Occurrence and properties of steryl ferulates and glycosides in wheat and rye

Cereal kernels are known to contain a number of minor components that possess beneficial health attributes. In this thesis rye and wheat were studied as sources of steryl ferulates and steryl glycosides and their behaviour in processing were evaluated. Further, enzymatic hydrolysis of these conjugates was studied, as well as the capacity of steryl ferulates to inhibit lipid oxidation at different temperatures. Steryl ferulates were shown to have a strong positive correlation with dietary fibre contents in milling fractions from the outer parts of the kernels obtained from a commercial scale mill. Highest contents of steryl ferulates were found in the bran in both cereals, with the content decreasing once moving towards the inner parts of the kernel. Variation in the contents of steryl ferulates was higher in wheat fractions than rye fractions. Steryl glycosides, on the other hand, had either negative or no correlation with dietary fibre, and the range of the steryl glycoside contents was much narrower than that of steryl ferulates in both cereals. There were significant differences in the sterol compositions of these steryl conjugates when compared with each other or with the total plant sterols in the corresponding fractions. Properties of steryl ferulates and steryl glycosides were evaluated after common processing methods and in enzymatic hydrolysis. Thermal and mechanical processing had only minor or no effects on the contents of steryl conjugates from rye and wheat bran. Enzymatic treatments on the other hand caused some changes, especially in the contents of glycosylated sterols. When steryl ferulates extracted from rye or wheat bran were subjected to enzymatic treatments by steryl esterase, significant differences in the rates of hydrolysis were observed between steryl ferulates from different sources with differing sterol compositions. Further, differences were also observed between enzymes from different sources. Steryl glycosides were shown to be hydrolysed by β-glucosidase (cellobiase) from A. niger, but less with β-glucosidases from other sources. Steryl ferulates showed good antioxidant activity at both moderate and high temperatures. In bulk and emulsion systems of methyl linoleate at 40°C steryl ferulates extracted from rye and wheat bran inhibited hydroperoxide formation much more effectively than synthetic steryl ferulates or those extracted from rice (γ-oryzanol), demonstrating that the sterol composition has an effect on the activity. At cooking (100°C) and frying temperatures (180°C) sitostanyl ferulate was shown to inhibit polymer formation significantly and, especially at 100°C, comparably to α-tocopherol. The rate of antioxidant degradation was slower for sitostanyl ferulate, showing higher heat stability than α-tocopherol. When evaluated as a mixture, no synergistic effect was observed between these two antioxidants. The data presented in this thesis provides information that may henceforth be applied when evaluating the intakes of steryl conjugates from cereal sources, as well as their possible influences as minor bioactive components. Wheat and rye both are good sources of steryl ferulates and steryl glycosides and, especially with steryl ferulates, what may be lost out to some other cereals on quantity is compensated with quality of the sterol composition.Viljatuotteet sisältävät useita bioaktiivisia yhdisteitä, kuten kasvisterolit, fenoliset hapot, tokoferolit, folaatit ja alkyyliresorsinolit, joiden pitoisuudet viljoissa saattavat olla verrattain pieniä, mutta joilla tiedetään olevan positiivisia vaikutuksia ihmisten aineenvaihdunnassa. Näiden ravinnon pienkomponenttien tutkimus on