Plant Lignans Secoisolariciresinol Research Articles

FLAXSEED MEAL FEEDING TO DAIRY COWS AS A STRATEGY TO IMPROVE MILK ENTEROLACTONE CONCENTRATION: A LITERATURE REVIEW

ABSTRACT: Flaxseed (Linum usitatissimum) is the richest source of the plant lignan secoisolariciresinol diglucoside (SDG). In mammals, including bovine, SDG is converted to the mammalian lignans enterolactone (EL) and enterodiol (ED) by the action of gastrointestinal microbes. There is a great deal of interest in promoting increased intakes of lignans in humans’ diet due to the potential health benefits of mammalian lignans, especially in the prevention of cardiovascular diseases, hypercholesterolaemia, breast and prostate cancers, and osteoporosis. Consumption of milk and dairy products enriched in EL could be an excellent strategy to increase the intake of lignans by humans. This literature review will focus on presenting feeding strategies capable to improve milk enterolactone concentration. Research has demonstrated the potential of flaxseed meal (FM) feeding to dairy cows as a strategy to improve milk EL concentration, therefore enhancing milk nutraceutical proprieties. A considerable number of studies have demonstrated that feeding vegetable lignans-rich sources, such as FM, to dairy cows improves EL in milk. Additionally, it has been reported that changes in the carbohydrate profile of FM-based diets fed to dairy cows can alter the output of milk EL. The application of animal nutrition as a tool to increase nutraceutical properties of milk (i.e. increased EL concentration) is a valuable strategy for promoting the association of milk with humans’ health benefits and is of great interest in contemporary society.
 Keywords: nutraceutical proprieties, bioactive compounds, lignans, dairy cows, disease risk reduction

The flaxseed lignan secoisolariciresinol diglucoside decreases local inflammation, suppresses NF\u03baB signaling, and inhibits mammary tumor growth

PurposeExposure to the polyphenolic plant lignan secoisolariciresinol diglucoside (SDG) and its metabolite enterolactone (ENL) has been associated with reduced breast cancer progression, particularly for estrogen receptor alpha (ERα)-negative disease, and decreased preclinical mammary tumor growth. However, while preclinical studies have established that SDG and ENL affect measures of progression in models of triple-negative breast cancer (TNBC, a subset of ERα-negative disease), the molecular mechanisms underlying these effects remain unclear.MethodsC57BL/6 mice were fed a control diet (control, 10% kcal from fat) or control diet + SDG (SDG, 100 mg/kg diet) for 8 weeks, then orthotopically injected with syngeneic E0771 mammary tumor cells (a model of TNBC); tumor growth was monitored for 3 weeks. The role of reduced NF-κB signaling in SDG’s anti-tumor effects was explored in vitro via treatment with the bioactive SDG metabolite ENL. In addition to the murine E0771 cells, the in vitro studies utilized MDA-MB-231 and MCF-7 cells, two human cell lines which model the triple-negative and luminal A breast cancer subtypes, respectively.ResultsSDG supplementation in the mice significantly reduced tumor volume and expression of phospho-p65 and NF-κB target genes (P < 0.05). Markers of macrophage infiltration were decreased in the distal-to-tumor mammary fat pad of mice supplemented with SDG relative to control mice (P < 0.05). In vitro, ENL treatment inhibited viability, survival, and NF-κB activity and target gene expression in E0771, MDA-MB-231, and MCF-7 cells (P < 0.05). Overexpression of Rela attenuated ENL’s inhibition of E0771 cell viability and survival.ConclusionsSDG reduces tumor growth in the E0771 model of TNBC, likely via a mechanism involving inhibition of NF-κB activity. SDG could serve as a practical and effective adjuvant treatment to reduce recurrence, but greater understanding of its effects is needed to inform the development of more targeted recommendations for its use.

