CS273141B1 - Productive strain of microorganism "aspergillus niger" for citric acid's surface production - Google Patents

Abstract

The subject of the invention is a productive strain of Aspergillus niger micro-organism, the mutant CA-179 (CCM 8037), for citric acid surface production, which gives higher yield than existing strains. Citric acid produced from the new strain can be used in the food and pharmaceutical industry and in consumer chemistry.

Description

(57) The subject matter of the invention is a production strain of Aspergillus niger, a mutant CA-179 (CCM 8037) for the surface production of citric acid, which achieves higher yields than well known strains. Citric acid made from a new strain is useful in the food, pharmaceutical and consumer chemistry industries.

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The present invention relates to a novel industrial strain (mutant) of the fungus Aspergillus niger, suitable for industrial application in a surface process for the production of citric acid.

The essence of the invention is that the strain designated CS-179 (CCM 3037) achieves increased yields, i.e. more efficient sugar utilization for molasses mashing by surface treatment at normal and elevated mash height in the fermentation dish than primarily the U-47 strain used previously. (CCM F-749), protected by copyright certificate No. 245 107.

Aspergillus niger industrial strains for the production of citric acid are assessed primarily in terms of production effect 'expressed as yield (conversion in% of sugar used to citric acid) or production expressed as g / m2 / day. Another indicator of the suitability of the strain is the sufficient sporulation of the fungus grown on a suitable sporulation medium (used for this strain according to the author's certificate no. 196 162), which is a prerequisite for obtaining quality vaccine material. The suitability of the strain is also assessed by the intensity of the fermentation process in progress, the rate of germination and young mycelium formation, the quality of the mycelial blanket produced, its weight, consistency and resistance to infectious microorganisms, see author's certificate No. 221 482.

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The type of production strain is also characterized by the simultaneous formation of secondary acids, especially oxalic acid, whose formation is influenced by the quality of the processed molasses and the process of preparation of the fermentation medium.

An important factor influencing the yield of the fermentation process is the composition of the fermentation medium, which depends on the quality of the processed raw material, molasses and the actual technological process of preparation, ie the method of dosing the additives and additives in its preparation, their amount, temperature regime, time sequence and sterilization method. U.S. Patent No. 121,832, U.S. Patent No. 130,943, Author's Certificates No. 180,276, 245,107 and 233,664).

The new CS-179 strain was gradually verified in the laboratory and in the plant with respect to the above criteria and compared with the U-47 strain used on various types of molasses. It has fully proved and demonstrated an increase in production activity over the same fermentation cycle time. Oxalic acid production is about 1-2% higher than strain U-47.

The present invention provides a novel production strain of Aspergillus niger (mutant) CS-179 deposited in Cs. Collection of Microorganisms - University of J. E. Purkyně in Brno, tř. Defenders of Peace No. 10 under the designation CCM 8037.

Morphological and physiological characteristics

Aspergillus niger CS-179 mutant grown on glycerine sporulation medium (Author's Certificate No 196 162) of the following composition: 125 g glycerin, 45 g malt extract, 15 g NaCl, 0.5 g NH4NO3, 0.08 g MgSO4 aq 0.0015 g of CuSO4 aq and 30 g of agar per liter of soil, at 32 DEG C., for 36 to 40 hours, form round colonies with a diameter of 0.5 to 0.7 cm, forming sporulating condidiophores in the center of the colony. From day 4 of cultivation, the sporulating agar area in the cultivation vessel is completely covered with a continuous sporulating mycelium with a yellowish, slightly wrinkled surface. After 8 days of cultivation, the culture forms a dense carpet of mycelia with abundant conidia formation, which can be easily separated mechanically or by suction and preserved.

The strain growth on the wort agar and Czapka agar appears to have similar characteristics with a 12 to 20 hour delay, with less sporulation.

Microscopic appearance: conidia are brown-black to black in color, smooth, spherical in shape with a diameter of 3 to 4 yum, conidiophores 2 to 3 mm high, conidial head of ύ 0.1 mm, spherical dark with colored primary and secondary rich-forming sterigms conidia after 60-70 hours cultivation at 32 ° C.

Mutant CS-179 cultured on production molasses medium with 15% sugar and optimal addition of potassium hexacyanate phosphoric acid and addition of ZnSO4 aq for 24 hours creates a continuous thin mycelium that gradually strengthens, whites and remains smoothly richly wrinkled throughout the surface fermentation with minimal sporulation.

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The CS-179 mutant, in contrast to the current U-47 strain, after a 9-day fermentation cycle · surface fermentation with a height of 9 to 12 cm molasses substrate achieves the following results under normal conditions:

% yield U-47 CS-179 75 to 80% 82 to 85 % oxalic acid formation (% by weight of generated U-47 3 to 11 % citric acid) CS-179 4 to 13 %

The demonstrated differences in the physiological properties of strains U-47 and CS-179 are so significant that the operational utilization of the new strain by increased yield in terms of production yields significant benefits.

The new production strain CS-179 was obtained by mutagenic treatment with UV light on the parental strain AS-68, deposited in the collection of strains of Lachema Kázeňjov.

The mutation used a Philips 30 germicidal lamp, emitting monochromatic light with a wavelength of 253.7 nm. The conidia suspension of the parent strain was obtained by flushing the sporulating culture with a sterile 0.5% tritone solution and filtering through a sparse frit to remove clusters of spores and hyphae residues. The suspension is diluted to a concentration of about 10 .mu.m / ml. 5 ml of this suspension was poured onto a Petri dish (0 10 cm) and irradiated with a UV lamp from a distance of 25 cm for 5 to 25 minutes at normal temperature and a radiation intensity of about 4.5 3 / m 2 / sec. Monosporic cultures grown on sporulated agar broth were obtained from surviving colonies and evaluated for citric acid production by molasses soil fermentation tests. The best and most active mutant designated CS-179 was then selected.

Example

The fermentation brewing substrate was prepared in the process digester by measuring the amount of molasses to the water substrate so that the solution contained 35% by weight. of sugar. The mixture was heated to 85 ° C, sulfuric acid was added to adjust the pH in the range of 6-6.6, potassium ferrocyanide in an amount of 4-6%. wt. per sugar used, phosphoric acid 0.2 to 0.6%. wt. and the solution was heated to boiling for 60 minutes. At the end of boiling, the solution was topped up with biologically harmless cold water to a final 14 wt. of sugar. After the addition of the prescribed amounts of formic acid and zinc sulfate, the fermentation solution was filled into the fermentation chamber dishes. The substrate height was 9 cm, the fermentation time was 9 days, the fermentation solution temperature was 30-35 ° C. The fermentation chambers were inoculated alternately with strains U-47 and CS-179.

The results below are the mean of the groups of 10 to 20 operating chambers with both strains fermenting 3 molasses.

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Molasses Strain Yield% h QX / KC AND U-47 77.2 5.9 CS-179 82.3 7.5 (B) U-47 77.6 8.9 CS-179 83.2 9.2 C U-47 • 80.8 7.9 CS-179 82.0 8.8

SUBJECT OF THE INVENTION

Claims (1)

Aspergillus niger production strain, mutant CS-179 (CCM 8037) for surface production of citric acid.