Vitamin feed supplement for fish based on chlorella vulgaris

. The obtained data on the cultivation of chlorella in a nutrient medium containing macro and microelements based on local raw materials practically corresponds in terms of efficiency to the traditional Tamiya nutrient medium. Thus, innovative technology has been developed for obtaining an inexpensive, highly effective vitamin feed additive based on Chlorella vulgaris to replace artificial vitamins in fish feeding. The resulting water-soluble complex of macro and microelements makes it possible to completely replace expensive chemical elements in the cultivation of Chlorella vulgaris . Thanks to this, you can get an inexpensive vitamin-protein supplement for feeding fish. The presence of a full complex of natural vitamins in Chlorella vulgaris makes it possible to abandon expensive artificial vitamins when feeding fish. The effectiveness of which is only 5-10%, when natural vitamins are absorbed by 100%. All this reduces the use of expensive premixes that are added to fish feed.


Introduction
Fish, like all living organisms, need a full range of vitamin supplements [1]. Vitamin deficiency occurs in fish if their feed lacks useful substances necessary for the body. As a result, fish begin to behave apathetically, and their appetite and growth process is disturbed [2]. In particular, the lack of vitamin B6 contributes to the formation and development of the nervous system.
As a result, its deficiency can lead to a violation of the mobility of the fish, in turn, vitamin B12 helps to increase the level of hemoglobin, an extremely important element due to which the body of fish is supplied with oxygen [3]. If the fish does not have the opportunity to receive the substance, the hemoglobin level will most likely begin to fall and the growth process will be disturbed. It has been shown that the regenerative abilities of the skeleton and its formation directly depend on vitamin C [4]. Also, this vitamin helps to strengthen the immune system, which protects underwater inhabitants from a host of diseases [5]. These examples show what important role vitamins play in the development of fish and their deficiency causes hypovitaminosis.
Hypovitaminosis is most often recorded in industrial-type fish farms: in salmon hatcheries, in trout farms, when growing various species in cages, pools, circulation plants, and also in aquarium fish. They cause significant damage by slowing down the growth and development of juvenile fish, reducing the resistance of their body to the effects of environmental factors and pathogens, as well as partial death of fish. To prevent this condition, artificial vitamins are used [6], as scientific studies have shown. Synthetic vitamins are "dead" substances that do not carry any energy, they are practically not absorbed by the body.
They have a crystalline structure that cannot be broken down and processed in the body. Synthetic vitamins do not contain the necessary components that help these vitamins to be absorbed. Without the presence of these components, artificial vitamins are of no value. At best, they are simply excreted from the body. A natural vitamin is a whole natural biological complex. It includes many active ingredients (bioflavonoids, tocopherols, etc.). This whole complex is closely interconnected and has its own special structure. A natural vitamin is a whole natural biological complex. It includes many active components (bioflavonoids, tocopherols, etc.). This whole complex is closely related to each other and has its own special structure.
Useful vitamins are contained in natural products. Vitamins, which are even produced from natural products, are just "dead" crystals. They are not active. And they can acquire biological activity only by taking the form of natural (natural) vitamins [7,8]. And this will happen only if they interact with other components that make up natural vitamins. Living organisms, like vitamins, do not synthesize these components. They must be fed. That is why artificial vitamins are absorbed in the body by only 5%, and 50% is excreted from the body, and the rest is deposited in the vessels, kidneys, and liver.
There is currently no such technology in the world that would make it possible to replicate the composition of a natural vitamin [9]. Therefore, attention was drawn to the natural sources of vitamin-producing microalgae chlorella. Chlorella synthesizes a significant number of various vitamins, surpassing all plant foods and crops and even yeast in this indicator [10]. For the experiment, a nutrient medium was used in its own modification consisting of the following elements g/l Na NO3 -0.3, and the rest was a suspension obtained after electro-hydraulic treatment of 500 g of soil in a volume of 1 liter.
For the disintegration of complex insoluble salts in the soil suspension, an electrohydraulic unit with a power of 0.5 kW was used ( Figure 2).
Soil suspension processing mode with the stored energy of 0.5 kJ, number of highvoltage pulses 300, specific energy consumption is 1 kWh/kg.
Technical characteristics of the electro-hydraulic installation: -Productivity, kg/h, not less than 3.5 -Number of electrodes, one pcs.
-Rated operating voltage, V 50000 -Rated stored energy, kJ, not more than 0.5 -Apparent power, kVA, no more than 1 -The average value of the current consumed from the network, A 12 -The power supply of the installation by voltage, V 220 The treated soil suspension was filtered through filter paper, the precipitate was removed, and the resulting filtrate was added to the photobioreactor. Chlorella cultivation time was 10 days. The number of obtained chlorella cells was counted using a Goryaev camera.

