Biomedical subjects

C L Hedley

Publications and source records attributed to C L Hedley.

12 recordsLinked to original sources

PubMed2003

Mutant alleles at the rugosus loci in pea affect seed moisture sorption isotherms and the relations between seed longevity and moisture content.

Pea (Pisum sativum L.) mutant near-isogenic lines (RRrbrb, rrRbRb, rrrbrb) with lower starch but higher lipid contents, brought about by lesions in the starch biosynthetic pathway, had seed moisture sorption isotherms displaced below that of the wild type (RRRbRb). The negative logarithmic relationship between seed longevity and seed storage moisture content (%, f.wt basis), determined in hermetic storage at 65 degrees C, also differed: longevity in the mutant near-isogenic lines was poorer and less sensitive to moisture content than in the wild type (i.e. C(W) was lower). The low-moisture-content limit (m(c)) to this relation also differed, being lower in the mutant near-isogenic lines (5.4-5.9%) than in the wild type (6.1%). In contrast, all four near-isogenic lines showed no difference (P >0.25) in the negative semi-logarithmic relationship between equilibrium relative humidity (ERH) and seed longevity. It is concluded that the effect of these alleles at the r and rb loci on seed longevity was largely indirect; a consequence of their effect on seed composition and hence on moisture sorption isotherms. However, this explanation could not be invoked at moisture contents below m(c) where differences in longevity remained substantial (RRRbRb double that of rrrbrb). Hence, these mutant alleles affected seed longevity directly at very low moisture contents.

Alleles↗

PubMed2002

Structural studies of starches with different water contents.

The proportion of double helices in starches from a series of pea [rb, rug4-b, rug3-a, and lam-c mutants, and the wild type (WT) parental line], potato and maize (normal and low amylose), and wheat (normal) lines, ranged from about 30-50% on a dry weight basis. In relatively dry starch powders, only about half of the double helices were in crystalline order, this proportion being higher for A-type than for B-type starches. Using starch from WT pea as an example, it was found that increasing water content results in an increase in total crystallinity. When the water content was raised to a level similar to that in excess water, the proportion of crystallinity was close to the proportion of double helices (DH). Measuring crystallinity in starches with a high water content is difficult using traditional methods such as x-ray diffraction. A method was developed, therefore, for determining starch structural characteristics in excess water by measuring the enthalpy of gelatinization transition in quasi-equilibrium differential scanning calorimetry (DSC) experiments. It is suggested that DH% = DeltaH(sp)/DeltaH(DH) x 100%, where DeltaH(sp) and DeltaH(DH) represent the specific enthalpies of gelatinisation transition, DeltaH(sp) being measured as J/g dry starch weight and DeltaH(DH) as J/g DH, in starch. Studies on potato and maize starches in excess water and in 0.6M KCl showed, respectively, that DeltaH(DH) was 36.3 and 35.6 J/g for B-type polymorphs and 33.0 and 35.0 J/g for A-type polymorphs. For C-type starches, such as those from pea, intermediate values of DeltaH(DH), related to the proportions A-/B-polymorphs, should be used. The type of crystallinity in starch can be determined by the shift in peak temperature for thermograms in excess water and in excess 0.6M KCl. For B-polymorphs this shift was found to be approximately 2-3 degrees C and for A-polymorphs approximately 7-12 degrees C. The ratio between ordered areas with both A- and B-polymorphs can be determined from the enthalpies of disruption of each area. These enthalpies can be obtained by deconvolution of bimodal thermograms produced by C-type starches in excess 0.6M KCl. This methodical approach can be applied to all starches that give a sharp gelatinisation thermogram in excess water. Using a range of methods, including DSC, it was found that starch granules from the mutant peas are constructed in a similar way to those from the WT, with B-polymorphs in the centre and A-polymorphs at the periphery of all granules. The proportion of A/B-polymorphs, however, differed between the mutants. It was found that in addition to increasing the total crystallinity, increasing the water content within the granules also resulted in an increase in the proportion of B-polymorphs.

Biopolymers↗

PubMed2001

The effect of water content on the ordered/disordered structures in starches.

(13)C cross-polarization magic angle spinning NMR has been used to study the ordered and disordered structures of starches with different water contents. The amorphous regions of starch have been shown to produce NMR patterns only if they are in a glassy state, the widths, positions, and areas of the peaks to some extent being dependent on the temperature and the water content of the starch. In the amorphous region, the peaks were all Gaussian in shape, while the peaks in the ordered regions had Lorentz profiles. Water contents in the range 10-50% did not influence the proportion of double helices in the starch. Decreasing the water content to 1-3%, however, resulted in a significant decrease in the proportion of double helices, the effect being greater in B- than in A-type starches. It is suggested that short-range order structures in starches (double helices) are stabilized by becoming part of long-range order structures (crystallites).