Short communication: The mammalian lignan enterolactone is absorbed by newborn dairy calves fed enterolactone-enriched milk

Flaxseed is the richest source of the plant lignan secoisolariciresinol diglucoside, which is converted to the mammalian lignans enterolactone (EL) and enterodiol by the gut microbiota of ruminants and humans. Enterolactone has been associated with improved animal and human health due to its antioxidant and anticarcinogenic properties. The objective of this study was to determine the pharmacokinetics of EL in newborn dairy calves fed milk replacer or EL-enriched milk. We hypothesized that newborn Holstein calves fed EL-enriched milk would have greater area under the curve and plasma concentration of EL compared with those fed milk replacer. On d 5 of life, calves were administered 2 L of milk replacer (n = 10; low-EL treatment: 123 nmol/L of EL) or 2 L of EL-enriched milk (n = 10; high-EL treatment: 481 nmol/L of EL) during the morning feeding (0700 h). Blood samples were taken from the jugular vein before (0 h) and at 0.5, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 10, 12, 24, and 48 h after oral administration of treatments. The area under the curve for the plasma concentration of EL was analyzed according to the trapezoidal rule between 0 and 12 h after treatment administration, and it was greater in high- (26 nmol/L × h) than low-EL calves (4.30 nmol/L × h). Similarly, the maximum concentration of EL in plasma was greater in high- (5.06 nmol/L) versus low-EL calves (1.95 nmol/L). Furthermore, the time after treatment intake to reach maximum plasma concentration of EL was faster in high- (4.31 h) compared with low-EL (4.44 h) treatment. Calves were able to absorb EL, indicating that EL-enriched milk can potentially be used as source of EL to pre-weaned ruminants.

Abstract 231: The polyphenolic plant lignan secoisolariciresinol diglycoside reduces mammary tumor growth, possibly via inhibition of local inflammatory signaling

Abstract Background: Secoisolariciresinol diglycoside (SDG) is a polyphenolic plant lignan found in flax and sesame seeds as well as legumes, whole grains, fruits and vegetables. It is metabolized by the gut bacteria into two major enterolignans: enterolactone (ENL) and enterodiol (END). These enterolignans have been associated with reduced breast cancer risk and progression in population studies as well as decreased tumor growth in preclinical models of breast cancer. Methods: The impact of SDG supplementation on tumor growth in a mouse model of basal-like breast cancer was examined. C57BL/6 mice were fed a control diet (10% kcal from fat) or control diet with SDG supplementation (100 mg/kg food) for eight weeks, then both groups were orthotopically injected with E0771 mammary tumor cells. An inflammatory signaling qPCR array (Qiagen) was performed on mammary tissue distal to tumor. Tumors were stained by immunohistochemistry (IHC) for Ki67 to measure proliferation levels and phospho-p65 to determine inflammatory signaling pathway activation. Tumors and mammary tissue were also stained for F4/80 to quantify macrophage infiltration. Serum level of hormones, adipokines, and cytokines were measured via luminex assay (Bio-Rad). Results: SDG supplementation significantly decreased tumor weight (p&amp;lt;0.05). SDG did not affect body weight or body fat percentage but did significantly decrease expression of F4/80, CRP, and other pro-inflammatory markers in the mammary tissue. IHC staining revealed no difference in tumor proliferation; however, SDG supplementation did reduce inflammatory signaling in the tumors, indicated by a significant decrease in phospho-p65 staining. However, serum cytokine levels were not significantly different between the groups. Tumors are currently being stained by IHC for cleaved caspase 3 to measure levels of apoptosis. In addition, cell culture experiments will be conducted to define the impact of ENL treatment on protumorigenic cross-talk between tumor cells, adipocytes, and macrophages. Specifically, the effects of conditioned media from adipocyte/macrophage co-cultures (with +/- ENL treatment) on mouse E0771 and human MDA-MB-231 tumor cell proliferation, migration and invasion will be examined. Conclusions: SDG supplementation reduced mammary tumor growth in association with SDG’s effects on local, but not systemic, inflammatory signaling. Citation Format: Claire G. Lineberger, Laura W. Bowers, Nikki A. Ford, Emily L. Rossi, Bruce K. Kimler, Carol J. Fabian, Stephen D. Hursting. The polyphenolic plant lignan secoisolariciresinol diglycoside reduces mammary tumor growth, possibly via inhibition of local inflammatory signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 231. doi:10.1158/1538-7445.AM2017-231

