The method of obtaining citric acid

 

(57) Abstract:

The invention relates to the microbiological industry and relates to a technology of production of edible organic acids, namely citric acid, by the method of deep fermentation. The method involves hydrolysis of starch suspension by liquefaction of a starch slurry with a solids content 26-36 wt. % using enzyme preparation bacterial-amylase at the rate of 0.5 to 1.5 units of amylase activity per 1 g of dry substance of the starch, the addition of mineral salts and fermentation using Aspergillus niger. Before adding salt starch suspension is maintained at a gauge pressure of 0.1 MPa for 30 minutes as the source of starch-containing raw material using purified starch suspension is an intermediate in the manufacture of starch concentration of solids 36%. The technical result consists in expanding the resource base and environmental clean technologies. table 1.

The invention relates to the microbiological industry and relates to a technology of edible organic acids, namely citric acid, by the method of deep fermentation.

Currently, the most common type is otoki of this raw material, as well as poor level of ecological purity of process upon receipt of citric acid on the raw materials, the challenge is to replace the molasses other types of raw materials suitable for fermentation them in deep conditions. One of such raw materials meet the specified requirements is starch, which is aimed at obtaining citric acid is subjected to hydrolysis followed by fermentation with the fungus-cyclotourists Spergillus niger.

The closest in technical essence to the proposed method is a method for citric acid using a nutrient medium containing starch, namely, that a 15% suspension of starch is first subjected to hydrolysis using a-amylase in the amount of 0.3% by weight of dry substance starch at 70oC for 45 min to obtain hydrolysis with dextrose equivalent DE10, add mineral salts at pH 3.0, sterilized under pressure 0,98105Pa add methanol to the amount of 2.0 vol.%, after which the mixture is fermented with the fungus Aspergillus niger with obtaining the target product.

In this way, as well as in other of these methods as a source of raw materials used dry starch, the cat is levym materials and its application in the above methods reduces economic efficiency, and, therefore, is their disadvantage. In addition, this technology does not allow to achieve ecological purity of process in connection with the use of methanol.

The technical result of the invention is to increase the yield of citric acid, reducing costs for preparation of hydrolyzed starch, the expansion of raw materials and the creation of environmentally friendly technologies citric acid.

This technical result is achieved in that in a method of producing citric acid-based medium, containing hydrolyzed starch and mineral salts, including the hydrolysis of starch suspension enzyme preparation of bacterial-amylase at high temperatures, the addition of mineral salts to the hydrolyzed starch and the subsequent fermentation of a nutrient medium mushroom-kislotoobrazoutei Aspergillus niger, lead the hydrolysis of a starch suspension containing 26 - 36 wt.% dry substances, enzyme drug bacterial-amylase, taken in an amount of 0.5 - 1.5 units amylolytic capacity per 1 g of dry substance of the starch, and before adding mineral salts withstand starch suspension at a pressure of 0.1 MPa for 30 mGy starch suspension - an intermediate in the manufacture of starch concentration of solids 36 wt.% or dry the starch content of 88 wt. % of dry matter, and as enzyme preparation bacterial-amylase drug use, 1 g of which has amylolytic ability (A. C.) 1000 units.

The method is illustrated by the following examples:

Example 1.

a) Preparation of conidia producer

Conidia of the fungus - producer Asp. niger is weighed into a sterile tube at the rate of 400 mg / 100 cm3placed in the environment for soaking conidia of the following composition, g 1 DM3:

Sugar - 50

Potassium dihydrophosphate - 0,16

Ammonium nitrate - 2,5

Magnesium sulfate heptahydrate - 0,25

A suspension of conidia in the flask is placed on the rocking chair with the speed of 160 min-1and incubated for 5 to 6 hours at a temperature of 32oC.

b) Preparation of nutrient media for cultivation of seed mycelium

12.4 g of dry starch dissolved in a preheated 50oC tap water and the volume was adjusted to 200 cm3getting 5 wt.%-percent starch suspension (calculated on dry substance starch), with vigorous stirring, heat the starch suspension to 60oC and type 2 wt.%-s ' solution of the farm is the mass of the dry substance of the starch, or 0.5 units A. S. /g of dry substance of the starch, with constant stirring for 45 min, bring the temperature up to 85oC and is kept in this mode, 90 minutes

Then the hydrolyzed starch is maintained at a gauge pressure of 0.1 MPa for 30 min, cooled to 45 - 50oC and sterile contribute 5 cm3solution of ammonium nitrate concentration of 10 wt.%, 0.5 cm3solution of sulphate of magnesium heptahydrate concentration of 10 wt.%, 0,32 cm3solution of potassium dihydrophosphate concentration of 10 wt.%.

