CN108251327B - Application of stenotrophomonas rhizophila DSM14405 in reduction of hexavalent chromium - Google Patents
Application of stenotrophomonas rhizophila DSM14405 in reduction of hexavalent chromium Download PDFInfo
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Abstract
The invention discloses an application of stenotrophomonas rhizophila DSM14405 in reduction of hexavalent chromium. The stenotrophomonas rhizophila DSM14405 survives in a slightly alkaline environment and is used for reducing hexavalent chromium into trivalent chromium, and the trivalent chromium is combined with hydroxyl to generate chromium hydroxide precipitate. The stenotrophomonas rhizophila DSM14405 of the invention has higher reduction speed on hexavalent chromium, the 32-hour reduction rate of the hexavalent chromium of 50mg/L basically can reach 100 percent, the tolerance concentration of the stenotrophomonas rhizophila DSM on the hexavalent chromium is higher, the thalli can grow when the concentration of the hexavalent chromium reaches 200mg/L, and the reduction rate in 48 hours can reach 47.6 percent.
Description
Technical Field
The invention relates to the technical field of biology, in particular to application of Stenotrophomonas rhizophila (Stenotrophoromonas rhizophila) DSM14405 in reduction of hexavalent chromium.
Background
Chromium salts are mainly used in metal processing and in tanning and electroplating industries. Chromate is also frequently added in industrial production in order to prevent the circulating water from corroding various equipments. In recent years, with the rapid development of economy in China, the consumption of chromium salt in industrial production is more and more, and the production waste residue is one of the most main heavy metal industrial pollution sources in the world at present. Therefore, chromium pollution in industrial waste water and exhaust gas is also becoming more serious. Chromium is mainly in two forms of trivalent chromium and hexavalent chromium in the environment, and the chemical behaviors and toxicity of the two forms are different. Wherein trivalent chromium is adsorbed on solid substances in water and deposited in the bottom mud, so that the harm is small; hexavalent chromium is mostly dissolved in water, is relatively stable, and has greater toxicity to human bodies and environmental hazards. Therefore, how to convert hexavalent chromium in wastewater into trivalent chromium which is easy to precipitate and less harmful is the key point of chromium pollution treatment at present.
In chromium pollution treatment, chemical and physical methods have the possibility of incomplete treatment or secondary pollution. Therefore, the microorganism treatment method is a popular method at present, and has the advantages of low treatment cost and no secondary pollution.
Disclosure of Invention
In view of the above, the invention aims to provide an application of stenotrophomonas rhizophila DSM14405 in reduction of hexavalent chromium, and the stenotrophomonas rhizophila DSM14405 can tolerate higher-concentration hexavalent chromium and reduce the hexavalent chromium into trivalent chromium, so that the content of the hexavalent chromium in soil and water is reduced, the risk of people for ingesting the hexavalent chromium is reduced, the absorption of plants to the chromium is reduced, and a technical method for microbial remediation of chromium pollution is enriched.
Based on the purpose, the invention provides the application of stenotrophomonas rhizophila DSM14405 in reduction of hexavalent chromium.
In some embodiments of the invention, the stenotrophomonas rhizophila DSM14405 survives in a more alkaline environment for reduction of hexavalent chromium to trivalent chromium, which combines with hydroxide to produce a chromium hydroxide precipitate.
The stenotrophomonas rhizophila DSM14405 can reduce hexavalent chromium in water into low-toxicity trivalent chromium through enzymatic reaction, and the stenotrophomonas rhizophila DSM14405 is suitable for living in a slightly alkaline environment, so that the trivalent chromium generated by the reduction reaction is combined with hydroxyl to generate chromium hydroxide precipitate, and the toxicity of chromium pollution in the environment can be further reduced.
In some embodiments of the invention, the stenotrophomonas rhizophila DSM14405 is used in the treatment of hexavalent chromium contaminated wastewater.
When repairing hexavalent chromium polluted soil, the stenotrophomonas rhizophila DSM14405 can be applied in the planting process of crops (such as rice, wheat, vegetables, fruits and the like), so that the content of hexavalent chromium in the soil can be effectively reduced, the absorption of chromium by plants is reduced, and the chromium is prevented from polluting food through edible organs of the plants.
