Alkylresorcinols Research Articles

Whole-Grain Rye Bread Consumption by Women Correlates with Plasma Alkylresorcinols and Increases Their Concentration Compared with Low-Fiber Wheat Bread ,

Whole-grain rye and wheat products are rich in lignans, the precursors of enterolactone (ENL) and alkylresorcinols (ARs), which are phenolic lipids. In this crossover trial, we examined the effect of whole-grain rye bread compared with low-fiber wheat bread on plasma AR levels. Women (n = 39) aged 59 +/- 0.94 y (mean +/- SEM) were advised to consume rye (214 +/- 7.1 g/d) or wheat (178 +/- 6.5 g/d) bread (20% of total daily energy intake) in random order for 8 wk. The test periods were separated by an 8-wk washout period. ARs with different homologues and ENL were measured in plasma collected at the beginning (habitual diet) and end of the test bread periods. AR concentrations were higher (P < 0.001) after the rye bread (352 +/- 24.7 nmol/L) and habitual diet (97.7 +/- 12.1 and 88.3 +/- 8.7 nmol/L) periods than after the wheat bread period (36.6 +/- 4.2 nmol/L). Plasma AR concentrations were correlated with intake of rye bread (r = 0.34, P = 0.037), and with insoluble fiber from the whole diet during the rye (r = 0.39, P = 0.013) and wheat (r = 0.32, P = 0.047) bread periods. The plasma AR 17:0/21:0 ratio differed after the rye (0.84 +/- 0.04) and wheat (0.53 +/- 0.08) bread diet periods (P < 0.001). These data strongly suggest that plasma ARs could be used as specific biomarkers for whole-grain rye intake, and their homologue pattern could be used as an indicator of the bread type consumed.

Effects of climacostol on normal and tumoral mammalian cell lines

Climacostol, 1,3‐dihydroxy‐5‐[(Z)‐2′‐nonenyl]benzene, is a natural toxin contained in the extrusomal cortical granules of the heterotrich ciliate Climacostomum virens. It is used for chemical defence against predators such as the raptorial ciliate Dileptus margaritifer and its cytotoxic activity has been assessed on several species of ciliates such as Didinium nasutum, Paramecium caudatum, and Blepharisma japonicum (Miyake et al. 2003, Europ. J. Protistol., 39:25–36). On the basis of its chemical structure, climacostol may be classified into the large group of natural compounds known as resorcinolic lipids, that show antimicrobial, antiparasitic, and antitumoral activities (Kozubek et al. 2003, Cell Moll. Biol. Lett., 6:351–355). To explore the possibility to use climacostol in medical applications, we examined the effects of chemically synthesized climacostol (Masaki et al. 2004, Tetrahedron, 60:7041–7048) on the growth and proliferation of tumoral and normal mammalian cell lines: (1) human promyelocytic leukaemia cells, HL60; (2) human squamous carcinoma cells, A431; and (3) non‐tumoral cells derived from mice Leydig cells, TM3. It was observed that (1) a concentration of 10 μg/ml of climacostol exerts a strong cytotoxic activity on all cell lines used; (2) at lower concentrations of 10 ng/ml and 1 ng/ml, the effect of climacostol is limited to the inhibition of the cell growth; and (3) the normal TM3 cells are more resistant to climacostol than the two tumoral HL60 and A431cell lines. The dose‐dependent cytotoxic effects of climacostol encourage further investigation on the potential use of this ciliate toxin as an anti‐cancer chemical.