Enterolactone Induces G1-phase Cell Cycle Arrest in Nonsmall Cell Lung Cancer Cells by Downregulating Cyclins and Cyclin-dependent Kinases

ABSTRACTFlaxseed is a rich source of the plant lignan secoisolariciresinol diglucoside (SDG), which is metabolized into mammalian lignans enterodiol (ED) and enterolactone (EL) in the digestive tract. The anticancer properties of these lignans have been demonstrated for various cancer types, but have not been studied for lung cancer. In this study, we investigated the anticancer effects of EL for several nonsmall cell lung cancer (NSCLC) cell lines of various genetic backgrounds. EL inhibited the growth of A549, H441, and H520 lung cancer cells in concentration- and time-dependent manners. The antiproliferative effects of EL for lung cancer cells were not due to enhanced cell death, but rather due to G1-phase cell cycle arrest. Molecular studies revealed that EL decreased mRNA or protein expression levels of the G1-phase promoters cyclin D1, cyclin E, cyclin-dependent kinases (CDK)-2, -4, and -6, and p-cdc25A; decreased phosphorylated retinoblastoma (p-pRb) protein levels; and simultaneously increased levels of p21WAF1/CIP1, a negative regulator of the G1 phase. The results suggest that EL inhibits the growth of NSCLC cell lines by downregulating G1-phase cyclins and CDKs, and upregulating p21WAF1/CIP1, which leads to G1-phase cell cycle arrest. Therefore, EL may hold promise as an adjuvant treatment for lung cancer therapy.

Plant lignan secoisolariciresinol suppresses pericardial edema caused by dioxin-like compounds in developing zebrafish: Implications for suppression of morphological abnormalities

Dioxins and dioxin-like compounds (DLCs) enter the body mainly through diet and cause various toxicological effects through activation of the aryl hydrocarbon receptor (AhR), a ligand activated transcription factor. Some plant extracts and phytochemicals are reported to suppress this transformation. However, most of these reports have been from in vitro experiments and few reports have been from in vivo experiments. In addition, there has been no report of foodstuffs that effectively prevent AhR-associated morphological abnormalities such as deformities caused by dioxins and DLCs in vivo. In this study, we show that secoisolariciresinol (SECO), a natural lignan-type polyphenolic phytochemical found mainly in flaxseed, has a rescuing effect, actually suppressing morphological abnormalities (pericardial edema) in zebrafish embryos exposed to 3,3′,4,4′,5-pentachlorobiphenyl (PCB126), a dioxin-like PCB congener. Importantly, the rescuing effect of SECO was still evident when it was applied 16 h after the beginning of exposure to PCB126. This study suggests that SECO may be useful as a natural suppressive agent for morphological abnormalities caused by dioxins and DLCs.