Results and discussion
Chosen as an object of research, chlorella is an active producer of biomass with a high percentage of complete protein, fats, carbohydrates, vitamins, etc. In total, more than 650 substances in concentrations far exceed their content in traditional foods: meat, milk, vegetables, and fruits. Used as a feed additive for fish and animals [11].
Chlorella is a unique natural, environmentally friendly feed additive for all types of animals and fish: cattle, chickens, poultry, rabbits, cats, dogs, silkworms, etc. In terms of nutritional value, this algae is not inferior to meat.
Chlorella synthesizes a significant number of various vitamins, surpassing all plant foods and crops and even yeast in this indicator. For example, in chlorella cells, more than in yeast, inositol was found by 1.5 times, biotin -by 2 times, pantothenic acid -by 1.3 times, para-aminobenzoic acid -by 2.9 times. Carotene is 7-10 times more than in rose hips or dried apricots. Also, these green algae, unlike higher plants, contains vitamin A in its pure form. Vitamin B 12 is not found in yeast or higher plants, but chlorella produces it. This alga in the process of photosynthesis is able to capture 10-12% of sunlight, while all other green plants are only 1-2% [12].
Because of this, her photosynthesis is more productive, and hence the reaction products (glucose, oxygen, and vitamins), chlorella forms more, on which its use as a feed additive is based. It should be noted that vitamin B12 is not found in yeast or higher plants, but chlorella produces it. If fish oil contains 6 vitamins, then chlorella contains at least 13. These are the following vitamins -B1, B2, B5, B6, B8, B9, B12, K, H, PP, E, D, A, and inositol [13]. In addition, the uniqueness of chlorella is given by 4 components, the presence of which is found only in chlorella or is present in it in very high concentrations, these are chlorophyll, chlorella growth factor, sporopollenin, and chlorelin.
Chlorophyll, similar in structure to hemoglobin, has a positive effect on the mucous membrane of the gastrointestinal tract, which is a natural barrier that protects against the penetration of toxic substances from the gastrointestinal tract directly into the bloodstream. Sporopollenin binds heavy metals and toxic substances, which are then excreted from the body in a natural way. Chlorella Growth Factor is a complex and unique substance found only in chlorella that promotes natural cell growth.
The chlorella growth factor consists of nucleic acids, a number of vitamins, sugars, a protein complex, enzymes, and glycoproteins, thanks to this factor, chlorella have the ability to divide at a rate of 40 cycles per day. The chlorella growth factor is useful for the intestinal flora of fish, it has been found that its presence in the body accelerates the reproduction of acidophilus bacteria by 4 times, which die when taking antibiotics or under E3S Web of Conferences 389, 03072 (2023) https://doi.org/10.1051/e3sconf/202338903072 UESF-2023 stress, and pathogenic fungi take their place. The 4th substance is chlorogenic, which successfully destroys pathogenic microflora.
It is effective against streptococci, staphylococci, and Escherichia coli, while it is a mild selective antibiotic. Unlike chemical antibiotics, chlorine acts only on pathogenic organisms, without disturbing the digestive microflora [5]. When considering how chlorella can replace a set of artificial vitamins, the daily requirement of fish for vitamins was considered. Comparative data are shown in Table 1. As can be seen from the data presented in the table, chlorella completely covers the need of fish for vitamins, and given that the biological activity of artificial vitamins is very low, then chlorella concentrate can fully provide fish with vitamins.
For the widespread introduction of chlorella as a feed vitamin supplement in fish farming, it is necessary to optimize the nutrient medium. In this regard, it is necessary to choose a simpler and more inexpensive nutrient medium without reducing the productivity of chlorella microalgae. It is known that the main costs in the industrial cultivation of microalgae fall on the cost of chemical reagents for the nutrient medium.
Therefore, it was proposed to use the soil as raw material, since it is known that during the complete chemical analysis of the soil, it contains almost all the elements of the periodic table [9]. To solve this problem, electrohydraulic tillage was carried out based on the electrohydraulic effect of Yutkin [1]. It was previously shown that during electrohydraulic crushing of rocks and other non-metallic materials, many chemical elements and their compounds that are part of these rocks pass into the water in the form of soluble compounds in amounts reaching 90-95% of their mass content in the original material [6].
The transition of chemical elements and their compounds into an aqueous solution is explained by the fact that during electrohydraulic treatment, the simultaneous influence and complex mechanism of all acting factors of the electrohydraulic effect lead to the breaking of sorption and peripheral chemical bonds and the formation of water-soluble ionic forms of compounds. The electric pulse unit is designed to generate short powerful pulses between electrodes in a liquid medium (voltage up to 70 kV, current strength of thousands of amperes, pulse duration of a fraction of a millisecond). The pulse frequency is from 1 to 3 Hz. Under such shock pressure, changes occur in complex soil salts in a liquid medium, while insoluble forms of phosphates become soluble, and chelate compounds are also formed that are easily absorbed by microalgae, such as chlorella.
It was found by analyzes that, as a result of processing, more than 30 chemical elements in the form of various compounds pass from the soil into a water-soluble state, and only tens of kilograms of these elements in soluble compounds are obtained from 1 ton of soil, instead of about 250 g with natural dissolution. The samples were processed in a specially designed experimental batch electrohydraulic crusher in the same mode: U=50 kV; C\u003d 0.1 uF; l=15 mm with the same ratio of solid and liquid phases in each experiment.
Thus, as a result of processing, more than 30 chemical elements in the form of various compounds pass from the soil into a water-soluble state. Further studies have shown that the process of decomposition of insoluble soil salts is caused by the complex action of a number of factors that make up the electrohydraulic effect. One of the factors at the moment of discharge in a liquid medium is shock waves that create a short-term hydrodynamic pressure of up to 100 thousand atmospheres [3]. Another factor is the specificity of obtaining an ultra-long discharge in water, which determines the predominance of OH anions in the discharge zone, which intensively transform into hydrogen peroxide, which, in turn, decomposes into H20 and O; in the aggregate, all this causes vigorous destruction and oxidation by the formed active atomic oxygen of previously insoluble and insoluble soil salts.
Under the conditions of electrohydraulic treatment, all the processes of destruction of complex soil salts, which occur extremely slowly in natural conditions, are realized almost instantly [4]. In this regard, an electrohydraulic plant for soil and water treatment was made. 500 grams of soil and 1.5 liters of water were loaded into the reactor chamber. Electrohydraulic treatment was carried out in the following mode U=50 kV; C\u003d 0.1 uF; l=15 mm, processing was carried out for 10 minutes. In the end, the resulting suspension of macro and microelements was filtered through filter paper for separation of silicon compounds.
At the next stages of the experiments, the efficiency of cultivating a strain of chlorella on Tamiya nutrient media was studied, as control and obtained nutrient medium containing macro and microelements based on local raw materials. The data obtained are presented in Table 2. Table 2. Data on the accumulation of chlorella biomass depending on the type of nutrient medium on the 14th day of cultivation.

