CN114084967A - Ganoderma lucidum fermented Shuanghuanglian mushroom residue for repairing water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene and application thereof - Google Patents
Ganoderma lucidum fermented Shuanghuanglian mushroom residue for repairing water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene and application thereof Download PDFInfo
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- CN114084967A CN114084967A CN202111336921.1A CN202111336921A CN114084967A CN 114084967 A CN114084967 A CN 114084967A CN 202111336921 A CN202111336921 A CN 202111336921A CN 114084967 A CN114084967 A CN 114084967A
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- Prior art keywords
- polycyclic aromatic
- aromatic hydrocarbon
- pyrene
- ganoderma
- ganoderma lucidum
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- FMMWHPNWAFZXNH-UHFFFAOYSA-N Benz[a]pyrene Chemical compound C1=C2C3=CC=CC=C3C=C(C=C3)C2=C2C3=CC=CC2=C1 FMMWHPNWAFZXNH-UHFFFAOYSA-N 0.000 title claims abstract description 124
- 125000005575 polycyclic aromatic hydrocarbon group Chemical group 0.000 title claims abstract description 71
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 title claims abstract description 64
- 239000009254 shuang-huang-lian Substances 0.000 title claims abstract description 15
- 240000008397 Ganoderma lucidum Species 0.000 title claims description 53
- 235000001637 Ganoderma lucidum Nutrition 0.000 title claims description 47
- 235000001674 Agaricus brunnescens Nutrition 0.000 title abstract description 24
- 241000222336 Ganoderma Species 0.000 claims abstract description 37
- 102000004190 Enzymes Human genes 0.000 claims abstract description 33
- 108090000790 Enzymes Proteins 0.000 claims abstract description 33
- 239000001963 growth medium Substances 0.000 claims abstract description 26
- 244000005700 microbiome Species 0.000 claims abstract description 20
- 238000000855 fermentation Methods 0.000 claims abstract description 17
- 230000004151 fermentation Effects 0.000 claims abstract description 17
- 239000007788 liquid Substances 0.000 claims abstract description 13
- 239000002994 raw material Substances 0.000 claims abstract description 4
- 241000233866 Fungi Species 0.000 claims description 15
- 239000007787 solid Substances 0.000 claims description 15
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 12
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 12
- 238000012258 culturing Methods 0.000 claims description 8
- 238000011218 seed culture Methods 0.000 claims description 8
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 claims description 6
- 240000008042 Zea mays Species 0.000 claims description 6
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 claims description 6
- 235000002017 Zea mays subsp mays Nutrition 0.000 claims description 6
- 235000005822 corn Nutrition 0.000 claims description 6
- 235000013312 flour Nutrition 0.000 claims description 6
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 6
- 235000019341 magnesium sulphate Nutrition 0.000 claims description 6
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 6
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 6
- 229910052757 nitrogen Inorganic materials 0.000 claims description 6
- 229910052698 phosphorus Inorganic materials 0.000 claims description 6
- 239000011574 phosphorus Substances 0.000 claims description 6
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 claims description 6
- LWIHDJKSTIGBAC-UHFFFAOYSA-K potassium phosphate Substances [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 claims description 6
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 claims description 5
- 239000011591 potassium Substances 0.000 claims description 5
- 229910052700 potassium Inorganic materials 0.000 claims description 5
- 229930006000 Sucrose Natural products 0.000 claims description 4
- 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 description 4
- 229930003270 Vitamin B Natural products 0.000 claims description 4
- 230000002378 acidificating effect Effects 0.000 claims description 4
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- 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 description 2
- 244000046052 Phaseolus vulgaris Species 0.000 claims description 2
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- 229940088598 enzyme Drugs 0.000 description 31
- 238000000034 method Methods 0.000 description 15
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 12
- 108010059896 Manganese peroxidase Proteins 0.000 description 11
- 108010029541 Laccase Proteins 0.000 description 10
- 108010054320 Lignin peroxidase Proteins 0.000 description 9
- 238000002474 experimental method Methods 0.000 description 9
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 8
- 238000001514 detection method Methods 0.000 description 7
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical group CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 6
- 229920005610 lignin Polymers 0.000 description 6
- 235000015097 nutrients Nutrition 0.000 description 6
- 239000002957 persistent organic pollutant Substances 0.000 description 6
- 239000000126 substance Substances 0.000 description 6
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- 239000007791 liquid phase Substances 0.000 description 4
- 239000012452 mother liquor Substances 0.000 description 4
- IZUPBVBPLAPZRR-UHFFFAOYSA-N pentachlorophenol Chemical compound OC1=C(Cl)C(Cl)=C(Cl)C(Cl)=C1Cl IZUPBVBPLAPZRR-UHFFFAOYSA-N 0.000 description 4
