NO162522B - AHEAD SOLUTION OF PRIOR PREPARED POLYMERS WITH REAL VISCOSITY CHARACTERISTICS AND PROCEDURE FOR AA INCREASING VISCITY IN A SOLUTION OF PRIOR PREPARED POLYMERS. - Google Patents

Description

The present invention relates to an aqueous solution

of pre-produced polymers with increased viscosity properties and a method for increasing the viscosity in a solution of pre-produced polymers.

As far as the present invention is concerned, all polymers can be divided into two classes: biopolymers and synthetic polymers. Biopolymers are polymers that are produced at least partially

show by the impact of biological processes, while synthetic polymers are produced by chemical methods.

Biopolymers can be obtained from a variety of natural plant sources, including tree exudates, seed extracts, seaweeds, starches and cellulose derivatives. Some biopolymers, such as certain polysaccharides, can be produced using microorganisms.

A well-known example of a microbial biopolymer is xanthan gum, which is an anionic heteropolysaccharide produced extracellularly from carbohydrate substances by means of organisms of the genus Xanthomonas, such as X. campe stris and X. begoniae. Xanthan and several other microbial biopolymers, such as the mushroom product scleroglucan that is sold

under the trademark "Polytran" and a polysaccharide obtained from the soil bacterium Erwinia marketed under the trademark "Zanflo"

is available in commercial quantities for such applications as printing inks and coatings, cosmetics, ceramics, paint thickeners, drilling muds, pharmaceuticals and food. Some microbial biopolymers have the ability to act as surfactants, such as e.g. surfactin produced by Bacillus subtilis. These surfactants are useful as flocculants, emulsifiers, demulsifiers, detergents, adhesives and liquids for use in enhanced oil recovery.

A microbial biopolymer is usually obtained either by fermentation or enzymatic synthesis. During fermentation, a microorganism that is genetically capable of building up a specific polymer is placed in a medium with the necessary nutrients and the necessary substrate. The polysaccharide is recovered from this medium. In enzymatic synthesis, microbial cells are removed from a cell culture so that a culture fluid containing the extracellular enzyme remains. The enzyme is then brought into contact with the substrate whereby the polysaccharide is produced. For further information regarding polysaccharides and their production, reference is made to "Microbial Polysaccharides", Kirk-Othmer Encyclopedia of Chemical Technology, 3rd ed., vol. 15, pages 439 - 58.

Synthetic polymers are produced by combining one or more types of monomeric substances under conditions suitable for polymerization. They are usually less complex than biopolymers as they usually consist of a regular sequence of simple monomer units.

Both biopolymers and synthetic polymers can be added to water or other solvents to form solutions that have beneficial properties, such as increased solution viscosity. The viscosity of these solutions can vary greatly depending on the amount of polymer in solution, molecular weight and the structure of the polymer, temperature, presence of salts (especially multivalent salts, such as magnesium and calcium) and other factors.

Persistent stability of polymer solutions under demanding conditions with regard to temperature, salt content, pressure, shear forces and the presence of oxygen and chemicals is important for some users. Some microorganisms

nisms, whether they naturally exist in the solution environment or are introduced by pollution, are also able to break down polymeric substances and thereby reduce the viscosity of the solution. E.g. Many microorganisms secrete enzymes (cellulase, amylase) that can hydrolyse polysaccharides into monosaccharides that can be used in the organism's metabolism. Synthetic polymers can also lose viscosity during microbial attack.

For the reasons stated above, one of the aims of the ongoing research is to produce stable polymer solutions.

The solution must be stable against the chemical, thermal and biological conditions encountered in various industrial processes. Of particular interest is the search for a polymer solution for use in enhanced oil recovery that retains its stability during pretreatment steps, injection

and through the oil-bearing formation.

For economic reasons, another aim is to achieve maximum viscosity at the lowest possible cost, i.e. the highest possible viscosity from the smallest possible amount of polymer in the solution.

The invention relates to an aqueous solution of pre-produced polymers with increased viscosity properties.

