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The varied appearances of starch particles in smears and paraffin sections.

Evidence is presented to show that maize starch particles, usually recognized in histological sections by their Maltese cross birefringence under cross-polarized light, lose this characteristic appearance when transected by the microtome blade. Such particles are, therefore, likely to be overlooked. Though maize starch usually enters the abdomen during surgery, the female genital tract is another possible route from sources such as contraceptive devices. Other types of starch particle have different shapes, sizes and surface markings and these may be distinguished from maize starch by light and electron optical methods.

Adult↗

Soluble starch synthase I: a major determinant for the synthesis of amylopectin in Arabidopsis thaliana leaves.

A minimum of four soluble starch synthase families have been documented in all starch-storing green plants. These activities are involved in amylopectin synthesis and are extremely well conserved throughout the plant kingdom. Mutants or transgenic plants defective for SSII and SSIII isoforms have been previously shown to have a large and specific impact on the synthesis of amylopectin while the function of the SSI type of enzymes has remained elusive. We report here that Arabidopsis mutants, lacking a plastidial starch synthase isoform belonging to the SSI family, display a major and novel type of structural alteration within their amylopectin. Comparative analysis of beta-limit dextrins for both wild type and mutant amylopectins suggests a specific and crucial function of SSI during the synthesis of transient starch in Arabidopsis leaves. Considering our own characterization of SSI activity and the previously described kinetic properties of maize SSI, our results suggest that the function of SSI is mainly involved in the synthesis of small outer chains during amylopectin cluster synthesis.

Amylopectin↗

Production of high-starch, low-glucose potatoes through over-expression of the metabolic regulator SnRK1.

Transgenic potato (Solanum tuberosum cv. Prairie) lines were produced over-expressing a sucrose non-fermenting-1-related protein kinase-1 gene (SnRK1) under the control of a patatin (tuber-specific) promoter. SnRK1 activity in the tubers of three independent transgenic lines was increased by 55%-167% compared with that in the wild-type. Glucose levels were decreased, at 17%-56% of the levels of the wild-type, and the starch content showed an increase of 23%-30%. Sucrose and fructose levels in the tubers of the transgenic plants did not show a significant change. Northern analyses of genes encoding sucrose synthase and ADP-glucose pyrophosphorylase, two key enzymes involved in the biosynthetic pathway from sucrose to starch, showed that the expression of both was increased in tubers of the transgenic lines compared with the wild-type. In contrast, the expression of genes encoding two other enzymes of carbohydrate metabolism, alpha-amylase and sucrose phosphate synthase, showed no change. The activity of sucrose synthase and ADP-glucose pyrophosphorylase was also increased, by approximately 20%-60% and three- to five-fold, respectively, whereas the activity of hexokinase was unchanged. The results are consistent with a role for SnRK1 in regulating carbon flux through the storage pathway to starch biosynthesis. They emphasize the importance of SnRK1 in the regulation of carbohydrate metabolism and resource partitioning, and indicate a specific role for SnRK1 in the control of starch accumulation in potato tubers.

Carboxylic Ester Hydrolases↗

Cyclodextrin glycosyltransferase may be the only starch-degrading enzyme in Bacillus macerans.

Cyclodextrin glycosyltransferase (CGTase) was released into the culture fluid by Bacillus macerans predominantly in the late stationary phase of growth and during autolysis in the presence of either glucose or starch as a carbon source. In both cases significant soluble intracellular enzyme activity could be observed in the early stationary phase, and a low non-soluble intracellular CGTase activity could be demonstrated also in the exponential growth phase in the presence of starch. At the end of the exponential phase the non-soluble specific intracellular enzyme activity was found to be constant with a value of 0.63 +/- 0.06 nkat/10(9) viable cells. Since amylase activity could not be detected in any intracellular or extracellular sample taken at any culture time, we conclude that cellbound CGTase is the only starch-digesting enzyme in growing B. macerans and, hence, may be fully responsible for the degradation of starch in the culture fluid.