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- KAKLDTSBHNFICH-UHFFFAOYSA-N 1,1,2,2,3,3,4,4,5-nonachloro-5-(2,2,2-trichloro-1-phenylethyl)cyclohexane Chemical compound ClC1(C(C(C(C(C1)(C(C(Cl)(Cl)Cl)C1=CC=CC=C1)Cl)(Cl)Cl)(Cl)Cl)(Cl)Cl)Cl KAKLDTSBHNFICH-UHFFFAOYSA-N 0.000 description 2
- 241000193830 Bacillus <bacterium> Species 0.000 description 2
- 241000186146 Brevibacterium Species 0.000 description 2
- 241001467578 Microbacterium Species 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
- OHDRQQURAXLVGJ-HLVWOLMTSA-N azane;(2e)-3-ethyl-2-[(e)-(3-ethyl-6-sulfo-1,3-benzothiazol-2-ylidene)hydrazinylidene]-1,3-benzothiazole-6-sulfonic acid Chemical compound [NH4+].[NH4+].S/1C2=CC(S([O-])(=O)=O)=CC=C2N(CC)C\1=N/N=C1/SC2=CC(S([O-])(=O)=O)=CC=C2N1CC OHDRQQURAXLVGJ-HLVWOLMTSA-N 0.000 description 2
- 230000001580 bacterial effect Effects 0.000 description 2
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- LHGVFZTZFXWLCP-UHFFFAOYSA-N guaiacol Chemical compound COC1=CC=CC=C1O LHGVFZTZFXWLCP-UHFFFAOYSA-N 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
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- 239000008055 phosphate buffer solution Substances 0.000 description 2
- 238000002203 pretreatment Methods 0.000 description 2
- 238000012216 screening Methods 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
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- 241000205585 Aquilegia canadensis Species 0.000 description 1
- 241000186063 Arthrobacter Species 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- 206010007269 Carcinogenicity Diseases 0.000 description 1
- 108010059892 Cellulase Proteins 0.000 description 1
- 241000037740 Coptis chinensis Species 0.000 description 1
- 235000019750 Crude protein Nutrition 0.000 description 1
- 101710121765 Endo-1,4-beta-xylanase Proteins 0.000 description 1
- 241000555682 Forsythia x intermedia Species 0.000 description 1
- 241000282414 Homo sapiens Species 0.000 description 1
- 240000001462 Pleurotus ostreatus Species 0.000 description 1
- 235000001603 Pleurotus ostreatus Nutrition 0.000 description 1
- 240000004534 Scutellaria baicalensis Species 0.000 description 1
- 235000017089 Scutellaria baicalensis Nutrition 0.000 description 1
- FEWJPZIEWOKRBE-UHFFFAOYSA-N Tartaric acid Natural products [H+].[H+].[O-]C(=O)C(O)C(O)C([O-])=O FEWJPZIEWOKRBE-UHFFFAOYSA-N 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- ZTOJFFHGPLIVKC-CLFAGFIQSA-N abts Chemical compound S/1C2=CC(S(O)(=O)=O)=CC=C2N(CC)C\1=N\N=C1/SC2=CC(S(O)(=O)=O)=CC=C2N1CC ZTOJFFHGPLIVKC-CLFAGFIQSA-N 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000000845 anti-microbial effect Effects 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 230000000711 cancerogenic effect Effects 0.000 description 1
- 125000004432 carbon atom Chemical group C* 0.000 description 1
- 231100000315 carcinogenic Toxicity 0.000 description 1
- 231100000260 carcinogenicity Toxicity 0.000 description 1
- 230000007670 carcinogenicity Effects 0.000 description 1
- 229940106157 cellulase Drugs 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000003344 environmental pollutant Substances 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 229960001867 guaiacol Drugs 0.000 description 1
- 238000003306 harvesting Methods 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 125000004435 hydrogen atom Chemical group [H]* 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
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- 108010062085 ligninase Proteins 0.000 description 1
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- 230000007886 mutagenicity Effects 0.000 description 1
- 231100000299 mutagenicity Toxicity 0.000 description 1
- HGASFNYMVGEKTF-UHFFFAOYSA-N octan-1-ol;hydrate Chemical compound O.CCCCCCCCO HGASFNYMVGEKTF-UHFFFAOYSA-N 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
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- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G18/00—Cultivation of mushrooms
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G18/00—Cultivation of mushrooms
- A01G18/20—Culture media, e.g. compost
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
- C02F3/342—Biological treatment of water, waste water, or sewage characterised by the microorganisms used characterised by the enzymes used
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
- C02F3/348—Biological treatment of water, waste water, or sewage characterised by the microorganisms used characterised by the way or the form in which the microorganisms are added or dosed
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2101/00—Nature of the contaminant
- C02F2101/30—Organic compounds
- C02F2101/32—Hydrocarbons, e.g. oil
- C02F2101/327—Polyaromatic Hydrocarbons [PAH's]
Abstract
