CN106367363B - Preparation and use methods of microbial inoculum for drainage basin hexavalent chromium pollution treatment - Google Patents

Preparation and use methods of microbial inoculum for drainage basin hexavalent chromium pollution treatment Download PDF

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CN106367363B
CN106367363B CN201610078160.7A CN201610078160A CN106367363B CN 106367363 B CN106367363 B CN 106367363B CN 201610078160 A CN201610078160 A CN 201610078160A CN 106367363 B CN106367363 B CN 106367363B
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hexavalent chromium
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孙莹
赵英
王新民
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Louis Ecosystem Engineering Co ltd
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Abstract

The invention discloses a production method of a Streptomyces purpureus (Streptomyces purpureus strain LYzhlm CGMCC No.11773) microbial inoculum and a use method for treating hexavalent chromium polluted water by using the product. The experiment for treating the hexavalent chromium polluted water by using the product can reduce the concentration of hexavalent chromium in the sewage from 1790mg/L to 0.08mg/L, completely reaches the discharge standard of hexavalent chromium and has the removal rate of 100 percent. The invention also discloses a production process of the strain, and a use condition and a method for treating hexavalent chromium polluted water, wherein the process is simple, has no secondary pollution compared with a chemical method, is convenient to use, and has wide market prospect.

Description

Preparation and use methods of microbial inoculum for drainage basin hexavalent chromium pollution treatment
Technical Field
The invention relates to the technical field of chromium pollution treatment, in particular to a preparation method and a use method of a microbial inoculum for treating hexavalent chromium pollution in a drainage basin.
Background
Chromium is a silvery-white hard metal, and has divalent, trivalent, and hexavalent compounds. The industrial application of chromium is wide, and mainly comprises metal processing, electroplating and leather making industries, and waste water and waste gas discharged by the industries are main pollution sources in polluted environment of drainage basins. Hexavalent chromium is highly toxic, and is a swallow poison/inhalation poison, and skin contact may cause allergy; more likely to cause genetic defects, inhalation may be carcinogenic, and there is a persistent risk to the environment. These are characteristic of hexavalent chromium, however, chromium metal, trivalent or tetravalent chromium does not have these toxicities.
The most common chromium pollution is water pollution, such as wastewater from chromium electroplating, and wastewater from industrial enterprises using chromium and its compounds in the tanning, pharmaceutical, and printing and dyeing industries, which mainly enters the environment in two valence states of Cr (III) and Cr (VI). According to the data, the tanning industry generally processes 1t of hides and discharges 50-60t of wastewater containing 410mg/L of chromium. The circulating cooling water used in oil refineries and chemical plants also has a high chromium content. The chromium content in the wastewater of the chromium plating plant is higher, and particularly, a large amount of chromium-containing wastewater is discharged when the electroplating solution is replaced. The pollution of chromium to water bodies is quite serious not only in China but also in all countries around the world. Chromium pollution is generally regarded as a key prevention and treatment object in all countries in the world. And the chromium slag is sodium dichromate and waste slag discharged in the production of metal chromium. When the chromium slag is stockpiled in the open air, a large amount of hexavalent chromium ions are dissolved, lost and seeped into the ground surface along with rainwater after long-term rainwater washing, so that underground water is polluted, rivers and lakes are polluted, and farmlands, aquatic products and human health are further harmed. An excess (over 10ppm) of hexavalent chromium is lethal to aquatic life. Experiments have shown that hexavalent chromium in contaminated drinking water can be carcinogenic. Hexavalent chromium compounds are commonly used in electroplating and the like, and are absorbed by cells of many tissues and organs in the body after animals drink water containing hexavalent chromium.
The bioremediation mechanism of the heavy metal polluted water body mainly comprises the fixation of heavy metals and the transformation of forms by microorganisms. The former is that the microorganism absorbs heavy metal ions through the charged cell surface, or actively absorbs the heavy metal ions by taking necessary nutrient elements, so as to enrich the heavy metal on the cell surface or inside the cell; the latter is to reduce heavy metal pollution by changing the form of heavy metals or reducing the bioavailability of heavy metals through the life activities of microorganisms, such as Cr6+Conversion to Cr3+And the toxicity is reduced, As, Hg, Se and the like are reduced into simple substances and volatilized, and the secretion of microorganisms has passivation effect on heavy metals and the like.