Bacterial transformation of dietary lignans in gnotobiotic rats

The bioactivity of lignans depends on their transformation by gut bacteria. The intestinal bacteria Clostridium saccharogumia, Eggerthella lenta, Blautia producta and Lactonifactor longoviformis convert the plant lignan secoisolariciresinol diglucoside via secoisolariciresinol (SECO) into the bioactive enterolignans enterodiol (ED) and enterolactone (EL). While the in vitro conversion of lignans by these bacteria has already been demonstrated, it is unclear whether this defined community is also capable of catalysing lignan transformation in vivo. We therefore associated germ-free rats with these four species. Germ-free rats served as a control. All animals were fed a diet containing 5% ground flaxseed. The caecal contents of rats associated with the four lignan-activating bacteria (ALB rats) contained SECO, ED and EL. The maximal EL formation rate from lignans in the pooled caecal contents of ALB rats was 7.52 nmol min(-1) g(-1) dry matter. The ALB rats excreted EL, but no SECO and ED, in their urine. The caecal contents of germ-free rats contained SECO, but no ED and EL. Their urine was devoid of lignans. Hence, the presence of enterolignans in the ALB rats, but not in the germ-free rats, demonstrates that this defined microbial community is capable of transforming plant lignans into EL in vivo.

Weekly excretion of the mammalian lignan enterolactone in milk of dairy cows fed flaxseed meal

Flaxseed meal (FM) is rich in the plant lignan secoisolariciresinol diglucoside (SDG) which is converted to the mammalian lignans enterodiol and enterolactone (EL) by ruminal microbiota. Feeding FM to dairy cows increases linearly EL concentration in milk but enterodiol is not detected. The objectives of the study were to determine the length of time to obtain peak EL concentration in the milk of dairy cows fed 20% FM and the length of time to return to EL baseline level in milk when cows are switched from high to low intake of flax SDG. A total of 12 multiparous lactating Holstein cows were assigned randomly to one of two feeding regimens: the control (CO) diet was fed for 6 weeks or the FM diet was fed from week 0 to 3 inclusive and then cows were switched to the control diet from week 3 to 6 inclusive. Milk samples were taken weekly for EL analysis. There was a significant interaction between feeding regimen and week for milk concentration of EL as a result of higher concentration of EL from week 1 to 3 for cows on the FM regimen compared with those on the CO regimen. Concentrations of milk EL on the FM regimen maintained uniform high levels from week 1 to 3 and they decreased significantly from week 3 to 4 when the CO diet was reintroduced in week 3. This study suggests that the conversion of SDG to the mammalian lignan EL and the transfer of EL to the mammary gland are well established after one week of feeding 20% FM in the diet of dairy cows and that milk concentration of EL returns to baseline level after one week of FM deprivation.

Daily Consumption for Six Weeks of a Lignan Complex Isolated from Flaxseed Does Not Affect Endothelial Function in Healthy Postmenopausal Women

The occurrence of menopause is associated with an increased risk of cardiovascular events, and this has partly been attributed to the decline in circulating levels of estrogen. A lignan complex rich in the plant lignan secoisolariciresinol diglucoside (SDG) was isolated from flaxseed. SDG is metabolized by the colonic microflora to the mammalian lignans enterodiol and enterolactone and is hypothesized to be cardioprotective due to their structural similarity to estrogen. The aim of this study was to investigate the effect of a lignan complex, providing 500 mg/d of SDG, on markers of endothelial function. Healthy postmenopausal women (n = 22) completed a randomized, double-blind, placebo-controlled, crossover study. Women consumed daily a low-fat muffin, with or without a lignan complex, for 6 wk, separated by a 6-wk washout period. Flow-mediated, endothelium-dependent vasodilatation (FMD) and nitroglycerine-mediated, endothelium-independent vasodilatation were measured at the end of each intervention period. The sum of Plasma nitrite and nitrate (NOx), endothelin-1 (ET-1), and asymmetric dimethylarginine (ADMA) were measured at the beginning and end of each intervention period. FMD was 3.6 ± 0.9% (mean ± SEM) after the lignan complex intervention period compared with 3.9 ± 0.7% after the placebo period (P = 0.72). Plasma concentrations of NOx, ET-1, and ADMA were not affected. We conclude that daily consumption for 6 wk of a low-fat muffin enriched with a lignan complex had no effect on endothelial function in healthy postmenopausal women.

