Seven new secondary metabolites isolated from roots of Lespedeza bicolor.

Chemical investigation of a methanol extract obtained from the roots of Lespedeza bicolor identified one new pterocarpene (1), three new pterocarpans (2-4), and three new arylbenzofurans (5-7), and two known compounds (8 and 9). Their structures were determined by interpretations obtained from combined UV, NMR, and HRTOFMS spectroscopic data. Furthermore, the absolute configurations of compounds 2 and 3 were established by the combination of electronic circular dichroism (ECD) calculations and NMR calculations with DP4+ probability analysis. All isolated compounds (1-9) were evaluated for cytotoxicity against the human lung carcinoma cell line A549 and the human hepatoma cell line Huh-7. Compound 4 showed antiproliferative activity against A549 cell line with IC50 value of 24.9 µM. Furthermore, compound 9 exhibited cytotoxicity against Huh-7 cell line with IC50 value of 68.7 µM.

Lespedeza bicolor extract supplementation reduced hyperglycemia-induced skeletal muscle damage by regulation of AMPK/SIRT/PGC1α-related energy metabolism in type 2 diabetic mice.

Lespedeza bicolor (LB) is known to have antidiabetic activities; however, the underlying molecular mechanisms of LB in hyperglycemia-induced skeletal muscle damage is unclear. Inflammation and oxidative stress caused by type 2 diabetes mellitus (T2DM) not only contributes to insulin resistance, but also promotes muscle atrophy via decreased muscle protein synthesis and increased protein degradation, leading to frailty and sarcopenia. In this study, we hypothesized that LB extract (LBE) supplementatin has an ameliorative effect on hyperglycemia-induced skeletal muscle damage by activation of 5' adenosine monophosphate-activated protein kinase (AMPK)/sirtuin (SIRT)/proliferator-activated receptor γ coactivator 1α (PGC1α)-associated energy metabolism in mice with T2DM. Diabetes was induced by a high-fat diet with a 2-time streptozotoxin injection (30 mg/kg body weight) in male C57BL/6J mice. After diabetes was induced (fasting blood glucose level ≥140 mg/dL), the mice were administered with LBE at a low dose (100 mg/kg/d) or high dose (250 mg/kg/d) by gavage for 12 weeks. LBE supplementation ameliorated glucose tolerance and hemoglobin A1c (%) in mice with T2DM. Moreover, LBE supplementation upregulated protein levels of insulin receptor subunit-1 and Akt accompanied by increased translocation of glucose transporter 4 in mice with T2DM. Furthermore, LBE increased mitochondrial biogenesis by activating SIRT1, SIRT3, SIRT4, and peroxisome PGC1α in diabetic skeletal muscle. Meanwhile, LBE supplementation reduced oxidative stress and inflammation in mice with T2DM. Taken together, the current study suggested that LBE could be a potential therapeutic to prevent skeletal muscle damage by regulation AMPK/SIRT/PGC1α-related energy metabolism in T2DM.

Biologically active polyphenolic compounds from Lespedeza bicolor.