The invention relates to lucid ganoderma fermentation Shuanghuanglian mushroom dregs for repairing water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene and application thereof, which can improve the metabolic activity of polycyclic aromatic hydrocarbon degrading bacteria in the environment and degrade polycyclic aromatic hydrocarbon by influencing the survival and degradation performance of microorganisms in the environment, repair the water environment polluted by polycyclic aromatic hydrocarbon BaP by extracellular enzyme and adsorption capacity contained in the mushroom dregs, effectively realize the protection problem of the water environment, inoculate lucid ganoderma seed liquid into Shuanghuanglian mushroom dregs culture medium, culture and collect lucid ganoderma, the remained culture medium residue is lucid ganoderma mushroom dregs, the lucid ganoderma mushroom dregs are added into the water body polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene in an acid-base environment with pH of 4.0-8.5 and at the temperature of 20-50 ℃ by weight volume of 5-10 percent, remove polycyclic aromatic hydrocarbon benzo [ a ] pyrene in the water body with the polycyclic aromatic hydrocarbon benzo [ a ] pyrene pollution concentration of 50mg/L, the invention has the advantages of rich raw materials, easy obtainment of mushroom dregs, convenient use and good effect, and is a great innovation in repairing the water body polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene.
Description
Technical Field
The invention relates to the technical field of water environment protection, in particular to ganoderma lucidum and coptis chinensis fermentation residues for repairing a water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene and application thereof.
Background
Polycyclic Aromatic Hydrocarbons (PAHs) are a very typical organic pollutant, are composed of two or more than two carbon atoms and hydrogen atoms, have high hydrophobicity, high stability and high intractable, and seriously threaten the health of human beings due to toxicity, mutagenicity and carcinogenicity of the PAHs. In the past 30 years, various methods and technologies are applied to PAHs pollution remediation, such as physical methods, chemical oxidation and biological remediation, and currently, research on a method for removing polycyclic aromatic hydrocarbons in a water environment has achieved certain results at home and abroad. The conventional control measures for effective treatment include adsorption, bioremediation, photolysis, advanced oxidation and the like.
Benzo [ a ] pyrene (BaP) is a polycyclic aromatic hydrocarbon consisting of five benzene rings, one of the most carcinogenic polycyclic aromatic hydrocarbons. It has very low water solubility, vapor pressure and high octanol water partition coefficient, BaP tends to be non-aqueous and low in microbial utilization, so BaP is more likely to accumulate in the environment. The PAHs in practical operation can be known by combining a large number of references
The removal process is mainly a biological method or a physical method, wherein the biological method is considered as an economic and effective method for degrading the polycyclic aromatic hydrocarbon in the polluted environment by using microorganisms, but researches show that the polycyclic aromatic hydrocarbon with more than four rings has strong antimicrobial activity degradation capability, and exogenous microorganisms are influenced by polluted environment factors (such as temperature, pH value, oxygen content, nutrient content and the like) to cause great difference of degradation effects of the polycyclic aromatic hydrocarbon in different media. It is generally accepted that pH is the main factor affecting the bioremediation efficiency of polycyclic aromatic hydrocarbons. Since the optimal pH for different species of microorganisms is different, pH affects the growth and metabolic activity of the microorganism. In addition, the temperature also has a crucial influence on a bioremediation system of the polycyclic aromatic hydrocarbon. In a proper temperature range, the activity of the microorganism increases with the increase of temperature, because it can promote the metabolism of the microorganism and the activity of enzyme, thereby accelerating the bioremediation process of the polycyclic aromatic hydrocarbon. In view of the limitation of microorganism reinforced repair, the invention utilizes the combination of biological method and physical method to improve the degradation effect of polycyclic aromatic hydrocarbon according to the defects of the prior art. The mushroom dregs are culture medium waste after mushroom production of edible mushrooms, and China is a main edible mushroom producing country and accounts for 75% of the total world production. The mushroom dregs contain various nutrient substances including organic matters, nitrogen, phosphorus and potassium and various microorganisms (such as bacillus, brevibacterium, arthrobacter and microbacterium), the nutrient substances in the mushroom dregs can be used for environment improvement, such as changing the pH value and the nutrient content in the environment, and the like, and the nutrient substances can promote the degradation of polycyclic aromatic hydrocarbon, and the mushroom dregs have the double advantages of biostimulation and biological strengthening. The biological stimulation is to activate microorganisms in the polluted environment by adding a nitrogen source and a phosphorus source so as to improve the degradation effect of the polycyclic aromatic hydrocarbon; the biological enhancement is a method for degrading PAHs by adding exogenous polycyclic aromatic hydrocarbon degrading bacteria on the basis of biological stimulation, and in addition, various extracellular enzymes including lignin peroxidase (LiP), manganese peroxidase (MnP) and Laccase (LAC) can be secreted in the growth and development processes of the edible fungi, the extracellular enzymes are accumulated in fungi residues, and the extracellular enzymes can also degrade organic pollutants.