In some embodiments of the invention, the stenotrophomonas rhizophila DSM14405 is used to remediate hexavalent chromium contaminated soil.
When the stenotrophomonas rhizophila DSM14405 disclosed by the invention is applied to treatment of hexavalent chromium polluted wastewater, the stenotrophomonas rhizophila DSM14405 can be inoculated in the hexavalent chromium polluted wastewater, and the stenotrophomonas rhizophila DSM14405 can reduce hexavalent chromium into trivalent chromium, so that the content of the hexavalent chromium is reduced, and the chromium polluted wastewater is repaired.
In some embodiments of the invention, the culture conditions of said stenotrophomonas rhizophila DSM14405 when reducing hexavalent chromium are: the temperature is 24-33 ℃, the pH is 6.0-8.0, the rotating speed is 120-210 rpm/min, and the concentration of hexavalent chromium is 10-100 mg/L.
In some embodiments of the invention, the culture conditions of said stenotrophomonas rhizophila DSM14405 when reducing hexavalent chromium are: the temperature is 30 ℃, the pH value is 7.5, the rotating speed is 180rpm/min, and the concentration of hexavalent chromium is 50 mg/L.
The stenotrophomonas rhizophila DSM14405 is purchased from American type culture Collection with the preservation number: ATCC BAA-473T。
The stenotrophomonas rhizophila DSM14405 has the advantages that the bacterial thallus is in a short rod shape and is arranged in a dispersed mode, the diameter of a bacterial colony is about 2-3mm, the bacterial colony is circular and yellow, the surface is smooth, and the edge is neat. Bacteria are acapsular, spore-free, gram-negative, and multiply by fission. The bacteria are heterotrophic bacteria, require oxygen in the growth process, do not require sunlight, have positive catalase reaction and negative oxidase reaction. The optimal growth temperature of the bacteria is about 30 ℃, and the optimal environment pH is about 7.0. The bacteria do not require the addition of growth factors and other nutrients during growth.
Compared with the prior art, the invention has the following beneficial effects:
the stenotrophomonas rhizophila DSM14405 has high reduction speed on hexavalent chromium, the 32-hour reduction rate of the hexavalent chromium of 50mg/L basically can reach 100 percent, the tolerance concentration of the stenotrophomonas rhizophila DSM14405 on the hexavalent chromium is high, the thalli can grow when the concentration of the hexavalent chromium reaches 200mg/L, and the reduction rate in 48 hours can reach 47.6 percent, so the stenotrophomonas rhizophila DSM14405 can be used for chromium pollution treatment.
Drawings
FIG. 1 shows stenotrophomonas rhizophila DSM14405 in culture medium Cr6+A comparison graph of growth curves at concentrations ranging from 0 to 1000 mg/L;
FIG. 2 shows stenotrophomonas rhizophila DSM14405 in culture medium Cr6+Cr with concentration in the range of 0-1000mg/L6+A reduction ratio comparison chart;
FIG. 3 shows stenotrophomonas rhizophila DSM14405 in culture medium Cr6+A comparison graph of growth curves at concentrations ranging from 0 to 200 mg/L;
FIG. 4 shows stenotrophomonas rhizophila DSM14405 in culture medium Cr6+Cr with a concentration in the range of 0-200mg/L6+A reduction ratio comparison chart;
FIG. 5 shows the different initial pH values for stenotrophomonas rhizophila DSM14405 in culture medium Cr6+The growth curve influence diagram at the concentration of 50 mg/L;
FIG. 6 shows the different initial pH values for stenotrophomonas rhizophila DSM14405 in culture medium Cr6+Cr at a concentration of 50mg/L6 +A reduction rate influence graph;
FIG. 7 shows the different rocking speeds of the rocking platforms for stenotrophomonas rhizophila DSM14405 in culture medium Cr6+The growth curve influence diagram at the concentration of 50 mg/L;
FIG. 8 shows the different rocking speeds of the rocking platforms for stenotrophomonas rhizophila DSM14405 in culture medium Cr6+Cr at a concentration of 50mg/L6+A reduction rate influence graph;
FIG. 9 shows different culture temperatures for stenotrophomonas rhizophila DSM14405 in culture medium Cr6+The growth curve influence diagram at the concentration of 50 mg/L;
FIG. 10 shows different culture temperatures for stenotrophomonas rhizophila DSM14405 in culture medium Cr6+Cr at a concentration of 50mg/L6+A reduction rate influence graph;
FIG. 11 shows the optimization of the various culture conditions for stenotrophomonas rhizophila DSM14405 in culture medium Cr6+Growth curve diagram and Cr at concentration of 50mg/L6+The reduction situation is shown in the figure, and the contrast is Cr without adding bacterial liquid6+The concentration changes.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below with reference to specific embodiments and the accompanying drawings.
