Separation of starch oligosaccharides by high temperature paper chromatography

French, Dexter; Mancusi, Joseph L.; Abdullah, Mukhtar; Brammer, Gary L.

doi:10.1016/s0021-9673(01)99480-4

Cited by 41 publications

(9 citation statements)

References 9 publications

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“…The standard contained a mixture of oligosaccharides-glucose (G1) to G,5. After development by the technique of French et al (6) and Robyt and French (19), the chromatogram was photographed on an X-ray viewer. A-G1 through G15 standard; B through J (incubation time and per cent conversion, calculated as maltose)-B, 0.5 hr and 6%; C, I hr and 10%; D, 2 hr and 17%; E, 5 hr and 32%; F, 12 hr and 51%; G, 19 hr and 60%; H, 35 hr and 69%; 1, 75 hr and 77%; J, 97 hr.…”

Section: Resultsmentioning

confidence: 99%

Extracellular Transglucosylase and α-Amylase of Streptococcus equinus

Boyer

Hartman

1971

J Bacteriol

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Culture filtrates of Streptococcus equinus 1091 contained a-amylase and transglucosylase. The effects of calcium carbonate, age of inoculum, concentration of maltose, and duration of the fermentation on a-amylase and transglucosylase production were determined. The extracellular a-amylase was purified 48-fold and was free of transglucosylase activity. The a-amylase (amylose substrate) required Clfor maximum activity; ethylenediaminetetraacetic acid (EDTA) partially inhibited activity, but CaCl2 prevented EDTA inhibition. The temperature optimum was 38 C at pH 7.0, and the pH optimum was 7.0 at 37 C in the presence of CaCl2. Predominant final products of amylose hydrolysis, in order of decreasing prevalence, were maltose, maltotriose, maltotetraose, and glucose. The a-amylase showed no evidence of multiple attack. The extracellular transglucosylase was purified 27-fold, but a small amount of a-amylase remained. Transglucosylase activity (amylose substrate) was not increased in the presence of CaCl2. The temperature optimum was 37 C at pH 6.5, and the pH optimum was 6.0 at 37 C. Carbohydrates that served as acceptors for the transglucosylase to degrade amylose were, in order of decreasing acceptor efficiency: D-glucose, D-mannose, L-sorbose, maltose, sucrose, and trehalose. The extracellular transglucosylase of S. equinus 1091 synthesized higher maltodextrins in the medium when the cells were grown in the presence of maltose.The action patterns and most of the other properties of the extracellular amylolytic enzymes of Streptococcus equinus have not been determined. Seeley and Dain (22) noted that, although S. equinus hydrolyzes starch on agar plates containing D-glucose, acid is not formed from starch in broth. They concluded that, since S. equinus ferments maltose, starch hydrolysis probably results in a nonfermentable intermediate. Dunican (L. K. Dunican, M.S. Thesis, Cornell University, 1960) found that, when S. equinus was grown in a medium containing 1% tryptone, 0.5% yeast extract, 0.2% K2HPO,, and S. equinus and S. bovis both possess group D antigen, but they are differentiated from the enterococci by several physiological characteristics. Smith and Shattock (25) studied the fermentation patterns of S. equinus and S. bovis extensively and suggested retention of the two species. 561 on August 5, 2020 by guest http://jb.asm.org/ Downloaded from 562

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Section: Resultsmentioning

confidence: 99%

Extracellular Transglucosylase and α-Amylase of Streptococcus equinus

Boyer

Hartman

1971

J Bacteriol

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“…c Reducing-value (RV) method: 10-min assays at pH 6 per minute. Amylase activities (0.9 Mg ofprotein per ml, 6.1 U of amylase activity per ml) were measured by using the reducing-value method with 10-min assays at 43°C in 0.067 M Sorensen phosphate buffer.…”

Section: Discussionmentioning

confidence: 99%

Purification and some properties of an extracellular alpha-amylase from Bacteroides amylophilus