The aqueous solution is characterized in that it is prepared by mixing, under conditions conducive to bacterial growth, an aqueous solution of previously prepared polymers with a bacterium selected from the group consisting of gram-negative rod bacteria, gram-positive rod bacteria and gram- positive sphere bacteria, without the ability to re-synthesize the polymers, but with the ability to increase the viscosity of the polymer solution, as this ability has been demonstrated using the following screening procedure:

(i) preparation of a culture of the bacterium, (ii) preparation of an aqueous solution of the previously prepared polymers and addition of nutrients and minerals required for bacterial growth, (iii) separation of the solution into two parts, a test- and a control part, both parts lacking substrate except for the pre-made polymers and thereby lacking a usable substrate to enable the synthesis of another polymer, (iv) inoculating the test solution with the culture of the bacterium and incubating the test and control solutions , and (v) measuring the viscosities of both the test and control solutions after incubation to determine the viscosity of the test solution relative to the viscosity of the control solution.

The invention further relates to a method for increasing the viscosity of a solution of pre-made polymers by mixing under conditions which favor bacterial growth,

(a) an aqueous solution of preformed poly-

more and

(b) a bacterium selected from the group consisting of gram-negative rod bacteria, gram-positive rod bacteria and gram-positive globule bacteria without the ability to re-synthesize the polymers, but with the ability to increase the viscosity of the polymer solution in the following screening method: (i) preparation of a culture of the bacterium, (ii) preparation of an aqueous solution of the previously prepared polymers and addition of nutrients and minerals required for bacterial growth, (iii) separation of the solution into two parts, a test part and a control part , both parts lacking substrate apart from the previously prepared polymers and thereby lacking a usable substrate to enable the synthesis of another polymer, (iv) inoculating the test solution with the culture of the microorganism and incubating the test-

and the control solutions, and

(v) measuring the viscosities of both the test and control solution after the incubation to determine the viscosity of the test solution relative to the viscosity of the control solution.

It has now been discovered that certain bacteria are able to increase the viscosity of a polymer solution even when the bacteria in question itself is genetically incapable of new synthesis of the polymer. The term "new synthesis" is used here to omit the known techniques where a bacterium builds up polymeric material (such as polysaccharides) from monomers and thereby increases the viscosity of a solution. As the present invention is defined, the bacterium cannot build up the selected polymer from monomers. This method is therefore best suited for previously prepared polymers (ie polymers that have previously been prepared either by chemical or microbial synthesis). E.g. a particular bacterium may be genetically unable to produce a polysaccharide, but is capable of altering a dissolution of the polysaccharide in a way that causes an increase in its viscosity.

Without intending to be bound by any particular theory, it is believed that the bacterium acts in a manner that increases the average molecular weight of the polymers in solution, possibly through the action of one or more enzymes. Below is a detailed description of suitable polymers, bacteria, viscosity changes and the method.

Polymers

The polymers that can be used in the present invention include all biopolymers and synthetic polymers that are compatible with water. It is preferred that the polymers are not only compatible with water, but that they are readily soluble in water as they will thereby be readily accessible to the bacteria and any accompanying extracellular enzymes in an aqueous medium. However, even partially soluble or water-insoluble polymers are considered to be within the scope of the present invention, provided that there is an interface between the polymer and the bacteria or the enzymes.

As far as the present invention is concerned, a "solution" of polymer means, in addition to dissolved polymer, any water-wettable polymer surface such as polymer suspensions, colloidal dispersions, polymer gels or emulsions, and wettable, solid polymer substances.

In the preferred embodiment of the invention, the polymer can be selected from among those known to be effective for a particular end use. E.g. a polyacrylamide can be chosen if the desired application is water treatment, increased oil extraction, mineral processing or wood pulp and paper processing.

Extensive information has been published regarding applications of polymer solutions. Water-soluble polymeric materials with long chains are widely used as flocculation aids in the separation of solids and liquids. The use of aminated starches, polyamines and polyacrylamides in industrial water treatment is summarized in "Water (Industrial)", Kirk-Othmer Encyclopedia of Chemical Technology, 2nd ed., vol. 22. Other polymeric flocculating agents are used in such processes as water clarification, sewage treatment, metal polishing, paper production, sugar refining, mineral extraction and food processing as discussed in "Flocculating Agents", Kirk-Othmer Encyclopedia of Chemical Technology, 3. ed., vol. 10.

Enhanced oil recovery (EOR) usually requires the injection of an aqueous fluid, such as water or salt water, to push the oil ahead through the formation to a production well. A polymer is often added to at least a portion of the injected fluid to form a slurry of increased viscosity. The polymer-thickened liquid travels with a smooth front and minimizes the bypassing and "finger-ing" that can otherwise occur during conventional waterflooding.

