Drug – bio-affecting and body treating compositions – Whole live micro-organism – cell – or virus containing – Fungus
Reexamination Certificate
1999-06-21
2002-09-03
Marx, Irene (Department: 1651)
Drug, bio-affecting and body treating compositions
Whole live micro-organism, cell, or virus containing
Fungus
C435S255100, C435S255400, C435S262500
Reexamination Certificate
active
06444204
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates generally to the field of bio-technologies and, more particularly, to microbiology and environmental protection, refers to the creation of a novel pool for cleaning oil-polluted soil, water and equipment, to the manufacture of a new biological preparation based on this pool designed to be used for the purification of water, industrial sewage, soil and other media from a wide range of petroleum products and directly from crude oil, and to the use of the obtained biological preparation for the efficient recovery of the above said oil-polluted media.
BACKGROUND OF THE INVENTION
The ever-growing volumes of oil output, transportation, refining and utilization result in the wide-spread pollution of water and soil areas with crude oil and oil products. Despite safety precautions taken, accidents intermittently occur at petroleum-extracting and oil-refining enterprises, pipelines, oil tankers, resulting in an environmentally hazardous oil and oil-product spillage. It is common knowledge that Nature itself possesses an efficient system for the recovery of oil-polluted areas by means of microorganisms capable of assimilating the hydrocarbons of oil. However, the microflora of Nature is no longer capable of effectively recovering million tons of oil finding every year its way into the environment. As a result, the mankind has found itself under the real threat of ecological disaster (see, L. R. Brown.
Chemical Engineering Progress
. (1987) Vol. 83, No. 10, pp. 35-40; “Oil-degrading microorganisms”).
Such traditional methods of recovery as mechanical, chemical and physical fail to provide a sufficient degree of recovery of oil-polluted water and soil. Besides, in many cases the application of these methods inflicts a more pronounced damage on the environment than oil spillage itself.
In view of the above, biological methods of recovery of oil-polluted media recently find the ever-growing application, including the use of oil-oxidizing microorganisms. Analysis of the development of bio-technologies relating to the recovery of oil-polluted water and soil ecosystems and equipment enables one to come to the conclusion that the processes based on the oil-product biodegradation under the action of microorganisms are ecologically efficient. In accordance with known bio-technologies, the polluted medium may be exposed to the action of biological preparations, including microorganisms either in the form of pure isolated cultures or pools of microorganisms, that is to say a combination or association of two or more organisms. However, it should be emphasized that from the viewpoint of achieving of a more complete utilization of biotechnological functionality of microorganisms—oil biodegrading agents, the application of pure isolated cultures of microorganisms is less advantageous than that of mixtures or associations of microorganisms which, in virtue of their specificity, are capable of utilizing a wider range of compositionally heterogeneous substrates (industrial sewage, soil pollution, etc.) and provide degradation of organic compounds at the expense of a combined action of several types of microorganisms. Thus a more pronounced ecological efficiency of biotechnological decontamination and recovery is attained, said biotechnological decontamination and recovery being carried out for a shorter time.
As a rule, known biological preparations for the recovery of oil-polluted areas are characterized either by a relatively low efficiency in virtue of the selectivity of monospecies cultures used in the biological preparation or by complexity of producing the biological preparation, which is associated with the process for adapting microorganisms to the co-existence when using a microorganism pool in the biological preparation concerned.
Thus, there has been known the strain Phodococcus sp. HX7 possessing oil-oxidizing properties, which is used when cleaning water and soil from oil pollution. In the treatment of the polluted areas with this bacteria, the usual dose of application is 1 gram of the cellular biomass per square meter of a treated area; or in the case of using a suspension of the biological preparation based on the above-cited bacteria it contains 1·10
7
-1·10
7
-1·10
8
cells in 1 milliliter (1·10
7
-1·10
8
cells/ml). In so doing, the application rate of the suspension is 1 liter per square meter (1 l m
2
) of an oil-polluted area.
However, the above-mentioned process for cleaning media from oil-pollution is insufficiently effective, since the use is made of a bacterial monospecies which limits a range of hydrocarbons to be purified. Besides, a process for growing such a culture is of long duration and may take as long as several days (see, Russian Patent No. 2,039,714, IPC C 02F 3/34, 1995).
Known in the prior art is the natural strain
Pseudomonas putida
-36 and its utilization for the recovery of oil-polluted soil (see, U.S. Pat. No. 4,822,490, IPC C 02F 3/34, 1989). In the known method of cleaning water and soil, a bacterial culture is applied to the oil-polluted area in the form of a biological preparation comprising the above-cited strain in admixture with a mineral fertilizer containing nitrogen in the nitrate form. The amount of the culture to be applied is at least 1·10
4
cells/mg, where the flow rate of an aqueous mixture is from 0.5·to 1.0 liter per square meter.
However, the degree of recovery from various kinds of oil attained when using the above said bacterial strain also remains insufficient. Moreover, the application of this strain has certain limitations. This is associated with the fact that the strain grown on a standard carbohydrate substrate hardly adapts itself to new conditions, which means the necessity of using new substrates from soil. Besides, the said strain
Pseudomonas putida
-36 possesses hemolytic and gelatinizing activity which prevents the development of the process of natural self-recovery by means of microorganisms. Furthermore, the use of the nitrate forms of nitrogen when applying the bacterial culture into soil results in an additional contamination of the environment with toxic nitrate ions.
There has been known a pool of microorganisms Rhodococcus sp. Bkmac—1500D, Rhodococcus maris VKM AC-1501D,
Rhodococcus crythropolis
VKM AC-1502D,
Pseudomonas stutzeri
VKM B- 1972D, Candida sp. VKM-Y-2778D and a biological preparation produced using this pool for cleaning water and soil polluted with crude oil and oil products.
However, the known pool and biological preparation are insufficiently effective when cleaning oil-polluted media and, moreover, characterized by a complicated production method because of a great number of various microorganisms constituting the pool. The industrial employment of this biological preparation comprising the said pool is difficult for cleaning purposes, since five strains constituting the pool differ very much in their physiological features and have different growth characteristics.
A known process for producing a biological preparation based on this pool comprises preparation of a solution of a liquid nutrient medium containing the sources of nitrogen, phosphorus, trace elements and liquid paraffins as a carbon source, followed by cultivation, for example in a fermenter, of an inoculation medium in the form of the above-mentioned pool of microorganisms, at a temperature of 28° C. and pH maintained at 6.5-7.0. In so doing, the process of cultivation is carried out under aerobic conditions, for example when the medium is bubbled with the air flow. Thereafter, a biomass to be cultured is concentrated by the separation method and dehydrated by means of the lyophilic or thermal-vacuum drying to a 10% humidity. A biological preparation thus obtained contains 4·10
11
-4·10
12
living cells per 1 gram and comprises both lipophilic and halophilic microorganisms. This biological preparation is capable of oxidizing petroleum products both in the zone of contacting with water and directly in an oil film and capable of removing oil pollution not only in the fresh but also in t
Avchieva Penker Babaevna
Kuznetsov Petr Alexandrovich
Knobbe Martens Olson & Bear LLP
Marx Irene
LandOfFree
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