PMID: 

Eur J Clin Nutr. 2015 Jan ;69(1):134-42. Epub 2014 Nov 5. PMID: 25369831

Abstract Title: 

Red clover isoflavones enriched with formononetin lower serum LDL cholesterol-a randomized, double-blind, placebo-controlled study.

Abstract: 

BACKGROUND: Although postmenopausal combined hormone replacement therapy reduces the risk of hip fracture, long-term use may be associated with an increased risk of breast cancer, and in women more than 10 years after menopause it is associated with an increased risk of cardiovascular disease. Isoflavones, because of preferential binding to estrogen receptor beta, may retain the beneficial effects on bone but lessen the adverse effects on the breast.OBJECTIVE: The objective of this study was to study the effects of an isoflavone obtained from red clover (Rimostil) on bone mineral density, and on low-density lipoprotein (LDL) cholesterol.DESIGN: In a double-blind, randomized, placebo-controlled trial, 50 mg of Rimostil was given to women who were menopausal for at least 1 year. Bone mineral density of the spine, femoral neck and forearm and serum LDL cholesterol were measured at baseline and at 6-month intervals. The duration of follow-up was 2 years.RESULTS: There was no beneficial effect of Rimostil on bone density at any site. There was a 12% fall in serum LDL cholesterol in the Rimostil-treated arm, which was significantly greater than the 2% drop seen in the control arm (P=0.005).

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PMID: 

Menopause. 2006 Mar-Apr;13(2):251-64. PMID: 16645539

Abstract Title: 

Clinical studies of red clover (Trifolium pratense) dietary supplements in menopause: a literature review.

Abstract: 

Red clover (Trifolium pratense L., Fabaceae) botanical dietary supplements have received much attention recently for their potential use in the treatment of menopause symptoms, maintenance/improvement of bone and cardiovascular health, and reported benign effects on the breast and endometrium. Literature searches of four computerized databases were run to identify clinical studies of red clover botanical dietary supplements. The manufacturer of the red clover products used in the majority of the studies was contacted for unpublished information and/or clarification regarding the chemical content of their products. Red clover studies were reviewed that pertained to women's health or menopause. Clinical evidence is presently lacking to support the efficacy of semipurified red clover isoflavone extracts for alleviation of climacteric vasomotor symptoms or reduction of low-density lipoprotein levels in the blood. Furthermore, the safety of use of red clover isoflavone supplements in patients with breast or endometrial cancer has not been established. Limited evidence suggests possible efficacy in maintenance of bone health and improvement of arterial compliance, a risk factor for atherosclerosis. This literature review covers red clover botanical dietary supplement clinical studies having a possible impact on the health care of mature and menopausal women, and provides historical perspective regarding the traditional uses of red clover.

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PMID: 

Am J Clin Nutr. 2017 Sep ;106(3):909-920. Epub 2017 Aug 2. PMID: 28768651

Abstract Title: 

Combined bioavailable isoflavones and probiotics improve bone status and estrogen metabolism in postmenopausal osteopenic women: a randomized controlled trial.

Abstract: 

Female age-related estrogen deficiency increases the risk of osteoporosis, which can be effectively treated with the use of hormone replacement therapy. However, hormone replacement therapy is demonstrated to increase cancer risk. Bioavailable isoflavones with selective estrogen receptor affinity show potential to prevent and treat osteoporosis while minimizing or eliminating carcinogenic side effects.In this study, we sought to determine the beneficial effects of a bioavailable isoflavone and probiotic treatment against postmenopausal osteopenia.We used a novel red clover extract (RCE) rich in isoflavone aglycones and probiotics to concomitantly promote uptake and a favorable intestinal bacterial profile to enhance isoflavone bioavailability. This was a 12-mo, double-blind, parallel design, placebo-controlled, randomized controlled trial of 78 postmenopausal osteopenic women supplemented with calcium (1200 mg/d), magnesium (550 mg/d), and calcitriol (25μg/d) given either RCE (60 mg isoflavone aglycones/d and probiotics) or a masked placebo [control (CON)].RCE significantly attenuated bone mineral density (BMD) loss at the L2-L4 lumbar spine vertebra (

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Int J Biol Macromol. 2020 Jan 21 ;148:750-760. Epub 2020 Jan 21. PMID: 31978472

Abstract Title: 

Extraction, purification, hypoglycemic and antioxidant activities of red clover (Trifolium pratense L.) polysaccharides.

