GB2058106A - Heteropolysaccharide S-88 - Google Patents

Abstract

A heteropolysaccharide called S-88 is prepared by fermentation of an unnamed Pseudomonas species, ATCC 31554. The polysaccharide has valuable properties as a thickening, suspending and stabilizing agent in aqueous systems. It is especially useful in formulating oil well drilling fluids and muds. Its chemical composition is within the following weight ranges of sugars: 10- 20% glucuronic acid; 10-30% mannose; 30-40% glucose; 35-45% rhamnose; the polymer also contains 3-7% acetyl groups.

Description

SPECIFICATION Heteropolysaccharide S-88 Background of the Invention Compound S-88 may be prepared by fermentation of a suitable nutrient medium with a hitherto undescribed organism, based on extensive taxonomic studies, which is an unnamed Pseudomonas species.

An unrestricted permanent deposit of this organism employed in making our heteropolysaccharide was made with the American Type Culture Collection on August21, 1979 under Accession No ATCC 31554.

Various classification keys for the genus Pseudomonas and the culture descriptions of Pseudomonas species are found in the 7th Edition of Bergey's Manual (Breed et al., (1957)) and the 8th Edition of Bergey's Manual (Doudoroff et al., (1974)), as well as by other schools in various publications; Hugh and Gilardi, 1974, Pseudomonas Manual of Clinical Microbiology, 2nd ed., Lennette petal., Eds., pp.250-269. lizuka et 1963, Attempt at Grouping the Genus Pseudomonas, J. Gen. App!. Microbiology 9:73-82; and Hendric et al., 1966, Identification of Certain Pseudomonas Species, Identification Methods for Microbiologists, Part A, Gibbs et al., Eds., pp. 1-7, Academic Press, New York.

These keys and descriptions were searched for a Pseudomonas species having morphological and cultural characteristics similar to those of ATCC 31554. The following considerations make the assignment of a new Pseudomonas species justified and necessary.

Description ofthe Strain A. Characteristics of Colonial Morphology Two types (typical and atypical) of colonies appear on both nutrient and YM agar. On nutrient agar, small translucent yellow colonies develop within 2 days at ambient tempetature; diameter reaches about 1 mm in 5 days. The colonies are round, smooth-edged, and convex. Typical colonies are more glistening and the surface texture is extremely hard; entire colonies are removed if pushed by a loop; however, atypical ones do not possess such characteristics.

On YM agar, the diameter of both typical and atypical colonies are about 7.9 mm in 5 days. Both are round, smooth, convex, opaque and yellow-pigmented. Typical colonies become more umbonate-shaped and atypical ones are more flattened. Concentric rings are more significant in the case of atypical colonies. The surface of typical colonies is more slimy, and an elastic membrane is formed with prolonged incubation.

B. Characteristics of Cell Morphology Strain S-88 is a gram-negative rod-shaped bacterium. There is no difference in the cell shape between typical and atypical strains, except with cell arrangement. Cells of a typical strain are seen in aggregation, as in a Zoogloea-like mass. On nutrient agar, the size of most cells are 0.5-0.6 by 2.0-2.5 item; tapered at one end.

Few cells are motile. Poly-ss-hydroxybutyrate granules can be seen in the cells with prolonged incubation.

Occasionally very long abnormal cells have been seen. Pleomorphism is common. Most cells are disintegrated within 5 days at ambient temperature.

On YM medium, cells are larger and longer (0.6-0.8 by 2.5-3.0 zm) with consistently-shaped rods. Most cells are in the arrangement of palisades. Poly-ss-hydroxybutyrate granules were accumulated. Cells become very long ( > 5 um) with prolonged incubation.

Flagella stain was extremely difficult because relatively few cells were flagellated, and formation of gum hindered observation of mode of flagellation. According to the methods of Iniss and Mayfield and a modified silver nitrate method, the majority of flagellated cells were monotrichously flagellated; insertion was polar and/or sub-polar.

C. Physiological and Biochemical Characteristics Cytochrome-oxidase weak or negative; catalase positve. Organism is capable of growth at 37"C, but not at 4"C and 41"C. No tolerance to 3% NaCI and pHs 4 and 12. No survival at 600C for 30 minutes.

