Lignan Metabolites Research Articles

Ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry analysis of In vitro metabolites of lignans from fructus forsythiae by Human Fecal Flora

Background: Phillyrin and (+)-pinoresinol 4-O-β-D-glucoside are the major active furofuran-type lignans in Fructus Forsythiae. The metabolic routes and metabolites of these two lignans are not well understood yet. Objective: In this study, we attempted to identify the human-intestine bacterial metabolites of lignans from Fructus Forsythiae. Materials and Methods: Two natural compounds, phillyrin and (+)-pinoresinol 4-O-β-D-glucoside were incubated with human fecal microflora in an anaerobic incubator for 72 h and the metabolites with highly sensitive ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) were analyzed. Results: As a result, nine metabolites were determined using a Syncronis™ C18column (particle size 1.7 mm) in a gradient elution system. These metabolites were then identified according to the mass fragmental mechanism, MS/MS fragment ions, and previous publications. The results of this study indicated that the major metabolites of furofuran-type lignans are through the processes of hydrolysis, demethylation, reduction, dehydroxylation, and oxidation. Conclusions: Lignans can be metabolized by intestinal microbiota and the intestinal bacteria play a critical role in the metabolism of components administered orally. Abbreviations used: GAM: General anaerobic medium; UPLC/Q-TOF-MS: Ultra-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry; ESI: Negative ion electrospray; RT: Retention time; TCM: Traditional Chinese Medicine.

Genistein and enterolactone in relation to Ki-67 expression and HER2 status in postmenopausal breast cancer patients.

Phytoestrogens (PE) may improve breast cancer prognosis by modifying tumor prognostic markers, such as cell proliferation marker Ki-67 and human epidermal growth factor receptor 2 (HER2). Epidemiological evidence linking lignans and isoflavones to Ki-67 and HER2 is limited. We examined associations between the major metabolites of lignans and isoflavones - enterolactone (ENL) and genistein (GEN) - respectively, and Ki-67 expression and HER2 in tumor tissue of breast cancer patients. Data from 1060 invasive breast cancer patients from the population-based MARIE study were used. Multivariate-adjusted linear (Ki-67 log-transformed) and quantile regression, and logistic regression analyses (HER2, Ki-67 dichotomized) were performed to calculate β estimates and ORs, respectively. Median post-diagnostic ENL and GEN concentrations were 19.5 and 4.8 nmol/L, respectively. Median Ki-67 was 12.0%, and 21.2% of the tumors were HER2+. After adjustment, there was an inverse association between GEN and Ki-67 at high expression levels (OR for Ki-67 ≥20% versus <20% of 0.93 (95%CI [0.87;0.99]) per 10 nmol/L GEN increment). Our findings indicate an inverse association between GEN and Ki-67 at high levels of Ki-67 expression. Additional investigations are recommended to confirm our findings and to further elucidate mechanisms linking PE metabolites to breast cancer survival.

Effect of Antibiotics and Diet on Enterolactone Concentration and Metabolome Studied by Targeted and Nontargeted LC-MS Metabolomics.

High plant lignan intake is associated with a number of health benefits, possibly induced by the lignan metabolite enterolactone (ENL). The gut microbiota plays a crucial role in converting dietary lignans into ENL, and epidemiological studies have shown that use of antibiotics is associated with lower levels of ENL. Here we investigate the link between antibiotic use and lignan metabolism in pigs using LC-MS/MS. The effect of lignan intake and antibiotic use on the gut microbial community and the pig metabolome is studied by 16S rRNA sequencing and nontargeted LC-MS. Treatment with antibiotics resulted in substantially lower concentrations of ENL compared with concentrations detected in untreated animals, whereas the plasma concentrations of plant lignans were unchanged. Both diet and antibiotic treatment affected the clustering of urinary metabolites and significantly altered the proportions of taxa in the gut microbiota. Diet, but not antibiotic treatment, affected the plasma lipid profile, and a lower concentration of LDL cholesterol was observed in the pigs fed a high lignan diet. This study provides solid support for the associations between ENL concentrations and use of antibiotics found in humans and indicates that the lower ENL concentration may be a consequence of the ecological changes in the microbiota.