Abstract: 

Hot water extraction was applied to extract red clover (Trifolium pratense L.) polysaccharides (RCP) and the extraction conditions were optimized using the response surface methodology (RSM). An RCP yield of 12.72 ± 0.14% was achieved under the optimum extraction conditions: extracting time of 95 min, extracting temperature of 93 °C, and solvent-material ratio of 21 mL/g. A component named RCP-1.1 with the molecular weight of 7528.81 kDa was purified from RCP. RCP-1.1 was composed of glucose, galacturonic acid, arabinose, and galactose, with molar percentages of 52.54, 1.04, 16.31, and 30.11%, respectively. At the determination concentration of 10 mg/mL, the α-glucosidase inhibition ability of RCP-1.1 reached 86.72% of that of acarbose. The scavenging rates of RCP-1.1 (3.0 mg/mL) for DPPH and ABTS radicals reached 91.82% and 98.95% of that of ascorbic acid (3.0 mg/mL), respectively. Based on these results, RCP-1.1 possesses the potential to be used as a natural hypoglycemic agent or an antioxidant.

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PMID: 

Maturitas. 2020 Feb ;132:7-16. Epub 2019 Nov 10. PMID: 31883666

Abstract Title: 

Effects of red clover (Trifolium pratense) isoflavones on the lipid profile of perimenopausal and postmenopausal women-A systematic review and meta-analysis.

Abstract: 

AIM: The aim of this systematic review and meta-analysis was to clarify the effect of a specific standardised extract of red clover (Trifolium pratense) on the lipid profile of perimenopausal and postmenopausal women.
METHODS: Medline (PubMed), EMBASE, and Cochrane Library electronic databases were searched for papers in English reporting randomized controlled trials published up to 2017. Reference lists from those papers were checked for further relevant publications. Studies were identified and reviewed for their eligibility for inclusion in this review. The changes from baseline in the levels of individual components of the lipid profiles were used to assess differences between the active treatment and placebo groups. Weighted mean differences and 95 % confidence intervals were calculated for continuous data using a random-effects model.
RESULTS: Ten eligible studies (twelve comparisons) with 910 peri- and postmenopausal women were selected for systematic review. The meta-analysis showed changes in serum levels: total cholesterol, -0.29 (95 % CI: -0.53 to -0.06) mmol/L [-11.21 (95 % CI: -20.49 to -13.92) mg/dL], p = 0.0136; LDL-cholesterol, -0.13 (95 % CI: -0.35 to 0.09) mmol/L [-5.02 (95 % CI: -13.53 to 3.48) mg/dL], p = 0.2418; triglycerides, -0.15 (95 % CI: -0.32 to 0.01) mmol/L [-13.28 (95 % CI: -28.34 to 0.88) mg/dL], p = 0.0592; and HDL-cholesterol, 0.14 (95 % CI: -0.08 to 0.36) mmol/L [5.41 (95 % CI: -3.09-13.92) mg/dL], p = 0.2103. TheIstatistic ranged from 87.95%-98.30 %, indicating significant heterogeneity.
CONCLUSIONS: The results suggest that a red clover extract is efficacious in reducing the concentrations of total cholesterol; however, changes in HDL-C, LDL-C and triglycerides are not as pronounced. Potentially, this means that women takingTrifolium pratense for menopausal symptoms can derive additional benefits from the plant's specific effect that corrects abnormal cholesterol levels. Additional studies are needed to assess its effects on post-menopausal women.