C) Preparation of culture medium for fermentation 90 g of dry starch dissolved in water heated to 50oC. the volume was adjusted to 300 cm3getting 26 wt.%-percent starch suspension (calculated on dry substance starch), which is heated with vigorous stirring to 60oC and type 2 wt.%-s ' solution of enzyme preparation of bacterial amylase in the number of 1.95 cm3that corresponds to 0.05 wt.% of the drug to the mass of dry substance starch or 0.5 units A. S./g of dry substance of the starch, with constant stirring for 45 min, bring the temperature up to 85oC and kept at this temperature for 90 minutes To stop the enzyme action of the drug hydrolyzate is heated daywalt at a gauge pressure of 0.1 MPa for 30 min, cool to 45 - 50oC and sterile contribute 12.5 cm3solution of ammonium nitrate concentration of 10 wt.%, 1.25 cm3solution of magnesium sulfate heptahydrate concentration of 10 wt.%, 0.8 cm3solution of potassium dihydrophosphate concentration of 10 wt. %, and diluted with sterile water to a concentration of fermented sugar 15010 g/DM3.

g) seed Growing mycelium

In a flask with a capacity of 750 cm3placed 50 cm3the nutrient medium and seeded on 10 cm3suspensions of conidia. The flask is placed on the rocking chair with the speed of 160 min-1and incubated for 48 hours at a temperature of 32oC.

d) Fermentation of a nutrient medium on the basis of hydrolyzed starch citric acid

In a flask with a capacity of 750 cm3placed 50 cm3the nutrient medium and seeded with its 10 cm3rising mycelium. The flask is placed on the rocking chair with the speed of 160 min-1and incubated for 6 days at a temperature of 32oC. After fermentation the culture of the fungus inactivate boiling and determine the technical and economic performance of the process.

The data are summarized in table.

Example 2.

a) Preparation of conidia producer, seed growing mycelium, Fermentas who have purified starch suspension, containing 36 wt.% dry substance starch.

b) Preparation of nutrient media for cultivation of seed mycelium

32 cm3purified starch suspension - semi-product of starch production bring tap water to a volume of 200 cm3getting to 5.8 wt.%-percent starch suspension. Further operations are performed according to example 1.

C) Preparation of culture medium for fermentation 250 cm3purified starch suspension - semi-product of the manufacture of starch is heated to 60oC under vigorous stirring enter 2 wt.%-s ' solution of enzyme preparation bacterial-amylase in the amount of 2.25 cm3that corresponds to 0.05 wt. % drug by weight dry substance of the starch, or 0.5 units A. S./g of dry substance starch. Further operations according to example 1.

The data are summarized in table.

Example 3.

a) Preparation of conidia producer, seed growing mycelium fermentation of a nutrient medium is conducted similarly to example 1.

b) Preparation of nutrient media for cultivation of seed mycelium

12.0 g of dry starch dissolved in a preheated 50oC tap water and the volume was adjusted to 200 cmZiya 60oC and type 2 wt.%-s ' solution of enzyme preparation bacterial-amylase in the amount of 0.8 cm3that corresponds to 0.15 wt.% of the drug to the mass of dry substance starch or 1.5 units A. S./g of dry substance starch.

Further operations are performed similarly to example 1.

C) Preparation of culture medium for fermentation 90 g of dry starch dissolved in water heated to 50oC, the volume was adjusted to 300 cm3getting 26 wt. % starch suspension with vigorous stirring and heated to 60oC enter 2 wt.%-s ' solution of enzyme preparation bacterial-amylase in the amount of 6.0 cm3that corresponds to 0.15 wt.% of the drug to the mass of dry substance starch or 1.5 units A. S./g of dry substance starch.