Whole-grain rye and wheat alkylresorcinols are incorporated into human erythrocyte membranes

Alkylresorcinols (AR), a group of phenolic lipids, exist in the human diet in whole-grain rye and wheat. They are absorbed by humans and have been quantified in plasma. In this 2-week study we assessed AR incorporation into human erythrocyte membranes. Nine subjects attended the study; four avoided whole-grain products for 1 week and then included whole-grain rye and wheat bread in the diet for the second week, four included whole-grain rye and wheat products in the diet during the whole follow-up and one followed a gluten-free diet. Plasma and erythrocyte membrane AR were analysed after weeks 1 and 2. Erythrocyte membrane AR concentrations increased an average of 231 nmol/l of packed erythrocytes (P=0.036) after consumption of whole-grain rye and wheat products. Plasma AR levels increased an average of 175 nmol/l (P=0.058). When intake of whole-grain products was constant, erythrocyte membrane and plasma AR levels remained stable. Long-chain AR were incorporated into erythrocyte membranes in a higher proportion compared to shorter-chain AR. This preliminary study shows that AR are incorporated into human erythrocyte membranes in vivo.

Alkylresorcinols as Markers of Whole Grain Wheat and Rye in Cereal Products

The alkylresorcinol (AR) content of Swedish wheat grain samples, as well as of cereal ingredients and cereal foods containing wheat and rye, was determined. The average total AR content in Swedish wheat was 412 microg/g (ranging between 227 and 639 microg/g), which is lower than that in Swedish rye analyzed in a previous study. The relative composition of AR homologues was consistent for wheat samples and differed markedly from that of rye. Notably, the ratio of the homologues C17:0/C21:0 was approximately 0.1 in wheat and approximately 1.0 in rye, indicating that it can be used to distinguish between those two cereals. The AR content in cereal foods commonly consumed in Sweden varied widely, from nondetectable levels in white wheat flour and products not containing the outer parts of wheat and/or rye to >900 microg/g in some whole grain rye products. AR content in cereal foods was calculated from their recipes using average AR values for the cereal ingredients determined in this study. As there was a good correlation between calculated and analyzed AR levels in cereal foods (R2 = 0.91), it is possible to estimate the proportion of whole grain wheat and/or rye in a given cereal product on the basis of AR content and C17:0/C21:0 ratio. ARs appear to be good markers of whole grain wheat and rye in foods, and their analysis may be an objective way to identify foods rich in whole grain wheat and/or rye or brans thereof.

1,3-dihydroxy-5-(tridec-4′,7′-dienyl)benzene: a new cytotoxic compound from Lithraea molleoides

A dichloromethane extract from the leaves of Lithraea molleoides (Anacardiaceae), an argentine medicinal plant, showed cytotoxicity on human hepatocellular carcinoma cell line. Bioassay guided fractionation of this extract led to the isolation of a new active 5-alkyl resorcinol: 1,3-dihydroxy-5-(tridec-4',7'-dienyl)benzene. Chemical structure was established based on spectroscopic data (UV, IR, MS, 1H-NMR, 13C-NMR, COSY). This compound presented cytotoxic activity on 3 human tumoral cell lines: hepatocellular carcinoma cell line-Hep G2 (IC50 +/- SD of 68 +/- 2 microM), mucoepidermoid pulmonary carcinoma cell line-H292 (IC50 +/- SD of 63 +/- 5 microM) and mammary gland adenocarcinoma cell line -MCF7 (IC50 +/- SD of 147 +/- 5).

The effect of merulinic acid on biomembranes

Merulinic acid (heptadecenylresorcinolic acid, resorcinolic acid) is one of the members of resorcinolic lipids, the natural amphiphilic long-chain homologues of orcinol (1,3-dihydroksy-5-methylbenzene). In the present study, membrane properties of merulinic acid were investigated. Merulinic acid exhibits strong haemolytic activity against sheep erythrocytes (EH 50 of 5±2 μM) regardless of the form of its application—direct injection into the erythrocyte suspension or injection as merulinic acid-enriched liposomes. The lysis of erythrocytes induced by merulinic acid was inhibited by the presence of divalent cations. The effectivity of the protection of erythrocytes was highest for Zn 2+ and weakest for Mn 2+. Merulinic acid at low concentrations also exhibits the ability for protection of cells against their lysis in hypoosmotic solutions. This protective effect is significant as, at 10 μM concentration of merulinic acid, the extent of osmotically induced cell lysis is reduced by approximately 40%. Merulinic acid induces increased permeability of liposomal vesicles. This effect was shown to be dependent on the composition of liposomal bilayer and it was stronger when lipid bilayer contained glycolipids (MGDG and DGDG) and sphingomyelin. Changes of TMA-DPH and NBD-PE fluorescence polarization show that the degree of merulinic acid incorporation into liposomal membrane is not very high. The polar “heads” of the molecules of investigated compounds are localized on the level of fatty acid's ester bonds in phospholipid molecules. Merulinic acid caused the increased fluorescence of the membrane potential fragile probe. This indicated an alteration of the surface charge and a decrease of the local pH at the membrane surface. This effect was visible in both low- and high-ionic strength environment. Merulinic acid causes also a decrease in activity of the membrane-bound enzyme acetylcholinesterase.