PMID: 

Cells. 2020 Feb 6 ;9(2). Epub 2020 Feb 6. PMID: 32041350

Abstract Title: 

Blocks Proliferation in Chronic Myelogenous Leukemia via the BCR-ABL/STAT5 Pathway.

Abstract: 

Some species of clover are reported to have beneficial effects in human diseases. However, little is known about the activity of the forage plant, or white clover, which has been recently found to exert a hepatoprotective action. Scientific interest is increasingly focused on identifying new drugs, especially natural products and their derivatives, to treat human diseases including cancer. We analyzed the anticancer effects ofin several cancer cell lines. The phytochemical components ofwere first extracted in a methanol solution and then separated into four fractions by ultra-high-performance liquid chromatography. The effects of the total extract and each fraction on cancer cell proliferation were analyzed by MTT assay and Western blotting.and, more robustly, its isoflavonoid-rich fraction showed high cytotoxic effects in chronic myelogenous leukemia (CML) K562 cells, with ICvalues of 1.67 and 0.092 mg/mL, respectively. The block of cell growth was associated with a total inhibition of BCR-ABL/STAT5 and activation of the p38 signaling pathways. In contrast, these strongly cytotoxic effects did not occur in normal cells. Our findings suggest that the development of novel compounds derived from phytochemical molecules contained inmight lead to the identification of new therapeutic agents active against CML.

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PMID: 

Nutrients. 2020 Jan 19 ;12(1). Epub 2020 Jan 19. PMID: 31963833

Abstract Title: 

The Effects ofL. Sprouts' Phenolic Compounds on Cell Growth and Migration of MDA-MB-231, MCF-7 and HUVEC Cells.

Abstract: 

Uncontrolled growth and migration and invasion abilities are common for cancer cells in malignant tumors with low therapeutic effectiveness and high mortality and morbidity. Estrogen receptorβ (ERβ), as a member of the nuclear receptor superfamily, shows potent tumor suppressive activities in many cancers. Phytoestrogens' structural resemblance to 17 β-estradiol allows their binding to ERβ isoform predominantly, and therefore, expression of genes connected with elevated proliferation, motility and invasiveness of cancer cells may be downregulated. Among polyphenolic compounds with phytoestrogenic activity, there are isoflavones fromL. (red clover) sprouts, containing high amounts of formononetin and biochanin A and their glycosides. To determine the source of the most biologically active isoflavones, we obtained four extracts from sprouts before and after their lactic fermentation and/orβ-glucosidase treatment. Our previous results of ITC (isothermal titration calorimetry) modelling and a docking simulation showed clover isoflavones' affinity to ERβ binding, which may downregulate cancer cell proliferation and migration. Thus, the biological activity ofsprouts' extracts was checked under in vitro conditions against highly invasive human breast cancer cell line MDA-MB-231 and non-invasive human breast cancer cell line MCF-7 cells. To compare extracts' activities acquired for cancer cells with those activities against normal cells, as a third model we choose human umbilical vein endothelial cells (HUVEC), which, due to their migration abilities, are involved in blood vessel formation. Extracts obtained from fermented sprouts at ICdosages were able to inhibit migration of breast cancer cells through their influence on intracellular ROS generation; membrane stiffening; adhesion; regulation of MMP-9, N-cadherin and E-cadherin at transcriptional level; or VEGF secretion. Simultaneously, isolated phenolics revealed no toxicity against normal HUVEC cells. In the manuscript, we proposed a preliminary mechanism accounting for the in vitro activity ofL. isoflavones. In this manner,sprouts, especially after their lactic fermentation, can be considered a potent source of biological active phytoestrogens and a dietary supplement with anti-cancer and anti-invasion properties.

PMID: 

Cardiovasc Hematol Agents Med Chem. 2020 Feb 5. Epub 2020 Feb 5. PMID: 32026788

Abstract Title: 

Therapeutic potential of biochanin-A against isoproterenol -induced myocardial infarction in rats.