Clostridium saccharogumia sp. nov. and Lactonifactor longoviformis gen. nov., sp. nov., two novel human faecal bacteria involved in the conversion of the dietary phytoestrogen secoisolariciresinol diglucoside

Two anaerobic bacteria involved in the conversion of the plant lignan secoisolariciresinol diglucoside were isolated from faeces of a healthy male adult. The first isolate, strain SDG-Mt85-3Db, was a mesophilic strictly anaerobic Gram-positive helically coiled rod. Based on 16S r RNA gene sequence analysis, its nearest relatives were Clostridium cocleatum (96.7% similarity) and Clostridium ramosum (96.6%). In contrast to these species, the isolate was devoid of alpha-galactosidase and -glucosidase and did not grow on maltose, melibiose, raffinose, rhamnose and trehalose. The hypothesis that strain SDG-Mt85-3Db represents a new bacterial species of the Clostridium cluster XVIII was confirmed by DNA–DNA hybridisation experiments. The G+C content of DNA of strain SDG-Mt85-3Db (30.7±0.8 mol%) was comparable with that of Clostridium butyricum, the type species of the genus Clostridium. The name Clostridium saccharogumia is proposed for strain SDG-Mt85-3Db (=DSM 17460 T=CCUG 51486 T). The second isolate, strain ED-Mt61/PYG-s6, was a mesophilic strictly anaerobic Gram-positive regular rod. Based on 16S rRNA gene sequence analysis, its nearest relatives were Clostridium amygdalinum (93.3%), Clostridium saccharolyticum (93.1%) and Ruminococcus productus (93.0%). The isolate differed from these species in its ability to dehydrogenate enterodiol. It also possessed alpha-arabinosidase and -galactosidase and had a higher G+C content of DNA (48.0 mol%). According to these findings, it is proposed to create a novel genus, Lactonifactor, and a novel species, Lactonifactor longoviformis, to accommodate strain ED-Mt61/PYG-s6. The type strain is DSM 17459 T (=CCUG 51487 T).

Phytoestrogens and Risk of Lung Cancer

To the Editor: In their study of phytoestrogens and risk of lung cancer, Dr Schabath and colleagues report on the intake of lignans in a US population. Their results, while interesting, are discrepant with those of other studies. In their article, the total intake of lignans was calculated as the sum of the plant lignans secoisolariciresinol and matairesinol and the mammalian enterolignans enterodiol and enterolactone. Enterolignans are not present in foods but can be determined via in vitro fermentation. Enterolignans are formed by bacteria in the colon at the expense of plant lignans. Summing plant lignans and enterolignans counts food contribution to the lignan intake twice. As a result, the estimates of total lignan intakes and the odds ratios based on these (in Tables 2, 4, and 6) may be biased. Apart from secoisolariciresinol and matairesinol, other plant lignans can also be converted to enterolignans. We determined the intake of secoisolariciresinol, matairesinol, lariciresinol, and pinoresinol in a Dutch population using a comprehensive food composition database and found a mean intake of 1240 μg/d. Although we included these additional plant lignans, which made up 75% of the total intake, this intake is still much lower than the 5400 μg/d (or approximately 5000 μg/d not including enterolignans) reported by Schabath et al. Regarding previously published intakes of secoisolariciresinol and matairesinol, daily intakes of 175 to 645 μg/d have been reported in a US population. Similar intakes were reported by us (303 μg/d) and by a Finnish group (434 μg/d). Boker et al reported higher intakes in a Dutch population (1110 μg/d), but they used category scores based on the highest values reported in the literature, which certainly leads to overestimation. In the study by Schabath et al, tea and coffee contributed 80% of the intake of secoisolariciresinol, whereas in the other studies their contribution never exceeded 44%. This high contribution suggests that the secoisolariciresinol contents of tea and coffee that were used are not correct. Pillow at al reported tea to contain 1090 μg/100 mL and coffee 520 μg/ 100 mL, whereas in our laboratory, contents obtained with liquid chromatography mass spectrometry were only 6 μg/ 100 mL for tea and 12 μg/100 mL for coffee. These discrepant lignan contents for coffee and tea may also have led to misclassification of participants and erroneous odds ratios.