Amylases↗

Effects of variously processed starch on pH of human dental plaque.

The aim of this investigation was to study the effect of starch in various cereal products on pH of human dental plaque in vivo as compared to availability to the alpha-amylase of starch in vitro. Three separate experimental series were performed including: 1) rye and wheat products, 2) wheat products processed under both mild and severe conditions, and 3) wheat products with and without added sucrose. The pH drop in dental plaque was studied after a mouthrinse in groups of 10 subjects. Solutions of glucose or sucrose were used as references. No differences were found between boiled rye and boiled wheat, nor between rye bread and wheat bread. Severely popped wheat resulted in the greatest pH drop, followed, in order, by drum-dried and extrusion-cooked products. Mildly steam-flaked and mildly dry-autoclaved products induced comparable small pH falls. Sucrose-sweetened wheat products resulted in lower plaque pH values compared to unsweetened products. This effect was most pronounced with a mildly processed sample. In all three series, the glucose and sucrose reference solutions showed the greatest pH drop. In conclusion, all processing methods increased the fermentability of starch in human dental plaque in vivo. Thus, the more severe the processing conditions the more prominent the pH falls. The pH drop in plaque in vivo correlated (r = 0.96 in Series 1 and r = 0.92 in Series 2) with the rate of starch hydrolysis with alpha-amylase in vitro.

Biological Availability↗

The production of alpha-cyclodextrin by enzymatic degradation of starch.

The maximum concentration of alpha-cyclodextrin for the enzymatic degradation of starch is limited to about 13.5 g X 1(-1). By addition of decanol, the equilibrium of the reaction system can be shifted towards an alpha-cyclodextrin yield of 50% even at high substrate concentrations. The main variables of the decanol process--pH, temperature, substrate quality, substrate, and enzyme concentration--have been studied. The cyclodextrin-glycosyltransferase from Klebsiella pneumoniae M5 al can preferentially be employed at pH 6 to 8, temperatures of 40 to 50 degrees C and a decanol concentration of 0.1 kg-1 starch. The dextrose equivalent of starch is important with respect to the maximum achievable starch concentration, but not with respect to the reaction. Under process conditions, the rate of alpha-cyclodextrin evolution is limited by the enzymatic reaction and not by mass transfer of decanol into the aqueous phase.

Cyclodextrins↗

Determination of the concentration of maltose- and starch-like compounds in drinking water by growth measurements with a well-defined strain of a Flavobacterium species.

The growth kinetics of Flavobacterium sp. strain S12 specialized in the utilization of glycerol, and a number of oligo- and polysaccharides were determined in batch-culture experiments at 15 degrees C in pasteurized tap water supplied with very low amounts of substrates. Kss for the growth on maltotriose, maltotetraose, maltopentaose, and maltohexaose were 0.03 microM or less and below those for glucose (1.5 microM) and maltose (0.16 microM). Kss for starch, amylose, and amylopectin were 8.4, 25.6, and 11.0 micrograms of C per liter, respectively. A yield of 2.3 X 10(7) CFU/micrograms of C on the oligo- and polysaccharides was calculated from the linear relationships observed between maximum colony counts in pasteurized tap water and the concentrations (usually below 25 micrograms of C per liter) of supplied compounds. The maximum colony counts of strain S12 grown in various types of raw water and tap water revealed that raw water contained only a few micrograms of maltose- and starch-like compounds per liter; in tap water the concentrations were all below 1 microgram of C and usually below 0.1 microgram of C per liter. The application of starch-based coagulant aids gave increased concentrations of maltose- and starch-like compounds in the water during treatment, but these concentrations were greatly reduced by coagulation and sedimentation, rapid sand filtration, and slow sand filtration.

Flavobacterium↗

Influence of elevated temperature on starch hydrolysis by enterotoxin-positive and enterotoxin-negative strains of Clostridium perfringens type A.