Aerobic; acid was produced frcm various carbohydrates, but not gas. Nitrate may be reduced. Litmus milk was reduced, but not peptonized. H2S was not produced (lead acetate method). Phosphatase and lipase were produced. Esculin and gelatin (very weakly) were hydrolyzed, but not starch and casein. ADH, LDC, and ODC were negative. Tolerant to 0.1% triphenyltetrazolium chloride. No growth on SS, Pseudosel, or MacConkey agar media. Acid was produced on slant; no growth in the butt of TSI medium.

D. Antibiotic Susceptibility Test The strain S-88 is susceptible to the following antibiotics: Carbenicillin . . (100 g) Gentamicin ... . .. . . (10 g) Chlortetracycline . ......

Penicillin ..... (10 units) Polymyxin B . . (300 units) Erythromycin .. . (15 g) Novobiocin. . (30 g) Kanamycin . . (30 g) Tetracycline . . (30 g) Neomycin ............. . .. (30 g) and not susceptible to: Streptomycin . . (10 g) Colistin . (10 g) E. Nutritional Characteristics Vitamins are not required for growth. Ammonium salts serve as sole nitrogen source. At least 26 out of 123 organic compounds are utilized as a sole source of carbon and energy; most are carbohydrates.

F. G+C Content of DNA The G+C moles % of the strain S-88 is 69.8.

G. Identification byAPland OXIIFERM Tube Systems The strain S-88 was not identified by either the API or OXI/FERM tube methods. This suggests that the organism could not be isolated from a clinical source.

H. Identification Since strain 5-88 is polar or sub-polalry monotrichously flagellated, the organism belongs to one of the members of genus Pseudomonas, jufging from the phenotypic characteristics. The G+C content of DNA (69.8 moles %) is in the range of the G+C moles % of the genus Pseudomonas.

TABLE 1 Biochemical and Other Miscellaneous Tests Employed for the Strain S-88 Oxidase: Kovac's + (weak) Hydrolysis of: Pathotech + (weak) Gelatin + (weak) Casein Catalase + Starch OF medium: Oxidative + Tween 80 + Fermentative - Pectin Gas from glucose - Alginate NT H2S production:TSI - Cellulose from cystine - Chitin Ammonium from peptone + DNA NT CB-Galactosidase (ONPG) - Esculin + Arginine dihydrolase Lysine decarboxylase - Growth on various media: Ornithinedecarboxylase - agar + Tryptophan deaminase NT MacConkey agar Phenylalanine deaminase - SS agar Urease - Mannitiol salt agar Indole - TCBSagar MR test Tinsdale tellurite VP test - blood agar Nitrate reduction - Pseudosel agar Nitrite reduction Denitrification NT Pigment production: N2-fixation: King A medium Growth in Burk's medium + King B medium Nitrogenase activity NT Malonate (oxidation) - Dye reaction:: Phosphatase + Congo red Haemolysis (sheep blood) - Nile blue NT Litmus milk: acid, reduction only 3-ketolactose production Survival at 60 C for 30 min. + TSI: Slant Acid Butt No Growth Gas Egg Yolk Reaction - = negative + = positive NT = not tested.

Fermentation Conditions: Heteropolysaccharide S-88 is produced during the aerobic fermentation of suitable aqueous nutrient media under controlled conditions via the inoculation with the organism of the unnamed Pseudomonas species. The media are usual media, containing source of carbon, nitrogen and inorganic salts.

In general, carbohydrates (for example, glucose, fructose, maltose, sucrose, xylose, mannitol and the like) can be used either alone or in combination as sources of assimilable carbon in the nutrient medium. The exact quantity of the carbohydrate source or sources utilized in the medium depend in part upon the other ingredients of the medium but, in general, the amount of carbohydrate usually varies between about 2% and 4% by weight of the medium. These carbon sources can be used individually, or several such carbon sources may be combined in the medium. In general, many proteinaceous materials may be used as nitrogen sources in the fermentation process. Suitable nitrogen sources include, for example, yeast hydrolysates, primary yeast, soybean meal, cottonseed flour, hydrolysates of casein, corn steep liquour, distiller's solubles or tomato paste and the like. The sources of nitrogen, either alone or in combination, are used in amounts ranging from about 0.5% to 0.2% by weight of the aqueous medium.

Among the nutrient inorganic salts which can be incorporated in the culture media are the customary salts capable of yielding sodium, potassium, ammonium, calcium, phosphate, sulfate, chloride, carbonate, and the like ions. Also included are trace metals such as cobalt, manganese, iron and magnesium.

