Observational data show an inverse association between the consumption of wholegrain foods, and inflammation and related diseases. Although the underlying mechanisms are unclear, wholegrains, and in particular the aleurone layer, contain a wide range of components with putative antioxidant and anti-inflammatory effects. We evaluated the effects of a diet high in wheat aleurone on plasma antioxidants status, markers of inflammation and endothelial function. In this parallel, participant-blinded intervention, seventy-nine healthy, older, overweight participants (45–65 years, BMI>25 kg/m2) incorporated either aleurone-rich cereal products (27 g aleurone/d), or control products balanced for fibre and macronutrients, into their habitual diets for 4 weeks. Fasting blood samples were taken at baseline and on day 29. Results showed that, compared to control, consumption of aleurone-rich products provided substantial amounts of micronutrients and phytochemicals which may function as antioxidants. Additionally, incorporating these products into a habitual diet resulted in significantly lower plasma concentrations of the inflammatory marker, C-reactive protein (P = 0·035), which is an independent risk factor for CVD. However, no changes were observed in other markers of inflammation, antioxidant status or endothelial function. These results provide a possible mechanism underlying the beneficial effects of longer-term wholegrain intake. However, it is unclear whether this effect is owing to a specific component, or a combination of components in wheat aleurone.

Fermentation of dietary fibre by the gut microflora may enhance levels of SCFA, which are potentially chemoprotective against colon cancer. Functional food containing wheat aleurone may prevent cancer by influencing cell cycle and cell death. We investigated effects of fermented wheat aleurone on growth and apoptosis of HT29 cells. Wheat aleurone, flour and bran were digested and fermented in vitro. The resulting fermentation supernatants (fs) were analysed for their major metabolites (SCFA, bile acids and ammonia). HT29 cells were treated for 24–72 h with the fs or synthetic mixtures mimicking the fs in SCFA, butyrate or deoxycholic acid (DCA) contents, and the influence on cell growth was determined. Fs aleurone was used to investigate the modulation of apoptosis and cell cycle. The fermented wheat samples contained two- to threefold higher amounts of SCFA than the faeces control (blank), but reduced levels of bile acids and increased concentrations of ammonia. Fs aleurone and flour equally reduced cell growth of HT29 more effectively than the corresponding blank and the SCFA mixtures. The EC50 (48 h) ranged from 10 % (flour) to 19 % (blank). Markedly after 48 h, fs aleurone (10 %) significantly induced apoptosis and inhibited cell proliferation by arresting the cell cycle in the G0/G1 phase. In conclusion, fermentation of wheat aleurone results in a reduced level of tumour-promoting DCA, but higher levels of potentially chemopreventive SCFA. Fermented wheat aleurone is able to induce apoptosis and to block cell cycle – two essential markers of secondary chemoprevention.

To elucidate the molecular aspects of cereal endosperm development, a good understanding of the histo-differentiation process is necessary. Reviewing the extensive history of lightand electron-microscopic investigations has revealed similarities between cereals with respect to cell ontogeny. Thus, using barley as a model and taking into account many developmental mutants in this and other species, a model for endosperm differentiation is proposed. Based on this model, molecular studies on endosperm gene-regulation are discussed and the available techniques for further investigations are considered.

Protein disulfide isomerase (PDI E.C. 5.3.4.1) catalyses the formation of disulfide bonds in secretory or cell-surface proteins during their biosynthesis. PDI activity is associated with the microsomal fraction of the cell and has been found in several mammalian tissues and in developing wheat endosperm.

The embryo and aleurone layer of germinating seeds of the wheat cultivar Galahad showed PDI activity associated with a microsome-enriched fraction of cell homogenates PDI activity in microsome-enriched fractions from aleurone layers of normally germinating seeds was significantly higher than in their abnormally germinating counterparts and diminished as germination proceeded.

Dry embryos and germinating embryos showed less PDI activity than aleurone layers. There was no significant difference in PDI activity between microsome-enriched fractions from embryos of normally and abnormally germinating seeds at 3 or 6 days of germination.

Wheat aleurone PDI was partially purified using gel filtration and had a molecular mass of 110–130 kDa and a monomeric molecular mass of ca. 57 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis.