Enterotoxin-positive (Ent+) and enterotoxin-negative (Ent-) strains of Clostridium perfringens were cultured in Duncan-Strong sporulation medium containing starch at 37 and 46 degrees C. At 37 degrees C, all strains degraded starch and sporulated well. However, only Ent- strains could hydrolyze starch, grow extensively, and sporulate at 46 degrees C. Growth, sporulation, and starch hydrolysis by Ent+ strains at 46 degrees C were equivalent to those obtained at 37 degrees C when alpha-amylase was added to the cultures during growth. The total amount of extracellular plus intracellular amylase in cultures of Ent+ strains was significantly less in cells incubated at 46 degrees C than in cells incubated at 37 degrees C. These results contradict an earlier report that Ent+ strains cannot sporulate well near their optimal growth temperature (R. G. Labbe and C. L. Duncan, Can. J. Microbiol. 20:1493-1501, 1974) and suggest that synthesis of alpha-amylase in Ent+ strains is regulated by temperature.

Amylases↗

Biodegradability and mechanical properties of poly-(beta-hydroxybutyrate-co-beta-hydroxyvalerate)-starch blends.

Wheat starch granules and poly-(beta-hydroxybutyrate-co-beta-hydroxyvalerate) [P(HB-co-HV), (19.1 mol% HV)] were blended at 160 degrees C. Increasing the starch content from 0 to 50% (wt/wt) decreased the tensile strength of P(HB-co-HV) from 18 MPa to 8 MPa and diminished flexibility as Young's modulus increased from 1,525 MPa to 2,498 MPa, but overall mechanical properties of the polymer remained in a useful range. A mixed microbial culture required more than 20 days to degrade 150-microns-thick samples of 100% P(HB-co-HV), whereas samples containing 50% (wt/wt) starch disappeared in fewer than 8 days. Starch granules degraded before P(HB-co-HV) did. Aerobic degradation proceeded more rapidly than anaerobic degradation.

Aerobiosis↗

Comparative characterization of complete and truncated forms of Lactobacillus amylovorus alpha-amylase and role of the C-terminal direct repeats in raw-starch binding.

Two constructs derived from the alpha-amylase gene (amyA) of Lactobacillus amylovorus were expressed in Lactobacillus plantarum, and their expression products were purified, characterized, and compared. These products correspond to the complete (AmyA) and truncated (AmyADelta) forms of alpha-amylase; AmyADelta lacks the 66-kDa carboxyl-terminal direct-repeating-unit region. AmyA and AmyADelta exhibit similar amylase activities towards a range of soluble substrates (amylose, amylopectin and alpha-cyclodextrin, and soluble starch). The specific activities of the enzymes towards soluble starch are similar, but the K(M) and V(max) values of AmyADelta were slightly higher than those of AmyA, whereas the thermal stability of AmyADelta was lower than that of AmyA. In contrast to AmyA, AmyADelta is unable to bind to beta-cyclodextrin and is only weakly active towards glycogen. More striking is the fact that AmyADelta cannot bind or hydrolyze raw starch, demonstrating that the carboxyl-terminal repeating-unit domain of AmyA is required for raw-starch binding activity.

Bacterial Proteins↗

Display of alpha-amylase on the surface of Lactobacillus casei cells by use of the PgsA anchor protein, and production of lactic acid from starch.

