The allocation method destructors of oil and oil products
The invention relates to the field of environmental protection and can be used for the restoration of contaminated land. The method includes sampling from contaminated surfaces, selection of hydrocarbon-oxidizing bacteria in liquid mineral medium with the addition of oil from contamination during incubation temperatures +5°C, +15°C, +25°C and +35°C, the selection of crops for replanting after 5, 10 and 15 days, re-seeding of the obtained cultures and further tested separately all atzelektronik microorganisms in fermenters under cultivation parameters, optimal for the selected microorganisms. Acquired bacterial preparations on the basis of indigenous microorganisms produce processing of oil-contaminated site. The method increases the likelihood of finding the most active representatives of the hydrocarbon-oxidizing microflora, utilizing hydrocarbons at low positive temperatures, reduces the cost of production of a biological product. The use of these biologics provides an opportunity to start the processing of oil-contaminated site in early spring and to clear the area of oil pollution in one VEGA is vlahuta a natural component of the soil system.
Scientists and experts is constantly working on the identification of the most effective strains capable of destructivity oil and its compounds.
In the all-Union collection of industrial microorganisms have a sufficient quantity of hydrocarbon-oxidizing bacteria, however, their optimal temperature indicator for Russia high - from +20 to +30°C, i.e., biological products based on them will actively destroy hydrocarbons only in the warm period of time. When the air temperature decreases the efficiency of biopreparations drops, restorative work on this end. Therefore, for the Central, especially in the Northern regions of Russia the allocation of oil-oxidizing microorganisms, their selection should be conducted in the direction of not only high destructive activity, but also adaptation to low positive temperatures and various oil fractions.
Recently, restoration of contaminated lands is widespread use of cultural liquid, based on aboriginal highly efficient hydrocarbon-oxidizing microorganisms.
The determining factor in this direction is the allocation method.
The main disadvantage of this method of allocation is that effective strains of hydrocarbon-oxidizing microorganisms isolated from the samples for a period of polluted more than 3 years, although reclamation work should be carried out immediately after the accident.
Markarova M. Y. (Use of hydrocarbon-oxidizing bacteria for remediation of oil-contaminated lands in the Far North. Diss. on saisc. academic degree Cand. Biol.-chem. Sciences.- Syktyvkar, 1999. - S. 44-45) offers a selection of hydrocarbon-oxidizing microflora conduct of oil polluted soil by re-sowing soil suspension on fresh pita bread medium and incubated at t+10°C. The flask was periodically shaken. 40 days to produce reseeding method of limiting dilutions of the suspension in a nutrient medium. Mineral medium is sterilized at 1 ATM for 1 hour, poured into Petri dishes and after solidification of the agar with a sterile spatula on the surface of the rubbed oil at 1 ml per 1 Petri dish. After germination on the surface of the agar separate colonies of microorganisms scatter them separately in Petri dishes on a nutrient medium.
The main disadvantage of this method of allocation nefteynaliva microorganisms is a very long incubation period of 40 days. Secondly, taken one temperature range - +10°C, although conditions in the Northern regions and to continue reclamation works at low positive temperatures, the incubation should be performed at lower temperatures.
As the prototype is set to the invention Vlasov S. A. and other “Method of cleaning soil from oil and oil products” (patent RF №2128703, With 12 N 1/26 At 09 With 1/10, 1999). Of oil-contaminated soils emit microorganisms, utilizing oil, then their produce in the fermenters and the culture fluid is treated with the place of pollution.
The main disadvantage of this izobreteny is high, secondly, the seeding is made from dilutions of 10-3and 10-4at that time, as an active destructors may remain in the first and second dilutions third, not taking into account the rate of appearance of colonies, fourth, incubation was carried out at a temperature of 30°C, i.e., when the air temperature decreases the biological product will lose its activity.
These deficiencies have been addressed in the proposed method of selection of effective destructors oil.
The essence of the method is performed: sampling from contaminated surface, then is the selection of hydrocarbon-oxidizing bacteria with subsequent reseeding of the obtained cultures and time separately all atzelektronik microorganisms in fermenters under cultivation parameters, optimal for the selected microorganisms. Breeding begins in liquid mineral medium with the addition of oil from the place of pollution, the incubation is carried out at different temperatures +5°C, +15°C, +25°C and +35°C, and the selection of crops for replanting carried out in 5, 10 and 15 days.
The progress: samples are dried to air-dry state, are chosen small stones, vegetable residues. Preparing m is - ,2 g CaCl2- 0.02 g FeCl3- 0,002 grams, water to 1 liter. One gram of sample is transferred into a sterile porcelain mortar, lightly moistened with saline environment and pounded to a paste-like state. Content is transferred into a flask with a volume of 750-1000 mm (mortar and pestle to rinse the salt environment, shifting the balance in the same bulb). Total volume of the mixture is brought to 100 ml flask submitted 1 ml of sterile raw oil with the specific location of contamination. Flasks are incubated at different temperatures:
- +5°C to highlight psychotomimetic active destructors carrying out the decomposition of oil and petroleum products at low positive temperatures;
- at +15°C to highlight effective aboriginal groups of bacteria that are actively acquiring oil in this temperature range;
- at +25°C for isolation of thermophilic microorganisms;
- at +35°C to highlight groups of bacteria that breed at high temperatures.