Carbohydrate Conformation↗

PubMed2001

Analysis of the raffinose family oligosaccharide pathway in pea seeds with contrasting carbohydrate composition.

Raffinose family oligosaccharides (RFOs) are synthesized by a set of galactosyltransferases, which sequentially add galactose units from galactinol to sucrose. The accumulation of RFOs was studied in maturing seeds of two pea (Pisum sativum) lines with contrasting RFO composition. Seeds of the line SD1 accumulated stachyose as the predominant RFO, whereas verbascose, the next higher homolog of stachyose, was almost absent. In seeds of the line RRRbRb, a high level of verbascose was accumulated alongside with stachyose. The increase in verbascose in developing RRRbRb seeds was associated with galactinol-dependent verbascose synthase activity. In addition, a galactinol-independent enzyme activity was detected, which catalyzed transfer of a galactose residue from one stachyose molecule to another. The two enzyme activities synthesizing verbascose showed an optimum at pH 7.0. Both activities were almost undetectable in SD1. Maximum activity of stachyose synthase was about 4-fold higher in RRRbRb compared with SD1, whereas the activities of galactinol synthase and raffinose synthase were only about 1.5-fold higher in RRRbRb. The levels of galactinol synthase and stachyose synthase activity were reflected by steady-state levels of corresponding mRNAs. We suggest that the accumulation of verbascose in RRRbRb was controlled by a coordinated up-regulation of the last steps of verbascose biosynthesis.

Disaccharides↗

PubMed2000

The rug3 locus of pea encodes plastidial phosphoglucomutase.

Two cDNA clones were isolated from pea (Pisum sativum L.) and their deduced amino acid sequences shown to have significant homology to phosphoglucomutases from eukaryotic and prokaryotic sources. The longer cDNA contained a putative transit-peptide-encoding sequence, supporting the hypothesis that the isolated clones represent the cytosolic and plastidial isoforms of phosphoglucomutase in pea. Plastid protein import assays confirmed that the putative plastidial isoform was targeted to the plastid stroma where it was proteolytically processed. Expression, co-segregation, linkage, and molecular analyses have confirmed that the rug3 locus of pea encodes plastidial phosphoglucomutase. Mutations at this locus result in a near-starchless phenotype of the plant.

Amino Acid Sequence↗

PubMed1999

Influence of genotype and processing on the in vitro rate of starch hydrolysis and resistant starch formation in peas (Pisum sativum L.).

The formation of resistant starch (RS) and the rate of starch hydrolysis were evaluated in vitro in a wild type of green-seeded pea genotype RRRbRb BC3 (33-Am) with 32.7% amylose content and in two mutants RRrbrb BC3 (23-Am) and rrRbRb BC3 (65-Am) with amylose contents of 23.3 and 65.1%, respectively. Pea samples were intact or homogenized and subjected either to autoclaving or to boiling at atmospheric pressure. The amount of RS (total starch basis) varied from 6.2 to 12.9% in the 23-Am products and from 31.2 to 33.4% in the 65-Am products. The RS level of the 33-Am product with a regular amylose content was 11.0%. Both the 23-Am and the 65-Am products were abundant sources of dietary fiber (39 and 34%, dry matter basis, respectively) versus 23% in the regular pea product. The amylose/amylopectin ratio was an important determinant of the rate of starch hydrolysis. The hydrolysis indices (HI) and predicted glycemic indices were lowest in the 65-Am peas (HI range = 42-59) as compared to the 23-Am peas (HI range = 53-84). It is concluded that the pea genotypes covered a wide range in starch availability, which is likely to affect nutritional parameters such as glycemic responses and colonic delivery of starch.

Amylose↗

PubMed1998

Mutations in the gene encoding starch synthase II profoundly alter amylopectin structure in pea embryos.

Mutations at the rug5 (rugosus5) locus have been used to elucidate the role of the major soluble isoform of starch synthase II (SSII) in amylopectin synthesis in the developing pea embryo. The SSII gene maps to the rug5 locus, and the gene in one of three rug5 mutant lines has been shown to carry a base pair substitution that introduces a stop codon into the open reading frame. All three mutant alleles cause a dramatic reduction or loss of the SSII protein. The mutations have pleiotropic effects on the activities of other isoforms of starch synthase but apparently not on those of other enzymes of starch synthesis. These mutations result in abnormal starch granule morphology and amylopectin structure. Amylopectin contains fewer chains of intermediate length (B2 and B3 chains) and more very short and very long chains than does amylopectin from wild-type embryos. The results suggest that SSII may play a specific role in the synthesis of B2 and B3 chains of amylopectin. The extent to which these findings can be extrapolated to other species is discussed.