A study of cross‐reactions between mango contact allergens and urushiol

The allergens causing mango dermatitis have long been suspected to be alk(en)yl catechols and/or alk(en)yl resorcinols on the basis of observed cross-sensitivity reactions to mango in patients known to be sensitive to poison ivy and oak (Toxicodendron spp.). Earlier, we reported the 3 resorcinol derivatives: heptadecadienylresorcinol (I), heptadecenylresorcinol (II) and pentadecylresorcinol (III); collectively named 'mangol', as mango allergens. In this study, we extracted the 1st 2 components (I and II) from the Philippine mango, adjusted them to 0.05% concentration in petrolatum and patch tested the components on 2 subjects with mango dermatitis. Both subjects reacted to I. 1 subject also elicited a weaker positive reaction to II. To investigate the cross-reaction between mangol and urushiol, we also patch tested the same subjects with urushiol. The subject sensitive to II reacted to urushiol. 6 subjects with a history of lacquer contact dermatitis and positive reactions to urushiol were similarly patch tested. 5 persons reacted to I. 2 subjects also exhibited a slower but positive reaction to II. This is the 1st report in which heptadec(adi)enyl resorcinols known to be present in mango have been shown to elicit positive patch test reactions in mango-sensitive patients.

5-n-Alkylresorcinols from Grains of Winter Barley (Hordeum vulgare L.)

The resorcinolic lipid content and homologue composition of winter barley grains harvested at two field locations were evaluated. Depending on the crop location, the predominant alkylresorcinols identified were 1,3-dihydroxy-5-n-heneicosylbenzene or 1,3-dihydroxy-5-n-pentacosylbenzene. Both resorcinol concentration and their homologue profiles were diverse in samples harvested at different fields indicating a prevailing role of the environment upon the alkylresorcinol biosynthesis in cereals.

Chromatographic analysis of alkylresorcinols and their metabolites

Alkylresorcinols (AR) are phenolic lipids present in high amounts in the bran fraction of wheat and rye. AR are of scientific interest as bioactive compounds, as markers for the presence of wholegrain or bran fractions of wheat and rye in food products, and as possible biomarkers of intake for wholegrain wheat and rye. This review discusses their extraction from cereal grains, food and biological fluids, and the various chromatographic methods used in their analysis.

Alkyl- and alkenylresorcinols of wheat grains and their chemotaxonomic significance.

Resorcinolic lipid contents and homologue compositions in extracts isolated from soft winter, soft spring and hard (durum) wheat grains were evaluated by both instrumental and chromatography means. Resorcinol concentrations determined in wheat were diverse and varied in samples harvested within two consecutive vegetative years, whereas their homologue profiles were found to be rather invariable. The predominant alkylresorcinols identified in wheat grains were saturated 1,3-dihydroxy-5-n-heneicosylbenzene and 1,3-dihydroxy-5-n-nonadecylbenzene. 1,3-Dihydroxy-5-n-heptadecylbenzene and 1,3-dihydroxy-5-n-tricosylbenzene were also determined, whereas 1,3-dihydroxy-5-n-pentadecylbenzene and 1,3-dihydroxy-5-n-pentacosylbenzene were present in these extracts only in spurious amounts. Furthermore, our results show that alk(en)ylresorcinols may be useful as chemotaxonomic markers for a distinction between soft and hard wheat plants. Cluster analysis with Ward's amalgamation algorithm and five different distance linkage types clearly discriminated particular wheats into species- and cultivar-specific clusters, whereas the use of principal component analysis allowed us to specify, which of the variables analysed were decisive. This approach may be useful for both plant breeders and taxonomists to classify wheat species/cultivars.

