CN113881594B - Optimized sulfate reducing bacteria culture medium and application thereof - Google Patents
Optimized sulfate reducing bacteria culture medium and application thereof Download PDFInfo
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- CN113881594B CN113881594B CN202111169520.1A CN202111169520A CN113881594B CN 113881594 B CN113881594 B CN 113881594B CN 202111169520 A CN202111169520 A CN 202111169520A CN 113881594 B CN113881594 B CN 113881594B
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- 241000894006 Bacteria Species 0.000 title claims abstract description 85
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 0.000 title claims abstract description 67
- 239000001963 growth medium Substances 0.000 title claims abstract description 40
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims abstract description 56
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 claims abstract description 41
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 claims abstract description 30
- 239000011593 sulfur Substances 0.000 claims abstract description 30
- 229910052717 sulfur Inorganic materials 0.000 claims abstract description 30
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims abstract description 28
- 229910052799 carbon Inorganic materials 0.000 claims abstract description 28
- 229910052757 nitrogen Inorganic materials 0.000 claims abstract description 28
- CSNNHWWHGAXBCP-UHFFFAOYSA-L magnesium sulphate Substances [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims abstract description 24
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 claims abstract description 18
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 18
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims abstract description 16
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims abstract description 15
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims abstract description 15
- 229930006000 Sucrose Natural products 0.000 claims abstract description 15
- 235000019270 ammonium chloride Nutrition 0.000 claims abstract description 15
- 239000008103 glucose Substances 0.000 claims abstract description 15
- LJCNRYVRMXRIQR-OLXYHTOASA-L potassium sodium L-tartrate Chemical compound [Na+].[K+].[O-]C(=O)[C@H](O)[C@@H](O)C([O-])=O LJCNRYVRMXRIQR-OLXYHTOASA-L 0.000 claims abstract description 15
- 235000011006 sodium potassium tartrate Nutrition 0.000 claims abstract description 15
- 239000005720 sucrose Substances 0.000 claims abstract description 15
- 229940041514 candida albicans extract Drugs 0.000 claims abstract description 14
- 239000012138 yeast extract Substances 0.000 claims abstract description 14
- 229940074439 potassium sodium tartrate Drugs 0.000 claims abstract description 13
- 239000001888 Peptone Substances 0.000 claims abstract description 12
- 108010080698 Peptones Proteins 0.000 claims abstract description 12
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 claims abstract description 12
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims abstract description 12
- 235000019341 magnesium sulphate Nutrition 0.000 claims abstract description 12
- 235000019319 peptone Nutrition 0.000 claims abstract description 12
- 239000000843 powder Substances 0.000 claims abstract description 11
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 claims abstract description 9
- 239000011668 ascorbic acid Substances 0.000 claims abstract description 9
- 235000010323 ascorbic acid Nutrition 0.000 claims abstract description 9
- 229960005070 ascorbic acid Drugs 0.000 claims abstract description 9
- IMBKASBLAKCLEM-UHFFFAOYSA-L ferrous ammonium sulfate (anhydrous) Chemical compound [NH4+].[NH4+].[Fe+2].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O IMBKASBLAKCLEM-UHFFFAOYSA-L 0.000 claims abstract description 9
- JVBXVOWTABLYPX-UHFFFAOYSA-L sodium dithionite Chemical compound [Na+].[Na+].[O-]S(=O)S([O-])=O JVBXVOWTABLYPX-UHFFFAOYSA-L 0.000 claims abstract description 9
- AKHNMLFCWUSKQB-UHFFFAOYSA-L sodium thiosulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=S AKHNMLFCWUSKQB-UHFFFAOYSA-L 0.000 claims abstract description 9
- 235000019345 sodium thiosulphate Nutrition 0.000 claims abstract description 9
- CYDQOEWLBCCFJZ-UHFFFAOYSA-N 4-(4-fluorophenyl)oxane-4-carboxylic acid Chemical compound C=1C=C(F)C=CC=1C1(C(=O)O)CCOCC1 CYDQOEWLBCCFJZ-UHFFFAOYSA-N 0.000 claims abstract description 8
- 239000011780 sodium chloride Substances 0.000 claims abstract description 8
- 239000001540 sodium lactate Substances 0.000 claims abstract description 8
- 235000011088 sodium lactate Nutrition 0.000 claims abstract description 8
- 229940005581 sodium lactate Drugs 0.000 claims abstract description 8
- 238000001514 detection method Methods 0.000 claims abstract description 7
- 239000000203 mixture Substances 0.000 claims description 38
- 238000012216 screening Methods 0.000 claims description 4
- 238000000926 separation method Methods 0.000 claims description 4