Further operations are performed similarly to example 1.

Example 4 (prototype)

a) Preparation of conidia producer, used in examples 1 - 3 similarly to example 1.

b) Preparation of hydrolyzed starch

150 g of dry starch dissolved in a preheated 50oC tap water and the volume was adjusted to 1000 cm3under vigorous stirring injected with 0.3% by weight of dry substance starch enzyme PR is Then the mixture is heated to 70oC and incubated with stirring for 45 minutes

Further, the hydrolyzate is heated to boiling and boiled for 10 min to inactivate the enzyme.

C) Preparation of culture medium for fermentation

The hydrolysate is cooled to a temperature of 30oC, bring 2 n sulfuric acid solution the pH value to 3.0 and add mineral salts ammonium nitrate and 1.5 g/DM3dihydrophosphate potassium - 0.15 g/cm3the magnesium sulfate heptahydrate and 0.2 g/DM3the ions of trivalent iron - 0.005 g/DM3then the nutrient solution is maintained at a gauge pressure of 0.1 MPa for 20 minutes

After cooling medium contribute to 2.0 vol.% methanol and the mixture is fermented similarly to example 1.

The technical result of the experiments presented in the examples 1 to 3 of the table, compared with the prototype consists in increasing the yield of citric acid to 81.6 - 87,2 wt.% and received the favorable conditions of the hydrolysis of starch suspension at a temperature of 85oC, the excess pressure of 0.1 MPa for 30 min, a smaller dose of enzymatic drug-amylase 0.5 - 1.5 units A. S./g of dry substance starch.

Increasing the dose of the drug-amylase greater than 1.5 units A. S./g of dry matter achieved in example 3.

Dose reduction of drug-amylase less than 0.5 units A. S./g of dry substance starch hydrolysis starch suspension containing 36 wt.% dry substances, also will not allow you to get the achieved yield of citric acid as in examples 1 - 3.

The increase in the concentration of dry substance of the starch in suspension above 36 wt.% leads to difficulties of mixing during the hydrolysis, the deterioration of the conditions of hydrolysis and reduction of the technical result.

Another technical result of the invention, namely the reduction of costs in the preparation stage of hydrolysis achieved in comparison with the prototype through the use of hydrolysis of a starch suspension containing 26 - 36 wt.% dry substance starch. In this case, for the preparation of the hydrolysis step requires the same capacity machines and the same labor, and by a known method, but increased by 1.7 - 2.5 times the amount of dry substance of the starch, which is processed in one process cycle.

The decrease in the concentration of dry substance of the starch in suspension is less than 26 wt. % leads to an undesirable increase in costs training hydrolysis step.

The application of this method avlyn the purity of the whole process, that also is a significant advantage.

The proposed method of producing citric acid as a feedstock is distributed as 25 wt.%-ing on dry substance starch suspension and purified starch suspension - intermediate starch syrup production, which leads to expansion of raw materials production of citric acid and at the same time is more effective than using a dry starch, such as, for example, in a known way, as it refers to cheaper raw materials and does not require additional preparation of the starch suspension. Along with this, you receive the possibility of organizing a workshop citric acid directly in the production of starch, which will greatly simplify and cheapen the process of obtaining citric acid due to the absence of problems associated with transportation of raw materials.

References

1. Patent N 125986 Poland, MKI 3 12 R 7/48. Sposob wytwarzania kwasu cytronowego na podlozu zavlerjacym skrobie/ Bolach E. W. Lesniak, J. Ziobrowski - N 215688 Declared 17.05.79, publ. 30.06.85.

A method of obtaining a citric acid-based medium, containing hydrolyzed starch and mineral salts, including the hydrolysis of starch to suspensibility starch and subsequent fermentation of a nutrient medium mushroom-kislotoobrazoutei Aspergillus niger, characterized in that conduct the hydrolysis of a starch suspension containing 26 - 36 wt.% dry substances, enzyme drug-amylase, taken in an amount of 0.5 - 1.5 units amylolytic capacity per 1 g of dry substance of the starch, and before adding mineral salts starch suspension is kept under positive pressure of 0.1 MPa for 30 minutes

 

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