Association of Polyphenol Biomarkers with Cardiovascular Disease and Mortality Risk: A Systematic Review and Meta-Analysis of Observational Studies.

Epidemiologic studies have suggested an inverse association between flavonoids and cardiovascular disease (CVD). However, the results might have been influenced by the use of dietary assessment methods, which are error prone. The aim of this paper was to systematically review and analyse the literature for evidence of associations between polyphenol biomarkers and CVD and mortality risk in observational studies. Eligible studies were identified through PubMed, Web of Science, and reference lists. Multivariable adjusted associations were extracted. Data were log-transformed and pooled using the random effects model. In total, eight studies were included, investigating 16 different polyphenol biomarkers in association with CVD and mortality. Blood and urine were used as biospecimens, and enterolactone, a lignan metabolite, was most often investigated. Three meta-analyses were conducted investigating the association between enterolactone, and all-cause and CVD mortality, and non-fatal myocardial infarction. A 30% and 45% reduced all-cause and CVD mortality risk were revealed at higher enterolactone concentrations. Furthermore, inverse associations were observed between polyphenol biomarkers and all-cause mortality, kaempferol, and acute coronary syndrome. There is evidence to suggest that enterolactone is associated with a lower CVD mortality risk. This emphasises the importance of the role of the microbiota in disease prevention. To strengthen the evidence, more studies are warranted.

Prediagnostic enterolactone concentrations and mortality among Danish men diagnosed with prostate cancer.

Evidence on the role of diet in relation to prostate cancer progression is sparse. Foods rich in lignans have shown beneficial effects on prostate cancer progression in both animal studies and small human intervention studies, including beneficial effects on prostate-specific antigen levels and tumour growth. The lignan metabolite, enterolactone, has further shown to slow prostate cancer cell growth in vitro. The aim was to investigate the association between prediagnostic enterolactone concentrations and mortality among men with prostate cancer.Subljects/Methods:Prediagnostic plasma concentrations of enterolactone from 1390 men diagnosed with prostate cancer from the Danish Diet, Cancer and Health cohort were related to all-cause or prostate cancer-specific death, using Cox proportional hazards models with follow-up time (from the date of diagnose until the date of death, emigration or end of follow-up by December 2013) as the underlying time axis. The hazard ratios for enterolactone concentrations assessed linearly by 20 nmol/l increments was 0.95 (0.90, 1.02) for all-cause mortality and 0.98 (0.92, 1.05) for prostate cancer-specific mortality. Categorisation of enterolactone concentrations into quartiles did not reveal a different pattern. No effect modifications by smoking, body mass index or sport were observed, and the associations did not differ by prostate cancer aggressiveness. We found no association between enterolactone concentrations and mortality among men diagnosed with prostate cancer.

Protocol for a 24-Week Randomized Controlled Study of Once-Daily Oral Dose of Flax Lignan to Healthy Older Adults.

BackgroundIncreased oxidative stress and inflammation are associated with aging, and contribute to an increased risk of chronic disease in older adults. Flaxseed lignans demonstrate antioxidant and anti-inflammatory activity, but their ability to reduce oxidative stress and inflammation markers in older adult populations has received limited investigation.ObjectiveThis is a chronic intervention trial of community-dwelling healthy older adults to examine the effects of a flaxseed lignan (secoisolariciresinol diglucoside; SDG) enriched supplement (BeneFlax) compared to a placebo. The primary aim was to demonstrate the safety of BeneFlax and confirm its anti-inflammatory efficacy on markers of oxidative stress and inflammation, and subsequent functional outcomes, including those associated with its anti-inflammatory efficacy. A secondary aim was to determine flaxseed lignan metabolite concentrations in blood.MethodsA double-blind randomized clinical trial was conducted. Subjects were healthy community-dwelling adults aged 60-80 years. Testing was performed at baseline, 8, 16, and 24 weeks. The 24-week intervention consisted of 600 milligrams (mg) of SDG daily or an equivalent amount (volume) of placebo. All participants received 1000 international units of vitamin D to ensure adequate vitamin D status. Measurements consisted of blood pressure, hematology, and tolerability for safety assessments; blood oxidative stress and inflammatory biomarkers for efficacy; and cognition, muscle strength, and pain as functional outcomes. Secondary endpoints of plasma levels of lignan metabolites were analyzed by mass spectrometry. Other tests, such as bone turnover markers and fecal levels of flax cyclolinopeptides, will be performed at a later date.ResultsThirty-two participants were recruited (19 intervention and 13 control) and all completed the trial. Numerous Health Canada-imposed exclusion criteria limited recruitment success. Analyses are ongoing, but the baseline data available for a number of parameters indicate no differences between treatment groups. Safety measures (vital signs) did not change from baseline and were not significantly different between treatment and placebo groups at 24 weeks.ConclusionsPreliminary results indicate that no safety concerns are associated with administering 600 mg SDG for 24 weeks to adults between the ages of 60 and 80 years.Trial RegistrationClinicaltrials.gov NCT01846117; https://clinicaltrials.gov/ct2/show/NCT01846117 (Archived by WebCite at http://www.webcitation.org/6nlDZNjmA)

