CN116764157A - Method for degrading toluene by enzyme oxidation - Google Patents

Abstract

The invention discloses a method for enzymatic oxidative degradation of toluene, which includes: extracting complex enzymes from plants, immobilizing the obtained enzymes, and then using air as an oxidant to oxidatively degrade toluene using the immobilized enzymes. This method has simple operation, mild conditions, low requirements for equipment, high activity of the immobilized enzyme catalyst, fast reaction speed, large processing capacity, long service life of the catalyst, and the immobilized enzyme still has high activity after being used for more than 1000 hours. This method can efficiently degrade toluene and is an environmentally friendly process.

Description

A method for enzymatic oxidative degradation of toluene

Technical field

The invention belongs to the field of enzyme technology, and specifically relates to a method for catalyzing the oxidative degradation of toluene by an immobilized enzyme.

Background technique

With the continuous development of industrial technology, more and more volatile organic compounds are emitted in the air, which has caused great harm to the environment and people's health. Therefore, people have begun to focus on the degradation of volatile organic compounds. The current methods for treating volatile organic compounds include physical adsorption, biodegradation, chemical catalysis and other methods. Among them, the methods that can effectively degrade volatile organic compounds mainly include microorganisms, enzymatic methods, and light. Catalysis method, plasma catalysis method, etc. Patent CN108514886 A discloses a silver-based catalyst for photothermal synergistic catalytic degradation of toluene. The use of precious metal catalysts in this catalyst increases production costs. Patent CN 109806743 A discloses a method and system for treating toluene-containing waste gas, which uses ozone to oxidatively degrade toluene. Ozone will cause secondary pollution. Patent CN 110152670 A discloses a catalyst for photothermal catalytic degradation of toluene. The degradation rate of toluene is more than 91%, but the illumination temperature is 125°C, the reaction temperature is high, the energy consumption is large, and toxic by-products are easily produced. product. Patent CN 112999845 A discloses a method for degrading toluene using a plasma power source. This method requires specific equipment, has high requirements for equipment, and consumes high energy. The ozone generated during the degradation process will also cause secondary pollution; patent CN110079472 A discloses a culture medium for isolating toluene-degrading bacteria. The cultured toluene-degrading bacteria can efficiently degrade toluene, but the reaction speed of microorganisms in processing toluene is slow and is greatly affected by the environment.

Biological enzyme treatment method is a method that uses the catalytic oxidation reaction of biological enzymes to degrade volatile organic compounds. This method uses enzymatic reactions to form free radicals, which then open complex chemical chains in volatile organic compounds, ultimately producing small molecule substances. The biological enzyme treatment method has the advantages of high degradation efficiency, mild reaction conditions, low requirements on equipment conditions, fast reaction speed, and wide application range for temperature, concentration and toxic substances.

Contents of the invention

In view of the above existing technical problems, the present invention provides a method for oxidative degradation of toluene with an immobilized enzyme. After immobilizing a complex enzyme extracted from plants, the toluene is catalytically oxidized, which can enable efficient oxidation and degradation of toluene, with a degradation rate of 99.5%.

In order to achieve the above objects, the present invention adopts the following technical solutions:

A method for enzymatic oxidative degradation of toluene, comprising the following steps:

(1) Extraction of complex enzyme: Take the leaves of the plant, crush them and add phosphate buffer solution for extraction, then centrifuge, and the supernatant is the complex enzyme solution;

The plant is any two or a mixture of two or more of white palm, tobacco, licorice, pothos, aloe vera, Sansevieria, dwarf palm, fig, and rubber tree;

(2) Immobilization of enzyme: Soak the clean and dry immobilized carrier in the composite enzyme solution obtained in step (1), then filter and dry to obtain the immobilized enzyme;

(3) Oxidative degradation of toluene: Place the immobilized enzyme obtained in step (2) into a reactor, and pass in air containing toluene at 20~80°C for reaction.

Preferably, the crushing method described in step (1) is: adding liquid nitrogen to cool and grind.

Preferably, the concentration of the phosphate buffer solution described in step (1) is 0.1~0.3 mol/L, and the pH is 5.5~10.5.