The flask periodically shaken. On the 5th, 10th, 15th day fixed visible changes in the environment (opacity, pigment layer of oil degassing, delamination, discoloration, appearance of products of saponification and others). The results of the laboratory awn education cell biomass and, as a rule, the time of cultivation of these strains is much lower. At the same time destructive of their properties are not always high, and especially oil, containing a high percentage of heavy fractions. The use of the biological product consisting mainly of bacteria that appeared on the fifth day, quite justified, as low destructive indicators are compensated by a high rate of bacterial growth.
Colonies that appeared on the tenth day, depending on the type of culture, have average harvest indicators in this Association are mainly active representatives of oil degrading, well utilizing both light and medium fractions of oil.
Group of bacteria that appeared on the fifteenth day, have a slow growth rate, their otsilindrovannye properties are on heavy oil fractions, it is difficult biodegradable.
From the flasks with the top layer select the eyedropper 0.5 cm3mixture and transferred into a test tube with 4.5 cm3salt environment. Done 2-3 serial tenfold dilution in the same environment. From each tube in 2 repetitions is seeding 0.1 cm3on cups: psychrophiles 22±2°C; for mesophilous 28±2°C; for thermoflow 37±2°C. Cups with sowing looking at daily. Shall count and description of colony morphology. All selected colonies isolated on nutrient agar method debilitating stroke”. The purity of selected crops checked visually. Appreciate cultural properties, the nature of growth of the colonies, their uniformity visually and by using a microscope. Pure bacterial culture plates avivausa tube with beveled agar, then incubated 3-5 days at appropriate temperatures. Selected representatives bacteria assign a registration number, make a log. Then conduct a study of the basic biological properties that determine their destructive activity.
The accumulation of the bacterial preparation is carried out in the fermentor MT-20 set for each culture the optimal parameters.
The application of this method for producing the bacterial preparation, consisting of the most effective destructors of oil and oil products, both economically and environmentally justified. The cost of production of the bacterial preparation on the basis of indigenous microorganisms is low, and the use of adapted to the source of sagra what about the budget.
The proposed method is used to allocate effective destructors oil, is part of the “remediation Technology neftezagryazneny lands on emergency stations and landfills by way of activation of the indigenous microflora”, approved by the Ministry of Natural Resources of the Russian Federation on February 21, 2002 No. 35-01-2/108. Positive conclusion of the State ecological examination approved by order of the Minister of Ministry of natural resources 13.02.2002, No. 70; “Technologies for remediation of contaminated land by way of activating the indigenous microorganisms in licensed areas OOO “LUKOIL-Western Siberia”, approved by the Ministry of Natural Resources of the Russian Federation September 2, 2002 No. 552. A positive conclusion of the State ecological expertise of 19 July 2002; “Technology for remediation of oil polluted soils in winter with the use of indigenous microorganisms destructors oil”, approved by the Ministry of Natural Resources of the Russian Federation September 2, 2002 №551. A positive conclusion of the State ecological expertise of 19 July 2002, developed by ZAO NTC Elite complex“.
The method has been applied in practice>P CLASS="ptx2">With oil-contaminated site LPDS “Torwali” Tyumen AA of Sibnefteprovod in the fall of 2001 were selected soil samples. In the Microbiology laboratory as previously described were isolated from psychrophilic, mesophilic and thermophilic aboriginal aftershave bacteria. In March 2002 prepared backpropagate. Upon the occurrence of positive air temperatures of oil-contaminated area was treated on the basis of psychotomimetic microorganisms, two weeks was repeated treatment with this drug. The results are presented in the table. The third spraying the herbicide on the basis of psychrophilic destructors oil held on may 15. Before drug treatment, consisting of mesophilic microorganisms - June 1, were selected soil samples at the amount of oil and oil products. The use of psychrophilic neftereklama bacteria allowed by this time spread out 85% of oil and oil products, while in the control volume decreased slightly with 194,7 to 173.2 g/kg of the Fourth processing of oil-contaminated site was conducted by the drug on the basis of mesophilous and on 1 July the site was cleared on 93,8%. The subsequent treatments n is isawanya psychrophilic nefteynaliva microorganisms allowed to begin remediation work in the early spring months and for one growing season to clear the area from oil pollution.
The allocation method destructors of oil and oil products, including sampling from contaminated surfaces, selection of hydrocarbon-oxidizing bacteria with subsequent reseeding of the obtained cultures and further tested separately all atzelektronik microorganisms in fermenters under cultivation parameters, optimal for the selected microorganisms, characterized in that the selection starts on liquid mineral medium with the addition of oil from the place of pollution, the incubation is carried out at different temperatures +5°C, +15°C, +25°C and +35°C and the selection of crops for replanting carried out in 5, 10 and 15 days.
FIELD: environment protection.
SUBSTANCE: method involves mechanical removal of excessive contaminants from surface and reducing toxicity of contaminated soil; sowing green manure crops and perennial grasses; preliminarily removing taking soil samples from contaminated surface and separating native contaminant destructors therefrom; after mechanical removal of contaminant from soil surface, providing sequential treatment of plot soil with water-soluble humates and at least double treatment with native contaminant destructors; mellowing soil; applying lime with mineral fertilizers such as saltpeter, double superphosphate and calcium chloride. Green manure crops are sown after application of fertilizers. Method further involves grinding green manure crops, plowing into soil and covering with soil by means of covering roll.
EFFECT: increased efficiency, reliable ecologically clean restoration of soil after contamination thereof.