viimeaikoina lisääntynyt merkittävästi, sillä niiden arvellaan tehostavan ravintokuidun positiivisia terveysvaikutuksia. Näitä "kuidun kanssamatkustajia" tiedetään löytyvän erityisesti täysjyväviljatuotteista sekä jyvän ulompia kuoriosia sisältävistä kuitupitoisista myllytysjakeista. Väitöskirjatutkimuksessa selvitettiin viljamateriaaleille tyypillisten steryyliferulaattien sekä steryyliglykosidien esiintymistä rukiissa ja vehnässä. Steryyliferulaatit ovat kasvisterolien ja fenolisen ferulahapon muodostamia estereitä ja steryyliglykosidit puolestaan kasvisterolien hiilihydraattijohdoksia. Pitoisuusmääritysten lisäksi tutkittiin erilaisten prosessointimenetelmien mahdollista vaikutusta sterolijohdosten pitoisuuksiin ja saatavuuteen leseistä, sekä sterolijohdosten entsymaattista pilkkoutumista. Lisäksi analysoitiin steryyliferulaattien hapettumisenestokykyä erilaisissa lämpötiloissa ja mallisysteemeissä. Sekä rukiin että vehnän steryyliferulaattipitoisuus korreloi vahvasti myllytysnäytteiden ravintokuitupitoisuuden kanssa, eli jyvien uloimmista kerroksista olevissa näytteissä, joissa kuitupitoisuus oli suuri, myös steryyliferulaattien pitoisuus oli suuri. Hajonta erilaisten myllytysnäytteiden steryyliferulaattipitoisuuksissa oli huomattavasti isompi erilaisten vehnä- kuin ruisnäytteiden välillä. Steryyliglykosidien pitoisuuksissa ei havaittu vastaavaa positiivista korrelaatiota kuitupitoisuuden kanssa ja pitoisuuksien vaihtelu eri näytteiden välillä oli huomattavasti pienempi kuin steryyliferulaattipitoisuuksien hajonta. Steryyliferulaattien ja steryyliglykosidien sterolikoostumukset poikkesivat sekä toisistaan että kokonaissterolikoostumuksesta merkittävästi. Prosessoinnin vaikutuksia steryyliferulaattien ja -glykosidien pitoisuuksiin tutkittiin erilaisten mekaanisten, lämpö- ja entsymaattisten käsittelyiden jälkeen. Tutkituista käsittelymenetelmistä mekaaniset ja lämpökäsittelyt eivät juurikaan muuttaneet sterolijohdosten pitoisuuksia ruis- ja vehnäleseissä. Sen sijaan entsymaattiset käsittelyt vaikuttivat erityisesti steryyliglykosidien pitoisuuksiin sekä yleisesti ottaen sterolien saatavuuteen leseistä. Prosessoinnin vaikutusten lisäksi leseestä eristettyjen steryyliferulaattien ja steryyliglykosidien entsymaattista pilkkoutumista tutkittiin mallikokeissa. Steryyliesteraasi-entsyymi pilkkoi rukiin ja vehnän steryyliferulaatteja tehokkaammin kuin riisistä eristettyjä steryyliferulaatteja, joiden sterolikoostumus on erilainen. Steryyliglykosidit hydrolysoituivat mikrobiperäisellä β-glukosidaasilla, mutta esimerkiksi mantelin β-glukosidaasi ei tutkimuksen koeolosuhteissa pilkkonut steryyliglykosideja. Hydrolysoitumisominaisuuksien lisäksi rukiin ja vehnän steryyliferulaatit osoittautuivat riisin steryyliferulaatteja tehokkaammiksi myös hapettumisen estäjinä eli antioksidantteina. Antioksidanttitehokkuutta arvioitiin erilaisissa rasvamalleissa (bulkkirasva ja emulsiot) ja lämpötiloissa (ml. keitto- ja paistolämpötilat). Steryyliferulaattien antioksidanttiteho oli alhaisempi kuin hyvin tunnetun antioksidantin, α-tokoferolin. Toisaalta steryyliferulaattien tuhoutuminen mallisysteemissä oli α-tokoferolin tuhoutumista hitaampaa. Steryyliferulaatin ja α-tokoferolin hapettumisenestokyvyssä ei havaittu synergististä yhteisvaikutusta. Väitöskirjatutkimuksessa saatuja tuloksia voidaan hyödyntää arvioitaessa steryyliferulaattien ja -glykosidien saantia ruis- ja vehnätuotteista, sekä arvioitaessa niiden ominaisuuksia ravinnon hyödyllisinä bioaktiivisina yhdisteinä. Ruis ja vehnä ovat erittäin hyviä steryyliferulaattien ja -glykosidien lähteitä ja erityisesti steryyliferulaattien kohdalla niiden optimaalinen sterolikoostumus antaa lisäarvoa verrattaessa eri viljalajeja steryyliferulaattien lähteenä