It should be noted that the media described in the examples are merly illustrative of the wide variety of media which may be employed, and are not intended to be limitative.

One important media characteristic is that when strain S-88 is grown under low Ca++ conditions, i.e., in deionized water, or an aqueous system having less than 200 ppm Ca++ ions, the resultant gum is readily soluble in solutions without gelling.

The fermentation is carried out at temperatures ranging from about 25"C to 35"C; however, for optimum results it is preferable to conduct the fermentation at temperatures of from about 28"C. to 32"C. The pH of the nutrient media for growing the Pseudomonas culture and producing the polysaccharide S-88 can vary from about 6 to 8.

Although the polysaccharide S-88 is produced by both surface and submerged culture, it is preferred to carry out the fermentation in the submerged state.

A small scale fermentation is conveniently carried out by inoculating a suitable nutrient medium with the culture, and aftertransferto a production medium permitting the fermentation to proceed at a constant temperature ofabout30cC on a shaker for several days.

The fermentation is initiated in a sterilized flask of medium via one or more stages of seed development.

The nutrient medium for the seed stage may be any suitable combination of carbon and nitrogen sources.

The seed flask is shaken in a constant temperature chamber at about 30"C for 1-2 days, or until growth is satisfactory, and some of the resulting growth is used to inoculate either a second stage seed or the production medium. Intermediate stage seed flasks, when used, are developed in essentially the same manner; that is, part of the contents of the flask from the last seed stage are used to inoculate the production medium. The inoculated flasks are shaken at a constant temperature for several days, and at the end of the incubation period the contents of the flasks are recovered by precipitation with a suitable alcohol such as isopropanol.

For large scale work, it is preferable to conduct the fermentation in suitable tanks provided with an agitator and a means of aerating the fermentation medium. According to this method, the nutrient medium is made up in the tank and sterilized by heating at temperatures of up to about 121"C. Upon cooling, the sterilized medium is inoculated with a previously grown seed of the producing culture, and the fermentation is permitted to proceed for a period of time as, for example, from 2 to 4 days while agitating and/or aerating the nutrient medium and maintaining the temperature at about 30"C. This method of producing the S-88 is particularly suited for the preparation of large quantites.

The product is recovered from the fermentation medium by precipitation with a suitable alchohol, such as isopropanol.

Heteropolysaccharide S-88 The heteropolysaccharide produced by an unmamed Pseudomonas species is composed principally of carbohydrate with 3-7% acetyl groups as the O-glycosidically linked ester.

The carbohydrate portion of the 5-88 polysaccharide contains 10-20% glucouronic acid; 10-30% mannose; 30-40% glucose; and 35-45% rhamnose.

Table II indicates specific analysis results on six fermentation samples, including triplicate analysis of two samples.

TABLE II Composition of S-88 Gum PROPRIETARY ORGANISMS Sample Glucouronic Pyruvate Acetyl Neutral Sugar Components (%) No. aAcid (%) p (%) Glu Man Fuc Rha Ara Protein (%) 1 11.8 0.1 3.2 33 22 -- 43 2 11.8 2 18.7 0.1 3.2 38 22 -- 39 1 11.9 3 18.0 0.1 7.0 38 24 -- 37 1 11.5 4 15.8 0.1 6.6 35 27 -- 36 2 11.7 5A 15.1 0.1 6.1 33 25 3 37 2 12.7 5B 15.1 0.1 6.1 38 19 -- 41 2 12.7 5C 15.1 0.1 6.1 37 23 1 37 2 12.7 6A 13.7 0.1 4.9 36 22 1 40 1 14.0 6B 13.7 0.1 4.9 41 13 1 44 1 14.0 6C 13.7 0.1 4.9 38 20 0 41 1 14.0 by by decarboxylation by colorimetric analysis The acetyl content of 5-10% was determined by treating a 0.2% aqueous solution of S-88 gum with an alkaline, hydroxylamine reagent followed by treatment with an acidic ferric chloride reagent [S. Hestrin (1949) J. Biol.

Chem. 180 pp. 249-261].

The neutral sugars of polysaccharide S-88 were determined by dissolving ten mg. of the product in 2 ml 2N H2SO4, and the mixture is heated at 100 C for 4 hours. The resulting solution is cooled, neutralized with barium hydroxide and the pH is brought to 5-6 with solid carbon dioxide. The resulting precipitate of barium sulfide is removed by centrifugation and the supernatent is concentrated to a syrup under reduced pressure.