Therefore, the method has wide application prospect in 'repairing' the industrial effluent polluted by the heavy metal, particularly for the water body with the heavy metal content below 100mg/L, the metal ions are removed by adopting other methods, the treatment is difficult or the cost is high, and the heavy metal ions can be effectively removed by using easily obtained and cheap raw materials, namely waste (such as fermentation byproducts) of certain industries or organisms (such as algae) which exist in large quantities in the nature as biological adsorbents.
The prior art needs to be improved and developed.
Disclosure of Invention
The invention aims to provide a preparation and application method of a microbial inoculum for drainage basin hexavalent chromium pollution treatment.
The technical solution of the invention is as follows: a preparation method of a microbial inoculum for treating hexavalent chromium pollution in a drainage basin is disclosed, wherein the microbial inoculum is Streptomyces puniceus strain LYzhlm, and the preservation number of the microbial inoculum is CGMCC No.11773, and the preparation method is characterized by comprising the following steps:
step 1, preparing a fermentation raw material: preparing a fermentation raw material according to the mass percentage of the liquid molasses, the industrial glucose 1.0-5%, the industrial yeast powder 0.5%, the bran powder 3.0% (the grinding requirement is less than 100 meshes), the malt extract 1.0%, the dipotassium hydrogen phosphate 0.3%, the sodium dihydrogen phosphate 0.1%, the ammonium sulfate 0.5% and the magnesium sulfate 0.05%;
step 2, sterilization: putting the raw materials in the step 1 into a fermentation tank for sterilization, wherein the sterilization temperature is 121 ℃, and the sterilization time is 0.5 hour;
step 3, fermentation: fermenting the fermentor containing the fermentation feedstock after sterilization; the fermentation temperature is controlled to be 30-32 ℃, the fermentation time is 96 hours, and during fermentation, the ventilation gas: the liquid aeration ratio is 1:0.5 in 0-24 hours, 1:1 in 24-96 hours, the tank pressure of a fermentation tank is 0.08Mpa, and the fermentation raw materials are stirred all the time after being put into the fermentation tank, wherein the stirring speed is 200 rpm;
and 4, checking the total number of bacteria of the fermentation broth produced by fermentation, filling and packaging to obtain a finished product.
And 5, adding the LYzhlm microbial inoculum into the hexavalent chromium polluted water, wherein the volume ratio of the microbial inoculum to the polluted water is 0.5-3.0: 100(L/L), controlling the pH value of the polluted water added with the microbial inoculum to be 5-9, controlling the temperature to be 25-35 ℃, and controlling the stirring speed to be 80-140 r/min.
And 6, the weight-volume ratio of the polluted water is 2.0:100 g/ml.
And 7, adding the microbial inoculum to obtain the polluted water with the pH value of 7.0.
And step 8, adding the microbial inoculum, and then heating the polluted water to 30 ℃.
The invention has the beneficial effects that:
the LYzhlm fermentation liquor has a strong hexavalent chromium reduction effect. The effect evaluation in the laboratory shows that the LYzhlm strain has strong performance of removing hexavalent chromium, and can reduce hexavalent chromium ions with the concentration of 100 mg/L-1000 mg/L to 0 under the conditions of oxygen and medium temperature, and the reduction rate is 100%. 2 times of removal experiments are carried out on the industrial hexavalent chromium polluted water, the initial concentrations of hexavalent chromium in the polluted water before the experiments are 1790mg/L and 972mg/L respectively, and the concentrations of hexavalent chromium in the polluted water treated by the method are reduced to 0.08mg/L and 0.046mg/L, so that the discharge standard of hexavalent chromium is reached. Compared with the prior art, the method can treat the hexavalent chromium-containing wastewater with higher concentration, and has high reduction efficiency.
Drawings
FIG. 1 is a curve for removing hexavalent chromium ions from hexavalent chromium wastewater treated by LYzhlm.
Detailed Description
The invention discloses a Streptomyces punicus strain LYzhlm capable of efficiently removing hexavalent chromium pollution, which is preserved in the common microorganism center of China general microbiological culture Collection management Committee on 12.2.2015, wherein the preservation number is CGMCC No.11773, and the Streptomyces punicus strain LYzhlm is named. Address: the institute of microbiology of western-style academy of sciences, north-Chen, Chaozhou, Chaoyang.
The 16S rDNA sequence is the sequence of the sequence table SEQ NO. 1.
The streptomycete for efficiently removing hexavalent chromium pollution is applied to the aspect of removing hexavalent chromium pollution.