Cereal Alkylresorcinols Elevate γ-Tocopherol Levels in Rats and Inhibit γ-Tocopherol Metabolism In Vitro

Alkylresorcinols (AR) are a class of amphiphilic phenolic lipids present in high amounts in wheat and rye bran. They have been reported to be both growth retarding and innocuous when fed to rats, and to have a broad range of bioactivities in vitro, suggested to be related to their ability to bind to proteins and modify membranes. This study was designed to test the effects of AR (purified from rye bran) on growth, tocopherol levels, and cholesterol levels in rats. Rats were fed 1 of 4 different levels of AR for 4 wk: 0 (control), 1, 2, and 4 g/kg diet. AR did not affect final body, liver, or lung weights. The AR diets increased the levels of gamma-tocopherol in liver and lungs (P < 0.05). To investigate whether AR could have increased gamma-tocopherol levels via inhibition of tocopherol-omega-hydroxylase, HepG2 cells were incubated with AR and the metabolism of gamma-tocopherol measured. AR significantly inhibited the conversion of gamma-tocopherol to its water-soluble hydroxychroman metabolite in vitro, indicating that AR may increase gamma-tocopherol levels via inhibition of tocopherol metabolism in vivo. The 4 g AR/kg diet decreased liver cholesterol (P < 0.001), but did not affect plasma lipids. AR were detected in the perirenal adipose tissue samples of rats fed AR, indicating that they can accumulate in the fatty tissues of rats. High levels of dietary AR moderately affect gamma-tocopherol, possibly via inhibition of tocopherol metabolism, and decrease liver cholesterol in rats.

Dietary Alkylresorcinols: Absorption, Bioactivities, and Possible Use as Biomarkers of Whole-grain Wheat- and Rye-rich Foods

The biologic and chemical properties of alkylresorcinols (ARs) have been reviewed previously, but there has been relatively little research or focus on the importance of ARs in food and diet. ARs represent a significant proportion of the phytochemicals present in wheat and rye, in which they normally exist in concentrations between 300 and 1500 microg/g. ARs are concentrated in the bran fraction of these cereals, and are therefore a significant component of food products rich in whole grain wheat and rye but not in products containing only refined cereal flour. In this review, we discuss the presence of ARs in food, methods of analysis, their absorption and role in the diet in light of their in vitro bioactivities, and their possible use as biomarkers of whole-grain wheat and rye intake.

Expedient Soxhlet extraction of resorcinolic lipids from wheat grains

The main objective of this study was to select the most suitable organic solvent for the extraction of alkylresorcinol fractions from wheat grains. Lipids were isolated in the Soxhlet apparatus. The solvents examined were acetone, chloroform, cyclohexane, diethyl ether, ethyl acetate and hexane. Each solvent yielded extracts that differed from each other in terms of resorcinolic lipid contents and homologue compositions as well as for the presence of contaminating ballast substances. The highest amounts of total resorcinolic lipids were found in acetone and ethyl acetate extracts, whereas as many as of 13 resorcinolic homologues were determined in the cyclohexane extract. Because of low contents of ballast substances and a complete absence of compounds cross-reacting with the diazonic salt Fast Blue B, the latter was suitable for direct qualitative and quantitative determinations of alkylresorcinol. The extraction using the Soxhlet apparatus and cyclohexane was shown to be faster and more versatile than previously published procedures.