A Lignan Complex Isolated from Flaxseed Does Not Affect Plasma Lipid Concentrations or Antioxidant Capacity in Healthy Postmenopausal Women ,

A lignan complex rich in the plant lignan secoisolariciresinol diglucoside (SDG) was isolated from flaxseed. SDG is metabolized by the colonic microflora to the mammalian lignans enterodiol (END) and enterolactone (ENL), and was hypothesized to reduce plasma lipid concentrations and improve antioxidant capacity. The aim of this study was to investigate the effects of a lignan complex, providing 500 mg/d of SDG, on serum concentration and urinary excretion of ENL, plasma lipids, serum lipoprotein oxidation resistance, and markers of antioxidant capacity. Healthy postmenopausal women (n = 22) completed a randomized, double-blind, placebo-controlled, crossover study. Women consumed daily a low-fat muffin, with or without a lignan complex, for 6 wk, separated by a 6-wk washout period. Serum ENL concentration, urinary ENL excretion, plasma concentrations of total cholesterol (TC), LDL cholesterol (LDL-C), HDL cholesterol (HDL-C), triacylglycerol (TAG), serum lipoprotein oxidation lag time, plasma Trolox-equivalent antioxidant capacity (TEAC), and ferric reducing ability of plasma (FRAP) were measured at the beginning and end of each intervention period. ENL concentrations in serum (P < 0.001) and ENL urinary excretion (P < 0.001) were significantly higher after the lignan complex intervention period compared with placebo. Plasma concentrations of TC, LDL-C, HDL-C, TAG, lipoprotein oxidation lag time, TEAC and FRAP were not affected. Daily consumption for 6 wk of a low-fat muffin enriched with a lignan complex significantly increased serum ENL concentrations and urinary ENL excretion in healthy postmenopausal women, but had no effect on plasma lipid concentrations, serum lipoprotein oxidation resistance, or plasma antioxidant capacity.

Intake of the Plant Lignans Secoisolariciresinol, Matairesinol, Lariciresinol, and Pinoresinol in Dutch Men and Women

Enterolignans (enterolactone and enterodiol) are phytoestrogens that are formed by the colonic microflora from plant lignans. They may reduce the risk of certain types of cancer and cardiovascular diseases. Initially, only secoisolariciresinol and matairesinol were considered to be enterolignan precursors, but recently, new precursors such as lariciresinol and pinoresinol were identified. We recently developed a lignan database including 4 major enterolignan precursors. We used this database to estimate lignan intake in a representative sample of Dutch men and women participating in the Dutch Food Consumption Survey, carried out in 1997-1998. Median total lignan intake among 4660 adults (19-97 y old) was 979 microg/d. Total lignan intake did not differ between men and women; thus, the lignan density of the diet was significantly higher (P < 0.001) in women than in men. Lignan intake was strongly skewed toward higher values (range 43-77584 microg/d, mean 1241 microg/d). Lariciresinol and pinoresinol contributed 75% to lignan intake, whereas secoisolariciresinol and matairesinol contributed only 25%. The major food sources of lignans were beverages (37%), vegetables (24%), nuts and seeds (14%), bread (9%), and fruits (7%). Lignan intake was significantly (P < 0.001) correlated with intake of dietary fiber (r = 0.46), folate (r = 0.39), and vitamin C (r = 0.44). Older persons, nonsmokers, vegetarians, and persons with a low BMI or a high socioeconomic status had higher lignan intakes than their counterparts. In brief, this study shows that the amount of enterolignan precursors in the diet has previously been largely underestimated.