Differential and directional estrogenic signaling pathways induced by enterolignans and their precursors.

Mammalian lignans or enterolignans are metabolites of plant lignans, an important category of phytochemicals. Although they are known to be associated with estrogenic activity, cell signaling pathways leading to specific cell functions, and especially the differences among lignans, have not been explored. We examined the estrogenic activity of enterolignans and their precursor plant lignans and cell signaling pathways for some cell functions, cell cycle and chemokine secretion. We used DNA microarray-based gene expression profiling in human breast cancer MCF-7 cells to examine the similarities, as well as the differences, among enterolignans, enterolactone and enterodiol, and their precursors, matairesinol, pinoresinol and sesamin. The profiles showed moderate to high levels of correlation (R values: 0.44 to 0.81) with that of estrogen (17β-estradiol or E2). Significant correlations were observed among lignans (R values: 0.77 to 0.97), and the correlations were higher for cell functions related to enzymes, signaling, proliferation and transport. All the enterolignans/precursors examined showed activation of the Erk1/2 and PI3K/Akt pathways, indicating the involvement of rapid signaling through the non-genomic estrogen signaling pathway. However, when their effects on specific cell functions, cell cycle progression and chemokine (MCP-1) secretion were examined, positive effects were observed only for enterolactone, suggesting that signals are given in certain directions at a position closer to cell functions. We hypothesized that, while estrogen signaling is initiated by the enterolignans/precursors examined, their signals are differentially and directionally modulated later in the pathways, resulting in the differences at the cell function level.

Nutrimetabolomics fingerprinting to identify biomarkers of whole grain wheat intake

Increased whole grain (WG) consumption has been inversely associated with the risk for developing some diet‐related disorders, such as type 2 diabetes, obesity, cancer, and cardiovascular diseases. Many epidemiological studies, however, have failed to generate consistent results on this topic due to a lack of accurate tools to assess dietary intake of whole grains. Nutritional epidemiology research needs more reliable and quantitative methods for measuring dietary intake of specific foods. Using a targeted LC/MS based metabolomic approach, we established the metabolic fingerprints of the major phytochemicals in WG wheat in human urine including metabolites of alkylresorcinols, benzoxazinoids, lignans, and phytosterols, as well as microbial‐derived metabolites. Using a non‐targeted LC/MS based global metabolomic approach, we investigated the impact of WG wheat bread intake on the endogenous urinary metabolome. Metabolic profiling revealed a number of differences relating to the consumption of WG wheat bread vs. refined grain wheat bread, such as 3,5‐dihydroxybenzoic acid, 2‐aminophenol sulfate, ferulic acid 4‐sulfate, 2‐acetamidophenol sulfate, and cysteine. Using the targeted and non‐targeted nutrimetabolomic approaches, we are able to develop multiple metabolites as the potential biomerkers of WG wheat intake.Support or Funding InformationUSDA‐NIFA grant 2012‐38821‐20012 to S. Sang

Flaxseed Lignan Metabolite, Enterolactone, Down‐regulated DNA Methyltransferase, Histone Deacetylase, and Methyl‐CpG‐binding Domain Protein Expression in Murine Adipocytes