Amylopectin↗

PubMed1996

Improved method for the analysis of alpha-galactosides in pea seeds by capillary zone electrophoresis. Comparison with high-performance liquid chromatography-triple-pulsed amperometric detection.

A capillary zone electrophoretic (CZE) method is described for the analysis of the raffinose family of oligosaccharides (RFO) in pea seeds. Extraction of RFO was carried out in 80% ethanol and the extract was passed through a Sep-Pak C18 cartridge. This proved to be an improvement on the currently accepted purification technique. High-quality electrophoregrams were obtained which allowed the separation and quantitation of sucrose, raffinose, stachyose and verbascose The CZE results were compared with those obtained by anion-exchange high-performance liquid chromatography coupled to a triple-pulsed amperometric detection (HPAC-PAD). The samples were obtained from four pea strains which were near-isogenic except for genes at the r and rb loci. A high degree of precision and reproducibility was obtained for the RFO compositions of all the pea strains. No statistically significant differences (p < or equal to 0.05) were found between the two analytical techniques using paired Student-t tests.

Chromatography, High Pressure Liquid↗

PubMed1996

Evolution and kinetics of monosaccharides, disaccharides and alpha-galactosides during germination of lentils.

The effect of light and seed rinsing during the germination of lentil seeds (Lens culinaris var. vulgaris, cultivar Magda-20) on the level of monosaccharides, disaccharides and alpha-galactosides (raffinose, ciceritol and stachyose) was investigated. The total soluble sugar content corresponded to about 9% of the mature seed weight, about 65% of which was alpha-galactosides. Germination brought about a large decrease in alpha-galactosides: 18% to 40% losses after 3 days and 100% after 6 days. However, glucose, which was not detected in ungerminated seeds, as well as fructose and sucrose gradually increased during germination. The content of alpha-galactosides decreased more rapidly when germinating seeds were given 6 h light per day, but under these conditions there was also a major reduction in the levels of fructose, glucose and sucrose. Seeds rinsed daily showed a greater reduction of alpha-galactosides and an increase in the levels of fructose, glucose and sucrose. When seeds were germinated for 10 days in the dark with daily rinsing, the content of alpha-galactosides decreased gradually during the first 4 days and they were not detected after 6 days. Under these conditions, fructose, glucose and sucrose, which represented about 3% of the mature seed weight, started increasing after day 2 and represented more than 13% of the germinated seed dry weight after 10 days.

Disaccharides↗

PubMed1996

Evolution of soluble carbohydrates during the development of pea, faba bean and lupin seeds.

Seeds of pea (Pisum sativum L. cv. Ergo), faba bean (Vicia faba ssp. minor Harz., cv. Tibo) and yellow pea lupin (Lupinus luteus L. cv. Juno) were sampled at different days after flowering (DAF) and their content of soluble carbohydrates was determined. Analysis of samples showed that myo-inositol, fructose, glucose, galactose and sucrose were found in high abundance early in development and their content decreased gradually during maturation. alpha-Galactosides, which includes the content of raffinose, stachyose and verbascose, started to appear later in seed development, at 37 DAF in peas, 40 DAF in faba beans and 45 DAF in lupins. Their accumulation increased considerably during seed growth, and the maximum content was obtained in mature seeds; 3.8% in peas, 4.5% in faba beans and 10.4% in lupins. Results obtained for these sugars during seed development were fitted to modelling curves in order to predict sugar content at different development stages.

Carbohydrates↗

PubMed1990

Storage protein gene expression is localised to regions lacking mitotic activity in developing pea embryos. An analysis of seed development in Pisum sativum XIV.

Storage protein gene expression has been studied in relation to mitotic activity to ascertain whether these processes are linked during embryo development in pea. Sections from immature pea embryos were probed by in situ hybridisation to show the pattern of vicilin storage protein gene expression. In addition, the location of mitotic cells was identified using fluorescence microscopy. Vicilin mRNA was first localised in the parenchyma cells of the upper adaxial region of the cotyledons. As the embryos increased in fresh weight, gene expression spread from this region, in a wave-like manner, down and across the cotyledons. The gene was only expressed in those regions of the embryo that lacked mitotic activity.

Fabaceae↗

PubMedDate not supplied

[Induction of delayed luminescence of pea photosystem II with genetically changed level of starch in the seeds].

We studied the curves of delayed luminescence induction in leaves of pea plants with mutations affecting the starch branching enzyme (r locus) and ADPG pyrophosphorylase (rb locus). In mutated pants, a 75% reduction of starch content in seeds was observed. The half-decay time of delayed luminescence intensity during induction for double mutants was shown to be longer than for wild type plants.

Luminescence↗