Determination of lignans in human plasma by liquid chromatography with coulometric electrode array detection

The presence of mammalian lignans, mainly enterolactone, in human plasma has been related to lower incidence of certain cancers and cardiovascular disease. The plant lignans secoisolariciresinol and matairesinol have been reported to be precursors of mammalian lignans, but recently other plant lignans relatively abundant in the diet have also been identified as precursors. To evaluate the importance and contribution of these new dietary precursors to the mammalian lignan formation in vivo, metabolic studies in human subjects must be carried out. For this purpose a method based on high-performance liquid chromatography using coulometric electrode array detection for the simultaneous determination of nine plant lignans and two mammalian lignans in human plasma was developed, validated, and shown to fulfill the reliability criteria.

Optimization of a liquid chromatography-tandem mass spectrometry method for quantification of the plant lignans secoisolariciresinol, matairesinol, lariciresinol, and pinoresinol in foods.

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the quantification of the four major enterolignan precursors [secoisolariciresinol, matairesinol, lariciresinol, and pinoresinol] in foods. The method consists of alkaline methanolic extraction, followed by enzymatic hydrolysis using Helix pomatia (H. pomatia) beta-glucuronidase/sulfatase. H. pomatia was selected from several enzymes based on its ability to hydrolyze isolated lignan glucosides. After ether extraction samples were analyzed and quantified against secoisolariciresinol-d8 and matairesinol-d6. The method was optimized using model products: broccoli, bread, flaxseed, and tea. The yield of methanolic extraction increased up to 81%, when it was combined with alkaline hydrolysis. Detection limits were 4-10 microg/(100 g dry weight) for solid foods and 0.2-0.4 microg/(100 mL) for beverages. Within- and between-run coefficients of variation were 6-21 and 6-33%, respectively. Recovery of lignans added to model products was satisfactory (73-123%), except for matairesinol added to bread (51-55%).

Studies on the metabolism of the plant lignans secoisolariciresinol and matairesinol.

The plant lignans secoisolariciresinol and matairesinol occur in numerous foods such as oil seeds, whole grains, vegetables, and fruits. We have studied the hitherto unknown oxidative metabolism of secoisolariciresinol and matairesinol in hepatic microsomes from untreated and Aroclor 1254-induced Wistar rats and from humans. Five oxidative metabolites of secoisolariciresinol and 10 oxidative metabolites of matairesinol were detected in rat liver microsomes, and their chemical structures were elucidated. The pathways in the metabolism of both secoisolariciresinol and matairesinol included aliphatic and aromatic hydroxylation, whereas oxidative demethylation was only observed for matairesinol. Human hepatic microsomes were able to metabolize secoisolariciresinol whereas matairesinol was only poorly metabolized. This study clearly shows that secoisolariciresinol and matairesinol are substrates of cytochrome P450-mediated metabolism. However, from preliminary experiments with rats dosed orally with secoisolariciresinol and matairesinol, it appears that the intestinal absorption and subsequent oxidative metabolism of these plant lignans occur only to a very small extent due to the highly efficient conversion of secoisolariciresinol and matairesinol to the mammalian lignans enterodiol and enterolactone by the gut microflora.

Differences in the antioxidant activity of plant and mammalian lignans

The antioxidative activities of the plant lignans secoisolariciresinol and matairesinol and the mammalian lignans enterodiol and enterolactone were investigated using the FRAP assay (Ferric Reducing/Antioxidant Power assay). The plant lignans showed a clearly higher antioxidant activity than the known antioxidant ascorbic acid. The mammalian lignans were far less active in this assay. Using 2-methoxy- p-cresol and m-cresol as structural model compounds for plant and mammalian lignans, the additional aromatic methoxy group was shown to be responsible for the different antioxidant activities.