Nutrition influences gene expression via epigenetic control mechanisms such DNA methylation and histone modification by DNA methyltransferase (DNMT) and histone deacetylase (HDAC), respectively. Oxidative stress caused by reactive oxygen species (ROS) damages DNA or cells which can be regulated by antioxidants. Flaxseed is the richest source of secoisolariciresinol diglucoside, which is metabolized into enterolactone by bacteria in the gut. Enterolactone has antioxidant activities which lower ROS induced oxidative damages. Thus, this study was conducted to determine the effects of enterolactone on DNMTs, HDACs, and methyl‐CpG‐binding domain protein (MBD) expression in ROS treated murine adipocytes. Enterolactone (0, 10, 50, or 100 μM) or control antioxidant, such as superoxide dismutase (100 U/mL) or catalase (100 U/mL), treated adipocytes (1×107) were treated with hydroxyl or superoxide radical, and then total RNA was extracted and purified using an RNeasy Mini kit. DNMTs, HDACs, and MBD2 expression was measured by real‐time quantitative PCR. The fold change in DNMTs, HDACs, and MBD2 expression was calculated using 2−ΔΔCT method. The results showed that enterolactone down‐regulates DNMTs, HDACs, and MBD2 expression of hydroxyl radical treated adipocytes. However, no significant effects of enterolactone on DNMTs, HDACs, and MBD2 gene expression were seen in superoxide radical treated adipocytes. How DNMTs, HDACs, and MBD2 expression of hydroxyl radical treated adipocytes was down‐regulated by enterolactone is unknown and needs further study.Support or Funding InformationThe research was funded by grants from the Department of Health, Nutrition, and Exercise Sciences and College of Human Development and Education at North Dakota State University.

Antidiabetic effect of enterolactone in cultured muscle cells and in type 2 diabetic model db/db mice.

Enterolactone (ENL) is formed by the conversion of dietary precursors like strawberry lignans via the gut microbiota. Urinary concentrations of lignan metabolites are reported to be significantly associated with a lower risk of Type 2 diabetes (T2D). In the present study, antidiabetic effect of ENL and its modes of action were studied in vitro and in vivo employing a rat skeletal muscle-derived cell line, L6 myocytes in culture, and T2D model db/db mice. ENL dose-dependently increased glucose uptake in L6 myotubes under insulin absent condition. This increase by ENL was canceled by compound C, an inhibitor of 5'-adenosine monophosphate-activated protein kinase (APMK). Activation (=phosphorylation) of AMPK and translocation of glucose transporter 4 (GLUT4) to plasma membrane in L6 myotubes were demonstrated by Western blotting analyses. Promotion by ENL of GLUT4 translocation to plasma membrane was also visually demonstrated by immunocytochemistry in L6 myoblasts that were transfected with glut4 cDNA-coding vector. T2D model db/db mice were fed the basal 20% casein diet (20C) or 20C supplemented with ENL (0.001 or 0.01%) for 6weeks. Fasting blood glucose (FBG) levels were measured every week and intraperitoneal glucose tolerance test (IPGTT) was conducted. ENL at a higher dose (0.01% in 20C) suppressed the increases in FBG levels. ENL was also demonstrated to improve the index of insulin resistance (HOMA-IR) and glucose intolerance by IPGTT in db/db mice. From these results, ENL is suggested to be an antidiabetic chemical entity converted from dietary lignans by gut microbiota.

Urine phyto-oestrogen metabolites are not significantly associated with risk of type 2 diabetes: the Singapore Chinese health study.