The hexavalent chromium Cr (VI) described in the following examples is present in the form of potassium dichromate (K)2Cr2O7)。
Example 1: discovery of hexavalent chromium reduction performance of stenotrophomonas rhizophila DSM14405
(1) The strain source is as follows: stenotrophomonas rhizophila DSM14405 is purchased from american type culture collection with accession number: ATCC BAA-473T。
(2) Culture medium
Tryptone Soy Broth (TSB): 17g/L tryptone, 3g/L soyase, 2.5g/L glucose, 5.0g/L sodium chloride, 2.5g/L dipotassium hydrogen phosphate, and pH7.3 +/-0.2.
Tryptone soy agar medium (TSA): 17g/L tryptone, 3g/L soyase, 2.5g/L glucose, 5.0g/L sodium chloride, 2.5g/L dipotassium hydrogen phosphate, 15g/L agar and pH7.3 +/-0.2.
Culture media containing different concentrations of Cr (vi): cr (VI) with the concentration of 5000mg/L is taken as mother liquor, and the mother liquor with the corresponding amount is added into the TSB culture medium to prepare Cr (VI) culture media with different final concentrations.
(3) Identification of whether stenotrophomonas rhizophila DSM14405 has capability of reducing Cr (VI) and growth condition of stenotrophomonas rhizophila DSM14405
Preparation of seed solution of stenotrophomonas rhizophila DSM14405
A loop of stenotrophomonas rhizophila DSM14405 to TSA medium is picked from a thawed glycerol strain liquid tube by using an inoculating loop to perform streak isolation culture. After the plate grows out colonies with proper size within 48h, selecting one colony to inoculate into TSB culture solution, culturing at 30 ℃ for 24h at 180rpm/min as seed solution, and storing at 4 ℃ for a short period of time for later use.
② the growth and Cr (VI) reduction ability determination of stenotrophomonas rhizophila DSM14405
Preparing TSB culture medium containing Cr (VI) of 0mg/L, 10mg/L, 20mg/L, 50mg/L, 100mg/L, 500mg/L and 1000mg/L respectively. Adding 1% (volume fraction) of seed solution, culturing in a shaking table at 30 deg.C at 180rpm/min, and measuring bacterial density (OD600) and Cr (VI) concentration every 24 hr until culturing for 72 hr. As shown in FIG. 1, the results of the growth curves show that the density of the bacteria in the medium at each Cr (VI) concentration was substantially constant after 24 hours of culture, and the bacteria in the medium containing 100mg/L or less of Cr (VI) showed a small difference from the control group, and were able to grow normally. And the OD600 value of the bacteria in the culture medium of Cr (VI) 500mg/L and 1000mg/L is not more than 1 all the time within 72h of culture, so that the Cr (VI) concentration is possibly too high, and the normal growth of the bacteria is inhibited.
As for the results of the Cr (VI) reducing ability, as shown in FIG. 2, the reduction rate reached 100% in the medium in which the initial Cr (VI) concentration was 10mg/L and 20mg/L after 24 hours of the culture; the reduction rate of the culture medium with the initial Cr (VI) concentration of 50mg/L is 92.4%. By culturing for 48 hours, the reduction rate in the culture medium with the initial concentration of Cr (VI) of 50mg/L and 100mg/L also reaches 100 percent. However, the reduction rate in the medium with initial Cr (VI) concentrations of 500mg/L and 1000mg/L was maintained at a low level throughout the 72h period of the culture assay. As a result, in this example, stenotrophomonas rhizophila DSM14405 showed a good effect on the growth and reduction of Cr (VI) at a concentration of 100mg/L or less.