The LYzhlm strain is derived from industrial sludge, and is subjected to indoor domestication, separation, purification and screening. Culturing LYzhlm strain in LB culture medium, and making the cell form be filamentous; after culturing for 4 days, the color of the colony is grey white, and the colony is round and unsmooth. The intrabasal hyphae did not break, and the aerial hyphae generally developed well to form long spore filaments. The intrabasal hyphae have many branches and produce a purplish red water-soluble pigment. The spore silk shape was microscopically observed. The spores are oval and have smooth surfaces.
The most common valence states of chromium in soil are trivalent chromium and hexavalent chromium, and in natural environment, hexavalent chromium is easily reduced to trivalent chromium by microorganisms, while trivalent chromium is difficult to be oxidized to hexavalent chromium. Hexavalent chromium in soils is commonly found as chromate (CrO)4 2-) And dichromate (Cr)2O4 2-) The product has high activity and strong toxic action on organisms. Trivalent chromium is mainly Cr (H) in soil, compared with hexavalent chromium2O)6 3+、Cr(H2O)2+、CrO2+The fertilizer has the advantages of existing forms, easy adsorption by soil or generation of sediment, low toxic and harmful effects on organisms and relatively small influence on the environment. The Streptomyces purpureus (Streptomyces purpureus strain LYzhlm) obtained by screening and domesticating changes the chemical form of heavy metal, fixes the heavy metal, reduces the bioavailability and reduces the harm of the heavy metal. Compared with physical and chemical repair technologies, the LYzhlm repair technology provides an efficient strain for treating the high pollution of hexavalent chromium due to the advantages of safety, non-destructiveness and economy.
The LYzhlm strain is amplified by PCR method using bacterial 16S rDNA universal primer. 16S rDNA partial fragment, PCR product sequencing to obtain the fragment sequence, sequencing the 16S rDNA (length 1.4kbp) of LYzhlm strain as shown in sequence table SEQ No. 1.
The homology between the 16S rDNA sequence of LYzhlm strain and Streptomyces aureofaciens is highest and reaches 99% by performing gene homology and similarity analysis by using BLAST tool on the sequence table SEQ NO.1 and performing BLAST search (see Table 1)
TABLE 1 BLAST search results Table
Figure GDA0002218362190000031
Figure GDA0002218362190000041
According to the BLAST analysis results, the strain belongs to the following biological taxonomy information:
domain：Bacteria；
phylum：Actinobacteria；
class：Streptomycetales；
order：Streptomycetaceae；
family：Streptomyces；
Genus：Streptomyces puniceus。
thus, the new strain was identified as Streptomyces purpureus (Streptomyces puniceus strain) and named Streptomyces purpureus (Streptomyces puniceus strain LYzhlm).
The preparation method of the strain comprises the following steps:
step 1, preparing a fermentation raw material: preparing a fermentation raw material according to the mass percentage of the liquid molasses, the industrial glucose 1.0%, the industrial yeast powder 0.5%, the bran powder 3.0% (the grinding requirement is less than 100 meshes), the malt extract 1.0%, the dipotassium phosphate 0.3%, the sodium dihydrogen phosphate 0.1%, the ammonium sulfate 0.5% and the magnesium sulfate 0.05%;
step 2, sterilization: putting the raw materials in the step 1 into a fermentation tank for sterilization, wherein the sterilization temperature is 121 ℃, and the sterilization time is 0.5 hour;
step 3, fermentation: fermenting the fermentor containing the fermentation feedstock after sterilization; the fermentation temperature is controlled to be 30-32 ℃, the fermentation time is 96 hours, and during fermentation, the ventilation gas: the liquid aeration ratio is 1:0.5 in 0-24 hours, 1:1 in 24-96 hours, the tank pressure of a fermentation tank is 0.08Mpa, and the fermentation raw materials are stirred all the time after being put into the fermentation tank, wherein the stirring speed is 200 rpm;
and 4, checking the total number of bacteria of the fermentation broth produced by fermentation, filling and packaging to obtain a finished product.
And 5, adding the LYzhlm microbial inoculum into the hexavalent chromium polluted water, wherein the volume ratio of the microbial inoculum to the polluted water is 0.5-3.0: 100(L/L), controlling the pH value of the polluted water added with the microbial inoculum to be 5-9, controlling the temperature to be 25-35 ℃, and controlling the stirring speed to be 80-140 r/min.