Disclosure of Invention
Aiming at the defects of the prior art and solving the defects of the prior art, the invention aims to provide the ganoderma lucidum fermented Shuanghuanglian mushroom dregs for repairing the water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene and the application thereof, the metabolic activity of polycyclic aromatic hydrocarbon degrading bacteria in the environment is improved and polycyclic aromatic hydrocarbon is degraded by influencing the survival and degradation performance of microorganisms in the environment, the water environment polluted by polycyclic aromatic hydrocarbon BaP is repaired by extracellular enzyme and adsorption capacity contained in the mushroom dregs, and the problem of protecting the water environment is effectively solved.
According to the technical scheme, lucid ganoderma fermentation Shuanghuanglian mushroom dregs for repairing a water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene are obtained, lucid ganoderma seed liquid is inoculated into a Shuanghuanglian dreg culture medium and cultured, lucid ganoderma is collected, the left culture medium dregs are the lucid ganoderma dregs, the lucid ganoderma dregs contain lignin decomposition enzyme, organic matters, nitrogen, phosphorus, potassium and a plurality of microorganisms, the lignin decomposition enzyme can secrete lignin peroxidase (LiP), manganese peroxidase (MnP) and Laccase (LAC) of a plurality of extracellular enzymes, and the extracellular enzymes have good degradation effects on various organic pollutants, particularly polycyclic aromatic hydrocarbon benzo [ a ] pyrene;
the ganoderma lucidum solid fermentation medium is prepared by taking 450g of Shuanghuanglian herb residues, 50g of corn flour, 15g of bran, 5g of gypsum, 5g of cane sugar, 0.06g of monopotassium phosphate, 0.06g of magnesium sulfate and 10.06g of vitamin B as raw materials;
the Shuanghuanglian decoction dregs refer to the residue left after decocting the composition consisting of the scutellaria baicalensis, the honeysuckle and the forsythia suspense (the known technology);
adding 5-10% of ganoderma lucidum dregs into a water body polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene under the conditions of acid-base environment with pH of 4.0-8.5 and temperature of 20-50 ℃, wherein the weight volume refers to g of the weight and mL of the volume, the survival and degradation performance of microorganisms for degrading polycyclic aromatic hydrocarbon benzo [ a ] pyrene can be effectively improved, polycyclic aromatic hydrocarbon benzo [ a ] pyrene in the water body with the polycyclic aromatic hydrocarbon benzo [ a ] pyrene pollution concentration of 50mg/L is removed, the ganoderma lucidum dregs especially show better degradation effect under the conditions of acidity pH of 4.5-5.5 and lower temperature of 20-25 ℃, the degradation rate of benzo [ a ] pyrene reaches more than 90% after 7 days, and the ganoderma lucidum dregs can be more widely applied to the repair of the water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene.
The invention has the advantages of rich raw materials, easy obtainment of mushroom dregs, convenient use and good effect, can effectively solve the problem of repairing the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene, is a great innovation on repairing the water body polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene, and has great economic and social benefits.
Detailed Description
The following detailed description of the embodiments of the present invention refers to the accompanying drawings.