Ageing of wheat seeds (Triticum aestivum cv. Galahad, 1986 harvest) during storage is accompanied by an increase in the proportion of abnormal seedlings produced when aged seed lots germinate. The response of aleurone layers from normally and abnormally germinated seeds to heat shock has been investigated. [14C]-amino acid incorporation into aleurone layers from 3-d normally and abnormally germinated seeds diminished after 4 h at 42°C but the aleurone layers were able to recover significantly during a 5-h period at 20°C following heat shock. [35S]-methionine pulse-labelling of aleurone proteins showed that α-amylase isoenzyme synthesis was abolished upon heat shock in aleurone layers for both normally and abnormally germinated seeds and that only aleurone layers from normally germinated seeds were capable of recommencing substantial α-amylase synthesis during the recovery period. Heat shock caused aleurone layers from abnormally germinated seeds to synthesize polypeptides not synthesized by aleurone layers from normally germinated seeds under the same conditions. The synthesis of certain polypeptides by aleurone layers from abnormally germinated seeds was enhanced at 20°C, 42°C and during recovery compared with normally germinated seeds. These results are discussed in terms of the control of gene expression during heat shock and recovery during seed ageing.

The development of (1→3,1→4)-β-glucan 4-glucanohydrolase (EC 3.2.1.73) has been examined in germinating wheat (Triticum aestivum cv. Millewa) grain, and in isolated aleurone layers and scutella. Activity is first detected in extracts of intact grain 2–3 days after the initiation of germination and thereafter increases until 6 days. In isolated aleurone layers, (1→3,1→4)-β-glucanase activity is secreted for up to 4 days. Treatment of aleurone layers with gibberellic acid (GA3) results in a 2-fold enhancement of secreted enzyme for the first 2 days, but activity decreases after 2 days. (1→3,1→4)-β-Glucanase secretion from GA3-treated aleurone layers clearly precedes the secretion of amylase and (1→4)-β-xylanase. Small but significant levels of cellulase are secreted from aleurone layers after GA3 treatment. Isolated scutella also secrete (1→3,1→4)-β-glucanase but GA3 has little apparent effect on secretion patterns from this tissue. Proteins secreted from excised aleurone layers and scutella, together with those in homogenates of intact, germinated grain, have been examined by Western blot analysis, using monoclonal antibodies against barley (1→3,1→4)-β-glucanases as probes. In each case the wheat (1→3,1→4)-β-glucanase is recognised only by the monoclonal antibody that is specific for barley isoenzyme El.

Cereal grain germination and early seedling growth involve the co-ordinated action of endosperm and embryo tissues to mobilize the storage reserves of the starchy endosperm. This mobilization is accomplished by hydrolases secreted from the aleurone and scutellar tissues. The breakdown products are then transported to the growing seedling by the scutellum. This resource-harvesting system is regulated at multiple levels. One well-defined aspect of control is brought about by the hormone gibberellin (GA). Gibberellin is released from the embryo upon imbibition and activates the aleurone cells. The secretory apparatus of the aleurone then proliferates, supporting increased hydrolase synthesis and secretion to degrade the starchy endosperm. The molecules that regulate this response to GA are now being increasingly characterized. Elements such as cGMP, calcium, calmodulin and protein kinases are well known as regulators in other eukaryotic cell types and are emerging as key control factors in the aleurone hormone response. However, superimposed upon this molecular regulatory system is another level of control, the structural pattern of tissues and stored macro- molecules that was laid down during grain development. It is the interaction of these structural motifs combined with the molecular regulatory mechanisms that ensure the appropriate timing and positioning of hydrolase production and endosperm reserve mobilization. This integrated control system ensures an extended release of nutrients to fuel early seedling growth.