- AVTYONGGKAJVTE-OLXYHTOASA-L potassium L-tartrate Chemical compound [K+].[K+].[O-]C(=O)[C@H](O)[C@@H](O)C([O-])=O AVTYONGGKAJVTE-OLXYHTOASA-L 0.000 claims description 2
- 239000001476 sodium potassium tartrate Substances 0.000 claims description 2
- 229910021653 sulphate ion Inorganic materials 0.000 claims 4
- 239000005864 Sulphur Substances 0.000 claims 1
- RWSOTUBLDIXVET-UHFFFAOYSA-N Dihydrogen sulfide Chemical group S RWSOTUBLDIXVET-UHFFFAOYSA-N 0.000 abstract description 8
- 229910000037 hydrogen sulfide Inorganic materials 0.000 abstract description 8
- 230000000694 effects Effects 0.000 abstract description 6
- 230000009286 beneficial effect Effects 0.000 abstract description 5
- 238000012544 monitoring process Methods 0.000 abstract description 3
- 235000016709 nutrition Nutrition 0.000 abstract description 3
- 238000005536 corrosion prevention Methods 0.000 abstract description 2
- 230000000813 microbial effect Effects 0.000 abstract description 2
- 230000035764 nutrition Effects 0.000 abstract description 2
- 230000002265 prevention Effects 0.000 abstract description 2
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 abstract 1
- 229960001031 glucose Drugs 0.000 abstract 1
- 229960004793 sucrose Drugs 0.000 abstract 1
- 239000003921 oil Substances 0.000 description 21
- 238000000034 method Methods 0.000 description 18
- 230000001580 bacterial effect Effects 0.000 description 8
- 239000002609 medium Substances 0.000 description 7
- 238000012360 testing method Methods 0.000 description 7
- 230000033558 biomineral tissue development Effects 0.000 description 6
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Chemical compound C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 description 4
- 238000012258 culturing Methods 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 230000018109 developmental process Effects 0.000 description 3
- 244000005700 microbiome Species 0.000 description 3
- 239000010779 crude oil Substances 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 239000003345 natural gas Substances 0.000 description 2
- 229920000642 polymer Polymers 0.000 description 2
- 208000005374 Poisoning Diseases 0.000 description 1
- LSNNMFCWUKXFEE-UHFFFAOYSA-N Sulfurous acid Chemical compound OS(O)=O LSNNMFCWUKXFEE-UHFFFAOYSA-N 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- -1 compound sulfate Chemical class 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000006477 desulfuration reaction Methods 0.000 description 1
- 230000023556 desulfurization Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 235000013601 eggs Nutrition 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000007789 gas Substances 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- KAEAMHPPLLJBKF-UHFFFAOYSA-N iron(3+) sulfide Chemical compound [S-2].[S-2].[S-2].[Fe+3].[Fe+3] KAEAMHPPLLJBKF-UHFFFAOYSA-N 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 239000003208 petroleum Substances 0.000 description 1
- 231100000572 poisoning Toxicity 0.000 description 1
- 230000000607 poisoning effect Effects 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000013049 sediment Substances 0.000 description 1
- 230000035939 shock Effects 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- DHCDFWKWKRSZHF-UHFFFAOYSA-N sulfurothioic S-acid Chemical compound OS(O)(=O)=S DHCDFWKWKRSZHF-UHFFFAOYSA-N 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 239000002341 toxic gas Substances 0.000 description 1
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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- C12N1/02—Separating microorganisms from their culture media
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Abstract
The invention discloses an optimized sulfate reducing bacteria culture medium and application thereof. The culture medium comprises the following components: carbon source, nitrogen source, sulfur source, sodium chloride, anhydrous calcium chloride, ascorbic acid, ferrous ammonium sulfate, sodium hydrosulfite and the balance of water; the carbon source is composed of one or more than two of sodium lactate, glucose, sucrose or potassium sodium tartrate; the nitrogen source comprises the following components: one or more than two of yeast extract powder, peptone or ammonium chloride; the sulfur source comprises the following components: anhydrous sodium sulfate, magnesium sulfate, sodium thiosulfate or a combination of two or more of elemental sulfur. The sulfate reducing bacteria culture medium can meet the nutrition requirements of various sulfate reducing bacteria in the oil field environment, and the optimized culture medium is beneficial to improving the detection and monitoring capability of the activity of the sulfate reducing bacteria in the oil field, and is beneficial to the prevention and control of the secondary hydrogen sulfide problem and the microbial corrosion prevention and control of the oil field.