Polymers considered to be useful in enhanced oil recovery (EOR) include guar gum, hydroxypropyl guar, sodium carboxymethylcellulose, hydroxyethylcellulose, carboxymethylhydroxyethylcellulose, xanthan gum, glucan, jonannes breadfruit gum, polyacrylamide, hydrolyzed polyacrylamide, poly(acrylic acid) and salts thereof, poly (2-acrylamido-2-methyl-propane-sulfuric acid)(2-salt) (AMPS), poly(acrylic acid coacrylate ester, poly(vinyl pyrrolidone), cellulose sulfate esters, poly-(ethylene oxide), poly(vinyl alcohol), polyamine, poly(vinyl acetate-comaleic anhydride) and poly(styrenesulfonic acid) and salts thereof Among these, hydroxyethylcellulose, carboxymethylhydroxyethylcellulose, xanthan gum and hydrolyzed polyacrylamide are preferred.

The concentration of the polymer solution used in

EOR is usually determined by the permeability of the rock and the viscosity of the oil in the formation. For economic reasons, the concentration is kept at the minimal effect levels which are usually between 20 ppm and 3000 ppm, more commonly 50 to 1000 ppm, and most commonly 200 to 600 ppm. Other formation conditions that must be taken into account when selecting an effective polymer are high temperatures, shear stresses, high salt content and extreme pH values.

Otherwise, polymers are used in oil fields during drilling, casting, fracturing, acidification, control of water production, prevention of sand production, clay stabilization, lost circulation and the like. Generally, guar, guarderi vats, cellulose-containing substances, xanthans, locust bean gums, starches and synthetic polymers such as e.g. polyacrylamide, as more fully described in Chatterji, J. and Borchards, J.K., "Applications of Water-Soluble Polymers in the Oil Field", J. Pet. Tech. , 2042 - 2056, (Nov. 1981).

Preferred polymers for mobility control agents in EOR have all or most of the following characteristics: 1. Good solubility in water with a high positive effect on solution viscosity; 2. Ability to withstand dissolved salts (salt water) in the aqueous solution; 3. High molecular weight for maximum viscosity effect; 4. Free from unusually high levels of branching or gels that can cause injection problems; 5. Stability under thermal, biological and shear stress influences in the formation environment and during the injection process; 6. Possibly cheap when produced in large volume or with a natural occurrence in large volume.

Measurement of viscosity

The viscosity of a water-soluble polymer in solution is a function of many factors, mainly molecular weight and electrolytic property. The higher the molecular weight of the polymer, the higher the viscosity. Charged polymers also have a tendency to lose viscosity in high salinity water due to neutralization of the charges by the ions in solution.

In the present invention, the classic definition of viscosity for Newtonian fluids will be used, where viscosity is equal to shear stress divided by the shear rate. Shear stress is further defined as the force per unit area required to produce the shear effect. The cutting speed is a measured value.

A suitable way to measure viscosity is to use a viscometer such as e.g. a Brookfield viscometer. Brookfield viscometers measure viscosity by measuring the force required to rotate a spindle in a liquid. Since practically all liquids become thinner as the temperature increases, and thicker as they cool, the temperature of the liquids being compared should be measured and, if possible, kept equal.

The LVT viscometer from Brookfield with the UL (ultra-low viscosity) adapter is particularly suitable for this application. The UL adapter provides amplification effects that enable measurements with a reproducibility of 0.2 centipoise (eps) in the ultra-low viscosity range from 0 to 10 eps. Calibration of the instrument gave an average error of -0.2 eps at both ends of the shear rate range (10 seconds, and 70 seconds). The shear rate of the instrument can be varied from 73.42 to 0.36 seconds covering the shear range encountered on in the formation during the flooding process (usually 10 seconds).

Other measuring devices such as the capillary viscometer, disk and cone viscometer, and concentric cylinders can be used if appropriate.

Some polymeric solutions will exhibit thixotropic properties, characterized by the ability of a solution to liquefy when subjected to shaking, stirring or other action, and then return to a gel upon standing. Common examples of this are mayonnaise and paint. Thixotropy is also desirable for polymer solutions used in enhanced oil recovery where the solution is more liquid to facilitate pumping, but becomes more viscous once it has entered the oil-bearing formation.