The invention relates to a ganoderma lucidum fungus residue for repairing a water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene, wherein the ganoderma lucidum fungus residue is obtained by fermenting residues collected by ganoderma lucidum by using Shuanghuanglian medicine residues as a base material by the following method:
(1) screening of mushroom dregs
And (3) carrying out lignin decomposition enzyme activity detection on the collected ganoderma lucidum dregs, oyster mushroom dregs and mushroom dregs, and screening out the dregs with higher lignin decomposition enzyme for a subsequent degradation test. 10.0g of the bacterial residues are weighed into a 250mL conical flask, 25mL of distilled water is added, then the conical flask is placed into a shaking table and is cultured for 1h at 37 ℃ and 180rpm in a shaking way. After shaking for 1h, taking 1mL of fungus residue liquid in a centrifuge tube, centrifuging at 12000rpm for 5min, and taking supernatant to detect ligninase activity;
through detection, the laccase activity of the ganoderma lucidum dregs is 468.2U/g, the manganese peroxidase activity is 699.52U/g, compared with other dregs, the lignin decomposition enzyme activity of the ganoderma lucidum dregs is higher, and the enzyme activity of the ganoderma lucidum dregs is higher than that reported in the existing documents.
And (3) laccase activity detection: the laccase activity is determined by adopting an ABTS method, the reaction system is 1mL, the buffer solution containing the tartaric acid with the pH value of 4.0 is 0.5mL, the distilled water is 0.38mL, the fermentation enzyme solution is 10 muL and the ABTS is 100 muL, the reaction is carried out in water bath at the temperature of 30 ℃ for 1min, the reaction lasts for 30s, the light absorption value is determined at the position of 420nm, and each group is repeated for 3 times. 1 enzyme activity unit is defined as the amount of enzyme required to oxidize 1 μmol abts per minute under the current reaction conditions.
Determination of enzyme activity of manganese peroxidase: the manganese peroxidase kit produced by Solarbio company is used, the determination reagent and the sample are sequentially added according to the kit use instruction, and the absorbance is determined at 465nm by a spectrophotometer. 1 enzyme activity unit is defined as the amount of enzyme required to oxidize 1nmol of guaiacol per gram of sample per minute as one enzyme activity unit.
(2) Ganoderma lucidum seed liquid culture
Inoculating the ganoderma lucidum strain into a seed culture medium, and placing the seed culture medium in a constant-temperature shaking table at 30 ℃ and 180rmp for shake culture for later use; making round hole with diameter of 0.5cm on Ganoderma plate, inoculating into liquid Ganoderma seed culture medium, culturing at 30 deg.C and 180rpm for 3 days;
the seed culture medium is as follows: 30g of corn flour, 15g of bean cake powder and 5g of bran, adding water, boiling for 1h, filtering by 8 layers of gauze, adding 20g of glucose, 0.2g of magnesium sulfate, 1g of monopotassium phosphate and 5g of yeast powder, fixing the volume to 1000mL, and sterilizing for 20min at 115 ℃;
(3) solid fermentation of ganoderma lucidum
Adding a ganoderma lucidum solid fermentation culture medium into the fungus bags, wherein the weight of each ganoderma lucidum fungus bag is 500g, adding water until the volume water content is 60%, sterilizing at 121 ℃ for 2h, and naturally cooling for later use;
the lucid ganoderma solid fermentation culture medium: 450g of Shuanghuanglian dregs of decoction, 50g of corn flour, 15g of bran, 5g of gypsum, 5g of cane sugar, 0.06g of monopotassium phosphate, 0.06g of magnesium sulfate and 10.06g of vitamin B;
inoculating the lucid ganoderma seed liquid into lucid ganoderma fungus bags, wherein the inoculation amount is 2% of the weight volume of a culture medium, the weight volume refers to that solid is counted by g, liquid is counted by mL, after inoculation is finished, the lucid ganoderma fungus bags are moved into a dark room to grow hyphae, the fungus bags full of hyphae are moved into an illumination incubator to carry out lucid ganoderma growth management, the temperature of the incubator is kept at 26-28 ℃, the humidity is kept at 85-90%, wind is avoided (ventilation is not needed) in the growing period of stipe, ventilation is carried out in time when the lucid ganoderma is sliced, the growing period of the illumination intensity stipe is 180-fold 220Lx, the slicing period is 450-fold 550Lx, the lucid ganoderma is harvested in 28-32 days, and solid fermentation culture medium residues in the fungus bags after lucid ganoderma harvesting are the lucid ganoderma fungus residues, namely the lucid ganoderma fungus residues.
The application of the ganoderma lucidum fungus dregs for repairing the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene obtained by the method in repairing the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene has the advantages that the initial pollution concentration of the polycyclic aromatic hydrocarbon benzo [ a ] pyrene is 50mg/L, the water environment is an acid-base environment with the pH value of 4.0-8.5, and the temperature is 20-50 ℃;
tests prove that the ganoderma lucidum dregs contain lignin-decomposing enzyme, organic matters, nitrogen, phosphorus, potassium, various microorganisms and the like, wherein the lignin-decomposing enzyme comprises lignin peroxidase (LiP), manganese peroxidase (MnP) and laccase (Lac), and the extracellular enzyme has a good degradation effect on various organic pollutants.