We investigated the ability of six prenylated prerocarpans, stilbenoid, and a new dimeric flavonoid, lespebicolin B, from stem bark as well as two 3-O-rutinosides and a mixture of 3-O-ß-D-glucosides of quercetin and kaempferol from flowers of Lespedeza bicolor to inhibit HSV-1 replication in Vero cells. Pretreatment of HSV-1 with polyphenolic compounds (direct virucidal effect) showed that pterocarpans lespedezol A2 (1), (6aR,11aR)-6a,11a-dihydrolespedezol A2 (2), (6aR,11aR)-2-isoprenyldihydrolespedezol A2 (4), and (6aR,11aR,3'R)-dihydrolespedezol A3 (5) significantly inhibited viral replication, with a selective index (SI) ≥10. Compound 4 possessed the lowest 50% - inhibiting concentration (IC50) and the highest SI values (2.6 µM and 27.9, respectively) in this test. (6aR,11aR)-2-Isoprenyldihydrolespedezol A2 (4) also had a moderate effect under simultaneous treatment of Vero cells with the tested compound and virus (IC50 and SI values were 5.86 µM and 12.4, respectively). 3-O-rutinosides of quercetin and kaempferol and a mixture of 3-O-ß-D-glucosides of quercetin and kaempferol (10 and 12) also showed significant virucidal activity, with SI values of 12.5, 14.6, and 98.2, respectively, and IC50 values of 8.6, 12.2, and 3.6, respectively. We also performed a quantitative structure-activity relationship (QSAR) analysis of data on the virucidal activity of polyphenolics with 4 < pIC50 < 6. It was found that the virucidal activity of these compounds depended on both the structure of the aromatic part and the conformation of geranyl and isoprenyl side chains of their molecules. These findings are correlated with the largest value of the principal moment of inertia (pmi) descriptor describing the geometry of molecules.

Preparation, characterization and anti-cancer activity of graphene oxide-­silver nanocomposite.

This work reported the preparation, characterization, cytotoxicity of green synthesized Lespedeza cuneate mediated silver nanoparticles (Lc-AgNPs) and graphene oxide­silver nanocomposite (GO-AgNComp) using Lc-AgNPs. The UV absorption spectrum at 419 nm indicated the successful formation of GO-AgNComp. The TEM analysis displayed the thin sheet of graphene decorated Lc-AgNPs in GO-AgNComp. Zeta potential was -13.2 mV for Lc-AgNPs and -30.5 mV for GO-AgNComp. The photothermal conversion efficiency was calculated as 31.09% for GO-AgNComp. The negatively charged zeta potential of GO-AgNComp enhanced its cellular penetration through enhanced permeability and retention (EPR) effect. The near-infrared laser (NIR) induced the anticancer activity of Lc-AgNPs and GO-AgNComp in human lung cancer cells (A549) and brain tumour (LN229). The results indicated that about 50% of A549 cells and LN229 cells were ablated by treatment of 24.73 ± 2.98 µg/mL and 27.34 ± 1.62 µg/mL of Lc-AgNPs, as well by 15.46 ± 2.31 µg/mL and 20.95 ± 1.35 µg/mL of GO-AgNComp respectively. Moreover, GO-AgNComp was not cytotoxic to normal mouse fibroblast cells (NIH3T3), but it caused the cancer cell death in A549 and LN229 through ROS generation, nuclear damage, and mitochondrial membrane potential (∆ψm) loss. This work reported the anticancer potential of GO-AgNComp, which deserves further study on the molecular elucidation of GO-AgNComp mediated human lung and tumour therapy.

Aviculin Isolated from Lespedeza cuneata Induce Apoptosis in Breast Cancer Cells through Mitochondria-Mediated Caspase Activation Pathway.

The global incidence of breast cancer has increased. However, there are many impediments to the development of safe and effective anticancer drugs. The aim of the present study was to evaluate the effect of aviculin isolated from Lespedeza cuneata (Dum. Cours.) G. Don. (Fabaceae) on MCF-7 human breast cancer cells and determine the underlying mechanism. Using the bioassay-guided isolation by water soluble tetrazolium salt (WST-1)-based Ez-Cytox assay, nine compounds (four lignan glycosides (1-4), three flavonoid glycosides (5-7), and two phenolic compounds (8 and 9)) were isolated from the ethyl acetate (EA) fraction of the L. cuneata methanolic extract. Of these, aviculin (2), a lignan glycoside, was the only compound that reduced metabolic activity on MCF-7 cells below 50% (IC50: 75.47 ± 2.23 µM). The underlying mechanism was analyzed using the annexin V Alexa Fluor 488 binding assay and Western blotting. Aviculin (2) was found to induce apoptotic cell death through the intrinsic apoptosis pathway, as indicated by the increased expression of initiator caspase-9, executioner caspase-7, and poly (ADP-ribose) polymerase (PARP). Aviculin (2)-induced apoptotic cell death was accompanied by an increase in the Bax/Bcl-2 ratio. These findings demonstrated that aviculin (2) could induce breast cancer cell apoptosis through the intrinsic apoptosis pathway, and it can therefore be considered an excellent candidate for herbal treatment of breast cancer.