Non-Newtonian fluids are those where a change in shear rate is not proportional to a change in shear stress. For such liquids, the ratio between shear rate and shear stress is sometimes determined at several shear rates. Although the polymeric solutions of the present invention may be non-Newtonian, shear rates obtained using a single set of shear stress conditions will usually be sufficient for use as a basis of comparison to determine any change in viscosity. If desired, additional measurements of shear speed can be made with different shear forces. For these purposes, all comparisons are thus made as if the fluids being measured behaved as Newtonian fluids. Procedures for determining viscosity are well within the professional knowledge of an analytical chemist, and more detailed guidelines can be found in textbooks, journals and instruction booklets from manufacturers of viscometers,

As far as the present invention applies, a change in measured viscosity of approx. 10% to be highly significant when comparing a solution of sample polymer with a control solution. As commonly available equipment is unable to accurately measure the weight of polymers with a molecular weight up to 1 million, the change in the polymer was measured indirectly by determining intrinsic viscosity.

The most common method for calculating the molecular weight of a polymer in solution is the use of "intrinsic viscosity" [N]. The intrinsic viscosity "[N]" can be related to the molecular weight of the polymer using the equation of Mark, Houwink and Sakurada:

[N] = KM<a>

where K and a are constants that are a function of polymer type, temperature and solvent. (This equation is discussed in more detail in Billmeyer (see below)).

The constants are first determined by measuring polymers of known molecular weight. The determination of [N] is done by means of capillary viscometry of dilute polymer solutions at several concentrations. Viscosity data is extrapolated to 0 concentration to give a value related to molecular weight at a constant polymer type, a constant solvent and a constant temperature.

Although the constant values are not known and the absolute values for molecular weight are unknown, the method is still useful as the value for [N] is directly proportional to a low power of the molecular weight. In a solvent-temperature-polymer system where the constants are not available, the values of [N] can be used to determine increases or decreases in molecular weight, but not the actual values. This concept is discussed further in a popular textbook (Billmeyer, Jr., F.W., Textbook of Polymer Science, 3rd ed., New York: Interscience Publishers. 1965; pp. 79-85). This method is satisfactory as one only needs to determine relative changes in molecular weight.

Bacteria

The bacteria that can be used in the present invention have the ability to increase or at least stabilize the viscosity of previously prepared polymers in solution.

In the broadest aspect of the invention, a suitable bacterium can be selected by choosing one or more candidates and carrying out the sorting test described below to determine the effect of the bacterium on the polymer solution. Current bacteria can be found almost anywhere. Soil samples and plants are logical places as well as ground and surface water. Availability of oxygen should not be a limiting factor in the selection, as both aerobic and anaerobic bacteria can be used. Existing cultures of bacteria can of course be tested with regard to applicability in the present invention.

Types of bacteria that can be used in the present invention include gram-negative aerobic rod bacteria, gram-negative facultatively anaerobic rod bacteria, gram-negative anaerobic rod bacteria and gram-positive rod bacteria. Additional information regarding the characteristics of each

of these varieties may be found in Bergy's Manual of Determinative Bacteriology, (8th ed.), R.E. Buchanan and N.E. Gibbons, Co-editors (1974.

Once the sample is obtained, the bacteria should be isolated

and propagated on one or more types of laboratory media using standard techniques.

Approach

The invention may be practiced using any preparation method suitable for microbiological synthesis. Procedures that are commonly used within the technique are divided into two categories: batchwise and continuous. In a batch method, all of the starting materials, including the bacteria, are placed in a container where they remain until the desired product is formed, after which the container is emptied and the product is separated. In a continuous process, the raw materials are added and the product removed at a constant rate. For the production and treatment of large quantities of materials, a continuous process is preferred, although batch processes are particularly useful when careful control over the process is desired.

It is characteristic that the method is carried out under conditions which are optimal for the growth of the bacteria, as the maximum results are obtained at a high metabolic rate. These optimal conditions usually correspond to those found in the bacteria's natural environment. If possible, the environment is copied, thereby obtaining the same temperature, pH, salinity, content of trace elements and other materials.

If the environment cannot be easily replicated, a standard nutrient medium is used to provide water, sugar, amino acids, vitamins, minerals and trace elements. Such media are available from suppliers of biological material.