Name of the nutrient medium Cell concentration in one ml of medium
Tamiya media (control) 28.0 million Nutrient medium containing macro and microelements based on soil suspension.

million
As shown by the data obtained on the cultivation of chlorella in a nutrient medium containing macro and microelements based on local raw materials, it practically corresponds in terms of effectiveness to the traditional Tamiya nutrient medium.

Conclusions
Based on the electro-hydraulic effect, where the soil is exposed to high-voltage discharges, a water-soluble complex of macro and microelements can be obtained. The resulting watersoluble complex of macro-and microelements makes it possible to completely replace expensive chemical elements in the cultivation of Chlorella vulgaris. The presence of a full complex of natural vitamins in Chlorella vulgaris makes it possible to abandon ineffective and expensive artificial vitamins when feeding fish.
Thus, the experimental data convincingly demonstrate the possibility of transferring the microalgae cultivation technology to the developed nutrient medium containing macro and microelements based on water-soluble chemical elements obtained from the soil. All this makes it possible to introduce a new technology of electro-hydraulic action of high-voltage discharges to develop alternative nutrient media for the cultivation of microalgae based on local raw materials, in particular soil. Due to this, the cost of cultivating valuable microalgae is significantly reduced, and the procedure for preparing a nutrient medium is simplified. Thus, an innovative technology has been developed for obtaining an inexpensive, highly effective vitamin feed additive based on (Chlorella vulgaris) to replace artificial vitamins in fish feeding.