We evaluated the relationship between urine concentrations of phyto-oestrogens (isoflavones and lignans) and risk of incident type 2 diabetes in middle-aged and elderly Chinese residing in Singapore. Urine metabolites of isoflavones and lignans were assayed by HPLC among 564 diabetes cases and 564 matched controls in a case-control study nested within the Singapore Chinese Health Study cohort. Participants were free of diagnosed diabetes, CVD and cancer at morning urine collections during 1999-2004. Cases were participants who reported to have physician-diagnosed diabetes at follow-up visits during 2006-2010, whereas controls were randomly selected among those who remained free of diabetes and were matched to the index cases by age, sex, dialect group and date of urine collection. Conditional logistic regression models were used to calculate OR and 95 % CI with adjustment for potential confounders. The mean age of the participants at the time of urine collection was 59·8 years, and the average interval between urine collection and diabetes diagnosis was 4·0 years. The multivariate-adjusted OR for diabetes were 1·00 (reference), 0·76 (95 % CI 0·52, 1·11), 0·78 (95 % CI 0·53, 1·14) and 0·79 (95 % CI 0·54, 1·15) across quartiles of urine isoflavones (P for trend=0·54), and were 1·00 (reference), 0·87 (95 % CI 0·60, 1·27), 1·10 (95 % CI 0·77, 1·56) and 0·93 (95 % CI 0·63, 1·37) for lignans (P for trend=0·93). The results were similar in men and women, as well as for individual metabolites of isoflavones (genistein, daidzein, glycitin and equol) or lignans (enterodiol and enterolactone). The present study did not find a significant association between urine phyto-oestrogen metabolites and risk of type 2 diabetes in Chinese adults.

Gene-to-metabolite network for biosynthesis of lignans in MeJA-elicited Isatis indigotica hairy root cultures.

Root and leaf tissue of Isatis indigotica shows notable anti-viral efficacy, and are widely used as “Banlangen” and “Daqingye” in traditional Chinese medicine. The plants' pharmacological activity is attributed to phenylpropanoids, especially a group of lignan metabolites. However, the biosynthesis of lignans in I. indigotica remains opaque. This study describes the discovery and analysis of biosynthetic genes and AP2/ERF-type transcription factors involved in lignan biosynthesis in I. indigotica. MeJA treatment revealed differential expression of three genes involved in phenylpropanoid backbone biosynthesis (IiPAL, IiC4H, Ii4CL), five genes involved in lignan biosynthesis (IiCAD, IiC3H, IiCCR, IiDIR, and IiPLR), and 112 putative AP2/ERF transcription factors. In addition, four intermediates of lariciresinol biosynthesis were found to be induced. Based on these results, a canonical correlation analysis using Pearson's correlation coefficient was performed to construct gene-to-metabolite networks and identify putative key genes and rate-limiting reactions in lignan biosynthesis. Over-expression of IiC3H, identified as a key pathway gene, was used for metabolic engineering of I. indigotica hairy roots, and resulted in an increase in lariciresinol production. These findings illustrate the utility of canonical correlation analysis for the discovery and metabolic engineering of key metabolic genes in plants.

A Prospective Investigation of the Association Between Urinary Excretion of Dietary Lignan Metabolites and Weight Change in US Women.

Results from animal studies have consistently suggested that lignans play a role in the regulation of in body weight, but evidence from human studies has been limited. We examined the associations between urinary excretion of enterolactone and enterodiol, the major intestinal microbial metabolites of dietary lignans, and 10-year prospective weight change using data from 2 well-characterized cohort studies of US women: the Nurses' Health Study (2000-2010) and Nurses' Health Study II (1997-2007). Urinary excretion levels of enterolactone and enterodiol were measured at baseline. Associations with prospective weight change were analyzed using a multivariable-adjusted linear mixed-effects model. We observed that women in the highest quartile of urinary excretion of total lignans had significantly lower baseline body mass indices (weight in kilograms divided by height in meters squared) (mean, 24.6, 95% confidence interval (CI): 23.9, 25.2) than did those in the lowest quartile (mean, 27.7, 95% CI: 27.0, 28.4; P for trend < 0.01). Compared with women in the lowest quartile of enterodiol excretion, those in the highest quartile gained 0.27 kg/year less weight (95% CI: 0.12, 0.41; P for trend < 0.01) during the 10-year follow-up. The association was borderline significant for enterolactone (for the fourth vs. first quartile, least square mean of weight change rate = -0.14 kg/year, 95% CI: -0.29, 0.00). Our data suggest that higher urinary excretion of lignan metabolites, especially enterodiol, is associated with modestly slower weight gain.