The optimum pH and temperature range for rapid growth, as well as the range of tolerable conditions, can be readily determined using common tests known to those skilled in the art. A more detailed description of these samples is reproduced in "Growth", Ralph N. Costilow, Ed. Manual of Methods for General Bacteriology, pp. 65-179 (1981).

Control procedure for bacterial viscosity increase

The following procedure can be used to check polymer blanks to determine if a given bacterium will grow

the viscosity of a solution of the polymer. It can too

used to control relevant bacteria to be-

vote whether they will increase the viscosity of a given polymer. The method can also be used to test completely new mixtures of polymers and bacteria.

A. Making culture. A culture of the bacterium is prepared in a suitable culture medium. The specific one

The pH and the ingredients in the medium will vary depending on the requirements of the particular bacteria. This culture is preferably incubated for 24 hours at the organism's optimal cultivation temperature.

B. Inoculation of the polymer solution. The polymer solution is preferably concentrated sufficiently to give measurable viscosities for the viscosity sample, but is sufficiently diluted that the amount of polymer is not toxic to the bacteria. The polymer solution may be sterilized by steam or other means to avoid the effects of competing growth of unwanted microbial contaminants. A solution of basal salts is optionally added to the polymer to ensure essential nutrients and minerals for the bacteria.

The growing cell culture can then be centrifuged, washed to remove any residues of culture medium and resuspended in a minimal amount of distilled water. To achieve optimal results, a cell count is performed to determine the concentration of cells. A measured amount of inoculum containing a given concentration of cells is then added to the solution containing the pre-prepared polymer. A control solution is also prepared which has the same concentration of polymer solution and basal salts, but which does not contain any bacteria. An equivalent volume of sterile distilled water is added to the control to match the volume after inoculum added to the solutions containing the pre-prepared polymer. C. Incubation of the solution. The control and the polymer solution with the microorganism are incubated at the optimal culture temperature for the bacteria, incubated at the optimal culture solution is removed at regular intervals to detect tendencies in any viscosity change. Definitive results are usually obtained after an incubation of 2 to 7 days.

After the incubation, the bacteria are removed by centrifugation (or other methods compatible with the polymer and the bacteria), as a bacteria-free

solution of the polymer remains.

D. Determination of viscosity changes

The viscosity of the bacteria-free solution can be determined using the methods discussed above, such as e.g. with a Brookfield viscometer with an ultra-low viscosity cell that provides viscosity data (centipoise) at different shear rates. The viscosity of the sample solution is compared with the viscosity of the control solution. If the viscosity of the test solution is greater than that of the control solution, the bacterium can be considered to be able to increase the viscosity of a solution of that polymer.

Several bacteria were checked to determine

their effect on polymer solutions, the following general procedure being followed except where otherwise specifically indicated.

Preparation of a medium. A basal salt medium was prepared

in a double strength concentration for later dilution with the appropriate polymer solution. The solution of basal salts consisted of the following:

The exact composition of the basal salt medium is usually not critical and it can be changed depending on the requirements of the bacterium being tested. However, the same medium with basal salts should be used throughout a similar series of viscosity tests as the presence of salts can affect the viscosity properties of a polymer in solution.

CaCl2" 2^0 was autoclaved separately to avoid precipitation with the other components during heating. After both solutions had cooled to room temperature, CaCl2 • 2H20 was added to the other components. The medium with basal salts was checked for purity by spread an aliquot on a plate of nutrient agar and incubate the plate at 30°C for 24 hours.

Making a culture. Bacteria and salts to be controlled were grown on slant media of nutrient agar and Sabouraud dextrose agar, respectively. The bacteria were incubated at 30°C for 24 hours, aerobically and/or anaerobically depending on their physiological characteristics. Culture purity was determined by gram staining and microscopic examination. 5 ml of the basal salt medium was added to each slant culture and the organisms were removed from the surface by vortexing. 5 hundredths of ml of the cell suspension was used as inoculum. Preparation of the polymer solution. A certain amount of the polymer solution was added to deionized water to obtain a concentration of 2400 ppm (w/v). The polymer solution was adjusted to pH 7.0 and was steam sterilized at 1.05 kg/cm<2> and 121°C for 60 minutes. The solution was then tested for purity as described for the solution of the basal salts.

