosis and cell cycle on A549 was confirmed subsequently. RESULTS: A new impecylone (Impecylone A) was isolated from the leaves of Patrinia villosa Juss, and its structures were established using 1D, 2D-NMR spectra and HR-ESI-MS. Impecylone A could selectivity inhibit HepG2 and A549 cell lines. The compound could induce apoptosis of A549 and arrest the cell cycle at G2/M phase in a dose-dependent manner. CONCLUSION: Impecylone A is a novel compound from Patrinia villosa Juss and could be a potential antitumor agent especially in the cell lines of A549.

Chemical constituents of Patrinia heterophylla Bunge and selective cytotoxicity against six human tumor cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Patrinia heterophylla Bunge, known as "Mu-Tou-Hui" in China, is distributed in most provinces and regions of China. As a traditional medicinal plant, which was first found in . In many traditional herbal books, there are records of "Mu-Tou-Hui" of treatment for uterine bleeding, cancer, swelling pain, leukemia, etc. However, there are few studies on the chemical constituents of Patrinia heterophylla Bunge. AIM OF THE STUDY: To investigate the chemical constituents of P. heterophylla and the basis of their antitumor activity. MATERIALS AND METHODS: 15 compounds were isolated from roots and rhizomes of P. heterophylla by repeating various column chromatography techniques, whose structures were determined by organic spectrum analysis methods and compared with published data. The cytotoxicities were evaluated by MTT assay on six cancer cell lines: human melanoma cell (A375), human hepatocellular carcinoma cell (SMMC-7721), human gastric cancer cell (SGC-7901), human cervical cancer cell (HeLa), human colon cancer cell (HCT-116), and human breast cancer cell (MDA-MB-231). The apoptosis-inducing activities of compounds 1, 5, 12 and 15 in A375 tumor cell determined by flow cytometry. RESULT: Five phenylpropanoids, ethyl caffeate (1), coniferaldehyde (5), trans-p-coumaryl aldehyde (6), caffeic acid methyl ester (12), and 3,4-dihydroxycinnamic acid (15), four acetophenones, 1-(2,4-dihydroxyphenyl) ethanone (2), 2',5'-Dihydroxyacetophenone (3), cynanchone A (8), and cynandione A (10), two phenols, vanillin (4) and catechol (9), two iridoids, sarracenin (7) and patriscabrol (11), one alkane, tetracosane (14), and one coumarin, scopoletin (13), were isolated from the EtOH extracts. Of them, compounds 1-10, 12 and 14-15 were isolated for the first time from the roots of P. heterophylla. Compounds 1 and 15 were reported for the first time with in vitro inhibitory activity against tumor cells. MTT assay showed that compounds 1, 5-9, 12-13 and 15 had selective cytotoxic activities (IC50 27.20-163.03 µM) against tumor cells. Apoptosis detected by flow cytometry revealed that compounds 1, 5, 12 and 15 can induce apoptosis for A375 at low concentrations when the concentrations of compounds 1, 5, 12 and 15 are the value of 14, 40, 34, 108 µM, the percentages of apoptotic cells were about 50%. CONCLUSIONS: Compounds 1-10, 12 and 14-15 were isolated for the first time from the P. heterophylla. This result enriches the previous studies on the chemical constituents of P. heterophylla. Compounds 1 and 15 were reported for the first time to have cytotoxic activities against tumor cells. Compounds 1, 5, 6, 7, 12, 15 showed cytotoxic activities against tumor cells. This result reveals that the active ingredient of P. heterophylla are composed of phenylpropanoids, iridoids and coumarins. This study provides some theoretical basis for the anti-tumor research of P. heterophylla.

Patrinia scabiosaefolia Inhibits Growth of 5-FU-Resistant Colorectal Carcinoma Cells via Induction of Apoptosis and Suppression of AKT Pathway.

OBJECTIVE: To investigate the effects of ethanol extract of Patrinia scabiosaefolia (EEPS) on chemo-resistance of colorectal cancer cells (CRC) and explore the possible molecular mechanisms. METHODS: 5-fluorouracil (5-FU)-resistant human colorectal carcinoma cell line (HCT-8/5-FU) and its parental cells HCT-8 were treated with EEPS (0, 0.25, 0.50, 1 or 2 mg/mL), or 5-FU (0, 100, 200, 400, 800 or 1600 µmol/L). The 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was performed to evaluate the cell viability. Cell density was observed by phase-contrast microscope, cell counting and colony formation assay were used to determine the cell proliferation of HCT-8/5-FU cells treated with 0, 0.5, 1 or 2 mg/mL EEPS. Cell apoptosis was determined by Hoechst staining. Western-blot was performed to detect the phosphorylation of AKT as well as the protein expression level of B-cell CLL/lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax). RESULTS: Compared with HCT-8 cells, MTT assay results indicated that HCT-8/5-FU cells were resistant to 5-FU treatment (P<0.05), and sensitive to EEPS treatment (P>0.05). Moreover, compared with untreated HCT-8/5-FU cells, 1 and 2 mg/mL of EEPS treatment significantly reduced cell density, cell number, inhibited cell survival (P<0.05), and induced apoptosis in HCT-8/5-FU cells. Furthermore, 1 and 2 mg/mL of EEPS significantly decreased the phosphorylation level of p-AKT and Bcl-2 protein expression, and increased the expression of Bax protein (P<0.05). CONCLUSION: EEPS is a promising therapeutic agent that may overcome chemo-resistance in cancer cells, likely through suppression of the AKT pathway and promotion of cancer cell apoptosis.

Inhibitory effect of Patrinia on BRL-3A cell apoptosis through the TLR4/PI3K/AKT/GSK3ß and TLR4/P38/JNK signaling pathways.

The present study investigated the inhibitory effect of Patrinia on lipopolysaccharide (LPS)-induced apoptosis of rat liver BRL­3A cells. A Cell Counting Kit­8 assay was performed to measure the effect of Patrinia on BLR­3A cell activities. A biochemical assay was employed to detect the release of lactate dehydrogenase (LDH) in BRL­3A cells induced by different doses of LPS. Based on the release rate of LDH, drug concentrations were set at 0.5, 1 and 2 g/l. Apoptotic morphology of cells was observed via Hoechst 33342 staining and flow cytometry was performed to detect apoptosis rates. Western blotting was performed to detect the expression of toll­like receptor 4 (TLR4), protein kinase B (AKT), phosphorylated (P)­AKTSer473, glycogen synthase kinase 3ß (GSK3ß), P­GSK3ßSer9, P38, P­P38, c­Jun N­terminal kinase (JNK), P­JNK, B­cell lymphoma­2 (Bcl­2), Bcl­2 associated X protein (Bax) and active­caspase­3 proteins. The translocation of GSK3ß was observed by immunofluorescence staining. Results revealed that Patrinia increases cell activities and inhibits apoptosis. The expression levels of TLR4, P­P38 and P­JNK were reduced, whereas the expression of P­AKTSer473 and P­GSK3ßSer9 were increased. Patrinia significantly reduced GSK3ß nuclear translocation induced by LPS, and significantly decreased the mRNA expression levels of Bax/Bcl­2 and caspase­3 in BRL­3A cells induced by LPS. In conclusion, Patrinia may significantly reduce apoptosis of BRL­3A induced by LPS via the TLR4/PI3K/AKT/GSK3ß and TLR4/P38/JNK signaling pathways, providing evidence for its potential use in liver disease therapy.