Polyphenolic Compounds from Lespedeza Bicolor Root Bark Inhibit Progression of Human Prostate Cancer Cells via Induction of Apoptosis and Cell Cycle Arrest.

From a root bark of Lespedeza bicolor Turch we isolated two new (7 and 8) and six previously known compounds (1-6) belonging to the group of prenylated polyphenols. Their structures were elucidated using mass spectrometry, nuclear magnetic resonance and circular dichroism spectroscopy. These natural compounds selectively inhibited human drug-resistant prostate cancer in vitro. Prenylated pterocarpans 1-3 prevented the cell cycle progression of human cancer cells in S-phase. This was accompanied by a reduced expression of mRNA corresponding to several human cyclin-dependent kinases (CDKs). In contrast, compounds 4-8 induced a G1-phase cell cycle arrest without any pronounced effect on CDKs mRNA expression. Interestingly, a non-substituted hydroxy group at C-8 of ring D of the pterocarpan skeleton of compounds 1-3 seems to be important for the CDKs inhibitory activity.

Antiproliferative Pterocarpans and Coumestans from Lespedeza bicolor.

Chromatographic purification of a methanol extract of the roots of Lespedeza bicolor led to the isolation of four new pterocarpans (1-4), two new coumestans (6 and 7), two new arylbenzofurans (8 and 9), and the known pterocarpan 1-methoxyerythrabyssin II (5). Their structures were identified using NMR spectroscopy, UV spectroscopy, and mass spectrometry. Cytotoxicity assays showed that compounds 1-9 exerted antiproliferative effects on blood cancer cells. Of these compounds, 1 and 6 induced mitochondrial depolarization and induced apoptosis in Jurkat cells. These compounds promoted cell death by inducing cell-cycle arrest at the G1 stage, reducing levels of BCL2, and increasing cleavage of PARP-1. These findings indicate that 1 and 6 are possible lead compounds for the treatment of human leukemia cells via intracellular signaling.

Cytotoxic polyphenolic compounds from Lespedeza bicolor stem bark.

Four new pterocarpans (6aR,11aR)-6a,11a-dihydrolespedezol A2 (2), (6aR,11aR)-2-isoprenyl-6a,11a-dihydrolespedezol A2 (3), (6aR,11aR,3'R)-6a,11a-dihydrolespedezol A3 (4), (6aR,11aR,3'S)-6a,11a-dihydrolespedezol A3 (5) and one new stilbenoid with 1,2-diketone fragment named bicoloketone (6) along with one previously known pterocarpen lespedezol A2 (1) have been isolated from Lespedeza bicolor stem bark using multistage column chromatography on polyamide and silica gel. The structures of the isolated polyphenolic compounds were determined by spectroscopic methods. The absolute configurations of 4 and 5 were determined by comparison of their electronic circular dichroism (ECD) spectra obtained experimentally and the spectra calculated using time-dependent density functional theory (TDDFT). The isolated compounds exhibited a moderate DPPH scavenging effect and ferric reducing power compared to the reference antioxidant quercetin. The cytotoxicity of compounds against three human cancer cell lines, HTB-19, Kyse-30, and HEPG-2, and two normal cell lines, RPE-1 and HEK-293, was tested using the MTT assay. Compound 3 showed the strongest cytotoxic activity against all cell lines (IC50 6.0-19.1 µM) compared with the positive control cisplatin. The other tested compounds possessed moderate cytotoxic activity against cancer cells.

Effects of Lespedeza Cuneata aqueous extract on testosterone-induced prostatic hyperplasia.