Aleurone layers of mature germinating barley (Hordeum vulgare, cv. Himalaya) grains respond to heat shock by synthesizing heat-shock proteins (HSPs) and by selectively suppressing the synthesis of proteins normally translated by endoplasmic reticulum (ER)-bound ribosomes. To determine if this also was the case during seed development, we investigated the synthesis of proteins translated by ER-bound ribosomes in heat-shocked aleurone layers isolated from developing barley grains. The optimal induction temperature for the heat shock response in developing aleurone layers was 37.5°C, and temperatures above 42°C inhibited translation. HSPs with apparent molecular masses of 71.1, 66.2, 57.8, 19.1 and 18.8 kDa were induced. Other studies have shown that, in gibberellic acid (GA)-induced aleurone layers from mature barley grains, these temperatures were 40°C and 45°C, respectively. Furthermore, in developing aleurone layers, mRNAs encoding proteins translated by ER-bound ribosomes (mRNAs for a lipid transfer protein and a putative amylase/protease inhibitor) remained stable during heat shock. The ER membranes themselves remained in stacks, but the lumen became distended with electron-dense material. Heat shock prevented the movement of proteins from the ER into the rest of the endomembrane pathway. In contrast, other studies show that in mature, GA-induced aleurone layers, heat shock dissociates ER stacks and blocks translation, but the processing of secretory proteins in the endomembrane pathway is not inhibited. The observation that the same tissue at different developmental stages may respond differently to heat shock indicates that components of the heat-shock response are developmentally regulated. This system provides an opportunity to better understand the nuances of the heat-shock response, especially the post-transcriptional gene regulatory mechanisms that occur.

Aleurone flour (ALF) is a rich source of natural folate (>500 μg/100 g wet weight). Our objective was to establish whether intake of ALF in man can significantly improve folate status and reduce plasma homocyst(e)ine. We performed a randomised, controlled intervention, of 16 weeks duration, in free-living healthy individuals (mean age 46–52 years). Participants were assigned to one of three groups: ALF, 175 g bread made with ALF and placebo tablet each day; PCS, 175 g bread made with pericarp seed coat (PCS) flour and placebo tablet each day (low-folate control); or FA, 175 g bread made with PCS flour and tablet containing 640 μg folic acid each day (high-folate control). The daily folate intake contributed by the bread and tablet was 233 μg in the PCS group, 615 μg in the ALF group and 819 μg in the FA group. The number of participants completing all phases of the PCS, ALF and FA interventions was twenty-five, twenty-five and eighteen, respectively. Plasma and red-cell folate increased significantly (P<0·0001) and plasma homocyst(e)ine decreased significantly (P<0·0001) in the ALF and FA groups only. Plasma folate and red-cell folate in the ALF group (mean, 95 % CI) increased from baseline values of 12·9 (9·9, 15·7) nmol/l and 509 (434, 584) nmol/l to 27·1 (22·5, 31·7) nmol/l and 768 (676, 860) nmol/l, respectively. Plasma homocyst(e)ine in the ALF group decreased from 9·1 (8·2, 10·0) μmol/l at baseline to 6·8 (6·2, 7·5) μmol/l after 16 weeks. In conclusion, moderate dietary intake of ALF can increase red-cell folate and decrease plasma homocyst(e)ine substantially.

The range of processes regulated by gibberellins (GAs) covers all aspects of the life history of the plant from seed germination to vegetative growth and flowering. In seeds there has been an intensive search, using the techniques of both biochemistry and cell biology, for the regulatory molecules linking GA perception to gene regulation and the events of germination. Although a GA receptor has yet to be identified, the site of perception has been localized to the plasma membrane. Calmodulin, Ca2+ and cGMP have also been identified as elements of the GA signal transduction pathway. These regulators parallel many of the signalling elements identified in the transduction of other signals such as phytochrome and ABA. Studies of GA-regulated gene expression, principally of the α-amylases of cereal aleurone, have identified core GA-responsive promoter elements, such as the gibberellin response element (GARE), box-1 and pyrimidine boxes, as well as elements that may lend specificity to GA-regulated expression, such as the Opaque-2-similar element (O2S), and TRE and CRE motifs. One of the most striking features of all of these studies of the molecular basis of GA action is the interaction of GA-dependent regulatory elements with those of other factors such as ABA. GA-response elements also appear to be conserved between disparate GA-response systems. For example, Myb transcription factors appear to regulate a multitude of GA-induced genes in cereal aleurone as well as to alter GA responses when expressed in Arabidopsis. Thus the study of GA signal transduction and response systems is highlighting the conservation of regulatory elements used by plants. These common factors, used by distinct signal transduction systems, provide a molecular basis for the integration of the GA signal with other growth regulators that is the hallmark of plant growth and development.