Effect of extrusion processing parameters on structure, texture and dietary fibre composition of directly expanded wholegrain oat-based matrices

Oat flour mixed with 30 g/100 g rice flour was extruded with a twin-screw extruder using a central composite orthogonal design. Temperatures (120 °C,140 °C, 160 °C) and moisture (14.5 g/100 g, 17.7 g/100 g, 20.6 g/100 g) were adjusted during extrusion, while screw speed was kept constant (400 rpm). Extrudates were analysed for structure (expansion, density, microstructure), texture (hardness), β-glucan (molecular weight and extractability), as well as fibre content. Expansion varied between 250 and 329%, density between 165 and 457 kg/m3 and hardness between 27 and 64 N. The response surface model showed that more expanded, less dense and less hard extrudates were achieved at low moisture, while high temperature resulted in lower density and hardness. Significant differences in β-glucan extractability were observed depending on extrusion conditions, with values ranging between 0.64 and 1.31 g/100 g. β-glucan extractability correlated with positively with porosity, and negatively with moisture content during extrusion, cell wall thickness and density. The results indicate that conditions that produce a more porous, crispier structure, also increases β-glucan extractability.ISSN:0460-1173ISSN:0023-6438ISSN:1096-112

Extraction of Pectin from Orange Peel Wastes as an Ingredient for Edible Films Containing Kabog Millet Flour

Fossil-based plastic is a popular material for food packaging. It can cause negative environmental consequences due to its low biodegradability. To address this challenge, a pectin-based edible plastic with added nutritional value by incorporating whole-grain kabog millet flour was prepared. The pectin in the films was extracted by microwave-assisted and enzymatic procedures from orange peel wastes. The extracted pectin was tested for its degree of esterification using Fourier-transform infrared spectroscopy, its molecular weight and behavior in aqueous solutions using size-exclusion chromatography, and its monosaccharide composition using ion-exchange chromatography. Biodegradable and edible pectin films were produced and tested for their mechanical properties: maximum strain, maximum stress, and water contact angle. The results showed a significant increase in hydrophobicity of the film surface by adding whole-grain kabog millet flour. The maximum strain of the film, however, was reduced to around 80% upon the addition of the whole-grain kabog millet flour. Enzymatically-extracted pectin increased the film hydrophobicity. Hydrophobic surfaces have higher water resistance; thus, the enzymatically-extracted pectin can be developed for further applications. Due to the low elasticity of the films, a possible application would be as direct coating of fruits and vegetables incorporating antioxidants or antimicrobials

UPLC-MS/MS Based Identification of Dietary Steryl Glucosides by Investigation of Corresponding Free Sterols

Dietary plant foods are characterized by a vast molecular diversity of glycosylated sterols (SG) that differ in the structure of the steryl backbone. The identification of these polar steryl conjugates represents a major challenge as they are structurally highly similar, and commercial standards are limited to a few naturally abundant species. Spectral databases do not yet contain MS/MS spectra of these sterol conjugates obtained by electrospray ionization (ESI), which would facilitate their reliable identification. Thus, this study aimed at providing novel information on ESI-MS/MS spectra of both abundant and minor SG found in foods. As a first step, however, free sterols (FS) were investigated for their fragmentation behavior as they share the same intermediate ion as SG. Pure SG were obtained from commercially available standard mixtures and minor SG were extracted from different food sources (oat bran, wheat bran, pumpkin seeds, melon, rapeseeds, and potato peel). ESI-MS/MS spectra of 15 FS were assessed and fragment ions reflective of structural features were identified and rationalized. Subsequently, 14 SG were identified at four different levels, while relative retention times from chromatographic separation and spectral features of FS served to identify five SG. Spectral data from FS were directly transferable to SG when analyzed as aglycone ions as shown by similarity scores while SG were characterized by shorter retention times in reverse phase chromatography and the additional analysis as sodiated adduct confirmed their glycosidic nature. Moreover, we report for the first time the occurrence of 24-methylenecholesterol and a 4-monomethyl sterol as glycosidic conjugates in higher plants. The presented data will serve as a valuable tool for SG profiling of foods by facilitating their identification

Western diet enhances intestinal tumorigenesis in Min/ plus mice, associating with mucosal metabolic and inflammatory stress and loss of Apc heterozygosity