Incubation. The solution of basal salts and the polymer solution were mixed in a volume ratio of 1:1, whereby a 5 ml sample solution was obtained. 5 hundredths of ml of the bacterial suspension (the inoculum) was added to each of the three reagent tubes. The tubes were incubated for 14 days under conditions described for the inoculum preparation. After incubation, each tube was examined for turbidity, which is an indication of bacterial growth.

Intrinsic viscosity. Results for comparative changes in molecular weights were used for these experiments. Low molecular weight metabolic products were not removed and were assumed to make no contribution to viscosity. Because of the washing process for removing the bacterial cells from the polymer solution, any change in viscosity can be attributed to a change in the polymer itself and not the presence of biological residues. The temperature at the intrinsic viscosity determination was 25°C.

Examples 1-4

A gram-negative rod bacterium characterized as a strain of the genus Enterobacter was isolated from oil field produced water and assigned the accession number ATCC 39553. In previous laboratory experiments, this strain had shown the ability to use the partially hydrolyzed polyacrylamide, "Dow Pusher 500". as its only carbon source for growth.

The media used for this experiment were prepared

in two parts. A first polymer solution was prepared by dissolving commercially available Dow Pusher 500® in distilled water at a concentration of 2400 ppm (w/v) and pH 7.0. The second solution was prepared in the same way but after purification of the polymer by alcohol extraction. Both solutions were stirred for 24 hours, weighed before sterilization and then autoclaved (sterilized) for 1 hour at 121°C and 1.05 kg/cm 2. Any water lost due to autoclaving was replaced by adding sterile distilled water until the polymer solutions reached their original weight.

A solution of basal salts was prepared at a double concentration as explained above. It contained 0.02% BBL yeast extract, 0.2% KH2PO4, 0.2%

K2HP04, 0.1% (NH^ 2S04°g °'02% CaC12 ' 2H2° 1 distilled water at pH 7.0. CaCl2 was prepared as a separate pre-solution and added to the solution of salts after both had been sterilized and cooled to room temperature.

The two separate solutions of Dow Pusher 500 and basal salts for this experiment were prepared by aseptically mixing 1:1 solutions of basal salt medium (2X) and 2400 ppm (w/v) polymer solution.

The culture of Enterobacter sp. ATCC 39553 was grown aerobically on a nutrient agar slant (Difco) at 30°C for 24 hours. The cells were harvested by adding 5 ml of sterile basal saline to the culture slant in the test tube and gently vortexing the tube. This cell suspension was used as inoculum for this experiment.

All bottles were incubated at 30°C for 14 days. Aerobic incubation included rotary shaking of the bottles at 200 rpm. Anaerobic incubation was carried out in an anaerobic chamber system

The control solution was incubated both aerobically and anaerobically.

After incubation, all sample solutions and controls were centrifuged (10,000 x g, 30°C, 20 min) and the supernatant was decanted aseptically into clean, sterile sample bottles. The viscosities of the solutions were measured.

The molecular weight of "Dow Pusher 500" was predicted to be 3.4 x 10<6>, based on published data (F.D. Martin and J.S. Ward, Polymer Preprints, vol. 22, no. 2, American Chemical Society Meeting, August, 1981 , p. 24). The viscosity of the control solution was also assumed to be constant. Detailed data showing measured viscosity, intrinsic viscosity and calculated molecular weight for the polymer in control solution A and examples 1 and 2 are available in table 1.

Compared to the control, the organism causes an increase

in viscosity or stabilizes the purified polymer against decomposition during the 14 days. Under anaerobic conditions, Enterobacter sp increases. ATCC 39553 the apparent molecular weight of "Dow Pusher 500" from 3.4 x IO<6> to 5.7 x IO<6>, an increase of approx. 68%. Based on changes in intrinsic viscosity, molecular weight calculations were made.

Examples 5-8

The above procedures were repeated to determine changes in intrinsic viscosity as discussed above.

All conditions were the same except where noted.

Table 2 summarizes the intrinsic viscosity results for these solutions incubated with Enterobacter sp.ATCC 39553 for 2 weeks. Relative viscosity increases were found in all samples tested. The greatest increase in solution viscosity was found in the samples that had been incubated anaerobically. As no control was available to compare the effects of a purified polymer during anaerobic incubation, the effects of purification were considered to be secondary. Examples 2 and 6 were therefore compared with control C, the closest comparison.