We developed a new cell surface engineering system based on the PgsA anchor protein from Bacillus subtilis. In this system, the N terminus of the target protein was fused to the PgsA protein and the resulting fusion protein was expressed on the cell surface. Using this new system, we constructed a novel starch-degrading strain of Lactobacillus casei by genetically displaying alpha-amylase from the Streptococcus bovis strain 148 with a FLAG peptide tag (AmyAF). Localization of the PgsA-AmyA-FLAG fusion protein on the cell surface was confirmed by immunofluorescence microscopy and flow cytometric analysis. The lactic acid bacteria which displayed AmyAF showed significantly elevated hydrolytic activity toward soluble starch. By fermentation using AmyAF-displaying L. casei cells, 50 g/liter of soluble starch was reduced to 13.7 g/liter, and 21.8 g/liter of lactic acid was produced within about 24 h. The yield in terms of grams of lactic acid produced per gram of carbohydrate utilized was 0.60 g per g of carbohydrate consumed at 24 h. Since AmyA was immobilized on the cells, cells were recovered after fermentation and used repeatedly. During repeated utilization of cells, the lactic acid yield was improved to 0.81 g per g of carbohydrate consumed at 72 h. These results indicate that efficient simultaneous saccharification and fermentation from soluble starch to lactic acid were carried out by recombinant L. casei cells with cell surface display of AmyA.

Enzymes, Immobilized↗

Glucose absorption from starch hydrolysates in the human jejunum.

The intestinal absorption and mucosal hydrolysis of a partial and a complete alpha-amylase hydrolysate of corn starch, simulating the normal intermediary and end products of luminal starch digestion, was studied using an in vivo steady state jejunal perfusion technique in normal human subjects. Alpha-amylase was excluded from the test segment by proximal balloon occlusion. Products of hydrolysis during intestinal perfusion were identified using gel permeation chromatography. Three isocaloric, isotonic sugar saline solutions containing 140 mM glucose, 70 mM maltose and the partial amylase hydrolysate of starch (51.5 +/- 1.4% of glucose content comprising glucose polymers of more than 10 glucose units) were perfused in the first study. Net glucose absorption during perfusion of the partial hydrolysate and free glucose was similar, but significantly faster from maltose (p less than 0.05). Hydrolysis of the polymer fraction containing more than 10 glucose units was significantly slower (29.5 +/- 2.0% of infused load) than the lower molecular weight fraction (56.4 +/- 3.8%, p less than 0.001). As net glucose absorption from the partial hydrolysate was similar to that from glucose, despite the slow hydrolysis of the higher molecular weight fraction, it seemed likely that oligosaccharides in the more rapidly hydrolysed lower molecular weight fractions were exerting a kinetic advantage on glucose absorption. This was confirmed in a second study, where glucose absorption from a complete amylase hydrolysate consisting predominantly of maltose, maltotriose and alpha-limit dextrins, occurred significantly faster (81.8 +/- 4.8 mmol/h/25 cm) than from isocaloric free glucose (55.8 +/- 4.9 mmol/h/25 cm, p less than 0.001). Chromatograms of intestinal aspirates suggested that (1->4), but not 1->6) linked oligosaccharides liberated during luminal and brush-border hydrolysis of dietary starch conferred a kinetic advantage on glucose absorption.

Adult↗

Constancy of glucose and starch fermentations by two different human faecal microbial communities.

The fermentation of glucose and corn starch by faecal suspensions from two subjects was examined over a three and a half year period. The substrate specificity and products of the faecal fermentations of each subject were relatively stable during this period and were significantly different between subjects. The major soluble end products of fermentation of glucose or starch were acetate, propionate, and butyrate. Hydrogen temporarily accumulated and was subsequently used in fermentations by both subjects. Hydrogen was used without methane production in fermentations of subject 1, but was used for methane formation in fermentations of subject 2. Although the rates of glucose fermentation were similar between both subjects; subject 1 produced a significantly greater molar ratio of propionate than did subject 2. The rate of fermentation of starch by faecal suspensions from subject 1 was faster than that of subject 2. The molar ratio of butyrate was greater for starch fermentations by subject 2, while the molar ratio of propionate was greater with subject 1. Significant differences were found between subjects in molar ratios and concentration of acetate and propionate and concentrations of butyrate in faeces.

Acetates↗

Starch and talc emboli in drug addicts' lungs.