Example 2: determination of optimum hexavalent chromium reduction concentration of stenotrophomonas rhizophila DSM14405
According to example 1, it was confirmed that stenotrophomonas rhizophila DSM14405 is superior in growth and reduction effects at a Cr (VI) concentration of 100mg/L or less. In this example, TSB medium containing Cr (VI) of 0mg/L, 50mg/L, 100mg/L and 200mg/L respectively was prepared, seed solution of stenotrophomonas rhizophila DSM14405 was inoculated into the TSB medium, and optimum hexavalent chromium reduction concentration of stenotrophomonas rhizophila DSM14405 was determined based on the growth and reduction effects of stenotrophomonas rhizophila DSM14405 in the TSB medium. The method comprises the following specific steps:
preparation of seed solution of stenotrophomonas rhizophila DSM14405
Inoculating the bacterial liquid cultured for 24h into TSB culture solution in an amount of 1% (volume fraction), culturing at 30 deg.C at 180rpm/min, continuously monitoring bacterial biomass growth in the culture solution until OD600 value of the culture solution reaches 1.0, using as seed solution, and storing at 4 deg.C for short term use.
② the growth of stenotrophomonas rhizophila DSM14405 and the determination of the optimum hexavalent chromium reduction concentration range
Preparing TSB culture medium containing Cr (VI) of 0mg/L, 50mg/L, 100mg/L and 200mg/L respectively. Adding 1% seed solution, culturing in shaking table at 30 deg.C at 180rpm/min, and measuring bacterial density (OD600) and Cr (VI) concentration every 8 hr until culturing for 48 hr. As shown in FIG. 3, the results of the growth curves show that the density of the growth of the bacteria was normal in the medium containing 50mg/L of Cr (VI) after 48 hours of cultivation. The growth of the bacteria was inhibited in the medium containing Cr (VI) at concentrations of 100mg/L and 200 mg/L.
As for the results of the Cr (VI) reducing ability, as shown in FIG. 4, the reduction rate reached 100% in the medium in which the initial concentration of Cr (VI) was 50mg/L after 32 hours of cultivation; the reduction rate of the medium with the initial Cr (VI) concentration of 100mg/L and 200mg/L is not completely reduced when the medium is cultured for 48 hours. In conclusion, in the embodiment, the growth condition and the reduction effect of stenotrophomonas rhizophila DSM14405 are optimal when the Cr (VI) concentration is 50 mg/L.
Example 3: effect of environmental conditions on the hexavalent chromium reducing potential of stenotrophomonas radicophila DSM14405
From example 2, it is understood that the growth and hexavalent chromium reduction effects of stenotrophomonas rhizophila DSM14405 are good when the Cr (VI) concentration is 50mg/L, and therefore the concentration of Cr (VI) is 50mg/L as a concentration to be studied when the culture conditions are optimized in this example.
(1) Influence of initial pH
The pH value of the TSB culture medium is adjusted to five levels of 6.0, 6.5, 7.0, 7.5 and 8.0 when the Cr (VI) concentration is 50mg/L, the temperature is 30 ℃, and the rotating speed is 180 rpm/min. Adding 1% of seed solution with OD600 value of 1.0 into culture medium, culturing for 36h, and measuring OD600 value and Cr (VI) concentration every 12 h. The effect of the initial pH on the growth of stenotrophomonas rhizophila DSM14405 is shown in FIG. 5, and at pH6.0 and 6.5, stenotrophomonas rhizophila DSM14405 did not grow substantially within 36 h; the relative lag time of pH7.0 is longer, but the bacterial density value can reach a little difference with pH7.5 at 36 h; the cell density values in the media of pH7.5 and pH8.0 increased faster in culture at the level of normal growth, and the cell density value at Ph8.0 increased faster than at pH7.5 and higher by 36 h.
The effect of the initial pH on the ability of stenotrophomonas rhizophila DSM14405 to reduce Cr (VI) is shown in FIG. 6, and at 36h, the groups at pH7.5 and pH8.0 reach a reduction rate of 100%; the group reduction rate of pH7.0 is higher but not up to 100%; the reduction rates were lower for the groups at pH6.0 and pH 6.5. In addition, the group with pH7.5 has a higher Cr (VI) reducing speed than that of the group with pH8.0 in the whole detection period, and the group with pH7.5 has a growth amount less than that of the group with pH8.0 in the detection period, so the consumed substances are less, therefore, the initial pH of the group with pH7.5 is the optimal pH value for reducing Cr (VI) by stenotrophomonas rhizophila DSM 14405.