The intrinsic viscosity data from Examples 1 to

4 and controls A and B are also shown in Table 2.

Examples 9 - 130

Eight bacterial strains were selected in addition to Enteroba cter sp. ATCC 39553 for testing. All had the ability to grow aerobically and/or anaerobically in different water-soluble polymers. The nine tribes are identified with the help of the following family names and deposition be<*>

Gram-negative rods: Gram-positive rods?

Gram-positive rods: Bacillus sp.ATCC 39556 The strains ATCC 39555, 39560 and 39558 were laboratory contaminants isolated from aqueous polymer solutions. All the other bacteria were isolated from water produced on oil fields and drilling mud.

All strains (with the exception of ATCC 39559) were grown aerobically in nutrient medium (Difco) at 30°C for 24 hours. Strain ATCC 39559 was incubated anaerobically at 30°C for 24 hours using nutrient solution containing thioglycolate.

After incubation, the bacteria were washed three times by centrifugation (5000 x g, 5°c, 10 min.) with sterile distilled water. The washed cells were resuspended in a minimal amount of sterile distilled water. Cell counts were performed on each cell suspension using a Petroff-Hausser counting chamber.

For this experiment, media were prepared in two parts. Polymer solutions were prepared in distilled water at a concentration of 2400 ppm (w/v) and pH 7.0. The polymers in Table 3 were used in the form in which they are purchased and after purification by extraction.

All solutions were stirred for 24 hours, weighed and then autoclaved (sterilized) for 1 hour at 121°C and 1.05 kg/cm 2 . Any water lost during the autoclaving was replaced by adding sterile distilled water until the polymer solutions reached its original weight.

A medium with basal salts was prepared as described in example 1. Eighteen different polymer media for this experiment were prepared by aseptically mixing 1:1 (volume/volume) solutions of medium with basal salts and 2400 ppm polymer solution. 50 ml aliquots of polymeric media were aseptically transferred to clean, sterile 150 ml Erlenmeyer flasks. Each flask (except the controls) was inoculated to a concentration of o lo 3 bacteria/ml with the relevant cell suspension. Controls were inoculated with equivalent volumes of sterile distilled water.

All flasks were incubated at 30°C for two weeks. Aerobic incubation included rotary shaking of the flasks at 200 rpm, and anaerobic incubation was carried out in an anaerobic chamber system. Controls were incubated aerobically.

After incubation, all sample solutions and controls were centrifuged (10,000 x g, 30°C, 20 min.) and the supernatant decanted aseptically into a clean, sterile sample bottle. Apparent viscosity determinations were made using a Brookfield viscometer. Table 4 shows the results from examples 9 - 130 and also includes data from examples 1 to 8.

Bacterial increase of viscosity in polymer solution

Several samples tested in this polymer/bacteria control showed increases in solution viscosity after 2 weeks of incubation. Some of the samples that showed bacterial increase of viscosity in polymer solution are reproduced in Table 5, grouped by the code designation of the bacterium.

Footnote to table 5

1 "A" means aerobic. "An" means anaerobic.

2 Intrinsic viscosity indicates the percentage change in the molecular weight of the polymer relative to the control.

3 "Dow Pusher 500". in

4 Purified by extraction with methanol.

7 Purified by extraction with acetonitrile.

The bacteria according to table 6 were before the administration

of the present patent application deposited with the American Type Culture Collection (ATCC) in Rockville, Maryland under an agreement whereby they will be available to the public after the issuance of the patent. Each bacterial culture deposited at ATCC has been assigned a unique number.

Claims (15)