Western-type diet (WD) is a risk factor for colorectal cancer, but the underlying mechanisms are poorly understood. We investigated the interaction of WD and heterozygous mutation in the Apc gene on adenoma formation and metabolic and immunological changes in the histologically normal intestinal mucosa of Apc(Min/+) (Min/ +) mice. The diet used was high in saturated fat and low in calcium, vitamin D, fiber and folate. The number of adenomas was twofold higher in the WD mice compared to controls, but adenoma size, proliferation or apoptosis did not differ. The ratio of the MM to wild-type allele was higher in the WD mice, indicating accelerated loss of Apc heterozygosity (LOH). Densities of intraepithelial CD3 epsilon(+) T lymphocytes and of mucosal FoxP3(+) regulatory T cells were higher in the WD mice, implying inflammatory changes. Western blot analyses from the mucosa of the WD mice showed suppressed activation of the ERK and AKT pathways and a tendency for reduced activation of the mTOR pathway as measured in phosphoS6/S6 levels. The expression of pyruvate dehydrogenase kinase 4 was up-regulated in both mRNA and protein levels. Gene expression analyses showed changes in oxidation/reduction, fatty acid and monosaccharide metabolic pathways, tissue organization, cell fate and regulation of apoptosis. Together, our results suggest that the high-risk Western diet primes the intestine to tumorigenesis through synergistic effects in energy metabolism, inflammation and oxidative stress, which culminate in the acceleration of LOH of the Apc gene. (C) 2016 Elsevier Inc. All rights reserved.Peer reviewe

DNA methylation changes and somatic mutations as tumorigenic events in Lynch syndrome-associated adenomas retaining mismatch repair protein expression

Background: DNA mismatch repair (MMR) defects are a major factor in colorectal tumorigenesis in Lynch syndrome (LS) and 15% of sporadic cases. Some adenomas from carriers of inherited MMR gene mutations have intact MMR protein expression implying other mechanisms accelerating tumorigenesis. We determined roles of DNA methylation changes and somatic mutations in cancer-associated genes as tumorigenic events in LS-associated colorectal adenomas with intact MMR. Methods: We investigated 122 archival colorectal specimens of normal mucosae, adenomas and carcinomas from 57 LS patients. MMR-deficient (MMR-D, n 49) and MMR-proficient (MMR-P, n 18) adenomas were of particular interest and were interrogated by methylation-specific multiplex ligation-dependent probe amplification and Ion Torrent sequencing. Findings: Promoter methylation of CpG island methylator phenotype (CIMP)-associated marker genes and selected colorectal cancer (CRC)-associated tumor suppressor genes (TSGs) increased and LINE-1 methylation decreased from normal mucosa to MMR-P adenomas to MMR-D adenomas. Methylation differences were statistically significant when either adenoma group was compared with normal mucosa, but not between MMR-P and MMR-D adenomas. Significantly increased methylation was found in multiple CIMP marker genes (1612, NEUROGI,CRABP1, and CDKN2A) and TSGs (SERPI and SFRP2) in MMR-P adenomas already. Furthermore, certain CRC-associated somatic mutations, such as KRAS, were prevalent in MMR-P adenomas. Interpretation: We conclude that DNA methylation changes and somatic mutations of cancer-associated genes might serve as an alternative pathway accelerating LS-associated tumorigenesis in the presence of proficient MMR. Fund: Jane and Aatos Erkko Foundation, Academy of Finland, Cancer Foundation Finland, Sigrid juselius Foundation, and HiL1FE. (C) 2019 Published by Elsevier B.V.Peer reviewe