And 6, the weight-volume ratio of the polluted water is 2.0:100 g/ml.
And 7, adding the microbial inoculum to obtain the polluted water with the pH value of 7.0.
And step 8, adding the microbial inoculum, and then heating the polluted water to 30 ℃.
Example 1 basis for detection
The method for measuring hexavalent chromium in sewage comprises the following steps:
dihydrazine diphenylcarbonate spectrophotometry for determining hexavalent chromium in water quality GB7467-1987
The method for measuring the total chromium in the sewage comprises the following steps:
method for measuring total chromium in water by oxidizing potassium permanganate and dihydrazide diphenyl carbonate spectrophotometry GB7466-1987
Example 2 the specific steps of the product of the invention for treating high concentration hexavalent chromium heavy metal contaminated water are as follows:
the product produced by the method is added into 1 of hexavalent chromium polluted water, and the detected concentration of the hexavalent chromium contained in the 1 of the polluted water is 1790 mg/L. The volume ratio of the product to the polluted water is 2.5:100(L/L), the pH value of the polluted water is adjusted to be 7.0, the temperature of the polluted water is controlled to be 30 ℃, the stirring speed is 100r/min, samples are taken at the treatment interval of 24 hours, and the concentration of total chromium and hexavalent chromium is measured. The sampling was stopped after the Cr (VI) concentration in the medium was below the detection limit.
Example 3 the specific steps of the product of the invention for treating hexavalent chromium heavy metal polluted water with medium difficulty are as follows:
the product produced by the method is added into 2 of the hexavalent chromium polluted water, and the detected hexavalent chromium concentration of the polluted water 2 is 972 mg/L. The volume ratio of the product to the polluted water is 2.0:100(L/L), the pH value of the polluted water is adjusted to be 7.0, the temperature of the polluted water is controlled to be 30 ℃, the stirring speed is 100r/min, samples are taken at the treatment interval of 24 hours, and the concentration of total chromium and hexavalent chromium is measured. The sampling was stopped after the Cr (VI) concentration in the medium was below the detection limit.
The data obtained for the removal of hexavalent chromium contaminated water by the process described in examples 2 and 3 is shown in table 2:
TABLE 2 result table for removing hexavalent chromium from waste water
Figure GDA0002218362190000051
Figure GDA0002218362190000061
As can be seen from Table 2, the method of the present invention has a significant improvement in the removal of hexavalent chromium from contaminated water by conventional microbiological methods.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
The above detailed description is specific to possible embodiments of the present invention, and the embodiments are not intended to limit the scope of the present invention, and all equivalent implementations or modifications that do not depart from the scope of the present invention are intended to be included within the scope of the present invention.
Figure GDA0002218362190000071
Sequence listing
<110> land area ecological engineering Co., Ltd
<120> streptomycete for efficiently removing hexavalent chromium pollution and application thereof
<160>1
<210> 1
<211> 1431
<212> DNA
<213> Artificial sequence
<400> 1
CTTACCTAACCGCGCTTACCATGCAGTCGACGATGAACCGCTTCGGTGGTGGATTAGTGGCGAACGGGTGAGTAACACGTGGGCAATCTGCCCTTCACTCTGGGACAAGCCCTGGAAACGGGGTCTAATACCGGATAATACTCCTGCCTGCATGGGTGGGGGTTGAAAGCTCCGGCGGTGAAGGATGAGCCCGCGGCCTATCAGCTTGTTGGTGGGGTAATGGCCTACCAAGGCGACGACGGGTAGCCGGCCTGAGAGGGCGACCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCACAATGGGCGAAAGCCTGATGCAGCGACGCCGCGTGAGGGATGACGGCCTTCGGGTTGTAAACCTCTTTCAGCAGGGAAGAAGCGCAAGTGACGGTACCTGCAGAAGAAGCGCCGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGGCGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGAGCTCGTAGGCGGCTTGTCACGTCGGATGTGAAAGCCCGGGGCTTAACCCCGGGTCTGCATTCGATACGGGCTAGCTAGAGTGTGGTAGGGGAGATCGGAATTCCTGGTGTAGCGGTGAAATGCGCAGATATCAGGAGGAACACCGGTGGCGAAGGCGGATCTCTGGGCCATTACTGACGCTGAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGTTGGGAACTAGGTGTTGGCGACATTCCACGTCGTCGGTGCCGCAGCTAACGCATTAAGTTCCCCGCCTGGGGAGTACGGCCGCAAGGCTAAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCAGCGGAGCATGTGGCTTAATTCGACGCAACGCGAAGAACCTTACCAAGGCTTGACATATACCGGAAAGCATCAGAGATGGTGCCCCCCTTGTGGTCGGTATACAGGTGGTGCATGGCTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGTTCTGTGTTGCCAGCATGCCTTTCGGGGTGATGGGGACTCACAGGAGACTGCCGGGGTCAACTCGGAGGAAGGTGGGGACGACGTCAAGTCATCATGCCCCTTATGTCTTGGGCTGCACACGTGCTACAATGGCCGGTACAATGAGCTGCGATGCCGTGAGGCGGAGCGAATCTCAAAAAGCCGGTCTCAGTTCGGATTGGGGTCTGCAACTCGACCCCATGAAGTCGGAGTTGCTAGTAATCGCAGATCAGCATTGCTGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACGTCACGAAAGTCGGTAACACCCGAAGCCGGTGGCCCAACCCCTGTGGGAGGAGCTTCGAAGTGACTTCTTGGGGTC

Claims (5)

1. A preparation method of a microbial inoculum for treating hexavalent chromium pollution in a drainage basin comprises the following bacterial strains Streptomyces puniceus strain LYzhlm, the preservation number of which is CGMCC No.11773, and the 16S rDNA sequence is the sequence of a sequence table SEQ No.1, and is characterized by comprising the following steps:
step 1, preparing a fermentation raw material: preparing a fermentation raw material according to the mass percentage of the liquid molasses and the water, wherein the liquid molasses is 5.0 percent, the industrial glucose is 1-5 percent, the industrial yeast powder is 0.5 percent, the bran powder is 3.0 percent, the crushing requirement is less than 100 meshes, the malt extract is 1.0 percent, the dipotassium hydrogen phosphate is 0.3 percent, the sodium dihydrogen phosphate is 0.1 percent, the ammonium sulfate is 0.5 percent, and the magnesium sulfate is 0.05 percent;
step 2, sterilization: putting the raw materials in the step 1 into a fermentation tank for sterilization, wherein the sterilization temperature is 121 ℃, and the sterilization time is 0.5 hour;
step 3, fermentation: fermenting the fermentor containing the fermentation feedstock after sterilization; the fermentation temperature is controlled to be 30-32 ℃, the fermentation time is 96 hours, and during fermentation, the ventilation gas: the liquid aeration ratio is 1:0.5 in 0-24 hours, 1:1 in 24-96 hours, the tank pressure of a fermentation tank is 0.08Mpa, and the fermentation raw materials are stirred all the time after being put into the fermentation tank, wherein the stirring speed is 200 rpm;
and 4, checking the total number of bacteria of the fermentation broth produced by fermentation, filling and packaging to obtain a finished product.
2. The use method of the microbial inoculum prepared by the method for preparing the microbial inoculum for treating the hexavalent chromium pollution of the drainage basin according to claim 1, is characterized in that: the LYzhlm microbial inoculum is added into hexavalent chromium polluted water, the volume ratio of the microbial inoculum to the polluted water is 0.5L-3.0L: 100L, the pH value of the polluted water added with the microbial inoculum is controlled to be 5-9, the temperature is controlled to be 25-35 ℃, and the stirring speed is 80-140 r/min.
3. The use method of the microbial inoculum prepared by the microbial inoculum preparation method for treating the hexavalent chromium pollution of the drainage basin according to claim 2, is characterized in that: the weight-volume ratio of the lyszhlm bacterial agent to the polluted water is 2.0:100 g/ml.
4. The use method of the microbial inoculum prepared by the microbial inoculum preparation method for treating the hexavalent chromium pollution of the drainage basin according to claim 2, is characterized in that: the pH value of the polluted water added with the microbial inoculum is 7.0.
5. The use method of the microbial inoculum prepared by the microbial inoculum preparation method for treating the hexavalent chromium pollution of the drainage basin according to claim 2, is characterized in that: the temperature of the polluted water after the microbial inoculum is added is 30 ℃.
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Z. Chen et al..Thioredoxin is involved in hexavalent chromium reduction in Streptomyces violaceoruber strain LZ-26-1 isolated from the Lanzhou reaches of the Yellow River.《International Biodeterioration & Biodegradation》.2014,第94卷第146-151页. *

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