[show abstract][hide abstract] ABSTRACT: Dough-derived cell wall fragments isolated by ultracentrifugation were largely derived from the starchy endosperm, with some fragments deriving from the aleurone and outer layers, as indicated by fluorescence microscopy. Dough mixing had little effect on the structure and composition of cell wall fragments compared to thin grain sections, as determined by Fourier transform infrared (FTIR) and 1H nuclear magnetic resonance (NMR) spectroscopy. These analyses confirmed that the fragments largely comprised water-unextractable arabinoxylan and β-glucan. FTIR microspectroscopy of dough-derived cell wall fragments prepared from five bread wheat cultivars showed that two largely comprised highly substituted arabinoxylan (cv. Manital and San Pastore), one comprised a mixture of low, medium, and highly substituted arabinoxylan (cv. Hereward), and the remaining two comprised a greater proportion of low substituted arabinoxylan (cv. Claire and Yumai 34). Yumai 34 yielded a greater mass of cell wall material, and its cell walls comprised a high proportion of medium substituted arabinoxylan. Such methods will allow for the impact of bakery ingredients and processing on endosperm cells, including the addition of xylanases, to be investigated in the future to ensure any potential health benefits arising from wheat breeding are realized in the food that reaches the consumer.
Journal of Agricultural and Food Chemistry 03/2013; · 2.91 Impact Factor
[show abstract][hide abstract] ABSTRACT: Two varieties each of spelt (Triticum aestivum var. spelta), durum wheat (Triticum turgidum var. durum), rye (Secale cereale), barley (Hordeum vulgare), oats (Avena sativa), einkorn (Triticum monococcum var. monococcum) and emmer (Triticum turgidum var. dicoccum) (all members of the Pooideae sub-family of grasses) were selected according to variation in their contents of soluble and/or total arabinoxylan (AX) determined during the HEALTHGRAIN diversity screen, together with one genotype of the related “model” grass species Brachypodium distachyon. The spatial distribution of low substituted (LS-AX) and highly substituted arabinoxylan (HS-AX) was determined using FT-IR spectroscopic mapping of transverse thin cross-sections consisting of cell walls only. Variation in cell wall AX composition was observed between the cereals, and compared with that observed for wheat (Triticum aestivum var. aestivum). One line of each cereal type was analysed in more detail using 1H NMR spectroscopy. The results of the two analyses were consistent, showing variation in the composition and structure of the endosperm cell wall AX that is consistent with the genetic relationships of the cereals studied.
Journal of Cereal Science 09/2012; 56(2):134–141. · 2.09 Impact Factor
[show abstract][hide abstract] ABSTRACT: The food-borne bacterial pathogen Campylobacter jejuni efficiently utilizes organic acids such as lactate and formate for energy production. Formate is rapidly metabolized via the activity of the multisubunit formate dehydrogenase (FDH) enzyme, of which the FdhA subunit is predicted to contain a selenocysteine (SeC) amino acid. In this study we investigated the function of the cj1500 and cj1501 genes of C. jejuni, demonstrate that they are involved in selenium-controlled production of FDH, and propose the names fdhT and fdhU, respectively. Insertional inactivation of fdhT or fdhU in C. jejuni resulted in the absence of FdhA and FdhB protein expression, reduced fdhABC RNA levels, the absence of FDH enzyme activity, and the lack of formate utilization, as assessed by (1)H nuclear magnetic resonance. The fdhABC genes are transcribed from a single promoter located two genes upstream of fdhA, and the decrease in fdhABC RNA levels in the fdhU mutant is mediated at the posttranscriptional level. FDH activity and the ability to utilize formate were restored by genetic complementation with fdhU and by supplementation of the growth media with selenium dioxide. Disruption of SeC synthesis by inactivation of the selA and selB genes also resulted in the absence of FDH activity, which could not be restored by selenium supplementation. Comparative genomic analysis suggests a link between the presence of selA and fdhTU orthologs and the predicted presence of SeC in FdhA. The fdhTU genes encode accessory proteins required for FDH expression and activity in C. jejuni, possibly by contributing to acquisition or utilization of selenium.
Journal of bacteriology 05/2012; 194(15):3814-23. · 3.94 Impact Factor
[show abstract][hide abstract] ABSTRACT: Fasting is one of the simplest metabolic challenges that can be performed in humans. We here report for the first time a comprehensive
analysis of the human “fasting metabolome” obtained from analysis of plasma and urine samples in a small cohort of healthy
volunteers, using nuclear magnetic resonance (NMR), gas chromatography- and liquid chromatography-mass spectrometry (GC-MS
and LC-MS). Intra- and inter-individual variation of metabolites was on measurement of four overnight fasting samples collected
from each volunteer over a four week period. One additional sample per volunteer was collected following a prolonged fasting
period of 36h. Amongst a total of 377 quantified entities in plasma around 44% were shown to change significantly in concentration
when volunteers extended fasting from 12 to 36h. In addition to known markers (plasma free fatty acids, glycerol, ketone
bodies) that reflect changes in the body’s fuel management under fasting conditions a wide range of “new” entities such as
α-aminobutyrate as well as other amino and keto acids were identified as fasting markers. Based on multiple correlations amongst
the metabolites and selected hormones in plasma such as leptin or insulin-like-growth-factor-1 (IGF-1), a robust metabolic
network with coherent regulation of a wide range of metabolites could be identified. The metabolomics approach described here
demonstrates the plasticity of human metabolism and identifies new and robust markers of the fasting state.