Structural determinants of plant lignans for the formation of enterolactone in vivo

The quantity of mammalian lignans enterolactone (ENL) and enterodiol (END) and of plant lignans secoisolariciresinol (SECO) and 7-hydroxymatairesinol (HMR) excreted in a 24-h rat urine sample was measured after a single p.o. dose of an equivalent quantity of secoisolariciresinol diglycoside (SDG), secoisolariciresinol (SECO), matairesinol (MR), 7-hydroxymatairesinol (HMR) and ENL. Plant lignans (SECO and HMR) were partially absorbed as such. The aglycone form of SECO was more efficiently converted into mammalian lignans END and ENL than the glycosylated form, SDG. Of plant lignans, MR produced the highest quantities of ENL: the quantity was over twofold compared with HMR or SDG. The majority of the animals, which had been given SECO, excreted higher quantities of END than ENL into urine, but ENL was the main lignan metabolite after SDG. The highest quantities of ENL in urine were measured after the administration of ENL as such. The (−)SECO isolated from Araucaria angustifolia was converted into (−)ENL only. The administration of (−)SDG, which was shown to produce (+)SECO, resulted in excretion of (+)ENL only and (−)HMR was converted into (−)ENL only. This confirmed that the absolute configurations at C8 and C8′ are not changed during the microbial metabolism. Whether the biological effects are enantiomer-specific, remains to be resolved.

Exposure to Flaxseed and its Purified Lignan Reduces Bone Strength in Young but Not Older Male Rats

Flaxseed is the richest source of the plant lignan secoisolariciresinol diglycoside (SDG), which is converted to the two major mammalian lignans, enterodiol (ED) and enterolactone (EL), by colonic bacteria. Because both ED and EL can produce biological effects similar to estrogen, exposure to lignans during early stages of development may adversely alter the normal development of bone in males since bone is a hormone-sensitive tissue. To determine whether early exposure to flaxseed or its lignan compromised the acquisition of bone mass or reduced bone strength, male offspring were exposed to one of three diets during lactation only (birth through postnatal day [PND] 21) via mother's milk or continuously from the start of lactation through to adolescence (PND 50) or young adulthood (PND 132). The diets were a basal diet (BD) that was devoid of phytoestrogens, BD containing 10% flaxseed, or BD containing the equivalent quantity of SDG present in a 10% flaxseed diet. To assess bone quantity, the bone mineral content (BMC) and bone mineral density (BMD) of femurs were assessed by dual-energy x-ray absorptiometry. Since the biomechanical properties of bone are indicators of the microarchitecture and thus bone quality, the biomechanical strength of femurs was assessed by three-point bending. At PND 50, ultimate bending stress and Young's modulus, measures of bone strength, were reduced among rats that received the 10% flaxseed diet from PND 0 through PND 50, while there were no marked differences in bone size, BMC, or BMD among groups. Interestingly, this effect does not appear to be due to the lignan in flaxseed, as continuous exposure to the diet containing the equivalent quantity of lignan (10 S diet) did not alter any measures of bone strength. In contrast to PND 50, bone strength did not differ among groups at PND 132, indicating that the compromise in bone strength was not sustained into early adulthood. Bone size, BMC, and BMD continued to be similar among treatment groups at PND 132. In conclusion, exposing male rats to a diet containing 10% flaxseed or an equivalent quantity of lignan either during lactation only or through to early adulthood is safe with respect to bone health, as measures of bone mass and strength were similar to control rats.

Method for Measurement of Dietary Secoisolariciresinol Using HPLC with Multichannel Electrochemical Detection

We describe a sensitive and specific assay for the determination of the plant lignan secoisolariciresinol, one of the main dietary precursors to the mammalian derived lignans, enterodiol and enterolactone. Quantification of secoisolariciresinol aglycone is achieved by reversephase high-performance liquid chromatography with multichannel electrochemical detection after hydrolysis of the glycoside moieties. This approach affords greater specificity than conventional ultraviolet detection and has a detection limit of 2.8 pmol. The method is ideally suited to the determination of secoisolariciresinol in processed flaxseed samples and can be used to assess the level of incorporation of flaxseed in fortified foods.