Preferably, the mass-to-volume ratio of the leaves of the plant described in step (1) to the phosphate buffer solution is 1:10 (g/mL).

Preferably, the centrifugal speed described in step (1) is 10,000 rpm.

Preferably, the centrifugation time described in step (1) is 10 minutes.

Preferably, the immobilized carrier described in step (2) is alumina.

Preferably, the soaking time described in step (2) is

Preferably, the drying temperature in step (2) is 60°C.

Preferably, the concentration of toluene in the toluene-containing air described in step (3) is 500 ppm~3000 ppm, more preferably 500 ppm~2500 ppm, and most preferably 1000 ppm~2500 ppm.

Preferably, for the reaction described in step (3), the flow rate of the toluene-containing air is 50 mL/min~600 mL/min, more preferably 100 mL/min~500 mL/min, and most preferably 100 mL/min. min~300 mL/min.

Preferably, the reaction temperature in step (3) is 20 to 60°C, and most preferably 20 to 40°C.

The method for degrading toluene of the present invention has the following advantages:

The reaction conditions are mild and the requirements for equipment are low.

Immobilized enzymes have high catalytic activity, fast reaction speed, and are applicable to a wide range of temperatures, concentrations, and toxic substances.

(3) The processing capacity is large, the catalyst has a long service life, and the immobilized enzyme still has high activity after being used for more than 1000 hours.

(4) This process can efficiently oxidize and degrade toluene and is an environmentally friendly process.

Description of drawings

Figure 1 is a scanning electron microscope image of the alumina support before loading.

Figure 2 is a scanning electron microscope image of alumina loaded with immobilized enzyme.

Figure 3 is a scanning electron microscope image of alumina loaded with immobilized enzyme after oxidative degradation of toluene.

Figure 4 shows the adsorption and desorption isotherms before the alumina carrier is loaded with immobilized enzyme.

Figure 5 shows the adsorption and desorption isotherms after the alumina carrier is loaded with immobilized enzyme.

Figure 6 shows the BJH pore size distribution diagram before the alumina carrier is loaded with immobilized enzyme.

Figure 7 shows the BJH pore size distribution diagram after the alumina carrier is loaded with immobilized enzyme.

Figure 8 shows the metachrotron analysis chart of the immobilized enzyme.

Implementation

The present description will be explained in more detail below by means of examples, which are not intended to limit the scope of the invention.

Example

Cut and mix the leaves of white palm, tobacco, licorice, pothos, aloe, Sansevieria, dwarf palm, fig and rubber tree, add liquid nitrogen to cool and then grind, then remove 0.1g of powder, add 1mL, 0.2mol/ L, phosphate buffer solution with pH 7.5 for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 0.1 h, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 20°C. The flow rate of air containing toluene was 50 mL/min. The initial concentration of toluene was. After passing through the column once, the toluene degradation rate was 62.4%. .

Example

Cut and mix the leaves of aloe vera, Sansevieria orchid and rubber tree, add liquid nitrogen to cool and grind them, then remove 0.1g of powder, add 1mL, 0.1mol/L, phosphate buffer solution with pH 6.5 for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 10 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube, the reaction temperature was 20°C, the flow rate of air containing toluene was 200 mL/min, and the initial concentration of toluene was

Example

Cut and mix the leaves of white palm, licorice, pothos, aloe, dwarf palm, fig and rubber tree, add liquid nitrogen to cool and then grind, then remove 0.1g of powder, add 1mL, 0.3mol/L, pH is 8.5 Soak in phosphate buffer solution. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 0.5 h, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 25°C. The flow rate of air containing toluene was 150 mL/min. The initial concentration of toluene was 700 ppm. After passing through the column once, the degradation rate of toluene was obtained. 85.0%.