Abstract 5542: Flaxseed and its lignan component protect from asbestos-induced inflammation in mice

Abstract Introduction/Background: Malignant mesothelioma (MM) is a highly lethal form of thoracic cancer with high mortality and poor treatment options. Development of MM has been linked directly to exposure to asbestos fibers, but with a long latency period. Recent studies have indicated that the pathogenesis of asbestos-induced cancers is due, in part, to chronic inflammation and oxidative tissue damage caused by persistent asbestos fibers. Whole grain flaxseed (FS) has known antioxidant, anti-inflammatory and cancer chemopreventive properties. Rationale: As a prelude to future chemoprevention studies, we tested the ability of oral FS and its lignan component, FLC which is enriched in the lignan secoisolariciresinol diglucoside (SDG), to prevent acute asbestos-induced inflammation and inflammatory cytokine release in MM-prone Nf2+/mut;Cdkn2a+/mut mice. Methods: Mice were given a single ip bolus of 400 mM of crocidolite asbestos. They were then placed on 10% FS or 10% FLC supplemented diets 1 day prior (-1) to or after (+1) asbestos instillation. All mice were evaluated 3 days after injection of asbestos for abdominal inflammation and proinflammatory cytokine release. The Nf2+/mut;Cdkn2a+/mut model was selected as it develops an accelerated form of MM when exposed to asbestos. Results: Using liquid chromatography and tandem mass spectrometry (LC/MS/MS), we showed that systemic levels (plasma) of flaxseed lignan metabolites (such as the mammalian lignans Enterolactone (EL) and Enterodiol (ED)) were comparable to those in other mouse models where FS was shown to be an effective chemopreventive agent. The numbers of macrophages (MF) and neutrophils (PMN) in peritoneal lavage fluid indicated that both FS and FLC blunted acute abdominal inflammation induced by asbestos. In addition, the levels of pro-inflammatory cytokines TNF alpha and IL-1 beta in lavage fluid were also decreased by the dietary agents. Conclusions: Our findings suggest that the known chemopreventive properties of FS and its lignan component appear to reduce short-term asbestos-induced inflammation and may thus prove to be a promising dietary agent in the chemoprevention of MM. Funding: This study was supported in part by Penn's SRP Center Grant (P42 ES027320), “Asbestos fate, exposure, remediation and adverse health effects” from the National Institute of Environmental Health (NIEHS) and 1R03CA180548-02 (MCS). Citation Format: Steven M. Albelda, Ralph A. Pietrofesa, Craig W. Menges, Joseph R. Testa, Melpo Christofidou-Solomidou. Flaxseed and its lignan component protect from asbestos-induced inflammation in mice. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5542. doi:10.1158/1538-7445.AM2015-5542

Age-dependency in the metabolism of flaxseed lignans by healthy adults

Flaxseed lignan metabolites, enterodiol (END) and enterolactone (ENL), have biological activities that may have potential therapeutic benefit through their antioxidative and oestrogenic properties. Circulating enterolignan concentrations are influenced by gut bacteria, which itself is subject to change with age. The plasma availability of END, ENL and total enterolignans (END + ENL) after 0 and 4-weeks of dietary milled flaxseed consumption (30 g/d) in healthy, younger (18–29 years) and older (45–69 years) adults was determined by using gas chromatography/mass spectrometry. Plasma total enterolignan concentrations increased in both younger and older adults, however, the distribution of END:ENL was lower in older adults (0.74) compared to younger adults (0.96). Post flaxseed treatment, ENL was detected in the plasma of 100% of the older adults, but only in 78% of the younger ones. The potential therapeutic benefit offered by enterolignans may be particularly important for ageing populations that are prone to chronic diseases.

Flaxseed reduces the pro-carcinogenic micro-environment in the ovaries of normal hens by altering the PG and oestrogen pathways in a dose-dependent manner.

