An improved FIA-system for measuring ?-amylase in cultivation media

Carlsen, Morten; Marcher, J.; Nielsen, Jens

doi:10.1007/bf00222839

Cited by 16 publications

(8 citation statements)

References 3 publications

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“…This is supported by the glucose-limited chemostat experiments where the specific a-amylase productivity was about two times higher than during the exponential growth phase where the glucose concentration was high. In the glucoselimited chemostat, the residual glucose concentration was about 2 mg 1-1 and from other chemostat experiments it has been found that glucose concentrations below 5 mg 1-1 do not repress a-amylase production (Carlsen, 1994). Since there was no repression of glucose in the chemostat experiment, it is obvious that there was induction by maltose since the specific productivity increased more than 60% upon changing to a maltose medium in the chemostat.…”

Section: Discussionmentioning

confidence: 80%

“…The third strain, CF2.1, is a morphological mutant of CF1.l that was isolated after treatment of CF1.l spores with nitrosoguanidine (NTG). Conidia from 6-8-d-old cultures (Carlsen, 1994) were inoculated into a defined batch medium containing (1-' ): 25.0 g glucose (monohydrate), 7-3 g (NH,),SO,, 1.5 g KH,PO,, 1.0 g MgSO,. 7H20, 1.0 g NaC1, 0.1 g CaC1,.2H20, 0.5 ml trace metal solution (14.3 g ZnSO,.…”

Section: Methodsmentioning

confidence: 99%

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Induction and repression of -amylase production in batch and continuous cultures of Aspergillus oryzae

1995

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have been measured during batch culture of a wild-type strain and two recombinant strains. The mean intracellular level for the two recombinant strains was about four to five times the level of the wild-type strain. The recombinant strains also had a higher a-amylase productivity, whereas the residence time of the intracellular a-amylase pool was approximately the same for the three strains. At high glucose concentrations there was a low constitutive synthesis of a-amylase, whereas at low glucose concentrations derepression resulted in an increased production rate. Shifts from a glucose-to a maltose-limited chemostat showed that maltose induces both the production and secretion of a-amylase. Finally, from immunoblots, botha glycosylated and an unglycosylated a-amylase have been detected.

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Section: Discussionmentioning

confidence: 80%

Section: Methodsmentioning

confidence: 99%

Induction and repression of -amylase production in batch and continuous cultures of Aspergillus oryzae

1995

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“…α-amylase The production of starch degrading enzymes on agar-solidified medium was detected by staining the surface with iodine solution 49 to visualize zones in which starch had been broken down. Amylase activity by mutants exhibiting the largest hydrolysis haloes were then tested in liquid cultures.…”

Section: Methodsmentioning

confidence: 99%

Random and direct mutagenesis to enhance protein secretion inAshbya gossypii

et al. 2013

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To improve the general secretion ability of the biotechnologically relevant fungus Ashbya gossypii, random mutagenesis with ethyl methane sulfonate (EMS) was performed. The selection and screening strategy followed revealed mutants with improved secretion of heterologous Trichoderma reesei endoglucanase I (EGI), native α-amylase and/or native β-glucosidase. One mutant, S436, presented 1.4- to 2-fold increases in all extracellular enzymatic activities measured, when compared with the parent strain, pointing to a global improvement in protein secretion. Three other mutants exhibited 2- to 3-fold improvements in only one (S397, B390) or two (S466) of the measured activities. A targeted genetic approach was also followed. Two homologs of the Saccharomyces cerevisiae GAS1, AgGAS1A (AGL351W) and AgGAS1B (AGL352W), were deleted from the A. gossypii genome. For both copies deletion, a new antibiotic marker cassette conferring resistance to phleomycin, BLE3, was constructed. GAS1 encodes an β-1,3-glucanosyltransglycosylase involved in cell wall assembly. Higher permeability of the cell wall was expected to increase the protein secretion capacity. However, total protein secreted to culture supernatants and secreted EGI activity did not increase in the Aggas1AΔ mutants. Deletion of the AgGAS1B copy affected cellular morphology and resulted in severe retardation of growth, similarly to what has been reported for GAS1-defficient yeast. Thus, secretion could not be tested in these mutants.

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“…The classical analysis of cr-amylase is based on measuring the degradation of starch either as the amount of reducing sugars produced or from the amount of starch which is hydrolysed. This method is simple, sensitive and gives reproducible results (Carslen et al, 1994). In the commercial Phadebas method, the release of chromophore groups from a starch derivative cross-linked to a matrix is monitored.…”

Section: Introductionmentioning

confidence: 99%

“…The analytical method based on measuring reducing sugars is not satisfactory for measuring cr-amylase in culture medium containing glucose. Several cr-amylase analysers based on this method have been reported (Carslen et al, 1994, Hansen, 1984. This method is a reliable method but is laborious and expensive (Marciniak and Kula, 1982).…”

Section: Introductionmentioning

confidence: 99%

Measurement of ?-amylase activity by Sequential Injection Analysis

1995

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A Sequential Injection Analysis @IA) system for monitoring cr-amylase activity is described. The SIA analyser is a further development of previously investigated Flow Injection Analysis (FIA) analyser. The analysis of a-amylase activity is based on monitoring the decoloration of an iodine-starch complex. Performances of the SIA analyser have been compared with the FIA analyser. A good agreement has been obtained between the SIA measurements and the FIA measurements.

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An improved FIA-system for measuring ?-amylase in cultivation media

Cited by 16 publications

References 3 publications

Induction and repression of -amylase production in batch and continuous cultures of Aspergillus oryzae

Induction and repression of -amylase production in batch and continuous cultures of Aspergillus oryzae

Random and direct mutagenesis to enhance protein secretion inAshbya gossypii

Measurement of ?-amylase activity by Sequential Injection Analysis

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