Description
Technical Field
The invention relates to the fields of microorganism quantity monitoring, microorganism detection and environmental microbiology. In particular to a culture medium which is suitable for sulfate reducing bacteria in oilfield produced water and is developed by optimizing the carbon source, nitrogen source and sulfur source nutritional ingredients in a compound sulfate reducing bacteria culture medium.
Background
The sulfate reducing bacteria can generate a large amount of H in the process of growth and propagation 2 S, which is the main cause of reservoir rancidity and presents a number of hazards to oil recovery: (1) Hydrogen sulfide is a colorless toxic gas with strong odor of the smelly eggs, and can cause harm to the health and even life of oilfield workers. 700ppm H 2 S can cause poisoning symptoms and even sudden shock death. 1000ppm of H 2 S is inhaled by people carelessly, so that people die immediately; (2) Hydrogen sulfide is acid gas, has strong corrosiveness to oil extraction pipelines, and can cause petroleum leakage, environmental pollution and economic loss when serious. (3) Metabolite H of sulfate-reducing bacteria 2 S can increase the sulfur content of crude oil and natural gas, reduce the quality and the oil reservoir value of the crude oil and the natural gas, and increase the subsequent desulfurization treatment cost; (4) In the chemical flooding process, sulfate reducing bacteria can cause the concentration of a polymer solution to be reduced, so that the polymer effect is affected, and the flooding technology is poor in application.
Sulfate-reducing bacteria in oil reservoirs have long been considered as the process of utilizing sulfur-containing compounds to grow and reproduce and produce large amounts of hydrogen sulfide, which is the main cause of rancidity in oil reservoirs. However, as the research is continued, many recent studies indicate that: 1. there are also some microorganisms in the reservoir environment that cannot utilize sulfate but can produce hydrogen sulfide. The method can utilize one or more sulfur-containing substrates such as elemental sulfur, sulfite, thiosulfate and the like to carry out growth and propagation and generate hydrogen sulfide; 2. the sulfate reducing bacteria have diversity, and the variety of the sulfate reducing bacteria among different oil fields is different.
Therefore, development of a culture medium which can effectively meet the growth, metabolism and propagation of various sulfate-reducing bacteria for different oil fields is needed. Further, the sulfate reducing bacteria which cause the oilfield spoilage can be deeply known, the number of the sulfate reducing bacteria in the oilfield environment can be effectively detected, the change of the number of the sulfate reducing bacteria can be monitored, and an effective control method for the oilfield spoilage can be found. Has important value for oil field development industry enterprises and is also an important challenge facing the oil field enterprises.
Disclosure of Invention
The invention aims to provide a culture medium suitable for growth and propagation of various sulfate reducing bacteria in an oilfield environment.