The sugars in the hydrolysate are tentatively identified by gas-liquid chromatography of their aldononitrile acetate derivatives on a Hewlett-Packard Model 5750 chromatograph using 3% by weight OV-225 on 80/100 mesh Gas Chrom Oat 210 C. The sugars are identified and quantitated by comparison with authentic standards [J.K. Baird, M.J. Holroyde, and D.C. Ellwood (1973) Carbohydr. Res. 27 pp.264-467].

The various neutral sugars of the polysaccharides were also characterized by use of descending paper chromatography on Whatman No. 1 chromatography paper using as the solvent the upper layer of pyridine:ethyl acetate:water (2:5:5). Chromatogramswere stained using silver nitrate dip and acid analine phthalate spray reagent. Component sugars were identified by co-chromatography with sugar standards and by the specific-color reaction with the analine phthalate reagent.

The glucouronic acid content of the polysaccharide was determined by two separate methods. In one method the sample was decarboxylated with 19% hydrochloric acid and the liberated carbon dioxide was trapped in standard sodium hydroxide and determined by back titration [B.L. Browning (1967) Methods of Wood Chemistry 11, pp. 632-633] and by the carbazole colorimetric method [T. Bitter and H.M. Muir (1962) Anal. Biochem. 4 pp.330-334].

Paper electrophoresis was used for the separation and tentative identification of the glucouronic acid present in the neutralized acid hydrolysate described above. Aliquots of this and known glucouronic acid standards were applied to Camag eiectrophoresis paper No. 68-011 and electrophoresis was carried out for 2.0 hours in a pH 2.7 buffer using a Camag Model HVE electrophoresis apparatus. Chromatograms were air dried and stained with silver nitrate dip reagent to locate the glucouronic acids being separated.

The polysaccharide S-88 imparts viscosity to an aqueous medium when dissolved in water in low concentrations. Because of this, its sensitivity to shear and overall rheology, it is useful as a thickening, suspending, emulsifying, stabilizing, lubricating, film-forming, or binding agent, especially in aqueous systems.In particular, it has uses in the following applications or products: adhesives, wall-joint cements, water-retentive grouts and mortars, spackling compounds, can sealing, boiler compounds, latex creaming, welding-rod fluxes, brazing pastes, ceramic glazes and extrusions, cleaners and polishes, toys, emusions (latex, asphalt, silicone), silver recovery, seed coatings, spray control for pesticides or herbicides, emulsifiable concentrated and flowable pesticides and herbicides, tobacco binders, water-based inks lithographic fountain solutions, leather finishes, hydro-mulching and hydro-seeding, textile printing and finishing, wet-end paper additives, wet-end paper retention and formation aid, anit-stick compounds, mold-release agents, liquid resins, slurry and packaged explosives, petroleum and water-well drilling muds, petroleum workover and completion fluids, petroleum stimulation fluids, cosmetics, pharmaceutical suspensions and emulsions.

Also this gum has utility in food systems such as jellies and other high sugar systems, beverages including citric acid based drinks, dairy products including ice cream and yogurt, salad dressings, dry mixes, icings, and glazes, syrups, puddings, farinaceous foods, canned and retorted foods, and bakery fillings.

A particularly valuable utility is in the field of petroleum and water-well drilling muds. More detailed examples illustrating this preferred use are found, infra.

Although S-88 gum possesses a general viscosity-imparting property, its particular profile of solution properties is a distinctive characteristic which enables it to be distinguished over other heteropolysaccharides.

In its dry form, the gum has a solids content of 85-90%, its most useful mesh size is 100% through 40 mesh, with no more than 30% through 325 mesh. It has a 1200-2500 cP at 1%, Brookfield, 60 rpm, and the following dial readings at 0.4 O concentration in 2% KCI tap water, Fann 35 (F = 02): 600 rpm > 120; 300 rpm > 92; 200 rpm > 76; 100 rprn > 70; 6 rpm > 32; and 3 rpm > 28.

The gum further has excellent heat stablility, and no viscosity loss upon autoclaving at 121"C and 15 psi for 15-20 minutes. It produces a firm gel with 15% NaOH and incubation at 80"C for two hours. It is incompatible with saturated CaCI2, ammonium polyphosphate, and 60% NH4NO3.

