CS245107B1 - New strain of Aspergillus niger CCM-F-749 - Google Patents

Abstract

The solution concerns an industrial strain (mutant) of the fungus species Aspergillus niger van Thieghem CCM-F-749 with high biochemical activity, which is capable of fermenting molasses substrates prepared by conventional methods with a citric acid yield that exceeds the citric acid production when using strains used so far. In addition, it is capable of fermenting higher layers and concentrations of fermentation medium with undiminished activity, and a finished product of higher quality and purity can be obtained from the produced fermentation liquors obtained from the fermentation process using the new strain. The production mycelium of the new strain shows higher resistance to attack by the infectious parasitic fungus of the genus Penicillium in industrial application.

Description

243107

BACKGROUND OF THE INVENTION The present invention relates to an industrial strain (mutant) of a fungus of the species Aspergillusniger van Thieghem with high biochemical activity, which is able to ferment copper substrates prepared by conventional procedures with a yield of citric acid, which exceeds the production of citric acid by use of the strains used so far. In addition, with the lowered activity of fermentation of higher layers and the concentration of fermentation media and of the fermentation liquors produced from the fermentation process using a new strain, a finished product of higher quality and purity can be obtained. The new strain strain mycelium appears to be more resistant to industrial infestation by Penicillium infectious parasitic fungus.

The new strain produces, in addition to citric acid, a small amount of oxalic acid in the range of 0-15% by weight. to the citric acid formed.

By the formation and composition of the fermentation medium, the byproduct formation can be influenced by limiting the formation of citric acid to the minimum necessary, possibly completely suppressing it.

Aspergillus niger strains, which are used in the production of citric acid by fermentation, are mainly evaluated for their productivity, i.e., the intensity of conversion of the incorporated sugar component of the medium to the desired metabolite - citric acid. Another prerequisite for the successful utilization of the selected strain is the possibility of obtaining the requisite amount of inoculum material, that is, sufficient production of conidia in the cultivation of the strain suitable for sporulating soil.

Among the industrially used strains are notable differences not only in the intensity of ongoing biological processes (germination rate, formation and character of mycelial matter, metabolic conversion of sugar component, formation of side acids), but also productivity of strains is significantly influenced by the properties and composition of the production media, particularly in strains used for the fermentation of molasses sugar.

It is known that in the molasses they have a considerable variation in their suitability for preparing the production medium of citric acid, due to their composition and related to the different content of stimulants or citric acid inhibitors. Also decisive is the process for preparing the fermentation medium, whereby a suitable process of operation guarantees maximum production of citric acid, corresponding to the possibilities of the strain used.

In general, strains which, in addition to citric acid, also form secondary acids (oxalic, gluconic), are less suitable and may cause difficulties in further processing of the fermented lye to the finished product. However, the criterion of citric acid production is critical to the suitability of the strain.

If one of the foreign acid strains exceeds the strain of the other, which does not form the side acid but the amount and yield is lower, the manufacturer chooses, in principle, a strain guaranteeing a higher production of citric acid and an undesirable foreign acid content through the modified elimine-technology. in a fermented substrate during processing.

The best production strain used up to now is 60 to 70 percent by weight on molasses processed by the process technology. citric acid (based on the quality of the molasses being processed), calculated on the carbonated sugar. By prolonging the fermentation time, it is not possible to efficiently ferment a higher layer of fermentation without a significant reduction in yield of more than 9 cm.

The new production line U 47 was obtained by mutagenic action of UV light on the parental strain H 16. The obtained mutant N 47 differs from the parental H 16 strain in the following physiological properties: a)% yield of citric acid from experiments with molasses substrate (scattering yields by molasses type): H 16 60 to 70 wt. KC on seeded sugar 47 85 to 90 wt. KC on seeded sugar b) Oxalic acid formation (% by weight citric acid): H16 15-20% by weight. U 47 3 to 8 wt. c) mycelial mass without significant difference, the U 47 mycelium shows high resistance to Penicillium infectious fungus; - Difficulty of citric acid and suppression of more oxalic acid formation to acceptable levels.