The Ganoderma residue can remove polycyclic aromatic hydrocarbon benzo [ a ] pyrene in acidic and alkaline water environment and at different temperatures, wherein the pH of the acid-base environment is 4.0-8.5, and the different temperatures are 20-50 deg.C.
The method comprises the following steps of culturing ganoderma lucidum dregs in different acid-base water environments (pH4.0-8.5), wherein the culture temperature is 37 ℃, the initial concentration of benzo [ a ] pyrene is 50mg/L, and after 5 days of degradation, liquid chromatography detection shows that the removal rate of benzo [ a ] pyrene is 60-80%, culturing the ganoderma lucidum dregs at different temperatures (20-50 ℃) and at pH6.0, the initial concentration of benzo [ a ] pyrene is 50mg/L, and after 5 days of degradation, liquid chromatography detection shows that the removal rate of benzo [ a ] pyrene is more than 90%.
The ganoderma lucidum dregs can efficiently degrade high molecular weight polycyclic aromatic hydrocarbon benzo [ a ] pyrene in acidic and alkaline environments and at different temperatures, wherein the pH value of the acidic and alkaline environments is 4.0-8.5, the different temperatures are 20-50 ℃, and the bioremediation of the high molecular weight polycyclic aromatic hydrocarbon benzo [ a ] pyrene in a water environment is effectively solved; the ganoderma lucidum dregs are culture medium waste after fruiting of edible fungi, contain various nutrient substances including organic matters, nitrogen, phosphorus and potassium and various microorganisms (such as bacillus, brevibacterium, arthrobacterium and microbacterium) which can promote degradation of polycyclic aromatic hydrocarbons, and have the triple advantages of adsorption, biostimulation and biological enhancement; in addition, various extracellular enzymes are secreted in the growth and development process of the edible fungi, including lignin peroxidase (LiP), manganese peroxidase (MnP) and laccase (Lac), the extracellular enzymes are accumulated in fungi residues, and the extracellular enzymes can also degrade organic pollutants, such as polycyclic aromatic hydrocarbons, pentachlorophenol, heptachlor, dichlorodiphenyl trichloroethane and the like; finally, the ganoderma lucidum dregs are different from the traditional culture medium formula, the culture medium matrix used by the dregs is a Shuanghuanglian dregs, the dregs are rich in crude protein, crude fiber, amino acid and other nutrients and insufficiently extracted medicinal active ingredients, and a material basis can be provided for the growth of microorganisms in the dregs matrix. And the experiment obtains very good beneficial technical effect, and the related data are as follows:
1. degradation experiment of ganoderma lucidum dregs on benzo [ a ] pyrene in water environment under different pH conditions:
phosphate buffer solutions with different pH values (4.0-8.5) are prepared. Preparing 1g/L benzo [ a ] pyrene standard mother liquor by using acetone, adding 1mL of benzo [ a ] pyrene mother liquor into a 100mL bottle with a plug, volatilizing the acetone at room temperature, adding 20mL of phosphate buffer solutions with different pH values after the acetone is volatilized, then adding 1.5g of ganoderma lucidum dregs, placing the benzo [ a ] pyrene aqueous solution without adding the ganoderma lucidum dregs as a contrast into a constant-temperature shaking table with the temperature of 30 ℃ and the rpm for degradation experiments, repeating the treatment for 3 times, and detecting the benzo [ a ] pyrene content after culturing for 5 days. Removal% = (control concentration-degraded sample concentration)/control concentration × 100%. The Biolog method measures the functional diversity of microorganisms in contaminated water.
The liquid phase sample pretreatment method comprises the following steps: extracting the extract phase with chromatographic grade n-hexane, adding 10mL of n-hexane into each sample, extracting, vortex shaking for 10min, standing until the sample is layered and stable, adding anhydrous sodium sulfate into the upper layer organic phase, and analyzing.
HPLC analysis: the content of PAHs is measured by an Agilent LC-1200 high performance liquid chromatograph. The sample injection amount is 20 muL, the separation column is ZORBAX SB-C18 column (0.46X 150mm, Agilent), the column temperature is 30 ℃, the ultraviolet detection wavelength is 254nm, the mobile phase is acetonitrile and water (volume ratio is 80%: 20%), and the flow rate is 2.0mL min-1。
The result shows that the removal rate of benzo [ a ] pyrene is more than 85% after 5 days of liquid phase detection (taking an average value). Biolog
The result shows that the microbial metabolic activity of the polluted water is obviously improved after the bacterial residues are added.