Abstract: 

BACKGROUND: This study determined the effect of Biochanin A (BCA) on isoproterenol (ISO) induced myocardial infarction (MI) in male Wistar rats.METHODS: Animals (Weighing 150-180 g) were divided into four groups, with six animals in each group and pretreated with BCA (10mg/kg body weight [BW] andɑ-tocopherol (60mg/kg BW for 30 days; and ISO (20mg/kg BW) was administrated subcutaneously on the day 31 and 32.RESULTS: ISO-induced MI rats demonstrated the significant elevation of serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, lactate dehydrogenase, creatine kinase-MB and cardiac troponin; however, concomitant pretreatment with BCA protected the rats from cardiotoxicity caused by ISO. Activities of antioxidant enzymes, such as superoxide dismutase, catalase, glutathione peroxidase, glutathione-S-transferase and glutathione reductase had significantly reduced in the heart with ISO-induced MI. Pretreatment with BCA produced a marked reversal of these antioxidant enzymes related to MI¬¬-induced by ISO.CONCLUSION: In conclusion, this study suggested that BCA exerts cardioprotective effects through modulating lipid peroxidation, enhancing antioxidants, and detoxifying enzyme systems.

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PMID: 

Cardiovasc Hematol Agents Med Chem. 2020 Feb 6. Epub 2020 Feb 6. PMID: 32031077

Abstract Title: 

Antiobesity effect of Biochanin-A: Effect on trace element metabolism in high fat diet-induced obesity in rats.

Abstract: 

BACKGROUND: Imbalanced diets have contributed to the increased prevalence of obesity and other metabolic disorders in the modern world including trace element metabolism. However, the underlying mechanisms are not fully understood.AIM AND OBJECTIVES: The present study investigated the effects of Biochanin A (BCA) on the changes in element metabolism induced by HFD-induced obese rats.METHODS: BCA was administered orally for 30 days to experimental obese rats. Changes in body weight, glucose, insulin resistance and lipid profiles of plasma as well as the level of trace elements (Fe, Zn, Mg and Cu) in various tissues (liver, kidney, heart and pancreas) and hepsidine and heme oxygenase were observed in experimental rats.RESULTS: The administration of BCA elicited a significant (p

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PMID: 

Microb Pathog. 2020 Jan ;138:103846. Epub 2019 Nov 4. PMID: 31698051

Abstract Title: 

Biochanin A protect against lipopolysaccharide-induced acute lung injury in mice by regulating TLR4/NF-κB and PPAR-γ pathway.

Abstract: 

Acute lung injury (ALI) is a serious respiratory syndrome featured with uncontrolled inflammatory response. Biochanin A has been showed to possess and anti-inflammatory effect. This study intended to explore the suppression of biochanin A on lipopolysaccharide (LPS)-induced ALI in mice. Seven hours later LPS-induced ALI model established, the indexes including, pathological changes, MPO activity, wet/dry ratio, proinflammatory cytokines TNF-α, IL-1β, and IL-6, production, as well as and TLR4/NF-κB and PPAR-γ signaling pathway expression were compared bwtween different groups. In addition, bronchoalveolar lavage fluid (BALF) was collected and the levels of total protein, inflammatory cells and TNF-α, IL-1β, and IL-6 were detected.The results revealed that LPS lead to significantly lung pathological injury, and damage of lung vascular permeability showing by higher lung wet/dry ratio and total protein levels in the BALF when compared to the control group mice. However, these changes significantly reversed by biochanin A. Moreover, the levels of inflammatory cells in BALF, proinflammatory cytokines TNF-α, IL-1β, and IL-6, in both lung and BALF were also dose-dependently reduced by biochanin A during ALI process. To investigate the anti-inflammatory mechanisms of biochanin A, we found that biochanin A significantly inhibited the activation of TLR4/NF-κB signaling pathway induced by LPS. Furthermore, the expression of PPAR-γ also markedly increased in the mice after treated with biochanin A. In conclusion, biochanin A alleviated LPS-induced ALI by inhibiting the inflammatory response, which was mediated via down-regulating the activation of TLR4/NF-κB signaling pathway and enhancing the expression of PPAR-γ.

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