The objective of the present study was to find the optimum dose of flaxseed that would decrease PG and alter oestrogen pathway endpoints implicated in ovarian cancer. In the study, four groups of fifty 1.5-year-old chickens were fed different amounts of flaxseed (0, 5, 10 or 15% of their total diet) for 4 months and were then killed to collect blood and tissues. Levels of flaxseed lignan metabolites, Enterolactone (EL) and Enterodiol (ED) were measured in the serum, liver and ovaries by liquid chromatography-MS/MS, and n-3 and n-6 fatty acid (FA) levels were measured by GC. The effects of the varied flaxseed doses were assessed by measuring levels of PGE2 and oestrogen metabolites (16-hydroxyestrone (16-OHE1) and 2-hydroxyestrone (2-OHE1)) as well as by analysing the expression of the oestradiol metabolising enzymes CYP3A4 (cytochrome p450, family 3, subfamily A, polypeptide 4), CYP1B1 (cytochrome p450, family 1, subfamily B, polypeptide 1) and CYP1A1 (cytochrome p450, family 1, subfamily A, polypeptide 1) and that of oestrogen receptor α (ERα) in the ovaries. The ratio of n-3:n-FA increased with an increase in flaxseed supplementation and corresponded to a dose-dependent decrease in cyclo-oxygenase-2 protein and PGE2 levels. EL and ED increased in the serum, liver and ovaries with increased concentrations of flaxseed. Flaxseed decreased the expression of ERα in the ovaries. The ratio of 2-OHE1:16-OHE1 in the serum increased significantly in the 15% flaxseed diet, and there was a corresponding increase in CYP1A1 in the liver and decrease in CYP3A4 in the ovaries. CYP1B1 mRNA also decreased with flaxseed diet in the ovaries. The 15% flaxseed-supplemented diet significantly decreased inflammatory PGE2, ERα, CYP3A4, CYP1B1 and 16-OHE1, but it increased CYP1A1 and 2-OHE1, which thus reduced the inflammatory and pro-carcinogenic micro-environment of the ovaries.

Bacterial metabolites of diet-derived lignans and isoflavones inversely associate with asthma and wheezing

Lignans and isoflavones are plant-derived chemicals with potent anti-inflammatory and antioxidant effects.1 Lignans are ubiquitous in Western diets, with flaxseeds containing the greatest amounts, while isoflavones are most abundant in soybeans (see Table E1 in this article's Online Repository at www.jacionline.org). The end product(s) of gut microbial metabolism of lignans is enterolactone2; metabolism of the isoflavone daidzein gives rise to equol and O-desmethylangolensin (O-DMA). Human studies demonstrate inverse associations between lignans and isoflavones and hormone-dependent cancers, cardiovascular disease, and osteoporosis,1 and improvements in asthma control are reported in those with high soy consumption.

Gut Microbiota Metabolites of Dietary Lignans and Risk of Type 2 Diabetes: A Prospective Investigation in Two Cohorts of U.S. Women

OBJECTIVETo examine urinary levels of enterolactone and enterodiol, intestinal microbial metabolites of dietary lignans, in relation to type 2 diabetes (T2D) risk.RESEARCH DESIGN AND METHODSUrinary concentrations of the lignan metabolites were assayed by liquid chromatography-mass spectrometry among 1,107 T2D and 1,107 control subjects in a nested case-control study conducted in participants from the Nurses’ Health Study (NHS) and NHSII. Subjects were free of diabetes, cardiovascular disease, and cancer at urine sample collection in 1995–2001. Incident self-reported T2D cases identified through 2008 were confirmed with a validated questionnaire.RESULTSIn both cohorts, T2D subjects had significantly lower concentrations of both enterolactone and enterodiol than control subjects. After multivariate adjustment for lifestyle and dietary risk factors of T2D, urinary concentrations of enterolactone were significantly associated with a lower risk of T2D (pooled odds ratio [OR] comparing the extreme quartiles 0.62 [95% CI 0.44, 0.88], P for trend = 0.003). Higher urinary concentrations of enterodiol were also marginally significantly associated with a lower T2D risk (pooled OR comparing extreme quartiles 0.67 [95% CI 0.48, 0.96], P for trend = 0.08). When concentrations of both metabolites were combined to reflect total lignan intake, the OR was 0.70 (95% CI 0.53, 0.92) for each SD increment of total lignan metabolites.CONCLUSIONSThese results indicate that lignan metabolites, especially enterolactone, are associated with a lower risk of T2D in U.S. women. Further studies are needed to replicate these findings and to explore potential mechanisms underlying the observed association.