CONTEXT: Lespedeza cuneata G. Don (Fabaceae), has been used as a traditional treatment of various diseases. There is a report L. cuneata effects on hormone replacement therapy for endocrine-related disease. However, studies related to benign prostatic hyperplasia (BPH) have not been investigated. OBJECTIVE: The effects of L. cuneata aqueous extract (LCW) on testosterone-induced prostatic hyperplasia (TPH) were examined. MATERIALS AND METHODS: Male Wistar rats (10 weeks, 330-350 g) were randomly divided to 6 groups (n = 6): Control group; TPH group (3 mg/kg, s.c, daily); TPH + LCW (25, 50, 100 mg/kg); TPH + Finasteride 10 mg/kg for 6 weeks. At the end of treatment, histological change of prostate, serum dihydrotestosterone (DHT) level, mRNA expression of 5α-reductase, inflammatory factors, proliferating cell nuclear antigen (PCNA) and fibroblast growth factor-2 (FGF-2) in prostate were examined. Then, LCW was treated with BPH-1, a human BPH cell line, at 25, 50, 100 µg/mL for 24 h and examine mRNA level of androgen receptor (AR) and prostate-specific antigen (PSA). In addition, the content of vicenin-2 was analyzed. RESULTS: LCW treatment of TPH inhibited serum DHT levels by 54.5, 51.2 and 54.1% and mRNA expression of 5α-reductase were inhibited 54.3, 61.3 and 73.6%, respectively. In addition, mRNA expression of inflammatory factors, PCNA and FGF-2 were decreased in the prostate of rats. Also, LCW attenuated mRNA level of AR and PSA in BPH-1 cell. The content of vicenin-2 in the LCW was analyzed to 0.89 mg/g. DISCUSSION AND CONCLUSIONS: Based on the results, LCW is a potential pharmacological candidate for the treatment of prostatic hyperplasia.

Effects of lespedeza condensed tannins alone or with monensin, soybean oil, and coconut oil on feed intake, growth, digestion, ruminal methane emission, and heat energy by yearling Alpine doelings.

Fifty-four Alpine doelings (initial BW and age of 31.7 ± 0.38 kg and 306 ± 1.9 d, respectively) were allocated to nine treatments individually fed for ad libitum intake of 25% concentrate and 75% forage diets (DM basis). Alfalfa was the forage in the control diet. Other diets contained Sericea lespedeza as the forage, with 1.25% DM of quebracho extract included in the concentrate fraction for a dietary condensed tannin level of 8.4%. Lespedeza treatments were no additive (L) and inclusion of monensin (I) at 22 mg/kg DM (L-I), soybean oil at 3% (L-S), coconut oil at 3% (L-N), I and 3% soybean oil (L-I-S), I and 3% coconut oil (L-I-N), 1.5% soybean oil and 1.5% coconut oil (L-S-N), and I, 1.5% soybean oil, and 1.5% coconut oil (L-I-S-N). The experiment was 12 wk with two 6-wk periods. Gas exchange was determined in weeks 6 and 12, and other measures occurred in weeks 5 and 11. The control diet offered averaged 2.67% nitrogen, 43.8% neutral detergent fiber, and 8.8% acid detergent lignin, and the L diet offered averaged 2.03% nitrogen, 42.8% neutral detergent fiber, and 13.2% acid detergent lignin. There were no treatment × period interactions for digestibilities (P ≥ 0.770) or methane emission (P ≥ 0.324). There were differences (P < 0.001) between the control treatment and diets with lespedeza in intake of DM (1.46, 1.23, 1.30, 1.18, 1.32, 1.10, 1.02, 1.20, and 1.01 kg/d; SEM = 0.059), digestibility of OM (57.4%, 50.9%, 51.8%, 52.7%, 50.3%, 52.1%, 52.1%, 51.9%, and 49.8%; SEM = 1.42), and digestibility of nitrogen (59.1%, 31.2%, 32.5%, 37.1%, 31.6%, 38.3%, 30.4%, 38.4%, and 34.1% for control, L, L-I, L-S, L-N, L-I-S, L-I-N, L-S-N, and L-I-S-N, respectively; SEM = 2.21). Ruminal methane emission was less (P < 0.001) for diets with lespedeza than for the control in MJ/d (1.36, 0.76, 0.84, 0.71, 0.71, 0.66, 0.65, 0.68, and 0.68; SEM = 0.048) and relative to intake of gross energy (5.92%, 3.27%, 3.49%, 3.19%, 2.84%, 2.91%, 3.20%, 3.20%, and 3.27%; SEM = 0.165) and digestible energy (11.19%, 6.98%, 7.40%, 6.38%, 5.90%, 5.69%, 6.37%, 6.38%, and 6.70% for control, L, L-I, L-S, L-N, L-I-S, L-I-N, L-S-N, and L-I-S-N, respectively; SEM = 0.400). In conclusion, the magnitude of effect of condensed tannins from lespedeza and quebracho extract on ruminal methane emission by Alpine doelings did not diminish over time and was not markedly influenced by dietary inclusion of monensin, soybean oil, or coconut oil.

