Citric acid and Aspergillus niger interact to produce high levels of citric acid. Citric acid is the largest organic acid consumed in the world and is formed as a mediator in the Kreb cycle. But it accumulates in larger amounts in Aspergillus niger, possibly due to metabolic abnormalities. This aspect of the fungus is used to produce fermentation of citric acid. It is also found as a natural product in many fruits, especially citrus fruits. Before the development of fermentation technology, citric acid was extracted from the juice of these fruits.Citric acid can also be made from glycerin, but it is very expensive Production of citric acid began with surface fermentation in 1923. Citric acid produced in 1929 was 5,000 tons, which increased to 4.0 tons by 1992. 60% of the citric acid produced is used in the food and beverage industries. A flavoring and preservative, while 10% is used in the pharmaceutical industry in the form of iron citrate, about 25% of citric acid is used in the chemical industry.
Various fermentation processes used to produce Citric acid and Aspergillus niger For the following reasons, Aspergillus niger is used in most processes this is a Applications of citric acid.
1. Easily cultivated.
2. Processes uniform biochemical properties.
3. Produces only small amounts of oxalic acid.
4. Produces a lot of citric acid.
The surface cultivation process using solid culture medium and the underwater cultivation process using reactor are briefly described here: Surface cultivation process Although this is an old process, it is still used. It is a kind of constant fermentation process.
Spore suspension is used as an inoculum to produce Citric acid and Aspergillus niger. The suitable and high-yielding A. niger strain is selected from a stock crop. The stock culture is inoculated at the level of the sporulating agent in the glass bottles. The bottles are incubated for 25-14 days at 25 ° C The composition of trace elements such as manganese, zinc or iron salts in the spore medium must be properly maintained. Otherwise they affect the performance of citric acid in actual fermentation. Hog suspension is obtained by suspending spores grown in suitable diluents such as water containing a moisturizer, sodium lauryl sulfate. In addition to the total number, the survival of the spore crop is critical.
The environment used in the production of citric acid must be a source of carbohydrates and inorganic salts. A variety of materials can be used as a carbon source. But in general, molasses, sucrose and beets are used as a source of carbon Sucrose is the best source of carbon tested among various organic substances. An environment with less than 15% sucrose has been reported to have a high yield of citric acid. When part of the sucrose is replaced by fructose or glucose, a decrease in citric acid function is observed. Commercially, beet molasses is also widely used as a carbon source in the production of citric acid using A niger Beetroot molasses, in addition to sugars, contains large amounts of inorganic salt. To eliminate these excess inorganic salts, it is treated with ferrocyanide or ferric cyanide before use in the preparation of the medium. Alternatively, inorganic salts are removed by passing beet molasses through a cation exchange resin Elements such as nitrogen, potassium, phosphorus and magnesium are also needed in addition to the carbon source in the environment. They are added as ammonium nitrate, potassium dihydrogen phosphate or potassium monohydrogen phosphate and magnesium sulfate to a minimum in the medium The presence of these elements in higher concentrations reduces the yield of citric acid and increases the yield of oxalic acid The pH of the environment should be adjusted to 3.5 – 3.5 using hydrochloric acid. Low pH has been reported as the most desirable case. Because it facilitates less pollution, more citric acid formation, suppression of oxalic acid formation and easy sterilization of the environment.
The production medium is placed in shallow containers so that a thin layer of medium with a depth of 1 to 2.5 cm is formed. Inoculation spores are added to the medium to float on its surface. This is achieved by appropriate adjusting devices. Incubation is done in incubation rooms at a temperature of 30-40 ° C During fermentation the temperature is kept constant at 30 ° C. Airflow ventilation is also important for gas exchange because if CO 2 in the atmosphere increases to 10%, citric acid production decreases. Within 24 hours after inoculation, germinating spores form a thin layer of mycelium on the surface of the nutrient solution. As a result of the adsorption of ammonium ions, the pH in the culture fluid reaches 1.5 to 2.