(2) Influence of the rotational speed of the rocking platforms
The initial pH of the culture medium is the pH value of the TSB culture medium without adjustment at the temperature of 30 ℃ and the Cr (VI) concentration of 50mg/L, and the rotating speeds of a shaking table are adjusted to four levels of 120rpm/min, 150rpm/min, 180rpm/min and 210 rpm/min. The seed solution with OD600 value of 1.0 is added into TSB culture medium according to the amount of 1% (volume fraction), and cultured for 36h, and the OD600 value and the Cr (VI) concentration are measured every 12 h. The effect of the shaker speed on the growth of stenotrophomonas rhizophila DSM14405 is shown in FIG. 7, and the growth of the bacteria at four speeds is relatively good, but the group growth at 180rpm/min and 210rpm/min is better than that of the other two groups.
The influence of the shaking table rotating speed on the Cr (VI) reducing capability of stenotrophomonas rhizophila DSM14405 is shown in figure 8, the reducing rate difference of four treatment groups is not large at 12h, and the reducing rate of the group with the shaking table rotating speed of 180rpm/min is slightly high; the data measured at 24h show that the group reduction rates at the rotation speeds of 180rpm/min and 210rpm/min are obviously higher than those of the other two groups, and the group reduction rate at the rotation speed of 210rpm/min is slightly higher than that of the group at the rotation speed of 180 rpm/min; at 36h, the reduction rate of the two groups of the rotating speeds of 180rpm/min and 210rpm/min reaches 100 percent, while the other two groups of the rotating speeds of 180rpm/min and 210rpm/min do not completely reduce Cr (VI), so that the rotating speeds of the shaking table of 180rpm/min and 210rpm/min are relatively more suitable for reducing Cr (VI) than the other two rotating speeds. In contrast, in the comparison between the rotation speeds of 180rpm/min and 210rpm/min, although the biological growth rate at the rotation speed of 210rpm/min is more than that at 180rpm/min, the difference between the Cr (VI) reduction rates at the two rotation speeds is not obvious. In conclusion, when the rotating speed of 180rpm/min is low in bacterial growth amount and consumed substances, the rotating speed of the shaking table is low, and the electric energy consumption is low, the reduction rate of Cr (VI) is close to that of 210 rpm/min. Therefore, the optimal rotation speed of the stenotrophomonas rhizophila DSM14405 for reducing Cr (VI) is determined to be 180rpm/min by combining feasibility and cost consideration in actual production.
(3) Effect of culture temperature
The initial pH of the culture medium is the pH value of the TSB culture medium without adjustment when the Cr (VI) concentration is 50mg/L and the rotation speed is 180rpm/min, and the culture temperature is adjusted to four levels of 24 ℃, 27 ℃, 30 ℃ and 33 ℃. The seed solution with OD600 value of 1.0 is added into the culture medium according to the amount of 1 percent (volume fraction), and the culture is carried out for 36 hours, and the OD600 value and the Cr (VI) concentration are measured every 12 hours. The effect of the culture temperature on the growth of stenotrophomonas rhizophila DSM14405 is shown in FIG. 9, the growth conditions of the bacteria at four culture temperatures are relatively good, but the group growth condition at the culture temperature of 30 ℃ is relatively better; the sudden increase in the density of bacteria measured at 36h in the group with an incubation temperature of 24 ℃ did not exclude the possibility of abnormalities.
The effect of the culture temperature on the Cr (VI) reducing ability of stenotrophomonas rhizophila DSM14405 is shown in FIG. 10, the reduction rates of the four treatment groups are relatively similar, but the reduction rate is a little higher than that of the other treatment groups when the culture temperature is 30 ℃, and the reduction rate of only the group Cr (VI) at the culture temperature of 30 ℃ reaches 100% when the culture is carried out for 36 h. In summary, the optimal temperature for the reduction of Cr (VI) by stenotrophomonas rhizophila DSM14405 was determined to be 30 ℃.