Lespedeza cuneata protects the endothelial dysfunction via eNOS phosphorylation of PI3K/Akt signaling pathway in HUVECs.

BACKGROUND: Lespedeza cuneata G.Don (LCE), which belongs to the genus Lespedeza (Leguminosae), is a traditional oriental medicine known to prevent diabetes and cardiovascular diseases. However, no scientific studies about the effectiveness of LCE, their responsible bioactive constituents, and its mechanisms against endothelial dysfunction have been performed. PURPOSE: This study was performed to investigate the role of LCE and its chemical components in ameliorating endothelial dysfunction. METHODS: The production of nitric oxide (NO) was evaluated after LCE treatment in HUVECs. Cell viability was measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reagent. Western blot analysis was performed to determine the protein expression of endothelial nitric oxide synthase (eNOS) and protein kinase B (PKB, also known as Akt) in human umbilical vein endothelial cells (HUVECs). RESULTS: Pretreatment with L-NAME and LY294002 significantly decreased the LCE-induced NO production, as well as eNOS and Akt phosphorylation. ß-Sitosterol and ß-Sitosterol 6'-linolenoyl-3-O-ß-D-glucopyranoside are the bioactive constituents increase NO production as well as eNOS phosphorylation. CONCLUSION: Our findings suggest that LCE increase NO production via eNOS phosphorylation of PI3K/Akt signaling pathway.

Lignan Glycosides and Flavonoid Glycosides from the Aerial Portion of Lespedeza cuneata and Their Biological Evaluations.

Lespedeza cuneata (Fabaceae), known as Chinese bushclover, has been used in traditional medicines for the treatment of diseases including diabetes, hematuria, and insomnia. As part of a continuing search for bioactive constituents from Korean medicinal plant sources, phytochemical analysis of the aerial portion of L. cuneata led to the isolation of two new lignan glycosides (1,2) along with three known lignan glycosides (3⁻7) and nine known flavonoid glycosides (8⁻14). Numerous analysis techniques, including 1D and 2D NMR spectroscopy, CD spectroscopy, HR-MS, and chemical reactions, were utilized for structural elucidation of the new compounds (1,2). The isolated compounds were evaluated for their applicability in medicinal use using cell-based assays. Compounds 1 and 4⁻6 exhibited weak cytotoxicity against four human breast cancer cell lines (Bt549, MCF7, MDA-MB-231, and HCC70) (IC50 < 30.0 µM). However, none of the isolated compounds showed significant antiviral activity against PR8, HRV1B, or CVB3. In addition, compound 10 produced fewer lipid droplets in Oil Red O staining of mouse mesenchymal stem cells compared to the untreated negative control without altering the amount of alkaline phosphatase staining.