The rate of bio-conversion of sugar to citric acid depends on the ratio of surface area to volume. If the ratio is lower, the yield of citric acid will be higher. In this method, the shallow pan has a lower surface-to-volume ratio, so a large area of mycelial mat is exposed to the shallow layer of the environment After 30 hours of fermentation, if the iron concentration is higher than oxalic acid and a yellowish pigment is formed, which in turn hinders the healing process. The fully developed mycelium floats in the form of a thick white layer floating on a liquid medium. Fermentation stops after 8-14 days Under these conditions, sugar is converted to citric acid. For this reason, this process is considered superior to the process of immersed culture. Yield per hour of this process reaches 1.2 – 1.5 kg of citric acid monohydrate per square meter of fermentation area.
Mycelium separates from fermented broth. Any intracellular citric acid in the mycelium is obtained by pressing the mycelium. The filtered broth is treated with calcium hydroxide. Filtered and washed. It is then treated with an equal volume of sulfuric acid to release citric acid. Calcium sulfate is formed in this process as a precipitate.The precipitate is removed by filtration. An impure solution of citric acid is obtained, which is decolorized by treatment with activated carbon and also mineralized Eventually pure crystals of citric acid are produced by evaporation. It is also recovered by alternative method with anti-current extraction method.
In this method, A. niger is made to be evenly distributed in the liquid production medium. Fermentation is generally done in large fermentation machines with a capacity of 4 kg per thousand gallons and is prepared with a mechanical stirrer and asparagus. Eighty percent of the world’s citric acid production is produced by this process. Production cost with this method is reduced by 25%. This includes lower labor costs, longer incubation periods, more energy consumption and more sophisticated techniques .Three factors are important for production in the process of immersion culture. They are the quality of the metal used to make fermentation, mycelial structure and oxygen delivery .Candida lipolytica , an alkane fungus can also be used in the production of citric acid under continuous fermentation. This product is 45% more than natural citric acid production.
Mycelium mats called pellets are used as inoculants for fermentation in this process. Suitable and high-yielding A. niger strains are selected from a stock culture. The spores in the seed fermenter cause germination. A nutrient solution containing 15% sugar from molasses is used in this seed fermenter. Cyanide ions are added to the medium to induce mycelium pellet formation.Pellet formation largely depends on the concentration of cyanide ions in the environment. Lower yield of citric acid occurs if cyanide ions are less concentrated.This is because lower concentrations of cyanide ions form natural mycelium instead of pellets. Spores germinate at 32 ° C and form pellets with a diameter of 0.2-0.5 mm within 24 hours. During this period the pH decreases to 4.3. These pellets are used as inoculants for production fermenters.
The medium used for the surface cultivation process is also used in this process.
Most fermentations used to produce citric acid are made in the range of 10 to 220 colts. They must be made of stainless steel to prevent heavy metals from leaching. Normal steel, if used in the manufacture of fermentation machines, at a low pH level of 1-2, may prevent the formation of citric acid.Small fermenters with a maximum capacity of 1000 liters, even if made of stainless steel. Due to the large surface / volume ratio, they should have a plastic cover. However, such a plastic coating is not required for large stainless steel fermenters.The mycelium pellets produced in the seed tank are transferred aseptically to the fermenter and incubated at a constant temperature of 30 ° C. The structure of the mycelium formed in the fermenter is critical to a successful production process. If the mycelium is loose, fibrous, with limited branches and lacks chlamydospores, little citric acid is produced.If the mycelium is in the form of pellets, optimal citric acid is produced. The ratio of iron to copper in the environment determines the nature of the mycelium.In some cases, production fermenters are inoculated directly with spores.Although A. niger requires relatively little oxygen, it is sensitive to hypoxia. During the fermentation process there should be a minimum oxygen concentration of 20 to 25% saturation. Short interruptions in oxygen supply stop production irreversibly. The aeration rate should be 0.2-1.0 volumes per minute. Stirring is not necessary during the acid production phase due to its low viscosity. Therefore, although some plants use agitators, air transfer reactors can also be used.Foaming is a problem in the process of immersion culture. However, it can be controlled by adding anti-foaming agents such as fat oil at frequent intervals. A floor chamber, 1/3 of the size of the fermentation volume, is required in both air lift and stirrers. Mechanical floor foam devices can also be used. The progress of the fermentation process is controlled by calculating the sugar and citric acid content in the regular fermentation.