2. Degradation experiment of ganoderma lucidum dregs on benzo [ a ] pyrene in water environment under different temperature conditions:
phosphate buffer solution with pH6.0 was prepared. Preparing 1g/L benzo [ a ] pyrene standard mother liquor by using acetone, adding 1mL of benzo [ a ] pyrene mother liquor into a 100mL bottle with a plug, volatilizing the acetone at room temperature, adding 20mL of phosphate buffer solution with pH of 6.0 after the acetone is volatilized, then adding 1.5g of ganoderma lucidum dregs and benzo [ a ] pyrene aqueous solution without the ganoderma lucidum dregs as a contrast, placing the mixture in constant-temperature shaking tables at different temperatures (20-50 ℃), performing degradation experiments at 180rpm, repeating the treatment for 3 times, and detecting the benzo [ a ] pyrene content after culturing for 5 days. Removal% = (control concentration-degraded sample concentration)/control concentration × 100%. The Biolog method measures the functional diversity of microorganisms in contaminated water.
The liquid phase sample pretreatment method comprises the following steps: extracting the extract phase with chromatographic grade n-hexane, adding 10mL of n-hexane into each sample, extracting, vortex shaking for 10min, standing until the sample is layered and stable, adding a certain amount of anhydrous sodium sulfate into the upper layer of organic phase, and analyzing.
HPLC analysis: the research adopts an Agilent LC-1200 high performance liquid chromatograph to measure the content of PAHs. The sample injection amount is 20 muL, the separation column is ZORBAX SB-C18 column (0.46X 150mm, Agilent), the column temperature is 30 ℃, the ultraviolet detection wavelength is 254nm, the mobile phase is acetonitrile and water (volume ratio is 80%: 20%), and the flow rate is 2.0mL min-1。
The result shows that the removal rate of benzo [ a ] pyrene is more than 90% after 5 days of liquid phase detection (taking an average value). The ganoderma lucidum dregs have good removal effect on high molecular weight polycyclic aromatic hydrocarbon under different temperature conditions, and Biolog results show that the microbial metabolic activity of the polluted water is obviously improved after the fungus dregs are added.
The invention obtains the same or similar effect with the experiment through repeated experiments, and the experiment does not detail and lists, and the experiment shows that the ganoderma lucidum dregs can effectively remove the polycyclic aromatic hydrocarbon benzo [ a ] pyrene with high molecular weight in the water solution. The fungus residue is predicted to have potential application value in the aspect of repairing high molecular weight polycyclic aromatic hydrocarbon under the water environment condition, can effectively solve the problem of repairing the water environment of polycyclic aromatic hydrocarbon benzo [ a ] pyrene pollution concentration of 50mg/L, the pH value of the water body is 4.0-8.5, and the temperature is 20-50 ℃, and realizes the protection of the water environment, compared with the prior art, the invention has the following advantages:
firstly, the invention utilizes the Shuanghuanglian herb residues to ferment the lucid ganoderma, realizes the green recycling of high added value by converting the herb residues through microorganisms, changes waste into valuable, and provides technical support for the modernization and healthy sustainable development of the traditional Chinese medicine industry.
Secondly, after the ganoderma lucidum is collected, the effective degradation of benzo [ a ] pyrene in the water environment is realized by using ganoderma lucidum fungus residues, the remediation of the polluted water body is realized, the fungus residues are changed into valuables, and a green circular economy mode is realized.
The mushroom dregs contain a large amount of components such as edible mushroom mycelium residues, extracellular enzymes, polysaccharides, crude fibers degraded by biological enzymes, lignin which is not completely utilized and the like, and are high-quality biomass resources. The nutrient components are even higher than those of the primary culture materials, so that the environment pollution condition can be optimized and improved, the metabolic activity of the original polycyclic aromatic hydrocarbon degrading bacteria in the environment can be improved, and the degrading efficiency can be further improved. Meanwhile, the mycelium in the mushroom dregs can secrete hormone substances and a series of extracellular enzymes for degrading lignocellulose in the growth and development process, wherein the extracellular enzymes comprise laccase, lignin peroxidase, cellulase, xylanase and the like, and the extracellular enzymes can degrade organic pollutants, such as polycyclic aromatic hydrocarbon, pentachlorophenol, heptachlor, dichlorodiphenyl trichloroethane and the like.