In order to achieve the above purpose, the invention adopts the following technical scheme: an optimized sulfate-reducing bacteria culture medium comprising the following components: carbon source, nitrogen source, sulfur source, sodium chloride 0.01-30 g/L, anhydrous calcium chloride 0.01-5 g/L, ascorbic acid 0.01-5 g/L, ferrous ammonium sulfate 0.01-5 g/L, sodium hydrosulfite 0.01-5 g/L and water balance.
The carbon source comprises the following components: sodium lactate 0.1-30 g/L, glucose 0.1-30 g/L, sucrose 0.1-30 g/L or potassium sodium tartrate 0.1-30 g/L.
The nitrogen source comprises the following components: one or more than two of yeast extract powder 0.01-15.0 g/L, peptone 0.01-15.0 g/L or ammonium chloride 0.01-15.0 g/L.
The sulfur source comprises the following components: anhydrous sodium sulfate 0.05-10 g/L, magnesium sulfate 0.1-20 g/L, sodium thiosulfate 0.01-10 g/L or elemental sulfur 0.01-5 g/L.
Preferably, the optimized sulfate reducing bacteria culture medium comprises the following carbon sources: glucose 0.45-1.35 g/L, sucrose 0.45-1.35 g/L and sodium potassium tartrate 1.1-3.3 g/L.
More preferably, the optimized sulfate reducing bacteria culture medium comprises the following carbon sources: glucose 0.90g/L, sucrose 0.90g/L, potassium sodium tartrate 2.2g/L.
Preferably, the above-mentioned optimized sulfate reducing bacteria culture medium, the nitrogen source composition is: yeast extract powder 0.25-0.75 g/L, peptone 0.25-0.75 g/L and ammonium chloride 0.5-1.5 g/L.
More preferably, the above-mentioned optimized sulfate reducing bacteria culture medium, the nitrogen source composition is: yeast extract powder 0.5g/L, peptone 0.5g/L, ammonium chloride 1.0g/L.
Preferably, the above-mentioned optimized sulfate reducing bacteria culture medium, the sulfur source composition is: anhydrous sodium sulfate 0.25-0.75 g/L, magnesium sulfate 0.7-2.1 g/L, sodium thiosulfate 0.1-0.3 g/L, and elemental sulfur 0.05-0.15 g/L.
More preferably, the optimized sulfate reducing bacteria culture medium comprises the following sulfur source components: the sulfur source comprises the following components: anhydrous sodium sulfate 0.75g/L, magnesium sulfate 2.1g/L, sodium thiosulfate 0.3g/L, and elemental sulfur 0.15g/L.
The sulfate reducing bacteria culture medium provided by the invention is applied to enrichment culture, separation screening and quantity detection of sulfate reducing bacteria.
The beneficial effects of the invention are as follows: the culture medium can realize enrichment culture, separation screening and quantity detection of various sulfate reducing bacteria. The culture medium can meet the growth, metabolism and propagation of various sulfate reducing bacteria in different oilfield environments. The sulfate reducing bacteria culture medium can meet the nutrition requirements of various sulfate reducing bacteria in the oil field environment, and the optimized culture medium is beneficial to improving the detection and monitoring capability of the activity of the sulfate reducing bacteria in the oil field, and is beneficial to the prevention and control of the secondary hydrogen sulfide problem and the microbial corrosion prevention and control of the oil field.
Detailed Description
Sulfate reducing bacteria counting method: and (3) carrying out gradient dilution on the water sample to be measured by using a sterile injector, and injecting the water sample into the sulfate reducing bacteria culture medium step by step, wherein three parallel samples are selected for each group. If black ferric sulfide sediment is generated at the bottom of the test bottle and is accompanied by hydrogen sulfide odor, positive reaction is indicated, and the sulfate reducing bacteria of the tested sample are counted by adopting an MPN method.
Example 1 Effect of optimized sulfate-reducing bacteria Medium (one) carbon Source composition on the Medium
1. Composition of sulfate-reducing bacteria medium:
carbon source, nitrogen source, sulfur source, sodium chloride 10g/L, anhydrous calcium chloride 0.1g/L, ascorbic acid 0.1g/L, ferrous ammonium sulfate 0.1g/L, sodium hydrosulfite 0.1g/L and water balance. Wherein,,
the composition of the carbon source is shown in Table 1.