Re-structuring of marine communities exposed to environmental change

Species richness is the most commonly used but controversial biodiversity metric in studies on aspects of community stability such as structural composition or productivity. The apparent ambiguity of theoretical and experimental findings may in part be due to experimental shortcomings and/or heterogeneity of scales and methods in earlier studies. This has led to an urgent call for improved and more realistic experiments. In a series of experiments replicated at a global scale we translocated several hundred marine hard bottom communities to new environments simulating a rapid but moderate environmental change. Subsequently, we measured their rate of compositional change (re-structuring) which in the great majority of cases represented a compositional convergence towards local communities. Re-structuring is driven by mortality of community components (original species) and establishment of new species in the changed environmental context. The rate of this re-structuring was then related to various system properties. We show that availability of free substratum relates negatively while taxon richness relates positively to structural persistence (i.e., no or slow re-structuring). Thus, when faced with environmental change, taxon-rich communities retain their original composition longer than taxon-poor communities. The effect of taxon richness, however, interacts with another aspect of diversity, functional richness. Indeed, taxon richness relates positively to persistence in functionally depauperate communities, but not in functionally diverse communities. The interaction between taxonomic and functional diversity with regard to the behaviour of communities exposed to environmental stress may help understand some of the seemingly contrasting findings of past research

DNA hypermethylation appears early and shows increased frequency with dysplasia in Lynch syndrome-associated colorectal adenomas and carcinomas

Abstract Background Lynch syndrome (LS) is associated with germline mutations in DNA mismatch repair (MMR) genes. The first “hit” to inactivate one allele of the predisposing MMR gene is present in every cell, contributing to accelerated tumorigenesis. Less information is available of the nature, timing, and order of other molecular “hits” required for tumor development. To this end, MMR protein expression and coordinated promoter methylation were examined in colorectal specimens prospectively collected from LS mutation carriers (n = 55) during colonoscopy surveillance (10/2011–5/2013), supplemented with retrospective specimens. Results Loss of MMR protein corresponding to the gene mutated in the germline increased with dysplasia, with frequency of 0 % in normal mucosa, 50–68 % in low-grade dysplasia adenomas, and 100 % in high-grade dysplasia adenomas and carcinomas. Promoter methylation as a putative “second hit” occurred in 1/56 (2 %) of tumors with silenced MMR protein. A general hypermethylation tendency was evaluated by two gene sets, eight CpG island methylator phenotype (CIMP) genes, and seven candidate tumor suppressor genes linked to colorectal carcinoma (CRC). Hypermethylation followed the same trend as MMR protein loss and was present in some low-grade dysplasia adenomas that still expressed MMR protein suggesting the absence of a “second hit.” To assess prospectively collected normal mucosa for carcinogenic “fields,” the specimen donors were stratified according to age at biopsy (50 years or below vs. above 50 years) and further according to the absence vs. presence of a (previous or concurrent) diagnosis of CRC. In mutation carriers over 50 years old, two markers from the candidate gene panel (SFRP1 and SLC5A8) revealed a significantly elevated average degree of methylation in individuals with CRC diagnosis vs. those without. Conclusions Our findings emphasize the importance and early appearance of epigenetic alterations in LS-associated tumorigenesis. The results serve early detection and assessment of progression of CRC

Cancer-Predicting Gene Expression Changes in Colonic Mucosa of Western Diet Fed Mlh1(+/-) Mice

Peer reviewe

Re-Structuring of Marine Communities Exposed to Environmental Change: A Global Study on the Interactive Effects of Species and Functional Richness

Species richness is the most commonly used but controversial biodiversity metric in studies on aspects of community stability such as structural composition or productivity. The apparent ambiguity of theoretical and experimental findings may in part be due to experimental shortcomings and/or heterogeneity of scales and methods in earlier studies. This has led to an urgent call for improved and more realistic experiments. In a series of experiments replicated at a global scale we translocated several hundred marine hard bottom communities to new environments simulating a rapid but moderate environmental change. Subsequently, we measured their rate of compositional change (re-structuring) which in the great majority of cases represented a compositional convergence towards local communities. Re-structuring is driven by mortality of community components (original species) and establishment of new species in the changed environmental context. The rate of this re-structuring was then related to various system properties. We show that availability of free substratum relates negatively while taxon richness relates positively to structural persistence (i.e., no or slow re-structuring). Thus, when faced with environmental change, taxon-rich communities retain their original composition longer than taxon-poor communities. The effect of taxon richness, however, interacts with another aspect of diversity, functional richness. Indeed, taxon richness relates positively to persistence in functionally depauperate communities, but not in functionally diverse communities. The interaction between taxonomic and functional diversity with regard to the behaviour of communities exposed to environmental stress may help understand some of the seemingly contrasting findings of past research