Ultrasonicated Lespedeza cuneata extract prevents TNF-α-induced early atherosclerosis in vitro and in vivo.

This study evaluated the use of ultrasonication to extract Lespedeza cuneata as a potential nutraceutical for preventing vascular inflammation. Ultrasonicated L. cuneata extract (ULCE) was prepared using 20% ethanol and 2 h of ultrasonication at room temperature, and its effects were investigated using relevant in vitro and in vivo models. ULCE suppressed tumor necrosis factor-alpha (TNF-α)-induced adhesion capacity, vascular cell adhesion protein 1 (VCAM-1) expression, and nuclear factor kappa-B (NF-κB) activity in human umbilical vein endothelial cells (HUVECs). ULCE also suppressed TNF-α-induced NF-κB signaling pathways and p65 translocation from the cytosol to the nucleus, as well as the mRNA expression of IL-1ß, IL-6, and TNF-α in HUVECs. Oral administration of ULCE suppressed TNF-α-induced monocyte infiltration into the intima and VCAM-1 expression, as well as the IL-1ß, IL-6, TNF-α, and monocyte chemoattractant protein-1 (MCP-1) mRNA expression in the main artery in mice. Among the compounds identified in the hydrolyzed ULCE, quercetin exhibited the strongest inhibitory effect against TNF-α-induced cell adhesion capacity. These results demonstrate that ULCE contains potent preventive factors against early atherosclerosis, which act by suppressing the NF-κB and VCAM-1 signaling axis.

PER, a Circadian Clock Component, Mediates the Suppression of MMP-1 Expression in HaCaT Keratinocytes by cAMP.

Skin circadian clock system responds to daily changes, thereby regulating skin functions. Exposure of the skin to UV irradiation induces the expression of matrix metalloproteinase-1 (MMP-1) and causes DNA damage. It has been reported both DNA repair and DNA replication are regulated by the circadian clock in mouse skin. However, the molecular link between circadian clock and MMP-1 has little been investigated. We found PERIOD protein, a morning clock component, represses the expression of MMP-1 in human keratinocytes by using a PER-knockdown strategy. Treatment with siPer3 alleviated the suppression of MMP-1 expression induced by forskolin. Results revealed PER3 suppresses the expression of MMP-1 via cAMP signaling pathway. Additionally, we screened for an activator of PER that could repress the expression of MMP-1 using HaCaT cell line containing PER promoter-luciferase reporter gene. Results showed Lespedeza capitate extract (LCE) increased PER promoter activity. LCE inhibited the expression of MMP-1 and its effect of LCE was abolished in knockdown of PER2 or PER3, demonstrating LCE can repress the expression of MMP-1 through PER. Since circadian clock component PER can regulate MMP-1 expression, it might be a new molecular mechanism to develop therapeutics to alleviate skin aging and skin cancer.

(-)-9'-O-(α-l-Rhamnopyranosyl)lyoniresinol from Lespedeza cuneata suppresses ovarian cancer cell proliferation through induction of apoptosis.