Example

Cut and mix the leaves of tobacco, licorice, aloe vera, dwarf palm and fig, add liquid nitrogen to cool and grind, then remove 0.1g of powder, add 1mL, 0.1mol/L, phosphate buffer solution with pH 10.5 for soaking . Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 1 hour, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube, the reaction temperature was 30°C, the flow rate of air containing toluene was 180 mL/min, and the initial concentration of toluene was 10

Example

Cut and mix the leaves of white palm, sansevieria, weeping fig and rubber tree, add liquid nitrogen to cool and grind, then remove 0.1g of powder, add 1mL, 0.2mol/L, pH 5.5 phosphate buffer solution for soaking . Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 2 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 35°C. The flow rate of air containing toluene was 200 mL/min. The initial concentration of toluene was 900 ppm. After passing through the column once, the degradation rate of toluene was obtained. 88.5%.

Example

Cut and mix the leaves of white palm, licorice, Sansevieria, dwarf palm, fig and rubber tree, add liquid nitrogen to cool and grind, then remove 0.1g of powder, add 1mL, 0.2mol/L, pH 7.5 Soak in phosphate buffer solution. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 2.5 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 20°C. The flow rate of air containing toluene was 250 mL/min. The initial concentration of toluene was 1000 ppm. After passing through the column once, the degradation rate of toluene was obtained. 91.3%.

Example

Cut and mix the leaves of tobacco, licorice, aloe, sansevieria, fig and rubber tree, add liquid nitrogen to cool and grind, then remove 0.1g of powder, add 1mL, 0.2mol/L, phosphate buffer with pH 7.5 Soak in the solution. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 3 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube, the reaction temperature was 40°C, the flow rate of air containing toluene was 100, and the initial concentration of toluene was 1500 ppm. After passing through the column once, the toluene degradation rate was 84.8%.

Example

Cut and mix the leaves of pothos, aloe, sansevieria and rubber tree, add liquid nitrogen to cool and then grind them, then remove 0.1g of powder, add 1mL, 0.2mol/L, phosphate buffer solution with pH 7.5 for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 3.5 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 20°C. The flow rate of air containing toluene was 300 mL/min. The initial concentration of toluene was 1800 ppm. After passing through the column once, the degradation rate of toluene was obtained. 92.6%.

Example

Cut and mix the leaves of tobacco, licorice, Sansevieria, dwarf palm and rubber tree, add liquid nitrogen to cool and grind, then remove 0.1g of powder, add 1mL, 0.2mol/L, phosphate buffer solution with pH 7.5. soak. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 4 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 45°C. The flow rate of air containing toluene was 350 mL/min. The initial concentration of toluene was 2000 ppm. After passing through the column once, the degradation rate of toluene was obtained. 91.3%.

Example

Cut and mix the leaves of white palm, licorice, aloe, Sansevieria, dwarf palm and fig. Add liquid nitrogen to cool and then grind. Then remove 0.1g of powder and add 1mL, 0.2mol/L, phosphoric acid with a pH of 7.5. Buffer solution for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 4.5 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 50°C. The flow rate of air containing toluene was 400 mL/min. The initial concentration of toluene was 2100 ppm. After passing through the column once, the degradation rate of toluene was obtained. 75.5%.

Example

Cut and mix the leaves of pothos, aloe vera and Sansevieria, add liquid nitrogen to cool and then grind them, then remove 0.1g of powder, add 1mL, 0.2mol/L, pH 7.5 phosphate buffer solution for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 5 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

Fill the immobilized enzyme into a glass reaction tube, the reaction temperature is 55°C, the flow rate of air containing toluene is 500 mL/min, and the initial concentration of toluene is 30

Example

Cut and mix the leaves of pothos and aloe vera, add liquid nitrogen to cool and then grind them. Then remove 0.1g of powder and add 1mL, 0.2mol/L, phosphate buffer solution with a pH of 7.5 for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 5.5 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 60°C. The flow rate of air containing toluene was 380 mL/min. The initial concentration of toluene was 2500 ppm. After passing through the column once, the degradation rate of toluene was obtained. 62.1%.

Example

Cut and mix the leaves of white palm, tobacco, pothos, aloe, dwarf palm, fig and rubber tree, add liquid nitrogen to cool and then grind, then remove 0.1g of powder, add 1mL, 0.2mol/L, pH is 7.5 Soak in phosphate buffer solution. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 6 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 65°C. The flow rate of air containing toluene was 6. The initial concentration of toluene was 2600 ppm. After passing through the column once, the toluene degradation rate was 58.7%.