Sterol Ferulates, Sterols, and 5-Alk(en)ylresorcinols from Wheat, Rye, and Corn Bran Oils and Their Inhibitory Effects on Epstein−Barr Virus Activation

Sterol ferulate, free sterol, and 5-alk(en)ylresorcinol constituents of wheat, rye, and corn bran oils were studied. Among the sterol ferulates, one novel compound, 24-methylenecholestanol ferulate (7), along with six known compounds, namely, 24-methylcholestanol ferulate (1), 24-methylcholesterol ferulate (2), 2-methyllathosterol ferulate (3), stigmastanol ferulate (4), sitosterol ferulate (5), and schottenol ferulate (6), were isolated and characterized. Five known free sterols, namely, 24-methylcholesterol (8), stigmastanol (9), sitosterol (10), schottenol (11), and stigmasterol (12), were isolated and identified. 5-Alk(en)ylresorcinols were found in wheat and rye bran oils but not in corn bran oil. Of these, one new compound, 5-n-(2'-oxo-14'-Z-heneicosenyl) resorcinol (19), and seven known compounds, namely, 5-n-heptadecyl- (13), 5-n-nonadecyl- (14), 5-n-heneicosyl- (15), 5-n-tricosyl- (16), 5-n-pentacosyl- (17), 5-n-(14'-Z-nonadecenyl)- (18), and 5-n-(2'-oxoheneicosyl)resorcinols (20), were isolated and characterized. These compounds were evaluated with respect to their inhibitory effects on the induction of Epstein-Barr virus early antigen (EBV-EA) by 12-O-tetradecanoylphorbol-13-acetate (TPA) in Raji cells, which is known to be a primary screening test for antitumor promoters. Four compounds, 1, 2, 4, and 11, showed potent inhibitory effects on EBV-EA induction.

Absorption of dietary alkylresorcinols in ileal-cannulated pigs and rats.

Alkylresorcinols (AR) are amphiphilic 1,3-dihydroxy-5-alkyl phenolic lipids. AR in food are only found in the outer layers of wheat and rye grains, and in whole grains are present at concentrations of 500-1000 microg/g. In wheat and rye, there are five main homologues, differing in the length of the odd-numbered alkyl chain (from seventeen to twenty-five C atoms long). Because AR may be bioactive, and might serve as biomarkers for these cereals, their absorption was investigated in model experiments with pigs and rats. Pigs with a cannula in the terminal ileum were fed four diets containing rye fractions with different levels of AR and the ileal effluents were analysed. The ileal recovery of AR was found to vary between 21 and 40 %, with no major difference between different chain-length homologues. The absorption of AR by rats was investigated by feeding (14)C-labelled heneicosylresorcinol (C21 : 0). Of the total activity, about 34 % was recovered in the urine, showing that the labelled AR was absorbed and metabolised by rats. AR were mostly cleared from rats by 60 h. It is concluded that AR are absorbed in the small intestine of single-stomached animals and excreted in metabolised form in the urine, and might contribute to the nutritional qualities of wholegrain wheat and rye diets.

Research Communication: Cereal Alkylresorcinols Are Absorbed by Humans

Currently there is no biomarker to link consumption of whole grain cereals and their observed health benefits. A candidate for a biomarker of whole grain wheat and rye intake is a class of phenolic lipids, the alkylresorcinols (AR). Studies to determine the uptake of AR in humans were carried out with a low fiber diet based on white wheat bread (AR free) and a high fiber diet based on rye bran-enriched bread (AR rich). For each diet, two meal frequencies were used: nibbling (7 small meals/d) and ordinary (3 large meals/d). Ten human ileostomy-operated subjects started with the AR-free diet for 2 wk, wk 1 on either nibbling or ordinary and wk 2 on the other meal frequency in a crossover design, followed by a 1-wk washout period, before the AR-rich diet performed as the AR-free diet. Food and ileostomy samples were analyzed for AR. Approximately 40% of AR were recovered in effluent from the small intestine, indicating that 60% of AR are taken up from or converted in the small intestine (ileal digestibility) with no difference between nibbling and ordinary meal frequencies. AR absorbed by humans may be of importance as bioactive compounds, or as a biomarker of whole grain wheat and rye intake.

Alkylresorcinols in cereals and cereal products.