Lespedeza cuneata (Dum. Cours.) G. Don. (Fabaceae), known as Chinese bushclover or sericea lespedeza, has been used in traditional medicine to treat diabetes, hematuria, and insomnia, and it has been reported that bioactive compounds from L. cuneata possess various pharmacological properties. However, there has been no study to determine the active compounds from L. cuneata with potential activity against ovarian cancer. This study aimed to isolate cytotoxic compounds from L. cuneata and identify the molecular mechanisms underlying the apoptosis pathway in ovarian cancer cells. Based on cytotoxic activity identified in the screening test, chemical investigation of the active fraction of L. cuneata led to the isolation of nine compounds including four lignanosides (1-4), three flavonoid glycosides (5-7), and two phenolics (8-9). Cytotoxicity and the molecular mechanism were examined by methyl thiazolyl tetrazolium (MTT) assay and Western blot analysis. Of the isolated compounds, (-)-9'-O-(α-l-rhamnopyranosyl)lyoniresinol (3) demonstrated the strongest effect in suppressing A2780 human ovarian carcinoma cell proliferation in a dose-dependent manner, with an IC50 value of 35.40 ±â€¯2.78 µM. Control A2780 cells had normal morphology, whereas cell blebbing, shrinkage, and condensation were observed after treatment with compound 3. Western blotting analysis showed that compound 3 inhibited A2780 human ovarian cancer cell viability by activating caspase-8, caspase-3, and PARP, which contributed to apoptotic cell death. These results suggest that (-)-9'-O-(α-l-rhamnopyranosyl)lyoniresinol (3) has potent anticancer activities against A2780 human ovarian carcinoma cells through the extrinsic apoptotic pathway. Therefore, (-)-9'-O-(α-l-rhamnopyranosyl)lyoniresinol is an excellent candidate for the development of novel chemotherapeutics.

A new phenyldilactone from Lespedeza cuneata.

A new phenyldilactone, maysedilactone B (1), together with twenty known compounds, were isolated from the aerial parts of Lespedeza cuneata. The structural elucidation of the isolated compounds was primarily based on HR-ESI-MS, IR and 1D and 2D NMR analyses. Compounds 1-8 and 15-21 were tested for cytotoxicity against four human tumor cell lines (A549, HCT116, SKOV3, and HepG2) using MTT method in vitro, while no significant activities were observed for the evaluated compounds.

Effect of replacing alfalfa with panicled-tick clover or sericea lespedeza in corn-alfalfa-based substrates on in vitro ruminal methane production.

Methane emissions from ruminant livestock contribute to total anthropogenic greenhouse gas emissions and reduce metabolizable energy intake by the animal. Condensed tannins (CT) are polyphenolic plant secondary compounds commonly produced by some perennial forage legumes that characteristically bind to protein, carbohydrates, and minerals. The degree to which CT may affect ruminant nutrition depends upon the concentration, structural composition, and biological activity of the CT. The objective of our experiment was to determine the effect of replacing alfalfa in a corn-alfalfa-based substrate with a legume containing CT on in vitro CH4 production and the dynamics of fermentation using an in vitro gas production technique. All fermented substrates contained 50% ground corn as the energy concentrate portion, whereas the forage portion (50%) of each diet was comprised of alfalfa (control) or some combination of alfalfa and sericea lespedeza (SL) or panicled-tick clover (PTC). Our treatments consisted of PTC or SL 15, 30, and 45, which corresponded with 15, 30, or 45% replacement of the diet (alfalfa component) with either PTC or SL. Substrates containing 45% PTC or SL reduced in vitro CH4 production. Treatments did not affect total gas production as compared with that of the control. Replacement of alfalfa with SL or PTC increased fermentable organic matter (FOM). The PTC treatment increased FOM by as much as 1.8% at the 45% replacement level, whereas FOM of SL 45 was increased by less than 1%. The replacement of alfalfa with PTC increased substrate nutritive value greater than replacement with SL. There were no correlations between any physicochemical constituent of the substrates and CH4 production. A combination of factors associated with the inclusion of PTC and SL contributed to the in vitro CH4 production, and CT in these forages was likely a major contributing factor. Further confirmation of these results on in situ or in vivo animal systems is required. If proven effective in an in vivo production scenario, replacement of commonly fed non-CT-containing legumes, such as alfalfa, with legumes containing CT might be a viable method to decrease the effect of animal agriculture on greenhouse gas production.

Lespedeza davurica (Lax.) Schindl. extract protects against cytokine-induced ß-cell damage and streptozotocin-induced diabetes.