Oilseeds ameliorate metabolic parameters in male mice, while contained lignans inhibit 3T3-L1 adipocyte differentiation in vitro

The focus was directed to the study of two of the most lignan-rich food sources: sesame and flaxseeds. Recent epidemiological and experimental evidences suggesting that these foods may improve metabolic functions underlying metabolic syndrome (MetS). To characterize the effect of these oilseeds on metabolic functions, we conducted an experimental study aimed at preventing adiposity and metabolic imbalance in a mouse model of high-fat diet (HFD)-induced MetS. Statistical analysis was performed by two-way analysis of variance test followed by post hoc Bonferroni analysis. We studied the effect of the oilseeds sesame and flaxseed on metabolic parameters in mice on a HFD. When the HFD was integrated with 20% of sesame or flaxseed flours, the mice showed a decrease in body fat, already at day 15, from time 0. The size of the adipocytes was smaller in epididymal fat, liver steatosis was inhibited, and insulin sensitivity was higher in mice on the supplemented diets. The supplemented diets also resulted in a significant increase in the serum levels of the lignan metabolites enterodiol and enterolactone compared with the controls. The expression of genes associated with the inflammatory response, glucose metabolism, adipose metabolism and nuclear receptor were altered by the oilseed-supplemented diets. Some of the most abundant lignans in these oilseeds were studied in 3T3-L1 preadipocyte cells and were effective in inhibiting adipocyte differentiation at the minimal dose of 1 nM. The consumption of sesame and flaxseed may be beneficial to decrease metabolic parameters that are generally altered in MetS.

Phyto-oestrogens and their metabolites in milk produced on two pastures with different botanical compositions

Phyto-oestrogens are a group of secondary plant metabolites that may bind to oestrogen receptors and exert oestrogenic or anti-oestrogenic effects in humans and can protect against cancer diseases. When ingested by dairy cows, phyto-oestrogens can be metabolised and transferred to the milk. The objective of this study was to assess the effects of grazing a recently established pasture containing red clover (Trifolium pratense L.) and an older pasture containing a variety of sown and unsown plant species on milk concentrations of phyto-oestrogens. Sixteen Norwegian Red dairy cows [mean (standard deviation); body weight 599 (45.1)kg, stage of lactation 73 (15.0) d in milk, milk yield 29.9 (2.90) kg/d at the start of the experiment] were divided into two groups and grazed either a short-term pasture (SP) or a long-term pasture (LP). The SP was representative of organically managed leys in Norway, which are frequently, approximately every third year, renewed by soil tillage and seeding, whereas LP was representative of organically managed grasslands that are less frequently renewed. The SP contained meadow fescue (Festuca pratensis Huds.) (mean 34%), timothy (Phleum pratense L.) (mean 19%), red clover (mean 28%), shepherd׳s-purse (Capsella bursa-pastoris (L.) Medik.) (mean 6%), pineappleweed (Matricaria matricarioides Porter ex Britton) (mean 5%) and scentless mayweed (Tripleurospermum perforatum (Mérat) Laínz) (mean 4%), and LP contained mainly white clover (Trifolium repens L.) (mean 21%), smooth meadowgrass (Poa pratensis L.) (mean 19%), timothy (mean 17%), meadow fescue (mean 15%), perennial ryegrass (Lolium perenne L.) (mean 6%), tufted hairgrass (Deschampsia cespitosa (L.) P. Beauv.) (mean 5%), northern dock (Rumex longifolius DC.) (mean 4%), common couch (Elytrigia repens (L.) Desv. Ex Nevski) (mean 4%), red clover (mean 3%) and dandelion (Taraxacum spp.) (mean 3%). In addition to a daily pasture allowance of 20kg dry matter per cow, supplements of 3.0kg barley (Hordeum vulgare L.) concentrate were fed. Herbage, concentrates and milk was sampled during the last week of three experimental periods and analysed for phyto-oestrogens using LC-MS/MS technology. Herbage from SP had 19 times higher concentration of isoflavones than herbage from LP, whereas only small differences were found for lignans. Milk produced on SP had 14 times higher concentrations of the mammalian isoflavonoid equol, and the concentrations of equol were higher than found in most other studies. This study confirms that grazing pastures containing red clover increases concentrations of isoflavones and especially equol in bovine milk compared to grazing pastures with other botanical composition. The higher milk concentrations of the lignan metabolite enterodiol in milk from SP compared to LP could not be related to differences in intake of the analysed lignans and may therefore be related to unidentified lignans.