In addition, the gum has the following profile of properties: 1. Viscosity and Shear A. Brookfieldl 1. 1.0% 60 rpm 1520 cPs 6 rpm 11,600 cPs Spindle No. 3 2. O.lO/o(ULadapter)a 29.5 cPs 3. 0.5 O Wells-Brookfield ea 9.6 sec -' 474 cPs B. Shearb 1. n@1.92sec-1 8512 cPs 2. n@9.6sec-1 2342 cPs 3. n @ 76.8 seed 320 cPs 4. n@384sec-1 64 cPs 5. n@384sec-1 64 cPs 6. n @ 9.6 sec-1 1958 cPs C. 40 C Storage 1450 cPs, No. 3 spindle a 60 rpm, no gelation, chunky flow.

2. Acid, Base, Heat Stability A. Stability 1. Acetic acid plus heat-- a. initialn: 1620cPs b. finaln: 2450cPs c. % change: +51% (gel-like) 2. 10% HCI plus heat- a. initialn: PptcPs b. finaln: PptcPs c. %change: Ppt% 3. 15% NaOH plus heat- a. initial n: 1869 cPs b. final n: Gel cPs 4. Heat only - a. initial n: 2061 cPs b. finaln: 2266cPs c. % change: + 10% B. pHEffect 1. 5%AceticAcid 2.69 pH 1741 cPsC 2. 5%NH4OH 10.99pH 1741 cPsC 3. S-88 solution is stable over the pH range 1.25-12.3 3. Salt and Dye Compatibility A. Salt 1. CaCl2(saturated) Precipitate 2. Amm. polyphosphate Precipitate 3. 60% NH4NO3 Precipitate 4. 1%Al2(SO4)3.18H2O Compatible 5. 1%CaCl2.2H2O Compatible 6. 1%KCl Compatible 7. 0.1%KCl 2214 cPsc 8. 2.5%KCl 922cPsc B. Dyes 1.Milling green Compatible 2. Methylene Blue Precipitate 4. Texture/Flow Properties Chunky, non- continuous flow, high viscosity, gel-like, gummy to the touch.

5. Synergism and EnzymesC Ohrnof 2hrnof 1 % n mixture mixture A. Guar 1600 cPs 1702 cPs 1741 cPs B. H.P. Guar 1715cPs 1741 cPs 1741 cPs C. CMC 832 cPs 1088 cPs 1665 cPs D. HEC 602 cPs 1229 cPs 998 cPs E. MAS 1-1 2253 cPs Expected Viscosity Synergism A. Guar 1900 cPs None B. H.P. Guar 1975 cPs None C. CMC 1375 cPs +21 D. HEC 1175cPs None 6. Milk Reactivity A. Dispersion: Poor-- small granular precipitate B. Whey off: 1 sot day C. Other observations: 7. Film Formation Gum pulls down unevenly, film formed, uneven consistency, slightly plastic, brittle.

aViscosity measured on a Brookfield Model LVF at 6 rpm with the No. 1 spindle and a UL adapter.

bAll measurements made on Wells-Brookfield micro-viscometer Model RVT-c/p.

CViscosity measured on a Wells-Brookfield micro-viscometer Model RVT-c/p at 9.6 sec~ Example 1 Fermentation Procedure for Producing Heterop olysaccharide S-88 A. Culture Maintenance The unnamed Pseudomonas organism, ATCC 31554, grows quite well on NA agar at an incubation temperature of 30"C. This organism produces a yellow carotenoid pigment. The colonies on NA are small (only 1-3 mm) by 48 hrs., are convex, and have a gelatinous texture. The typical colony has a tendency to stick tenaciously to the agar surface, Occasionally, a morphological variant may develop which is easy to spot on NA. The variant has a flat colony and does not stick tenaciously to the agar surface. This variant was found to have a decreased activity of S-88 gum production.

B. Seed Preparation Flask seeds are prepared in YM broth incubated at 30"C.

In this medium the culture will typically give flocculant-type growth followed by viscosity increases with a granular-type appearance. The YM seeds are then used at 24-30 hrs. to inoculate seed medium which is the same as final fermentor medium, except that the phosphate concentration is increased to 0.5%. One-gallon fermentors are used as seed vessels for the 20L and 70L fermentors.