The alkylresorcinol (AR) content of 8 commonly consumed cereals, 125 Triticum cultivars, milling fractions of wheat and rye, bread, and other cereal products was analyzed. ARs were found in wheat (489-1429 microgram/g), rye (720-761 microgram/g), triticale (439-647 microgram/g), and barley (42-51 microgram/g), but not in rice, oats, maize, sorghum, or millet. One durum wheat variety was found to have an exceptionally low level of ARs (54 microgram/g) compared to other durum wheat varieties (589-751 microgram/g) and Triticumspecies analyzed. The AR content of milling fractions closely followed the ash content and could be used as a marker of the presence of bran in flour. Using hot 1-propanol extraction, all ARs could be extracted from bread, contrary to previous studies which suggested that ARs were destroyed during baking. Cereal products varied greatly in AR content, with those containing wheat bran or whole rye having the highest content.

Rice seedling whole exudates and extracted alkylresorcinols induce stress-response in Escherichia coli biosensors.

A set of Escherichia coli sensor strains was used to evaluate the stress exerted on surrounding bacteria by germinating rice seed exudates. These biosensor strains contain Vibrio fischeri luxCDABE genes fused to the promoters of different genes involved in bacterial responses to environmental stresses. They provided clear evidence for a stress exerted by rice exudates, as shown by the induction of the universal stress protein gene uspA as well as genes of the heat shock regulon, grpE, lon and dnaK. The oxidative stress gene katG, and the post-transcriptional ompF regulator encoded by micF were also activated. The lack of derepression of recA, uvrA and alkA indicated that damage to the DNA was not induced in the E. coli strains tested. Interestingly, resorcinolic lipids extracted from rice root seedlings induced the same promoters as whole exudates, suggesting that these compounds may contribute to the stress exerted by seedling exudates. The results obtained with E. coli biosensors thus indicate that, in vivo, exudates may also exert a selective pressure on root-colonizing bacteria.

FT‐Raman Spectra of Unsoaked and NaOH‐Soaked Wheat Kernels, Bran, and Ferulic Acid

ABSTRACTThe sodium hydroxide (NaOH) test for determining wheat color class depends on the observation that on soaking in NaOH, red wheat turns a darker red and white wheat turns straw yellow. To understand the mechanism of this test, Raman spectra of wheat bran, wheat starch, ferulic acid, and whole kernels of wheat, before and after NaOH soak, were studied. The major observable components in the whole kernel were that of starch, protein, and ferulic acid, perhaps esterified to arabinoxylan and sterols. When kernels are soaked in NaOH, spectral bands due to ferulic acid shift to lower energy and show a slightly reduced intensity that is consistent with deprotonation of the phenolic group and extraction of a portion of the ferulic acid into solution. Other phenolic acids, alkyl resorcinols, and flavonoids observed in the NaOH extracts of wheat by HPLC were not observed in the Raman spectra. Wheat bran accounts for most of the ferulic acid in the whole kernel, as indicated by the increased intensity of the doublet at 1,631 and 1,600 cm‐1 in the bran. The intense starch band at 480 cm‐1 in whole kernel wheat was nearly absent in the wheat bran.

Process-induced changes on bioactive compounds in whole grain rye.

Manufacturing of healthy wholegrain foods demands knowledge of process-induced changes in macro-, micro- and non-nutrients. The high content of dietary fibre is a challenge in relation to good product texture and sensory quality. The stability and bioavailability of bioactive compounds have a marked influence on the health effects of cereal foods. It was confirmed that sterols, folates, tocopherols and tocotrienols, alkylresorcinols, lignans, phenolic acids and total phenolics are concentrated in the bran layers of the rye grain, and are only present at low levels in the flour endosperm. The levels of folate and easily-extractable phenolic compounds increase in germination and sourdough baking, but there are negligible changes in the levels of sterols, lignans and alk(en)ylresorcinols. The levels of tocopherols and tocotrienols are reduced during the sourdough fermentation. In conclusion, many of the bioactive compounds in wholegrain rye are stable during food processing, and their levels can even be increased with suitable processing.