For the cultivation of the U 47 strain, the glycerine sporulating soil of the following composition proved to be successful: 125 g glycerin, 45 g malt extract, 15 g over, 0.5 g NH 4 NO 3, 0.06 g MgSO 4 aq, 0.0015 CuSCU aq and 30 g agar per liter After 8 days of growth on this soil, at a culture temperature of 32 ° C, strain U 47 produces a continuous growth rate of culture with a conidiogram length of 2 to 3 mm, a conidial head of medium size, a dark color. Primary sterigms of 20 to 30 μΐη are formed all around the circumference, and from the third day, conidia chains are formed on short (6 to 10 μm) secondary sterigamers, which are 3–4 μého in diameter, smooth in shape. dark brown to black.

After 5 days of cultivation (32 ° C) on the same soil, giant colonies with a diameter of 60 to 65 mm with a non-spacing edge zone of 3-5 mm are formed. 243107

Under the same conditions, the parental strain H16 produces longer conidiophores of 3 to 4 mm, the primary and secondary sterigms are uneven against U 47, the conidia chain development of 3 to 4 µm is richer, the conidia color is dark brown. Older cultures (from day 8), unlike U 47, tend to produce sterile secondary hyphae (graying culture).

The giant colonies of H16 reach visual and dimensional U 47o parameters 20 to 24 hours earlier.

Strain U 47 does not require special fermentation molasses medium and commonly used techniques of preparation (2, 3, 4, 5) can be used for its application. For the U 47 strain, the sporulating glycerine soil used for the current L-1 production strain has been proven. The generally used practice is to compare the new strain obtained either with the parenteral strain from which it was obtained, for comparison with the currently used strain in production, it is preferable to operate and use it.

The new U 47 strain outperforms citric acid production with both the current L-1 (10-15% w / w) and the H16 parental while reducing the more oxalic acid formation. U 47 is given in Table 1 above.

Strain

Laboratory fermentation conditions oxalic acid / KC yield / citric acid (% w / w) (% w / w)

Fermentation operating conditions oxalic acid / KC yield / citric acid (% w / w) L-l 70.2 0 H-16 65.0 6.4 U 47 89.3 2.7

The amount of oxalic acid formed can be influenced by the composition of the fermentation medium, for example by changing the dose of phosphoric acid, potassium hexacyanoferrate or by changing the process of preparing the molasses medium. The strain relies reliably in the sub-substrate 6 to 7 and the wide range of potassium ferrocyanide doses. .

The new strain is able to ferment the molasses substrate containing 14 wt. sugar 68.3 0 61.9 17.1 82.8 8.1 at a height higher than the usual 9 cm, with higher efficiency than the currently used strain L-1. In addition to this ability, it appears to be more suitable for fermentation of substrates with a higher sugar concentration than the usual 14% by weight.

The property of the new strain during the same fermentation period (9 days) of fermentation of a higher layer of molasses substrate results from the results given in Table 2. a) Citric acid yield in% by weight Substrate height (cm) 9 10 11 12 strain L -menene U 47 63,6 61,2 57,0 75,1 74,2 68,0 55,5 66,0 b) amount of citric acid formed (g / m per day) substrate height (cm) 9 strain Ll 953 strain U 47 1126

On the sporulation soil (4), the new production strain U 47 grows at 32-35 ° C very well and after 8 to 9 days of cultivation produces a dense carpet of sporulating mycelias rich in conidia formation, which can be separated by easy mechanical scraping or aspiration. The conidia can be preserved either in sterile sand or in a washed sterile carborafine. In the course of growth, to the early cultivation of still non-sporulating young washes- 10 11 12 1 023 1 053 1 110 1 240 1 257 1 320 lium is colored with a yellow pigment, which is also secreted into the cultivation soil.

The conidiophore strains are 2 to 3 mm long, conidial heads medium-sized, black. Round vesicles carry a radius of 20 to 30 microns in length across the entire circumference, and chain-horse chains are formed on shorter secondary sterigms (6 to 10 microns). which are spherical, smooth, dark brown to black, 3 to 4 ftm in diameter

Claims (1)

245107 A new production strain of CCM-F-749 was obtained by mutagenic action of UV light at a dose of 20,000 arg / mm 2 on spores of the parental strain. After selection and production assays of monosporic cultures, the most active mutant was obtained and designated as strain U 47. The novel strain Asperglllus niger van Thieghem CCM-F-749 suitable for the industrial production of citric acid. Severografla, n. P., Závod 7, Most Price 2,40 Kčs