The nitrogen source comprises the following components: 1.0g/L yeast extract powder and 1.0g/L ammonium chloride.
The composition of the sulfur source is as follows: anhydrous sodium sulfate 0.5g/L and magnesium sulfate 2.0g/L.
2. And (3) detecting sulfate reducing bacteria:
the method comprises the following steps: and measuring sulfate reducing bacteria in the water sample extracted from the northwest oil field. According to the composition of the sulfate reducing bacteria culture mediums of different carbon sources in step 1, the sulfate reducing bacteria culture mediums of different carbon sources are prepared by a strict anaerobic method, so that the mineralization degree, pH and temperature of the sulfate reducing bacteria culture mediums of different carbon sources are the same as the mineralization degree, pH and temperature of water samples taken from a northwest oilfield actually sampled. And (3) dripping water samples extracted from a certain northwest oil field into sulfate reducing bacteria culture mediums with different carbon sources, and culturing for 7 days. The sulfate-reducing bacteria of the test samples were counted by the MPN method, and the results are shown in Table 1.
Table 1: bacterial amount statistics in sulfate reducing bacteria culture media with different carbon sources
| Carbon source | Bacterial count (individual/ml) | |
| 1 | Sodium lactate 3.5g/L | 4.5×10 3 |
| 2 | Glucose 2.8g/L | 6.0×10 3 |
| 3 | Sucrose 2.67g/L | 4.5×10 3 |
| 4 | Potassium sodium tartrate 6.6g/L | 6.0×10 3 |
| 5 | Sodium lactate 1.2 g/L+glucose 0.90 g/L+sucrose 0.90g/L | 4.5×10 3 |
| 6 | Sodium lactate 1.2 g/L+glucose 0.90 g/L+potassium sodium tartrate 2.2g/L | 6.0×10 3 |
| 7 | Sodium lactate 1.2 g/L+sucrose 0.90 g/L+potassium sodium tartrate 2.2g/L | 6.0×10 3 |
| 8 | Glucose 0.90 g/L+sucrose 0.90 g/L+potassium sodium tartrate 2.2g/L | 7.5×10 3 |
As can be seen from Table 1, the most enriched bacteria were cultivated with the composition of group 8 carbon sources, and therefore, the preferred composition of carbon sources in the present invention is: glucose + sucrose + potassium sodium tartrate.
(di) Effect of Nitrogen Source composition on the Medium
1. Composition of sulfate-reducing bacteria medium:
carbon source, nitrogen source, sulfur source, sodium chloride 10g/L, anhydrous calcium chloride 0.1g/L, ascorbic acid 0.1g/L, ferrous ammonium sulfate 0.1g/L, sodium hydrosulfite 0.1g/L and water balance. Wherein,,
the composition of the carbon source is as follows: glucose 0.90 g/L+sucrose 0.90 g/L+potassium sodium tartrate 2.2g/L.
The nitrogen source composition is shown in table 2.
The composition of the sulfur source is as follows: anhydrous sodium sulfate 0.5g/L and magnesium sulfate 2.0g/L.
2. And (3) detecting sulfate reducing bacteria:
the method comprises the following steps: and measuring sulfate reducing bacteria in the water sample extracted from the northwest oil field. According to the composition of the sulfate reducing bacteria culture mediums of different nitrogen sources in step 1, the sulfate reducing bacteria culture mediums of different nitrogen sources are prepared by a strict anaerobic method, so that the mineralization degree, pH and temperature of the sulfate reducing bacteria culture mediums of different nitrogen sources are the same as the mineralization degree, pH and temperature of water samples extracted from a northwest oilfield actually sampled. And (3) dripping water samples extracted from a certain northwest oil field into sulfate reducing bacteria culture mediums with different nitrogen sources, and culturing for 7 days. The sulfate-reducing bacteria of the test samples were counted by the MPN method, and the results are shown in Table 2.
