Charlotte Burton

Modification of Standard Enzymatic Protocol to a Cost-Efficient Format for Mixed-Linkage (1→3,1→4)-β-D-Glucan Measurement

ABSTRACT The current enzymatic assay approach (AACC International Approved Method 32-23) for the measurement of mixed-linkage β-glucan in small grains was modified to a cost-efficient and high-throughput format without compromising the accuracy of the results. Ten barley (Hordeum vulgare L.) genotypes used in the study represented a wide range of β-glucan content levels. A reduced reaction volume is used in the new protocol to adapt to a 96-well plate format. The volume of key components lichenase and β-glucosidase were reduced to 25% of the volume required in the original protocol and the cost per sample was reduced to 22% of that in the original protocol. Labor cost was also decreased to 25% of the original protocol as a result of format changes. The accuracy of the measurement from the modified protocol was comparable to the current standard enzymatic procedure. β-Glucan measurement accuracy of the modified and original protocols were also compared using 21 oat (Avena sativa L.) samples. The results in...

Modified Orcinol–Ferric Chloride Colorimetric Assay for Water-Extractable Arabinoxylan in Wheat

Arabinoxylans are an important quality component for which a simple and rapid analysis method useful for plant breeding needs to be developed. Bial’s orcinol–ferric chloride colorimetric method was modified to allow simultaneous analysis of at least 24 samples in a single run and in small volumes to measure water-extractable arabinoxylan (WEAX) in wheat meal. The modified parameters were reduction to 6N HCl, increased concentrations of orcinol and 10% ferric chloride, and a total volume of 500 µL. The sample turbidity was reduced by rapid cooling in an ice water bath followed by addition of 500 µL of 95% ethanol postincubation. The WEAX results from this protocol were compared with GC/MS values. The wheat varieties of Alta Blanca, Eltan, and Hatton were overestimated by 4.3, 1.5, and 0.9%, respectively, whereas the varieties of Eden, Darwin, Finley, and Boundary were underestimated by 14.4, 8.9, 11.3, and 3.6%, respectively. The modified procedure is useful to determine a range of WEAX concentrations in w...

Molecular and Chemical Characterization of a New Waxy Allele in Barley (Hordeum vulgare L.)

ABSTRACT Barley m38 mutant was selected for its high level of mixed-linkage (1,3),(1,4)-β-d-glucan (MLG) in the grain. This elevated level of MLG was found to be associated with decreased amylose accumulation as well as other chemical composition alterations. Molecular characterization results revealed m38 as a new allele of the Waxy gene, encoding an endosperm-specific granule-bound starch synthase I (GBSSI). Additional mapping data from amylose phenotype and GBSSI gene specific markers supported the conclusion of the GBSSI mutation in m38. The m38 locus contains a nucleotide alteration that would result in the substitution of glycine at position 263 with serine in the putative adenosine-5′-diphosphate-glucose binding domain. This amino acid substitution alters loop structures on the exterior surface of the folded protein and may affect its enzyme activity. Characterizations of m38 in this report provide for a new allele of the Waxy gene and additional evidence of pleiotropic effects on other chemical co...