Lespedeza has been used for the management of diabetes in folklore medicine. The purpose of this study is to investigate the protective effects of the methanol extract of Lespedeza davurica (LD) on cytokine-induced ß-cell damage and streptozotocin- (STZ-) induced diabetes. RINm5F cells were treated with interleukin- (IL-) 1ß and interferon- (IFN-) γ to induce pancreatic ß-cell damage. The exposure of LD extract significantly decreased cell death, nitric oxide (NO) production, nitric oxide synthase (iNOS) expression, and nucleus factor-kappa B (NF-κB) p65 activation. Antidiabetic effects of LD extract were observed by oral glucose tolerance test (OGTT) in normal rats and by checking the biochemical, physiological, and histopathological parameters in STZ-induced diabetic rats. In OGTT, glucose clearance levels improved by oral treatment of LD extract. The water intake, urine volume, blood glucose, and serum TG, TC, TBARS, and DPP-IV levels were significantly decreased, and liver glycogen content was significantly increased by treatment of LD extract (250 mg/kg BW) in STZ-induced diabetic rats. Also, insulin immunoreactivity of the pancreases was increased in LD extract administrated rats compared with diabetic control rats. These results indicate that LD extract may protect pancreatic ß-cell damage and regulate the blood glucose in STZ-induced diabetic rats.

Hepatoprotective effect of flavonoid glycosides from Lespedeza cuneata against oxidative stress induced by tert-butyl hyperoxide.

The aerial parts of Lespedeza cuneata G. Don, perennial legume native to Eastern Asia, have been used therapeutically in traditional Asian medicine to protect the function of liver, kidneys and lungs. However, little is known about the pharmaceutical effect of extracts from this plant. In the present study, the aerial parts of L. cuneata were used to prepare an ethanol extract, which was then tested for hepatoprotective effects against injury by tert-butyl hyperoxide (t-BHP). At a dose of 20 µg/mL, the ethanol extract significantly protected HepG2 cells against the cytotoxicity of t-BHP. Further fractionation of the extract with ethyl acetate allowed the isolation of five flavonoid compounds that were structurally identified by ¹H and ¹³C NMR spectroscopy as isovitexin, hirsutrin, trifolin, avicularin and quercetin. Hirsutrin, avicularin and quercetin (10 µM) showed clear hepatoprotective activity against injury by t-BHP in HepG2 cells, whereas isovitexin and trifolin showed no protective effects. The observed hepatoprotective effect of the investigated compounds showed a high correlation with radical scavenging activity, which followed the structure-activity relationships of the flavonoid aglycones.

The melanin synthesis inhibition and radical scavenging activities of compounds isolated from the aerial part of Lespedeza cyrtobotrya.

The EtOAc fraction of Lespedeza cyrtobotrya showed mushroom tyrosinase inhibitory and radical scavenging activity. Four active compounds were isolated based on LH-20 chromatography and HPLC, and the structures were elucidated on the basis of their LC-MS and NMR spectral data, as 2-(2,4-Dihydroxyphenyl)-6-hydroxybenzofuran (1), eriodictyol-7-O-glucopyranoside (2), haginin A (3), and dalbergioidin (4), respectively. 2-(2,4-Dihydroxyphenyl)-6-hydroxybenzofuran (1) showed mushroom tyrosinase inhibitory activity with an IC50 value of 5.2 micronM and acted as a competitive inhibitor. Furthermore, 37.3 micronM of compound 1 reduced 50 % of the melanin content on a human melanoma (MNT-1) cells. The radical scavenging activity of 2-(2,4-dihydroxyphenyl)-6-hydroxybenzofuran (1), eriodictyol-7-O-glucopyranoside (2), haginin A (3), and dalbergioidin (4) was shown with IC50 values of 11.0, 24.5, 9.0 and 36.5 micronM in an ABTS system and with IC50 values of 42.7, 36.0, 37.7 and 61.7 micronM in a DPPH system, respectively. The mushroom tyrosinase inhibitory activity of EtOAc fraction of Lespedeza cyrtobotrya was contributed by compound 1, 3 and 4, and radical scavenging activity of it was contributed by compound 1-4.