The lungs of eight drug addicts dying as a consequence of their habit have been examined. All showed the presence of small amounts of talc emboli and five the presence of starch emboli. Talc was invariably associated with a marked foreign body reaction which was insignificant in association with starch. Animal experiments showed very rapid (90% in 24 hours) removal of maize starch emboli; such rapid removal in man would explain the lack of a foreign body response. Quantitation of the amount of starch present in lungs from two of the cases gave values of 1.5 and 5.2 g. The higher amount could have been a contributory factor in the sudden death of the addict. The amounts of talc seen were not sufficient to be of clinical significance.

Acetaminophen↗

Genetic diversity of maize kernel starch-synthesis genes with SNAPs.

Measuring genetic diversity in populations of a crop species is very important for understanding the genetic structure of and subsequently improving the crop species by genetic manipulation. Single-nucleotide amplified polymorphisms (SNAPs) among and within maize populations of waxy, dent, and sweet corns at 25 single-nucleotide polymorphism (SNP) sites in 6 kernel starch-synthesis genes (sh2, bt2, su1, ae1, wx1, and sh1) were determined. Because of the intensive selection of some favorable alleles in starch-synthesis genes during the breeding process, and the resultant strong linkage disequilibrium (LD), the number of haplotypes in each population was far less than expected. Subsequent phenetic clustering analysis with the SNAPs indicated that the dent, waxy, and sweet corns formed distinct subclusters, except in a few incidences. LD was surveyed among SNAPs of intragenic, intergenic, and intrachromosomal SNPs in whole and subpopulations, which revealed that some SNAPs showed high LD with many other SNAPs, but some SNAPs showed low or no significant LD with others, depending on the subpopulation, indicating that these starch genes have undergone different selection in each subpopulation during the breeding process. Because the starch synthesis genes used in this study are important in maize breeding, the genetic diversity, LD, and accessions having rare SNAP alleles might be valuable in maize improvement programs.

Alleles↗

Relationship between fermentations and calcium in the cecum of rats fed digestible or resistant starch.

The present studies were undertaken to investigate the effects of the dietary calcium (Ca) level on the Ca balance and on large intestine physiology in rats fed diets providing carbohydrate as digestible wheat starch (DS) or resistant starch (RS). Resistant starch was a high-amylose maize starch. The Ca level ranged from a marginally sufficient level (3 g/kg) to a level in excess of nutritional requirements (6 g/kg). The cecal fermentations were more developed in rats fed RS diets and they were affected by the dietary Ca level, compared to rats fed the DS diets. In rats fed a low Ca RS diet, the cecal fermentations were more acidic than with a high Ca level and there was an accumulation of lactic acid together with depressed concentrations of short-chain fatty acids. There was a considerable accumulation of Ca and phosphate (Pi) in the cecum of rats fed the 6 g/kg level; on the other hand, the concentrations of minerals (Ca, Pi) were very low in the cecum of rats fed the low Ca RS diet. The proportion of soluble Ca in the cecum was markedly enhanced by active fermentations, especially in rats fed the high Ca RS diet (16.3 mmol/l). In rats fed the RS diet, there was a substantial absorption of Ca from the cecum, which was relatively proportional to the dietary Ca level. The above data suggest that in these rats, Ca absorption in the large intestine made a substantial contribution to the digestive balance, probably at the expense of absorption in the upper part of the intestine.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

The inhibitory effect of hydrolytic products of starch on in vitro colonization by Streptococcus mutans.

Synthetic culture medium containing potato starch and 1 gm% sucrose, with and without human parotid saliva, were pre-incubated. Filtrates from these mixtures were then incubated with Streptococcus mutans. S. mutans colonization on glass rods was suppressed in the cultures using filtrates from saliva and starch interaction. No inhibition was noted in filtrates pre-treated with parotid saliva in the absence of starch. The results suggest that the hydrolysis of starch by saliva yields by-products which inhibit in vitro S. mutans colonization.

Adhesiveness↗