CS262877B1 - Strain of microorganism aspergillus niger ccm 80045 - Google Patents

Abstract

The solution relates to a new strain of Aspergillus niger CCM 8004 that it produces on a simple defined medium acid gluconic high speed and degree conversion, with minimal unwanted content substances and pigments. Strain. has use in fermentation production of gluconic acid.

Description

The present invention relates to a novel strain of Aspergillus niger mutant N-XI which produces gluconic acid on a simple mineral medium with high production rate and selectivity, without significant amounts of colored substances, and exhibits increased resistance to gluconic acid salts in the fermentation medium.

The ability of the Aspergillus niger strain to produce gluconic acid has been known for over 40 years (Liil, L. L., Organic Acids, In Irimary Iroducts of I.etabolism, Academic Iress, kev / York 197 &). This property is utilized in the fermentative production of gluconic acid, its salts and glucono-T-lactone. the literature suggests that the conversion of glucose to gluconic acid takes place on a multi-component mineral medium with the addition of other nutrient components (corn steep liquor, yeast autolysate, etc.), which, together with a higher biomass concentration (3 to 5 grams per liter) the resulting product contains a significant amount of ballast substances (components of the production medium, substances released from biomass - pigments, secondary metabolites, etc.) and places higher economic demands on obtaining a highly pure product which is needed eg for pharmaceutical or food use, culture catalogs hub u nás (Catalog of Cultures, Charles University, Iraha 1276 ·, Catalog of Cultur - Yungi Czechoslovak collection of microorganisms,

YIPPEE. University Female, Jrno 1S & 2) disclose Aspergillus niger species without the ability to selectively produce gluconic acid. We do not have high-producing strains of fungi capable of highly selective production of gluconic acid.

- 2,282,877

This drawback removes the invention, which is a novel strain of Aspergillus niger mutant N-XI. The strain is deposited in the Czechoslovak Collection of Microorganisms' University J.E. Purkinje in Srn (Peace Defender Class) under the designation CCM 8004. The new strain was obtained by treating a water suspension of Aspergillus niger spores at a concentration of 10 spores per milliliter with the addition of 1% (w / v) Tween 80. radiation for 20 to 30 minutes. The spore suspension was then spilled on wort agar and cultured at 25-30 ° C. A spore suspension was prepared from the cultured culture and after suitable dilution was spilled onto wort agar. Individual grown colonies of the microorganism culture were evaluated by fermentation broths, both in terms of gluconic acid production and tolerance to high concentrations of gluconate salts and pigment formation. The Aspergillus niger eofcant h-kl strain thus obtained was passaged several times on a high glucose liquid medium and maintained in a conventional manner. Among the main advantages of such a selected. Aspergillus niger mutant strains h-kl include high · gluconic acid production rate - 1.2 to 1.6 / w / v utilized carbohydrate per hour at yields of 94 to 98 / theoretical, but biomass concentration compared to available literature data (3 to 5 g dry weight of mycelium / liter of medium) was substantially lower (0.5 to 1.0 g per liter of medium). These values are below 3 o cno (λιλοα ^ θρι defined mineral medium o assimilable carbon source and nutrient salts, which together with lower biomass concentration allows to obtain the desired product of high purity with a minimum content of ballast and pigmenting substances. capability even at high concentrations of glu282,877 konate salts in the fermentation medium (more than 430 kg / m @ 2).

The new Aspergillus niger mutant N-kl strain grows well on conventional culture media used to cultivate microscopic fungi (Ozapek-Dox agar, wort agar, glucose agar and yeast extract .apod.). Even at 28 ° C, colonies grow linearly on Czapek -Dox agar on average

4.3 cm in 72 hours, on wort agar (5 ° Bgj 3.1 cm. On Czapek-cox agar, colonies are flat, low, initially formed by white airy mycelium. Sporulation color from center from yellow to light to dark brown, on malt agar dark brown to black, colony is radially creased on the underside, yellow-black, abundant sporulation on the whole colony occurs at 6 to 7 don, at that time a fine secondary mycelium is formed on the colony. conidiophore is 1 to 2 mm high, most fructificone structures have conidiophore 1.5 mm high, conidiophore width is 1 Šířka, 5 µm in diameter, conidial head is 230yum in diameter, average conidia size is 4.2 / The viability of the spores in long-term storage in activated carbon suspension, in paraffin oil or in the lyophilized state is very good.

Aspergillus niger strain, a k-kl mutant, can be used for the highly efficient fermentation preparation of gluconic acid, glucone-6-lactone, glucone- ^17- lactone or gluconate salts, which have a wide range of uses (eg pharmaceutical, food, chemical industry) ). The invention is illustrated by the following non-limiting examples.

- 4 example 1

282 877

ITa water suspension of Aspergillus niger joints at a concentration of 10 ° spores / ml with the addition of 1 / (v / v) Tween oO is treated

-beams from a source of θ 30ο for 20 to 30 minutes. The suspension is poured onto the wort agar xotome and allowed to resuspend in a thermostate at 25430 ° C. Thus, in sporulated cultures, a spore suspension in 0.9% sodium chloride is prepared, which is diluted so that after spilling on wort agar, individual colonies of the microorganism culture are grown after 2 < 44 days and evaluated by fermentation tests for gluconic acid production. and increased resistance to salts of gluconic acid. Thus, the Aspergillus niger strain, the mutant h-kl, was selected.

Iřiklad 2

Prepare 2.5 L of culture medium containing 200 g of glucose, C, 39 g of ammonium hydrogen phosphate, C, 19 g of potassium dihydrogen phosphate and 0.16 g of magnesium sulfate heptahydrate per liter. the sterile culture medium in a 5 L fermenter is inoculated with spores with "

Aspergillus niger, mutant -.-hl to a spore concentration of 0.5-1.10 ° / ml and cultured at 30 ° C, with stirring at 350 rpm and aerating 1.2 volumes of air per volume of fermentation medium per minute. During fermentation, the pH of the medium is maintained at 6.5 by adding 48% (w / v) sodium hydroxide solution. Glucose was transformed in the production phase to gluconic acid at a rate of 1.3-1.4 g (w / v) glucose per hour, with a maximum biomass concentration of no more than 0.9 grams of dry calcium per liter of fermentation medium, after fermentation, it contains more than 233g of sodium gluconate per liter, which is 96 / theoretical. Total fermentation time is maximum 29 hours.

Example. 3

An isolated strain has the property

282 877 that its mycelium after separation from the fermentation medium can be used for the fermentation preparation of gluconic acid required for the growth of xnycelium. 170 g of this solution are added to a 5 liter fermentation vessel of 2.5 liter of this solution and mixed with mycelium several times in succession to save glucose time in 1 liter containing recovered fermentation medium after the formation or in Example 2. The fermentation conditions are straightforward.

2. The total fermentation time is 13 hours and sodium is obtained, which is 97 hours of theoretical value.

Claims (1)

Aspergillus niger strain CCM 8004, producing gluconic acid.