Table 2: bacterial quantity statistics in sulfate reducing bacteria culture medium with different nitrogen sources
| Nitrogen source | Bacterial count (individual/ml) | |
| 1 | Yeast extract powder 1.0g/L and ammonium chloride 1.0g/L | 7.5×10 3 |
| 2 | Peptone 1.0 g/L+ammonium chloride 1.0g/L | 9.5×10 3 |
| 3 | Yeast extract powder 0.5g/L, peptone 0.5g/L and ammonium chloride 1.0g/L | 1.15×10 4 |
As can be seen from Table 2, the group 3 nitrogen source composition, the most bacteria were enriched, and therefore, the preferred nitrogen source composition of the present invention is: yeast extract, peptone and ammonium chloride.
(III) Effect of Sulfur Source composition on the Medium
1. Composition of sulfate-reducing bacteria medium:
carbon source, nitrogen source, sulfur source, sodium chloride 10g/L, anhydrous calcium chloride 0.1g/L, ascorbic acid 0.1g/L, ferrous ammonium sulfate 0.1g/L, sodium hydrosulfite 0.1g/L and water balance. Wherein,,
the composition of the carbon source is as follows: glucose 0.90 g/L+sucrose 0.90 g/L+potassium sodium tartrate 2.2g/L.
The nitrogen source comprises the following components: yeast extract 0.5 g/L+peptone 0.5 g/L+ammonium chloride 1.0g/L.
The sulfur source composition is shown in table 3.
2. And (3) detecting sulfate reducing bacteria:
the method comprises the following steps: and measuring sulfate reducing bacteria in the water sample extracted from the northwest oil field. According to the composition of the sulfate reducing bacteria culture mediums of different sulfur sources in step 1, the sulfate reducing bacteria culture mediums of different sulfur sources are prepared by a strict anaerobic method, so that the mineralization degree, pH and temperature of the sulfate reducing bacteria culture mediums of different sulfur sources are the same as the mineralization degree, pH and temperature of water samples extracted from a northwest oilfield actually sampled. And (3) dripping water samples extracted from a certain northwest oil field into sulfate reducing bacteria culture mediums with different sulfur sources, and culturing for 7 days. The sulfate-reducing bacteria of the test samples were counted by the MPN method, and the results are shown in Table 3.
Table 3: bacterial quantity statistics in sulfate reducing bacteria culture medium of different sulfur sources
As can be seen from Table 3, the group 4 sulfur source composition, the number of bacteria enriched in culture is the greatest, and therefore, the preferred sulfur source composition of the present invention is: anhydrous sodium sulfate + magnesium sulfate + sodium thiosulfate + elemental sulfur.
(IV) performing orthogonal experiments with preferred carbon source composition, nitrogen source composition and sulfur source composition
Three-factor three-level design of carbon source composition, nitrogen source composition and sulfur source composition is shown in table 4, and the rest of the culture medium comprises 10g/L of sodium chloride, 0.1g/L of anhydrous calcium chloride, 0.1g/L of ascorbic acid, 0.1g/L of ferrous ammonium sulfate and 0.1g/L of sodium hydrosulfite.
TABLE 4 three factors three levels L 9 (3 4 ) Orthogonal test meter
Three factors and three levels L designed in Table 4 9 (3 4 ) Performing an orthogonal test, and obtaining from the test resultsAs is known, the maximum bacterial amount of sulfate reducing bacteria in a certain northwest oil field is measured by an MPN method by using an A2B2C3 culture medium, and the bacterial amount of the sulfate reducing bacteria reaches 4.5X10 5 And each ml. The composition of the optimized sulfate reducing bacteria culture medium is as follows: glucose 0.90g/L, sucrose 0.90g/L, potassium sodium tartrate 2.2g/L, yeast extract 0.5g/L, peptone 0.5g/L, ammonium chloride 1.0g/L, anhydrous sodium sulfate 0.75g/L, magnesium sulfate 2.1g/L, sodium thiosulfate 0.3g/L, elemental sulfur 0.15g/L, sodium chloride 10g/L, anhydrous calcium chloride 0.1g/L, ascorbic acid 0.1g/L, ferrous ammonium sulfate 0.1g/L, sodium hydrosulfite 0.1g/L.