C. Final Fermentor Medium The following medium gives acceptable results in both 20L and 70L fermentors.

Glucose . 3.0 % K2HPO4 0.05% AMP 0.05% NH4NO3 0.09% MgSO4.7H2O 0.01% Fe++ 1 ppm HoLe salts 1 ml L An agitation rate set at 500 rpm in both the 20L and 70Lfermentors is desirable. Fermentation times can range from 45-70 hrs. with beer viscosity ranging from 3000 cps to 5000 cps. Conversion efficiencies vary from 31-52% with 3% glucose. Small amounts of commercially available antifoam agent can be used.

Gram stains made from S-88 fermentation beer showed gram-negative cells approximately 1 .25it x 2.Sit in size with dark staining polar bodies.

HoLe salts are a trace element solution containing tartrate, magnesium molybdate, CoCI3, ZnCl2, CuC12, boric acid, manganese chloride and ferrous sulfate.

When a low calcium product is desired, a 30L fermentor medium is as follows: 30L Fermentor Medium for Low-Calcium Deionized water Glucose . 3.0 % K2HPO4 . ..... ............... 0.05% AMP .......... . 0.05% MgSO4.7H2O . ..... ....... 0.02% NH4NO3 ..... ..... ............. 0.09% Yeast extract ........ ...... ...... 0.01% HoLe salts . . ..... ........... . 40ml Vitamin mix . ..... ... 25ml Fe++ ....... ...:::. . 1 ppm Ca++ . ..... ...... 2 ppm The vitamin mixture is a mixture of 1 ulmI each of thiamine, cyanocobalamin, patothenate, riboflavin, nicotinic acid, choline, and pyridoxamine; 0.05 pIml folic acid and p-aminobenzoic acid; and 0.005 p/ml biotin.

D. Recovery Fermentation beer is pasteurized at 167 F for 10-15 min. Due to the excellent heat stability exhibited by this product, higher pasteurization temperatures with shorter holding times should be acceptable. Good fibers are typically produced under precipitation conditions giving 58-60% spent IPA.

E. Drying All product recovered thus far has been dried at 50-55 C for up to one hour in a forced-air tray dryer.

The properties of S-88 gum produced during these fermentation conditions have been given above.

When the low calcium S-88 gum is prepared, it shows a similar chemical analysis and property profile to S-88, but has an unusual response in NaCI solution. Although the initial viscosity is 0% is slightly low, a stable viscosity is maintained up to 6%, see Table Ill.

TABLE Ill Salt Response: NaCI NaCI Concentration, wt. % 0 2 4 6 8 10 15 20 600" 13.4 14.2 14.8 13.8 11.4 10.2 9.5 9.0 300" 20.1 21.4 22.0 20.6 20.2 19.7 19.4 19.0 200 26.0 26.1 27.1 25.4 24.3 24.3 23.8 23.6 100" 41.7 42.0 42.3 41.2 40.6 39.6 39.1 38.5 332 332 339 342 326 289 276 254 232 578 577 586 563 510 482 441 410 viscosity, cP at Fann 35 rpm Formulations using S-88 As noted above, S-88 gum, both in regular or calcium form, can be used in fresh- or salt-water drilling muds.

A typical formulation for a fresh-water mud is as follows: S-88 .. 1.01 lbs Bentonite . . 10.0 lbs Fresh Water . 1.0 bbl Fann Viscosity Data: Speed (rpm) 3 6 100 200 300 600 Dial Reading 7.6 7.9 16.8 21.5 25.6 35.3 pH = 8.3; API Filtrate = 12.0 ml Another formula, for a salt-water mud, is: S-88 . . . . 1.0 lb Sea Water . . 1.Obbl Fann Viscosity Data: Speed (rpm) 3 6 100 200 300 600 Dial Reading 2.8 3.2 9.0 12.8 15.6 23.6 pH = 7.2

Claims (4)

1. Heteropolysaccharide S-88, which contains 10-20% glucouronic acid; 10.30% mannose; 30-40% glucose; 35-45% rhamnose; and 3-7% acetyl groups, prepared by fermentation under controlled conditions of culture ATCC 31554, a Pseudomonas species.

2. The compound claimed in claim 1, containing less than 200 ppm Ca++ ions.

3. The compound claimed in claim 1 or 2, for use as a thickening, suspending, emulsifying, stabilizing, lubricating, film-forming, or binding agent in an aqueous system.

4. A drilling med containing the compound claimed in claim 1 or 2.