McWethy¹,

Hartman²

1977

J Bacteriol

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A medium was developed to obtain maximum yields of extracellular amylase from Bacteroides amylophilus 70. Crude enzyme preparations, obtained by ammonium sulfate precipitation of cell-free broth, contained six amylolytic isoenzymes that were detected by isoelectric focusing and polyacrylamide gel electrophoresis. One of these amylases was purified by diethylaminoethyl-Sephadex A-50 ion-exchange chromatography and Sephadex G-200 gel filtration techniques. Some properties of the purified extracellular a-amylase were: optimum pH, 6.3; optimum temperature, 43°C; pH stability range, 5.8 to 7.5; isoelectric point, pH 4.6; molecular weight, 92,000 (by sodium dodecyl sulfatedisc gel electrophoresis); and sugars causing inhibition, cyclomaltoheptaose, cyclomaltohexaose, and aD -phenylglucoside. In addition, Ca2+ and Co2+ were strong activators, and Hg2+ was a strong inhibitor; all other cations were slightly stimulatory. Dialysis against 0.01 M ethylenediaminetetraacetic acid caused a 58% loss of activity that was restored to 92% of the original by the addition of 0.04 M Ca2+. The enzyme affected a blue-value-reducing-value curve characteristic of alpha-type amylases. The relative rates of hydrolysis of amylose, soluble starch, amylopectin, and dextrin were 100, 97, 92, and 60%, respectively; Michaelis constants for these substrates were 18.2, 18.7, 18.2, and 16.7 ,umol of D-glucosidic bond/liter, respectively. The enzyme degraded maize (corn) starch granules to some extent and had relatively little activity on potato starch granules. Digestion of starch in the rumen is essential to maximum utilization of feed grains by the ruminant. To gain an understanding of starch digestion in the rumen, attempts have been made to purify and characterize rumen amylases. At present, however, only three rumen amylases, produced by Streptococcus bovis (10, 33, 35), Clostridium butyricum (10, 35), and Bacteroides amylophilus (S. S. Rahman, Ph.D. thesis, University of British Columbia, Vancouver, 1970, Diss. Abstr. 32:77), have been partially characterized. B. amylophilus produces extracellular amylases and may play an important role as a starch digester in the rumen (9). This report presents data on the production, purification, and characterization of this amylase and defines its possible role in rumen starch breakdown. MATERIALS AND METHODS Organism and cultural conditions. The organism used in this study was B. amylophilus 70, kindly ' Joumal paper no. J-8471 of the Iowa Agriculture and

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“…Enzyme with radiolabelled and nonradiolabelled substrate was incubated at 40°C and pH 6.0 (Universal buffer, 0.1 M). The saccharide components of these digests were separated and purified by ascending paper chromatography at 85°C (French et al 1965). Saccharides were visualised by aniline phthalate spray reagent and the relevant spots cut from the paper.…”

Section: Preparation Of Reducing-end Radiolabelled Maltoolandosaccharidesmentioning

confidence: 99%

The high maltose-producing α-amylase of the thermophilic actinomycete, Thermomonospora curvata

Collins

Kelly

Fogarty

et al. 1993

Appl Microbiol Biotechnol

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The alpha-amylase of Thermomonospora curvata catalyses the formation of very high levels of maltose from starch (73%, w/w) without the attendant production of glucose. The enzyme was produced extracellularly in high yield during batch fermentation in a 5-1 fermentor. Purification was achieved by ammonium sulphate fractionation, Superose-12 gel filtration and DEAE-Sephacel ion-exchange chromatography. The enzyme exhibited maxima for activity at pH 6.0 and 65 degrees C, had a relative molecular mass of 60,900-62,000 and an isoelectric point at 6.2. The exceptionally high levels of maltose produced and the unique action pattern exhibited on starch and related substrates indicate a very unusual maltogenic system. The predominance of maltose as the final end-product may be explained by the participation of reactions other than simple hydrolysis and the preferential cleavage of maltotriose from higher maltooligosaccharides. The enzyme exhibits very low affinity for maltotriose (Km = 7.7 x 10(-3) M) and its conversion to maltose is achieved by synthetic followed by hydrolytic events, which result in the very high levels of maltose observed and preclude glucose formation. This system is distinguished from other very high maltose-producing amylases by virtue of its high temperature maximum, very low affinity for maltotriose and the absence of glucose in the final saccharide mixture.

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Separation of starch oligosaccharides by high temperature paper chromatography

Cited by 41 publications

References 9 publications

Extracellular Transglucosylase and α-Amylase of Streptococcus equinus

Extracellular Transglucosylase and α-Amylase of Streptococcus equinus

Purification and some properties of an extracellular alpha-amylase from Bacteroides amylophilus

The high maltose-producing α-amylase of the thermophilic actinomycete, Thermomonospora curvata

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