(fifth) comparative example
Conventional culture medium was used: 2.5g/L of sodium lactate, 1.0g/L of yeast extract powder, 1.0g/L of ammonium chloride, 0.5g/L of anhydrous sodium sulfate, 2.0g/L of magnesium sulfate, 0.1g/L of anhydrous calcium chloride, 0.1g/L of ascorbic acid, 0.1g/L of ferrous ammonium sulfate and 0.1g/L of sodium hydrosulfite. Detecting sulfate reducing bacteria in northwest oil field, wherein the bacterial amount of the sulfate reducing bacteria is only 4.5 multiplied by 10 measured by MPN method 3 And each ml.
Therefore, the sulfate reducing bacteria culture medium can more effectively detect the quantity of sulfate reducing bacteria in the oil field environment. Is suitable for the application of enrichment culture, separation screening, quantity detection and the like of sulfate reducing bacteria, and has important value for oilfield development industry enterprises.
Claims (8)
1. An optimized sulfate reducing bacteria culture medium, characterized in that the sulfate reducing bacteria culture medium comprises the following components: carbon source, nitrogen source, sulfur source, sodium chloride 0.01-30 g/L, anhydrous calcium chloride 0.01-5 g/L, ascorbic acid 0.01-5 g/L, ferrous ammonium sulfate 0.01-5 g/L, sodium hydrosulfite 0.01-5 g/L and water balance;
the carbon source comprises the following components: 0.1 to 30g/L of sodium lactate, 0.1 to 30g/L of glucose, 0.1 to 30g/L of sucrose and 0.1 to 30g/L of potassium sodium tartrate;
the nitrogen source comprises the following components: one or more than two of yeast extract powder 0.01-15.0 g/L, peptone 0.01-15.0 g/L and ammonium chloride 0.01-15.0 g/L;
the sulfur source comprises the following components: 0.05 to 10g/L of anhydrous sodium sulfate, 0.1 to 20g/L of magnesium sulfate, 0.01 to 10g/L of sodium thiosulfate and 0.01 to 5g/L of elemental sulfur.
2. An optimised sulphate reducing bacteria culture medium according to claim 1 wherein the carbon source is of the composition: glucose 0.45-1.35 g/L, sucrose 0.45-1.35 g/L and sodium potassium tartrate 1.1-3.3 g/L.
3. An optimised sulphate reducing bacteria culture medium according to claim 2 wherein the carbon source is of the composition: glucose 0.90g/L, sucrose 0.90g/L, potassium sodium tartrate 2.2g/L.
4. An optimised sulphate reducing bacteria culture medium according to claim 1 wherein the nitrogen source is of the composition: yeast extract powder 0.25-0.75 g/L, peptone 0.25-0.75 g/L and ammonium chloride 0.5-1.5 g/L.
5. The optimized sulfate-reducing bacteria culture medium of claim 4, wherein the nitrogen source composition is: yeast extract powder 0.5g/L, peptone 0.5g/L, ammonium chloride 1.0g/L.
6. An optimised sulphate reducing bacteria culture medium according to claim 1 wherein the sulphur source is of the composition: anhydrous sodium sulfate 0.25-0.75 g/L, magnesium sulfate 0.7-2.1 g/L, sodium thiosulfate 0.1-0.3 g/L, and elemental sulfur 0.05-0.15 g/L.
7. The optimized sulfate-reducing bacteria culture medium of claim 6, wherein the sulfur source composition is: anhydrous sodium sulfate 0.75g/L, magnesium sulfate 2.1g/L, sodium thiosulfate 0.3g/L, and elemental sulfur 0.15g/L.
8. Use of the sulfate-reducing bacteria culture medium according to any one of claims 1-7 for enrichment culture, separation screening and quantity detection of sulfate-reducing bacteria.
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