Meanwhile, the ganoderma lucidum dregs can also absorb certain benzo [ a ] pyrene pollutants, have triple functions of absorption, biological enhancement (degradation) and biological stimulation, are a great innovation on repairing polycyclic aromatic hydrocarbon benzo [ a ] pyrene pollution, show better degradation effect on benzo [ a ] pyrene under subacidity and lower temperature, have the removal rate of polycyclic aromatic hydrocarbon benzo [ a ] pyrene of more than 90 percent, provide powerful technical support for repairing polycyclic aromatic hydrocarbon benzo [ a ] pyrene polluted water, effectively protect water environment, and have huge economic and social benefits.
Claims (4)
1. A Ganoderma lucidum dreg for repairing water environment polluted by polycyclic aromatic hydrocarbon benzo [ a ] pyrene is characterized in that the dreg is prepared by inoculating Ganoderma lucidum seed liquid into a Ganoderma lucidum solid fermentation culture medium, culturing, collecting Ganoderma lucidum, and obtaining the residue of the Ganoderma lucidum solid fermentation culture medium, wherein the residue of the Ganoderma lucidum solid fermentation culture medium is Ganoderma lucidum dreg which contains lignin-decomposing enzyme, organic matters, nitrogen, phosphorus, potassium and various microorganisms;
the ganoderma lucidum solid fermentation medium is prepared from 450g of Shuanghuanglian herb residues, 50g of corn flour, 15g of bran, 5g of gypsum, 5g of cane sugar, 0.06g of monopotassium phosphate, 0.06g of magnesium sulfate and 10.06g of vitamin B.
2. The preparation method of the ganoderma lucidum dregs for repairing the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene, which is claimed in claim 1, is characterized by comprising the following steps:
(1) and culturing lucid ganoderma seed liquid:
inoculating the ganoderma lucidum strain into a seed culture medium, and placing the seed culture medium in a constant-temperature shaking table at 30 ℃ and 180rmp for shake culture for later use; making round hole with diameter of 0.5cm on Ganoderma plate, inoculating into liquid Ganoderma seed culture medium, culturing at 30 deg.C and 180rpm for 3 days;
the lucid ganoderma seed culture medium comprises: 30g of corn flour, 15g of bean cake powder and 5g of bran, adding water, boiling for 1h, filtering by 8 layers of gauze, adding 20g of glucose, 0.2g of magnesium sulfate, 1g of monopotassium phosphate and 5g of yeast powder, fixing the volume to 1000mL, and sterilizing for 20min at 115 ℃;
(2) solid fermentation of glossy ganoderma
Adding a ganoderma lucidum solid fermentation culture medium into the fungus bags, wherein the weight of each ganoderma lucidum fungus bag is 500g, adding water until the volume water content is 60%, sterilizing at 121 ℃ for 2h, and naturally cooling for later use;
the ganoderma lucidum solid fermentation medium is prepared by taking 450g of Shuanghuanglian herb residues, 50g of corn flour, 15g of bran, 5g of gypsum, 5g of cane sugar, 0.06g of monopotassium phosphate, 0.06g of magnesium sulfate and 10.06g of vitamin B as raw materials;
inoculating the lucid ganoderma seed liquid into lucid ganoderma fungus bags, wherein the inoculation amount is 2% of the weight volume of a culture medium, the weight volume refers to that solid is counted by g, liquid is counted by mL, after inoculation is finished, the lucid ganoderma fungus bags are moved into a dark room to grow hyphae, the fungus bags full of the hyphae are moved into an illumination incubator to carry out lucid ganoderma growth management, the temperature of the incubator is kept at 26-28 ℃, the humidity is 85-90%, wind is avoided in the growing period of stipe, ventilation is carried out in time when the lucid ganoderma is sliced, the growing period of the stipe is 180-fold of 220Lx in illumination intensity, the slicing period is 450-fold of 550Lx, the lucid ganoderma is harvested in 28-32 days, and solid fermentation culture medium residues in the fungus bags after the lucid ganoderma are harvested are lucid ganoderma fungus residues.
3. The application of the ganoderma lucidum dregs for repairing the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene in the claim 1 in repairing the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene, wherein the initial pollution concentration of the water body polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene is 50mg/L, the water environment is an acid-base environment with the pH value of 4.0-8.5, and the temperature is 20-50 ℃.
4. The application of the ganoderma lucidum dregs for repairing the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene in repairing the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene according to claim 3, wherein the initial pollution concentration of the water environment polluted by the polycyclic aromatic hydrocarbon benzo [ a ] pyrene is 50mg/L, the water environment is an acidic environment with the pH value of 4.0-5.5, and the temperature is 20-25 ℃.
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