KeywordsFasting response–Metabolite profiling–Subject variability
[show abstract][hide abstract] ABSTRACT: A combination of enzyme mapping, FT-IR microscopy and NMR spectroscopy was used to study temporal and spatial aspects of endosperm cell wall synthesis and deposition in developing grain of bread wheat cv. Hereward. This confirmed previous reports that changes in the proportions of the two major groups of cell wall polysaccharides occur, with beta-glucan accumulating earlier in development than arabinoxylan. Changes in the structure of the arabinoxylan occurred, with decreased proportions of disubstituted xylose residues and increased proportions of monosubstituted xylose residues. These are likely to result, at least in part, from arabinoxylan restructuring catalysed by enzymes such as arabinoxylan arabinofurano hydrolase and lead to changes in cell wall mechanical properties which may be required to withstand stresses during grain maturation and desiccation.
[show abstract][hide abstract] ABSTRACT: Previous studies using spectroscopic imaging have allowed the spatial distribution of structural components in wheat endosperm cell walls to be determined. FT-IR microspectroscopy showed differing changes in arabinoxylan (AX) structure, during grain development under cool/wet and hot/dry growing conditions, for differing cultivars (Toole et al. in Planta 225:1393-1403, 2007). These studies have been extended using Raman microspectroscopy, providing more details of the impact of environment on the polysaccharide and phenolic components of the cell walls. NMR studies provide complementary information on the types and levels of AX branching both early in development and at maturity. Raman microspectroscopy has allowed the arabinose:xylose (A/X) ratio in the cell wall AX to be determined, and the addition of ferulic acid and related phenolic acids to be followed. The changes in the A/X ratio during grain development were affected by the environmental conditions, with the A/X ratio generally being slightly lower for samples grown under cool/wet conditions than for those from hot/dry conditions. The degree of esterification of the endosperm cell walls with ferulic acid was also affected by the environment, being lower under hot/dry conditions. The results support earlier suggestions that AX is either delivered to the cell wall in a highly substituted form and is remodelled through the action of arabinoxylan arabinofuranohydrolases or arabinofuranosidases, or that low level substituted AX are incorporated into the wall late in cell wall development, reducing the average degree of substitution, and that the rate of this remodelling is influenced by the environment. (1)H NMR provided a unique insight into the chemical structure of intact wheat endosperm cell walls, providing qualitative information on the proportions of mono- and disubstituted AX and the levels of branching of adjacent units. The A/X ratio did not change greatly with either the development stage or the growth conditions, but the ratio of mono- to disubstituted Xylp residues increased markedly (by about fourfold) in the more mature samples, confirming the changes in branching levels determined using FT-IR. To the best of our knowledge, this is the first time that intact endosperm cell walls have been studied by (1)H NMR.
[show abstract][hide abstract] ABSTRACT: To discriminate orange juice from grapefruit juice in a context of fraud prevention, (1)H NMR data were submitted to different treatments to extract informative variables which were then analysed using multivariate techniques. Averaging contiguous data points of the spectrum followed by logarithmic transformation improved the results of the data analysis. Moreover, supervised variable selection methods gave better rates of classification of the juices into the correct groups. Last, independent-component analysis gave better classification results than principal-component analysis. Hence, ICA may be an efficient chemometric tool to detect differences in the (1)H NMR spectra of similar samples, and so may be useful for authentication of foods.
Analytical and Bioanalytical Chemistry 02/2008; 390(1):419-27. · 3.66 Impact Factor
[show abstract][hide abstract] ABSTRACT: This study investigates the use of high resolution 1H NMR as a suitable alternative to the standard chromatographic method for the determination of adulteration of orange juice (Citrus sinensis) with grapefruit juice (Citrus paradisi) based on flavonoid glycoside content. Fifty-nine orange juices (OJ), 23 grapefruit juices (GJ) and 10 blends (OG), obtained from local retail outlets were used to assess the performance of the 1H NMR method. The work presented here introduces the Evolving Window Zone Selection (EWZS) function that holds promise for the automatic detection of spectral regions tailored to discriminate predefined groups. This technique was applied on the pre-processed 1H NMR spectra of the 92 juices. Independent Component Analysis (ICA) is a good alternative to Principal Component Analysis (PCA) for recovering linearly-mixed unobserved multidimensional independent signals and has been used in this study to build supervised models that classify the samples into three categories, OJ, GJ, OG. The regions containing the known flavonoid glycoside markers were selected as well as another zone containing the signals of sucrose, alpha-glucose and other components that were tentatively attributed. ICA was applied on three different groups of selected variables and showed good results for both discrimination and interpretation of the signals. Up to 97.8% of the juices were correctly attributed. This method gave better results than the commonly used PCA method. In addition, the time required to carry out the 1H NMR analysis was less than half the time of the standard chromatographic method.