From examples 1-3, the optimal culture conditions for the stenotrophomonas rhizophila DSM14405 in the reduction of hexavalent chromium are: rotation speed of 180rpm/min, initial pH7.5, culture temperature of 30 ℃ and Cr (VI) concentration of 50mg/L, under the optimal culture conditions, stenotrophomonas rhizophila DSM14405 was cultured to obtain the growth curve and Cr (VI) reduction results as shown in FIG. 11, wherein the control is the Cr (VI) reduction rate without adding bacteria in the culture medium with Cr (VI) concentration of 50 mg/L. As can be seen from FIG. 11, after the optimization, 50mg/L Cr (VI) can be completely reduced within 28 hours, which is faster than the original 32 hours, while the reduction rate can only reach about 10% in the blank without adding bacteria. Cr (VI) in the blank without adding bacteria can also be reduced because Cr (VI) is an oxidant stronger than oxygen, and can oxidize organic matters in the culture medium at normal temperature, but the oxidation speed is slower. Therefore, in conclusion, it can be determined that stenotrophomonas rhizophila DSM14405 can promote the reduction speed of Cr (VI) so that Cr (VI) can be reduced to Cr (III) more quickly.
In conclusion, the stenotrophomonas rhizophila DSM14405 has a high hexavalent chromium reduction speed, the hexavalent chromium reduction rate of 50mg/L can reach 100% in 32 hours basically, the tolerance concentration of the stenotrophomonas rhizophila DSM14405 to the hexavalent chromium is high, the thalli can grow when the hexavalent chromium concentration reaches 200mg/L, and the reduction rate of 48 hours can reach 47.6%, so the stenotrophomonas rhizophila DSM14405 can be used for chromium pollution treatment.
Those of ordinary skill in the art will understand that: the discussion of any embodiment above is meant to be exemplary only, and is not intended to intimate that the scope of the disclosure, including the claims, is limited to these examples; within the idea of the invention, also technical features in the above embodiments or in different embodiments may be combined and there are many other variations of the different aspects of the invention as described above, which are not provided in detail for the sake of brevity. Therefore, any omissions, modifications, substitutions, improvements and the like that may be made without departing from the spirit and principles of the invention are intended to be included within the scope of the invention.
Claims (1)
1. The application of stenotrophomonas rhizophila DSM14405 in reduction of hexavalent chromium is characterized in that the stenotrophomonas rhizophila DSM14405 is used for reducing hexavalent chromium into trivalent chromium, and the trivalent chromium is combined with hydroxyl to generate chromium hydroxide precipitate;
using the stenotrophomonas rhizophila DSM14405 in hexavalent chromium polluted wastewater treatment;
using the stenotrophomonas rhizophila DSM14405 for repairing hexavalent chromium contaminated soil;
the stenotrophomonas rhizophila DSM14405 survives in a slightly alkaline environment, and the pH value of the environment is 7.5-8.0;
the culture conditions of the stenotrophomonas rhizophila DSM14405 in the reduction of hexavalent chromium are as follows: the temperature is 30 ℃, the pH value is 7.5, the rotating speed is 180rpm/min, and the concentration of hexavalent chromium is 50 mg/L.
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| CN110643551B (en) * | 2019-11-18 | 2022-03-15 | 吉林大学 | Stenotrophomonas rhizophila S11 for preventing and treating rice blast and application thereof |
| WO2021114784A1 (en) * | 2019-12-11 | 2021-06-17 | 山东科技大学 | Strain capable of tolerating high concentration of chromium, and microbial agent used for hexavalent chromium pollution control |
| CN111533585A (en) * | 2020-04-22 | 2020-08-14 | 中国科学院生态环境研究中心 | Application of stenotrophomonas rhizophila in improvement of rhizosphere soil and promotion of plant growth |
| CN112592850B (en) * | 2020-12-08 | 2022-06-21 | 北京农业生物技术研究中心 | Stenotrophomonas for promoting growth and development of lily and/or antagonizing lily pathogenic bacteria and application thereof |
| CN114480219B (en) * | 2022-03-15 | 2023-07-18 | 中南大学 | Microbial functional flora for improving content of ginkgolide in ginkgo leaves and preparation and application thereof |
| CN115287241B (en) * | 2022-09-15 | 2023-06-09 | 南昌航空大学 | Modified hexavalent chromium reducing bacteria and preparation method and application thereof |
| CN115805232B (en) * | 2022-11-29 | 2023-10-20 | 西南科技大学 | Application method of encapsulated bacteria for efficiently treating hexavalent chromium polluted soil |
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