Example

Cut and mix the leaves of tobacco, sansevieria and fig. Then add liquid nitrogen to cool and grind them. Then remove 0.1g of powder and add 1mL, 0.2mol/L, phosphate buffer solution with pH 7.5 for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 6.5 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 70°C. The flow rate of air containing toluene was 450 mL/min. The initial concentration of toluene was 2700 ppm. After passing through the column once, the degradation rate of toluene was obtained. 49.6%.

Example

After cutting and mixing the leaves of white palm and pothos, add liquid nitrogen to cool and grind, then remove 0.1g of powder, add 1mL, 0.2mol/L, phosphate buffer solution with pH 7.5 for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 7 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 75°C. The flow rate of air containing toluene was 550 mL/min. The initial concentration of toluene was 2800 ppm. After passing through the column once, the degradation rate of toluene was obtained. 43.7%.

Example

Cut the leaves of tobacco and dwarf palm into pieces, mix them, add liquid nitrogen to cool them and then grind them. Then remove 0.1g of the powder and add 1mL, 0.2mol/L, phosphate buffer solution with a pH of 7.5 for soaking. Then centrifuge at 10,000 rpm for 10 min, and take the supernatant as the enzyme stock solution.

Wash the alumina carrier with deionized water, shake it in ultrasonic for 20 minutes to remove surface impurities, and then dry it in a drying oven; completely immerse the pretreated alumina carrier in the enzyme stock solution, soak for 8 hours, and filter. It was then dried at 60°C to obtain the immobilized enzyme.

The immobilized enzyme was filled into a glass reaction tube. The reaction temperature was 80°C. The flow rate of air containing toluene was 600 mL/min. The initial concentration of toluene was 3000 ppm. After passing through the column once, the degradation rate of toluene was obtained. 40.1%.

Claims (9)

1. A method for degrading toluene by enzymatic oxidation, which is characterized by comprising the following steps:

(1) Extraction of complex enzyme: pulverizing leaves of plants, adding phosphate buffer solution for extraction, and centrifuging to obtain supernatant as complex enzyme solution;

the plant is any two or more than two of radix et rhizoma Nardostachyos, tobacco, glycyrrhrizae radix, scindapsus aureus, aloe, herba Polygoni Cuspidati, petiolus Trachycarpi, ficus benincasae and rubber tree;

(2) Immobilization of enzymes: soaking the clean and dried immobilized carrier in the complex enzyme solution obtained in the step (1), and then filtering and drying to obtain immobilized enzyme;

(3) Oxidative degradation of toluene: and (3) placing the immobilized enzyme obtained in the step (2) into a reactor, and introducing air containing toluene at 20-80 ℃ for reaction.

2. The method of claim 1, wherein the comminuting in step (1) is by: adding liquid nitrogen, cooling, and grinding.

3. The method according to claim 1, wherein the concentration of the phosphate buffer solution in the step (1) is 0.1-0.3 mol/L and the pH is 5.5-10.5.

4. The method of claim 1, wherein the mass to volume ratio of leaf to phosphate buffer solution of the plant in step (1) is 1:10 (g/mL).

5. The method of claim 1, wherein the immobilization support of step (2) is preferably alumina.

6. The method of claim 1, wherein the temperature of the drying in step (2) is 60 ℃.

7. The method according to claim 1, wherein the toluene concentration in the toluene-containing air in step (3) is 500 ppm to 3000 ppm, more preferably 500 ppm to 2500 ppm, most preferably 1000 ppm~2500 ppm.

8. The method according to claim 1, wherein the flow rate of the air containing toluene in step (3) is 50 mL/min to 600 mL/min, more preferably 100 mL/min to 500 mL/min, most preferably 100 mL/min to 300 mL/min.

9. The method according to claim 1, wherein the temperature of the reaction in step (3) is 20-60 ℃, most preferably 20-40 ℃.