1. Aqueous solution of pre-made polymers with increased viscosity properties, characterized in that it is produced by mixing, under conditions conducive to bacterial growth, an aqueous solution of pre-produced polymers with a bacterium selected from the group consisting of gram-negative rod bacteria, gram-positive rod bacteria and gram-positive globule bacteria, without the ability to re-synthesize the polymers, but with the ability to increase the viscosity of the polymer solution, as this ability has been demonstrated by means of the following screening procedure: (i) preparation of a culture of the bacterium, (ii) preparation of an aqueous solution of the previously prepared polymers and addition of nutrients and minerals required for bacterial growth, (iii) separating the solution into two parts, a test and a control part, where both parts lack substrate apart from the previously produced polymers and thereby lack a usable substrate to enable the synthesis of another polymer, (iv) inoculation of the test solution with the culture of the bacterium and incubation of the test and control solutions, and (v) measuring the viscosities in both test and control solutions gene after incubation to determine the viscosity of the test solution relative to the viscosity of the control solution. 2. Process for increasing the viscosity of a solution of pre-made polymers, characterized by mixing under conditions favoring bacterial growth, (a) an aqueous solution of pre-prepared polymers and (b) a bacterium selected from the group consisting of gram-negative rods, gram-positive rods and gram-positive globules incapable of re-synthesis of the polymers, but with the ability to increase the viscosity of the polymer solution in the following screening procedure: (i) preparation of a culture of the bacterium, (ii) preparation of an aqueous solution of the previously prepared polymers and addition of nutrients and minerals which is required for bacterial growth, (iii) separating the solution into two parts, a test and a control part, both parts lacking substrate apart from the pre-made polymers and thereby lacking a usable substrate to enable the synthesis of another polymer , (iv) inoculating the test solution with the culture of the microorganism and incubating the test and control solutions, and (v) must measuring the viscosities in both the test and control solution after the incubation to determine the viscosity of the test solution in relation to the viscosity of the control solution. 3. Method according to claim 2, characterized in that a bacterium selected from Enterobacter sp. is used. ATCC 39553, Enterobacter sp. ATCC 39555, Citrobacter sp. ATCC 39560, Pseudomonas sp. ATCC 39558, Pseudomonas sp. ATCC 39554, Staphylococcus sp. ATCC 39557, Staphylococcus sp. ATCC 39552, Bacillus sp.. ATCC 39556 and Peptococcus sp. ATCC 39559. 4. Method according to claim 2, characterized in that a polymer selected from the group consisting of polyacrylamide, hydrolyzed polyacrylamide, polyvinyl alcohol, xanthan gum, poly(2-acrylamido-2-methyl-propanesulfonic acid), hydroxyethylcellulose, poly(acrylic acid), carboxymethylcellulose and glucan. 5. Method according to claim 4, characterized in that a polymer selected from the group consisting of hydrolyzed polyacrylamide, polyvinyl alcohol, poly(2-acrylamido-2-methyl-propane-sulfonic acid), xanthan gum, poly (acrylic acid) and glucan is used. 6. Method according to claim 5, characterized in that a polymer selected from the group consisting of hydrolysed polyacrylamide, poly(acrylic acid) and glucan is used. 7. Method according to claim 6, characterized in that hydrolyzed polyacrylamide is used as polymer. 8. Method according to claim 6, characterized in that poly(acrylic acid) is used as polymer. 9. Method according to claim 6, characterized in that glucan is used as polymer. 10. Method according to claim characterized in that hydrolyzed polyacrylamide is used as polymer, and that a bacterium selected from the group consisting of Enterobacter sp. ATCC 39555, Pseudomonas sp. ATCC 39558, Pseudomonas sp. ATCC 39554 and Enterobacter sp. ATCC 39553. 11. Method according to claim 2, characterized in that polyvinyl alcohol is used as polymer, and that a bacterium selected from the group consisting of Enterobacter sp. ATCC 39555 and Pseudomonas sp, ATCC 39558. 12. Method according to claim 2, characterized in that poly(2-acrylamido-2-methyl-propanesulfonic acid) is used as polymer, and that a bacterium selected from the group consisting of Enterobacter sp. ATCC 39555, Staphylococcus sp. ATCC 39557 and Enterobacter sp. ATCC 39553. 13. Method according to claim 2, characterized in that xanthan gum is used as polymer, and that a bacterium selected from the group consisting of Enterobacter sp. ATCC 39555 and Bacillus sp. ATCC 39556. 14. Method according to claim 2, characterized in that glucan is used as polymer and that a bacterium selected from the group consisting of Enterobacter sp. ATCC 39555, Bacillus sp. ATCC 39556, Pseudomonas sp. ATCC 39558, Pseudomonas sp. ATCC 39554, Enterobacter sp. ATCC 39553 and Citrobacter sp. ATCC 39560. 15. Method according to claim 2, characterized in that poly(acrylic acid) is used as polymer, and that a bacterium selected from the group consisting of Pseudomonas sp. ATCC 39558 and Enterobacter sp. ATCC 39553.